Clinical trial NCT04221230
Conditions: Major Depressive Disorder
Keywords: NMRA-335140 Neumora NMRA-140 Navacaprant
Sponsor: Neumora Therapeutics, Inc.
Start date: 2020-01-03; Completion date: 2022-06-23
Enrollment: 204 participants; Allocation: RANDOMIZED; Intervention model: PARALLEL

Brief summary:
A proof of concept (POC) study evaluating the impact of NMRA-335140 (BTRX-335140) relative to placebo on symptoms of major depressive disorder (MDD) in adult participants with MDD and symptoms of anhedonia and anxiety following 8 weeks of double-blind treatment as assessed by the HAMD-17 Scale.

Detailed description:

Arms: arm 1: NMRA-335140 will be administered. arm 2: Placebo
Interventions: intervention 1: NMRA-335140 intervention 2: Placebo
Intervention descriptions: intervention 1: Active Drug intervention 2: Placebo

Protocol documents:
Prot_000.pdf: 
BTRX-335140 
BlackThorn Therapeutics, Inc. 
Clinical Study Protocol Amendment 7 
Date: 24Jan2022 
 
Confidential 
Page 1 of 88 
 
 
CLINICAL STUDY PROTOCOL: K2-MDD-201 
 
 
Protocol Title: 
A Phase 2a, Randomized, Double-blind, Placebo-
controlled Proof of Concept Study to Evaluate the 
Effects of Oral BTRX-335140 Versus Placebo in 
Subjects With Major Depressive Disorder 
Brief Title: 
Proof of Concept Phase 2a, Double-blind Study in 
Major Depressive Disorder with BTRX-335140 vs 
Placebo 
Investigational Product: 
BTRX-335140 
Phase of Development: 
Phase 2a 
Sponsor: 
BlackThorn Therapeutics, Inc. 
1700 Owens Street, Suite 535 
San Francisco, CA 940158 
Telephone: 866-258-5277 
Medical Monitor: 
, 
 
Executive Medical Director 
Medical Affairs 
Worldwide Clinical Trials 
Telephone: 
 
Medical Project Leader: 
, 
 
VP, Clinical Development 
BlackThorn Therapeutics, Inc. 
Telephone: 
 
Amendment 7 
(Version/Effective Date): 
8.0/24 January 2022 
 
The information contained in this document, particularly unpublished data, is the property or under 
control of BlackThorn Therapeutics, Inc., and is provided to you in confidence as an investigator, 
potential investigator or consultant, for review by you, your staff, and an applicable Institutional 
Review Board or Independent Ethics Committee. The information is only to be used by you in 
connection with an authorized clinical study of the investigational drug described in the protocol. You 
will not disclose any of the information to others without written authorization from BlackThorn, 
except to the extent necessary to obtain informed consent from those persons to whom the drug may 
be administered. 
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BTRX-335140 
BlackThorn Therapeutics, Inc. 
Clinical Study Protocol Amendment 7 
Date: 24Jan2022 
 
Confidential 
Page 2 of 88 
 
SPONSOR SIGNATURE 
Protocol Title: 
A Phase 2a, Randomized, Double-blind, Placebo-controlled Proof 
of Concept Study to Evaluate the Effects of Oral BTRX-335140 
Versus Placebo in Subjects With Major Depressive Disorder 
Protocol Number: 
K2-MDD-201 
 
 
This Protocol Amendment 7, Version 8.0 has been reviewed and approved by the Sponsor. 
 
 
 
Company/Sponsor signatory 
, 
 
VP Clinical Development 
BlackThorn Therapeutics, Inc. 
Tel:
e-mail:
 
Date 
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BTRX-335140 
BlackThorn Therapeutics, Inc. 
Clinical Study Protocol Amendment 7 
Date: 24Jan2022 
 
Confidential 
Page 3 of 88 
 
INVESTIGATOR AGREEMENT 
Protocol Title: 
A Phase 2a, Randomized, Double-blind, Placebo-controlled 
Proof of Concept Study to Evaluate the Effects of Oral 
BTRX-335140 Versus Placebo in Subjects With Major 
Depressive Disorder 
Protocol Number: 
K2-MDD-201 
 
 
This Protocol Amendment 7, Version 8.0 has been reviewed and approved by the Sponsor. 
 
 
The signature below constitutes agreement with this protocol and attachments, and provides 
assurance that this study will be conducted according to all stipulations of the protocol, 
including all statements regarding confidentiality, and according to national and local legal 
and regulatory requirements and applicable regulations and guidelines. 
 
Principal Investigator: 
 
Address: 
 
Signature: 
 
Date: 
 
 
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BTRX-335140 
BlackThorn Therapeutics, Inc. 
Clinical Study Protocol Amendment 7 
Date: 24Jan2022 
 
Confidential 
Page 4 of 88 
 
SUMMARY OF CHANGES 
This section describes significant changes made in Version 8.0 of the protocol. Changes to 
punctuation, formatting (ie, changes in numbering, word order, location, etc.), the table of 
contents, and the list of abbreviations and definitions of terms are not listed in the summary 
of changes table (Table 1). 
Table 1: 
Summary of Changes 
Section Number and Title 
Description of Change 
Rationale 
Synopsis – Statistical Methods, 
Section 9 Statistical 
Considerations, Efficacy 
Analyses 
Changed alpha, power, and 
confidence interval (CI) for 
efficacy analyses to reflect 2-sided 
alpha=0.05; power of 84% and 
95% CIs 
To decrease the chance of a Type I 
error. 
Synopsis – Study Design, 
Schedule of Procedures and 
Assessments (footnotes c, h, and 
t), Section 3.1, Section 6.3.6, 
Section 7.6, Section 7.7.2, 
Section 7.8 
Clarified wording around the Week 
12 corneal specular microscopy, 
which should be completed for all 
subjects (ie, completers and early 
termination) unless consent is 
withdrawn. Also added 
clarification that additional 
ophthalmologic examinations 
(standard ophthalmologic exams 
and/or corneal specular 
microscopies) may be conducted at 
any time during the study to assess 
subject safety. 
In the event of early termination, 
every attempt should be made to 
get a subject to come back for 
corneal specular microscopy at 
12 weeks after the first dose of 
study drug.  
Added text to reflect potential of 
unscheduled ophthalmologic and/or 
corneal specular microscopies to 
assess safety  
Section 3.2.2, Section 3.2.3, 
Section 9.3.3.2, Section 9.3.5 
Moved CGI-S from an exploratory 
to a secondary endpoint. 
Supports the secondary objectives; 
consistency with the SAP 
Synopsis – Statistical Methods, 
Section 9.3.3.2 
Added description for analysis of 
CGI-S using generalized 
estimating equations (GEE) 
Consistency with the SAP 
Synopsis – Statistical Methods, 
Section 9.3.3.2 
Revised statistical text for 
secondary efficacy analyses to 
allow for ANCOVA as 
appropriate. 
Clarity and consistency with the 
SAP 
Synopsis – Study Endpoints, 
Section 3.2.2, Section 9.3.3.2, 
Section 9.3.5 
Moved HAMD-17 response rates 
from an exploratory endpoint to a 
secondary endpoint  
Consistency with the SAP 
Section 9.3.3.2 
Added bullet for CGI-I 
Inadvertently missed 
Section 9.3.6 
Clarified text describing subgroup 
analyses 
Consistency with the SAP 
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BTRX-335140 
BlackThorn Therapeutics, Inc. 
Clinical Study Protocol Amendment 7 
Date: 24Jan2022 
 
Confidential 
Page 5 of 88 
 
SYNOPSIS 
Name of Finished Product: BTRX-335140 
Protocol Number: K2-MDD-201 
Phase of Development: 2a 
Protocol Title: A Phase 2a, Randomized, Double-blind, Placebo-controlled Proof of Concept Study to 
Evaluate the Effects of Oral BTRX-335140 Versus Placebo in Subjects With Major Depressive Disorder 
Number of Planned Subjects: Approximately 180 subjects randomized with treatment 
Study Sites: Approximately 40 study centers in the United States (US) 
Duration of Study: Approximately 24 months 
Study Objectives: 
Primary Objectives: 
To establish Proof of Concept (POC) for BTRX-335140, a kappa opioid receptor antagonist, by evaluating 
the impact of BTRX-335140 relative to placebo on symptoms of major depressive disorder (MDD) in adult 
subjects with MDD and symptoms of anhedonia and anxiety following 8 weeks of double-blind treatment as 
assessed by the Hamilton Depression Rating Scale (HAMD-17) 
Secondary Objectives: 
1. To evaluate the effects of BTRX-335140 on self-reported anhedonia in adult subjects with MDD 
2. To evaluate the effects of BTRX-335140 on anxiety-related symptoms in adult subjects with MDD 
3. To evaluate the effects of BTRX-335140 on functional impairment in adult subjects with MDD 
4. To evaluate the safety and tolerability of BTRX-335140 in adult subjects with MDD 
Study Design: Study K2-MDD-201 is an 8-week, randomized, double-blind, placebo-controlled, 
parallel-group, multicenter study to evaluate the effects of BTRX-335140 on symptoms of depression in 
adult subjects with MDD and symptoms of anhedonia and anxiety after 8 weeks of double-blind treatment. 
The study will consist of a 7- to 28-day screening period, an 8-week active treatment period (during which 
subjects will receive either BTRX-335140 or placebo), and a 4-week Safety Follow-up period. 
Subjects will sign an informed consent form (ICF) and will then enter the screening period to ensure they 
meet the required inclusion and exclusion criteria, including diagnosis and stability of depressive symptoms 
and absence of psychiatric and medical conditions that would preclude study participation. The 28-day 
screening period may be extended to 35 days (after consultation with and approval by the medical monitor), 
if necessary, for reasons including, but are not limited to, discontinuation of a subject’s current inefficacious 
medications or to obtain an ophthalmologic examination. 
In the event that an investigator requests to re-screen a subject that previously screen failed 
Study K2-MDD-201, the medical monitor and Sponsor will consider allowing re-screening on a case-by-case 
basis. 
After eligibility is confirmed by the medical monitor, and before randomization, baseline ophthalmologic 
examinations (standard and corneal specular microscopy) must be performed. Subjects who meet the 
Diagnostic and Statistical Manual of Mental Disorders Fifth Edition (DSM5) diagnostic criteria for MDD 
and all other eligibility criteria for the study will be randomized in a 1:1 ratio to 1 of 2 treatment arms 
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BTRX-335140 
BlackThorn Therapeutics, Inc. 
Clinical Study Protocol Amendment 7 
Date: 24Jan2022 
 
Confidential 
Page 6 of 88 
 
(placebo or BTRX 335140 once daily [QD]) until approximately 180 subjects are randomized. Beginning at 
Visit 2 (Baseline), subjects will receive treatment with BTRX-335140 80 mg QD or placebo for 8 weeks. 
During the Safety Follow-up, all subjects will receive a telephone call from study site personnel 
approximately 2 weeks after discontinuation of study drug for assessment of AEs, concomitant medication 
usage, and CSSRS; and at Week 12, subjects will have corneal specular microscopy performed by a study 
site affiliated ophthalmologist. The overall duration in the study for each subject is expected to be 
approximately 14 to 16 weeks. Additional ophthalmologic examinations (standard ophthalmologic exams 
and/or corneal specular microscopies) may be conducted to monitor subject safety. 
Possible Changes in Conduct of Study: Study sites will maintain, at a minimum, coronavirus disease 2019 
(COVID-19) risk mitigation procedures as per regulatory and local guidelines and/or laws. Allowed 
modifications to the conduct of the study, in the event of site staff and/or patient restrictions due to 
COVID-19, are outlined in Appendix 1 to the protocol. 
Duration of Treatment: 8 weeks 
Study Population: 
Inclusion criteria: 
Subjects are eligible to be included in the study only if they meet all the following criteria:  
1. Are adult men or women 18 to 65 years of age (inclusive) at informed consent 
2. Have a primary DSM-5 diagnosis of MDD, with prominent symptoms of anhedonia confirmed by 
Structured Clinical Interview for DSM-5 Disorders, Clinical Trials Version (SCID-5-CT) 
a. 
The current episode must have started at least 3 weeks prior to screening but no more than 
12 months before the screening visit. 
b. Have not failed 2 or more courses of antidepressant treatment in the current episode  
c. 
No more than a 3-point change in HAMD-17 between screening and baseline 
d. Have sufficient history or an independent report to confirm that symptoms are causing functional 
impairment or clinically significant distress  
3. Meet the blinded-rule list based on clinical scale criteria 
4. Body mass index (BMI) of 18 to 40 kg/m2 (inclusive) 
5. Medically stable (in the opinion of the investigator and Sponsor/Sponsors delegate) based on medical 
history, vital signs, clinical laboratory tests, and 12-lead electrocardiogram (ECG) performed at 
screening and baseline 
6. Agree to the following birth control: 
a. 
Nonvasectomized men must agree to use a condom with spermicide, if sexually active during the 
study, until 90 days after the last dose of study drug administration. No restrictions are required for 
a vasectomized man, provided his vasectomy was performed 4 months or more prior to the first 
dose of study drug. A man who has been vasectomized less than 4 months prior to the first dose of 
study drug must follow the same restrictions as a nonvasectomized man. Additionally, men must 
refrain from sperm donation during study treatment and for at least 90 days following the last dose 
of study drug. 
b. Women of child-bearing potential (women not surgically sterilized and between menarche and 2 
years postmenopausal) must have a negative serum pregnancy test at screening and a negative urine 
pregnancy test at enrollment and agree to use reliable birth control (eg, oral contraceptives or 
Norplant®; a reliable double barrier method of birth control [diaphragms with contraceptive jelly; 
cervical caps with contraceptive jelly; condoms with contraceptive foam]; intrauterine devices; 
partner with vasectomy; or abstinence) during the study and for 10 days following the last dose of 
the study drug (BTRX-335140 or placebo). Women will be considered surgically sterile, if they 
have had tubal ligation, bilateral salpingo oophorectomy, or a hysterectomy. 
 
Note: Abstinence will be allowed if, in the investigator’s judgement, it is determined that the subject 
is reliable, that abstinence is the preferred and usual lifestyle of the subject, and that abstinence will 
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BTRX-335140 
BlackThorn Therapeutics, Inc. 
Clinical Study Protocol Amendment 7 
Date: 24Jan2022 
 
Confidential 
Page 7 of 88 
 
be continued for the duration of the study including the 10 days (women) or 90-day period (men) 
following last dose of study drug as noted above. 
c. 
Or engaged exclusively in a non-heterosexual relationship 
7. Willing and able to give written informed consent to participate 
8. Able to understand and comply with instructions in English 
9. Are judged by the investigator to be reliable and agree to keep all appointments for clinic visits, tests, 
and procedures, including venipuncture, and examinations required by the protocol 
Exclusion Criteria: 
Subjects will be excluded from the study if they meet any of the following criteria: 
1. Have a history of any of the following DSM-5 disorders within the specified timeframe: 
a. 
Currently or in the past year: diagnosis of personality disorder, attention deficit disorder/attention 
deficit hyperactivity disorder, anorexia nervosa, or bulimia nervosa. Subjects with comorbid 
generalized anxiety disorder, social anxiety disorder, or panic disorder for whom MDD is 
considered the primary diagnosis are not excluded. 
b. Lifetime: diagnosis of bipolar 1 or 2, schizophrenia, obsessive compulsive disorder, or post-
traumatic stress disorder 
2. Have a history of substance or alcohol use disorder (AUD), per DSM-5 criteria, within the past year 
3. Are actively suicidal (eg, any suicide attempts within the past 12 months) or are at serious suicidal risk 
as indicated by any current suicidal intent, including a plan, as assessed by the Columbia Suicide 
Severity Rating Scale (C-SSRS) (score of “YES” on suicidal ideations item 4 or 5 within 3 months prior 
to Visit 1 [Screening]) and/or based on clinical evaluation by the investigator; or are homicidal, in the 
opinion of the investigator 
4. Have a history or signs of Cushing’s disease, Addison’s Disease, primary amenorrhea or other evidence 
of significant disorders of the hypothalamus-pituitary-adrenal axis 
5. Have any other clinically significant medical or psychiatric condition or circumstance prior to 
randomization that, in the opinion of the investigator, or Sponsor could affect subject safety, preclude 
evaluation of response, interfere with the ability to comply with study procedures, or prohibit completion 
of the study, such as acute stress disorder, adjustment disorder, impulse control disorder, uncontrolled 
diabetes mellitus, renal or hepatic impairment, coronary artery disease, evidence of significant active 
cardiac, respiratory, or hematologic disease, cancer with <5-year remission (basal cell carcinoma is not 
excluded), chronic pain, fibromyalgia, gastric bypass, lap band placement, or any other significant 
gastrointestinal condition 
6. Have had prior seizures (other than remote history of childhood febrile seizure) or other condition that 
would place the subject at increased risk of seizures or is taking anticonvulsants for seizure control 
7. Have a history of serious head injury (eg, skull fracture, cerebral contusion, or trauma resulting in 
prolonged unconsciousness), intracranial neoplasm, or hemorrhage 
8. Have ever had electroconvulsive treatment, vagal nerve stimulation, or treatment with ketamine or 
esketamine for MDD 
9. Have initiated transcranial magnetic stimulation, psychotherapy (such as Cognitive Behavioral Therapy) 
or have had a change in psychotherapy, or other non-drug therapies (such as acupuncture or hypnosis) 
within 4 weeks prior to Visit 1 (Screening) or at any time during the acute phase of the study 
10. Have a visual or physical motor impairment that could interfere with subject’s ability to perform study 
assessments, as assessed by the investigator 
11. Have alanine aminotransferase (ALT) or aspartate aminotransferase (AST) levels ≥2 x upper limit of 
normal (ULN) or a bilirubin level 1.5 x ULN unless due to a documented history of Gilbert’s syndrome 
12. Estimated glomerular filtration rate (eGFR) ≤60 mL/min/1.73m2 as calculated by the Chronic Kidney 
Disease Epidemiology Collaboration [CKD-EPI] 2009 creatinine equation at Visit 1 (Screening) 
13. Positive hepatitis C virus (HCV) antibody (Ab), hepatitis B surface antigen (HBs Ag), hepatitis A virus 
(HAV) IgM antibody (HAV-Ab [IgM]) or human immunodeficiency virus (HIV) test at Visit 1 
(Screening) 
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BTRX-335140 
BlackThorn Therapeutics, Inc. 
Clinical Study Protocol Amendment 7 
Date: 24Jan2022 
 
Confidential 
Page 8 of 88 
 
14. Have a thyroid-stimulating hormone (TSH) level of <0.9 x lower limit of normal (LLN) or >1.2 x ULN 
on or off stable treatment for hyperthyroidism or hypothyroidism; if TSH is abnormal, evaluate reflex 
Free T3 and Free T4. If reflex testing is normal, the assessment of normal thyroid function will be 
determined based on the judgement of the investigator, following discussion with the medical monitor. 
15. Have any other clinically significant abnormalities (significant would include laboratory deviations 
requiring acute medical intervention or further medical evaluation) in laboratory results at screening, 
including clinical chemistries, hematology, and urinalysis, and any clinical information that, in the 
judgment of the investigator or Sponsor, should preclude a subject’s participation at study entry 
16. Exclusionary ECG abnormalities obtained at Visit 1 (Screening) or Visit 2 (Baseline) are QT interval 
corrected using Fridericia’s formula (QTcF) >450 msec in males or >470 msec in females, complete 
bundle branch block, evidence of myocardial infarction or ischemia, and predominantly nonsinus 
conducted rhythms. Other abnormalities can be exclusionary at the discretion of the principal 
investigator or medical monitor. See Section 6.3.5 of protocol for guidance on ECG interpretations. 
17. Have a positive urine drug screen for amphetamines, barbiturates, cocaine, methadone, opioids, 
propoxyphene, tetrahydrocannabinol (THC), phencyclidine, or a positive blood alcohol level assessed by 
breathalyzer at Visit 1 (Screening) and Visit 2 (Baseline). For occasional (1 to 2 times per month 
maximum) cannabis users only, 1 retest is allowed and subject must agree to abstain from use for the 
duration of the study; a positive second test is exclusionary. 
18. Have any use, by history, of Salvinorin A 
19. Use of the following concomitant medications (contact the Sponsor-designated medical monitor to 
determine eligibility when in doubt):  
a. 
Psychoactive medication including stimulants, benzodiazepines and anxiolytics, oral antipsychotics, 
mood stabilizers/anticonvulsants (carbamazepine, lamotrigine, etc.), lithium, antidepressants, 
S-adenosylmethionine, melatonin, agomelatine, and hypnotics/sedatives within 5 half-lives or 
14 days (whichever is longer) of Visit 2 (Baseline) 
b. Fluoxetine and irreversible monoamine oxidase inhibitors within 4 weeks of Visit 2 (Baseline) 
depot antipsychotics within 2 months of Visit 2 (Baseline) 
c. 
Opioid agonists and antagonists 
20. Are currently taking or have taken within 5 half-lives of Visit 2 (Baseline) any medications or 
supplements 
 
 
21. Are women who are either pregnant or breastfeeding 
22. Have participated (received study treatment) in a clinical study or any other type of medical research 
judged by the investigator or Sponsor to be scientifically or medically incompatible with this study 
within 30 days prior to Visit 1 (Screening). Contact the Sponsor-designated medical monitor to 
determine eligibility when in doubt. 
23. Have participated in multiple interventional clinical studies, such that, in the opinion of the investigator, 
the subject is not a suitable candidate for participation 
24. Have previously completed or withdrawn from this study or any other study investigating 
BTRX-335140 
25. Are investigator site personnel directly affiliated with this study, and/or their immediate families. 
Immediate family is defined as a spouse, parent, child, or sibling, whether biological or legally adopted. 
26. Are employees of the Sponsor or are employees of any third-party organizations (TPOs) (eg, laboratory 
staff, study vendors and transportation providers) involved in study who require exclusion of their 
employees 
27. Has any of the following: 1) useful vision in only 1 eye from a pre-existing ophthalmic disease or 
amblyopia; 2) a corneal transplant in either eye; 3) corneal dystrophy or family history of corneal 
dystrophy; 4) severe dry eye syndrome [keratitis sicca]; 5) will not or cannot cooperate with ophthalmic 
examination requiring pupillary dilation (includes history of severe adverse reaction to mydriatic agents 
or untreated narrow angle glaucoma). Note: The following ocular disorders are allowed: cataracts, prior 
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BTRX-335140 
BlackThorn Therapeutics, Inc. 
Clinical Study Protocol Amendment 7 
Date: 24Jan2022 
 
Confidential 
Page 9 of 88 
 
cataract surgery, glaucoma (narrow angle glaucoma is allowed if definitively treated with laser 
peripheral iridectomy), macular degeneration, or ocular changes associated with diabetes mellitus or 
multiple sclerosis. 
Investigational Product, Dose, and Mode of Administration: A starting dose of BTRX-335140 80 mg 
(1 capsule) QD at Visit 2 (Baseline), orally and this dose will be maintained for the subsequent 8 weeks.  
Reference Therapy, Dose, and Mode of Administration: Placebo will be assigned as 1 capsule QD at 
Visit 2 (Baseline), orally and will be maintained for the subsequent 8 weeks. Placebo capsules will consist of 
inactive ingredients and look identical to BTRX-335140.  
Study Assessments:  
Efficacy: 
x 
HAMD-17 
x 
Clinical Global Impression Scale – Severity (CGI-S) 
x 
Clinical Global Impression of Improvement (CGI-I) 
x 
Snaith-Hamilton Pleasure Scale (SHAPS) 
x 
Hamilton Anxiety Rating Scale (HAM-A) 
x 
Hospital Anxiety and Depression Scale (HADS) subscales (ie, anxiety subscale [HADS-A] and 
depression subscale [HADS-D]) 
x 
Sheehan Disability Scale (SDS) 
x 
x 
x 
Safety: Safety assessments include adverse events (AEs), serious adverse events (SAEs), adverse events of 
special interest (AESIs), vital signs, weight and BMI, clinical laboratory tests, physical, ECG, and 
ophthalmologic examinations (standard and corneal specular microscopy), C-SSRS, and Clinical Opiate 
Withdrawal Scale (COWS). 
 
will also 
be obtained; all assessments are optional 
 
Study Endpoints: 
Primary Efficacy: The primary efficacy endpoint is the change from baseline to Week 8 on the HAMD-17 
score in the Efficacy population. 
Secondary: Secondary endpoints will include the following:  
x 
CGI-I score assessed at each postbaseline timepoint 
x 
Change from baseline to each timepoint assessed in SHAPS score 
x 
Change from baseline to each timepoint assessed in HADS subscales (ie, HADS-A and HADS-D) 
scores 
x 
Change from baseline to each timepoint assessed in HAM-A total score 
x 
Change from baseline to each timepoint assessed in SDS score 
x 
Change from baseline to end of study in CGI-S scores 
x 
HAMD-17 response rate 
x 
Assessment of safety through the occurrence of treatment-emergent AEs (TEAEs), SAEs, and 
AESIs; change from baseline in vital signs measurements, body weights, BMI, clinical laboratory 
test results, C-SSRS scores, COWS scores; physical examination findings, ECG results, and 
ophthalmologic examination findings 
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BlackThorn Therapeutics, Inc. 
Clinical Study Protocol Amendment 7 
Date: 24Jan2022 
 
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Page 10 of 88 
 
All efficacy endpoints will also be summarized by visit, including change from baseline information. 
Statistical Methods: All statistical comparisons will be made at a 2-sided alpha of 0.05. No adjustment for 
multiplicity is planned. SAS Version 9.4 or higher will be used to analyze data.  
Subjects will be analyzed according to randomized treatment assignment for efficacy assessments and 
according to treatment received for safety assessments. 
Primary Efficacy Analysis: The primary efficacy analysis is a comparison of mean reduction in HAMD-17 
scores between BTRX-335140 and placebo at Week 8. The analysis will be conducted using a Mixed-
Models Repeated Measures (MMRM) with change from baseline HAMD-17 score as the dependent variable 
and will include treatment group, Week, and Week by treatment group interaction as factors, and baseline 
HAMD-17 score as covariate. Variance estimation will be based on an unstructured covariance matrix. The 
model-based Week 8 least square (LS) means, standard errors, 95% confidence intervals (CIs), and p-values 
will be reported. 
Secondary Efficacy Analysis: Comparisons between BTRX-335140 versus placebo for the change from 
baseline in score at each timepoint assessed for the secondary endpoints will be conducted using MMRM 
analyses as specified above or analysis of covariance (ANCOVA), as appropriate. For CGI-I an additional 
analysis comparing the percentage of subjects with responses of ‘much improved’ or ‘very much improved’ 
at each timepoint between BTRX-335140 and placebo will be performed using generalized estimating 
equations (GEE). Response rates for decrease in HAMD-17 (e.g., the percentage of subjects with ≥ 50% 
decrease in HAMD-17 from baseline to Week 4 and Week 8) will be compared between BTRX-335140 and 
placebo using Cochran-Mantel-Haenszel tests. 
 
Safety Analyses: Safety data, including AEs, safety laboratory results, physical examination results, vital 
signs, suicidality, COWS scores, ECGs, and ophthalmologic examination findings will be summarized by 
treatment group and/or listed. 
 
 
 
 
 
 
 
 
 
 
 
 
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BlackThorn Therapeutics, Inc. 
Clinical Study Protocol Amendment 7 
Date: 24Jan2022 
 
Confidential 
Page 11 of 88 
 
SCHEDULE OF PROCEDURES AND ASSESSMENTS 
Possible Changes in Conduct of Study: Study sites will maintain, at a minimum 
coronavirus disease 2019 (COVID-19) risk mitigation procedures as per regulatory and local 
guidelines and/or laws. Allowed modifications to the conduct of the study, if needed, are 
outlined in Appendix 1 to the protocol. 
 
