Clinical trial NCT03763175
Conditions: Irritable Bowel Syndrome With Constipation
Keywords: Irritable Bowel Syndrome with Constipation Lovastatin SYN-010
Sponsor: Cedars-Sinai Medical Center
Start date: 2018-12-24; Completion date: 2020-10-09
Enrollment: 59 participants; Allocation: RANDOMIZED; Intervention model: PARALLEL

Brief summary:
Irritable bowel syndrome (IBS) is a gastrointestinal (GI) syndrome characterized by chronic abdominal pain and altered bowel habits in the absence of any organic cause. The symptoms of IBS not only adversely affect a patient's health-related quality of life (QoL), but also place a significant financial burden on society due to reduced work productivity and increased use of healthcare-related resources. Patients with IBS frequently complain of abdominal bloating and increased gas production in the form of flatulence or belching. The prevalence in North America and Europe is approximately 10-15%. Irritable bowel syndrome affects all ages and genders however there is a 2:1 female predominance in North America. Irritable bowel syndrome is classified into 4 subtypes based on stool pattern: IBS with constipation (IBS-C), IBS with diarrhea, mixed IBS, and un-subtyped IBS. Irritable bowel syndrome with constipation is defined as the presence of hard or lumpy stools with ≥ 25 percent of bowel movements and loose or watery stools with \< 25% of bowel movements.

SYN-010 is a modified release, oral formulation of lovastatin being developed for the treatment of IBS-C. The SYN-010 program is based predominantly on research by Dr. Mark Pimentel and collaborators hypothesizing that reduction in intestinal methane (methane) production can reverse constipation and improve global symptoms in IBS-C. Methane production in humans is due to methanogenic archaea in the intestine, predominantly Methanobrevibacter smithii (M. smithii). Methane, the key product of anaerobic respiration of methanogens, had been perceived to produce no ill effects in humans aside from gaseous distention. However, several research groups worldwide have shown that a significant percentage of patients with IBS-C excrete methane, and elevated methane production by methanogens correlates with constipation and related symptoms in both IBS-C and chronic idiopathic constipation. A direct causative role for methane in IBS-C was demonstrated in a recent case report, wherein a woman undergoing fecal microbiota transplantation (FMT) for C. difficile infection unknowingly received stool containing a high concentration of methanogens. The FMT recipient rapidly developed severe symptoms of IBS-C that were subsequently reversed by ablation of methane production.

Detailed description:
SYN-010 has previously been evaluated in consecutive Phase 2a clinical trials. Sixty-three (63) IBS-C patients with high breath methane (\>10 ppm) at screening were enrolled in a multicenter, randomized, controlled, double-blinded clinical trial (RCT) in which they received SYN-010 21 mg, SYN-010 42 mg or Placebo once daily for 4 weeks. Fifty-four (54) subjects who completed the RCT continued into an open-label extension (EXT) in which all subjects received SYN-010 42 mg once daily for an additional 8 weeks.

The SYN-010 Phase 2a studies were intentionally designed as mechanistic proof-of-concept studies, wherein reductions in breath methane were employed as a rapid and cost-effective means by which to determine if SYN-010 could be effective in treating an underlying cause of symptoms in IBS-C. Breath methane was reduced relative to baseline in SYN-010 treatment groups, and lower breath methane levels correlated with an increased number of complete spontaneous bowel movements (CSBMs) at week 12, consistent with the proposed methane-inhibiting action of lovastatin lactone.

Since lovastatin has not previously been used to treat IBS-C patients, the SYN-010 Phase 2a studies were also focused on the safety of the SYN-010 dosage form. Daily doses of SYN-010 were well-tolerated by IBS-C patients over the 12-week treatment period (at least 8 weeks of SYN-010 42 mg). SYN-010 did not cause clinically meaningful or persistent changes in serum liver and muscle markers in IBS-C patients at daily doses of 21 mg and 42 mg. Modest decreases from baseline in lipid parameters observed after 7 days of SYN-010 21 mg or 42 mg had largely faded by 28 days and were not evident after 12 weeks of dosing. Very few adverse events were reported over 12 weeks of SYN-010 treatment and all were of mild or moderate intensity. No serious adverse events were reported and there were no incidences of drug-related diarrhea, which is an important potential benefit of SYN-010 as an IBS-C therapy.

Although the Phase 2a studies were not prospectively powered for formal statistical evaluation of clinical endpoints, compelling improvements in CSBMs, abdominal pain, and bloating were observed in SYN-010 treatment groups. These clinical findings have been presented in multiple public forums and a panel of clinical advisors affirmed that the Phase 2a data validate the need to evaluate optimal dosing of SYN-010 in a larger patient population over a longer dosing period.

Based on the potential clinical benefit observed in Phase 2a, this Phase 2b clinical study will evaluate in more detail the clinical effects of two dose strengths of SYN-010 administered over a longer treatment period (12 weeks per FDA guidelines) to a larger number of IBS-C patients. The study will seek more definitive evidence regarding potential symptom improvements and safety in IBS-C patients. The study also seeks to provide new information of relevance to both efficacy and safety by measuring changes to the microbiome in patient stool samples; serum level of cytokine markers of inflammation; and expanded breath gas measurements.

SYN-010 is a hydroxypropyl methylcellulose (HPMC) capsule filled with enteric-coated tablets from which lovastatin is released at different intestinal pH values. The tablets are designed to pass through the stomach unchanged then release a small amount of lovastatin into the duodenum and the majority of the lovastatin dose into the ileocecal junction and colon. The amount of lovastatin to be released into the small and large intestine is anticipated to be consistent with the relative levels of methane-producing archaea in each location of the intestine.

Methane production (methanogenesis) is a ubiquitous process in the human intestine, disposing of hydrogen and other by-products formed during bacterial fermentation. Methane production in humans is almost entirely due to the archeon M. smithii. Elevated intestinal methane production reduces intestinal motility and is a cause of constipation, pain and bloating in IBS-C.

Interest in lovastatin as a potential inhibitor of methanogenesis originated with recognition that the rate-limiting step in the synthesis of archaeal lipid membranes is catalyzed by HMG-CoA reductase (HMGR) which is the target enzyme for cholesterol-lowering statins. Subsequent in vitro studies (described above) and computational studies have since determined that HMGR is not the target for lovastatin anti-methanogenic activity; rather, lovastatin lactone appears to exert a direct effect on methanogenesis enzymes. The mechanism by which lovastatin lactone inhibits methane production by M. smithii has not been conclusively determined; however, detailed computational studies showed that lovastatin lactone may competitively inhibit F420-dependent methylenetetrahydromethanopterin dehydrogenase (mtd), an enzyme that is integral to the M. smithii methanogenesis pathway.

Methanogenic archaea reside predominantly in the human colon, with lower methanogen levels measured in the small intestine of some patients. Methane production at both sites contributes to reduced gastrointestinal motility and rat studies suggest that the ileocecal region may be of particular significance. SYN-010 utilizes a dual-pulse release profile to deliver a portion of the lovastatin dose to the small intestine and the majority of the dose to the ileocecal junction and colon where the most methane-producing organisms are found; the relative amounts of lovastatin released into the small and large intestine are consistent with the anticipated relative levels of methanogens in each location.

Lovastatin lactone exerts its methane-reducing effect in the intestinal lumen and systemic absorption of lovastatin is not required for its therapeutic effect in IBS-C.

Arms: arm 1: Enrolled subjects will be screened and randomized into the three study arms in a 1:1:1 ratio. 50 subjects with constipation-predominant irritable bowel syndrome (IBS-C) will be administered a 12-week course of lovastatin (21 mg PO QD). Study activities will be the same across all three arms. arm 2: Enrolled subjects will be screened and randomized into the three study arms in a 1:1:1 ratio. 50 subjects with constipation-predominant irritable bowel syndrome (IBS-C) will be administered a 12-week course of lovastatin (42 mg PO QD). Study activities will be the same across all three arms. arm 3: Enrolled subjects will be screened and randomized into the three study arms in a 1:1:1 ratio. 50 subjects with constipation-predominant irritable bowel syndrome (IBS-C) will be administered a 12-week course of placebo. Study activities will be the same across all three arms.
Interventions: intervention 1: SYN-010 21 mg intervention 2: SYN-010 42 mg intervention 3: Placebo
Intervention descriptions: intervention 1: 21 mg lovastatin will be administered to patients diagnosed with IBS-C to evaluate whether the medication and dose is effective in increasing frequency of completely spontaneous bowel movements. The secondary objectives of the study are to assess the role of lovastatin lactone in increasing overall stool frequency and reducing abdominal pain severity, bloating severity, and rescue medication use. Additional endpoints include improvement in reported adequate relief and the effect of the study drug in lowering exhaled methane levels. intervention 2: 42 mg lovastatin will be administered to patients diagnosed with IBS-C to evaluate whether the medication and dose is effective in increasing frequency of completely spontaneous bowel movements. The secondary objectives of the study are to assess the role of lovastatin lactone in increasing overall stool frequency and reducing abdominal pain severity, bloating severity, and rescue medication use. Additional endpoints include improvement in reported adequate relief and the effect of the study drug in lowering exhaled methane levels. intervention 3: A placebo will be administered to patients diagnosed with IBS-C to evaluate whether the medication and dose is effective in increasing frequency of completely spontaneous bowel movements. The secondary objectives of the study are to assess the role of lovastatin lactone in increasing overall stool frequency and reducing abdominal pain severity, bloating severity, and rescue medication use. Additional endpoints include improvement in reported adequate relief and the effect of the study drug in lowering exhaled methane levels.

Protocol documents:
Prot_000.pdf: 
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Evaluation of the EfficAcy and Safety of SinglE, Daily Oral Doses  
of SYN-010 Compared to Placebo in Adult Patients with  
Irritable Bowel Syndrome with Constipation (EASE-DO) 
 
 
 
 
Medically Associated Science and Technology (MAST) Program 
Cedars-Sinai Medical Center 
Los Angeles, CA 
 
 
 
 
 
 
 
 
Investigator Sponsored Research (ISS) Protocol  
Product Name: SYN-010 
Chemical Name:  Lovastatin Lactone Modified Release Capsules 
USAN (United States Adopted Name): Lovastatin 
Chemical formula: 1S-[1α(R*),3α,7α,8α(2S*,4S*),8aα]]-1,2,3,7,8,8a-hexahydro-3,7-dimethyl-8-[2-
(tetrahydro-4-hydroxy-6-oxo-2H-pyran-2-yl) ethyl]-1-naphthalenyl 2-methylbutanoate (C24H36O5) 
NCT03763175 
 


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Table of Contents 
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Background .................................................................................... Error! Bookmark not defined. 
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Summary of findings from pre-clinical studies with potential clinical significance .............. Error! 
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3.3 
Rationale of the study .................................................................... Error! Bookmark not defined. 
3.4 
SYN-010 (lovastatin lactone modified release capsules) ............... Error! Bookmark not defined. 
3.4.1 
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3.4.2 
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4. 
Study objectives ..................................................................................... Error! Bookmark not defined. 
4.1 
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4.3 
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5.1 
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5.3 
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Statistical methods and determination of sample size .......................... Error! Bookmark not defined. 
6.1 
General considerations .................................................................. Error! Bookmark not defined. 
6.2 
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Analysis populations ...................................................................... Error! Bookmark not defined. 
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Demographics and baseline characteristics ................................... Error! Bookmark not defined. 
6.6 
Efficacy analyses ............................................................................ Error! Bookmark not defined. 
6.7 
Safety analyses ............................................................................... Error! Bookmark not defined. 
6.8 
Determination of sample size ........................................................ Error! Bookmark not defined. 
7. 
Study population .................................................................................... Error! Bookmark not defined. 
7.1 
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7.2 
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7.2.1 
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7.2.2 
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7.3 
Excluded therapies and medications, previous and concomitant . Error! Bookmark not defined. 
7.4 
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7.5 
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7.6 
Screen failures ................................................................................ Error! Bookmark not defined. 
8. 
Treatments ............................................................................................. Error! Bookmark not defined. 
8.1 
Randomization ............................................................................... Error! Bookmark not defined. 
8.2 
Investigational product .................................................................. Error! Bookmark not defined. 
8.2.1 
SYN-010 ...................................................................................... Error! Bookmark not defined. 
8.3 
Comparator treatment to be administered ................................... Error! Bookmark not defined. 
8.3.1 
Placebo ....................................................................................... Error! Bookmark not defined. 
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Full description and handling of treatments.................................. Error! Bookmark not defined. 
8.4.1 
SYN-010 ...................................................................................... Error! Bookmark not defined. 
8.5 
Blinding .......................................................................................... Error! Bookmark not defined. 
8.5.1 
Emergency unblinding by the investigator ................................ Error! Bookmark not defined. 
8.6 
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Storage ....................................................................................... Error! Bookmark not defined. 
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Destruction and return .............................................................. Error! Bookmark not defined. 
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Treatment compliance ................................................................... Error! Bookmark not defined. 
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Prior and concomitant therapy ...................................................... Error! Bookmark not defined. 
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9.3.1 
ROME IV criteria ......................................................................... Error! Bookmark not defined. 
9.3.2 
Demographics and medical history ............................................ Error! Bookmark not defined. 
9.3.3 
Physical exam ............................................................................. Error! Bookmark not defined. 
9.3.4 
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12-lead ECG ................................................................................ Error! Bookmark not defined. 
9.3.6 
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9.3.9 
Stool samples ............................................................................. Error! Bookmark not defined. 
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Breath gases ............................................................................... Error! Bookmark not defined. 
9.3.11 
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Pregnancies ................................................................................ Error! Bookmark not defined. 
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Further safety ............................................................................. Error! Bookmark not defined. 
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Archiving ........................................................................................ Error! Bookmark not defined. 
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12.1 
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Laboratory tests ............................................................................. Error! Bookmark not defined. 
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Daily questionnaire ........................................................................ Error! Bookmark not defined. 
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Quality of life questionnaire .......................................................... Error! Bookmark not defined. 
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Medical history .............................................................................. Error! Bookmark not defined. 
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1. Project summary 
Compound No. 
 