Period 
Screening 
Baseline 
Study Drug Treatment 
Safety Follow-up  
 
Visit 
1b 
2 
3 
4 
5 
6 
TCa 
Ocular 
Exam 
 
Days –7 
to –28 
Day 1 
Day 15 
(±2 d) 
Day 29 
(±2 d) 
Day 43 
(±2 d) 
Day 57/ 
EOTc 
(±2 d) 
Procedure and 
Assessments 
NA 
Week 0 
Week 2 
Week 4 
Week 6 
Week 8 
Week 10 
Week 12 
Informed consent 
X 
 
 
 
 
 
 
 
Inclusion/exclusion 
criteria 
X 
Xd 
 
 
 
 
 
 
Medical/psychiatric 
history 
X 
 
 
 
 
 
 
 
Demographics 
X 
 
 
 
 
 
 
 
Vital signse 
X 
X 
X 
X 
X 
X 
 
 
Physical examinationf 
X 
 
 
 
 
 
 
 
Body weightg 
X 
X 
 
X 
 
X 
 
 
12-Lead ECG 
X 
X 
 
X 
 
X 
 
 
Standard 
ophthalmologic 
examinationh 
 
Xi, j 
 
 
 
X 
 
 
Corneal specular 
microscopyh 
 
Xi 
 
 
 
 
 
Xt 
Prior/concomitant 
medication/therapy 
X 
X 
X 
X 
X 
X 
X 
 
Randomization 
 
X 
 
 
 
 
 
 
Clinical laboratory tests 
(hematology, 
chemistry, urinalysis)  
X 
X 
X 
X 
X 
X 
 
 
Serum/urine pregnancy 
testk 
X 
X 
X 
X 
X 
X 
 
 
UDS (onsite and central 
laboratory) and 
breathalyzer alcohol 
screen 
X 
X 
X 
X 
X 
X 
 
 
 
 
 
 
Thyroid function tests  
X 
 
 
 
 
 
 
 
Viral serology 
X 
 
 
 
 
 
 
 
Hepatic safety testingn 
 
SCID-5-CT  
X 
 
 
 
 
 
 
 
CGI-S  
X 
X 
 
 
 
X 
 
 
CGI-I  
 
X 
X 
X 
X 
X 
 
 
C-SSRSo 
X 
X 
X 
X 
X 
X 
X 
 
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Period 
Screening 
Baseline 
Study Drug Treatment 
Safety Follow-up  
 
Visit 
1b 
2 
3 
4 
5 
6 
TCa 
Ocular 
Exam 
 
Days –7 
to –28 
Day 1 
Day 15 
(±2 d) 
Day 29 
(±2 d) 
Day 43 
(±2 d) 
Day 57/ 
EOTc 
(±2 d) 
Procedure and 
Assessments 
NA 
Week 0 
Week 2 
Week 4 
Week 6 
Week 8 
Week 10 
Week 12 
HAMD-17 
X 
X 
 
X 
 
X 
 
 
HAM-A  
X 
X 
 
X 
 
X 
 
 
 
 
SDS 
 
X 
 
X 
X 
X 
 
 
HADS 
 
X 
X 
X 
X 
X 
 
 
SHAPS 
X 
X 
 
X 
 
X 
 
 
 
 
 
 
 
 
COWS 
X 
X 
X 
X 
X 
X 
 
 
Adverse eventsp 
 
X 
X 
X 
X 
X 
X 
 
Complete IWRS 
X 
X 
X 
X 
X 
X 
 
 
Dispense study drug 
 
Xq 
Xq 
Xq 
Xq 
 
 
 
Study drug 
administration 
 
Xr 
 
Xs 
 
 
 
 
Study drug compliance 
 
 
X 
X 
X 
X 
 
 
BMI = body mass index; CRF=case report form; CGI-I = Clinical Global Impression of Improvement; CGI-S = Clinical 
Global Impression of Severity; COWS = Clinical Opiate Withdrawal Scale; C-SSRS = Columbia Suicide Severity Rating 
Scale; d = day; DSM = Diagnostic and Statistical Manual of Mental Disorders; ECG = electrocardiogram; eCRF=electronic 
case report form; EOT = End of Treatment; HADS = Hospital Anxiety and Depression Scale; HAM-A = Hamilton Anxiety 
Rating Scale; HAMD-17 = Hamilton Rating Scale for Depression – 17-Item Version; IWRS = Interactive Web-response 
System;
; NA = not applicable; 
 
 SDS = Sheehan Disability Scale; SCID-5-CT = Structured Clinical Interview for DSM-5 
Disorders, Clinical Trials Version; SHAPS = Snaith-Hamilton Pleasure Scale;
TC = telephone call; UDS=urine drug screen. 
a 
Study site personnel will contact subjects by telephone for Safety Follow-up assessments (adverse events, concomitant 
medication usage, and C-SSRS) approximately 2 weeks after discontinuation of study drug. 
b 
Visit 1 procedures may be completed over 2 days, if necessary. 
c 
Subjects who discontinue early will complete an EOT visit <2 days after their last dose of study drug; efficacy 
assessments are to be performed only if the subject has remained on study drug. For subjects who do not return, the site 
should complete a C-SSRS if they are aware of a potential suicide-related thought or behavior by other 
communications. The 12-week corneal specular microscopy examination should still be conducted approximately 12 
weeks after the first dose of study drug (Section 7.7.2).  
d 
Inclusion/exclusion criteria must be reconfirmed for eligibility prior to randomization. 
e 
Collect vital signs measurements (respiratory rate first, then blood pressure and pulse) after subject has been supine for 
10 minutes; subjects should then stand for 3 minutes before obtaining standing vital signs measurements [see 
Section 6.3.4]. Oral temperature can be assessed in any position. 
f 
A full physical examination involving an evaluation of cardiovascular, respiratory, gastrointestinal, neurological 
(examination of cranial nerves, motor system, sensory, and reflexes), dermatological, and musculoskeletal systems, and 
include general appearance, skin, head, neck, eyes, ears, nose, throat, lungs, heart, abdomen, back, lymph nodes, 
extremities, and vascular status will be conducted at the screening visit. Further physical examination will be conducted 
as needed. (Section 6.3.3). 
g 
At Screening, collect height for BMI calculation. 
h 
Ophthalmologic examinations will be performed by a site-affiliated ophthalmologist. Details for the standard 
ophthalmologic examination and corneal specular microscopy will be described in a separate ocular monitoring 
manual.  Additional standard ophthalmologic exams and / or corneal specular microscopies may be conducted to assess 
subject safety. 
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i 
Baseline ophthalmologic examinations (standard and corneal specular microscopy) must be performed before 
randomization. 
j 
Prior to Visit 3 (Week 2) (between Days 7 and 14 [+2 days]), a standard ophthalmologic examination will be 
performed. 
k 
Serum pregnancy test is to be performed at screening visit only and urine pregnancy tests will be performed by the site 
at all subsequent visits. 
l 
 
 
 
m 
Optional assessment; subject refusal to perform will not prohibit study participation. 
n 
Unscheduled hepatic monitoring testing may be performed at any time on the basis of clinical laboratory testing results, 
after consultation with the Sponsor-designated medical monitor (see Section 6.3.2.2). 
o 
The C-SSRS Baseline version will be used at Screening and “Since Last Visit” version will be used for all subsequent 
visits. 
p 
Adverse events reported before first dose of study drug to be recorded as medical history. Serious AEs will be recorded 
on the AE eCRF from the time of informed consent.  
q 
Subjects will be instructed to take their study drug daily in the morning and preferably with food. 
r 
Subjects will take their first dose of study drug (BTRX-335140 or placebo) under the supervision of clinical study 
personnel during the baseline visit. 
s 
 
t 
The Week 12 corneal specular microscopy should be completed for all subjects (ie, completers and early termination) 
unless consent is withdrawn. The corneal specular microscopy should be conducted approximately 12 weeks after the 
first dose of study drug; exceptions to be discussed with the medical monitor.  
  
 
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Date: 24Jan2022 
 
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TABLE OF CONTENTS 
CLINICAL STUDY PROTOCOL: K2-MDD-201 ...................................................................1 
SPONSOR SIGNATURE ..........................................................................................................2 
INVESTIGATOR AGREEMENT.............................................................................................3 
SUMMARY OF CHANGES .....................................................................................................4 
SYNOPSIS .................................................................................................................................5 
SCHEDULE OF PROCEDURES AND ASSESSMENTS .....................................................11 
TABLE OF CONTENTS .........................................................................................................14 
LIST OF TABLES ...................................................................................................................18 
LIST OF FIGURES .................................................................................................................18 
LIST OF APPENDICES ..........................................................................................................18 
LIST OF ABBREVIATIONS ..................................................................................................19 
1 
INTRODUCTION .................................................................................................22 
1.1 
Background Information ..................................................................................22 
1.1.1 
Major Depressive Disorder ..................................................................22 
1.1.2 
Treatments............................................................................................22 
1.1.3 
BTRX-335140......................................................................................23 
1.2 
Rationale for Study Design, Doses, and Control Groups ................................25 
1.3 
Potential Risks and Benefits ............................................................................27 
2 
OBJECTIVES ........................................................................................................28 
2.1 
Primary Objective ............................................................................................28 
2.2 
Secondary Objectives.......................................................................................28 
 
 
3 
INVESTIGATIONAL PLAN AND ENDPOINTS ...............................................29 
3.1 
Description of Study Design ............................................................................29 
3.2 
Study Endpoints ...............................................................................................30 
3.2.1 
Primary .................................................................................................30 
3.2.2 
Secondary .............................................................................................30 
 
 
3.3 
Study Duration .................................................................................................31 
4 
STUDY ENROLLMENT AND WITHDRAWAL ...............................................32 
4.1 
Study Population ..............................................................................................32 
4.1.1 
Inclusion Criteria .................................................................................32 
4.1.2 
Exclusion Criteria ................................................................................33 
4.2 
Method of Assigning Subjects to Treatment ...................................................36 
4.3 
Blinding............................................................................................................36 
4.4 
Subject Withdrawal ..........................................................................................36 
4.5 
Early Termination of Subjects From the Study ...............................................38 
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5 
STUDY TREATMENTS .......................................................................................39 
5.1 
Description of Treatments................................................................................39 
5.2 
Manufacturing, Packaging, and Labeling ........................................................39 
5.3 
Storage .............................................................................................................39 
5.4 
Study Drug Administration ..............................................................................39 
5.5 
Study Drug Compliance ...................................................................................40 
5.6 
Study Drug Accountability ..............................................................................40 
5.7 
Prior and Concomitant Therapy .......................................................................40 
6 
STUDY PROCEDURES AND ASSESSMENTS .................................................42 
6.1 
Medical History and Demographics ................................................................42 
6.1.1 
Medical History ...................................................................................42 
6.1.2 
Demographics and Baseline Characteristics ........................................42 
6.2 
Efficacy Assessments.......................................................................................43 
6.2.1 
Hamilton Rating Scale for Depression – 17-item Version ..................43 
6.2.2 
Snaith Hamilton Pleasure Scale ...........................................................43 
6.2.3 
Clinical Global Impression Scale – Severity .......................................43 
6.2.4 
Clinical Global Impression Scale – Improvement ...............................43 
6.2.5 
Hamilton Anxiety Rating Scale ...........................................................43 
6.2.6 
 
6.2.7 
Hospital Anxiety and Depression Scale ...............................................44 
6.2.8 
 
6.2.9 
Sheehan Disability Scale......................................................................44 
6.2.10 
 
6.2.11 
6.3 
Safety Assessments ..........................................................................................45 
6.3.1 
Adverse Events ....................................................................................45 
6.3.2 
Clinical Laboratory Tests .....................................................................45 
6.3.2.1 
Sample Collections .....................................................................45 
6.3.2.2 
Hepatic Safety and Enhanced Hepatic Testing ...........................47 
6.3.2.3 
Urine Drug Screen ......................................................................48 
6.3.2.4 
6.3.2.5 
6.3.3 
Physical Examinations .........................................................................50 
6.3.4 
Vital Signs ............................................................................................50 
6.3.5 
Electrocardiography .............................................................................50 
6.3.6 
Ophthalmologic Examinations.............................................................51 
6.3.7 
Suicidal Ideation/Suicidality ................................................................52 
6.3.8 
Opiate Withdrawal ...............................................................................52 
6.4 
 
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7 
TIMING OF PROCEDURES AND ASSESSMENTS .........................................54 
7.1 
Screening Period (Day –28 to Day–7) .............................................................54 
7.2 
Visit 2: Baseline (Day 1)..................................................................................55 
7.3 
Visit 3: Week 2 (Day 15) .................................................................................57 
7.4 
Visit 4: Week 4 (Day 29) .................................................................................57 
7.5 
Visit 5: Week 6 (Day 43) .................................................................................59 
7.6 
Visit 6: Week 8 (Day 57)/End of Treatment ....................................................59 
7.7 
Safety Follow-up ..............................................................................................61 
7.7.1 
Telephone Call (Week 10) ...................................................................61 
7.7.2 
Ocular Examination (Week 12) ...........................................................61 
7.8 
Early Termination ............................................................................................61 
8 
SAFETY MONITORING AND REPORTING.....................................................62 
8.1 
Adverse Events ................................................................................................62 
8.1.1 
Definition of an Adverse Event ...........................................................62 
8.1.2 
Definition of a Serious Adverse Event ................................................62 
8.1.3 
Definition of Suspected Adverse Reaction ..........................................63 
8.1.4 
Definition of Serious Suspected Adverse Reaction .............................63 
8.2 
Classification of Adverse Events .....................................................................63 
8.2.1 
Severity of Adverse Events ..................................................................63 
8.2.2 
Relationship to Study Drug ..................................................................64 
8.3 
Time Period and Frequency for Event Assessment and Follow-up .................64 
8.4 
Adverse Events of Special Interest ..................................................................65 
8.5 
Reporting Procedures .......................................................................................65 
8.5.1 
Reporting Serious Adverse Events and Adverse Events of Special 
Interest..................................................................................................65 
8.5.2 
Regulatory Reporting Requirements ....................................................66 
8.5.3 
Overdose ..............................................................................................67 
8.5.4 
Pregnancy Reporting ............................................................................67 
8.6 
Safety Oversight...............................................................................................67 
9 
STATISTICAL CONSIDERATIONS...................................................................68 
9.1 
General Considerations ....................................................................................68 
9.2 
Analysis Populations ........................................................................................68 
9.2.1 
Safety Population .................................................................................68 
9.2.2 
Efficacy Population ..............................................................................68 
9.2.3 
Per-protocol Population .......................................................................68 
9.2.4 
 
9.3 
Statistical Analysis Methods ............................................................................69 
9.3.1 
Disposition ...........................................................................................69 
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9.3.2 
Demographics and Baseline Characteristics ........................................69 
9.3.3 
Efficacy Analyses ................................................................................69 
9.3.3.1 
Primary Efficacy Analysis ..........................................................69 
9.3.3.2 
Secondary Efficacy Analyses .....................................................69 
9.3.4 
Safety Analysis ....................................................................................70 
9.3.4.1 
Adverse Events ...........................................................................70 
9.3.4.2 
Adverse Events of Special Interest .............................................70 
9.3.4.3 
Vital Signs ...................................................................................70 
9.3.4.4 
Physical Examinations ................................................................70 
9.3.4.5 
Clinical Laboratory Tests ............................................................71 
9.3.4.6 
Electrocardiography ....................................................................71 
9.3.4.7 
Ophthalmology ...........................................................................71 
9.3.4.8 
Columbia Suicide Severity Rating Scale ....................................71 
9.3.4.9 
Clinical Opiate Withdrawal Scale ...............................................71 
9.3.4.10 
Treatment Compliance with Study Drug ....................................71 
9.3.5 
 
9.3.6 
Subgroup Analyses ..............................................................................72 
9.4 
Interim Analysis ...............................................................................................72 
9.5 
 
10 
QUALITY ASSURANCE AND QUALITY CONTROL .....................................74 
11 
REGULATORY AND ETHICAL CONSIDERATIONS .....................................75 
11.1 
Regulatory Considerations ...............................................................................75 
11.2 
Ethical Conduct of the Study ...........................................................................75 
11.3 
Institutional Review Board Approval ..............................................................75 
11.4 
Informed Consent Process ...............................................................................75 
11.5 
Confidentiality .................................................................................................76 
12 
STUDY ADMINISTRATION...............................................................................77 
12.1 
Clinical Monitoring ..........................................................................................77 
12.2 
Source Documents and Record Retention .......................................................77 
12.3 
Management of Protocol Amendments and Deviations ..................................77 
12.3.1 
Protocol Modifications.........................................................................78 
12.3.2 
Protocol Violations and Deviations .....................................................78 
12.4 
Financial Disclosure.........................................................................................78 
12.5 
Stopping Criteria: Suspension or Termination of Study or Investigational 
Site ...................................................................................................................78 
12.6 
Publication and Information Disclosure Policy ...............................................79 
13 
REFERENCES ......................................................................................................80 
 
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LIST OF TABLES 
Table 1: 
Summary of Changes ...................................................................................4 
Table 2: 
Clinical Laboratory Tests ...........................................................................46 
Table 3: 
Hepatic Monitoring Tests ..........................................................................48 
 
LIST OF FIGURES 
Figure 1: 
Study Design ..............................................................................................29 
 
LIST OF APPENDICES 
Appendix 1: 
Guidance for Virtual Visits (Via Telephone).............................................84 
Appendix 2: 
 ....87 
 
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LIST OF ABBREVIATIONS 
Abbreviation 
or Term 
Definition 
Ab 
antibody 
ADD 
attention deficit disorder  
ADHD 
attention deficit hyperactivity disorder 
AE 
adverse event 
ALT 
alanine aminotransferase 
APA 
American Psychiatric Association 
AESI(s) 
adverse event(s) of special interest 
AST 
aspartate aminotransferase 
ANCOVA 
analysis of the covariance 
AUC 
area under the curve 
AUD 
alcohol use disorder 
BMI 
body mass index 
BP 
blood pressure 
CAT 
Clinical Assessment Technology 
CFR 
Code of Federal Regulations 
cGCP 
current Good Clinical Practice 
CGI-I 
Clinical Global Impression of Improvement 
CGI-S 
Clinical Global Impression of Severity 
CI 
confidence interval 
CIOMS 
Council for International Organizations of Medical Sciences 
CKD-EPI 
Chronic Kidney Disease Epidemiology Collaboration 
Cmax 
maximum plasma drug concentration 
CNS 
central nervous system 
COVID-19 
coronavirus disease 2019  
COWS 
Clinical Opiate Withdrawal Scale 
CREB 
cAMP response binding protein 
CRF 
case report form 
CRO 
Contract Research Organization 
C-SSRS 
Columbia Suicide Severity Rating Scale 
DSM-5 
Diagnostic and Statistical Manual of Mental Disorders Fifth Edition 
DSM-IV-TR 
Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition, Text 
Revision 
EC 
Ethics Committee 
ECG 
electrocardiogram, electrocardiographic 
eCRF 
electronic case report form 
EDC 
electronic data capture͒ 
eGFR 
estimated glomerular filtration rate 
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Abbreviation 
or Term 
Definition 
FDA 
Food and Drug Administration 
GAD 
generalized anxiety disorder 
GEE 
generalized estimating equations 
HADS 
Hospital Anxiety and Depression Scale 
HADS-A 
Hospital Anxiety and Depression Scale – Anxiety Subscale 
HADS-D 
Hospital Anxiety and Depression Scale – Depression Subscale 
HAM-A 
Hamilton Anxiety Rating Scale 
HAMD-17 
Hamilton Depression Rating Scale – 17-item Version 
HAV 
hepatitis A virus 
HAV-Ab (IgM)  hepatitis A virus antibody 
HBs Ag 
hepatitis B surface antigen 
HCV 
hepatitis C virus 
HIPAA 
Health Insurance Portability and Accountability Act 
HIV 
human immunodeficiency virus 
HPMC 
hydroxypropyl methylcellulose 
IB 
investigator’s brochure 
IC50 
half-maximal inhibitory concentration 
ICC 
intraclass correlation coefficient 
ICF 
informed consent form 
ICH 
International Council for Harmonisation of Technical Requirements for 
Pharmaceuticals for Human Use 
IEC 
Independent Ethics Committee 
IgM 
immunoglobulin M 
IND 
US Investigational New Drug Application 
IRB 
Institutional Review Board 
IWRS 
interactive web-response system 
KOR 
kappa opioid receptor 
KORA 
kappa opioid receptor antagonist 
LC-MS/MS 
liquid chromatography-tandem mass spectrometry  
LLN 
lower limit of normal 
LS Means 
least-square means 
MedDRA 
Medical Dictionary for Regulatory Activities 
MDD 
major depressive disorder 
MMRM 
mixed-models repeated measures 
NOAEL 
no-observed-adverse-effect-level 
PDF 
portable document format 
PET 
positron emission tomography 
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Abbreviation 
or Term 
Definition 
 
 
 
 
POC 
proof of concept 
PP 
per-protocol 
PTSD 
post-traumatic stress disorder 
QD 
once daily 
QTcF 
Fridericia’s correction method for QT Interval 
REB 
Research Ethics Board 
RO 
receptor occupancy 
SAE 
serious adverse event 
SAP 
statistical analysis plan 
SCID-5-CT 
Structured Clinical Interview for DSM-5 Disorders, Clinical Trials Version 
SD 
standard deviation 
SDS 
Sheehan Disability Scale 
SHAPS 
Snaith-Hamilton Pleasure Scale 
SIGH-A 
Structured Interview Guide for the Hamilton Anxiety Rating Scale 
SIGH-D 
Structured Interview Guide for the Hamilton Depression Rating Scale 
SNRI 
serotonin norepinephrine reuptake inhibitor 
SOP 
Standard Operating Procedure 
SSRI 
selective serotonin reuptake inhibitor 
 
 
SUSAR 
serious unexpected suspected adverse reaction 
 
 
TC 
telephone call 
TEAE 
treatment-emergent adverse event 
THC 
tetrahydrocannabinol 
TPO 
third-party organization 
TSH 
thyroid-stimulating hormone 
UDS 
urine drug screen 
ULN 
upper limit of normal 
US 
United States 
WHO 
World Health Organization 
WHODrug 
World Health Organization Drug classification 
 
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1 
INTRODUCTION 
1.1 
Background Information 
1.1.1 
Major Depressive Disorder 
Major depressive disorder (MDD) is the most common mood disorder and imposes 
considerable economic and humanitarian suffering, such as decreased quality of life, 
functional impairment, and increased mortality rate. Currently, MDD is the leading cause of 
disability and afflicts approximately 322 million people worldwide (4.4% of the global 
population), with prevalence increasing 18% between 2005 and 2015 (World Health 
Organization [WHO] 2017). By 2020, depressive disorders are expected to be the second 
highest cause of morbidity in the world (Murray and Lopez 2006) and have been predicted to 
become the leading cause of disease burden by the year 2030 (WHO 2004). The lifetime 
prevalence of MDD is approximately 16.6% in the United States (US) (Kessler et al 2003). 
Episodes of MDD are often chronic and recurrent, with a relapse chance rate of 55% to 90% 
for individuals who experienced one or two prior depressions. More than 80% of the 
individuals who experience a second episode and who are not treated will experience a third 
episode within 3 years (Thase and Sullivan 1995). 
A clinical diagnosis of MDD is made based on the continuous presence of at least 5 of 
9 symptoms over at least 2 weeks. One of these symptoms, as defined in the Diagnostic and 
Statistical Manual of Mental Disorders, 5th Edition (DSM-5), must be either depressed mood 
or anhedonia, which is defined as diminished interest or pleasure in response to rewarding 
stimuli (APA 2013). This diagnostic classification assigns equal importance to depressed 
mood and anhedonia thus highlighting anhedonia as a core feature of the disorder. The 
classification system also acknowledges that anhedonia may be present in a subset of 
individuals with MDD. Estimates of significant anhedonia among patients with MDD are 
rare; 1 report describes an anhedonia rate of approximately 37% among a sample of 
65 individuals diagnosed with MDD (Pelizza and Ferrari 2009). Moreover, published data 
indicate that the presence of anhedonia may represent a distinct physiologically based subset 
of patients with MDD, as evidence from functional neuroimaging assessments indicates that 
anhedonia is linked to disruptions in central reward system functioning and is an important 
aspect of depression pathophysiology (Keedwell et al 2005). This is an important finding 
from the standpoint of therapeutic interventions because the presence of anhedonic 
symptoms has been found to be associated with poorer longitudinal symptom progression 
among patients with depression (Morris et al 2009) and is a major predictor of suicide 
(Fawcett et al 1990; Winer et al 2014). 
1.1.2 
Treatments 
The current mainstays of pharmacological treatments for depression include the use of 
selective serotonin reuptake inhibitors (SSRIs) (Fava et al 2007; Zupancic and Guilleminault 
2006) and serotonin-norepinephrine reuptake inhibitors (SNRIs) (Stahl et al 2005), which 
provide reasonable treatment options; however, these therapies are not effective in all 
patients, and are often associated with undesirable side effects, such as weight gain and 
sexual dysfunction. The lack of efficacy and the adverse events (AEs) associated with the use 
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of these antidepressants lead to high levels of treatment discontinuation (Zajecka 2000). Even 
with multiple consecutive treatments, only a small proportion of patients remain 
asymptomatic (Rush 2007). Thus, there continues to be a substantial unmet medical need for 
new antidepressants with greater response rates and improved tolerability. 
The presence of anhedonia is associated with difficulty in treating MDD. Research findings 
indicate that available therapies such as SSRIs do not target depression-related motivational 
and reward-processing deficits sufficiently (APA 2000; Dunlop and Nemeroff 2007; McCabe 
et al 2010; Nutt et al 2007; Price et al 2009; Shelton and Tomarken 2001) and that anhedonic 
symptoms predict inadequate treatment response (Spijker et al 2001). 
The presence of anhedonia is associated with inadequate treatment response to antidepressant 
drugs (McMakin et al 2012; Uher et al 2012) and potentially also to psychological treatments 
(Craske et al 2016). Unfortunately, research findings indicate that available therapies such as 
SSRIs do not target depression-related motivational and reward-processing deficits 
sufficiently (APA 2000; Dunlop and Nemeroff 2007; McCabe et al 2010; Nutt et al 2007; 
Price et al 2009; Shelton and Tomarken 2001). For these reasons, recent efforts have been 
directed towards developing therapies that effectively target anhedonia, such as ketamine and 
Positive Affect Treatment (Craske et al 2016; Lally et al 2014). 
1.1.3 
BTRX-335140 
BTRX-335140 is a selective kappa opioid receptor (KOR) antagonist that is being evaluated 
for the treatment of neurobehavioral disorders. Research studies carried out in animals and a 
proof of mechanism study carried out in humans suggest that medications which block KORs 
have the potential for being effective new treatments for patients with mood and anxiety 
spectrum disorders (Krystal et al 2020). The KOR system is an important mediator of the 
negative effects of stress-induced alterations on mood, emotional regulation, and motivation. 
This system has been a target for drug development in neurobehavioral disorders where 
dysregulation of stress signaling is associated with increased vulnerabilities. It is thought that 
KOR antagonists may improve anhedonia and reduce the vulnerabilities associated with 
stress. 
In laboratory animals, stress or repeated exposure to drugs of abuse triggers intracellular 
events involving cAMP response binding protein (CREB) in the nucleus accumbens – an 
area of the mesolimbic dopamine system known to be involved in reward and motivation 
(Carlezon and Krystal 2016). This increased CREB activity leads to elevated expression of 
the opioid peptide dynorphin (Bruchas et al 2009), which in preclinical models causes core 
symptoms of depression and anxiety. In preclinical models of depression and anxiety such as 
the forced swim test or tail suspension test, KOR antagonism blocks the biochemical and 
behavioral response to stress, leading to antidepressant and anxiolytic behavioral effects 
(Carlezon and Krystal 2016). In preclinical studies of addiction behavior, KOR antagonists 
prevent stress-induced drug-seeking behavior (Beardsley et al 2005). 
In humans, KOR agonists, including the plant-derived Salvinorin A, have been reported to 
trigger symptoms of dysphoria and anxiety (Pfeiffer et al 1986; Walsh et al 2001) suggesting 
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that antagonists may play a beneficial role in the treatment of anxiety disorders and 
anhedonic symptoms in depression. 
In the recently completed NIMH FAST-MAS Phase 2 Proof of concept (POC) study, patients 
with mood and anxiety disorders (Diagnostic and Statistical Manual of Mental Disorders, 
Fourth Edition, Text Revision [DSM-IV-TR] diagnostic criteria for: MDD, Bipolar 1 or 2 
Depressed, Generalized Anxiety Disorder [GAD], Social Phobia, Panic Disorder, or 
Post-Traumatic Stress Disorder [PTSD]) who had high levels of anhedonia (Snaith-Hamilton 
Pleasure Scale [SHAPS] >20) were treated with a KOR antagonist (KORA) or placebo for 
8 weeks. In that study, the group receiving the KORA treatment showed greater ventral 
striatal activation (a neural target related to hedonic response) during anticipation of both 
gain and loss during an incentivized test of working memory as well as a significant 
improvement in their anhedonia score as evaluated by the SHAPS compared to those patients 
who received placebo (Krystal et al 2020). 
Nonclinical studies indicate that the risk for cardiovascular, respiratory, and central nervous 
system (CNS) AEs in the BTRX-335140 clinical program is low. Body weight loss or 
decreased body weight gain associated with decreased food intake was observed across 
species tested. BTRX-335140 was nonmutagenic in in vitro Ames and chromosomal 
mutation assays. 
 
 
 Ocular phototoxicity (corneal edema), with confirming 
histopathologic findings was observed at the 100 mg/kg/day dose. 
 
 
 
No phototoxicity was noted in the 
retina or any other ocular structure at any dose level. 
BTRX-335140 is orally bioavailable across animal species studied. BTRX-335140 is a 
 
 
Based on nonclinical studies, BTRX-335140 is not anticipated to significantly inhibit or 
induce
at concentrations relevant in vivo. 
 
 
The first-in-human Phase 1 study in healthy adult subjects evaluated single and multiple 
ascending doses of BTRX335140. Treatment with BTRX-335140 exhibited a favorable 
pharmacokinetic (PK) profile, and BTRX-335140 was well tolerated. There were no deaths, 
serious adverse events (SAEs), or subject discontinuations due to AEs in this study. Overall, 
AEs were infrequent, and the majority of events were Grade 1 in intensity. In the 
multiple-dose phase, 13 (54%) subjects experienced a total of 42 treatment-emergent adverse 
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events (TEAEs) following BTRX-335140 administration, and all reported TEAEs in the 
multiple-dose phase were Grade 1 in severity. The most common events reported were 
pruritus and fatigue. Grade 1 pruritus was reported 4 times among 3 (13%) subjects overall, 
with 1 subject receiving 20 mg BTRX-335140 and 2 subjects receiving 160 mg 
BTRX-335140 daily for 10 days. A total of 3 (13%) subjects each experienced a single 
TEAE of Grade 1 fatigue, including 2 subjects who received 80 mg BTRX-335140 and 
1 subject who received 160 mg BTRX-335140 daily for 10 days. All remaining AEs were 
reported by 2 or fewer subjects each in both the single- and multiple-dose phases. Following 
multiple-dose administration of BTRX-335140, the following AEs were considered at least 
possibly related to the study treatment: abnormal dreams (40 mg and 80 mg), fatigue (80 mg 
and 160 mg), headache (80 mg and 160 mg), sleep disorder (40 mg), asthenia, chills, 
dizziness, nausea, somnolence (80 mg), generalized pruritus, macular rash, tremor (160 mg), 
and papule (placebo). Overall, there were no remarkable findings in the remaining safety 
assessments in the study for clinical laboratory parameters, vital sign measurements, 
electrocardiogram (ECG) findings, and Columbia Suicide Severity Rating Scale (C-SSRS) 
scores. Overall, safety findings were comparable for BTRX-335140 and placebo and there 
were no trends observed in safety data with respect to ascending single and multiple 
BTRX-335140 doses. 
 
After 
10 days of once-daily oral dosing, some accumulation (up to 3.9-fold for AUC and up to 
4.6-fold for Cmax exposures) of BTRX-335140 in plasma is observed and steady state is 
reached by Day 10. The administration of a high-fat diet with BTRX-335140 resulted in an 
increase in exposure by 17% (AUC) and 12% (Cmax) compared to fasted conditions. 
The second Phase 1 study used positron emission tomography (PET) imaging study in 
healthy adult males to assess KOR Occupancy in the human brain using the radiotracer 
[11C]LY2795050 following single ascending oral doses of BTRX-335140 showed tracer 
uptake was reduced by treatment with BTRX-335140, and the relationship between KOR 
Occupancy and BTRX-335140 concentration was IC50 = 9.85 ±1.40 ng/mL. 
1.2 
Rationale for Study Design, Doses, and Control Groups 
The current study is the first Phase 2 study designed to establish POC for BTRX-335140 by 
evaluating the impact of treatment with BTRX-335140 relative to treatment with matching 
placebo on symptoms of MDD in adult patients. 
The randomization, study drug blinding, and placebo controls will serve to minimize the 
chance of bias in the implementation and execution of the study. Randomization to treatment 
assignment reduces the likelihood that results will reflect selection bias. Having both 
investigators and subjects blinded to treatment assignment (double-blind) controls for 
expectation bias relative to any particular treatment condition. A placebo group is included to 
control the subject and investigator expectations of improvement during the study that could 
confound interpretation of pharmacological effects of treatment.  
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Double-blind, placebo-controlled studies are the standard in assessment of efficacy in 
randomized clinical studies evaluating subjects with conditions such as MDD. There have 
been estimates of placebo response rates in MDD clinical studies between 30% and 50% 
therefore it is important to use a placebo comparator despite the risk of a possible worsening 
of symptoms. Certain factors are associated with improvement in placebo-treated subjects. 
These include therapeutic effects secondary to interaction with a medical professional, 
misdiagnosis of the psychiatric disorder, clinicians overestimating baseline symptom severity 
or symptom change, and regression to the mean in subjects who began the study with more 
severe symptoms of illness (Fava et al 2003). 
A starting dose of 80 mg of BTRX-335140 was selected for this study on the basis of 
pharmacokinetic/pharmacodynamic modeling from the PK and PET receptor occupancy 
(RO) studies that indicates that this dose should provide approximately 80–90% RO at steady 
state. The study dose of 80 mg BTRX 335140 as well as a higher dose of 160 mg have been 
given once daily (QD) for 10 consecutive days and was well-tolerated in healthy subjects 
(Study K2-ADS-101). Pharmacokinetics of a single 80 mg dose were similar under fed and 
fasted conditions. The brain KOR occupancy of a single 160 mg dose of BTRX-335140 is 
approximately 90% (Study K2-HV-102). Human PK modeling based on drug exposure levels 
in Study K2-ADS-101 indicates that this same level of occupancy will be achieved with an 
80-mg daily dose at steady state. This occupancy level is projected to be sufficient to produce 
a KORA mechanism-based effect in an MDD population. 
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1.3 
Potential Risks and Benefits 
Information about the known and expected benefits, risks, and reasonably anticipated AEs 
with BTRX-335140 treatment may be found in the Investigator’s Brochure (IB). No data are 
available on the long-term effect of BTRX-335140 on fertility; therefore, the possibility of a 
negative impact on fertility cannot be excluded. 
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2 
OBJECTIVES 
2.1 
Primary Objective 
To establish POC for BTRX-335140, a KORA, by evaluating the impact of BTRX-335140 
relative to placebo on symptoms of MDD in adult subjects with MDD and symptoms of 
anhedonia and anxiety following 8 weeks of double-blind treatment as assessed by the 
Hamilton Depression Rating Scale (HAMD-17). 
2.2 
Secondary Objectives 
• To evaluate the effects of BTRX-335140 on self-reported anhedonia in adult subjects 
with MDD 
• To evaluate the effects of BTRX-335140 on anxiety-related symptoms in adult subjects 
with MDD 
• To evaluate the effects of BTRX-335140 on functional impairment in adult subjects with 
MDD 
• To evaluate the safety and tolerability of BTRX-335140 in adult subjects with MDD 
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3
INVESTIGATIONAL PLAN AND ENDPOINTS
3.1
Description of Study Design
Study K2-MDD-201 is an 8-week, randomized, double-blind, placebo-controlled, 
parallel-group, multicenter study to evaluate the effects of BTRX-335140 on symptoms of 
depression in adult subjects with MDD and symptoms of anhedonia and anxiety after 
8 weeks of double-blind treatment. The study will consist of a 7- to 28-day screening period, 
an 8-week active treatment period (during which subjects will receive either BTRX-335140 
or placebo), and a 4-week Safety Follow-up period (Figure 1).
Figure 1:
Study Design
OE = ophthalmologic examinations; TC = telephone contact; Wk = Week.
Note: To reduce subject burden, subjects may be allowed to perform 1 virtual visit at Week 2 (Visit 3), 4
(Visit 4), or 6 (Visit 5). Virtual visits will be performed according to procedures specified in Appendix 1.
Subjects will sign an informed consent form (ICF) and will then enter the screening period to 
ensure they meet the required inclusion and exclusion criteria, including diagnosis and 
stability of depressive symptoms and absence of psychiatric and medical conditions that 
would preclude study participation. The 28-day screening period may be extended up to a 
maximum of 35 days (after consultation with and approval by the medical monitor), if 
necessary, for reasons including, but are not limited to, discontinuation of a subject’s current 
inefficacious medications or to obtain an ophthalmologic examination. Some of the 
enrollment criteria in this study are based on analyses of depression data by BlackThorn
using explainable artificial intelligence to inform about predictors of response to a KORA. 
Sites will be provided with a tablet computer preprogramed with an algorithm to confirm 
subject eligibility on the clinical assessment scales. In addition, for those subjects who are 
deemed to meet eligibility criteria, data will be requested to allow review by the medical 
monitor and the Clinical Assessment Technology (CAT) team to monitor data quality, and 
the ophthalmologic examinations (standard ophthalmologic and corneal specular 
microscopy) will be performed prior to randomization.
Screening
Placebo or 80 mg BTRX-335140 for 8 weeks
Double-blind Treatment
Baseline
Wk 4
Wk 8
Wk 2
Wk 6
Safety Follow-up
Day 1
Wk 10 (TC)
Day -28 to Day-7
Wk 12 (OE)
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In the event that an investigator requests to re-screen a subject that previously screen failed 
Study K2-MDD-201, the medical monitor and Sponsor will consider allowing re-screening 
on a case-by-case basis. 
Subjects who meet the DSM-5 diagnostic criteria for MDD and all other eligibility criteria 
for the study will be randomized in a 1:1 ratio to 1 of 2 treatment arms (placebo or 
BTRX-335140 QD) until approximately 180 subjects are randomized. Beginning at Visit 2 
(Baseline), subjects will receive treatment with BTRX-335140 80 mg QD or placebo for 
8 weeks. During the Safety Follow-up, all subjects will receive a telephone call (TC) from 
study site personnel approximately 2 weeks after discontinuation of study drug for 
assessment of AEs, concomitant medication usage, and C-SSRS; and at Week 12, subjects 
will have corneal specular microscopy performed by a study site-affiliated ophthalmologist. 
The overall duration in the study for each subject is expected to be approximately 14 to 
16 weeks.  
 
The Week 12 corneal specular microscopy should be completed for all subjects (ie, 
completers and early termination) unless consent is withdrawn. The corneal specular 
microscopy should be conducted approximately 12 weeks after the first dose of study drug; 
subjects who terminate the study early would need to plan for the corneal specular 
microscopy at 12 weeks after the first dose of study drug. Any exceptions are to be discussed 
with the medical monitor.  
 
Additional ophthalmologic examinations (standard ophthalmologic exams and/or corneal 
specular microscopies) may be conducted to assess subject safety.  
 
Details of study procedures and assessments are provided in Section 6; for details on the 
timings of study assessments refer to Section 7 and the Schedule of Procedures and 
Assessments. 
Study sites will maintain, at a minimum, coronavirus disease 2019 (COVID-19) risk 
mitigation procedures as per regulatory and local guidelines and/or laws. Allowed 
modifications to the conduct of the study, in the event of site staff and/or patient restrictions 
due to COVID-19, are outlined in Appendix 1. 
3.2 
Study Endpoints 
3.2.1 
Primary 
The primary efficacy endpoint is the change from baseline to Week 8 on the HAMD-17 score 
in the Efficacy population. 
3.2.2 
Secondary 
• Clinical Global Impression of Improvement (CGI-I) score assessed at each postbaseline 
timepoint 
• Change from baseline to each timepoint assessed in SHAPS scores 
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• Change from baseline to each timepoint assessed in Hospital Anxiety and Depression 
Scale (HADS) subscales (ie, anxiety subscale [HADS-A] and depression subscale 
[HADS-D] scores) 
• Change from baseline to each timepoint assessed in Hamilton Anxiety Rating Scale 
(HAM-A) total score 
• Change from baseline to each timepoint assessed in Sheehan Disability Scale (SDS) 
scores 
• Change from baseline to end of study in Clinical Global Impression Scale – Severity 
(CGI-S) scores 
• HAMD-17 response rate 
• Assessment of safety through the occurrence of TEAEs, SAEs, and adverse events of 
special interest (AESIs); change from baseline in vital signs measurements, body 
weights, body mass index (BMI), clinical laboratory test results, C-SSRS scores, Clinical 
Opiate Withdrawal Scale (COWS) scores; physical examination findings, ECG results, 
and ophthalmologic examination findings. 
 
3.3 
Study Duration 
The overall duration of the study is expected to be up to approximately 24 months. 
The expected participation period for a subject is approximately 14 to 16 weeks, including up 
to 28 days screening, approximately 8 weeks of double-blind treatment, and 4 weeks for 
safety follow–up. 
The end of the study is defined as the date when the last scheduled study procedure, as 
outlined in the Schedule of Procedures and Assessments, is performed for the last subject. 
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4 
STUDY ENROLLMENT AND WITHDRAWAL 
4.1 
Study Population 
Approximately 180 subjects will be enrolled in the study at approximately 40 study centers in 
the US. 
4.1.1 
Inclusion Criteria 
Subjects must fulfill all of the following inclusion criteria to be eligible for participation in 
the study: 
1. Are adult men or women 18 to 65 years of age (inclusive) at informed consent 
2. Have a primary DSM-5 diagnosis of MDD, with prominent symptoms of anhedonia 
confirmed by Structured Clinical Interview for DSM-5 Disorders, Clinical Trials Version 
(SCID-5-CT) 
a. The current episode must have started at least 3 weeks prior to screening but no more 
than 12 months before the screening visit. 
b. Have not failed 2 or more courses of antidepressant treatment in the current episode  
c. No more than a 3-point change in HAMD-17 between screening and baseline 
d. Have sufficient history or an independent report to confirm that symptoms are 
causing functional impairment or clinically significant distress  
3. Meet the blinded-rule list based on clinical scale criteria 
4. BMI of 18 to 40 kg/m2 (inclusive) 
5. Medically stable (in the opinion of the investigator and Sponsor/Sponsors delegate) based 
on medical history, vital signs, clinical laboratory tests, and 12-lead ECG performed at 
screening and baseline 
6. Agree to the following birth control:  
a. Nonvasectomized men must agree to use a condom with spermicide, if sexually 
active during the study, until 90 days after the last dose of study drug administration. 
No restrictions are required for a vasectomized man, provided his vasectomy was 
performed 4 months or more prior to the first dose of study drug. A man who has 
been vasectomized less than 4 months prior to the first dose of study drug must 
follow the same restrictions as a nonvasectomized man. Additionally, men must 
refrain from sperm donation during study treatment and for at least 90 days following 
the last dose of study drug. 
b. Women of child-bearing potential (women not surgically sterilized and between 
menarche and 2 years postmenopausal) must have a negative serum pregnancy test at 
screening and a negative urine pregnancy test at enrollment and agree to use reliable 
birth control (eg, oral contraceptives or Norplant®; a reliable double barrier method of 
birth control [diaphragms with contraceptive jelly; cervical caps with contraceptive 
jelly; condoms with contraceptive foam]; intrauterine devices; partner with 
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vasectomy; or abstinence) during the study and for 10 days following the last dose of 
the study drug (BTRX-335140 or placebo). Women will be considered surgically 
sterile, if they have had tubal ligation, bilateral salpingo oophorectomy, or a 
hysterectomy. 
 
Note: Abstinence will be allowed if, in the investigator’s judgement, it is determined 
that the subject is reliable, that abstinence is the preferred and usual lifestyle of the 
subject, and that abstinence will be continued for the duration of the study including 
the 10 days (women) or 90-day period (men) following the last dose of study drug as 
noted above. 
c. Or engaged exclusively in a non-heterosexual relationship 
7. Willing and able to give written informed consent to participate 
8. Able to understand and comply with instructions in English 
9. Are judged by the investigator to be reliable and agree to keep all appointments for clinic 
visits, tests, and procedures, including venipuncture, and examinations required by the 
protocol 
4.1.2 
Exclusion Criteria 
Subjects will be excluded from participating in this study if they meet any of the following 
criteria: 
1. Have a history of any of the following DSM-5 disorders within the specified timeframe: 
a. Currently or in the past year: diagnosis of personality disorder, attention deficit 
disorder/attention deficit hyperactivity disorder, anorexia nervosa, or bulimia nervosa. 
Subjects with comorbid GAD, social anxiety disorder, or panic disorder for whom 
MDD is considered the primary diagnosis are not excluded 
b. Lifetime diagnosis of bipolar 1 or 2, schizophrenia, obsessive compulsive disorder, or 
post-traumatic stress disorder 
2. Have a history of substance or alcohol use disorder (AUD), per DSM-5 criteria, within 
the past year 
3. Are actively suicidal (eg, any suicide attempts within the past 12 months) or are at serious 
suicidal risk as indicated by any current suicidal intent, including a plan, at screening or 
baseline as assessed by the C-SSRS (score of “YES” on suicidal ideations item 4 or 5 
within 3 months prior to Visit 1 [Screening]) and/or based on clinical evaluation by the 
investigator; or are homicidal, in the opinion of the investigator 
4. Have a history or signs of Cushing’s disease, Addison’s Disease, primary amenorrhea or 
other evidence of significant disorders of the hypothalamus-pituitary-adrenal axis  
5. Have any other clinically significant medical or psychiatric condition or circumstance 
prior to randomization that, in the opinion of the investigator, or Sponsor could affect 
subject safety, preclude evaluation of response, interfere with the ability to comply with 
study procedures, or prohibit completion of the study, such as acute stress disorder, 
adjustment disorder, impulse control disorder, uncontrolled diabetes mellitus, renal or 
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hepatic impairment, coronary artery disease, evidence of significant active cardiac, 
respiratory, or hematologic disease, cancer with <5-year remission (basal cell carcinoma 
is not excluded), chronic pain, fibromyalgia, gastric bypass, lap band placement, or any 
other significant gastrointestinal condition 
6. Have had prior seizures (other than remote history of childhood febrile seizure) or other 
condition that would place the subject at increased risk of seizures or is taking 
anticonvulsants for seizure control 
7. Have a history of serious head injury (eg, skull fracture, cerebral contusion, or trauma 
resulting in prolonged unconsciousness), intracranial neoplasm, or hemorrhage 
8. Have ever had electroconvulsive treatment, vagal nerve stimulation, or treatment with 
ketamine or esketamine for MDD 
9. Have initiated transcranial magnetic stimulation, psychotherapy (such as Cognitive 
Behavioral Therapy) or have had a change in psychotherapy, or other non-drug therapies 
(such as acupuncture or hypnosis) within 4 weeks prior to Visit 1 (Screening) or at any 
time during the acute phase of the study 
10. Have a visual or physical motor impairment that could interfere with the subject’s ability 
to perform study assessments, as assessed by the investigator 
11. Have alanine aminotransferase (ALT) or aspartate aminotransferase (AST) levels 
≥2 x upper limit of normal (ULN) or a bilirubin level 1.5 x ULN unless due to a 
documented history of Gilbert’s syndrome 
12. Estimated glomerular filtration rate (eGFR) ≤60 mL/min/1.73m2 as calculated by the 
Chronic Kidney Disease Epidemiology Collaboration [CKD-EPI] 2009 creatinine 
equation (Levey et al 2009) at Visit 1 (Screening) 
13. Positive hepatitis C virus (HCV) antibody (Ab), hepatitis B surface antigen (HBs Ag), 
hepatitis A virus (HAV) IgM antibody (HAV-Ab [IgM]) or human immunodeficiency 
virus (HIV) test at Visit 1 (Screening) 
14. Have a thyroid-stimulating hormone (TSH) level of <0.9 x lower limit of normal (LLN) 
or >1.2 x ULN on or off stable treatment for hyperthyroidism or hypothyroidism; if TSH 
is abnormal, evaluate reflex Free T3 and Free T4. If reflex testing is normal, the 
assessment of normal thyroid function will be determined based on the judgement of the 
investigator, following discussion with the medical monitor. 
15. Have any other clinically significant abnormalities (significant would include laboratory 
deviations requiring acute medical intervention or further medical evaluation) in 
laboratory results at screening, including clinical chemistries, hematology, and urinalysis, 
and any clinical information that, in the judgment of the investigator or Sponsor, should 
preclude a subject’s participation at study entry 
16. Exclusionary ECG abnormalities obtained at Visit 1 (Screening) or Visit 2 (Baseline) are 
QT interval corrected using Fridericia’s formula (QTcF) >450 msec in males or 
>470 msec in females, complete bundle branch block, evidence of myocardial infarction 
or ischemia, and predominantly nonsinus conducted rhythms. Other abnormalities can be 
exclusionary at the discretion of the principal investigator or medical monitor. See 
Section 6.3.5 for guidance on ECG interpretations. 
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17. Have a positive urine drug screen for amphetamines, barbiturates, cocaine, methadone, 
opioids, propoxyphene, tetrahydrocannabinol (THC), phencyclidine, or positive blood 
alcohol level assessed by breathalyzer at Visit 1 (Screening) and Visit 2 (Baseline). For 
occasional (1 to 2 times per month maximum) cannabis users only, 1 retest is allowed and 
subject must agree to abstain from use for the duration of the study; a positive second test 
is exclusionary. 
18. Have any use, by history, of Salvinorin A 
19. Use of the following concomitant medications (contact the Sponsor-designated medical 
monitor to determine eligibility when in doubt):  
a. Psychoactive medication including stimulants, benzodiazepines and anxiolytics, oral 
antipsychotics, mood stabilizers/anticonvulsants (carbamazepine, lamotrigine, etc.), 
lithium, antidepressants, S-adenosylmethionine, melatonin, agomelatine, and 
hypnotics/sedatives within 5 half-lives or 14 days (whichever is longer) of Visit 2 
(Baseline) 
b. Fluoxetine and irreversible monoamine oxidase inhibitors within 4 weeks of Visit 2 
(Baseline); depot antipsychotics within 2 months of Visit 2 (Baseline) 
c. Opioid agonists and antagonists 
20. Are currently taking or have taken within 5 half-lives of Visit 2 (Baseline) any 
medications or supplements that are
 
 
 
21. Are women who are either pregnant or breastfeeding 
22. Have participated (received study treatment) in a clinical study or any other type of 
medical research judged by the investigator or Sponsor to be scientifically or medically 
incompatible with this study within 30 days prior to Visit 1 (Screening). Contact the 
Sponsor-designated medical monitor to determine eligibility when in doubt.  
23. Have participated in multiple interventional clinical studies, such that, in the opinion of 
the investigator, the subject is not a suitable candidate for participation 
24. Have previously completed or withdrawn from this study or any other study investigating 
BTRX-335140 
25. Are investigator site personnel directly affiliated with this study, and/or their immediate 
families. Immediate family is defined as a spouse, parent, child, or sibling, whether 
biological or legally adopted. 
26. Are employees of the Sponsor or are employees of any third-party organizations (TPOs) 
(eg, laboratory staff, study vendors and transportation providers) involved in study who 
require exclusion of their employees 
27. Has any of the following: 1) useful vision in only 1 eye from a pre-existing ophthalmic 
disease or amblyopia; 2) a corneal transplant in either eye; 3) corneal dystrophy or family 
history of corneal dystrophy; 4) severe dry eye syndrome [keratitis sicca]; 5) will not or 
cannot cooperate with ophthalmic examination requiring pupillary dilation (includes 
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history of severe adverse reaction to mydriatic agents or untreated narrow angle 
glaucoma). Note: The following ocular disorders are allowed: cataracts, prior cataract 
surgery, glaucoma (narrow angle glaucoma is allowed if definitively treated with laser 
peripheral iridectomy), macular degeneration, or ocular changes associated with diabetes 
mellitus or multiple sclerosis. 
4.2 
Method of Assigning Subjects to Treatment 
Subjects who meet all inclusion criteria and are subsequently not excluded by the exclusion 
criteria will be randomized to double-blind treatment at Visit 2 (Baseline) using an 
interactive web-response system (IWRS). Subjects will be randomized in a 1:1 ratio to 1 of 
2 treatment arms (placebo or BTRX-335140 QD). 
The IWRS will be used to assign bottles containing double-blind study drug or placebo to 
each subject. Further details will be provided in the Pharmacy manual. 
4.3 
Blinding 
This is a double-blind study. Subjects, site personnel, and the Sponsor will be blinded to 
treatment. In the event there is a safety emergency, the investigator is requested, where 
possible, to contact the medical monitor or the Sponsor before unblinding; however, if there 
is insufficient time to do so, the investigator may perform emergency unblinding for SAEs 
through the IWRS as described in Section 8.1.2. 
4.4 
Subject Withdrawal 
A subject’s participation in this study is voluntary, and the subject may refuse to participate 
or withdraw from the study at any time for any reason. The investigator also has the right to 
withdraw a subject from the study for any reason. The investigator should make a reasonable 
effort to ascertain the primary reason for withdrawal and document that reason in the 
electronic case report form (eCRF). 
Possible reasons for early discontinuation are as follows: 
• Investigator/physician decision 
- 
The investigator/physician decides that the subject should be withdrawn from the 
study for any reason, eg, the subject is significantly noncompliant with the study drug 
regimen (see Section 5.5; subjects with study drug compliance ≤70% over any 
2-week visit interval). 
- 
After Visit 2 (Baseline), the subject is found to have met an exclusionary criterion or 
not met an entry criterion required for study participation. 
- 
The subject is enrolled in any other clinical study involving an investigational product 
or is enrolled in any other type of medical research judged not to be scientifically or 
medically compatible with this study. 
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• Subject Decision 
- 
The subject requests to be withdrawn from the study for any reason. If the reason to 
request withdrawal is an AE this should be documented as the primary reason for 
discontinuation. 
• Sponsor Decision  
- 
An investigator, site personnel performing assessments, or subject is unblinded (see 
Section 4.3). 
• Adverse Event  
- 
The subject experiences an AE that would necessitate discontinuation of study drug. 
Any subject who is discontinued from study drug due to AEs or clinically significant 
laboratory abnormalities will be followed until resolution or clinical stabilization. If 
this decision is made because of an SAE or a clinically significant laboratory value, 
the study drug is to be discontinued and appropriate measures are to be taken. The 
Sponsor or its medical monitor designee is to be alerted immediately (see 
Section 8.5.1). 
- 
Discontinuation should be considered by the investigator after consultation with the 
Sponsor or its medical monitor designee when a subject has the following clinically 
significant abnormal laboratory findings: 
◦ 
ALT or AST >8 x ULN 
◦ 
ALT or AST >5 x ULN for more than 2 consecutive weeks 
◦ 
ALT or AST >3 x ULN and total bilirubin level >2 x ULN 
◦ 
ALT or AST >3 x ULN and prothrombin time (or INR) >1.5 x ULN 
◦ 
ALT or AST >3 x ULN with the appearance of fatigue, nausea, vomiting, right 
upper-quadrant pain or tenderness, fever, rash, and/or eosinophilia (>5 %). 
- 
If the subject is thought to be at risk of harm to him/herself or others at any time 
during the study, a further psychiatric assessment by the investigator must be 
conducted. This includes subjects who score 3 or above on item 11 of the HAMD-17. 
Subjects must be discontinued from study participation and offered appropriate 
treatment and care if they:  
◦ 
are assessed as homicidal 
◦ 
are actively suicidal (any suicidal ideation with intent or specific plan or any 
suicide attempt) or at serious suicidal risk 
◦ 
if suicidal gestures are present  
- 
Any subject who is found to be dependent on, or abusing opioids, should be 
withdrawn from the study and provided with appropriate medical care. If at any time 
during the study a score of 5 or more is obtained on the COWS, the investigator (or 
designee) must determine if the subject is experiencing opioid withdrawal (see 
Section 6.3.8). 
- 
The investigator remains responsible for following, through an appropriate healthcare 
option, AEs that caused the subject to discontinue study drug. 
• Unsatisfactory Therapeutic Effect 
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- 
If the investigator determines that a subject requires treatment with another 
therapeutic agent for MDD and that this alternative treatment cannot be safely 
delayed until completion of the current study, discontinuation from the study will 
occur prior to introduction of the new agent. 
• Loss to Follow-Up 
- 
For subjects who are lost to follow-up, all steps performed to contact them (dates of 
telephone calls, registered letters, etc.) will be documented in the source documents. 
The investigators are required to make at least 3 attempts to contact the subject for a 
follow-up safety visit.  
• Pregnancy 
- 
Study drug treatment will be discontinued for female subjects who become pregnant 
during study. See Section 8.5.4 for additional information. 
• Other Reasons 
- 
If a subject misses, or is expected to miss, more than 1 sequential clinic visit, the 
investigator must contact the medical monitor to discuss continuation in the study. If 
it is determined that the subject must be discontinued the reason for termination, 
eg, termination related to COVID-19, must be documented as such in EDC (see 
Appendix 1). 
4.5 
Early Termination of Subjects From the Study 
For subjects who discontinue study drug, every attempt should be made to complete the 
End-of-Treatment Visit assessments within 2 days after their last dose of study drug 
(Section 7.8). The 12-week corneal specular microscopy examination should be completed 
for subjects who discontinue participation in the study unless consent for this safety 
procedure is withdrawn. The corneal specular microscopy should be conducted 
approximately 12 weeks after the first dose of study drug; exceptions are to be discussed with 
the medical monitor.  
 
Additional ophthalmologic examinations (standard ophthalmologic exams and/or corneal 
specular microscopies) may be conducted to monitor subject safety. 
 
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5 
STUDY TREATMENTS 
5.1 
Description of Treatments 
The drug product BTRX-335140 is supplied for clinical study use as capsules and  
 
 
 
 
  
The placebo capsule product consists of a capsule size and color that matches the active drug 
product contains 
 exclusively. 
5.2 
Manufacturing, Packaging, and Labeling 
BTRX-335140 and placebo capsules and packaging will be identical to maintain investigator 
and subject blinding. Clinical study materials will be labeled according to the regulatory 
requirements. Each bottle will contain 16 capsules of study drug with a unique 5-digit 
identification number. 
5.3 
Storage 
 consistent with 
the product label. The drug product is stable when stored according to the labeling 
instructions. 
5.4 
Study Drug Administration 
This study involves a comparison of BTRX-335140 versus placebo taken orally QD for 
8 weeks. Treatment will be initiated at 80 mg (1 capsule) QD and this dose will be 
maintained for all 8 weeks of double-blind treatment. Subjects should be instructed to 
swallow capsules whole and not crush or chew and to store the capsules in their original 
packaging at the temperature listed on the package label. Subjects will be instructed to take 
their first dose of study drug (BTRX-335140 or placebo) under the supervision of clinical 
study personnel during the baseline visit and to try to take their study medication at 
approximately the same time each day, preferably in the morning with food. 
Subjects will also be instructed to refrain from taking their investigational study drug at home 
on the morning of Visit 4 (to allow PK sample to be withdrawn predose). Study drug will be 
administered at the site from the bottle dispensed at Visit 4. 
 