Name of Active Ingredient of 
Investigational Agent 
Route of Administration 
 
SYN-010 lovastatin lactone modified release capsules 
 
21mg given orally once daily 
42mg given orally once daily 
Title of Study 
Evaluation of the EfficAcy and Safety of SinglE, Daily Oral Doses of 
SYN-010 Compared to Placebo in Adult Patients with Irritable Bowel 
Syndrome with Constipation (EASE-DO) 
Study Center 
Cedars-Sinai Medical Center 
8700 Beverly Boulevard 
Los Angeles, CA 90048 
Phase of Clinical Development Phase II 
Indication 
Irritable Bowel Syndrome with Constipation (IBS-C) 
Control Agent 
Name of Active Ingredient of 
Control Agent 
Route of Administration 
Reference product is Placebo in matching presentation to be given 
orally once-daily. 
Clinical Study Phase 
Phase II 
Study Objectives 
Primary: 
Change from baseline in the weekly average number of CSBMs during 
the 12-week Treatment Period for SYN-010 relative to placebo. 
Secondary: 
Safety and tolerability of SYN-010 21 mg and 42 mg in IBS-C patients 
treated for 12 weeks. 
For SYN-010 relative to placebo: 
• 
Proportion of overall stool frequency responders. 
• 
Proportion of overall bloating responders. 
• 
Proportion of patients using rescue medication. 
• 
Proportion of overall responders in the SYN-010 21 mg and 42 
mg treatment groups versus placebo. 
• 
Change from baseline for SYN-010, relative to placebo, in: 
o Weekly average score for worst abdominal pain at Weeks 
1 through 12. 
o Weekly average number of CSBMs at Weeks 1 through 
12. 
o Weekly average number of SBMs at Weeks 1 through 12. 
o Weekly average bloating score at Weeks 1 through 12. 
o Weekly average stool consistency (BSFS) at Weeks 1 
through 12. 
Exploratory: 
For SYN-010 relative to placebo: 
• 
Proportion of patients with adequate relief. 


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• 
Change from baseline for SYN-010 in patient reported 
outcomes (PRO) using validated questionnaires in the REDCap 
system. 
• 
Breath gas area-under-the-curve (AUC) (methane, hydrogen, 
and hydrogen sulfide) on lactulose breath test at EOS 
compared to Screening. 
• 
Spot breath tests at Randomization, Day 28, and Day 56. 
• 
Comparison of serum cytokine levels at baseline and EOS – 
measuring for potential immune or inflammatory signals to 
include the key cytokines of the Th1/Th2/Th17 pathways:  IL-2, 
IL-4, IL-5, IL-9, IL-10, IL-13, IL-17, GM-CSF and IFN-γ. Plus TNF-α, 
IL-12, IL-6, the two chemokines IL-8 and MCP-1, and the gut 
cytokine IL-25. 
• 
Microbiome measurements in stool:  
o Bacteria (16S rRNA) 
o Methanogens (qPCR) 
Study Design 
Single-center, randomized, double-blind, placebo-controlled clinical 
trial comprising three periods (Figure 1): 
1. Screening and Pre-treatment Period (up to 17 days prior to the 
first dose of the study drug), 
2. Treatment Period comprising Randomization (Day 1) and 12 
weeks of dosing (with clinic visits at Weeks 4, 8, and 12), and 
3. End-of-Study contact between 24 and 48 hours after Week 12 
for patients who complete the Treatment Period. 
 
Figure 1: Clinical study design 
Main Inclusion Criteria 
1. Male or female participants aged between 18 and 65 years 
inclusive. 


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2. Patient must be willing and able to participate in the study for 
the required duration, understand and sign the informed 
consent form (ICF), and be willing to comply with all protocol-
related visits and procedures. 
3. Patient has had IBS-C symptoms (as defined by Rome IV 
diagnostic criteria) for at least 6 months prior to diagnosis.  
4. Patient has an average score of ≥ 3.0 for daily abdominal pain 
at its worst (11-point numerical rating scale [NRS]) during and 
up to the 17 days immediately before randomization (i.e. Pre-
treatment Period). 
5. Patient has an average of ≤ 3 CSBMs per week or ≤ 5 SBMs per 
week during the 17 days immediately before randomization 
(i.e. Pre-treatment Period). 
6. Patient has a breath methane level ≥ 10 ppm on a lactulose 
breath test administered at Screening. 
7. Patient may be on a stable, continuous regimen of fiber or 
probiotics one month before the Screening Visit; however, they 
must maintain a stable dose regimen through Week 12. 
8. Patient must agree to refrain from starting a new diet, 
changing stable dose of supplemental fiber, or changing 
exercise pattern that may affect IBS-C symptoms from the time 
of Screening through the end of the study. If the patient takes 
food products that are strong inhibitors of cytochrome P450 3A 
(CYP3A) (e.g. grapefruit juice, Seville orange juice, St. John’s 
Wort), he/she must agree to refrain from taking these from the 
time of Screening through the end of the study. 
9. Patient must agree to use an acceptable method of 
contraception from the time of signing the ICF to 30 days after 
the final dose of study drug if the patient is a sexually active 
female of child-bearing potential (defined as any female who 
has experienced menarche and who is NOT permanently sterile 
or postmenopausal. Postmenopausal is defined as 12 
consecutive months with no menses without an alternative 
medical cause). Adequate contraceptive measures include: oral 
contraceptives (stable use for two or more cycles before 
Screening); intrauterine device; Depo-Provera®; Norplant® 
System implants; partner with a vasectomy; double-barrier 
birth control (e.g. use of a condom plus diaphragm or condom 
plus either contraceptive sponge, foam, or jelly); or abstinence. 
According to drug research standards, male patient must agree 
to use an acceptable method of contraception and refrain from 
donating sperm from the time of signing the ICF to 90 days 
after the final dose of study drug. 


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10. A female patient of child-bearing potential must be non-
pregnant and non-lactating and have negative pregnancy tests 
at the Screening Visit and on Day 1 prior to dosing with study 
drug. 
11. Patients must be able to understand and be willing to sign the 
written informed consent form. A signed informed consent 
form must be appropriately obtained prior to the conduct of 
any trial-specific procedure. 
12. Willing and able to comply with the protocol, including follow-
up visits and examinations. 
Main Exclusion Criteria 
1. Patient has loose (mushy) or watery stools for > 25% of their 
bowel movements (BMs) during the 12 weeks before Screening 
or during the Screening and Pre-treatment Periods. 
2. Patient has a history of cathartic colon, laxative, or enema 
abuse. 
3. Patient has a history of ischemic colitis. 
4. Patient has a history of pelvic floor dysfunction. 
5. Patient has a history of bariatric surgery for the treatment of 
obesity. 
6. Patient has a history of surgery to remove a segment of the 
gastrointestinal (GI) tract at any time before the Screening 
Visit. 
7. Patient has any history of myopathy, rhabdomyolysis, chronic 
myalgia, or familial history of hereditary muscular disorders. 
8. Patient has been diagnosed with or has a family history of 
familial adenomatous polyposis, hereditary nonpolyposis 
colorectal cancer, or any other form of familial colorectal 
cancer. 
9. Patient currently has any structural abnormality of the GI tract 
or a disease or condition that can affect GI motility, or any 
unexplained and clinically significant symptoms such as lower 
GI bleeding, rectal bleeding, heme-positive stool, iron-
deficiency anemia, weight loss, or systemic signs of infection. 
10. Patient has had any previous surgery involving the abdomen, 
pelvis, or retroperitoneal region during the last 12 months 
prior to Screening, with the exclusion of laparoscopic 
gallbladder surgery or appendix removal. 
11. Patient has a history of diverticulitis or any chronic condition 
that could be associated with abdominal pain or discomfort 
and could confound the assessments in the study (e.g. 
inflammatory bowel disease, chronic pancreatitis, polycystic 
kidney disease, ovarian cysts, endometriosis, or lactose 
intolerance). 


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12. Patient has history of severe renal insufficiency defined as an 
actual or estimated glomerular filtration rate of < 30 
mL/min/1.73 m2 within the 6 months prior to Screening Visit. 
13. Patient has any history of a medical condition or a concomitant 
medical condition that, in the opinion of the investigator, 
would compromise the patient’s ability to complete the study 
safely or could confound the assessments in this study (e.g. 
uncontrolled hypothyroidism). 
14. Patient is known to have elevated liver enzyme levels 
(aspartate aminotransferase [AST], alanine aminotransferase 
[ALT], alkaline phosphatase [ALP]) or creatine kinase levels that 
are ≥ 1.5 times the upper limit of normal (ULN) that have not 
been resolved within the 4 weeks prior to consent and/or 
these elevated levels are present at the Screening Visit 
laboratory assessment. 
15. Patient has any abnormal laboratory results, electrocardiogram 
(ECG) findings, or physical examination findings deemed 
clinically significant by the investigator during the Screening 
Period. 
16. Within 14 days prior to the Screening Visit, patient has used 
concomitant medications that are: (1) moderate-to-strong 
inhibitors of cytochrome P450 3A (CYP3A) and/or organ anion 
transporting polypeptide (OATP)1B1 (e.g. cyclosporine, 
verapamil, dronedarone, diltiazem, amiodarone, itraconazole, 
ketoconazole, posaconazole, voriconazole, clarithromycin, 
telithromycin, human immunodeficiency virus protease 
inhibitors, boceprevir, telaprevir, nefazodone, erythromycin, 
cobicistat-containing products); (2) other concomitant 
medications that are excluded from the lovastatin label (e.g. 
rifampin, colchicine, ranolazine); or (3) metformin or GLP-1 
agonists. Patients should not take any of these concomitant 
medications during the treatment phase of the study without 
contacting the investigator. 
17. Patient has hypersensitivity to statins; or has used any statins, 
fibrates, > 1 g/day of niacin, or gemfibrozil within the 3 months 
prior to the Screening Visit. 
18. Patient reports current chronic or frequent use of drugs known 
to cause constipation (e.g. narcotics) for the 3 months prior to 
Screening. 
19. Patient has taken over-the-counter IBS treatments (e.g. 
laxatives) or proton pump inhibitors within 3 days prior to the 
Screening Visit. 
20. Certain drugs used for the treatment of IBS (e.g. low dose 
tricyclic antidepressants) may be allowed at the discretion of 
the Medical Monitor provided the patient remains on a stable 
dose for one month prior to the Screening Visit and throughout 
the study with the exception of tegaserod, lubiprostone, 


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linaclotide, metoclopramide, prucalopride, domperidone, 
plecanatide, CandiBactin, Atrantil, Allimax/Allimed within 2 
weeks prior to the Screening Visit or antibiotics within 2 
months prior to the Screening Visit. 
21. Patient has used an opioid chronically or frequently within the 
3 months prior to the Screening Visit and for the duration of 
the study. 
22. Patient is currently enrolled in, or plans to enroll in, another 
clinical study or has used any investigational drug or device 
within 1 month before signing the ICF through the completion 
of the study. 
23. Patient has previously participated in a SYN-010 study. 
24. Patient has a history of alcohol or drug abuse within the 12 
months prior to the Screening Visit. 
Additional Screening and 
Inclusion 
During the Screening and Pre-treatment Period, patients will be 
provided an ID number and password-protected access to the REDCap 
website to record Baseline parameters (such as weekly average worst 
abdominal pain score and weekly number of CSBMs). During this 
period, patients will also (i) complete a lactulose breath test including 
analysis of methane, hydrogen, hydrogen sulfide and carbon dioxide; 
(ii) provide a stool sample for analysis of the microbiome; and (iii) 
provide a blood sample for analysis of cytokines. Patients will be 
eligible for randomization if they have a breath methane level of ≥ 10 
ppm on the lactulose breath test and fully respond to the PRO 
questionnaires in the REDCap system on at least 11 of 14 days of the 
Screening and Pre-treatment Period. 
On Day 1, eligible patients will be randomized in a 1:1:1 ratio to receive 
study drug (SYN 010 21 mg, SYN-010 42 mg, or placebo). Study drug will 
be administered during the Treatment Period once daily at bedtime for 
up to 12 weeks. Rescue medication, bisacodyl, magnesium citrate, 
magnesium hydroxide, or lactulose, will be allowed for severe 
constipation (i.e. when symptoms become intolerable).  Dosage and 
frequency should be followed according to package insert. For the 
Randomization Visit only, rescue medication will not be allowed on the 
day before, the day of, or the calendar day after the Randomization 
Visit. 
Patients will be asked to refrain from starting a new diet, changing 
stable dose of supplemental fiber and/or changing their exercise 
patterns that may affect IBS-C symptoms from the time of Screening 
throughout the duration of the study. Patients will also be asked to 
refrain from taking food products that are strong inhibitors of 
cytochrome P450 3A (CYP3A) (e.g. grapefruit juice, Seville orange juice, 
St. John’s Wort), from the time of Screening throughout the duration 
of the study. 


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Treatment Schedule and Dose 
Adjustment and 
Management 
Oral administration of one (1) capsule each day at bedtime. 
Criteria for Evaluation  
Study Endpoints 
Three analysis populations are defined for this study. The Intent to 
Treat (ITT) patient population will consist of all patients who are 
randomized. The full analysis set will consist of all randomized patients 
who receive at least 1 dose of study drug. The full analysis set will be 
used for the efficacy and safety analyses. Efficacy analysis will be 
conducted according to the randomized treatment the patients 
receive, and safety analysis will be conducted according to the actual 
treatment the patients receive. 
The per-protocol set will consist of all patients in the full analysis set 
who do not have major protocol deviations as specified in the statistical 
analysis plan. 
Plan for Statistical Analysis 
Efficacy: 
The analysis for the change from baseline in weekly number of CSBMs 
will be performed using the analysis of covariance (ANCOVA) with 
treatment group and the Baseline value as covariates using the ITT 
analysis set. The primary analysis will be conducted using the observed 
values. For missing data, two sensitivity analyses will be conducted by 
using the multiple imputation and last observation carried forward 
(LOCF) methods. Details will be included in the statistical analysis plan 
(SAP).   
For secondary and exploratory analyses, the Chi-squared test will be 
used to determine differences between treatment groups for 
categorical variables. For continuous-type variables, analysis of 
covariance (ANCOVA) methods will be used with treatment group and 
the corresponding baseline value as covariates. For the responder 
endpoints, the analysis will be conducted by imputing the missing data 
as “non-responders”. A sensitivity analysis will be conducted by 
imputing the missing data as “responders”.  All statistical tests will be 
conducted at a 2-sided significance level of 5% unless otherwise 
specified. 
Due to the COVID-19 pandemic, study enrollment was 
temporarily stopped, at which time 53 patients had been 
randomized to study drug treatment groups.  A futility interim 
analysis will be conducted on these 53 patients (49 have 
completed the study and 4 have withdrawn/early terminated).  
Safety: 
All AEs will be analyzed in terms of descriptive statistics and qualitative 
analysis. Adverse events will be listed for each patient and summarized 
by system organ class (SOC) and preferred term (PT) using the Medical 
Dictionary for Regulatory Activities (MedDRA). Safety laboratory data, 
ECGs, physical examination findings and vital signs measurements will 
be listed for each patient and presented descriptively by treatment 
group time points. 