 
Subjects who cannot tolerate 1 capsule daily will be discontinued from participation in the 
study. 
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5.5 
Study Drug Compliance 
Each subject will be instructed to return all study drug packaging and unused material to the 
study site at each visit. The study site will keep a record of all drug dispensed to and returned 
by the subjects throughout the study.  
Subject compliance with BTRX-335140 or placebo will be assessed by direct questioning 
and counting of returned capsules. 
For subjects who demonstrate noncompliance (subject has taken <80% of the prescribed 
dosage for a visit interval), investigative sites must counsel subjects on the importance of 
study drug compliance. Any missed study drug or extra dose of study drug will be recorded 
as a protocol deviation.  
Subjects with study drug (BTRX-335140 or placebo) compliance ≤70% over any 2-week 
visit interval (beginning with the Visit 2 [Baseline] to Visit 3 [Week 2] interval) will require 
early discontinuation. 
5.6 
Study Drug Accountability 
The study drug provided for this study will be used only as directed in the study protocol. In 
accordance with current Good Clinical Practice (cGCP), investigators are required to 
maintain accurate and up-to-date records of all study drugs to permit reconciliation. The 
investigator or designee must maintain adequate records of distribution, including the date 
received, number and units received, manufacturer, lot numbers, dispensing, and return or 
destruction of all study drug (ie, accountability or dispensing logs). 
All study drug records must be up to date and accurate for inspection by the site’s clinical 
monitor and/or auditor. Instructions for returns, disposal, or destruction will be provided in 
the study reference manual and/or pharmacy manual. 
If there are any issues during the course of the study related to the quality of the study drug, 
the investigator, clinical site pharmacist or pharmacy designee should contact the Sponsor or 
designated Contract Research Organization (CRO). 
5.7 
Prior and Concomitant Therapy 
All nonpsychiatric medications, including over-the-counter medications and supplements, 
and also significant nonpharmacological therapies, taken by the subjects from the time of 
screening (ie, signing of informed consent) through the end of the Safety Follow-up period 
(ie, last contact) will be recorded in the subject’s eCRF. 
Previous and current MDD psychiatric therapy history (pharmacologic and 
nonpharmacologic) during the current episode will be recorded separately. Concomitant 
therapies will be reviewed for disallowed medications. 
BTRX-335140 is a 
 
. This 
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restriction includes 
 
(Appendix 2). However, nonsystemic (eg, topical creams, eye drops, 
mouthwashes) applications are permissible. 
In vitro, BTRX-335140 is an
 
 
In general, concomitant medications with primary CNS activity are not allowed in the study. 
The investigator should instruct the subject to notify the study site about any new 
medications he/she takes and about any significant nonpharmacological therapies 
administered after the start of the study drug (eg, acupuncture, hypnosis etc.). 
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6 
STUDY PROCEDURES AND ASSESSMENTS 
Study procedures and their timing are summarized in the Schedule of Procedures and 
Assessments. Investigator/clinician-rated scales and tasks will be completed and/or 
administered by qualified and trained site raters who meet the training requirements and 
qualifications of the scale or task standards set by the Sponsor and training vendors. 
Subject-rated assessments will be recorded using a tablet computer. The SCID and COWS 
are not included on the tablet; they are available only in paper form.  
Rater consistency is critical in assessing the HAM-D, HAM-A, CGI-S, and CGI-I. Therefore, 
every attempt should be made, where possible, to have the same rater assess the subject at 
each visit to provide consistency in these ratings. It is important that the site strives to 
maintain rater consistency for the primary endpoint, the HAM-D. The CGI-S and CGI-I rater 
should also be the same rater and should take into account all available information in the 
assessment, particularly when the CGI rater is not rating the HAM-D and/or HAM-A. 
6.1 
Medical History and Demographics 
Medical history and demographics will be collected at Screening and will be reviewed at the 
baseline visit. 
6.1.1 
Medical History 
A complete medical history will be obtained at screening and will be reviewed prior to 
randomization to ensure subjects qualify for the study. Confirmation of the subject’s MDD 
will also be obtained. 
Psychiatric history (including a lifetime history of substance or alcohol abuse or dependence) 
will be assessed using the SCID-5-CT. The SCID-5-CT will be completed by qualified and 
trained investigator site raters. The interview begins with an overview section, which is 
abridged to entail questions relevant to the current clinical study. 
Ophthalmologic history will be obtained at screening to assess general ocular disorders and 
specifically whether the subject has any of the following exclusionary conditions: 1) useful 
vision in only 1 eye from a pre-existing ophthalmic disease or amblyopia; 2) a corneal 
transplant in either eye; 3) corneal dystrophy or family history of corneal dystrophy; 4) 
severe dry eye syndrome [keratitis sicca]; or 5) history of severe adverse reaction to 
mydriatic agents. If a subject has narrow angle glaucoma, then there must be confirmation of 
definitive treatment with laser iridectomy as verified by the subject’s ophthalmologist in 
order to be enrolled. 
Nonserious AEs reported before the first dose of study drug will be recorded as medical 
history. 
6.1.2 
Demographics and Baseline Characteristics 
Demographic information and baseline characteristics collected at Screening will include 
age, sex, race, ethnicity, height, weight, and BMI. 
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6.2 
Efficacy Assessments 
6.2.1 
Hamilton Rating Scale for Depression – 17-item Version 
The HAM-D 17-item version (Hamilton 1967) will be the primary endpoint to evaluate 
changes in depression severity with treatment. This interviewer-administered semi-structured 
interview (Structured Interview Guide for the Hamilton Depression Rating Scale [SIGH-D], 
Williams 1988) is one of the most widely used instruments in depression treatment studies 
and its reliability and validity has been extensively studied. The HAMD-17 scoring ranges 
from 0 to 52 with individual items ranging from 0-4 points and 0-2 points to capture 
symptom severity. The HAMD-17 is validated for videoconference and telephone assessment 
(Kobak et al 2008). 
6.2.2 
Snaith Hamilton Pleasure Scale 
The SHAPS is a 14-item self-report instrument which measures anhedonia (Snaith et al 
1995). It has been shown to be valid and reliable in normal and clinical samples, with 
adequate construct validity, satisfactory test-retest reliability (intraclass correlation 
coefficient [ICC] = 0.70), and high internal consistency (Cronbach’s alpha of 0.94) (Franken 
et al 2007). The scale will be completed by the subject and reviewed by site personnel 
qualified to oversee completeness. Each of the 14 items has a set of 4 responses, 2 of which 
endorse agreement (Definitely Agree, Agree) and 2 of which endorse disagreement 
(Disagree, Strongly Disagree). A total score can be derived by summing the responses items 
answered with strongly agree are coded as 1 while a strongly disagree response will be coded 
as 4. Therefore, scores on the SHAPS can range from 14 to 56, with higher scores 
corresponding to higher levels of anhedonia. 
6.2.3 
Clinical Global Impression Scale – Severity 
The CGI-S is a clinician-rated instrument that measures the severity of depression at the time 
of assessment. This rating is based upon observed and reported symptoms, behavior, and 
function in the past 7 days. The score should reflect the average severity level across the 7 
days. The CGI-S is scored on a 7-point scale where a score of 1 indicates that the subject is 
“normal, not at all ill” a score of 4 indicates that the subject is “moderately ill,” and a score 
of 7 indicates that the subject is “among the most extremely ill subjects” (Guy 1976). 
6.2.4 
Clinical Global Impression Scale – Improvement 
The CGI-I scale (Guy 1976) is a clinician-rated instrument that measures the improvement of 
the subject’s symptoms. It is a 7-point scale where a score of 1 indicates that the subject is 
“very much improved,” a score of 4 indicates that the subject has experienced “no change,” 
and a score of 7 indicates that the subject is “very much worse.” 
6.2.5 
Hamilton Anxiety Rating Scale 
The HAM-A is administered to assess severity of anxiety, its improvement during the course 
of treatment, and the timing of such improvement (Hamilton 1960). This instrument will be 
completed by qualified and trained investigator site raters based on a semi-structured 
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interview (Structured Interview Guide for the Hamilton Anxiety Rating Scale [SIGH-A], 
Williams 1988) his/her assessment of the subject. The scale consists of 14 items. Each item is 
rated on a scale of 0 (feeling not present) to 4 (very severe prevalence of the feeling). The 
HAM-A total score is the sum of the 14 items and the score ranges from 0 to 56. 
6.2.7 
Hospital Anxiety and Depression Scale 
The HADS measures levels of anxiety and depression without regards to somatic symptoms 
(Zigmond and Snaith 1983). This self-report scale consists of 14 items. Seven of the items 
are used to evaluate anxiety and 7 evaluate depression. Each item on the questionnaire is 
scored from 0 to 3. Therefore, the anxiety subscale (HADS-A) and the depression subscale 
(HADS-D) each range from 0 to 21. Higher scores indicate higher levels of anxiety and 
depression, respectively. 
6.2.9 
Sheehan Disability Scale 
The Sheehan Disability Scale (SDS) was developed to assess functional impairment in 
3 related domains of work/school, social, and family life. It is a brief self-rated tool where the 
subject rates the 3 items on an anchored 10-point visual analogue scale (Sheehan 1983). 
Following the subject’s self-assessment, the rater should review the Work/School and Days 
Lost items to confirm appropriateness of the selected rating by evaluating the impact of 
MDD symptoms across those sectors. SDS may be administered via telephone assessment; 
however, the subject must be provided with a portable document format (PDF) copy of the 
assessment for reference of the visual analogue scale when selecting their ratings. 
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6.3 
Safety Assessments 
6.3.1 
Adverse Events 
All AEs will be evaluated from the time the subject provides written informed consent 
through the last Safety Follow-up TC. Nonserious AEs reported before the first dose of study 
drug will be recorded as medical history (see Section 6.1.1). Serious AEs will be recorded on 
the AE eCRF from the time of informed consent. Additional safety information, including 
the definition of an AE/SAE/AESI and reporting requirements is provided in Section 8. 
Clinically significant findings for laboratory results, vital signs, ECGs, ophthalmologic 
examinations, and abnormal physical examination should be recorded as AEs; a clinical 
diagnosis, rather than the changes in laboratory analyte or other assessment should be 
recorded. 
6.3.2 
Clinical Laboratory Tests 
6.3.2.1 
Sample Collections 
A central laboratory will be used for safety laboratory tests. Standard clinical laboratory tests 
will be performed at times specified in the Schedule of Procedures and Assessments. Repeat 
testing of samples may be permitted during screening, after review and approval by the 
medical monitor. 
Abnormal laboratory values will be graded by the investigator as “clinically significant” or 
“not clinically significant,” where applicable. Clinically significant abnormal laboratory 
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values will be reported as AEs as per Section 6.3.1 above). Investigators may repeat 
laboratory tests for any parameter that is abnormal and/or clinically significant. 
Samples collected for safety laboratory tests will be destroyed within 60 days of receipt of 
confirmed test results. Tests are run and confirmed promptly whenever scientifically 
appropriate. When scientific circumstances warrant, however, it is acceptable to retain 
samples to batch the tests run, or to retain the samples until the end of the study to confirm 
that the results are valid. Certain samples may be retained for a longer period, if necessary, to 
comply with applicable laws, regulations, or laboratory certification standards. 
Table 2 lists the specific laboratory analyses that will be performed for this study. 
Table 2: 
Clinical Laboratory Tests 
Hematologya:  
Hemoglobin   
Hematocrit  
Erythrocyte count (red blood cell [RBC])  
Mean cell volume (MCV)  
Mean cell hemoglobin concentration (MCHC)  
Leukocytes (white blood cell [WBC])  
Neutrophils, segmented 
Absolute Neutrophil Count (ANC) 
Lymphocytes  
Monocytes 
Eosinophils 
Basophils 
Platelets 
Prothrombin timeb 
Chemistrya: Serum concentrations of: 
Sodium 
Potassium  
Bicarbonate  
Chloride 
Total bilirubin 
Direct bilirubin 
Alkaline phosphatase 
Alanine aminotransferase/serum glutamic pyruvic 
transaminase (ALT/SGPT) 
Aspartate aminotransferase/serum glutamic oxaloacetic 
transaminase (AST/SGOT) 
Gamma-glutamyl transferase (GGT) 
Blood urea nitrogen (BUN) 
Creatinine 
Uric acid 
Phosphorous 
Calcium 
Glucose (random) 
Albumin 
Total cholesterol 
Creatine kinase (CK)c 
Magnesium 
Amylase 
Lipase 
Urinalysisa: 
Specific gravity  
pH  
Protein  
Glucose  
Ketones  
Blood  
Urine leukocyte esterase 
Urobilinogen 
Urine Drug Screen and Breathalyzer Alcohol Screena 
Amphetamines 
Barbiturates 
Benzodiazepines 
Delta-9-tetrahydrocannabinol (THC) 
Cocaine 
Ethyl alcohol (via breathalyzer) 
Opioids 
Phencyclidine 
Propoxyphene 
Methadone 
Viral Serologyd 
Hepatitis B Surface Antigen (HBs Ag) 
Hepatitis B Core Antibody (HBc Ab) 
Hepatitis C Antibody (HC Ab) 
Hepatitis A Antibody (HAV-Ab [IgM]) 
Human immunodeficiency virus (HIV) 
Thyroid functiond: 
Thyroid-stimulating hormone (TSH); if abnormal perform 
Free T3 and Free T4 reflex testing 
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Pregnancy Test (females of child bearing potential 
only)e 
 
 
 
a 
Performed at each visit. 
b 
To be conducted at baseline only, unless indicated for hepatic monitoring testing (see Table 3) 
c 
Creatine kinase is to be fractionated if CK results >1000 IU/L. 
d 
Screening only. 
e 
Serum pregnancy test at Visit 1 (Screening) is to be performed by a Sponsor-designated laboratory; urine pregnancy 
tests will be performed at all subsequent visits by the site. 
f 
Baseline only; optional collection. 
Note: All safety laboratory tests will be analyzed by a Sponsor-designated laboratory, unless otherwise specified. 
 
6.3.2.2 
Hepatic Safety and Enhanced Hepatic Testing 
Bilirubin, ALT, and AST will be measured at each scheduled visit (as part of the scheduled 
clinical laboratory tests). If a subject has elevated ALT or AST (>3 x ULN) or elevated total 
bilirubin (>2 x ULN), the following procedures must be performed: 
1. Contact the medical monitor/Sponsor to assess whether the subject should be 
discontinued from study drug (see Section 4.4). 
2. Perform clinical laboratory testing according to the list of hepatic monitoring tests in 
Table 3 within 48 to 72 hours. The frequency of repeat testing to be performed will be 
determined in consultation with the medical monitor/Sponsor. In general, repeat 
laboratory testing should be performed 2 to 3 times per week; frequency of retesting can 
decrease to once a week or less if, the laboratory abnormality(ies) stabilizes and/or 
subject is asymptomatic. 
3. Provide additional history from the subject including: 
- 
history of concomitant medication (prescription, nonprescription, over-the-counter, 
herbal supplements, dietary supplements) 
- 
alcohol use, recreational drug use, and special diets 
- 
recent illnesses, exposures, and exposures to environmental chemical agents 
These results will be reviewed in a timely fashion by the Sponsor-designated medical 
monitor, along with the subject’s clinical case, to determine possible cause. 
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Table 3: 
Hepatic Monitoring Tests 
Hepatic Hematologya 
Hemoglobin 
Hematocrit   
RBC  
WBC  
Neutrophils, segmented 
Lymphocytes   
Monocytes 
Eosinophils 
Basophils 
Platelets 
Haptoglobin 
Hepatic Coagulationa 
Prothrombin time, INR 
Autoimmune Hepatitis Antibodiesa,b,c  
Anti-smooth muscle antibody 
Anti-nuclear antibody 
Hepatic Chemistrya 
Total bilirubin 
Direct bilirubin 
Alkaline phosphatase 
ALT 
AST 
GGT 
CPK 
Albumin 
Hepatic Serologiesa,b,c 
Hepatitis A antibody Total 
HAV-Ab [IgM] 
HBs Ag  
Hepatitis B surface antibody (anti-HBs) 
HBc Ab  
HCV Ab  
Hepatitis E antibody, IgG 
Hepatitis E antibody, IgM 
Ab = antibody; ALT = alanine aminotransferase; AST = aspartate aminotransferase; CPK = creatine phosphokinase; 
GGT = Gamma-glutamyltransferase; HAV-Ab [IgM] = hepatitis A virus antibody; HBc Ab = hepatitis B core antibody; 
HBs Ag = hepatitis B surface antigen; HCV Ab = hepatitis C virus antibody; Ig = immunoglobulin; INR = international 
normalized ratio; RBC = red blood cells; WBC = white blood cells. 
a 
Assayed by a Sponsor-designated or local laboratory. 
b 
Reflex/confirmation dependent on regulatory requirements and/or testing availability. 
c 
Autoimmune hepatic antibodies and hepatic serologies need only be obtained at the time of the first hepatic monitoring 
test. Additional testing will be directed by the medical monitor/Sponsor. 
 
6.3.2.3 
Urine Drug Screen 
The urine drug screen for drugs of abuse will include amphetamines, barbiturates, 
benzodiazepines, THC, cocaine, opioids, phencyclidine, propoxyphene, and methadone. This 
test will be done at the site using the dip sticks provided as well as at the central laboratory. 
For occasional (1 to 2 times per month maximum) cannabis users only, 1 retest is allowed; a 
positive second test is exclusionary. 
The site must contact the Sponsor-designated medical monitor or designee for preapproval 
before a retest of drugs other than THC that presented as positive; a retest will be permitted if 
the positive results can be explained by a valid prescription, and if there is no evidence of a 
substance use disorder. If the results from an allowed repeat urine drug screen prior to 
randomization are negative, the subject may be randomized. The investigator must be 
satisfied that the subject has been appropriately counseled regarding the restrictions of study 
participation and that the subject intends to comply. If the repeat results are positive, the 
subject must be excluded from the study. A second retest is not admissible. 
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If a subject tests positive following randomization, the site must contact the 
Sponsor-designated medical monitor or designee, to discuss the subject’s continued 
participation in the study. In the event that the subject is allowed to continue in the study, the 
investigator will document the discussion with the medical monitor and counsel the subject 
regarding study restrictions. 
Blood alcohol level will be assessed by breathalyzer testing. 
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6.3.3 
Physical Examinations 
A full physical examination will be performed at Screening and will include an evaluation of 
cardiovascular, respiratory, gastrointestinal, neurological (examination of cranial nerves, 
motor system, sensory, and reflexes), dermatological, and musculoskeletal systems and 
include general appearance, skin, head, neck, eyes, ears, nose, throat, lungs, heart, abdomen, 
back, lymph nodes, extremities, and vascular status. Additional physical examinations will be 
performed as needed. 
Height and weight will be measured during the study as indicated in the Schedule of 
Procedures and Assessments. Height should be measured only once at screening with no 
shoes for correct measurement. The BMI will be calculated based on height measured at 
screening and weight measured at Visits 1, 2, 4, and 6. 
6.3.4 
Vital Signs 
Vital signs (respiratory rate, blood pressure [BP], and pulse) will be measured at each visit 
throughout the screening and treatment periods of the study in supine and standing positions. 
Subjects should be supine for 10 minutes before vital signs (respiratory rate first, then BP and 
pulse) are measured (respiratory rate and pulse should be assessed for 30 seconds, then 
doubled to record breaths/minute and beats/minute, respectively). Subjects will then be asked 
to stand for 3 minutes prior to measurement of standing respiratory rate, BP, and pulse. Oral 
temperature can be measured in any position at each visit throughout the study. 
6.3.5 
Electrocardiography 
For each subject, single 12-lead digital ECGs will be collected after the subject has rested for 
5 minutes. The ECGs will initially be interpreted by a qualified physician (the investigator or 
qualified designee) at the site as soon after the time of ECG collection as possible, and 
ideally while the subject is still present, to determine whether the subject meets entry criteria 
at the relevant visit(s) and for immediate subject management, should any clinically relevant 
findings be identified.  
After enrollment, clinically significant increases in the QTcF interval from baseline (eg, 
QTcF ≥500 msec and/or QTcF increased ≥60 msec) or other clinically significant 
quantitative or qualitative changes from baseline should be reported as AEs. The subject will 
be assessed by the investigator for symptoms (eg, palpitations, near syncope, syncope) and to 
determine whether the subject can continue study drug. The investigator or qualified 
designee is responsible for determining if any change in subject management is needed and 
must document his/her review of the ECG printed at the time of evaluation for each time 
point.  
All digital ECGs will be electronically transmitted to a designated central ECG laboratory. A 
cardiologist at the central ECG laboratory will then conduct a full over-read. A report based 
on data from this over-read will be issued to the investigative site. All data from the 
over-reads will be placed in the Sponsor database for analytical and study report purposes. 
Any clinically significant finding that was present on the fully over-read ECG will be 
reported to the investigator and to the Sponsor. 
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BlackThorn Therapeutics, Inc. 
Clinical Study Protocol Amendment 7 
Date: 24Jan2022 
 
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The cardiologist’s ECG interpretation will be used for inclusion/exclusion determination for 
the screening ECG; the baseline ECG will be based on the investigator review and 
interpretation of the machine read for inclusion/exclusion determination at Visit 2 (Baseline). 
Exclusionary QT interval criteria, corrected using Fridericia’s formula (QTcF), at Visit 1 
(Screening) or Visit 2 (Baseline), are QTcF >450 msec in males or >470 msec in females. In 
the event that the QTcF is greater than the specified cutoffs and the investigator (in 
consultation with the medical monitor) determines that additional ECGs may aid in the 
further evaluation to determine enrollment at either screening or baseline visits, triplicate 
ECGs may be obtained. The average of the triplicate QTcF values may be used to evaluate 
eligibility at screening or baseline. If the average exceeds the protocol-specified cutoff, then 
the subject is not eligible for enrollment. At baseline, if central review subsequently 
determines the subject is not eligible by the cardiologist’s interpretation, the subject must be 
bought in to discontinue. 
When there are differences in ECG interpretation between the investigator (or qualified 
designee) and the cardiologist at the central ECG laboratory, the investigator (or qualified 
designee) should consult with the cardiologist at the central ECG laboratory. The investigator 
(or qualified designee) will be responsible for the medical management of the subject. 
Interpretations from the cardiologist at the central ECG laboratory will be used for data 
analysis and report writing purposes.  
The investigator (or qualified designee) must document his/her review of ECGs printed at the 
time of collection, the final over-read ECG report issued by the central ECG laboratory, and 
any alert reports. 
6.3.6 
Ophthalmologic Examinations 
Ophthalmologic examinations will be conducted in this study as follows (further details will 
be described in an ophthalmologic examination manual): 
1. Standard ophthalmologic examinations: to include best corrected visual acuity 
assessment, slit-lamp examination of both eyes including assessment of intraocular 
pressure, as well as dilation of the pupils to examine the optic nerve and retina 
2. Corneal specular microscopy: noncontact or contact specular microscopy for qualitative 
and quantitative examination of the central corneal endothelium (cell count, cell shape, 
density, and morphology), with image capture 
After confirmation that the subject has met all eligibility criteria, and is approved for 
randomization, the 2 examinations will be performed and will serve as baseline 
ophthalmologic examinations. The study site will refer the subject to an affiliated 
ophthalmologist. Ophthalmologic examinations will be conducted as per the Schedule of 
Procedures and Assessments and Section 7. Any ocular abnormalities at baseline should be 
recorded in medical history. Corneal specular microscopy results will be sent to a designated 
expert ophthalmology center for review and analysis. Additional examinations may be 
required in the event a subject has an ocular AESI (see Section 8.4). 
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Clinical Study Protocol Amendment 7 
Date: 24Jan2022 
 
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Any subject who discontinues from the study early must return to the clinic for the follow-up 
ophthalmologic examinations 12 weeks from the start of study drug. 
6.3.7 
Suicidal Ideation/Suicidality 
Suicide-related events (behavior and/or ideation) will be assessed and evaluated at every 
scheduled and unscheduled visit with the administration of the C-SSRS (Posner 2007). The 
C-SSRS captures the occurrence, severity, and frequency of suicide-related thoughts and 
behaviors during the assessment period. The scale includes suggested questions to solicit the 
type of information needed to determine if a suicide-related thought or behavior occurred. 
The C-SSRS was developed by the Columbia group to prospectively categorize 
suicide-related events. The 2009 “Baseline” version will be used at Visit 1 (Screening), 
evaluating past year and past 3 months, and the 2009 “Since Last Visit” version will be used 
for all subsequent visits. 
Subjects who score 3 or above on Item 11 of the HAMD-17 must be further evaluated to 
determine is they are at risk or harm to themselves or others (see Section 4.4). If a 
suicide-related event is identified at any time during the study, a thorough evaluation should 
be performed by the investigator or suitably qualified study physician and appropriate 
medical care provided. In some patients taking antidepressants, worsening of depression, 
suicidal events (suicidal thinking and/or behavior), or unusual changes in behavior have been 
reported, especially, at the beginning of the drug therapy, at the time of dose changes, or 
early after treatment discontinuation. It is important that subjects are instructed to notify their 
doctor immediately if they have any distressing thoughts or feelings at any time related to 
harm to either self or others. 
6.3.8 
Opiate Withdrawal 
Opiate withdrawal will be assessed at screening and at all subsequent treatment visits through 
Week 8 using the COWS. The COWS is a clinician-administered instrument used to rate 
11 common opiate withdrawal signs or symptoms (Wesson and Ling 2003). The total score 
of the 11 items will then be used to assess a subject’s level of opiate withdrawal and to 
determine their level of physical dependence on opioids. This assessment provides an 
additional safeguard to ensure no subjects are enrolled with opiate withdrawal or for whom 
withdrawal may begin during the course of the study. 
The COWS will provide an ongoing assessment of subjects for any symptoms suggestive of 
opiate withdrawal. If at any time during the study, a score of 5 or more is obtained on the 
COWS, the investigator (or designee) must determine if the subject is experiencing opioid 
withdrawal. Any subject who is found to be dependent on opioids, abusing opioids, or 
experiencing opioid withdrawal should be withdrawn from the study and provided with 
appropriate medical care. 
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7 
TIMING OF PROCEDURES AND ASSESSMENTS 
Information on study procedures and assessments is provided in Section 6. The timing for all 
assessments is provided in the Schedule of Procedures and Assessments. 
Possible COVID-19 related changes in conduct of study: Study sites will maintain, at a 
minimum, COVID-19 risk mitigation procedures as per regulatory and local guidelines 
and/or laws. Allowed modifications to the conduct of the study, in the event of site staff 
and/or patient restrictions due to COVID-19, are outlined in Appendix 1 of the protocol. In 
addition, to reduce subject burden, subjects may take Visit 3 (Week 2), 4 (Week 4), or 
5 (Week 6) as a virtual visit according to procedures outlined in Appendix 1. 
7.1 
Screening Period (Day –28 to Day–7) 
Before any screening procedure is performed, subject informed consent must be obtained. 
Reinforce the need for the subject to cover skin, and to wear sunscreen and sunglasses when 
exposed to the sun, to minimize the potential risk of sun damage. After obtaining the 
subject’s informed consent, the investigator will conduct/administer the screening 
assessments. Administration of the assessments in the order listed is strongly recommended. 
If any given criterion is not met, the screening should be stopped without completing any of 
the subsequent assessments. 
1. Have subject sign ICF 
2. Enter subject information in IWRS 
3. Demographic details, height and weight 
4. Breathalyzer for blood alcohol level 
5. Subject and investigator will complete the following study eligibility criteria on the tablet 
supplied: 
a. HAMD-17 
b. HAM-A 
c. Allow subject to complete SHAPS; reviewed by site personnel qualified to oversee 
completeness 
d. 
 
e. C-SSRS ‘Baseline’ version with lookback to past year and past 3 months 
f. CGI-S 
g. 
 
6. Medical/psychiatric history review 
7. Complete the SCID-5-CT (including history of substance or alcohol abuse or 
dependence) 
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Clinical Study Protocol Amendment 7 
Date: 24Jan2022 
 
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8. Prior/concomitant therapies review and reporting 
9. Previous and current MDD psychiatric therapy/pharmacologic/nonpharmacologic history  
10. Physical examination 
11. Vital signs (respiratory rate, BP, and pulse [supine and standing]; temperature) 
12. COWS 
13. 12-lead ECG 
14. Blood and urine collection for central laboratory analysis (including urine drug screen for 
all subjects and serum pregnancy test for women of child-bearing potential, and blood for 
thyroid function tests, hepatitis screening, and hepatic monitoring) 
The screening visit may be completed over 2 days for scheduling convenience 7 to 28 days 
prior to Visit 2 (Baseline). The 28-day screening period may be extended to 35 days (after 
consultation with and approval by the medical monitor), if necessary, for reasons including, 
but not limited to, discontinuation of a subject’s current inefficacious medications or to 
obtain an ophthalmologic examination. A retest will be allowed for occasional cannabis users 
(ie, 1 to 2 times/month) only. 
The investigator will review the results of the central laboratory tests for inclusion/exclusion 
criteria and alert the Sponsor-designated medical monitor to confirm eligibility. The 
investigator will be notified of the medical monitor’s decision of the subject’s eligibility prior 
to Visit 2 (Baseline). No subject is to be randomized without prior approval by the CAT team 
and medical monitor. 
After eligibility is confirmed, and before randomization (Visit 2), the following assessments 
must be performed: 
1. Standard ophthalmologic examination 
2. Corneal specular microscopy 
7.2 
Visit 2: Baseline (Day 1) 
The following order of procedures is recommended to prioritize eligibility and safety 
assessments: 
1. Breathalyzer test for blood alcohol level 
2. Subject and investigator will complete the following study eligibility criteria on the tablet 
supplied: 
a. HAMD-17 
b. HAM-A 
c. Allow subject to complete SHAPS; reviewed by site personnel qualified to oversee 
completeness 
d. 
 
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BlackThorn Therapeutics, Inc. 
Clinical Study Protocol Amendment 7 
Date: 24Jan2022 
 
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e. C-SSRS ‘Since Last Visit’ version 
f. 
 
 
g. Allow subject to complete HADS; reviewed by site personnel qualified to oversee 
completeness 
h. Allow subject to complete SDS; reviewed by site personnel qualified to oversee 
completeness 
i. CGI-I and CGI-S 
j. 
 
 
3. Concomitant therapies review and reporting 
4. Physical examination if indicated 
5. Weight 
6. Vital signs (respiratory rate, BP, and pulse [supine and standing]; temperature) 
7. COWS 
8. 12-lead ECG 
9. Urine drug screen for all subjects and urine pregnancy test for women of child-bearing 
potential 
The investigator will confirm the subject’s eligibility prior to completing the remaining 
randomization visit procedures. After confirmation, it is recommended that the remaining 
procedures be completed in the following order: 
1. Complete randomization via IWRS 
2. Collect blood and urine samples for central laboratory analysis,
 
3. Dispense allocated study drug, and instruct subject on the importance of compliance and 
returning the dispensed bottle with any unused study drug at each visit. Instruct all 
subjects to take 1 capsule of their assigned study drug daily in the morning and preferably 
with food; administer the first dose of study drug, which should be taken under the 
supervision of clinical study personnel. It is recommended that subjects remain in the 
clinic for 1 hour after taking study drug. 
4. AE reporting 
The investigator will review the results of the central laboratory tests for safety prior to the 
next visit. 
Prior to Visit 3 (Week 2) (between Days 7 and 14 [+2 days]), a standard ophthalmologic 
examination should be performed (refer to the Schedule of Procedures and Assessments, 
footnote “j” and Section 6.3.6). 
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Clinical Study Protocol Amendment 7 
Date: 24Jan2022 
 
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7.3 
Visit 3: Week 2 (Day 15) 
The following order of procedures is recommended to prioritize safety and efficacy 
assessments: 
1. Breathalyzer test for blood alcohol level 
2. Subject and investigator will conduct/administer the following assessments on the tablet 
supplied: 
a. Allow subject to complete HADS; reviewed by site personnel qualified to oversee 
completeness 
b. C-SSRS ‘Since Last Visit’ version 
c. CGI-I 
3. Concomitant therapies review and reporting 
4. AE reporting 
5. Physical examination if indicated 
6. Vital signs (respiratory rate, BP, and pulse [supine and standing]; temperature) 
7. COWS 
8. The subject-returned investigational study drug will be accounted for and subject will be 
reinstructed if found to be out of compliance 
9. Complete IWRS 
10. Blood and urine collection for central laboratory analysis (including urine drug screen for 
all subjects and urine pregnancy test for women of child-bearing potential). Note: blood 
and urine may be collected earlier in the visit. 
11. Dispense allocated study drug and instruct subject on the importance of compliance and 
returning the dispensed bottle with any unused study drug at each visit.  
12. Instruct all subjects to take 1 capsule of their assigned study drug daily in the morning 
and preferably with food. 
The investigator will review the results of the central laboratory tests for safety in a timely 
manner prior to the next visit. 
7.4 
Visit 4: Week 4 (Day 29) 
The following order of procedures is recommended to prioritize safety and efficacy 
assessments: 
1. Breathalyzer test for blood alcohol level 
2. 
 
3. Administer 1 capsule from the bottle assigned to the subject. 
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Date: 24Jan2022 
 
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4. Subject and investigator will conduct/administer the following assessments on the tablet 
supplied: 
a. HAMD-17 
b. HAM-A 
c. Allow subject to complete SHAPS; reviewed by site personnel qualified to oversee 
completeness 
d. 
 
e. Allow subject to complete HADS; reviewed by site personnel qualified to oversee 
completeness 
f. Allow subject to complete SDS; reviewed by site personnel qualified to oversee 
completeness 
g. C-SSRS ‘Since Last Visit’ version 
h. CGI-I 
5. Concomitant therapies review and reporting 
6. AE reporting 
7. Physical examination, if indicated 
8. Weight 
9. Vital signs (respiratory rate, BP, and pulse [supine and standing]; temperature) 
10. COWS 
11. 12-lead ECG 
12. The subject-returned investigational study drug will be accounted for and subject will be 
reinstructed if found to be out of compliance. 
13. Complete IWRS 
14. Blood and urine collection for central laboratory analysis (including urine drug screen for 
all subjects and urine pregnancy test for women of child-bearing potential) 
15. Dispense allocated study drug and instruct subject on the importance of compliance and 
returning the dispensed bottle(s) with any unused study drug at each visit. 
16. Remind all subjects to take 1 capsule of their assigned study drug daily in the morning 
and preferably with food starting the day after their clinic visit and continuing until their 
next clinic visit 
17. 
 
The investigator will review the results of the central laboratory tests for safety prior to the 
next visit. 
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Clinical Study Protocol Amendment 7 
Date: 24Jan2022 
 
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7.5 
Visit 5: Week 6 (Day 43) 
The following order of procedures is recommended to prioritize safety and efficacy 
assessments: 
1. Breathalyzer test for blood alcohol level 
2. Subject and investigator will conduct/administer the following assessments on the tablet 
supplied: 
a. Allow subject to complete HADS; reviewed by site personnel qualified to oversee 
completeness 
b. Allow subject to complete SDS; reviewed by site personnel qualified to oversee 
completeness 
c. C-SSRS ‘Since Last Visit’ version 
d. CGI-I 
3. Concomitant therapies review and reporting 
4. AE reporting 
5. Physical examination if indicated 
6. Vital signs (respiratory rate, BP, and pulse [supine and standing]; temperature) 
7. COWS 
8. The subject-returned investigational study drug will be accounted for and subject will be 
reinstructed if found to be out of compliance 
9. Complete IWRS 
10. Blood and urine collection for central laboratory analysis (including urine drug screen for 
all subjects and urine pregnancy test for women of child-bearing potential) 
11. Dispense allocated study drug and instruct subject on the importance of compliance and 
returning the dispensed bottle with any unused study drug at each visit. 
12. Remind all subjects to take 1 capsule of their assigned study drug daily in the morning 
and preferably with food 
The investigator will review the results of the central laboratory tests for safety prior to the 
next visit.  
7.6 
Visit 6: Week 8 (Day 57)/End of Treatment 
Special effort will be made by the site to encourage subjects who discontinue from the study 
early to complete all Visit 6 (Week 8) safety assessments (within 2 days of their last dose of 
study drug), the Safety Follow-up TC, and ophthalmologic examinations (Section 7.8); 
efficacy assessments are to be performed only if the subject has remained on study drug. If 
the subject does not attend the discontinuation visit, the C-SSRS should be completed if the 
site has become aware of a potential suicide-related thought or behavior by other 
communications.  
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Date: 24Jan2022 
 
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Additional ophthalmologic examinations (standard ophthalmologic exams and/or corneal 
specular microscopies) may be conducted to monitor subject safety. 
The following order of procedures is recommended to prioritize safety and efficacy 
assessments: 
1. Breathalyzer test for blood alcohol level 
2. Subject and investigator will conduct/administer the following assessments on the tablet 
supplied: 
a. HAMD-17 
b. HAM-A 
c. Allow subject to complete SHAPS; reviewed by site personnel qualified to oversee 
completeness 
d. 
 
e. 
 
f. Allow subject to complete HADS; reviewed by site personnel qualified to oversee 
completeness 
g. Allow subject to complete SDS; reviewed by site personnel qualified to oversee 
completeness 
h. C-SSRS ‘Since Last Visit’ version 
i. CGI-I and CGI-S 
j. 
 
 
k. 
 
3. Concomitant therapies review and reporting  
4. AE reporting 
5. Physical examination if indicated 
6. Weight 
7. Vital signs (respiratory rate, BP, and pulse [supine and standing]; temperature) 
8. COWS 
9. 12-lead ECG 
10. Standard ophthalmologic examination (± 2 days) (refer to Section 6.3.6) 
11. The subject-returned investigational study drug will be accounted for. 
12. Complete IWRS 
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13. Blood and urine collection for central laboratory analysis (including urine drug screen for 
all subjects and urine pregnancy test for women of child-bearing potential) 
14. 
 
The investigator will review the results of the central laboratory tests for safety in a timely 
manner. 
7.7 
Safety Follow-up 
7.7.1 
Telephone Call (Week 10) 
Subjects will receive a TC from the study site approximately 2 weeks after the last dose of 
study drug to inquire about AEs and concomitant medication usage, and will administer the 
C-SSRS. 
7.7.2 
Ocular Examination (Week 12) 
The final ocular examination for corneal specular microscopy is to be conducted 
approximately 12 weeks (+2 days) after the first dose of study drug (including both subjects 
who complete the study and subjects who discontinue early from the study); exceptions to be 
discussed with medical monitor.  
 
Additional ophthalmologic examinations (standard ophthalmologic exams and/or corneal 
specular microscopies) may be conducted to monitor subject safety. 
7.8 
Early Termination 
Subjects who discontinue early from the study should be encouraged to complete the End of 
Treatment (Visit 6) safety assessments within 2 days after their last dose of study drug; 
efficacy assessments are to be performed only if the subject has remained on study drug. If 
the subject does return for an End of Treatment visit, the C-SSRS ‘Since Last Visit’ version 
should be completed if the site has become aware of a potential suicide-related thought or 
behavior by other communications (see Section 7.6). 
Subjects who discontinue early from the study should plan to complete the Week 12 corneal 
specular microscopy (see Section 7.7.2).  
 
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8 
SAFETY MONITORING AND REPORTING 
Investigators are responsible for the detection and documentation of events that meet the 
definition of an AE, an SAE, suspected adverse reaction, serious suspected adverse reaction, 
unanticipated problems, or AESIs, as provided in this protocol. 
Investigators must review the BTRX-335140 IB to be aware of the safety-related events, 
which may be anticipated with its use. Investigators will also be versed in the latest standard 
of care guidelines. 
8.1 
Adverse Events 
8.1.1 
Definition of an Adverse Event 
An AE is defined as any untoward medical occurrence associated with the use of a drug in 
humans, whether or not considered drug related. 
An AE may be any unfavorable and unintended sign, symptom, illness, clinically significant 
abnormal laboratory value or ECG finding, or other untoward medical occurrence that 
appears or worsens in a subject during a clinical study. This definition does not imply a 
causal relationship between the AE and the study drug. 
Adverse events beginning with the initial dose of study drug or during the subsequent 
duration of their enrollment in the study will be considered TEAEs. Treatment-emergent AEs 
will be recorded on the AE eCRF to include the event, date of onset, whether the AE is 
associated with an episode of self-harm, severity, frequency, seriousness, date of resolution, 
action taken with respect to the AE (eg, discontinue study drug, begin concomitant 
medication, begin nonpharmacological treatment, etc.), outcome, and relationship to the 
study drug. All TEAEs related to study drug will be followed to a satisfactory resolution or 
until the event becomes stable or can be explained by another known cause (ie, concurrent 
condition or medication). 
Medical conditions present or AEs occurring prior to the first dose of study drug will be 
captured as medical history in the eCRF. Any AE occurring during the study that is related to 
a pre-existing condition or event that worsens in intensity or frequency after the first dose of 
study drug will be recorded as a TEAE.  
Lack of drug effect is not an AE in clinical studies because the purpose of the clinical study 
is to establish drug effect. 
8.1.2 
Definition of a Serious Adverse Event 
Serious AEs will be recorded on the AE eCRF from the time of informed consent. An SAE is 
any AE that results in 1 of the following outcomes, regardless of the investigator’s opinion of 
causation: 
• Death 
• Initial or prolonged inpatient hospitalization 
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- 
Surgeries planned prior to signing the ICF will not be considered SAEs. However, 
worsening of the underlying medical condition during the study will be considered an 
AE and must be captured as serious if any SAE-defining outcomes occur as a result. 
• A life-threatening experience (that is, immediate risk of dying) 
• Persistent or significant disability/incapacity 
• Congenital anomaly/birth defect 
• Other medically important serious event as determined by the investigator (for example, 
an AE that jeopardizes the subject and may require medical or surgical intervention to 
prevent 1 of the outcomes listed in this definition) 
 
The following events do not meet the definition of an SAE:  
• Hospitalization for elective treatment of a pre-existing condition that does not worsen 
from baseline 
• Hospitalizations for a standard procedure for study drug administration and routine 
monitoring of the studied indication not associated with any deterioration in condition 
• Social or convenience admission to a hospital 
• Prolongation of a hospitalization for social or convenience reasons not associated with 
the occurrence of an AE 
• Hospitalization or an emergency room visit that lasts less than 24 hours that does not 
meet the criteria of an important medical or a life-threatening event 
 
8.1.3 
Definition of Suspected Adverse Reaction 
A suspected adverse reaction is defined as any AE for which there is a reasonable possibility 
that the AE was caused by the study drug. 
8.1.4 
Definition of Serious Suspected Adverse Reaction 
A serious suspected adverse reaction is any suspected adverse reaction that is determined to 
be serious, based on the definition of an SAE in Section 8.1.2. 
8.2 
Classification of Adverse Events 
8.2.1 
Severity of Adverse Events 
The investigator will assess the severity of each AE based on his/her clinical judgment using 
one of the following categories: 
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• Mild: Usually transient and may require only minimal treatment or therapeutic 
intervention. The event does not generally interfere with usual activities of daily living.  
• Moderate: Usually alleviated with additional specific therapeutic intervention. The event 
interferes with usual activities of daily living, causing discomfort but poses no significant 
or permanent risk of harm to the subject. 
• Severe: Interrupts usual activities of daily living, or significantly affects clinical status, or 
may require intensive therapeutic intervention. 
8.2.2 
Relationship to Study Drug 
The investigator will assess the relationship (ie, causality) of each AE to study drug based on 
his/her clinical judgment. The investigator’s assessment of an AE’s relationship to study drug 
is part of the documentation process, but it is not a factor in determining what is or is not 
reported in the study. If there is any doubt as to whether a clinical observation is an AE, the 
event should be reported. All AEs must have their relationship to study agent assessed. The 
Sponsor’s assessment of relationship may differ from the investigator’s assessment. 
Relationship to study drug will be assessed according to the following guidelines: 
• Possibly related: The AE is known to occur with the study drug, there is a reasonable 
possibility that the study drug caused the AE, or there is a temporal relationship between 
the study drug and event. Reasonable possibility means that there is evidence to suggest a 
causal relationship between the study drug and the AE. 
• Unlikely related: There is not a reasonable possibility that the administration of the 
study drug caused the event, there is no temporal relationship between the study drug and 
event onset, or an alternate etiology has been established. 
For initially reporting SAEs, even in situations in which minimal information is available, it 
is important that the investigator make an assessment of causality for every event. The 
causality assessment is 1 of the criteria used when determining regulatory reporting 
requirements. The investigator may change his or her opinion of causality in light of 
follow-up information and amend the SAE information accordingly in the eCRF or the SAE 
reporting form, as applicable. 
8.3 
Time Period and Frequency for Event Assessment and Follow-up 
All AEs from the time of informed consent through the last follow-up visit will be recorded 
in each enrolled subject’s eCRF (see Section 6.3.1). For subjects who prematurely 
discontinue from the study, AEs will continue to be recorded through the last follow-up visit. 
For subjects who are found to be ineligible for the study during the Screening period and are 
not enrolled (ie, Screening failures), AEs that are SAEs will be captured. Adverse events 
beginning with the initial dose of study drug or during the subsequent duration of their 
enrollment in the study will be considered TEAEs. Treatment-emergent AEs will be recorded 
on the AE eCRF to include the event, date of onset, whether the AE is associated with an 
episode of self-harm, severity, frequency, seriousness, date of resolution, action taken with 
respect to the AE (eg, discontinue study drug, begin concomitant medication, begin 
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nonpharmacological treatment, etc.), outcome, and relationship to the study drug. All TEAEs 
related to study drug will be followed to a satisfactory resolution or until the event becomes 
stable or can be explained by another known cause (ie, concurrent condition or medication). 
• All SAEs that occur after the informed consent is signed will be captured. Serious AEs 
reported after a subject has taken the last dose of study drug will be collected in the 
pharmacovigilance system for 30 days after the last dose of study drug. Thereafter, only 
SAEs that the investigator feels were related to the study drug or a protocol procedure 
must be reported. 
8.4 
Adverse Events of Special Interest 
Adverse events of special interest (serious or nonserious) are TEAEs of particular safety 
importance and are required to be reported to the sponsor immediately (and no more than 
24 hours after learning of the event, see Section 8.5 for reporting requirements). 
Adverse events of special interest for this study are based on nonclinical findings of 
skin-related and ocular phototoxicity (see Section 1.1.3). Reported TEAEs that are classified 
in the system organ classes (SOCs) listed below will be considered AESIs: 
• Skin and subcutaneous tissue disorders 
• Eye disorders 
Nonserious AEs in the above SOCs that occur prior to study drug dosing will be recorded in 
medical history. 
Investigators will monitor for dermatologic or ocular events and for any change in visual 
acuity. 
8.5 
Reporting Procedures 
8.5.1 
Reporting Serious Adverse Events and Adverse Events of Special Interest 
All SAEs and AESIs must be reported on a study-specific SAE/AESI form in addition to the 
AE eCRF by electronic data capture (EDC). Study site personnel must alert the 
Sponsor-designated medical monitor of the SAE/AESI and must submit the completed 
study-specific SAE/AESI form to
 
 within 24 hours of the site’s awareness of the 
SAE/AESI. 
The investigator must also enter the SAE/AESI information into the eCRF as soon as 
possible thereafter. 
In the initial email, the investigator must provide to the Sponsor the SAE/AESI form, 
completed to the greatest extent possible and the following pages from the eCRF: 
• AE record 
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• Medical history 
• Prior and concomitant medications 
Also, the following documents are to be forwarded: any laboratory results, diagnostic test 
results, or medical reports relevant to the SAE/AESI. 
The investigator and supporting personnel responsible for subject care should discuss with 
the Sponsor-designated medical monitor or designee any need for supplemental 
investigations of SAEs/AESIs. The results of these additional assessments conducted must be 
reported to the Sponsor-designated medical monitor.  
In the event of a medical emergency in which knowledge of the subject’s treatment 
assignment may influence their clinical care the investigator has the option to unblind the 
subjects’ treatment assignment via the IWRS system. The investigator should make every 
effort to contact the medical monitor prior to unblinding unless this would adversely delay 
appropriate medical care. The medical monitor will not be unblinded, they will only provide 
assistance to the investigator. The reasons for unblinding must be noted in the source 
documents. The investigator must not disclose the subject’s treatment assignment to anyone 
who does not need the information due to their direct involvement in the subject’s clinical 
care. Disposition of subjects who are unblinded due to a medical emergency will be 
determined following discussion with the Sponsor. 
In the event of death, every effort should be made to obtain a death certificate and if possible, 
an autopsy report. If the cause of death is unknown, death will be recorded as the event.  
The investigator is responsible for safety reporting in compliance with their IRB. 
8.5.2 
Regulatory Reporting Requirements 
The investigator must promptly report all SAEs to the Sponsor in accordance with the 
procedures detailed in Section 8.5.1. The Sponsor has a legal responsibility to notify, as 
appropriate, both the local regulatory authority and other regulatory agencies about the safety 
of a product under clinical investigation. Prompt notification of SAEs by the investigator to 
the appropriate project contact for SAE receipt is essential so that serious suspected adverse 
reactions that are either unexpected or observed with increasing occurrence be reported and 
legal obligations and ethical responsibilities regarding the safety of other subjects are met. 
Investigator letters are prepared according to Sponsor policy and are forwarded to the 
investigators as necessary. An investigator letter is prepared for a serious unexpected 
suspected adverse reaction (SUSAR) that is attributable to study drug. The purpose of the 
investigator letter is to fulfill specific regulatory and cGCP requirements regarding the 
product under investigation. 
The investigator, or responsible person according to local requirements, must comply with 
requirements related to the reporting of SAEs to the IRB. 
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The Sponsor is responsible for informing IRB, investigators, and regulatory authorities of 
any finding that could adversely affect the safety of subjects or affect the conduct of the 
study. Events will be reported to regulatory authorities in accordance with expedited 
reporting requirements. 
8.5.3 
Overdose 
Overdose is defined as an accidental or intentional administration of more than the intended 
dose of study drug. In the event of an overdose of study drug, the investigator should use 
clinical judgment in treating the overdose and contact the Sponsor-designated medical 
monitor, or designee. The investigator should refer to the relevant document(s) for detailed 
information regarding warnings, precautions, contraindications, AEs, and other significant 
data pertaining to the study drug. Such documentation may include, but not be limited to the 
IB. 
8.5.4 
Pregnancy Reporting 
If a female subject or the female partner of a treated male subject becomes pregnant, the 
subject must notify the investigator within 24 hours of learning of the pregnancy. The 
investigator must make every effort to ensure that the subject or the pregnant female is aware 
of the need to notify her healthcare provider regarding her male partner’s participation in this 
clinical study and his potential exposure to BTRX-335140. 
The study site must complete a pregnancy form and send to the Sponsor or designee within 
24 hours of learning of the pregnancy. The study site will make every effort to follow the 
pregnancy until outcome is known. 
8.6 
Safety Oversight 
The Sponsor-designated medical monitor and the study team will monitor blinded safety data 
throughout the course of the study as detailed in the medical monitoring plan. This will 
include review of AEs, SAEs, AESIs, ECG findings, clinical laboratory results, vital signs, 
ophthalmologic findings, and other safety data. 
In the event that safety monitoring uncovers an issue that needs to be addressed by 
unblinding at the group level, a data monitoring committee will be formed to protect the 
integrity of data and to evaluate additional analyses of the safety data. 
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9 
STATISTICAL CONSIDERATIONS 
9.1 
General Considerations 
Full details on the statistical methodology to be used will be included in a statistical analysis 
plan (SAP) to be developed prior to study unblinding. 
For the purpose of all safety analyses where applicable, baseline is defined as the last 
measurement prior to the start of study drug administration. For efficacy analyses, baseline is 
defined as the arithmetic mean of pretreatment measurements ie, Screening and Baseline, 
prior to the first administration of study drug, or as the last measurement prior to the first 
administration of study drug if only single value is available. 
Continuous endpoints will be summarized with n, mean, standard deviation (SD), median, 
minimum and maximum. In addition, change from baseline and percent change from baseline 
values will be calculated at each time point and summarized descriptively. For categorical 
endpoints, descriptive summaries will include counts and percentages.  
No adjustments will be made for multiple comparisons.  
All statistical comparisons will be 2-sided with alpha of 0.05.  
SAS version 9.4 or higher will be used to analyze data.  
9.2 
Analysis Populations 
The safety analysis will be conducted on the Safety Population. Efficacy analyses will be 
conducted on the Efficacy population and the per-protocol (PP) population. The Efficacy 
population will be used for the primary efficacy analysis. 
 
 Subjects will be analyzed according to randomized treatment assignment for 
efficacy assessments and according to treatment received for safety assessments. 
9.2.1 
Safety Population 
The Safety population is defined as all subjects who received study drug and will be used to 
summarize safety data. 
9.2.2 
Efficacy Population 
The Efficacy population includes all randomized subjects who received at least 1 dose of 
study drug, have a baseline HAMD-17 assessment value, and have at least 1 post-baseline 
assessment. This population will be used to summarize the primary and other efficacy data. 
9.2.3 
Per-protocol Population 
The PP population includes subjects in the efficacy population who did not experience any 
major protocol deviations that impact primary efficacy endpoint analysis. 
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9.2.4 
 
 
9.3 
Statistical Analysis Methods 
9.3.1 
Disposition 
Frequency counts and percentages of subjects who were screened, randomized, completed 
the treatment, or discontinued early will be presented by treatment group. Reasons for 
discontinuation will be summarized. 
9.3.2 
Demographics and Baseline Characteristics 
Demographic and baseline characteristics including age, sex, BMI, race, ethnicity, height, 
weight, CGI-S, baseline HAMD-17, duration of current depressive episode, antidepressant 
treatment during the current episode, and other medical history, physical examinations, and 
prior and concomitant medications will be summarized using descriptive statistics. Prior and 
concomitant medications will be summarized by World Health Organization Drug 
Classification (WHODrug) for Drug Statistics Methodology. 
9.3.3 
Efficacy Analyses 
9.3.3.1 
Primary Efficacy Analysis 
The primary efficacy analysis is a comparison of mean reduction in HAMD-17 scores 
between BTRX-335140 and placebo at Week 8. The analysis will be conducted using a 
Mixed-Models Repeated Measures (MMRM) with change from baseline in HAMD-17 score 
as the dependent variable and will include treatment group, Week, and Week by treatment 
group interaction as factors, and baseline HAMD-17 score as covariate. Variance estimation 
will be based on an unstructured covariance matrix. The model-based Week 8 least square 
(LS) means, standard errors, 95% confidence intervals (CIs), and p-values will be reported. 
9.3.3.2 
Secondary Efficacy Analyses 
Similar to the method described above, a MMRM model will be calculated for the change 
from baseline to each timepoint assessed for comparisons between BTRX-335140 and 
placebo for the measures listed below. The model-based time points’ LS means, standard 
errors, 95% CIs, and p-values will be reported. Alternatively, ANCOVA may be used, as 
appropriate. The following measures will be analyzed: 
• CGI-I 
• CGI-S 
• SHAPS 
• HADS subscales: HADS-A and HADS-D 
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• HAM-A total score 
• SDS 
For CGI-I, an additional analysis comparing the percentage of subjects with responses at 
Week 4 and Week 8 of ‘much improved’ or ‘very much improved’ between BTRX-335140 
and placebo will be performed using generalized estimating equations (GEE). The model will 
include treatment, baseline CGI-S score, week, and week by treatment group interaction. An 
exchangeable working correlation structure will be assumed.  
The change from baseline to Week 8 in CGI-S will be estimated using an ANCOVA model, 
with main effects for treatment group and a covariate adjustment for baseline score. 
The response rates (the percentage of subjects with ≥ 50% decrease in HAMD-17 from 
baseline to Week 4 and Week 8) will be compared between BTRX-335140 and placebo-
treated groups using Cochran-Mantel-Haenszel test. Additional categories may be explored.  
. 
9.3.4 
Safety Analysis 
Safety data, including AEs, safety laboratory results, physical examination results, , vital 
signs, suicidality, COWS scores, and ECG will be summarized by treatment group (placebo 
and BTRX-335140) and/or listed. 
9.3.4.1 
Adverse Events 
Adverse events occurring after the first dose of study drug will be considered to be TEAEs. 
The incidence of TEAEs will be summarized by seriousness, severity, and causality to study 
drug. Treatment-emergent AEs leading to discontinuation will also be summarized. 
Adverse events with onset before first study dose and after last study dose will be listed. 
9.3.4.2 
Adverse Events of Special Interest 
Adverse events of special interest for this study are based on nonclinical findings of 
skin-related and ocular phototoxicity (see Section 1.1.3 and Section 8.4). AESIs will be 
summarized by SOC/preferred term.  
9.3.4.3 
Vital Signs 
Changes from baseline will be summarized for temperature, pulse (supine and standing), 
respiratory rate (supine and standing), and BP (supine and standing) by treatment and visit.  
9.3.4.4 
Physical Examinations 
Clinically significant physical examination abnormalities will be listed. 
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9.3.4.5 
Clinical Laboratory Tests 
Changes from baseline will be summarized for each laboratory test by study treatment and 
visit. 
9.3.4.6 
Electrocardiography 
Electrocardiographic parameters will be listed. Clinically significant abnormalities on ECGs 
will be listed by subject and visit. 
9.3.4.7 
Ophthalmology 
Ophthalmologic endpoints will be summarized. Clinically significant abnormalities on 
ophthalmologic exam will be listed by subject and visit. 
9.3.4.8 
Columbia Suicide Severity Rating Scale 
Suicidality will be listed for all subjects. 
9.3.4.9 
Clinical Opiate Withdrawal Scale 
COWS scale items and total scores, along with the numbers of subjects with a score of ≥5, 
will be summarized at each visit by treatment group. 
9.3.4.10 
Treatment Compliance with Study Drug 
A subject is defined to be compliant if he/she has taken ≥80% and <120% of the number of 
doses expected to be taken during the study overall. Noncompliant subjects will be listed by 
treatment group. 
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9.3.6 
Subgroup Analyses 
A subgroup analysis based on gender may be performed. Subgroup analyses based on other 
characteristics will be considered and included in the SAP.  
Efficacy endpoint analyses, which exclude the subjects (n=24) enrolled at the time of the 
study enrollment shutdown due to COVID-19, will be performed (Section 9.5). Details of 
these analyses will be included in the SAP. 
The impact of the efficacy endpoints ascertainment (in-person vs telephone) will be explored 
in additional analyses, if a large imbalance in ascertainment method between treated and 
placebo subjects is present at study visits. Details of these analyses will be included in the 
SAP. 
9.4 
Interim Analysis 
No interim analyses are planned for this study. 
 