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2. General information 
STUDY TITLE:  Evaluation of the EfficAcy and Safety of SinglE, Daily Oral Doses of SYN-010 Compared 
to Placebo in Adult Patients with Irritable Bowel Syndrome with Constipation (EASE-DO) 
SPONSOR INSTITUTION: Cedars-Sinai Medical Center – Medically Associated Science and Technology 
(MAST) 
INDICATION: Irritable Bowel Syndrome with Constipation (IBS-C) 
Investigational drug(s) and 
route of administration: 
SYN-010 21mg, 42mg, Placebo  
Study purpose: 
Proof-of-concept, dose finding, health economics 
Clinical study phase: 
II 
Version no.: 
3.0 
Date: 
9 Jan. 2019     
Amendment no.: 
23411 
Date: 
09 Apr 2019     
Institution study no.: 
54792 
The study will be conducted in compliance with the protocol, ICH-GCP and any applicable regulatory 
requirements. 
Confidential 
This protocol submitted to Synthetic Biologics is the property of the investigator named as Principal 
Investigator within this document. The information contained herein is confidential and is intended 
solely for the guidance of the clinical investigation and submission to Synthetic Biologics for an appeal 
for financial or clinical drug supply support. Reproduction or disclosure of this document whether in 
part or in full   to parties not associated with the clinical investigation, or its use for any other purpose, 
without the prior written consent of the Principal Investigator is not permitted. Any changes or 
amendments to the conduct of this protocol must be resubmitted in writing to Synthetic Biologics. 
Throughout this document, symbols indicating proprietary names (, TM) are not displayed.  Hence, the 
appearance of product names without these symbols does not imply that these names are not protected. 
Sample Size Rationale 
A total of 150 adult IBS-C patients: 50 in the placebo group and 50 in 
each of the SYN-010 21 mg and SYN-010 42 mg treatment groups.  The 
sample size calculation is based on the following assumptions: the 
weekly CSBM change from baseline is 0.5 for the placebo group, and 
1.5 for each active group, with common standard deviation of 1.5, the 
study will have 90% power for each SYN-010 group compared with the 
placebo by using the t-test with alpha=0.05. 


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Principal Investigator: 
Ali Rezaie, MD, MSc 
Director, GI Motility 
Director, Bioinformatics 
Medically Associated Science and Technology (MAST) Program 
Cedars-Sinai Medical Center 
Thalians Building 
8730 Alden Drive 
Second Floor East  
Los Angeles, CA 90048 
Phone: 310-423-0617 (Clinical) 
Phone: 310-423-1955 (Laboratory)  
Email:  ali.rezaie@cshs.org 
 
Data Management and 
Coordinating Center: 
 
 
Christine Chang, RN 
Research Nurse 
Medically Associated Science and Technology (MAST) Program 
Cedars-Sinai Medical Center 
Thalians Building 
8730 Alden Drive 
Second Floor West 
Los Angeles, CA 90048 
Phone: 310-423-0617 (Clinical) 
Phone: 310-423-1955 (Laboratory)  
Email:  christine.chang@cshs.org 
 
Rashin Sedighi, RN, PhD 
Clinical Research Coordinator 
Medically Associated Science and Technology (MAST) Program 
Cedars-Sinai Medical Center 
Thalians Building 
8730 Alden Drive 
Second Floor West 
Los Angeles, CA 90048 
Phone: 310-423-0617 (Clinical) 
Phone: 310-423-1955 (Laboratory)  
Email:  rashin.sedighi@cshs.org 
 
Signatures 
The undersigned confirm that they are the authors of this protocol and agree to conduct this trial in 
accordance with all stipulations of the protocol and in accordance with International Conference on 
Harmonisation (ICH) and all applicable local Good Clinical Practice (GCP) guidelines, including the 
Declaration of Helsinki  


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__________________________________ 
__________________ 
Ali Rezaie, MD MSc 
 
 
 
 
Date  
Principal Investigator 
 
__________________________________ 
__________________ 
Christine Chang, RN 
 
Date 
Study Coordinator 
 
 
 
 


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3. Introduction 
3.1 
Background 
Irritable bowel syndrome (IBS) is a gastrointestinal (GI) syndrome characterized by chronic abdominal 
pain and altered bowel habits in the absence of any organic cause. The symptoms of IBS not only 
adversely affect a patient's health-related quality of life (QoL), but also place a significant financial 
burden on society due to reduced work productivity and increased use of healthcare-related 
resources1,2. Patients with IBS frequently complain of abdominal bloating and increased gas 
production in the form of flatulence or belching. The prevalence in North America and Europe is 
approximately 10-15%3,4. Irritable bowel syndrome affects all ages and genders however there is a 
2:1 female predominance in North America5. Irritable bowel syndrome is classified into 4 subtypes 
based on stool pattern: IBS with constipation (IBS-C), IBS with diarrhea, mixed IBS, and un-subtyped 
IBS.  Irritable bowel syndrome with constipation is defined as the presence of hard or lumpy stools 
with ≥ 25 percent of bowel movements and loose or watery stools with < 25% of bowel movements6. 
SYN-010 is a modified release, oral formulation of lovastatin being developed for the treatment of 
IBS-C. The SYN-010 program is based predominantly on research by Dr. Mark Pimentel (Director, 
Medically Associated Science and Technology Program, Cedars-Sinai Medical Center) and 
collaborators7,8,9 hypothesizing that reduction in intestinal methane (methane) production can 
reverse constipation and improve global symptoms in IBS-C. Methane production in humans is due to 
methanogenic archaea in the intestine, predominantly Methanobrevibacter smithii (M. smithii).  
Methane, the key product of anaerobic respiration of methanogens, had been perceived to produce 
no ill effects in humans aside from gaseous distention. However, several research groups worldwide 
have shown that a significant percentage of patients with IBS-C excrete methane, and elevated 
methane production by methanogens correlates with constipation and related symptoms in both IBS-
C and chronic idiopathic constipation7,8,9,10,11,12,13. A direct causative role for methane in IBS-C was 
demonstrated in a recent case report, wherein a woman undergoing fecal microbiota transplantation 
(FMT) for C. difficile infection unknowingly received stool containing a high concentration of 
methanogens. The FMT recipient rapidly developed severe symptoms of IBS-C that were 
subsequently reversed by ablation of methane production14. 
Further details can be found in the Investigator's Brochure, which contains comprehensive 
information on the investigational product. 
3.2 
Summary of findings from pre-clinical studies with potential clinical significance 
A series of preclinical studies have sought to understand the mechanism by which methane exerts its 
effects on gastrointestinal motility. The precise mechanism remains under investigation; however, 
these studies indicate that methane may exert its actions in the intestine by modulation of the 
cholinergic pathway of the enteric nervous system15. In vivo studies in dogs and ex vivo studies using 
guinea pig ileal strips showed that methane stimulates non-propagating contractions, preventing 
propulsion of materials through the intestinal tract16,17. 
The active ingredient of SYN-010 is lovastatin lactone, which was first approved by the FDA in 1987; it 
is a cholesterol-lowering agent (Mevacor®, Merck & Co., Inc.)18 and has a history of safe use at oral 
doses up to 80 mg/day. In its cholesterol-lowering indication, lovastatin lactone is a prodrug and 
must be converted to the β-hydroxyacid form to inhibit the enzyme HMG-CoA reductase (HMGR). 
The preclinical safety of lovastatin lactone and β-hydroxyacid was established in detailed preclinical 
and toxicology studies conducted by Merck19. 


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Interest in lovastatin as a potential inhibitor of methanogenesis originated with recognition that the 
rate-limiting step in the synthesis of lipid membranes in the methanogen M. smithii is catalyzed by 
HMGR. As detailed below, it has since been determined that HMGR is not the target for lovastatin 
anti-methanogenic activity and the methane reducing effects of SYN-010 can be separated from 
potential systemic effects such as cholesterol lowering. 
The ability of lovastatin to inhibit methane production was established in ex vivo studies investigating 
the effects of nine statins (including lactone and β-hydroxyacid forms) on methane production by 
stool samples from IBS-C patients. In these studies, lovastatin lactone was identified as the only 
effective statin inhibitor of methane production, while the cholesterol-lowering metabolite lovastatin 
β-hydroxyacid was ineffective. In fact, the β-hydroxyacid forms of pravastatin, simvastatin, 
rosuvastatin and atorvastatin were all ineffective inhibitors of methane production in his system. 
These experiments also demonstrated the dose-dependence of methane inhibition by lovastatin 
lactone and a head-to-head comparison of lovastatin lactone, lovastatin lactone-diol and lovastatin β-
hydroxyacid affirmed that the lactone form of lovastatin is the anti-methanogenic species20. 
An additional important feature of lovastatin as a methane lowering agent is that neither lovastatin 
lactone nor the β-hydroxyacid appear to be microbicidal. Neither species eradicated M. smithii or 
bacteria in the intestine of rats administered daily oral doses of each compound for 10 days21. 
3.3 
Rationale of the study 
SYN-010 has previously been evaluated in consecutive Phase 2a clinical trials. Sixty-three (63) IBS-C 
patients with high breath methane (>10 ppm) at screening were enrolled in a multicenter, 
randomized, controlled, double-blinded clinical trial (RCT) in which they received SYN-010 21 mg, 
SYN-010 42 mg or Placebo once daily for 4 weeks. Fifty-four (54) subjects who completed the RCT 
continued into an open-label extension (EXT) in which all subjects received SYN-010 42 mg once daily 
for an additional 8 weeks. 
The SYN-010 Phase 2a studies were intentionally designed as mechanistic proof-of-concept studies, 
wherein reductions in breath methane were employed as a rapid and cost-effective means by which 
to determine if SYN-010 could be effective in treating an underlying cause of symptoms in IBS-C. 
Breath methane was reduced relative to baseline in SYN-010 treatment groups, and lower breath 
methane levels correlated with an increased number of complete spontaneous bowel movements 
(CSBMs) at week 12, consistent with the proposed methane-inhibiting action of lovastatin lactone22. 
Since lovastatin has not previously been used to treat IBS-C patients, the SYN-010 Phase 2a studies 
were also focused on the safety of the SYN-010 dosage form. Daily doses of SYN-010 were well-
tolerated by IBS-C patients over the 12-week treatment period (at least 8 weeks of SYN-010 42 mg). 
SYN-010 did not cause clinically meaningful or persistent changes in serum liver and muscle markers 
in IBS-C patients at daily doses of 21 mg and 42 mg. Modest decreases from baseline in lipid 
parameters observed after 7 days of SYN-010 21 mg or 42 mg had largely faded by 28 days and were 
not evident after 12 weeks of dosing23. Very few adverse events were reported over 12 weeks of SYN-
010 treatment and all were of mild or moderate intensity. No serious adverse events were reported 
and there were no incidences of drug-related diarrhea, which is an important potential benefit of 
SYN-010 as an IBS-C therapy. 
Although the Phase 2a studies were not prospectively powered for formal statistical evaluation of 
clinical endpoints, compelling improvements in CSBMs, abdominal pain, and bloating were observed 
in SYN-010 treatment groups. These clinical findings have been presented in multiple public forums 


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and a panel of clinical advisors affirmed that the Phase 2a data validate the need to evaluate optimal 
dosing of SYN-010 in a larger patient population over a longer dosing period. 
Based on the potential clinical benefit observed in Phase 2a, this Phase 2b clinical study will evaluate 
in more detail the clinical effects of two dose strengths of SYN-010 administered over a longer 
treatment period (12 weeks per FDA guidelines) to a larger number of IBS-C patients. The study will 
seek more definitive evidence regarding potential symptom improvements and safety in IBS-C 
patients. The study also seeks to provide new information of relevance to both efficacy and safety by 
measuring changes to the microbiome in patient stool samples; serum level of cytokine markers of 
inflammation; and expanded breath gas measurements. 
3.4 
SYN-010 (lovastatin lactone modified release capsules) 
3.4.1 
Description 
SYN-010 is a hydroxypropyl methylcellulose (HPMC) capsule filled with enteric-coated tablets from 
which lovastatin is released at different intestinal pH values. The tablets are designed to pass 
through the stomach unchanged then release a small amount of lovastatin into the duodenum and 
the majority of the lovastatin dose into the ileocecal junction and colon.  The amount of lovastatin 
to be released into the small and large intestine is anticipated to be consistent with the relative 
levels of methane-producing archaea in each location of the intestine24. 
Each tablet in the SYN-010 dosage form contains 7 mg of lovastatin combined with USP excipients 
and coated with two different EUDRAGIT® enteric polymers that dissolve at either pH 5.5 
(duodenal release; DR) or pH 7.0 (ileocecal release; ICR). 
• 
SYN-010, 21 mg (D1-I2) comprises an opaque, green, size 1 HPMC capsule containing 1 x pH 
5.5-coated tablet (DR) and 2 x pH 7.0-coated tablets (ICR). 
• 
SYN-010, 42 mg (D1-I5) comprises an opaque, green, size 1 HPMC capsule containing 1 x pH 
5.5-coated tablet (DR) and 5 x pH 7.0-coated tablets (ICR). 
3.4.2 
Mechanism of action 
Methane production (methanogenesis) is a ubiquitous process in the human intestine, disposing of 
hydrogen and other by-products formed during bacterial fermentation. Methane production in 
humans is almost entirely due to the archeon M. smithii. Elevated intestinal methane production 
reduces intestinal motility and is a cause of constipation, pain and bloating in IBS-C.  
Interest in lovastatin as a potential inhibitor of methanogenesis originated with recognition that 
the rate-limiting step in the synthesis of archaeal lipid membranes is catalyzed by HMG-CoA 
reductase (HMGR) which is the target enzyme for cholesterol-lowering statins. Subsequent in vitro 
studies (described above) and computational studies have since determined that HMGR is not the 
target for lovastatin anti-methanogenic activity; rather, lovastatin lactone appears to exert a direct 
effect on methanogenesis enzymes. The mechanism by which lovastatin lactone inhibits methane 
production by M. smithii has not been conclusively determined; however, detailed computational 
studies showed that lovastatin lactone may competitively inhibit F420-dependent 
methylenetetrahydromethanopterin dehydrogenase (mtd), an enzyme that is integral to the M. 
smithii methanogenesis pathway25. 


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Methanogenic archaea reside predominantly in the human colon, with lower methanogen levels 
measured in the small intestine of some patients. Methane production at both sites contributes to 
reduced gastrointestinal motility and rat studies suggest that the ileocecal region may be of 
particular significance21. SYN-010 utilizes a dual-pulse release profile to deliver a portion of the 
lovastatin dose to the small intestine and the majority of the dose to the ileocecal junction and 
colon where the most methane-producing organisms are found; the relative amounts of lovastatin 
released into the small and large intestine are consistent with the anticipated relative levels of 
methanogens in each location. 
Lovastatin lactone exerts its methane-reducing effect in the intestinal lumen and systemic 
absorption of lovastatin is not required for its therapeutic effect in IBS-C. The pharmacokinetic 
profile of SYN-010 has been evaluated in a repeated-dose study in heathy volunteers. Two of the 
most therapeutically relevant findings from the PK study were: 
• 
Conversion of the anti-methanogenic lovastatin lactone to the cholesterol-lowering β-
hydroxyacid metabolite was significantly reduced in the SYN-010 formulation, and this was 
reflected in reduced systemic levels of the β-hydroxyacid and an absence of clinically 
meaningful or persistent SYN-010 effects on serum liver, lipid and muscle markers; 
• 
Conversion of the anti-methanogenic lovastatin lactone to the cholesterol-lowering β-
hydroxyacid metabolite was significantly reduced in the SYN-010 formulation, and this was 
reflected in reduced systemic levels of the β-hydroxyacid and an absence of clinically 
meaningful or persistent SYN-010 effects on serum liver, lipid and muscle markers. 
In addition to the pharmacokinetic study, sparse PK sampling was conducted in IBS-C patients 
during Phase 2a clinical testing. After daily administration of SYN-010 21 mg or SYN-010 42 mg, 
plasma trough levels of both lovastatin lactone and β-hydroxyacid were low and variable, such that 
≥ 50% of patients had undetectable plasma levels of each lovastatin species after both 7 and 28 
days of treatment23. Reduced systemic levels of lovastatin species and high concentrations of 
lovastatin lactone in the stool are compelling findings regarding the use of SYN-010 as an intestine-
acting anti-methanogenic therapy. 
4. Study objectives 
4.1 
Primary 
Change from baseline in the weekly average number of CSBMs during the 12-week Treatment 
Period for SYN-010 relative to placebo. 
4.2 
Secondary 
Safety and tolerability of SYN-010 21 mg and 42 mg in IBS-C patients treated for 12 weeks. 
For SYN-010 relative to placebo: 
• 
Proportion of overall stool frequency responders. 
• 
Proportion of overall bloating responders. 
• 
Proportion of patients using rescue medication. 


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• 
Proportion of overall responders in the SYN-010 21 mg and 42 mg treatment groups versus 
placebo. 
• 
Change from baseline for SYN-010, relative to placebo, in: 
o Weekly average score for worst abdominal pain at Weeks 1 through 12. 
o Weekly average number of CSBMs at Weeks 1 through 12. 
o Weekly average number of SBMs at Weeks 1 through 12. 
o Weekly average bloating score at Weeks 1 through 12. 
o Weekly average stool consistency (BSFS) at Weeks 1 through 12. 
4.3 
Exploratory 
For SYN-010 relative to placebo: 
• 
Proportion of patients with adequate relief27. 
• 
Change from baseline for SYN-010 in patient reported outcomes (PRO) using validated 
questionnaires in the REDCap system. 
• 
Breath gas area-under-the-curve (AUC) (methane, hydrogen, and hydrogen sulfide) on 
lactulose breath test at EOS compared to Screening. 
• 
Spot breath tests at Randomization, Day 28, and Day 56. 
• 
Comparison of serum cytokine levels at baseline and EOS – measuring for potential immune 
or inflammatory signals to include the key cytokines of the Th1/Th2/Th17 pathways:  IL-2, IL-
4, IL-5, IL-9, IL-10, IL-13, IL-17, GM-CSF and IFN-γ. Plus TNF-α, IL-12, IL-6, the two 
chemokines IL-8 and MCP-1, and the gut cytokine IL-25. 
• 
Microbiome measurements in stool of bacteria (16S rRNA) and methanogens (qPCR). 
5. Study design and investigational plan 
5.1 
Pre-treatment period 
Screening and Pre-treatment Period (up to 17 days prior to the first dose of study drug): 
During the Screening and Pre-treatment Period, patients will be provided an ID number and 
password-protected access to the REDCap website to record baseline parameters (such as weekly 
average worst abdominal pain score and weekly number of CSBMs). During this period, patients will 
also (i) complete a lactulose breath test including analysis of methane, hydrogen, hydrogen sulfide, 
and carbon dioxide; (ii) provide a stool sample for analysis of the microbiome; and (iii) provide a 
blood sample for analysis of cytokines and baseline tests. Patients will be eligible for randomization 
if they have a breath methane level of ≥ 10 ppm on the lactulose breath test and fully respond to 
the PRO questionnaires in the REDCap system on at least 11 of 14 days of the Screening and Pre-
treatment Period. 
5.2 
Treatment phase 
Treatment Period comprising Randomization (Day 1) and 12 weeks of dosing (with clinic visits at 
Weeks 4, 8, and 12). 
On Day 1, eligible patients will be randomized in a 1:1:1 ratio to receive study drug (SYN-010 21 mg, 
SYN-010 42 mg, or placebo). Study drug will be administered during the Treatment Period and taken 
once daily at bedtime for up to 12 weeks. 


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5.3 
Post-treatment period or follow-up period 
End-of-Study contact between 24 and 48 hours after Week 12 for patients who complete the 
Treatment Period. 
6. Statistical methods and determination of sample size 
6.1 
General considerations 
A statistical analysis plan (SAP) describing all the statistical analyses to be completed will be finalized 
prior to database lock. Unless otherwise specified, for numeric data, descriptive statistics will 
include the number of patients with data to be summarized (n), mean, standard deviation (SD), 
median, minimum (min), and maximum (max). Categorical data will be presented using frequency 
counts and percentages. All statistical tests will be conducted at a 2-sided significance level of 5% 
unless otherwise specified. 
6.2 
Disposition of patients 
Patient disposition will be summarized for the study, along with reasons for early termination. 
Numbers (%) of patients in each analysis population will be tabulated. 
6.3 
Protocol deviations 
All protocol deviations will be reviewed and documented before database lock. Protocol deviations 
will be recorded by the site staff, and study monitor. Major protocol deviations will be defined in the 
SAP before unblinding the study. 
6.4 
Analysis populations 
Three analysis populations are defined for this study:  
• 
The randomized patient population will consist of all patients who are randomized; 
• 
The full analysis set will consist of all randomized patient who receive at least one dose of 
study drug. The full analysis set will be used for the efficacy and safety analyses. Efficacy 
analysis will be conducted according to the randomized treatment the patients receive, and 
safety analysis will be conducted according to the actual treatment the patients receive; 
• 
The per-protocol set will consist of all patients in the full analysis set who do not have major 
protocol deviations as defined in the SAP. 
6.5 
Demographics and baseline characteristics 
Demographics and baseline characteristics will be summarized by treatment for the full analysis set. 
Demographics and baseline characteristics of the patient sample will be described using summary 
statistics. 
6.6 
Efficacy analyses 
The primary endpoint is the change from baseline in the weekly average number of CSBMs during 
the 12-week Treatment Period. 
The secondary endpoints are defined as below:  


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• 
Proportion of overall responders during the 12-week Treatment Period (overall 12-week 
responders).  An overall 12-week responder is defined as a patient with a weekly response 
in at least 50% of the weeks of treatment (6 of 12 weeks).  A weekly response is defined as a 
decrease in the patient’s weekly average score for worst abdominal pain in the past 24 
hours of at least 30% compared to baseline and a stool frequency increase of 1 or more 
CSBMs per week compared with baseline. 
• 
Proportion of overall stool frequency responders during the 12-week Treatment Period. An 
overall stool frequency responder is defined as a patient with a weekly stool frequency 
response in at least 50% of the weeks of treatment (6 of 12 weeks). A weekly stool 
frequency response is defined as a stool frequency increase of 1 or more CSBMs per week 
compared with baseline, with abdominal pain unchanged or improved compared with 
baseline. 
• 
Proportion of overall abdominal pain intensity responders during the 12-week Treatment 
Period. An overall abdominal pain intensity responder is defined as a patient with a weekly 
abdominal pain intensity response in at least 50% of the weeks of treatment (6 of 12 
weeks). A weekly response abdominal pain intensity response is defined as a decrease in the 
patient’s weekly average score for worst abdominal pain in the past 24 hours of at least 30% 
compared to baseline, with stool frequency unchanged or improved compared with 
baseline. 
• 
Proportion of overall bloating responders during the 12-week Treatment Period. An overall 
bloating responder is defined as a patient with a weekly bloating response in at least 50% of 
the weeks of treatment (6 of 12 weeks). A weekly bloating response is defined as a weekly 
average bloating score of at least 30% improvement compared to baseline, with stool 
frequency unchanged or improved compared with baseline. 
• 
Proportion of patients using rescue medication. 
Additional secondary endpoints are change from baseline.  The change from baseline at Visit X is 
defined as Value at Visit X – Value at Baseline. The percent change from baseline is defined as 
(change from baseline)/baseline value. If the baseline value = 0 or missing, then the percent change 
from baseline will be missing. 
• 
Change from baseline in weekly average score for worst abdominal pain at Weeks 1 through 
12. 
• 
Percent change from baseline in weekly average score for worst abdominal pain at Weeks 1 
through 12. 
• 
Change from baseline in weekly average number of CSBMs at Weeks 1 through 12. 
• 
Change from baseline in weekly average number of SBMs at Weeks 1 through 12. 
• 
Change from baseline in the weekly average bloating score at Weeks 1 through 12. 
• 
Change from baseline in weekly stool consistency score (BSFS) at Weeks 1 through 12. 
• 
Proportions of overall abdominal pain intensity responders with different cutoff points for 
the % improvement from baseline in weekly average score for worst abdominal pain, i.e., ≥ 
10%, 20%, 40%, 50%, 60% and 70%. 
• 
Proportions of overall stool frequency responders with different cutoff points for the 
improvements from baseline in the weekly number of CSBMs ≥ 2, 3, 4, 5, and 6. 
• 
Proportions of patients with improvements of ≥ 10%, 20%, 30%, 40%, 50%, 60%, and 70% 
from baseline in the weekly average bloating score in at least 50% of the weeks of 
treatment (6 of 12 weeks). 


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The exploratory endpoints are defined as below:  
• 
Proportion of patients with adequate relief26. 
• 
Change from baseline for SYN-010 in patient reported outcomes (PRO) using validated 
questionnaires in the REDCap system. 
• 
Change from baseline in the area-under-the-curve (AUC) of breath CH4 production, based on 
the 120-minute lactulose breath test. 
• 
Change from baseline in breath CH4 production based on a single-point breath CH4 test. 
The efficacy analyses will be conducted based on the full analysis set and per-protocol set. The 
analysis for the primary endpoint will be performed using the analysis of covariance (ANCOVA) with 
treatment group and the baseline value as covariates. The primary analysis will be conducted using 
the observed values. For missing data, two sensitivity analyses will be conducted by using the 
multiple imputation and last observation carried forward (LOCF) methods. Details will be included in 
the SAP.   
For missing data imputation, the primary analysis will be conducted based on the following 
imputation method: If a subject drops out of the study or otherwise does not report efficacy data for 
a particular treatment period week, the subject will not be considered a responder for that week. 
The weekly response will not be considered a responder if there are more than three days with 
missing assessments during a week.  
For the responder endpoints, the analysis will be conducted by imputing the missing data as “non-
responders”. Two sensitivity analyses for the primary endpoint will be conducted: (1) missing data 
for the responders will be imputed as “responders”; (2) the overall responders are defined based on 
the last observation carried forward (LOCF) for missing values of the weekly average score for worst 
abdominal pain and/or weekly average number of CSBMs. 
For the analyses of the secondary and exploratory endpoints, the Chi-squared test will be used to 
determine differences between each active treatment group versus placebo group for categorical 
variables. P-values and 95% Newcombe-Wilson confidence intervals (CIs) for treatment differences 
will be presented. For continuous-type variables, analysis of covariance (ANCOVA) methods will be 
used with treatment group and the corresponding baseline value as covariates. P-values and least 
squares (LS) treatment means and their differences will be presented, accompanied by 95% CIs for 
each time point.   
A fixed sequence testing procedure will be used for the statistical testing of the primary and 
secondary endpoints. More specifically, the primary endpoint will be tested at the two-sided alpha = 
0.05 level first. If statistical significance is not achieved for the primary endpoint, then the testing 
procedure will stop, and the secondary endpoints will not be tested formally. If statistical 
significance is achieved for the primary endpoint, then the secondary endpoints will be tested 
formally in the order of the list in section 4.2. The first listed secondary endpoint will be tested at 
the two-sided alpha = 0.05 level. If statistical significance is not achieved, then the testing procedure 
will stop. If statistical significance is achieved for the first listed secondary endpoint, then the 
following secondary endpoint will be tested at the two-sided alpha = 0.05 level. The testing 


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procedure will continue for the testing of the secondary endpoints until statistical significance is not 
achieved for a secondary endpoint. 
Due to the COVID-19 pandemic, study enrollment was temporarily stopped, at which time a total of 
53 patients had been randomized to study drug treatment groups. 49 of these patients have 
completed the study and 4 have withdrawn or early terminated.  A futility interim analysis will be 
conducted based on the data from these 53 patients.  A recommendation to stop enrollment of the 
study will be made by the unblinded statistician(s) if, for both dose groups, the conditional power is 
less than 20% based on the primary endpoint AND the difference of the overall 12-week responder 
rates is less than 2%.  The interim analysis will be conducted by a group of people that are not 
involved in the daily activities of the study.  Details will be included in the SAP. 
6.7 
Safety analyses 
All AEs, whether considered drug-related or not, will be reported with a diagnosis, start/stop dates, 
action taken, whether treatment was discontinued, any corrective measures taken, outcome, and 
other possible causes.  For all events, the relationship to treatment and the intensity of the event 
will be determined by the investigator. 
All AEs will be analyzed in terms of descriptive statistics and qualitative analysis. Treatment 
emergent adverse events will be listed for each patient and summarized by system organ class (SOC) 
and preferred term (PT) using the Medical Dictionary for Regulatory Activities (MedDRA). 
Safety laboratory data, ECGs, physical examination findings, and vital signs measurements will be 
listed for each patient and presented descriptively by treatment group and time point. 
Safety variables may include but not limited to the following: laboratory changes (complete blood 
counts, electrolytes, chemistry, and coagulation), changes in vital signs (blood pressure, heart rate, 
respiratory rate, and temperature), and ECG and, in some instances, as produced at the 
investigator’s discretion. 
6.8 
Determination of sample size 
The sample size calculation is based on the following assumptions: the weekly CSBM change from 
baseline is 0.5 for the placebo group, and 1.5 for each active group, with common standard 
deviation of 1.5; 50 patients in each group will have 90% power for each SYN-010 group compared 
with the placebo by using the t-test with alpha=0.05. 
7. Study population 
7.1 
Study population and statistical assumptions 
A total of 150 adult IBS-C patients will be enrolled in a single site: 50 in the Placebo group and 50 in 
each of the SYN-010 21 mg and SYN-010 42 mg treatment groups.  The sample size calculation is 
based on the t-test to achieve 90% power for each SYN-010 group when compared with the placebo 
group. 