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10 
QUALITY ASSURANCE AND QUALITY CONTROL 
Quality assurance and quality control systems will be implemented and maintained with 
Standard Operating Procedures (SOPs) by the Sponsor and/or its designee(s), as appropriate, 
to ensure that the clinical study is conducted and the data are generated, documented 
(recorded), and reported in compliance with the protocol, International Council for 
Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH)-GCP 
E6(R2) guidelines, and applicable regulatory requirements. The accuracy, completeness, and 
reliability of the study data presented to the Sponsor, however, are the responsibility of the 
investigator. The investigator or designee must record all required data using the prespecified 
data collection method defined by the Sponsor or its designee.  
The study will be monitored regularly by the Sponsor (Section 8.6) and may be audited or 
inspected by the Sponsor (or designee), IRB, and/or regulatory authorities at any time during 
the study or after study completion. In the event of an audit, the investigator agrees to allow 
the Sponsor, representatives of the Sponsor, the competent authority, or other regulatory 
agencies direct access to all study records. The investigator will immediately notify the 
Sponsor of all audits or inspections scheduled by any regulatory authority and promptly 
forward copies of any audit or inspection reports received to the Sponsor. 
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11 
REGULATORY AND ETHICAL CONSIDERATIONS 
11.1 
Regulatory Considerations 
This study will be conducted in accordance with consensus ethics principles derived from 
international ethics guidelines, including the Declaration of Helsinki and Council for 
International Organizations of Medical Sciences (CIOMS) International Ethical Guidelines, 
the ICH GCP Guideline [E6], and other applicable laws and regulations. 
The Sponsor certifies that this study is being conducted under an active US Investigational 
New Drug (IND) application.  
Some of the obligations of the Sponsor will be assigned to a CRO. 
11.2 
Ethical Conduct of the Study 
This study will comply with the requirements that are enunciated in the European Clinical 
Trial Directive 2001/20/EC and in the US Code of Federal Regulations (CFR). 
The Sponsor or its representatives must approve all ICFs before they are submitted to the 
IRB and are used at investigative sites(s). All ICFs must be compliant with the ICH guideline 
on cGCP.  
Documentation of IRB approval of the protocol and the ICF must be provided to the Sponsor 
before the study may begin at the investigative site(s). The IRB(s) will review the protocol as 
required.  
Any member of the IRB who is directly affiliated with this study as an investigator or as site 
personnel must abstain from the IRB’s vote on the approval of the protocol. 
11.3 
Institutional Review Board Approval  
This study will be conducted in full compliance with the IRB regulations in 21 CFR 56. 
Before enrollment of subjects into the study, the protocol, informed consent documents and 
any subject related advertising will be reviewed and approved by the appropriate IRB and 
regulatory authority. Amendments to the protocol will be submitted to the same IRB and 
regulatory authority review requirements as the original protocol. The investigator will 
promptly notify the IRB and Sponsor of any SAEs or of any other information that might 
affect the safe use of the investigational product during the study. Institutional Review Board 
approvals positive opinions and regulatory authorities’ approvals must be sent to the 
Sponsor, or its designee, before initiation of the study or before an amendment is instituted. 
All correspondence with the IRB and the regulatory authority must be retained in the study 
regulatory files. 
11.4 
Informed Consent Process 
Written informed consent from each subject must be obtained before any study-specific 
Screening or Baseline period evaluations are performed. If any new information becomes 
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available that might affect patients’ willingness to participate in the study, or if any 
amendments to the protocol require changes to the informed consent/assent form, the 
Sponsor will provide Investigators with a revised informed consent/assent form. One copy of 
the signed informed consent documents will be given to the subject; the investigator will 
retain the original copies of these documents.  
The informed consent document, as prepared by the Sponsor or designee, must be reviewed 
and approved by the IRB before initiation of the study. The informed consent document must 
contain the basic required elements of consent and additional elements, as applicable, as 
specified in the 21 CFR 50.25. 
The investigator is responsible for ensuring that the subject understands the potential risks 
and benefits of participating in the study, including answering any questions the subject may 
have throughout the study and sharing in a timely manner any new information that may be 
relevant to the subject’s willingness to continue his or her participation in the study.  
The ICF will be used to explain the potential risks and benefits of study participation to the 
subject in simple terms before the subject is entered into the study, and to document that the 
subject is satisfied with his or her understanding of the risks and benefits of participating in 
the study and desires to participate in the study.  
The investigator is responsible for ensuring that informed consent is given by each subject. 
This includes obtaining the appropriate signatures and dates on the ICF prior to the 
performance of any protocol procedures and prior to the administration of study drug. 
11.5 
Confidentiality 
All information regarding the nature of the proposed investigation that is provided to the 
investigator by the Sponsor, the Sponsor’s designee, or the study monitor, with the exception 
of information that is required by law or regulations to be disclosed to the IRB/IEC, the 
subject’s parent(s) or legal guardian(s) or the appropriate regulatory authority, must be kept 
in confidence by the investigator in accordance with current Health Insurance Portability and 
Accountability Act (HIPAA) standards and/or European standards.  
The anonymity of participating subjects will be maintained to the extent required by 
applicable laws and in accordance with current HIPAA standards. Subjects may be 
referenced by their initials and an assigned subject identification number on the CRFs and 
other data collected by the Sponsor. The investigator must maintain all documents related to 
the study that identify the subject (eg, the signed informed consent document) in strict 
confidence, except to the extent necessary to allow auditing by the appropriate regulatory 
authorities, the IRB/IEC, the study monitor, or the Sponsor or its representatives. 
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12 
STUDY ADMINISTRATION 
12.1 
Clinical Monitoring 
Electronic CRFs will be completed for all study subjects enrolled in the study. At scheduled 
monitoring visits, eCRFs will be verified against source documentation. In the event that 
on-site scheduled monitoring cannot be conducted, the monitoring of data in the eCRF will 
be detailed in the clinical monitoring plan. Upon completion of this review, finalized eCRFs 
will be submitted to the Sponsor’s designated CRO. Any subsequent changes to the eCRFs 
will be performed in accordance with the CRO’s SOPs for editing and clarifying eCRFs. 
Data entry will be performed through username and password-protected access to a secure 
database. All data will be entered using electronic eCRFs. Internally developed programs for 
plausibility, consistency, and out-of-range data fields will supplement the review of the data. 
A 100% manual review of AEs, drug accountability and termination summary data, will be 
performed by Data Management personnel. The Medical Dictionary for Regulatory 
Activities (MedDRA) coding thesaurus will be used to classify AEs and medical history, and 
the WHODrug classification will be used to code medications. 
After all subjects complete the study and data discrepancies are resolved, and prior to study 
unblinding, protocol deviations during both enrollment and study execution will be reviewed. 
Significant protocol deviations and procedural discrepancies will be discussed and major 
deviations will be identified and reported. 
12.2 
Source Documents and Record Retention 
Essential study documents are among the critical documents required before study 
enrollment is to occur. Essential documents, as well as supplemental information such as the 
IB, Pharmacy Manual, CRF Completion Guidelines, final protocol, as specified in the 
Clinical Operations Manual and/or Regulatory Binder, must be kept on-site in a designated 
study site file.  
The study site files will also contain, including but not limited to, subject accountability 
records, drug accountability (receipt/dispensing) records, Sponsor/investigator 
correspondence, IRB/IEC correspondence, deviations, biological sample records, and SAE 
and IND safety reports/Safety Alert Letters/SUSARs. 
12.3 
Management of Protocol Amendments and Deviations 
All processes and procedures defined in this protocol must be adhered to. Emergency 
departures from the protocol that eliminate an apparent immediate hazard to a particular 
subject and that are deemed by the investigator as crucial for the safety and wellbeing of that 
subject may be instituted for that subject only and documented as deviations. The 
investigator will contact the medical monitor as soon as possible regarding such a deviation. 
These departures do not require preapproval by the IRB/IEC; however, the IRB and medical 
monitor must be notified in writing as soon as possible in accordance with the IRB policies 
after the departure has been made. 
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Study records must be retained for at least 2 years after the last approval of a marketing 
application in an ICH region and until there are no pending or contemplated marketing 
applications in an ICH region or until at least 2 years have elapsed since the formal 
discontinuation of clinical development of the investigational product. However, these 
documents should be retained for a longer period if required by applicable regulatory 
requirements or if agreed to in the Clinical Trial Agreement. It is the responsibility of the 
Sponsor to inform the site as to when these documents no longer need to be retained. 
12.3.1 
Protocol Modifications 
The protocol cannot be modified except in a formal protocol amendment by the Sponsor. 
12.3.2 
Protocol Violations and Deviations 
Protocol deviations are a change, divergence, or departure from the study design or 
procedures defined in this protocol. An Important Protocol Deviation is a deviation that may 
significantly impact the completeness, accuracy, and/or reliability of the study data or that 
may significantly affect a subject’s rights, safety, or well-being. The investigator will notify 
the IRB of any protocol deviations as required by IRB guidelines and site requirements. 
Protocol deviations will be documented at the site and in the Sponsor files. In the event of an 
Important Protocol Deviation, the site will notify the Sponsor or designee. The Sponsor is 
responsible for notifying the regulatory authorities of any protocol deviations, as required. 
12.4 
Financial Disclosure 
Investigators are required to inform the Sponsor of all disclosable financial interests or 
arrangements (including those of their spouse and dependent children), prior to study 
initiation at the site, at study completion, and 1 year after study completion in accordance 
with 21 CFR Part 54. In addition, the investigator or sub-investigators must promptly notify 
the Sponsor if there are any reportable changes that occur during the above-described period.  
Disclosable financial interests or arrangements, or the absence thereof will be recorded on the 
Financial Disclosure for Clinical Investigators Form. 
Any investigator(s) added as investigational staff to the Food and Drug Administration 
(FDA) 1572 form must complete the Financial Disclosure for Clinical Investigators Form at 
the start of his/her participation in the study. The Financial Disclosure for Clinical 
Investigators Form for any investigator(s) leaving the study prior to completion will also be 
obtained. 
12.5 
Stopping Criteria: Suspension or Termination of Study or Investigational 
Site 
The Sponsor may suspend or terminate the study, or part of the study, at any time for any 
reason. If the study is suspended or terminated, the Sponsor will ensure that applicable sites, 
regulatory agencies, and IRBs/ECs are notified as appropriate. 
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If the Sponsor, investigator, or officials from regulatory agencies discover conditions arising 
during the study that indicate that the study should be halted or that a study site should be 
closed, this action may be taken after appropriate consultation between the Sponsor and 
investigator(s). 
12.6 
Publication and Information Disclosure Policy 
The information that is developed during the conduct of this clinical study is considered to be 
strictly confidential. This information may be disclosed only as deemed necessary by 
BlackThorn. At the conclusion of this clinical study, a clinical study report will be prepared. 
In addition, a manuscript may be prepared for publication in a reputable scientific journal 
under the direction of the Sponsor. BlackThorn will publish and communicate the clinical 
study results, irrespective of positive or negative findings. Data generated for this study will 
be exclusively owned by BlackThorn, as detailed in the Clinical Trial Agreement. The study 
will be registered on ClinicalTrials.gov. After completion of the study, results will be 
disseminated through ClinicalTrials.gov. 
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13 
REFERENCES 
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Beardsley PM, Howard JL, Shelton KL, et al. Differential effects of the novel kappa opioid 
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Bruchas MR, Land BB, Lemos JC, et al. CRF1-R activation of the dynorphin/kappa opioid 
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Carlezon WA Jr, Krystal AD. Kappa-Opioid Antagonists for Psychiatric Disorders: From 
Bench to Clinical Trials. Depress Anxiety. 2016 Oct;33(10):895-906. Doi: 
10.1002/da.22500. Review. PMID: 27699938 
Craske MG, Meuret AE, Ritz T, et al. Treatment for Anhedonia: A Neuroscience Driven 
Approach. Depress Anxiety. 2016;33(10):927-38. 
Dunlop BW, Nemeroff CB. The Role of Dopamine in the Pathophysiology of Depression. 
Arch Gen Psychiatry. 2007;64(3)327-37. 
Fava GA, Tomba E, Grandi S. The road to recovery from depression-don’t drive today with 
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Fava M, Evins AE, Dorer DJ, et al. The problem of the placebo response in clinical trials for 
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Guy W. ECDEU assessment manual for psychopharmacology-revised. Rockville, MD: 
National Institute of Mental Health, Psychopharmacology Research Branch, Division of 
Extramural Research Programs. 1976:217-22. 
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Clinical Study Protocol Amendment 7 
Date: 24Jan2022 
 
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Hamilton M. A rating scale for depression. J Neurol Neurosurg Psychiatry. 1960;23:56-62. 
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Depression and anxiety. 2008;25(11):913-9. 
Krystal AD, Pizzagalli DA, Smoski M, et al. A randomized proof-of-mechanism trial 
applying the ‘fast-fail’ approach to evaluating κ-opioid antagonism as a treatment for 
anhedonia. Nat Med 2020:26:760-768. https://doi.org/10.1038/s41591-020-0806-7. 
Lally N, Nugent AC, Luckenbaugh DA, et al. Anti-anhedonic effect of ketamine and its 
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Levey AS, Stevens LA, Schmid CH, et al. A new equation to estimate glomerular filtration 
rate. Ann Intern Med. 2009;150(9):604-12. 
McCabe C, Mishor Z, Cowen PJ, et al. Diminished neural processing of aversive and 
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McMakin DL, Olino TM, Porta G, et al. Anhedonia predicts poorer recovery among youth 
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Morris BH, Bylsma LM, Rottenberg J. Does emotion predict the course of major depressive 
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Murray CJ, Lopez AD. The global burden of disease: a comprehensive assessment of 
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Nutt D, Demyttenaere K, Janka Z, et al. The other face of depression, reduced positive affect: 
the role of catecholamines in causation and cure. J. Psychopharmocol. 2007;21(5): 461-71. 
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BlackThorn Therapeutics, Inc. 
Clinical Study Protocol Amendment 7 
Date: 24Jan2022 
 
Confidential 
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Pelizza L and Ferrari A. Anhedonia in schizophrenia and major depression: state or trait? 
Ann. of Gen. Psychiatry. 2009;8:22. 
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Psychol Med. 2012;42(5):967–80. 
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1983;67(6):361–70. 
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CNS Drugs. 2006;20(12):981–92. 
 
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APPENDIX 1: 
GUIDANCE FOR VIRTUAL VISITS (VIA TELEPHONE) 
The guidance below is intended for subjects who have been successfully enrolled and 
randomized into the study and who elect to conduct a virtual visit to reduce subject burden or 
who are unable to attend in-person clinic visits due to COVID-19 restrictions. 
Every reasonable effort should be made to ensure that the subjects continue on study per 
protocol; however, in the event that a subject cannot come to the clinic after the baseline visit 
due to self-quarantine, local restrictions, illness, or other reasons as described in the protocol, 
the site personnel may conduct a virtual (by telephone contact) visit. Clinical laboratory tests 
are to be performed at a minimum every 4 weeks. Up to 2 nonsequential virtual visits 
between Visits 3 and 5 may be allowed. If the subject is unable to return for Visit 6, the 
subject should return to the site, at their earliest convenience, for an End of Treatment visit 
(see Sections 4.4 and 7.8). 
Conducting Study Procedures for the Virtual Visit 
The following procedures are to be completed via the telephone (see Table A1-1): 
• The following clinician-completed assessments can be performed via the telephone: 
HAM-A, HAMD-17, CGI-S, CGI-I, and C-SSRS. Raters will need to document, in the 
tablet, if data were collected via phone (vs in person). 
• The following subject-rated scales will be conducted over the phone:
, HADS, SDS, 
, and SHAPS. Subjects must have the PDF copy of the SDS with them during the 
assessment. Following the subject’s self-assessment on the SDS, the rater should review 
the Work/School and Days Lost items to confirm appropriateness of the selected ratings 
by evaluating the impact of MDD symptoms across those sectors. Site personnel will 
enter the information directly into the tablet on behalf of the subject and will indicate, in 
the tablet, that this information was captured over the phone. 
• The following will also be discussed with the subject: AEs and concomitant medications, 
and alcohol and drug use; details will be captured in the source data. As the COWS 
cannot be fully administered by phone (as some items require direct observations), in 
capturing AEs site staff should be alert to symptoms that require further enquiry to 
determine if there is any possibility of opioid withdrawal. 
• Study drug compliance will be discussed with the subject; however, pill count will be 
completed at the time of the next visit when the subject returns to the clinic. 
The above assessments are required to be done as listed. 
 
 
When subjects cannot come into the clinic and a virtual visit is performed, all other 
assessments including laboratory testing, vital signs measurements, and ECG will be noted as 
unable to be completed. 
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Table A1-1: Procedures for Virtual Visit 
Period 
Virtual Visit 
Visit 
3 – 5  
Guidance 
Procedure and Assessments 
 
 
Vital signs 
NA 
 
Physical examination 
NA 
 
Body weight 
NA 
 
12-Lead ECG 
NA 
 
Standard ophthalmologic 
examination 
NA 
 
Corneal specular microscopy 
NA 
 
Concomitant medication/therapy 
X 
Review concomitant medications and therapy; 
document in source  
Clinical laboratory tests 
(hematology, chemistry, urinalysis)  
NA 
Every effort should be made to have the subject 
return to the site to obtain clinical laboratory 
tests at Visit 3 
Serum/urine pregnancy test 
NA 
 
Urine drug screen and breathalyzer 
alcohol screen 
NA 
Review recreational drug use and alcohol 
consumption document; document in source 
 
 
 
 
CGI-S  
X 
Rater to assess over the telephone 
CGI-I  
X 
Rater to assess over the telephone 
C-SSRS 
X 
Rater to assess over the telephone 
HAMD-17 
X 
Rater to assess over the telephone 
HAM-A  
X 
Rater to assess over the telephone 
 
 
 
SDS 
X 
Ensure the subject has access to a PDF copy of 
the scale. Rater to enter subject scores. 
Following this assessment, the rater should 
review the Work/School and Days Lost items to 
confirm appropriateness of the selected ratings 
by evaluating the impact of MDD symptoms 
across those sectors. 
HADS 
X 
Rater to enter subject scores 
SHAPS 
X 
Rater to enter subject scores 
 
 
 
 
 
 
 
 
 
COWS 
NA 
 
Adverse events 
X 
Investigator to assess; in capturing AEs site 
staff should be alert to symptoms that require 
further enquiry to determine if there is any 
possibility of opioid withdrawal. 
Complete IWRS 
X 
To be done after the subject has confirmed 
inability to attend the visit in person, then the 
study drug will be sent to the subject’s home 
Dispense study drug 
X 
Dispense as per site specific policy 
Study drug compliance 
X 
Discuss compliance with subject; pill count to 
be performed at time of the return 
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Clinical Study Protocol Amendment 7 
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AE = adverse event; CGI-I = Clinical Global Impression of Improvement; CGI-S = Clinical Global Impression of Severity; 
COWS = Clinical Opiate Withdrawal Scale; C-SSRS = Columbia Suicide Severity Rating Scale; ECG = electrocardiogram; 
HADS = Hospital Anxiety and Depression Scale; HAM-A = Hamilton Anxiety Rating Scale; HAMD-17 = Hamilton Rating 
Scale for Depression - 17-Item Version; IWRS = Interactive Web-response System; MDD = major depressive disorder; 
NA = not applicable; PDF = portable document format; 
 
; SDS = Sheehan Disability Scale; SHAPS = Snaith-Hamilton Pleasure Scale; 
. 
 
Dispensing Study Drug 
Subjects may choose to pick up the study drug from the site if that option is available at the 
site. If subjects are not able to come to the site, the IWRS will be completed in a timely 
manner to ensure that the subject gets the next bottle of study drug, which will be couriered 
to the subject’s home prior to the current supply running out. Delivery of the study drug will 
be tracked by the courier and documentation will be provided to the site. The subject will 
retain their current bottle of medication and any unused supplies, and the prior and new bottle 
will be returned at their next clinic visit. The courier documentation will be maintained in the 
pharmacy file to track the distribution of the product, and the subject will be instructed to 
confirm the receipt of the study drug. This will be documented in the source documents for 
this subject. 
If a subject is required, or is predicted to need, to miss more than 1 sequential clinic 
visit, they will be instructed to discontinue study drug and asked to return to the clinic 
for safety evaluation when appropriate and safe to do so. 
End of Study Visit 
At the early termination visit (see Section 7.8), End of Treatment assessments (see 
Section 7.6) will be conducted at the clinic. The reason for termination will be documented in 
EDC and, if appropriate, note that termination was due to the COVID-19 situation (see 
Section 4.4). 
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The listed 
 below are not exhaustive. See also: 
http://www.fda.gov/drugs/developmentapprovalprocess/developmentresources/druginteractio
nslabeling/ucm093664.htm. 
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Table A2-1: 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
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SAP_001.pdf: 
BlackThorn Therapeutics, Inc. 
Statistical Analysis Plan Addendum
K2-MDD-201 
 
Version 1.0; 06 October 2022
 
Confidential                                                           
Page 1 of 9 
 
 
 
 
STATISTICAL ANALYSIS PLAN ADDENDUM 
BlackThorn Therapeutics, Inc. 
K2-MDD-201 
 
 
 
 
Protocol Title: 
A Phase 2a, Randomized, Double-blind, Placebo-controlled 
Proof of Concept Study to Evaluate the Effects of Oral BTRX-
335140 Versus Placebo in Subjects With Major Depressive 
Disorder 
Protocol Version 
and Date: 
Amendment 7 Version 8.0; 24 January 2022 
Sponsor: 
BlackThorn Therapeutics, Inc. 
450 Arsenal Way, Suite 200 
Watertown, MA 02472 
Prepared By: 
 
Precision for Medicine 
6005 Hidden Valley Road, Suite 170 
Carlsbad, CA 92011 
Document Version 
and Date: 
Version 1.0; 06 October 2022 
 






BlackThorn Therapeutics, Inc. 
Statistical Analysis Plan Addendum
K2-MDD-201 
 
Version 1.0; 06 October 2022
 
Confidential                                                           
Page 4 of 9 
2 TABLE OF CONTENTS 
 
1 
Statistical Analysis Plan Addendum Approval ........................................................2 
2 
Table of Contents ........................................................................................................4 
3 
List of Abbreviations ..................................................................................................6 
4 
Introduction.................................................................................................................7 
5 
Statistical Methods ......................................................................................................7 
5.1 
General Methodology ............................................................................................7 
5.1.1 
Analysis Visit Window .....................................................................................7 
5.1.2 
Baseline Definition ...........................................................................................7 
5.2 
Efficacy Evaluation ................................................................................................8 
5.2.1 
Measurements of Treatment Compliance .....................................................8 
5.2.2 
Sensitivity Analyses of the Primary Efficacy Endpoint................................8 
5.2.3 
Subgroup Analyses of the Primary Efficacy Endpoint ................................8 
5.2.4 
Subgroup Analyses of the Secondary Efficacy Endpoints ...........................9 
5.3 
Safety Evaluation ...................................................................................................9 
5.3.1 
Vital Signs, Physical Findings, and Other Observations Related to 
Safety .................................................................................................................9 
 
 
 


BlackThorn Therapeutics, Inc. 
Statistical Analysis Plan Addendum
K2-MDD-201 
 
Version 1.0; 06 October 2022
 
Confidential                                                           
Page 5 of 9 
TABLE OF TABLES 
Table 1 
List of Abbreviations .................................................................................6 
Table 2 
Analysis Windows for Standard Ophthalmologic Examination by 
Visit..............................................................................................................7 
 
 


BlackThorn Therapeutics, Inc. 
Statistical Analysis Plan Addendum
K2-MDD-201 
 
Version 1.0; 06 October 2022
 
Confidential 
                                           Page 6 of 9 
 
3 LIST OF ABBREVIATIONS  
Table 1 
List of Abbreviations  
Abbreviation 
Definition  
ANCOVA 
analysis of covariance 
CGI-I 
Clinical Global Impression of Improvement 
CGI-S 
Clinical Global Impression of Severity 
ET 
end of treatment 
eCRF 
electronic case report form 
HADS 
Hospital Anxiety and Depression Scale 
HAM-A 
Hamilton Anxiety Rating Scale 
 
 
HAMD-17 
Hamilton Depression Rating Scale - 17-item Version 
LOCF 
last observation carried forward 
MDD 
major depressive disorder 
MMRM 
mixed-models repeated measures 
 
 
SAP 
statistical analysis plan 
SDS 
Sheehan Disability Scale 
SHAPS 
Snaith-Hamilton Pleasure Scale 
 
 
 
 
 


BlackThorn Therapeutics, Inc. 
Statistical Analysis Plan Addendum
K2-MDD-201 
 
Version 1.0; 06 October 2022
 
Confidential 
                                           Page 7 of 9 
 
4 INTRODUCTION 
 
Detailed descriptions of the methods and presentation of data analyses proposed for 
BlackThorn Therapeutics, Inc. Protocol K2-MDD-201 (A Phase 2a, Randomized, 
Double-blind, Placebo-controlled Proof of Concept Study to Evaluate the Effects of Oral 
BTRX-335140 Versus Placebo in Subjects With Major Depressive Disorder) 
Amendment 7 Version 8.0 (24 January 2022) are described in the original statistical 
analysis plan (SAP), version 2.0, 03 June 2022 (hereafter referred to as “the main SAP”).  
 
The purpose of this addendum is to provide clarifications to the definitions and 
methodology related to the analyses described in the main SAP.  
5 STATISTICAL METHODS 
5.1 
General Methodology 
5.1.1 
Analysis Visit Window 
For efficacy and safety analyses by visit, assessments at scheduled and unscheduled visits 
will be mapped to the appropriate analysis window as detailed in Table 3 and Table 4 of 
the main SAP. The analysis visit windows for the standard ophthalmologic examination 
at Visit 3 (Week 2) and Visit 4 (Week 4) from Table 4 of the main SAP will instead be 
mapped as shown in Table 2. 
Table 2 
Analysis Windows for Standard Ophthalmologic Examination by 
Visit 
Period 
Screening 
Baseline 
Study Drug Treatment 
Safety Follow-
up 
Visit 
1 
2 
3 
4 
5 
6 
 
 
 
Days –7 to 
–28 
Day 1 
Day 15 
(±2 d) 
Day 29 
(±2 d) 
Day 43 
(±2 d) 
Day 57  
(±2 d) 
Day  
71 
Day 
85 
Analysis Visit 
Name 
 
Baseline 
Week 
2 
Week  
4 
Week 
6 
Week  
8 
Week 
10 
Week 
12 
Standard 
ophthalmologic 
examination 
NA 
≤ 1 Pre-
dose 
[2, 14] 
NA 
NA 
≥ 15 
 
NA 
 
NA 
5.1.2 
Baseline Definition 
For efficacy and safety analyses presented by study visit, the main SAP defines the 
baseline value as “the last measurement prior to the start of study drug administration”. 
The baseline value will differ for summaries of Hamilton Depression Rating Scale - 17-
item Version (HAMD-17), Clinical Global Impression of Improvement (CGI-I), Snaith-
Hamilton Pleasure Scale (SHAPS), Hospital Anxiety and Depression Scale (HADS), 
Hamilton Anxiety Rating Scale (HAM-A), Sheehan Disability Scale (SDS), Clinical 
Global Impression of Severity (CGI-S), 
 
 and 


BlackThorn Therapeutics, Inc. 
Statistical Analysis Plan Addendum
K2-MDD-201 
 
Version 1.0; 06 October 2022
 
Confidential 
                                           Page 8 of 9 
 
presented by study visit, in which case the baseline value will be 
defined as the last measurement prior to or within 1 hour of the first administration of 
study drug. For all other efficacy and safety analyses presented by study visit, the 
baseline value will be defined as stated in the main SAP. 
5.2 
Efficacy Evaluation 
5.2.1 
Measurements of Treatment Compliance 
As stated in the main SAP, percentage compliance to the study treatment regimen will be 
determined as the total dose actually taken divided by the expected dose received, 
multiplied by 100. The expected dose received will be calculated as (the date of last dose 
of study drug – the date of first dose of study drug + 1 day) x 1 capsule/day. If the date of 
last dose is missing or unknown, the date of last dose will instead be equal to the date of 
last available visit prior to Visit 7 or the date of study completion/termination, whichever 
is earlier. The date of the last available visit prior to Visit 7 is defined as the date of the 
last available visit between Visit 2 (Baseline) and Visit 6 (Week 8)/ET (end of treatment). 
Any bottles dispensed on or after the date of last dose will be excluded from the 
calculation of total dose actually taken. 
In the case of missing values, the following calculations will differ from the main SAP. If 
the number of capsules taken is missing, but the corresponding number of capsules 
dispensed and returned are not missing, then the number of capsules taken will be 
calculated as the number of capsules dispensed minus the number of capsules returned. If 
both the number of capsules returned and the number of capsules taken are missing, then 
it will be assumed that 0 capsules were returned and all capsules dispensed were taken. In 
addition, if the number of capsules dispensed is 0, then the corresponding number of 
capsules taken and returned will be set to 0.  
Dosing compliance will be summarized using descriptive statistics, by treatment group, 
based on the Efficacy Population. As an exception rule, the compliance rate for subjects 
with ≤ 14 days exposure duration will not be calculated, as the compliance may not be 
calculated reliably for such short duration. 
5.2.2 
Sensitivity Analyses of the Primary Efficacy Endpoint 
The main SAP describes the sensitivity analyses that will be performed. The Last 
Observation Carried Forward (LOCF) Imputation technique will be performed as 
follows: the last observed non-missing post-baseline value will be used to fill in missing 
values at Week 8, regardless of when the missing value occurred. The imputed dataset 
generated with the LOCF technique will be used as input data in the analysis of 
covariance (ANCOVA) analysis.  
5.2.3 
Subgroup Analyses of the Primary Efficacy Endpoint 
The primary efficacy endpoint (change from baseline to Week 8 in HAMD-17 total 
score) will be summarized using the subgroups described in the main SAP. Additionally, 


BlackThorn Therapeutics, Inc. 
Statistical Analysis Plan Addendum
K2-MDD-201 
 
Version 1.0; 06 October 2022
 
Confidential 
                                           Page 9 of 9 
 
the mixed-models repeated measures (MMRM) analysis described for the primary 
efficacy endpoint in the main SAP will be repeated for all subgroups.  
5.2.4 
Subgroup Analyses of the Secondary Efficacy Endpoints 
The secondary efficacy endpoint SHAPS will be summarized using the subgroups 
described in the main SAP. HAM-A total score will instead be presented by baseline 
SHAPS Total Score (14 to 56) (< median and ≥ median) and SHAPS Original Total 
Score (0 to 14) (< median and ≥ median). 
5.3 
Safety Evaluation 
5.3.1 
Vital Signs, Physical Findings, and Other Observations Related to Safety 
5.3.1.1 
Ophthalmology Examination 
The main SAP states “Standard ophthalmologic examination and corneal specular 
microscopy examination results will be summarized by presenting shift from baseline to 
end of study results as normal, abnormal not clinically significant, and abnormal 
clinically significant.” This summary will be presented by visit.   
Additionally, the subjects who have endothelial dystrophy medical history prior to 
treatment used for the subgroup analysis will be identified by having a preferred term of 
“Corneal Dystrophy” collected on the Medical and Psychiatric History Other Than MDD 
(major depressive disorder) electronic case report form (eCRF) prior to date of first dose 
of study drug. 
 