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7.2 
Eligibility criteria 
Patients must meet all of the following inclusion and exclusion criteria to be included in the study. 
7.2.1 
Inclusion criteria 
1. 
Male or female participants aged between 18 and 65 years inclusive. 
2. Patient must be willing and able to participate in the study for the required duration, 
understand and sign the informed consent form (ICF), and be willing to comply with all 
protocol-related visits and procedures. 
3. Patient has had IBS-C symptoms (as defined by Rome IV diagnostic criteria) for at least 6 
months prior to diagnosis.  
4. Patient has an average score of ≥ 3.0 for daily abdominal pain at its worst (11-point numerical 
rating scale [NRS]) during and up to the 17 days immediately before randomization (i.e. Pre-
treatment Period). 
5. Patient has an average of ≤ 3 CSBMs per week or ≤ 5 SBMs per week during the 17 days 
immediately before randomization (i.e. Pre-treatment Period). 
6. Patient has a breath methane level ≥ 10 ppm on a lactulose breath test administered at 
Screening. 
7. Patient may be on a stable, continuous regimen of fiber or probiotics one month before the 
Screening Visit; however, they must maintain a stable dose regimen through Week 12. 
8. Patient must agree to refrain from starting a new diet, changing stable dose of supplemental 
fiber, or changing exercise pattern that may affect IBS-C symptoms from the time of 
Screening through the end of the study. If the patient takes food products that are strong 
inhibitors of cytochrome P450 3A (CYP3A) (e.g. grapefruit juice, Seville orange juice, St. John’s 
Wort), he/she must agree to refrain from taking these from the time of Screening through 
the end of the study. 
9. Patient must agree to use an acceptable method of contraception from the time of signing 
the ICF to 30 days after the final dose of study drug if the patient is a sexually active female of 
child-bearing potential (defined as any female who has experienced menarche and who is 
NOT permanently sterile or postmenopausal. Postmenopausal is defined as 12 consecutive 
months with no menses without an alternative medical cause). Adequate contraceptive 
measures include: oral contraceptives (stable use for two or more cycles before Screening); 
intrauterine device; Depo-Provera®; Norplant® System implants; partner with a vasectomy; 
double-barrier birth control (e.g. use of a condom plus diaphragm or condom plus either 
contraceptive sponge, foam, or jelly); or abstinence. According to drug research standards, 
male patient must agree to use an acceptable method of contraception and refrain from 
donating sperm from the time of signing the ICF to 90 days after the final dose of study drug. 
10. A female patient of child-bearing potential must be non-pregnant and non-lactating and have 
negative pregnancy tests at the Screening Visit and on Day 1 prior to dosing with study drug. 
11. Patients must be able to understand and be willing to sign the written informed consent 
form. A signed informed consent form must be appropriately obtained prior to the conduct 
of any trial-specific procedure. 
12. Willing and able to comply with the protocol, including follow-up visits and examinations. 


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7.2.2 
Exclusion criteria 
Patients who meet any of the following criteria will be excluded from this study: 
1. Patient has loose (mushy) or watery stools for > 25% of their bowel movements (BMs) during 
the 12 weeks before Screening or during the Screening and Pre-treatment Periods. 
2. Patient has a history of cathartic colon, laxative, or enema abuse. 
3. Patient has a history of ischemic colitis. 
4. Patient has a history of pelvic floor dysfunction. 
5. Patient has a history of bariatric surgery for the treatment of obesity. 
6. Patient has a history of surgery to remove a segment of the gastrointestinal (GI) tract at any 
time before the Screening Visit. 
7. Patient has any history of myopathy, rhabdomyolysis, chronic myalgia, or familial history of 
hereditary muscular disorders. 
8. Patient has been diagnosed with or has a family history of familial adenomatous polyposis, 
hereditary nonpolyposis colorectal cancer, or any other form of familial colorectal cancer. 
9. Patient currently has any structural abnormality of the GI tract or a disease or condition that 
can affect GI motility, or any unexplained and clinically significant symptoms such as lower GI 
bleeding, rectal bleeding, heme-positive stool, iron-deficiency anemia, weight loss, or 
systemic signs of infection. 
10. Patient has had any previous surgery involving the abdomen, pelvis, or retroperitoneal region 
during the last 12 months prior to Screening, with the exclusion of laparoscopic gallbladder 
surgery or appendix removal. 
11. Patient has a history of diverticulitis or any chronic condition that could be associated with 
abdominal pain or discomfort and could confound the assessments in the study (e.g. 
inflammatory bowel disease, chronic pancreatitis, polycystic kidney disease, ovarian cysts, 
endometriosis, or lactose intolerance). 
12. Patient has history of severe renal insufficiency defined as an actual or estimated glomerular 
filtration rate of < 30 mL/min/1.73 m2 within the 6 months prior to Screening Visit. 
13. Patient has any history of a medical condition or a concomitant medical condition that, in the 
opinion of the investigator, would compromise the patient’s ability to complete the study 
safely or could confound the assessments in this study (e.g. uncontrolled hypothyroidism). 
14. Patient is known to have elevated liver enzyme levels (aspartate aminotransferase [AST], 
alanine aminotransferase [ALT], alkaline phosphatase [ALP]) or creatine kinase levels that are 
≥ 1.5 times the upper limit of normal (ULN) that have not been resolved within the 4 weeks 
prior to consent and/or these elevated levels are present at the Screening Visit laboratory 
assessment. 
15. Patient has any abnormal laboratory results, electrocardiogram (ECG) findings, or physical 
examination findings deemed clinically significant by the investigator during the Screening 
Period. 
16. Within 14 days prior to the Screening Visit, patient has used concomitant medications that 
are: (1) moderate-to-strong inhibitors of cytochrome P450 3A (CYP3A) and/or organ anion 
transporting polypeptide (OATP)1B1 (e.g. cyclosporine, verapamil, dronedarone, diltiazem, 
amiodarone, itraconazole, ketoconazole, posaconazole, voriconazole, clarithromycin, 
telithromycin, human immunodeficiency virus protease inhibitors, boceprevir, telaprevir, 
nefazodone, erythromycin, cobicistat-containing products); (2) other concomitant 
medications that are excluded from the lovastatin label (e.g. rifampin, colchicine, ranolazine); 
or (3) metformin or GLP-1 agonists. Patients should not take any of these concomitant 


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medications during the treatment phase of the study without contacting the investigator. 
17. Patient has hypersensitivity to statins; or has used any statins, fibrates, > 1 g/day of niacin, or 
gemfibrozil within the 3 months prior to the Screening Visit. 
18. Patient reports current chronic or frequent use of drugs known to cause constipation (e.g. 
narcotics) for the 3 months prior to Screening. 
19. Patient has taken over-the-counter IBS treatments (e.g. laxatives) or proton pump inhibitors 
within 3 days prior to the Screening Visit. 
20. Certain drugs used for the treatment of IBS (e.g. low dose tricyclic antidepressants) may be 
allowed at the discretion of the Medical Monitor provided the patient remains on a stable 
dose for one month prior to the Screening Visit and throughout the study with the exception 
of tegaserod, lubiprostone, linaclotide, metoclopramide, prucalopride, domperidone, 
plecanatide, CandiBactin, Atrantil, Allimax/Allimed within 2 weeks prior to the Screening Visit 
or antibiotics within 2 months prior to the Screening Visit. 
21. Patient has used an opioid chronically or frequently within the 3 months prior to the 
Screening Visit and for the duration of the study. 
22. Patient is currently enrolled in, or plans to enroll in, another clinical study or has used any 
investigational drug or device within 1 month before signing the ICF through the completion 
of the study. 
23. Patient has previously participated in a SYN-010 study. 
24. Patient has a history of alcohol or drug abuse within the 12 months prior to the Screening 
Visit. 
7.3 
Excluded therapies and medications, previous and concomitant 
• 
Prior use of SYN-010. 
• 
Concurrent use of another investigational drug or device therapy (i.e. outside of study 
treatment) during or within 4 weeks of trial entry (signing of the informed consent form). 
• 
Major surgery within 30 days prior to start of study drug. 
7.4 
Removal of patients from therapy or assessment 
The investigator may discontinue treating a patient with a study drug or withdraw the patient from 
the study at any time for safety or administrative reasons. The patient may decide to discontinue 
study drug or withdraw from the study at any time for any reason. The reason for discontinuation 
must be documented. 
7.5 
Withdrawal of patients from the study 
Patients must be withdrawn from the trial (treatment and procedures) for the following reasons:  
• 
Patient withdraws consent from study treatment and study procedures. A patient must be 
removed from the trial at his/her own request or at the request of his/her legally acceptable 
representative. At any time during the trial and without giving reasons, a patient may 
decline to participate further. The patient will not suffer any disadvantage as a result. 
• 
Patient is lost to follow-up. 
• 
Death. 
• 
Pregnancy. Lovastatin is a pregnancy Category X drug and should not be taken during 
pregnancy. Female subjects who become pregnancy must immediately discontinue taking 
the study drug. Subjects will be instructed that known or suspected pregnancy occurring 


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during the study should be confirmed and reported to the investigator, who will then 
withdraw the female subjects from the study without delay. Protocol-required procedures 
for study discontinuation and follow-up must be performed on the subject unless 
contraindicated by pregnancy. The investigator should also be notified of any pregnancy 
occurring during the study and for 30 days after completion of the study. Within 24 hours of 
awareness of any pregnancy associated with study drug exposure during the study and for 
30 days after the last dose of study drug, the investigator must record the event on the 
supplied pregnancy form and send it to the safety vendor. Any pregnancy will be followed to 
term or termination, and the status of mother and child will be reported on the pregnancy 
form after delivery. If delivery occurs after study closure, the outcome should be reported 
directly to the Sponsor. 
Patients may be withdrawn from the study for the following reasons: 
• 
The patient is non-compliant with study drug, trial procedures, or both, including the use of 
concomitant medications not prescribed by the study protocol.   
• 
If, in the investigator's opinion, continuation of the trial would be harmful to the patient's 
well-being. 
• 
The development of a secondary infection or other condition.  
• 
Development of an intercurrent illness or situation which would, in the judgment of the 
investigator, significantly affect assessments of clinical status and trial endpoints. 
• 
Use of illicit drugs or other substances that may, in the opinion of the investigator, have a 
reasonable chance of contributing to toxicity or otherwise skewing trial result. 
 
Any patient removed from the trial will remain under medical supervision until discharge or transfer 
is medically acceptable. 
In all cases, the reason for withdrawal must be recorded in the CRF and in the patient's medical 
records. 
7.6 
Screen failures 
Patients withdrawn during the Screening Period due to abnormal ECG, liver function tests, creatine 
kinase level, opiate use, or pregnancy will not be allowed to repeat the Screening Period. All other 
patients withdrawn during the Screening Period will be reviewed and evaluated on an individual 
basis and may be allowed to re-enter the Screening Period at the discretion of the investigator and 
the study team. 
8. Treatments 
8.1 
Randomization 
Randomization will occur after a questionnaire collection period of at least 14 days, with compliance 
in fully responding to questions on at least 11 of the 14 days before the start of the Treatment 
Period. 
After the successful completion of the Screening and Pre-treatment Period, eligible subjects will be 
randomly assigned in a 1:1:1 ratio to the following treatment groups: SYN-010 21 mg, SYN-010 42 
mg, or placebo. 


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Patients will be assigned to the protocol specified treatments using a unique identification number 
which will be assigned when a patient is evaluated for inclusion into the study.  
Randomization will be conducted through the research pharmacist using an online generator 
(www.sealedenvelope.com). Randomization will be in blocks of 6. 
8.2 
Investigational product 
8.2.1 
SYN-010 
SYN-010 will be supplied by Synthetic Biologics, Inc. as a 21 mg or 42 mg capsule of lovastatin in a 
modified-release formulation. 
SYN-010 is a capsule containing enteric-coated tablets of lovastatin designed to release drug in 
both the duodenal (pH 5.5) and ileocecal (pH 7.0) regions of the GI tract. 
8.3 
Comparator treatment to be administered 
8.3.1 
Placebo 
The placebo is a capsule containing enteric-coated tablets without drug product that will be 
identical in appearance to SYN-010 and packaged in identical containers. The placebo will be 
supplied by Synthetic Biologics, Inc. 
8.4 
Full description and handling of treatments 
8.4.1 
SYN-010 
SYN-010 will be supplied as a 21 mg or 42 mg capsule of lovastatin in a modified release 
formulation. SYN-010 is a green, size 1 HPMC capsule containing either 3 (21 mg) or 6 (42 mg) 
enteric-coated tablets of lovastatin (each 7 mg) designed to release drug in both the duodenal (pH 
5.5) and ileocecal (pH 7.0) regions of the GI tract. The placebo is an HPMC capsule containing 
enteric coated tablets without drug product that will be identical in appearance to SYN-010 and 
packaged in identical containers. 
 