 
 






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BlackThorn Therapeutics, Inc. 
Statistical Analysis Plan
K2-MDD-201 
 
Version 2.0; 03 June 2022 
 
Confidential                                                           
Page 1 of 45 
 
 
 
 
STATISTICAL ANALYSIS PLAN 
BlackThorn Therapeutics, Inc. 
K2-MDD-201 
 
 
 
 
Protocol Title: 
A Phase 2a, Randomized, Double-blind, Placebo-controlled 
Proof of Concept Study to Evaluate the Effects of Oral BTRX-
335140 Versus Placebo in Subjects With Major Depressive 
Disorder 
Protocol Version 
and Date: 
Amendment 7 Version 8.0; 24 January 2022 
Sponsor: 
BlackThorn Therapeutics, Inc. 
450 Arsenal Way, Suite 200 
Watertown, MA 02472 
Prepared By: 
 
Precision for Medicine 
6005 Hidden Valley Road, Suite 170 
Carlsbad, CA 92011 
Document Version 
and Date: 
Version 2.0; 03 June 2022 
 






BlackThorn Therapeutics, Inc. 
Statistical Analysis Plan
K2-MDD-201 
 
Version 2.0; 03 June 2022 
 
Confidential                                                           
Page 4 of 45 
2 TABLE OF CONTENTS 
 
1 
Statistical Analysis Plan Approval ............................................................................2 
2 
Table of Contents ........................................................................................................4 
3 
List of Abbreviations ..................................................................................................7 
4 
Introduction...............................................................................................................10 
5 
Study Objectives .......................................................................................................10 
5.1 
Primary Study Objective .....................................................................................10 
5.2 
Secondary Study Objectives................................................................................10 
5.3 
 
6 
Investigational Plan ..................................................................................................11 
6.1 
Overall Study Design ...........................................................................................11 
6.2 
Schedule of Assessments ......................................................................................12 
6.3 
Treatments ............................................................................................................12 
6.3.1 
Treatments Administered .............................................................................12 
6.3.2 
Method of Assigning Subjects to Treatment Groups .................................12 
6.4 
Efficacy and Safety Variables .............................................................................12 
6.4.1 
Efficacy Variables ..........................................................................................12 
6.4.2 
Safety Variables .............................................................................................15 
6.4.3 
Pharmacokinetic Variable.............................................................................21 
6.5 
Data Quality Assurance .......................................................................................21 
7 
Statistical Methods ....................................................................................................21 
7.1 
General Methodology ..........................................................................................21 
7.1.1 
Reporting Conventions ..................................................................................22 
7.1.2 
Analysis Visit Window ...................................................................................22 
7.1.3 
Data Handling Rules ......................................................................................24 
7.1.4 
Standard Calculations ...................................................................................24 
7.1.5 
Baseline Definition .........................................................................................26 
7.1.6 
Multiple Comparisons/Multiplicity ..............................................................26 
7.2 
Analysis Populations ............................................................................................26 
7.3 
Study Subjects ......................................................................................................27 
7.3.1 
Disposition of Subjects ...................................................................................27 
7.3.2 
Important Protocol Deviations .....................................................................27 
7.4 
Efficacy Evaluation ..............................................................................................27 
7.4.1 
Datasets Analyzed ..........................................................................................27 
7.4.2 
Demographic and Other Baseline Characteristics ......................................27 
7.4.3 
Measurements of Treatment Compliance ...................................................28 
7.4.4 
Primary Efficacy Endpoint Analysis Methods ............................................29 
7.4.5 
Sensitivity Analyses of the Primary Efficacy Endpoint..............................30 
7.4.6 
Subgroup Analyses of the Primary Efficacy Endpoint ..............................30 
7.4.7 
Secondary Endpoint Analysis Methods .......................................................31 
7.4.8 
Subgroup Analyses of the Secondary Efficacy Endpoints .........................32 
7.4.9 
 
7.4.10 
Interim Analyses and Data Monitoring .......................................................34 
7.4.11 
Additional supportive analyses to assess impact of COVID-19 .................34 


BlackThorn Therapeutics, Inc. 
Statistical Analysis Plan
K2-MDD-201 
 
Version 2.0; 03 June 2022 
 
Confidential                                                           
Page 5 of 45 
7.4.12 
Plasma Concentrations ..................................................................................34 
7.5 
Safety Evaluation .................................................................................................34 
7.5.1 
Adverse Events ...............................................................................................35 
7.5.2 
Extent of Exposure .........................................................................................36 
7.5.3 
Deaths, Other Serious Adverse Events, and Other Significant 
Adverse Events ...............................................................................................36 
7.5.4 
Clinical Laboratory Evaluation ....................................................................36 
7.5.5 
Vital Signs, Physical Findings, and Other Observations Related to 
Safety ...............................................................................................................37 
7.6 
 
7.7 
Changes in the Conduct of the Study or Planned Analyses .............................41 
8 
Reference List ............................................................................................................41 
Appendix A: Schedule of Procedures and Assessments ...............................................42 
Appendix B: Steps for Multiple Imputation ..................................................................44 
 
 
 


BlackThorn Therapeutics, Inc. 
Statistical Analysis Plan
K2-MDD-201 
 
Version 2.0; 03 June 2022 
 
Confidential                                                           
Page 6 of 45 
TABLE OF TABLES 
Table 1 
List of Abbreviations .................................................................................7 
Table 2 
Clinical Laboratory Tests........................................................................18 
Table 3 
Analysis Windows for Efficacy Analyses ...............................................23 
Table 4 
Analysis Windows for Safety Analyses by Visit ....................................24 
Table 5 
Potentially Clinically Significant Laboratory Results ..........................37 
 
 


BlackThorn Therapeutics, Inc. 
Statistical Analysis Plan 
K2-MDD-201 
 
Version 2.0; 03 June 2022 
 
Confidential 
                                           Page 7 of 45 
 
3 LIST OF ABBREVIATIONS  
Table 1 
List of Abbreviations  
Abbreviation 
Definition  
Ab 
antibody 
AE 
adverse event 
AESI 
adverse event of special interest 
ALT/SGPT 
alanine aminotransferase 
ANC 
absolute neutrophil count 
ANCOVA 
analysis of covariance 
AST/SGOT 
aspartate aminotransferase 
ATC 
Anatomical Therapeutic Chemical 
BMI 
body mass index 
BP 
blood pressure 
BUN 
blood urea nitrogen 
C-SSRS 
Columbia Suicide Severity Rating Scale 
CGI-I 
Clinical Global Impression of Improvement 
CGI-S 
Clinical Global Impression of Severity 
CI 
confidence interval 
CK 
creatine kinase 
CPK 
creatine phosphokinase 
COWS 
Clinical Opiate Withdrawal Scale 
COVID-19 
coronavirus disease 2019  
CSM 
Corneal Specular Microscopy 
CSR 
clinical study report 
CV 
coefficient of variation 
DSM-5 
Diagnostic and Statistical Manual of Mental Disorders Fifth Edition 
ECG 
electrocardiogram 
eCRF 
electronic case report form 
GGT 
gamma-glutamyltransferase 
GLMM 
generalized linear mixed model 
HADS 
Hospital Anxiety and Depression Scale 
HADS-A 
Hospital Anxiety and Depression Scale - Anxiety Subscale 
HADS-D 
Hospital Anxiety and Depression Scale - Depression Subscale 
HAM-A 
Hamilton Anxiety Rating Scale 
HAMD-17 
Hamilton Depression Rating Scale - 17-item Version 
HAV-Ab 
hepatitis A virus antibody 
HBc Ab 
hepatitis B core antibody 


BlackThorn Therapeutics, Inc. 
Statistical Analysis Plan 
K2-MDD-201 
 
Version 2.0; 03 June 2022 
 
Confidential 
                                           Page 8 of 45 
 
Abbreviation 
Definition  
HBs Ag 
hepatitis B surface antigen 
HC Ab 
hepatitis C antibody 
HIV 
human immunodeficiency virus 
HPMC 
hydroxypropyl methylcellulose 
ICF 
informed consent form 
ICH 
International Council for Harmonisation 
IgM 
immunoglobulin M 
INR 
International normalized ratio 
IWRS 
interactive web-response system 
KORA 
kappa opioid receptor antagonist 
LOCF 
last observation carried forward 
LS 
least-square 
MCV 
mean cell volume 
MCHC 
mean cell hemoglobin concentration 
MDD 
major depressive disorder 
MedDRA 
Medical Dictionary for Regulatory Activities 
MI 
multiple imputation 
MMRM 
mixed-models repeated measures 
OD 
oculus dextrus 
OS 
oculus sinister 
PC 
pre COVID 
PCS 
potentially clinically significant 
 
POC 
proof of concept 
PP 
Per-Protocol 
QD 
once daily 
QTcB 
corrected QT interval using Bazett’s correction 
QTcF 
corrected QT interval using Fridericia’s correction 
RBC 
red blood cells 
SAE 
serious adverse event 
SAP 
statistical analysis plan 
SD 
standard deviation 
SDS 
Sheehan Disability Scale 


BlackThorn Therapeutics, Inc. 
Statistical Analysis Plan 
K2-MDD-201 
 
Version 2.0; 03 June 2022 
 
Confidential 
                                           Page 9 of 45 
 
Abbreviation 
Definition  
SE 
standard error 
SHAPS 
Snaith-Hamilton Pleasure Scale 
SI 
Système International 
SIGH-A 
Structured Interview Guide for the Hamilton Anxiety Rating Scale 
SIGH-D 
Structured Interview Guide for the Hamilton Depression Rating 
Scale 
SOC 
system organ class 
TEAE 
treatment-emergent adverse event 
TESAE 
treatment-emergent serious adverse event 
THC 
delta-9-tetrahydrocannabinol 
TSH 
thyroid stimulating hormone 
ULN 
upper limit of normal 
WBC 
white blood cell 
WHODrug 
World Health Organization Drug classification   
 
 
 






BlackThorn Therapeutics, Inc. 
Statistical Analysis Plan 
K2-MDD-201 
 
Version 2.0; 03 June 2022 
 
Confidential 
                                           Page 12 of 45 
 
6.2 
Schedule of Assessments 
For the complete schedule of assessments, refer to Appendix A: Schedule of Procedures 
and Assessments. 
6.3 
Treatments 
6.3.1 
Treatments Administered 
Treatment is initiated at 1 capsule of study drug (80mg BTRX-335140 or placebo) QD 
and this dose is maintained for all 8 weeks of double-blind treatment. Subjects are 
instructed to swallow capsules whole and not crush or chew and to store the capsules in 
their original packaging at the temperature listed on the package label. Subjects are 
instructed to take their first dose of study drug under the supervision of clinical study 
personnel during the baseline visit and to try to take their study medication at 
approximately the same time each day, preferably in the morning with food. 
6.3.2 
Method of Assigning Subjects to Treatment Groups 
Subjects who meet all inclusion criteria and are not excluded by the exclusion criteria are 
randomized to double-blind treatment at Visit 2 (Baseline) using an interactive web-
response system (IWRS). Subjects are randomized in a 1:1 ratio to 1 of 2 treatment arms 
(placebo or BTRX-335140 QD).  
The IWRS is used to assign bottles containing double-blind study drug or placebo to each 
subject. Further details are provided in the Pharmacy manual. 
6.4 
Efficacy and Safety Variables 
6.4.1 
Efficacy Variables 
6.4.1.1 
Primary Efficacy Variable 
The primary efficacy endpoint is the change from baseline to Week 8 on the HAMD-17 
score in the Efficacy population.  
This instrument is completed based on a semi-structured interview (Structured Interview 
Guide for the Hamilton Depression Rating Scale [SIGH-D], Williams 1988). The 17-item 
HAM-D comprises individual ratings related to the following symptoms: depressed mood 
(sadness, hopeless, helpless, worthless), feelings of guilt, suicide, insomnia (early, 
middle, late), work and activities, retardation (slowness of thought and speech; impaired 
ability to concentrate; decreased motor activity), agitation, anxiety (psychic and somatic), 
somatic symptoms (gastrointestinal and general), genital symptoms, hypochondriasis, 
loss of weight, and insight. The HAMD-17 scoring ranges from 0 to 52 with individual 
items ranging from 0-4 points and 0-2 points where higher scores corresponding to higher 
levels of symptom severity. 


BlackThorn Therapeutics, Inc. 
Statistical Analysis Plan 
K2-MDD-201 
 
Version 2.0; 03 June 2022 
 
Confidential 
                                           Page 13 of 45 
 
6.4.1.2 
Secondary Efficacy Variables 
Secondary efficacy endpoints include the following: 
x HAMD-17 response rate at Week 4 and Week 8. The response rate is the 
percentage of subjects with ≥ 50% decrease from baseline in HAMD-17 score.  
x CGI-I. CGI-I score assessed at each postbaseline timepoint compared to baseline. 
The CGI-I scale (Guy 1976) is a clinician-rated instrument that measures the 
improvement of the subject’s symptoms. It is a 7-point scale where a score of 1 
indicates that the subject is “very much improved,” a score of 4 indicates that the 
subject has experienced “no change,” and a score of 7 indicates that the subject is 
“very much worse.” For analysis purposes, a CGI-I response indicating symptom 
improvement will be defined as “much improved” or “very much improved”. 
x SHAPS. Change from baseline to each timepoint assessed in SHAPS scores. The 
SHAPS is a 14-item self-report instrument which measures anhedonia (Snaith et 
al 1995). Each of the 14 items has a set of 4 responses, 2 of which endorse 
agreement (Definitely/Strongly Agree, Agree) and 2 of which endorse 
disagreement (Disagree, Strongly Disagree). There are two scoring methods.  
o SHAPS Original Total Score (0 to 14): The original method assigned a 
score of 1 to either of the disagreement responses (Disagree, Strongly 
Disagree) and a score of 0 to either of the agreement responses 
(Definitely/Strongly Agree, Agree). Therefore, original scores on the 
SHAPS can range from 0-14 with higher scores corresponding to higher 
levels of anhedonia.  
o SHAPS Total Score (14 to 56): The second method derives a total score 
by summing the item responses where Definitely/Strongly Agree 
responses score 1 point, Agree responses score 2 points, Disagree 
responses score 3 points, and Strongly Disagree responses score 4 points. 
Therefore, scores on the SHAPS can range from 14 to 56 by the second 
method, with higher scores corresponding to higher levels of anhedonia. 
Both methods will be used for analyses. 
x HADS. Change from baseline to each timepoint assessed in HADS total score and 
HADS subscales (ie, anxiety subscale [HADS-A] and depression subscale 
[HADS-D] scores). The HADS measures levels of anxiety and depression without 
regards to somatic symptoms (Zigmond and Snaith 1983). This self-report scale 
consists of 14 items. Seven of the items are used to evaluate anxiety and 7 
evaluate depression. Each item on the questionnaire is scored from 0 to 3. 
Therefore, the anxiety subscale (HADS-A) and the depression subscale (HADS-
D) each range from 0 to 21.  
x HAM-A. Change from baseline to each timepoint assessed in HAM-A total score. 
This instrument will be completed based on a semi-structured interview 






BlackThorn Therapeutics, Inc. 
Statistical Analysis Plan 
K2-MDD-201 
 
Version 2.0; 03 June 2022 
 
Confidential 
                                           Page 16 of 45 
 
becomes stable or can be explained by another known cause (ie, concurrent condition or 
medication). 
Medical conditions present or AEs occurring prior to the first dose of study drug will be 
captured as medical history in the eCRF. Any AE occurring during the study that is 
related to a pre-existing condition or event that worsens in intensity or frequency after the 
first dose of study drug will be recorded as a TEAE.  
The investigator will assess the severity of each AE based on his/her clinical judgment 
using one of the following categories: 
Mild: Usually transient and may require only minimal treatment or therapeutic 
intervention. The event does not generally interfere with usual activities of daily 
living.  
Moderate: Usually alleviated with additional specific therapeutic intervention. The event 
interferes with usual activities of daily living, causing discomfort but poses no 
significant or permanent risk of harm to the subject. 
Severe: Interrupts usual activities of daily living, or significantly affects clinical status, or 
may require intensive therapeutic intervention. 
The investigator will assess the relationship (ie, causality) of each AE to study drug based 
on his/her clinical judgment. Relationship to study drug will be assessed according to the 
following guidelines: 
Possibly related: The AE is known to occur with the study drug, there is a reasonable 
possibility that the study drug caused the AE, or there is a temporal relationship 
between the study drug and event. Reasonable possibility means that there is evidence 
to suggest a causal relationship between the study drug and the AE. 
Unlikely related: There is not a reasonable possibility that the administration of the 
study drug caused the event, there is no temporal relationship between the study drug 
and event onset, or an alternate etiology has been established. 
6.4.2.2 
Serious Adverse Events 
An SAE is any AE that results in 1 of the following outcomes, regardless of the 
investigator’s opinion of causation: 
x Death 
x Initial or prolonged inpatient hospitalization 
o Surgeries planned prior to signing the ICF will not be considered SAEs. 
However, worsening of the underlying medical condition during the study 
will be considered an AE and must be captured as serious if any SAE-
defining outcomes occur as a result. 


BlackThorn Therapeutics, Inc. 
Statistical Analysis Plan 
K2-MDD-201 
 
Version 2.0; 03 June 2022 
 
Confidential 
                                           Page 17 of 45 
 
x A life-threatening experience (that is, immediate risk of dying) 
x Persistent or significant disability/incapacity 
x Congenital anomaly/birth defect 
x Other medically important serious event as determined by the investigator (for 
example, an AE that jeopardizes the subject and may require medical or surgical 
intervention to prevent one of the outcomes listed in this definition) 
The following events do not meet the definition of an SAE:  
x Hospitalization for elective treatment of a pre-existing condition that does not 
worsen from baseline 
x Hospitalizations for a standard procedure for study drug administration and 
routine monitoring of the studied indication not associated with any deterioration 
in condition 
x Social or convenience admission to a hospital 
x Prolongation of a hospitalization for social or convenience reasons not associated 
with the occurrence of an AE 
x Hospitalization or an emergency room visit that lasts less than 24 hours that does 
not meet the criteria of an important medical or a life-threatening event 
6.4.2.3 
Adverse Events of Special Interest 
AESIs (serious or nonserious) are TEAEs of particular safety importance as identified by 
the Sponsor. Reported TEAEs that are classified in the system organ classes (SOCs) 
listed below will be considered AESIs: 
x Skin and subcutaneous tissue disorders 
x Eye disorders 
Nonserious AEs in the above SOCs that occur prior to study drug dosing will be recorded 
in medical history. 
6.4.2.4 
Laboratory Parameters 
A central laboratory will be used for safety laboratory tests. Standard clinical laboratory 
tests are performed at times specified in the Schedule of Procedures and Assessments 
section of the clinical study protocol. Repeat testing of samples may be permitted during 
screening, after review and approval by the medical monitor. 


BlackThorn Therapeutics, Inc. 
Statistical Analysis Plan 
K2-MDD-201 
 
Version 2.0; 03 June 2022 
 
Confidential 
                                           Page 18 of 45 
 
Abnormal laboratory values are graded by the investigator as “clinically significant” or 
“not clinically significant,” where applicable. Clinically significant abnormal laboratory 
values will be reported as AEs. Investigators may repeat laboratory tests for any 
parameter that is abnormal and/or clinically significant. 
Table 2 lists the specific laboratory analyses that will be performed for this study. 
Table 2 
Clinical Laboratory Tests 
Hematologya:  
Hemoglobin 
Hematocrit  
Erythrocyte count (red blood cell [RBC])  
Mean cell volume (MCV)  
Mean cell hemoglobin concentration (MCHC)  
Leukocytes (white blood cell [WBC])  
Neutrophils, segmented 
Absolute Neutrophil Count (ANC) 
Lymphocytes  
Monocytes 
Eosinophils 
Basophils 
Platelets 
Prothrombin timeb 
Chemistrya: Serum concentrations of: 
Sodium 
Potassium  
Bicarbonate  
Chloride 
Total bilirubin 
Direct bilirubin 
Alkaline phosphatase 
Alanine aminotransferase/serum glutamic pyruvic 
transaminase (ALT/SGPT) 
Aspartate aminotransferase/serum glutamic 
oxaloacetic transaminase (AST/SGOT) 
Gamma-glutamyl transferase (GGT) 
Blood urea nitrogen (BUN) 
Creatinine 
Uric acid 
Phosphorous 
Calcium 
Glucose (random) 
Albumin 
Total cholesterol 
Creatine kinase (CK)c 
Magnesium 
Amylase 
Lipase 
Urinalysisa: 
Specific gravity  
pH  
Protein  
Glucose  
Ketones  
Blood  
Urine leukocyte esterase 
Urobilinogen 
Urine Drug Screen and Breathalyzer Alcohol 
Screena 
Amphetamines 
Barbiturates 
Benzodiazepines 
Delta-9-tetrahydrocannabinol (THC) 
Cocaine 
Ethyl alcohol (via breathalyzer) 
Opioids 
Viral Serologyd 
Hepatitis B Surface Antigen (HBs Ag) 
Hepatitis B Core Antibody (HBc Ab) 
Hepatitis C Antibody (HC Ab) 
Hepatitis A Antibody (HAV-Ab [IgM]) 
Human immunodeficiency virus (HIV) 




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6.4.2.7 
Electrocardiography 
ECG is performed at Screening, Baseline (Week 0), Week 4, and Week 8. For each 
subject, single 12-lead digital ECGs are collected after the subject has rested for 5 
minutes. All digital ECGs are electronically transmitted to a designated central ECG 
laboratory. Any clinically significant finding that was present on the fully over-read ECG 
is reported to the investigator and to the Sponsor. 
The results include ventricular rate, PR interval, RR interval, QRS duration, QT interval, 
QTcB interval (corrected QT interval using Bazett’s correction), QTcF interval (corrected 
QT interval using Fridericia’s correction), interpretation (normal, abnormal) and 
evaluation. 
The corrected QT interval will be corrected for respiratory rate using Fridericia’s 
correction: QTcF = QT/RR0.33 and Bazett’s correction: QTcF = QT/RR0.5. 
6.4.2.8 
Ophthalmologic Examination 
 
The following ocular assessments are performed, using these standard clinical operating 
procedures, at specified visits, on both eyes of each subject:  
 
1. Standard Ophthalmologic Examinations: performed at Screening/Baseline, 
between post-dose days 7-15, and at end of treatment/Week 8.  
2. Corneal Specular Microscopy (CSM): noncontact or contact specular microscopy 
for qualitative and quantitative examination of the central corneal endothelium 
(cell count, cell shape, density, and morphology), with image capture at 
Screening/Baseline and 12 weeks following first dose. 
 
Local investigators assess and record the results of items 1 and 2 on the examination 
worksheet. For both the standard ophthalmologic examination and CSM, an assessment 
of normal/abnormal is captured as well as whether the abnormal result is clinically 
significant or not. CSM results are sent to 
 where the 
designated ophthalmology expert will review. Additional ophthalmologic examinations 
(standard ophthalmologic exams and/or corneal specular microscopies) may be 
conducted at any time during the study to assess subject safety. 
 
The final ocular examination for corneal specular microscopy is conducted approximately 
12 weeks (+2 days) after the first dose of study drug (including both subjects who 
complete the study and subjects who discontinue early from the study). 
6.4.2.9 
Suicidal Ideation/Suicidality 
Suicide-related events (behavior and/or ideation) are assessed and evaluated at every 
scheduled and unscheduled visit with the administration of the C-SSRS (Posner 2007). 
The C-SSRS captures the occurrence, severity, and frequency of suicide-related thoughts 
and behaviors during the assessment period. The 2009 “Baseline” version is used at Visit 


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1 (Screening), evaluating past year and past 3 months, and the 2009 “Since Last Visit” 
version is used for all subsequent visits. 
6.4.2.10 
Opiate Withdrawal 
Opiate withdrawal is assessed at screening and at all subsequent treatment visits through 
Week 8 using the COWS. The COWS is a clinician administered instrument used to rate 
11 common opiate withdrawal signs or symptoms (Wesson and Ling 2003). The total 
score of the 11 items can then be used to assess a subject's level of opiate withdrawal and 
to determine their level of physical dependence on opioids. This assessment provides an 
additional safeguard to ensure no subjects are enrolled with opiate withdrawal or for 
whom withdrawal may begin during the course of the study. Scores range from 0 to 48 
with higher scores indicating greater severity of opioid withdrawal symptoms. 
6.4.3 
 
 
 
6.5 
Data Quality Assurance 
Report summaries will be generated using validated Base SAS£ software, version 9.4 or 
higher, on a PC or server-based platform. Additional validated software may be used to 
generate analyses, as needed.  
All SAS programs that create outputs or supporting analysis datasets will be validated by 
a second statistical programmer or biostatistician. At a minimum, validation of programs 
will consist of a review of the program log, review of output or dataset format and 
structure, and independent confirmatory programming to verify output results or dataset 
content. Additionally, all outputs will undergo a review by a senior level team member 
before finalization. 
The content of the source data will be reviewed on an ongoing basis by project statistical 
programmers and statisticians. Data will be checked for missing values, invalid records, 
and extreme outliers through defensive programming applications, analysis-based edit 
checks, and other programmatic testing procedures. All findings will be forwarded to the 
project data manager for appropriate action and resolution.  
7 STATISTICAL METHODS 
7.1 
General Methodology 
Data will be analyzed by Precision for Medicine biostatistics personnel. Statistical 
analyses will be reported with tables, figures, and listings, presented in rich text format, 
and using recommended ICH numbering. Output specifications for all tables, figures, and 
listings will be in conformance with guidelines specified by the ICH in Appendix 7 of the 
Electronic Common Technical Document Specification (Apr 2003).  


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7.1.1 
Reporting Conventions 
Tables and figures will be summarized by treatment group. Tables summarizing 
demographics and other baseline characteristics will also include a column for all 
subjects combined. In general, all data collected and any derived data will be presented in 
subject data listings for all randomized subjects except for the screening failures listing 
which will be presented for only screen failures and the informed consent and eligibility 
criteria listing which will be presented for all screened subjects. Listings will be ordered 
by treatment group, subject number, and assessment or event date. The treatment groups 
will be displayed in the same order as they appeared in the summary tables. The 
treatment group presented in safety listings will be based on the actual assignment. The 
treatment group presented in all other listings will be based on the planned assignment, 
unless otherwise noted. 
In general, continuous variables will be summarized to indicate the study population 
sample size (N), number of subjects with available data (n), mean, SD, median, 25th (Q1) 
and 75th (Q3) quartiles, minimum, and maximum values. Categorical variables will be 
summarized by the population size (N), number of subjects with available data (n), 
number of subjects in each category, and the percentage of subjects in each category. 
Unless otherwise noted, the denominator to determine the percentage of subjects in each 
category will be based on the number of subjects with available data. The denominator 
for percentages for incidence data (such as adverse events) will be based on the number 
of subjects in the analysis population. Select ordinal data may be summarized using both 
descriptive statistics and counts and percentages of subjects in each category, as 
appropriate. 
7.1.2 
Analysis Visit Window 
For efficacy analyses and safety analysis by visit, assessments at scheduled and 
unscheduled visits will be mapped to the appropriate analysis window as detailed in 
Table 3 and Table 4. In the event where more than one visit falls in the same analysis 
window, the following rules will be used in sequence to determine the record that will be 
analyzed: 
x If there is a scheduled visit in the analysis window, then the scheduled visit’s data 
will be used. 
x If there is no scheduled visit in the analysis visit window, the data closest to the 
scheduled day/time visit will be used. 
x If there is no scheduled visit in the analysis visit window and there is a tie 
between the visits with regards to the number of days/hours before and after the 
scheduled day, the later data will be used. 
Visit windows will not apply to listings. All data will be included in subject listings. 
Windowing will be applied to the data prior to any missing data calculations. 
 