8.4.1.1    General warning  
SYN-010 capsules should not be chewed or opened. 
The active ingredient of SYN-010 (lovastatin) is currently listed as a pregnancy category X drug 
and should not be taken during pregnancy. Female subjects who become pregnant must 
immediately discontinue taking study drug. Subjects will be instructed that known or suspected 
pregnancy occurring during the study should be confirmed and reported to the investigator, 
who will then withdraw the female subjects from the study without delay. The investigator 
should also be notified of any pregnancy occurring during the study but confirmed after 
completion of the study. 
Clinical studies with the commercially-available cholesterol-lowering lovastatin formulation 
Mevacor® (Merck) showed that lovastatin, like other inhibitors of HMG-CoA reductase, 
occasionally causes myopathy manifested as muscle pain, tenderness or weakness with creatine 
kinase (CK) above ten times the upper limit of normal. Myopathy sometimes takes the form of 


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rhabdomyolysis with or without acute renal failure secondary to myoglobinuria, and rare 
fatalities have occurred. The risk of myopathy is increased by high levels of HMG-CoA reductase 
inhibitory activity in plasma. SYN-010 is designed to limit systemic exposure of HMG-CoA 
reductase inhibiting lovastatin species and no myopathy or rhabdomyolysis were observed in 
Phase 1 or Phase 2a clinical trials with up to 12 weeks of SYN-010 42 mg dosing. Transient and 
reversible increases in serum CK were observed in two subjects treated with SYN-010 42 mg for 
8 weeks in the Phase 2a clinical trial; however, these were deemed unrelated to study drug by 
the investigator and returned to normal without discontinuation of SYN-010. 
Clinical studies with the commercially-available cholesterol-lowering lovastatin formulation 
Mevacor® (Merck) also found persistent increases (to more than 3 times the upper limit of 
normal) in serum transaminases occurred in 1.9% of adult patients who received lovastatin for 
at least one year in early clinical trials. When the drug was interrupted or discontinued in these 
patients, the transaminase levels usually fell slowly to pretreatment levels. The increases usually 
appeared 3 to 12 months after the start of therapy with lovastatin and were not associated with 
jaundice or other clinical signs or symptoms.  SYN-010 is designed to limit systemic exposure 
to lovastatin species and SYN-010 21 mg and 42 mg doses did not cause clinically meaningful or 
persistent alterations to serum transaminase levels in Phase 1 or Phase 2a clinical trials. 
8.4.1.2    Dose handling 
All study drug has been manufactured according to Good Manufacturing Practice for Medicinal 
Products and the relevant regulatory requirements. The study drug will be distributed by PCI 
Pharma Services. All packaging and labeling operations have been performed according 
to Good Manufacturing Practice for Medicinal Products and the relevant regulatory 
requirements. 
Study drug at the clinical study sites should be stored at controlled room temperature in a 
locked area accessible only to appropriate study personnel. Study drug bottles, and any unused 
study drug, will be retained by the study site until final reconciliation is completed, then 
destroyed by the study site according to local procedures. The investigator must provide an 
explanation for any missing bottles or missing capsules. Any missing bottles or capsules must be 
documented in the applicable source documentation and Accountability Log. 
8.4.1.3    Dose calculation 
SYN-010 doses were selected based on previous clinical experience in Phase 1 and Phase 2a 
clinical studies. These studies showed that SYN-010 provided continuous delivery of lovastatin 
lactone throughout the colon, and stool concentrations for the SYN-010 21 mg and 42 mg doses 
were equivalent to concentrations that inhibited methane production by ~60% and ~90% 
respectively in stool samples from an IBS-C patient in vitro. Moreover, these SYN-010 doses did 
not cause significant or persistent systemic effects on blood chemistry, hematology or 
urinalysis parameters over 12 weeks of dosing in Phase 2a clinical trials. 
8.4.1.4    Dose administration 


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The study drugs, SYN-010 21 mg or 42 mg (active) or placebo (control), will be administered 
orally once daily at bedtime for 12 weeks. 
8.4.1.5    Dose modification 
No dose adjustments will be allowed. 
8.5 
Blinding 
In compliance with applicable regulations, in the event of a SUSAR completed for patients on 
investigational drug SYN-010, the patient’s treatment code will be unblinded and reported to the 
health authorities.  SUSARs involving placebo would not be required to be submitted to the health 
authorities, however local and institutional regulations should be followed.  Any safety reports 
completed, do not need to be unblinded before submitting to other investigators.  Submission of 
safety reports to the ethics committee should follow local reporting regulations. 
8.5.1 
Emergency unblinding by the investigator 
Unblinding may occur for emergency purposes only. The investigator should note that the 
occurrence of a serious adverse event or progressive disease should not routinely precipitate the 
immediate unblinding of the label. If unblinding is necessary for the treatment of a patient for a 
serious adverse event, every attempt should be made to contact the investigator prior to 
unblinding. If this is not feasible, then the investigator must be contacted within 24 hours of 
unblinding. 
8.6 
Drug logistics and accountability 
All study drugs will be stored at the investigational site in accordance with Good Clinical Practice 
(GCP) and Good Manufacturing Practices (GMP) requirements and the instructions given by the 
clinical supplies department of the Institution and will be inaccessible to unauthorized personnel. 
8.6.1 
Storage 
All investigational product will be kept in a locked area with access limited to study staff only. The 
locked area will be maintained under controlled temperature conditions at 20oC-25oC (68oF-77oF), 
and away from light and moisture. 
8.6.2 
Accountability 
The investigator, or a responsible party designated by the investigator, must maintain a careful 
record of the inventory and disposition of the agent(s) (investigational or free of charge) using the 
Drug Accountability Record or another comparable drug accountability form.  
8.6.3 
Destruction and return 
At the end of the study, unused supplies of SYN-010 and other investigational agents should be 
destroyed appropriately and according to institutional policies. Destruction will be documented in 
the Drug Accountability Record Form. 


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8.7 
Treatment compliance 
Upon receipt of the study drug, the investigator, or a responsible party designated by the 
investigator, will conduct an inventory of the supplies and verify that study drug supplies are 
received intact and in the correct amounts. A full drug accountability log will be maintained at the 
study site at all times. All discrepancies must be accounted for and documented. 
Subjects will be expected to return all unused study drug and empty containers to the site at each 
clinic visit, and the number of capsules dispensed and returned will be documented by study staff.  
The study personnel will account for all drugs dispensed and returned. All unused supplies of SYN-
010 and other investigational agents will be destroyed appropriately and according to institutional 
policies. Destruction will be documented in the Drug Accountability Record form. 
Subjects who are not compliant with treatment protocol, including laboratory evaluations, will be 
subject to withdrawal from the study at the discretion of the investigator.  
8.8 
Prior and concomitant therapy 
Any prior or concomitant therapy that the patient may be taking and may be necessary for the 
patient’s welfare taken within two weeks prior to the start of the study and during the study must 
be recorded in the patient’s source documentation and in the CRF (including start/stop dates, dose 
frequency, route of administration, and indication). 
The patient may be on a stable, continuous regimen of fiber, or probiotics one month before the 
Screening Visit; however, he/she must maintain a stable dose regimen through Week 12. 
Additionally, certain drugs used for the treatment of IBS (e.g. low dose tricyclic antidepressants) may 
be allowed at the discretion of the Medical Monitor provided the patient remains on a stable dose 
for one month prior to the Screening Visit and throughout the study with the exception of 
tegaserod, lubiprostone, linaclotide, metoclopramide, prucalopride, domperidone, plecanatide, 
CandiBactin, Atrantil, Allimax/Allimed within 2 weeks prior to the Screening Visit or antibiotics 
within 2 months prior to the Screening Visit. 
Rescue medication will be allowed for severe constipation (i.e. when symptoms become 
intolerable). For guidance purposes for this protocol, intolerable symptoms may be when a subject 
has not had a complete spontaneous bowel movement (CSBM) in the preceding three days with an 
abdominal pain score of ≥ 6 on an 11-point (0-10) scale each day during that time period. However, 
intolerability is ultimately a subjective judgment, and principal investigators and their patients make 
the final decision regarding what is intolerable and requires use of the rescue medication. For the 
Randomization Visit only, rescue medication will not be allowed the day before, the day of, or the 
calendar day after the Randomization Visit. If rescue medication is taken during this time, it must be 
recorded as a protocol deviation and the subject should not be randomized. The use of protocol-
specified rescue medication and non-protocol-specified laxatives will be recorded in the subject 
record. 
Permitted rescue medications include bisacodyl, magnesium citrate, magnesium hydroxide, and 
lactulose. Dosage and frequency should be followed according to package insert on rescue 
medication and should not exceed recommended doses as approved by package insert. 


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9. Procedures and variables 
9.1 
Schedule of assessments  
Table 1: Schedule of assessments 
X = required; O = only if directed by symptoms or reported AE; ET = early termination 
 
 
 
Visit No. 
1 
-- 
2 
3 
4 
5 
EOS 
Type 
Screen 
Pre-treatment Randomize 
 
 
 
Call/Visit 
Days 
-17 to -14 
-17 to -1 
1 
28 
56 
84 or ET 
85 
Window (Days) 
-- 
-- 
-- 
±5 
±5 
±5 
+1 
Participant Information 
 
 
 
 
 
 
 
Informed Consent1 
X 
-- 
-- 
-- 
-- 
-- 
-- 
Inclusion/Exclusion1 
X 
-- 
-- 
-- 
-- 
-- 
-- 
IBS-C Criteria (ROME IV)1 
X 
-- 
-- 
-- 
-- 
-- 
-- 
Demographics2 
X 
-- 
-- 
-- 
-- 
-- 
-- 
Medical History3  
X 
-- 
X 
-- 
-- 
-- 
-- 
Prior/Concomitant 
Medications 
X 
-- 
X 
X 
X 
X 
X 
Diary compliance/ 
eligibility to randomize4 
-- 
X 
X 
-- 
-- 
-- 
-- 
Physical 
 
 
 
 
 
 
 
Physical Exam5 
X 
-- 
X 
O 
O 
X 
O 
Vital Signs6 
X 
-- 
X 
X 
X 
X 
O 
12-Lead ECG7 
X 
-- 
 
-- 
-- 
X 
-- 
Drug Dispensing 
 
 
 
 
 
 
 
Drug Dispensing 
-- 
-- 
X 
X 
X 
-- 
-- 
Accountability/Returns 
-- 
-- 
-- 
X 
X 
X 
-- 
Adverse Events 
 
 
 
 
 
 
 
AEs 
-- 
-- 
X 
X 
X 
X 
X 
Blood Samples 
 
 
 
 
 
 
 
Clinical Chemistry (CMP) 
X 
-- 
X 
X 
-- 
X 
-- 
Lipid Panel 
 
 
X 
 
 
X 
 
Cytokine Analysis 
-- 
-- 
X 
X 
-- 
X 
-- 
Hematology 
X 
-- 
 
 
-- 
X 
-- 
Serum Pregnancy8 
X 
-- 
-- 
-- 
-- 
-- 
-- 
Creatine Kinase (CK) 
X 
-- 
-- 
O 
O 
O 
O 
Urine Samples 
 
 
 
 
 
 
 


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Urine Drug Screen 
X 
-- 
-- 
-- 
-- 
-- 
-- 
Urine Pregnancy8 
-- 
-- 
X 
X 
X 
X 
-- 
Patient Reported 
Outcomes (REDCap) 
 
 
 
 
 
 
 
Diary Dispensing or 
Return9 
X 
-- 
-- 
-- 
-- 
X 
-- 
Diary Compliance10 
-- 
X 
X 
X 
X 
X 
-- 
EQ-5D-5L QoL 
Questionnaire11 
-- 
-- 
X 
-- 
-- 
X 
-- 
Stool Samples12 
 
 
 
 
 
 
 
Bacteria (16S rRNA) 
-- 
 
X 
-- 
-- 
X 
-- 
Methanogens (qPCR) 
-- 
 
X 
-- 
-- 
X 
-- 
Breath Gases 
 
 
 
 
 
 
 
Lactulose breath test13 
X 
-- 
-- 
-- 
-- 
X 
-- 
Single-point breath test 
-- 
-- 
X 
X 
X 
-- 
-- 
1Informed consent, inclusion/exclusion criteria, and ROME IV may be reviewed and signed prior to Visit 1.  
2Date of birth, gender, race/ethnicity. 
3Patients will complete the information in the REDCap system during the Screening Period. 
4Patients will be eligible for randomization if they fully respond to the PRO questions on at least 11 of 14 days. 
5General examination, including head, ears, eyes, nose, throat, endocrine, cardiovascular, respiratory, abdomen, 
skin, neurological, extremities, and musculoskeletal features. 
6Body weight, blood pressure (requires the patient to be semi-supine for at least 5 minutes prior to the test) height 
(Screening Visit only), radial pulse rate, respiratory rate, and temperature. 
712-lead ECG requires the participant to be supine for at least 5 minutes prior to the test. 
8All female participants of child-bearing potential. 
9Provide and train the patient to use the REDCap system with questionnaire. 
10Ongoing review and confirmation of patient PRO questionnaire compliance. Patients will complete the 
information contained in the REDCap system each day, assessing stool frequency, SBMs, CSBMs, and use of rescue 
medication or other laxative use; stool consistency using the Bristol Stool Chart; abdominal pain assessment using 
visual NRS scale (0-10) and visual Likert scale (0-4) for straining and bloating assessment. Questionnaires will also 
be completed weekly for the PAC-SYM questionnaire and Adequate Relief question. 
11EQ-5D-5L questionnaire will be completed in the REDCap system on Day 1 prior to study drug administration and 
at Week 12 or upon early termination. 
12Pre-treatment stool sample can be collected any time during the week preceding and including Day 1 and will 
only be analyzed if the patient is randomized into the study; Day 84 stool sample can be collected any time during 
the week preceding and including Day 84. 


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13Patients are fasted overnight prior to lactulose breath testing. Lactulose breath test measures hydrogen, 
methane, hydrogen sulfide, and carbon dioxide levels. 
9.2 
Timing of assessments 
 
9.3 
Assessment parameters 
Subjects who provide written consent to participate will be screened for inclusion in the study. 
Consent to participate must be obtained prior to completing any study procedures and may be 
obtained prior to the Screening Visit. 
9.3.1 
ROME IV criteria 
The investigator or designee will assess whether the subject meets the following ROME IV IBS 
criteria28: 
Subject has had IBS symptoms at least six months prior to diagnosis and is experiencing 
abdominal pain, on average, at least one day per week for the three months preceding the 
Screening Visit and associated with two or more of the following: 
• 
Abdominal pain or discomfort related to defecation; 
• 
Abdominal pain or discomfort associated with a change in frequency of stool; and/or 
• 
Abdominal pain or discomfort associated with a change in form (appearance) of stool. 
On symptomatic days, subject reports more than 25% of stools are Type 1 or 2 and less than 
25% of stools are Type 6 or 7 on the Bristol Stool Form Scale. 
In addition to the ROME IV criteria, subjects must meet the inclusion and exclusion criteria listed 
above.  


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9.3.2 
Demographics and medical history 
The study staff will record the subject’s sex, date of birth, and ethnic origin. 
Medical history findings (i.e. previous diagnoses, diseases or surgeries) meeting all criteria listed 
below will be collected: 
• 
Not pertaining to the study indication; 
• 
Start before signing of the informed consent; 
• 
Considered relevant to the study; 
• 
Differentiated from adverse events. 
Findings will be recorded in REDCap via the medical history questionnaire (Appendix 13.6). 
9.3.3 
Physical exam 
Complete physical exams will be performed during the Screening Visit and Visits 2 and 5 by the 
investigator or a physician co-investigator. The general examination will include evaluation of the 
head, ears, eyes, nose, throat, endocrine, cardiovascular, respiratory, abdomen, skin, neurological, 
extremities, and musculoskeletal systems. Abnormalities noted at the Screening Visit will be 
recorded as medical history. Any clinically significant change from the Screening Visit will be 
recorded as an adverse event. 
9.3.4 
Vital signs 
Vital sign measurements, including body weight, blood pressure, radial pulse rate, respiratory rate, 
and temperature will be recorded at all clinic visits except the End-of-Study Call/Visit. Vital signs will 
be collected after the subject has been semi-supine for at least five minutes prior to the test. Height 
will be recorded at the Screening Visit only. 
9.3.5 
12-lead ECG 
A standard 12-lead ECG will be performed by a trained research nurse during the Screening Visit and 
Visit 5. ECGs will be interpreted by a trained cardiologist. 
9.3.6 
Blood draw 
Blood will be drawn by a trained research nurse and collected for hematology, clinical chemistry, 
lipid, cytokine, and creatine phosphokinase. Laboratory analyses will be performed in the central 
clinical laboratory and MAST research laboratory at Cedars-Sinai Medical Center. Reference ranges 
will be supplied by the clinical laboratory and used by the investigator to assess the laboratory 
results for clinical significance and pathological changes. Blood samples will be used for serum 
pregnancy tests in women of childbearing potential. 
Clinical chemistry (CMP) will be assessed on Visits 1 (Screening Visit), 2, 3, and 5. Creatine kinase 
(CK) will be assessed during Visit 1, however may be assessed as needed during the remaining study 
visits as indicated by the development of symptoms suggestive of myopathy and/or rhabdomyolysis. 


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Blood will be drawn during all visits except Visit 4 and the End-of-Study Call/Visit; however, blood 
may be drawn on Visit 4 and/or the End-of-Study Call/Visit as dictated by occurrence of subject 
symptom or adverse events. 
9.3.7 
Urine samples 
Subjects will be asked to provide up to one cup of urine for drug and toxicology analysis during the 
Screening Visit. Urine samples will also be collected for pregnancy tests from women of childbearing 
potential during Visits 2, 3, 4 and 5. 
9.3.8 
Daily diary and questionnaires 
A study diary will be completed each day assessing stool frequency, SBMs, CSBMs, and the use of 
rescue medication (Appendix 13.3). Stool consistency will be assessed using the Bristol Stool Chart 
(Appendix 13.2). Severity of abdominal pain Will be assessed via visual NRS scale (0-10), while 
severity of bloating and straining will be assessed via Likert scale (0-4)  
The PAC-SYM questionnaire and adequate relief question (Appendix 13.4) will be completed weekly. 
The EQ-5D-5L questionnaire (Appendix 13.5) will be completed on Day 1 prior to study drug 
administration and at Week 12 or upon early termination.  
 Subjects will be instructed on how to use REDCap to complete the assessments. During the 
Screening and Pre-treatment Period, subjects must complete the daily assessment for at least 14 
days and have full responses to the questions on 11 or more of the 14 days before the start of the 
Treatment Period. 
9.3.9 
Stool samples 
Subjects will be asked to provide up to one cup of stool for analysis in the MAST research laboratory. 
Stool samples will be analyzed for total bacterial and methanogen levels. 
9.3.10 Breath gases 
9.3.10.1    Lactulose breath test 
Lactulose breath tests will be performed during the Screening Visit and Visit 5. Subjects will be 
asked to undergo a 24-hour preparatory period just prior to the test. Subjects will maintain a 
bland diet during the first 12 hours and fast during the remaining 12. A baseline breath test will 
be collected by exhaling into disposable collection bag with a volume of 750 mL. Following the 
baseline breath sample, subjects will consume 10 g lactulose dissolved in water. Subsequent 
breath samples will be collected every 15 minutes over the remainder of the 120-minute test. 
Breath samples will be analyzed for hydrogen, methane, carbon dioxide, and hydrogen sulfide 
levels immediately after collection via gas chromatograph. The gas chromatograph will be used 
as approved. 
9.3.10.2    Methane breath test 
Methane breath tests will be performed during Visits 2, 3, and 4. Subjects will exhale into a 
disposable collection bag with a volume of 750 mL. Breath samples will be analyzed immediately 
after collection via gas chromatograph. The gas chromatograph will be used as approved. 


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9.3.11 Adverse events 
Investigators should refer to the Safety Information section of the current IB for SYN-010, including 
the DCSI (development core safety information), for the expected side effects of SYN-010.  As with 
any agent, there is always the potential for unexpected AEs, including hypersensitivity reactions.  
The IB will be updated if any new relevant safety data are obtained. 
Therapeutic monitoring should be performed following dose selection of SYN-010 in a manner 
consistent with the local clinical standard of care.  In general, patients should be closely monitored 
for side effects of all concomitant medications regardless of the path of drug elimination. 
All concomitant medications must be recorded in the patient’s source documentation. 
Patients must be carefully monitored for AEs.  This monitoring also includes clinical laboratory tests.  
Adverse events should be assessed in terms of their seriousness, intensity, and relationship to the 
study drug treatment. 
The institutional sponsor and investigator are responsible to comply with the local regulation and 
legislation for adverse events reporting. 
9.3.11.1    Definitions 
Adverse event (AE) 
In a clinical study, an AE is any untoward medical occurrence (i.e. any unfavorable and 
unintended sign [including abnormal laboratory findings], symptom or disease) in a patient or 
clinical investigation patient after providing written informed consent for participation in the 
study.  Therefore, an AE may or may not be temporally or causally associated with the use of an 
investigational product. 
A surgical procedure that was planned prior to the start of the study by any physician treating 
the patient should not be recorded as AE. The clinical manifestation of any failure of expected 
pharmacological action (lack of efficacy) is not recorded as an AE if it is already reflected as a 
data point captured in the CRF.  If, however, the event fulfills any of the criteria of an SAE, it 
must be recorded as and AE and reported as an SAE. 
• 
Conditions that started before signing of informed consent and for which no symptoms 
or treatments are present until signing of informed consent are recorded as medical 
history (e.g. seasonal allergy without acute complaints). 
• 
Conditions that started before signing of informed consent and for which symptoms or 
treatment are present after signing of informed consent, at unchanged intensity, are 
recorded as medical history (e.g. allergic pollinosis). 
• 
Conditions that started or deteriorated after signing of informed consent will be 
documented as adverse events. 
 
 


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Serious adverse event (AE) 
An SAE is classified as any untoward medical occurrence that, at any dose, meets any of the 
following criteria: 
• 
Results in death. 
• 
Is life-threatening. 
o The term ‘life-threatening’ in the definition refers to an event in which the 
patient was at risk of death at the time of the event, it does not refer to an event 
which hypothetically might have caused death if it were more severe. 
• 
Requires inpatient hospitalization or prolongation of existing hospitalization. 
o A hospitalization or prolongation of hospitalization will not be regarded as an SAE 
if at least one of the following exceptions is met: 
▪ 
The admission results in a hospital stay of less than 12 hours. 
▪ 
The admission is pre-planned. 
▪ 
The admission is not associated with an AE. 
o Results in persistent or significant disability / incapacity. 
o Disability means a substantial disruption of a person’s ability to conduct normal life’s 
functions. 
▪ 
Is a congenital anomaly/birth defect. 
▪ 
Is another medically important serious event as judged by the 
investigator. 
9.3.11.2    Reporting of serious adverse events 
The definition of serious adverse events (SAEs) is given above in section 7.3.1.1.  
Each serious adverse event must be followed up until resolution or stabilization, by submission 
of updated reports to the designated person. The investigator must promptly report to the 
Institutional Review Board all events, incidents, information, and outcomes that represent a 
possible unanticipated problem involving risks to subjects or others. An isolated laboratory 
abnormality that is assigned as serious, according to the investigator is not reportable as an SAE, 
unless the investigator assesses that the event meets standard ICH criteria for an SAE. Baseline 
laboratory abnormalities that are part of the disease profile should not be reported as an SAE, 
specifically when they are allowed or not excluded by the protocol inclusion/exclusion criteria.   
When required, and according to local law and regulations, serious adverse events that are 
determined by the Investigator to be related to the investigational drug must be reported by the 
investigator or the institution safety officer to the Ethics Committee and Regulatory Authorities. 
Per the emergency unblinding section above (section 6.4.1), if SAE is considered to be related to 
the treatment, the Cedars-Sinai safety group will unblind the subject and determine if the 
subject was administered investigational drug and if so, will proceed with expedited reporting to 
the FDA.    
If the SUSAR requires expedited safety reporting, MedWatch Form should be submitted within 
the timelines specified in 21 CFR 312.32 using: 


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• 
A MedWatch form available at http://www.fda.gov/medwatch/ 
 
Copies of all MedWatch reports shall be sent electronically within 15 days to: 
 
Electronic Mailbox:  
Michael Kaleko, MD, PhD (Research & Development): mkaleko@syntheticbiologics.com   
Amy Sloan (Regulatory Affairs): asloan@syntheticbiologics.com 
Address: 
9605 Medical Center Drive, Suite 270 
Rockville, MD  20850 
  
Phone:   
(240) 238-3862 
 At the conclusion of the study, all safety reports and associated data should be forwarded to 
Dr. Kaleko at Synthetic Biologics. 
9.3.12 Pregnancies 
The investigator must report any pregnancy occurring in a study patient, or in his partner, during the 
patient’s participation in this study.  The report should be submitted within the same timelines as an 
SAE, although a pregnancy per se is not considered an SAE.   
For a study patient, the outcome of the pregnancy should be followed up carefully, and any 
abnormal outcome of the mother or the child should be reported. 
For the pregnancy of a study patient’s partner, all efforts should be made to obtain similar 
information on course and outcome, patient to the partner’s consent. 
9.3.13 Further safety 
Individual subject stopping criteria: If any subject demonstrates (i) clinical signs of potential serious 
liver injury, hepatotoxicity, and/or hyperbilirubinemia jaundice, or (ii) myopathy/rhabdomyolysis 
with or without acute renal failure secondary to myoglobinuria, or (iii) a hypersensitivity reaction 
determined by the investigator to be associated with treatment, that subject should be withdrawn 
from therapy.  
An increase in serum aminotransferases to > 3x ULN will be followed by repeat testing within 48 to 
72 hours to confirm the abnormality and determine whether they are increasing or decreasing. If an 
abnormality develops during the course of the study that meets any of the following criteria, the 
subject should be withdrawn: 
• 
ALT or AST > 8x ULN; 
• 
ALT or AST > 5x ULN for more than 2 weeks; 
• 
ALT or AST > 3x ULN and (total bilirubin > 2x ULN or INR > 1.5); or 
• 
ALT or AST > 3x ULN with the appearance of fatigue, nausea, vomiting, right upper quadrant 
pain or tenderness, fever, rash, and/or eosinophilia (> 5%).  


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If an ECG abnormality develops during the course of the study that meets the following criteria, the 
subject should be withdrawn: 
• 
Any QTc interval > 500 milliseconds, correction by Fridericia’s formula; 
• 
Any QTc interval increase from baseline > 60 milliseconds, correction by Fridericia’s formula; 
• 
New onset of arrhythmias such as ventricular tachycardia, multifocal ectopic tachycardia, 
ventricular triplets, supraventricular tachycardia, atrial fibrillation, second- or third-degree 
heart block, or bradyarrhythmias; or 
• 
New onset of ECG signs of cardiac ischemia such as, but not limited to, ST segment 
abnormalities (elevation or depression). 
Study stopping criteria: A blinded safety review of liver function tests will be conducted when 25%, 
50%, and 75% of the subjects have been enrolled. If 5% of the enrolled subjects in the study have 
persistent (e.g. confirmed on a repeated test and not resolved after 1 week) elevated liver function 
tests (AST, ALT) > 3xULN or experience myalgia or myositis with elevated CPK of grade 3 (severe), 
the investigator or designee will convene an independent drug safety group to assess the safety 
profile of the investigational drug for this study. In addition, any hepatocellular injury meeting the 
definition of Hy’s Law will require study enrollment to be halted until an independent drug safety 
group is convened and their recommendation is received. Any potential Hy’s Law case will be 
handled as a suspected unexpected serious adverse reaction. 
9.3.13.1    Progressive disease 
If progressive disease leads to signs and symptoms that meet the criteria for an SAE 
(i.e., hospitalization, disability, death, or important medical event), the signs and symptoms 
should be reported as an SAE and not the underlying progressive disease. 
9.3.13.2    Death 
If any patient dies during the trial or within 30 days of the End-of-Treatment Visit, the 
investigator or the institution’s safety officer will document and record the cause of death in 
detail using the institutions SAE reporting procedure.  The investigator or institution safety 
officer will complete the appropriate MedWatch report criteria and provide notification to  
Synthetic Biologics immediately. 
10. Data handling and quality assurance 
10.1 Data recording 
The investigator will be responsible for assuring that all the required data will be collected and 
properly documented. It is the expectation that all data has source documentation available at the 
site. The site must implement processes to ensure compliance with Good Clinical Practice (GCP).  
10.2 Monitoring 
The investigator will appoint two monitors outside of the study team. The monitors are responsible 
for checking the quality of data and ensuring that all activities adhere to the study protocol. 
Additionally, the monitors ensure that legal and ethical requirements as stated in local laws and the 


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principles of GCP are being followed. Further information about the monitoring plan is included on a 
separate document titled “Data Safety Monitoring Plan”. 
10.3 Data processing 
10.3.1 Data collection 
The study staff will enter the data required by the protocol into an electronic database from the 
source documents (e.g. medical records and study-specific data capture tools as needed). Data 
entered will be reviewed for completeness and accuracy. 
10.3.2 Data handling and quality assurance 
The investigator is required to prepare and maintain adequate and accurate case histories designed 
to record all observations and other data pertinent to the study for each study participant. All 
information recorded in the database for this study must be consistent with the subjects’ source 
documentation (i.e. medical records). 
The final data will be analyzed in accordance with the SAP.  
10.4 Audit and inspection 
Inspections by regulatory health authority representatives (i.e. FDA and IEC[s]/IRB[s]) are possible.  
The investigator should notify Synthetic Biologics immediately if a notification of inspection is 
received from the FDA regarding this study. 
10.5 Archiving 
Essential documents shall be archived safely and securely in such a way that ensures that they are 
readily available upon authorities’ request. 
Patient (hospital) files will be archived according to local regulations and in accordance with the 
maximum period of time permitted by the hospital, institution or private practice. 
11. Premature termination of the study 
• 
If risk-benefit ratio becomes unacceptable owing to: 
o Safety findings from this study (e.g. SAEs); 
o Results of parallel clinical studies; 
o Results of parallel animal studies (on e.g. toxicity, teratogenicity, carcinogenicity, or 
reproduction toxicity). 
• 
If the study conduct (e.g. recruitment rate, drop-out rate, data quality, protocol compliance) does 
not suggest a proper completion of the trial within a reasonable timeframe.  
The investigator has the right to close his/her center at any time. 
For any of the above closures, the following applies: 