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Table 3 
Analysis Windows for Efficacy Analyses 
Period 
Screening 
Baseline 
Study Drug Treatment 
Safety Follow-up 
Visit 
1 
2 
3 
4 
5 
6 
 
 
 
 
Days –7 
to –28 
Day 1 
Day 15 
(±2 d) 
Day 29 
(±2 d) 
Day 43 
(±2 d) 
Day 57  
(±2 d) 
Day  
71 
Day 
85 
Analysis 
Visit Name 
 
Baseline
* 
Week 
2 
Week  
4 
Week  
6 
Week  
8 
 
Week 
10 
 
Week 
12 
CGI-I 
HADS 
NA 
≤ 1 Pre-
dose 
[2, 22] 
[23, 36] 
[37, 50] 
[51, 71] 
 
NA 
 
NA 
HAMD-17 
HAM-A 
SHAPS 
 
NA 
≤ 1 Pre-
dose 
NA 
[16, 43]  
NA 
[44, 71]  
 
NA 
 
NA 
CGI-S 
 
NA 
≤ 1 Pre-
dose 
NA 
NA 
NA 
[44, 71] 
 
NA 
 
NA 
SDS 
NA 
≤ 1 Pre-
dose 
NA 
[23, 36] 
[37, 50] 
[51, 71]  
 
NA 
 
NA 
* Baseline for analysis visit refers to baseline value for efficacy analysis defined in Section 7.1.5 


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Table 4 
Analysis Windows for Safety Analyses by Visit 
Period 
Screening 
Baseline 
Study Drug Treatment 
Safety Follow-
up 
Visit 
1 
2 
3 
4 
5 
6 
 
 
 
 
Days –7 to 
–28 
Day 1 
Day 15 
(±2 d) 
Day 29 
(±2 d) 
Day 43 
(±2 d) 
Day 57  
(±2 d) 
Day  
71 
Day 
85 
Analysis Visit 
Name 
 
Baseline 
Week 
2 
Week  
4 
Week 
6 
Week  
8 
Week 
10 
Week 
12 
Vital signs 
Clinical 
laboratory tests 
COWS 
NA 
≤ 1 Pre-
dose 
[2, 22] 
[23, 36] 
[37, 
50] 
[51, last 
dose 
+30] 
 
NA 
 
NA 
Body weight 
ECG 
NA 
≤ 1 Pre-
dose 
NA 
[2, 43] 
NA 
[44, last 
dose 
+30] 
 
NA 
 
NA 
Standard 
Ophthalmologic 
examination 
NA 
≤ 1 Pre-
dose 
NA 
[2, 14] 
NA 
≥ 15 
 
NA 
 
NA 
Corneal specular 
microscopy 
NA 
≤ 1 Pre-
dose 
NA 
NA 
NA 
NA 
 
NA 
 
≥2 
C-SSRS 
Nominal 
 ≤ 1 Pre-
dose 
[2, 22] 
[23, 36] 
[37, 
50] 
[51, 64] 
[65, last 
dose 
+30] 
NA 
7.1.3 
Data Handling Rules 
Unless otherwise noted, values reported as greater than or less than some quantifiable 
limit (e.g., “< 1.0”) will be summarized with the sign suppressed in summary tables and 
figures, using the numeric value reported. Data will display on subject listings to include 
the sign. 
7.1.4 
Standard Calculations 
Where appropriate, the calculated study day of each assessment or event will be 
presented with the assessment or event date on subject data listings, where study day will 
be determined as: 
x The assessment/event date minus the date of first dose of study drug, if the 
assessment/event date is prior to the date of first dose; and 
x The assessment/event date minus the date of first dose of study drug, plus one, if 
the assessment/event date is on or after the date of first dose. 
Start and stop dates will be imputed when partial dates are present as needed to determine 
treatment emergent events and concomitant medications/procedures. No imputation will 


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be done for a completely missing start/stop date or for subjects who did not receive study 
treatment. 
Start dates with a missing day but which have month and year provided will be imputed 
such that: 
x If the provided month and year match the month and year for that subject’s first 
dose date, then the Day 1 date will be used 
x In all other cases the 1st of the month will be used with the provided month and 
year 
Start dates with a missing day and month but which have year populated will be imputed 
such that: 
x If the provided year matches the year for that subject’s first dose date, then the 
first dose date will be used 
x In all other cases the 1st of January will be used with the provided year 
Stop dates will be imputed as follows: 
x Missing day with a provided year and month will use the last day of the month 
x Missing day and month with provided year will use December 31 
If the imputed stop date is greater than the last study date for the subject, then the 
imputed date will be replaced with the last known subject date. 
The date of the start of the current depressive episode and date of diagnosis of MDD will 
be imputed when the month or day are missing as follows: 
x Missing day is set to 1 if the same year and month as the date of assessment. 
Otherwise it is set to 15 
x Missing month and day are set to Jan 1 if the same year as the date of assessment. 
Otherwise it is set to July 1. 
Other variables requiring calculations will be derived using the following formulas: 
x Days: A duration between two dates expressed in days will be calculated using 
the following conventions: 
o Later date – earlier date + 1, if the earlier date is on or after the date of 
first dose of study drug; or 
o Later date – earlier date, if the earlier date is prior to the date of first dose 
of study drug. 
x Months: A duration expressed in months will be calculated by dividing the 
duration in days by (365.25 / 12); 


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x Years: A duration expressed in years will be calculated by dividing the duration 
in days by 365.25; 
x Change from Baseline: Change from baseline will be calculated as the 
post-baseline value minus the baseline value; 
x Percentage Change from Baseline: Percentage change from baseline will be 
calculated as the change from baseline divided by the baseline value, multiplied 
by 100. 
7.1.5 
Baseline Definition 
For efficacy and safety analyses presented by study visit, the baseline value will be 
defined as the last measurement prior to the start of study drug administration. 
7.1.6 
Multiple Comparisons/Multiplicity 
The prespecified primary efficacy analysis will be conducted using a two-sided test at the 
α=0.05 level of significance. All other analyses will be also conducted with no 
adjustments for multiple comparisons.  
7.2 
Analysis Populations 
The analysis populations are defined as follows:  
x The Safety population is defined as all subjects who received study drug and will 
be used for summaries of safety parameters. Subjects will be analyzed according 
to treatment received. 
x The Efficacy population includes all randomized subjects who received at least 1 
dose of study drug, have a baseline HAMD-17 assessment total score, and have at 
least 1 post-baseline HAMD-17 assessment total score. Subjects will be analyzed 
according to randomized treatment assignment. This population will be used for 
the primary analysis and for all other efficacy analyses. 
x The Per-Protocol (PP) population includes subjects in the efficacy population 
except those who experienced an important protocol deviation that is determined 
to substantially impact efficacy analysis, or did not have a HAMD-17 score at 
Week 8 analysis visit, or whose compliance at the end of the study was less than 
70%. Subjects will be analyzed according to randomized treatment assignment. 
This population will be used for supportive efficacy analyses. The list of 
important protocol deviations for PP population will be classified by the sponsor 
before the final database lock and the unblinding of the study. 


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7.3 
Study Subjects 
7.3.1 
Disposition of Subjects 
Subject disposition will be summarized for all subjects by treatment group and over all 
subjects combined. Summaries will include the number and percentage of subjects 
screened, randomized, in each analysis population, completing the treatment, completing 
the study, and discontinuing the study early by the primary reason for discontinuation. 
Subject disposition will also be summarized separately for each study site. 
Subject completion status, date of study completion/discontinuation, study day of 
discontinuation, and reason for discontinuation will be listed. Inclusion and exclusion 
eligibility will be listed separately. 
7.3.2 
Important Protocol Deviations 
Important protocol deviations will be summarized by treatment group and over all 
subjects combined for the Efficacy Population. Important protocol deviations are protocol 
deviations captured on-study that are deemed by the Sponsor to potentially impact the 
efficacy or safety conclusions of the study.  
Important protocol deviation rules will be determined and appropriately categorized prior 
to database lock and prior to breaking the blind of the treatment group assignments. The 
number and percentage of subjects with any important protocol deviations as well as the 
number and percentage of subjects with deviations within each category will presented.  
The important protocol deviations will be listed by treatment group and subject. 
7.4 
Efficacy Evaluation 
7.4.1 
Datasets Analyzed 
All efficacy summaries will be produced on the Efficacy Population; select efficacy 
summaries (including HAMD-17, response rate of HAMD-17, subgroup analysis of 
HAMD-17 (<22 vs ≥ 22), SHAPS, CGI-I) will also be produced on the PP Population. A 
data listing of subjects excluded from the Efficacy Population or PP Population, to 
include the reason for exclusion, will be presented. All data listings will be listed for all 
randomized or treated subjects.  
7.4.2 
Demographic and Other Baseline Characteristics 
Demographic variables including age, sex, ethnicity and race will be summarized by 
treatment group and over all subjects combined for the Safety, Efficacy, and PP 
Populations. Age at the time of informed consent, as collected on the eCRF, will be 
summarized using descriptive statistics. Sex, ethnicity, and race will be summarized with 
the number and percentage of subjects in each parameter category (including the number 
missing). 


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Baseline characteristics include height, weight, BMI, baseline CGI-S score, HAMD-17 
score, SHAPS scores by both scoring methods, HAM-A score,
HADS subscale and total scores,
 SDS total score, duration of current 
depressive episode, time since first diagnosis of depression, MDD medication use for the 
current episode of MDD prior to the start of study drug administration, non-
pharmacological procedures for the current episode of MDD, and medical and psychiatric 
history other than MDD. Height, weight, BMI, baseline CGI-S score, HAMD-17, SHAPS 
scores by both scoring methods, HAM-A score,
HADS total and 
subscale scores, 
 SDS total score, duration of current depressive 
episode, and time since first diagnosis of depression will be summarized using 
descriptive statistics by treatment group and over all subjects combined for the Safety, 
Efficacy, and PP Populations.   
Medications for the current episode of MDD prior to the start of study drug 
administration will be coded using World Health Organization Drug Classification 
(WHODrug) for Drug Statistics Methodology (B3 Global) September 2019 version. For 
the MDD medication summary, the number and percentage of subjects receiving any 
medication will be summarized by treatment group, as will the number and percentage 
receiving any medication by Anatomical Therapeutic Chemical (ATC) drug class and 
generic drug name. Subjects reporting use of more than one medication at each level of 
summarization (any medication received, ATC class, and generic drug name) will be 
counted only once. ATC class terms will be displayed by descending order of incidence, 
as will generic drug names within each ATC class. 
Medical and psychiatric history other than MDD will be presented only for the Safety 
Population. Medical and psychiatric history conditions will be coded using the Medical 
Dictionary for Regulatory Activities (MedDRA, version 22.1). Medical and psychiatric 
history will be summarized using frequency counts and percentages by treatment group, 
system organ class and preferred term.  
All other baseline characteristics except MDD non-pharmacologic procedures will be 
summarized by treatment group and over all subjects combined for the Safety, Efficacy, 
and PP Populations. Demographic and baseline characteristic information will be listed 
for all randomized or treated subjects. 
7.4.3 
Measurements of Treatment Compliance 
Percentage compliance to the study treatment regimen will be determined as the total 
dose actually taken divided by the expected dose received, multiplied by 100. The 
expected dose received will be calculated as (the date of last dose of study drug – the date 
of first dose of study drug + 1) x 1 capsule/day. If the date of last dose is not collected or 
missing, the date of last dose will be equal to the date of Visit 6/EOT or date of study 
completion/termination, whichever is earlier.  
The total dose actually taken will be determined as the difference between the sum of all 
capsules dispensed before the date of last dose and the sum of all capsules returned. If 
number of capsules taken is not missing, the number of capsules returned will be 


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calculated as the number of capsules dispensed minus the number of capsules taken. If 
both the number of capsules returned and the number of capsules taken are missing, then 
it will be assumed that 0 capsules were returned. In addition, if the number of capsules 
dispensed is 0, then the corresponding number of capsules returned will be set to 0, if 
applicable.  
Dosing compliance will be summarized using descriptive statistics, by treatment group, 
based on the Efficacy Population. As an exception rule, the compliance rate for subjects 
with < 14 days exposure duration will not be calculated, as the compliance may not be 
calculated reliably for such short duration. A subject is defined to be compliant if he/she 
has taken ≥80% and <120% of the number of doses expected to be taken during the study 
overall. The number and percentages of subjects who are compliant and non-compliant 
within each treatment group will be summarized. 
7.4.4 
Primary Efficacy Endpoint Analysis Methods 
The primary efficacy endpoint is the change from baseline to Week 8 on the HAMD-17 
score. The estimand for the primary efficacy endpoint is the difference in means between 
treatment groups in the change of HAMD-17 scores from baseline to Week 8 for all 
subjects in the Efficacy population. For efficacy analyses, baseline is defined as the latest 
measurement prior to the first administration of study drug. The null hypothesis to be 
tested is that there is no difference between BTRX-335140 and placebo: 
 
H0:  PA = PB;  
 
Where PA and PB represent the mean values for BTRX-335140 and placebo, respectively. 
The alternate hypothesis to be tested is that the treatment group means differ: 
 
H1:  PA ɽ PB;  
The analysis will be conducted using a mixed-models repeated measures (MMRM) with 
change from baseline in HAMD-17 score as the dependent variable and will include 
treatment group, week, and week by treatment group interaction as factors, and baseline 
HAMD-17 score as covariate. Variance estimation will be based on an unstructured 
covariance matrix. If the unstructured covariance matrix results in a lack of convergence, 
the heterogeneous autoregressive variance-covariance structure will be used.   
The primary analysis will compare the groups at Week 8. The primary results obtained 
from the model will be the estimated treatment difference at Week 8. Significance will be 
assessed based on a two-sided test at the α=0.05 level of significance. In addition, the 
estimated treatment difference at Week 4 will be provided. The model-based least square 
(LS) means, standard errors, 95% CIs, and 2-sided p-values will be reported. 
In addition, descriptive summary of HAMD-17 and each individual item will be 
generated by treatment group and by visit. 


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7.4.5 
Sensitivity Analyses of the Primary Efficacy Endpoint 
Sensitivity analyses will be used to investigate the impact of important protocol 
deviations or missing data for the primary efficacy endpoint assessment (ie, HAMD-17 
total score). The primary efficacy analysis will be repeated using 
x The per-protocol population. 
x If ≥10% of subjects have missing data for the primary efficacy endpoint 
assessment (i.e., HAMD-17 total score), the following will be conducted: 
o Rubin’s rules for multiple imputation (MI) with SAS Proc MI and 
MIANALYSIS for the efficacy population. See Appendix B : Steps for 
Multiple Imputation for further details.  
o Last Observation Carried Forward (LOCF) Imputation technique, ie, the 
last observed non-missing value will be used to fill in missing values at a 
later point in the study, regardless of when the missing value occurred. 
The imputed dataset generated with the LOCF technique will be used as 
input data in the ANCOVA. The efficacy population will be used. 
7.4.6 
Subgroup Analyses of the Primary Efficacy Endpoint 
The primary efficacy endpoint (change from baseline to Week 8 in HAMD-17 total 
score) will be summarized using the following subgroups: 
x Gender (male and female) 
x Baseline HAMD-17 scores: < 22 vs ≥ 22 
x by baseline SHAPS Total Score (14 to 56) (< median and ≥ median) and SHAPS 
Original Total Score (0 to 14) (< median and ≥ median). 
Summaries by subgroup will only be produced if there are at least 20 subjects in the 
category of interest. Additional subgroup analyses may be performed post-hoc, as 
appropriate. 
The primary endpoint (HAMD-17 total score) will also be summarized in subgroups of 
the Efficacy Population that include all subjects screened on or before March 16, 2020 
(labeled as PreCOVID) versus the subjects screened after March 16, 2020 (labeled as 
PostCOVID).  


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7.4.7 
Secondary Endpoint Analysis Methods  
Secondary endpoints including the change from baseline to each timepoint assessed for 
comparisons between BTRX-335140 and placebo for the measures listed below, will be 
analyzed using an MMRM model for treatment group, week, and week by treatment 
group interaction as fixed factors and a covariate adjustment for baseline score. For CGI-
I, the baseline CGI-I score will not be included as a covariate, as the baseline assessment 
of CGI-I refers to the comparison with screening visit. For all other measures the baseline 
score included as a covariate will be the baseline score for that measure. Variance 
estimation will be based on an unstructured covariance matrix unless there is a lack of 
convergence in which case a heterogeneous autoregressive variance-covariance structure 
will be used.  
x CGI-I at each post-baseline visit 
x Change in SHAPS from baseline at each post-baseline visit 
x Change in HADS total score and HADS subscales: HADS-A and HADS-D from 
baseline at each post-baseline visit 
x Change in HAM-A total score from baseline at each post-baseline visit 
x Change in SDS from baseline at each post-baseline visit 
The response rates (the percentage of subjects with ≥ 50% decrease in HAMD-17 from 
baseline to Week 4 and Week 8) will be compared between BTRX-335140 and placebo-
treated groups. The response rates, response rate difference and corresponding 95% Wald 
CI, and Cochran-Mantel-Haenszel p-value will be reported. 
The change at Week 8 from baseline in CGI-S will be estimated using an analysis of the 
covariance (ANCOVA) model with main effects for treatment group and a covariate 
adjustment for baseline CGI-S score. The model-based LS means, standard errors, 95% 
CIs, and 2-sided p-values will be reported. 
 
For SDS, analysis will be performed for the three SDS items separately and the SDS total 
score as the sum of the individual domains. Note that the work/school domain may be not 
applicable for all subjects and therefore the analysis of SDS total score will be performed 
for the subjects with all 3 domain scores and the checkbox of “No work or school” is 
NOT marked. In addition, a sensitivity analysis of SDS total score will be conducted for 
all subjects, where the SDS total score will be calculated as (sum of social and family 
life)*30/20 for the subjects for whom the checkbox of “No work or school” is marked. 
For CGI-I, an additional analysis will be performed to compare the symptom 
improvement rate, i.e. the percentage of subjects with responses at each post-baseline 
visit of ‘much improved’ or ‘very much improved’ (i.e. CGI-I score ≤2) between BTRX-
335140 and placebo. The symptom improvement rate, improvement rate difference and 
corresponding 95% Wald CI, and Cochran-Mantel-Haenszel p-value will be reported. 








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7.5.1 
Adverse Events 
TEAEs are defined as those AEs with onset after the first dose of study drug or existing 
events that worsened after the first dose during the study and that occur prior to 30 days 
after last dose of treatment. TEAEs will be summarized by treatment group. Events 
reported with a partial onset date (e.g., month and year are reported but the day is 
missing) will be considered to be treatment-emergent unless a partial date clearly 
indicates that it occurred prior to first dose of study treatment or more than 30 days after 
last dose of treatment. 
Verbatim terms on case report forms will be mapped to preferred terms and system organ 
classes using MedDRA version 22.1.   
Summaries that are displayed by system organ class and preferred terms will be ordered 
by descending incidence of system organ class and preferred term within each system 
organ class. Summaries displayed by preferred term only will be ordered by descending 
incidence of preferred term. Summaries of the following types will be presented: 
x Overall summary of number of unique TEAEs and treatment-emergent serious 
adverse events (TESAEs) and subject incidence of TEAEs meeting various 
criteria; 
x Subject incidence of TEAEs by MedDRA system organ class and preferred term; 
x Subject incidence of TEAEs by MedDRA preferred term; 
x Subject incidence of TEAEs by severity grade, MedDRA system organ class, and 
preferred term; 
x Subject incidence of TEAEs related to study drug, MedDRA system organ class, 
and preferred term;  
x Subject incidence of TEAEs related to study drug by MedDRA preferred term; 
x Subject incidence of TEAEs leading to discontinuation of study drug by 
MedDRA system organ class and preferred term; 
x Subject incidence of severe TEAEs related to study drug by MedDRA system 
organ class and preferred term;  
x Subject incidence of SAEs by MedDRA system organ class and preferred term; 
x Subject incidence of AESIs by MedDRA system organ class and preferred term. 
x Subject incidence of AESIs by severity grade, MedDRA system organ class, and 
preferred term; and 


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x Subject incidence of AESIs related to study drug, MedDRA system organ class, 
and preferred term 
At each level of summarization (e.g., any AE, system organ class, and preferred term), 
subjects experiencing more than one TEAE will be counted only once. In the summary of 
TEAEs by severity grade, subjects will be counted once at the highest severity reported at 
each level of summarization.  
When summarizing the related adverse events, the adverse events with relationship of 
unlikely related, possibly related, and missing are counted. 
All adverse events will be presented in data listings by treatment group, subject, and 
event date. Serious AEs, AEs leading to discontinuation of the study drug, and AEs 
leading to death will be presented in separate data listings. 
7.5.2 
Extent of Exposure 
Extent of exposure to study treatment will be summarized for the Safety Population by 
treatment group. The duration of exposure will be presented in days and calculated as the 
date of last dose of study drug minus the date of first dose of study drug, plus one. If the 
date of last dose is not collected, the date of last dose will be equal to the date of Visit 
6/EOT or date of study completion/termination, whichever is earlier. The total dose 
received is calculated as (sum of all capsules dispensed before date of last dose – sum of 
all capsules returned) x 80mg. If the number of capsules taken is not missing, the number 
of capsules returned will be calculated as the number of capsules dispensed minus the 
number of capsules taken. If both the number of capsules returned and the number of 
capsules taken are missing, then it will be assumed that 0 capsules were returned. In 
addition, if the number of capsules dispensed is 0, then the corresponding number of 
capsules returned will be set to 0, if applicable. Duration of exposure and total dose 
received (mg) will be summarized using descriptive statistics.  
7.5.3 
Deaths, Other Serious Adverse Events, and Other Significant Adverse 
Events 
All deaths during the study, including the post treatment follow-up period, will be listed 
by treatment group and subject, to include the primary cause of death. Treatment-
emergent serious AEs and other significant AEs, including those that led to 
discontinuation of the study drug, will be provided in separate subject data listings. 
7.5.4 
Clinical Laboratory Evaluation 
All descriptive summaries of laboratory results will be based on data analyzed by the 
central laboratory and presented in Système International (SI) units, as suggested by the 
Center for Biologics Evaluation and Research and the Center for Drug Evaluation and 
Research Position on Use of SI Units for Lab Tests (Oct 2013). All data will be included 
in by-subject data listings. Laboratory measurements identified as abnormal (ie, outside 
the normal range) will also be listed separately.  




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7.5.5 
Vital Signs, Physical Findings, and Other Observations Related to Safety 
7.5.5.1 
Vital Signs 
Vital signs will include pulse (supine and standing), systolic/diastolic blood pressure 
(supine and standing), respiratory rate, body temperature, height, weight, and BMI will 
be listed by treatment group, subject, and visit date. Vital sign parameter measurements 
will be summarized by treatment group. Descriptive statistics will be presented for results 
and change from baseline at each visit where parameters were scheduled to be collected.  
The number and percentage of subjects with a PCS measurement in supine will be 
presented by treatment group and study visit. PCS measurements include systolic blood 
pressure ≤ 90 mmHg, systolic blood pressure ≥ 180 mmHg, diastolic blood pressure ≤ 50 
mmHg, diastolic blood pressure ≥ 105 mmHg, heart rate ≤ 40 bpm, and heart rate ≥ 120 
bpm. PCS measurements in supine will also be listed. 
Additionally, the number and percentage of subjects with an orthostatic systolic BP 
decrease > 20 mmHg or diastolic BP decrease > 10 mmHg from supine to standing 
position at any post-baseline measurement during the study will be summarized and 
listed. 
7.5.5.2 
12-Lead Electrocardiogram 
Twelve-Lead ECG interval parameters including heart rate, PR, RR, QRS, QT, QTc 
interval calculated using the Fridericia method (QTcF), and QTc interval calculated using 
the Bazett method (QTcB) will be summarized by treatment group. Descriptive statistics 
will be presented for observed values and changes from baseline at each visit where 
parameters were scheduled to be collected. 
The number and percentage of subjects with a PCS QTcF result will be presented by 
treatment group and study visit. PCS results at post-baseline visits include QTcF ≥ 500 
msec, QTcF change from baseline ≥ 60 msec, QTcF change from baseline ≥ 30 msec. 
PCS results at post-baseline visits will also be listed. 
7.5.5.3 
Ophthalmology Examination 
Standard ophthalmologic examination and corneal specular microscopy examination 
results will be summarized by presenting shift from baseline to end of study results as 
normal, abnormal not clinically significant, and abnormal clinically significant. Clinically 
significant abnormalities on ophthalmologic exam will be listed by treatment group, 
subject, and visit date.  
The baseline, post-baseline and percent (%) change from baseline of cell density and 
coefficient of variation (CV) results from the central reading group for CSM will be 
summarized descriptively for each eye (oculus dextrus [OD] and oculus sinister [OS]). 
The number and percentage of subjects with percent decrease from baseline in cell 
density >= 5%, 10%, and 15% will be summarized for each eye and for both eyes. 
Results from the central reading group for CSM will also be presented in a data listing. 


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Additional subgroup summaries will be performed by three device types (Konan, Nidek 
and Tomey) and by age groups (18-29, 30-39, 40-49, 50-59, 60-65). All the above 
summaries will be repeated based on the set of subjects excluding the subjects who had 
endothelial dystrophy medical history prior to treatment. 
7.5.5.4 
Physical Examination 
Abnormal results of the physical examination will be presented in subject data listings by 
treatment group, subject, and visit date. 
7.5.5.5 
Prior and Concomitant Medications 
Medications will be coded using WHODrug (B3 Global) September 2019 version. 
Medications entered on the eCRF will be mapped to ATC drug class (level 4) and drug 
name. If ATC drug class level 4 is not available, then ATC drug class level 3 will be 
utilized. If ATC drug class levels 4 and 3 are not available, then ATC drug class level 2 
will be utilized. If ATC drug class levels 4, 3, and 2 are not available, then ATC drug 
class level 1 will be utilized. 
Prior and concomitant medications will be summarized separately and the study phase of 
each medication will be determined programmatically based on medication start and end 
dates. A prior medication is defined as any medication administered and stopped prior to 
the date of the first dose of study drug. A concomitant medication is defined as any 
medication administered on or after the date of the first dose of study drug. If it cannot be 
determined whether a medication was received after the start of study drug dosing, it will 
be considered concomitant. 
For both prior and concomitant medications summaries, the number and percentage of 
subjects receiving any medication will be summarized by treatment group, as will the 
number and percentage receiving any medication by ATC drug class and generic drug 
name. Prior medications will also be summarized over all subjects combined. Subjects 
reporting use of more than one medication at each level of summarization (any 
medication received, ATC class, and generic drug name) will be counted only once. ATC 
class terms will be displayed by descending order of incidence, as will generic drug 
names within each ATC class. Prior and concomitant medications will be listed 
separately. 
7.5.5.6 
Prior and Concomitant Non-Pharmacological Procedures 
Non-pharmacological procedures will be listed by treatment group, subject, and date. 
7.5.5.7 
Columbia Suicide Severity Rating Scale 
Suicidality data collected on the C-SSRS at screening, baseline, and by visit during the 
treatment period will be listed for all subjects. Listings will include all data collected on 
the C-SSRS.  
At screening visit, the number and percentage of subjects with a response of ‘Yes’ to: 








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Period 
Screening 
Baseline 
Study Drug Treatment 
Safety Follow-up  
 
Visit 
1b 
2 
3 
4 
5 
6 
TCa 
Ocular 
Exam 
 
Days –7 to 
–28 
Day 1 
Day 15 
(±2 d) 
Day 29 
(±2 d) 
Day 43 
(±2 d) 
Day 57/ 
EOTc 
(±2 d) 
Procedure and Assessments 
NA 
Week 0 
Week 2 
Week 4 
Week 6 
Week 8 
Week 10 
Week 
12 
SHAPS 
X 
X 
 
X 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
COWS 
X 
X 
X 
X 
X 
X 
 
 
Adverse eventsp 
 
X 
X 
X 
X 
X 
X 
 
Complete IWRS 
X 
X 
X 
X 
X 
X 
 
 
Dispense study drug 
 
Xq 
Xq 
Xq 
Xq 
 
 
 
Study drug administration 
 
Xr 
 
Xs 
 
 
 
 
Study drug compliance 
 
 
X 
X 
X 
X 
 
 
BMI = body mass index; CRF=case report form; CGI-I = Clinical Global Impression of Improvement; CGI-S = 
Clinical Global Impression of Severity; COWS = Clinical Opiate Withdrawal Scale; CSSRS = Columbia Suicide 
Severity Rating Scale; d = day; DSM = Diagnostic and Statistical Manual of Mental Disorders; ECG = 
electrocardiogram; eCRF=electronic case report form; EOT = End of Treatment; HADS = Hospital Anxiety and 
Depression Scale; HAM-A = Hamilton Anxiety Rating Scale; HAMD-17 = Hamilton Rating Scale for Depression – 
17Item Version; IWRS = Interactive Web-response System;
 NA = not applicable; 
 
; 
; 
; SDS = Sheehan Disability 
Scale; SCID5CT = Structured Clinical Interview for DSM-5 Disorders, Clinical Trials Version; SHAPS = 
SnaithHamilton Pleasure Scale; 
; TC = telephone call; UDS=urine drug screen. 
a 
Study site personnel will contact subjects by telephone for Safety Follow-up assessments (adverse events, 
concomitant medication usage, and C-SSRS) approximately 2 weeks after discontinuation of study drug. 
b 
Visit 1 procedures may be completed over 2 days, if necessary. 
c 
Subjects who discontinue early will complete an EOT visit <2 days after their last dose of study drug; efficacy 
assessments are to be performed only if the subject has remained on study drug. For subjects who do not return, 
the site should complete a C-SSRS if they are aware of a potential suicide-related thought or behavior by other 
communications. The 12-week corneal specular microscopy examination should still be conducted approximately 
12 weeks after the first dose of study drug (Section 7.7.2 of the protocol).  
d 
Inclusion/exclusion criteria must be reconfirmed for eligibility prior to randomization. 
e 
Collect vital signs measurements (respiratory rate first, then blood pressure and pulse) after subject has been 
supine for 10 minutes; subjects should then stand for 3 minutes before obtaining standing vital signs 
measurements [see Section 6.3.4 of the protocol]. Oral temperature can be assessed in any position. 
f 
A full physical examination involving an evaluation of cardiovascular, respiratory, gastrointestinal, neurological 
(examination of cranial nerves, motor system, sensory, and reflexes), dermatological, and musculoskeletal 
systems, and include general appearance, skin, head, neck, eyes, ears, nose, throat, lungs, heart, abdomen, back, 
lymph nodes, extremities, and vascular status will be conducted at the screening visit. Further physical 
examination will be conducted as needed. (Section 6.3.3 of the protocol). 
g 
At Screening, collect height for BMI calculation. 
h 
Ophthalmologic examinations will be performed by a site-affiliated ophthalmologist. Details for the standard 
ophthalmologic examination and corneal specular microscopy will be described in a separate ocular monitoring 
manual.  Additional standard ophthalmologic exams and / or corneal specular microscopies may be conducted to 
assess subject safety. 
i 
Baseline ophthalmologic examinations (standard and corneal specular microscopy) must be performed before 
randomization. 
j 
Prior to Visit 3 (Week 2) (between Days 7 and 14 [+2 days]), a standard ophthalmologic examination will be 
performed.