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• 
Closures should occur only after consultation between involved parties. 
• 
All affected institutions (e.g. IEC[s]/IRB[s], competent authority[ies], study center, head of study 
center) must be informed as applicable according to local law. 
• 
In case of a partial study closure, ongoing patients, including those in post study follow-up, must 
be taken care of in an ethical manner. 
12. Ethical and legal aspects 
12.1 Ethical and legal conduct of the study 
The procedures set out in this protocol, pertaining to the conduct, evaluation, and documentation of 
this study, are designed to ensure that the investigator abide by Good Clinical Practice (GCP) 
guidelines and under the guiding principles detailed in the Declaration of Helsinki. The study will also 
be carried out in keeping with applicable local law(s) and regulation(s). 
Documented approval from appropriate IEC(s)/IRBs will be obtained for all participating centers 
before start of the study, according to GCP, local laws, regulations and organizations. When 
necessary, an extension, amendment or renewal of the EC/IRB approval must be obtained and also 
forwarded to Synthetic Biologics. 
Strict adherence to all specifications laid down in this protocol is required for all aspects of study 
conduct; the investigator may not modify or alter the procedures described in this protocol.   
Modifications to the study protocol will not be implemented by the investigator without discussion 
and agreement by Synthetic Biologics.  However, the investigator may implement a deviation from, 
or a change of, the protocol to eliminate an immediate hazard(s) to the trial patients without prior 
IEC/IRB/Synthetic Biologics approval/favorable opinion.  As soon as possible, the implemented 
deviation or change, the reasons for it and if appropriate the proposed protocol amendment should 
be submitted to the IEC/IRB/head of medical institution.  Any deviations from the protocol must be 
explained and documented by the investigator. 
The investigator is responsible for the conduct of the clinical trial at the site in accordance with Title 
21 of the Code of Federal Regulations and/or the Declaration of Helsinki. The investigator is 
responsible for personally overseeing the treatment of all study patients. The investigator must 
assure that all study site personnel, including sub-investigators and other study staff members, 
adhere to the study protocol and all FDA/GCP/NCI regulations and guidelines regarding clinical trials 
both during and after study completion. 
The investigator will be responsible for assuring that all the required data will be collected and 
properly documented. 
12.2 Patient information and consent 
This section must adhere to the following regulations as applicable: 
• 
Section 4.8 of the ICH E6 Guideline for Good Clinical Practice. 
• 
Health Insurance Portability and Accountability Act (HIPAA). 
The following standard text should be adapted as needed: 


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Each patient/legal representative or proxy consenter will have ample time and opportunity to ask 
questions and will be informed about the right to withdraw from the study at any time without any 
disadvantage and without having to provide reasons for this decision. 
Only if the patient/legal representative or proxy consenter voluntarily agrees to sign the informed 
consent form and has done so, may he/she enter the study. Additionally, the investigator and other 
information provider (if any) will personally sign and date the form. The patient/legal representative 
or proxy consenter will receive a copy of the signed and dated form. 
The signed informed consent statement is to remain in the investigator site file or, if locally required, 
in the patient’s note/file of the medical institution. 
In the event that informed consent is obtained on the date that baseline study procedures are 
performed, the study record or patient’s clinical record must clearly show that informed consent was 
obtained prior to these procedures. 
If the patient is not capable of providing a signature, a verbal statement of consent can also be given 
in the presence of an impartial witness (independent of Synthetic Biologics and the investigator). This 
is to be documented by a signature from the informing physician as well as by a signature from the 
witness. 
For minors or adults under legal protection, consent shall be given by the legal guardian(s). The 
consent of a minor or adult under legal protection shall also be requested where such a person is 
able to express his/her own will. His/her refusal or the withdrawal of his/her consent may not be 
disregarded. 
In emergency situations, when prior consent of the patient is not possible, the consent of the 
patient’s legal representative(s) or proxy consenter, if present, should be requested. The patient 
should be informed about the study as soon as possible and his/her consent to continue the study 
should be requested. 
The informed consent form and any other written information provided to patients/legal 
representatives or proxy consenters will be revised whenever important new information becomes 
available that may be relevant to the patient’s consent, or there is an amendment to the protocol 
that necessitates a change to the content of the patient information and/or the written informed 
consent form. The investigator will inform the patient/legal representative or proxy consenter of 
changes in a timely manner and will ask the patient to confirm his/her participation in the study by 
signing the revised informed consent form. Any revised written informed consent form and written 
information must receive the IEC/IRB’s approval/favorable opinion in advance of use. 
12.3 Publication policy 
Synthetic Biologics recognizes the right of the investigator to publish results upon completion of the 
study.  All investigator-initiated studies are the property of the investigator and publications 
generated from such studies are at the discretion of the investigator. Synthetic Biologics encourages 
investigators to publish the results of all studies supported through the Investigator Sponsored Study 
(ISS) Program.  


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Investigators are encouraged to submit a draft manuscript of supporting publication(s) or abstract(s) 
for a courtesy review prior to submission of the final version for publication or congress presentation. 
This will be reviewed promptly and will not be withheld unreasonably.   
The investigator should ensure that the information regarding the study be publicly available on the 
internet at www.clinicaltrials.gov. 
12.4 Confidentiality 
All records identifying the patient will be kept confidential and, to the extent permitted by the 
applicable laws and/or regulations, will not be made publicly available. 
Should direct access to medical records require a waiver or authorization separate from the patient’s 
statement of informed consent, the Investigator will obtain such permission in writing from the 
appropriate individual. 
13. Appendices 
13.1 Laboratory tests 
CLINICAL CHEMISTRY 
HEMATOLOGY 
URINE SAMPLE 
Electrolytes 
Full Blood Exam 
Drug Screen (Screening) 
Bicarbonate 
Hematocrit 
Amphetamine 
Calcium, Total 
Hemoglobin 
Barbiturates 
Chloride 
Packed Cell Volume 
Benzodiazepines 
   Potassium 
Platelet Count 
Cocaine 
Sodium 
Mean Cell Volume 
Narcotics 
Lipids 
Mean Cell Hemoglobin 
Other Tests 
  Cholesterol, Total 
Mean Cell Hemoglobin Conc. 
Urine Pregnancy2 
Low Density Lipoprotein (LDL) 
Red Blood Cell count 
 
Triglycerides (TG) 
White Blood Cell Counts (total and 
differential) 
 
Liver Function Tests 
 
 
   Albumin 
 
 
Alanine Aminotransferase (ALT) 
 
 
Alkaline Phosphatase (ALP) 
 
 
Aspartate Aminotransferase (AST) 
 
 
Bilirubin, Total 
 
 
Protein 
 
 
Metabolic Parameters 
 
 
Glucose 
 
 
Renal Function Tests 
 
 
   Creatinine 
 
 
Urea Nitrogen (BUN) 
 
 
Other Tests 
 
 
   Creatine kinase (CK) 
 
 
Serum Pregnancy (bHCG; Screening)¹ 
 
 
¹Only women of child-bearing 
potential. 
²Urine pregnancy test can utilize a 
dipstick. 
 
 


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13.2 Bristol stool chart 
 
13.3 Daily questionnaire 
Did you have a bowel movement today? 
If yes, did it feel complete? 
What did it look like? Please use the Bristol Stool Chart as a reference. 
Did you have more than one bowel movement today? 
If yes, did it feel complete? 
What did it look like? 
Did you have more than two bowel movements today? 
If yes, did it feel complete? 
What did it look like? 
Did you take a laxative today? 


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If yes, please specify: Protocol-recommended rescue medication (bisacodyl, magnesium citrate, 
magnesium hydroxide, or lactulose) or Other laxative (oral, suppository, or enema) 
If other, please specify the name of the laxative taken. 
How severe has straining been during your bowel movements over the last 24 hours. 
0 
None (absent) 
1 
Mild (not very severe) 
2 
Moderate (somewhat severe) 
3 
Severe 
4 
Very Severe 
 
Using the scale above, please rate the worst level of abdominal pain you have felt over the past 24 
hours.  
 
Bloating is a condition in which the belly (abdomen) feels full and tight or may look or feel 
swollen/distended. Patients may experience abdominal fullness, pressure, or a sensation of trapped 
gas. 
Please rate the worst level of bloating you have felt over the last 24 hours. 
0 
None (absent) 
1 
Mild (not very severe) 
2 
Moderate (somewhat severe) 
3 
Severe 
4 
Very Severe 
13.4 PAC-SYM 
This questionnaire asks you about your constipation symptoms in the past week. Answer each 
question according to your symptoms, as accurately as possible. There are no right or wrong answers. 
For each symptom below, please indicate how severe your symptoms have been during the past 
week. If you have not had the symptom during the past week, tick 0. If the symptom seemed mild, 
tick 1. If the symptom seemed moderate, tick 2. If the symptom seemed severe, tick 3. If the 
symptom seemed severe, tick 4. Please be sure to answer every question. 
How severe have each of these symptoms been in the past week? 
 
Absent  
(0) 
Mild  
(1) 
Moderate 
(2) 
Severe  
(3) 
Very Severe 
(4) 


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Discomfort in your 
stomach 
 
 
 
 
 
Pain in your stomach 
 
 
 
 
 
Bloating in your stomach 
 
 
 
 
 
Stomach cramps 
 
 
 
 
 
Painful bowel movements 
 
 
 
 
 
Rectal burning during or 
after a bowel movement 
 
 
 
 
 
Rectal bleeding or tearing 
during or after a bowel 
movement 
 
 
 
 
 
Incomplete bowel 
movement, as though you 
didn’t “finish” 
 
 
 
 
 
Stools that were too hard 
 
 
 
 
 
Stools that were too small 
 
 
 
 
 
Straining or squeezing to 
try to pass stools 
 
 
 
 
 
Feeling like you had to 
pass a stool but you 
couldn’t (false alarm) 
 
 
 
 
 
 
Overall, have you had adequate relief of your IBS symptoms over the last 7 days? 
 
13.5 Quality of life questionnaire 
Under each heading, please check the ONE box that best describes your health TODAY. 
MOBILITY 
I have no problems in walking about 
I have slight problems in walking about 
I have moderate problems in walking about 
I have severe problems in walking about 
I am unable to walk about 
SELF-CARE 
I have no problems washing or dressing myself 
I have slight problems washing or dressing myself 
I have moderate problems washing or dressing myself 


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I have severe problems washing or dressing myself 
I am unable to wash or dress myself 
USUAL ACTIVITIES (e.g. work, study, housework, family, or leisure activities) 
I have no problems doing my usual activities 
I have slight problems doing my usual activities 
I have moderate problems doing my usual activities 
I have severe problems doing my usual activities 
I am unable to do my usual activities 
PAIN/DISCOMFORT 
I have no pain or discomfort  
I have slight pain or discomfort 
I have moderate pain or discomfort 
I have severe pain or discomfort 
I have extreme pain or discomfort  
ANXIETY/DEPRESSION 
I am not anxious or depressed  
I am slightly anxious or depressed 
I am moderately anxious or depressed 
I am severely anxious or depressed 
I am extremely anxious or depressed 
PLEASE RATE YOUR OVERALL HEALTH TODAY 
The scale is from 0 to 100, where 0 is the WORST health you can imagine and 100 is the BEST health 
you can imagine. 
 
13.6 Medical history 
SECTION 1: DEMOGRAPHICS 
Age 
Gender 
Height (inches) 
Weight (pounds) 
DEFINITIONS OF RACIAL CATEGORIES: 
White:  
A person having origins in any of the original peoples of Europe, North Africa, or the Middle East. 
Black or African American:  
A person having origins in any of the Black racial groups of Africa. 


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Asian:   
A person having origins in any of the original peoples of the Far East, Southeast Asia, or the 
Indian subcontinent including, for example, Cambodia, China, India, Japan, Korea, Malaysia, 
Pakistan, the Philippine Islands, Thailand, and Vietnam. 
American Indian or Alaska Native:  
A person having origins in any of the original peoples of North and South America, including 
Central America, and who maintains tribal affiliation or community attachment. 
Native Hawaiian or Other Pacific Islander: 
A person having origins in any of the original peoples of Hawaii, Guam, Samoa, or other Pacific 
Islands. 
Please choose all that apply. 
Please indicate whether you are of Hispanic ethnicity. 
DEFINITION: 
Hispanic or Latino: 
A person of Cuban, Mexican, Puerto Rican, South or Central American, or other Spanish culture or 
origin regardless of race. 
SECTION 2: MEDICAL HISTORY 
What year was your constipation diagnosed? 
Have you ever had an obstruction (blockage) of your bowel? 
Have you ever had a fistula (abnormal tube-like connection between two organs)? 
Have you had a perforation (cut/hole) in your bowel? 
Have you had a fissure (tear in the skin) or abscess (pocket of inflamed tissue filled with pus) 
around the anus? 
SECTION 3: SURGICAL HISTORY 
Have you had an appendectomy (removal of the appendix)? 
If yes, date of your appendectomy: 
 Have you had a cholecystectomy (removal of the gallbladder)? 
If yes, date of your cholecystectomy: 
Have you had a bowel resection (removal of part of the bowel)? 
If yes, date of your bowel resection: 
SECTION 4: SOCIAL HISTORY 
Do you exercise? 
Type of exercise? Please choose all that apply. 
Aerobic 
Strength training 
Yoga/stretching 
How would you describe your level of physical activity? 


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Inactive: Rare physical work or exercise 
Active: Moderate physical work or leisure activities, such as running, cycling, swimming, or 
aerobics, for less than 30 minutes per day and less than 3 days per week 
Very Active: Strenuous physical work or activities, such as running, cycling, swimming, or 
aerobics, for more than 30 minutes per day and at least 3 days per week 
Do you, or did you, drink alcohol? 
If yes, what type of alcohol do you drink? 
How many standard drinks do you have per week? 
1 glass of wine = 1.5 standard drinks 
1 bottle of beer = 1 standard drink 
1 oz. hard liquor = 1 standard drink 
Do you, or did you ever, smoke? 
If yes, what year did you start? 
Did you quit? 
If yes, what year did you quit? 
On average, how many cigarettes do you, or did you, smoke per week? 
Do you “vape” (use e-cigarettes)? 
Are you exposed to secondhand smoke? 
Do you, or did you ever, smoke marijuana? 
If yes, what year did you start? 
Did you quit? 
If yes, what year did you quit? 
Do you use recreational drugs (not including marijuana)? 
If yes, what type of recreational drugs do you use? 
Have you ever used recreational drugs in the past? 
If yes, what type of recreational drugs did you use in the past? 
SECTION 5: ADDITIONAL MEDICAL HISTORY 
(For female subjects only.) 
Have you ever been pregnant? 
If yes, how many times have you been pregnant? 
Have you ever taken hormone replacement therapy? 
If yes, how many years have you been taking/did you use hormone replacement therapy? 
Do you take birth control pills? 
Are you menopausal? 
If yes, natural or surgical menopause? 
Are you pre-menopausal? Please choose all that apply. 
No 


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Yes, hot flashes 
Yes, night sweats 
Yes, vaginal dryness 
Have you had a hysterectomy? 
If yes, what type of operation did you have? 
Uterus 
Uterus + one ovary 
Uterus + both ovaries 
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SAP_001.pdf: 
NCT03763175