Phase 1/1b Study of Redirected Autologous T Cells Engineered to Contain an Anti CD19 and Anti CD20 scFv Coupled to CD3ζ and 4-1BB Signaling Domains in Patients With Relapsed and/or Refractory CD19 or CD20 Positive B Cell Malignancies
Phase: PHASE1
Conditions: Lymphoma, Non-Hodgkin, Lymphoma, B-Cell, Chronic Lymphocytic Leukemia, Small Lymphocytic Lymphoma
Sponsor: Medical College of Wisconsin

 
 
 
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CLINICAL STUDY PROTOCOL 
Phase 1/1b Study of Redirected Autologous T Cells Engineered to Contain an Anti CD19 and 
Anti CD20 scFv Coupled to CD3ζ and 4-1BB Signaling Domains in Patients with Relapsed 
and/or Refractory CD19 or CD20 Positive B Cell Malignancies 
 
 
Short Protocol Title  
Phase 1/1b Study of CAR-20/19-T Cells in Patients with Relapsed Refractory B Cell 
Malignancies  
 
IND Number 17518 
 
Clinical Trials.gov Number: NCT03019055 
 
 
Principal Investigator 
Nirav Shah, MD MSHP  
 
 
 
 
 
 
Version 5 
6/18/2019 
 
 
 
 
Proprietary and Confidential 
The information in this document is considered privileged and confidential, and may not be 
disclosed to others except to the extent necessary to obtain Institutional Review Board approval 
and informed consent, or as required by federal and state laws. Persons to whom this 
information is disclosed should be informed that this information is privileged and confidential 
and that it should not be further disclosed. 

 
 
 
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MCW Protocol No.:  PRO00028724.   
FDA IND No.: 17518 
Principal Investigator (PI) 
Nirav Shah, MD MS 
Assistant Professor of Medicine 
Medical College of Wisconsin 
9200 W. Wisconsin Ave 
Milwaukee, WI 53226 
Phone: 414-805-4600 
Fax: 414-806-6815 
Email address: nishah@mcw.edu 
 
Principal Lab Investigator 
Bryon D. Johnson, PhD 
Professor of Medicine, Hem/Onc 
Director MCW BMT Laboratories 
Medical College of Wisconsin 
8700 Watertown Plank Road 
Milwaukee, WI, 53226 
Phone 414-805-6145 
Email: bjohnson@mcw.edu 
 
Co-Principal Investigator  
Parameswaran Hari, MD MS 
Professor 
Hematology/Oncology 
Medical College of Wisconsin 
9200 W. Wisconsin Ave 
Milwaukee, WI 53226 
Phone: 414-805-4600 
Fax: 414-806-6815 
Email: phari@mcw.edu 
 
Biostatistician: 
Mei-Jie Zhang, PhD 
Professor 
Division of Biostatistics 
Medical College of Wisconsin 
9200 W. Wisconsin Ave 
Milwaukee, WI 53226 
Email: meijie@mcw.edu 
 
Co-Investigator 
Timothy S. Fenske MD 
Professor of Medicine 
Medical College of Wisconsin 
9200 W. Wisconsin Ave 
Milwaukee, WI 53226 
Phone: 414-805-4600 
Fax: 414-806-6815 
tfenske@mcw.edu 
 
 
 
 
Co-Lab Investigator 
Fenlu Zhu, PhD 
Assistant Professor of Medicine 
Medical College of Wisconsin 
8700 Watertown Plank Road 
Milwaukee, WI, 53226 
Phone 414-805-6145 
fzhu@mcw.edu 
 
Co-Investigator 
Mehdi Hamadani MD 
Professor of Medicine 
Medical College of Wisconsin 
9200 W. Wisconsin Ave 
Milwaukee, WI 53226 
Phone: 414-805-4600 
Fax: 414-806-6815 
mhamdani@mcw.edu 
 
 

 
 
 
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PROTOCOL SUMMARY 
Title 
Phase 1/1b Study of Redirected Autologous T Cells Engineered to 
Contain an Anti CD19 and Anti CD20 scFv Coupled to CD3ζ and 4-
1BB Signaling Domains in Patients with Relapsed and/or Refractory 
CD19 or CD20 Positive B Cell Malignancies 
Protocol Number 
PRO00028724 
IND # 
17518 
Principal Investigator  Nirav Shah, MD MSHP 
Assistant Professor, Department of Medicine 
Study Sites 
Froedtert Hospital & Medical College of Wisconsin Cancer Center 
Clinical Trial Phase 
Phase 1/1b 
Study Disease 
CD19 or CD20 Positive B Cell Malignancies including CLL/SLL and 
B-cell NHL 
Inclusion Criteria 
 
1. Diagnosis of B-cell NHL or CLL/SLL: Patients must be aged≥18 
years with relapsed, refractory disease and no available curative 
options that meet clinical criteria to initiate treatment. 
2. Patients with B-cell NHL or CLL/SLL must have either CD19 or 
CD20 positive disease on most recent biopsy performed (a 
repeat biopsy is not mandatory for this study except as noted 
below). A minimum of 5% CD19 or CD20 positivity by 
immunohistochemistry or flow cytometry on prior or repeat 
biopsy is required.  
3. Absolute CD3+ T cell count ≥50/mm3  
4. MRI brain and Lumbar Puncture with CSF analysis by cytology 
and flow cytometry without evidence of CNS involvement ONLY 
in patients with history of CNS involvement 
5. Measurable disease must have been documented within 4 weeks 
of the time of consent defined as the following by disease specific 
subtype: 
a. B-cell NHL: Active disease defined as nodal lesions greater 
than 20 mm in the long axis or extranodal lesions >10 mm in 
long and short axis or bone marrow involvement that is biopsy 
proven 
b. CLL/SLL: Active disease by either bone marrow, peripheral 
flow cytometry, or CT and/or PET imaging with nodal disease 
6. Patients should have failed at least two lines of a standard 
treatment and meet disease specific criteria detailed below: 
a. CLL/SLL: measurable disease as defined above that has relapsed 
after at least one line of chemo-immunotherapy and progressed 
or intolerant to ibrutinib monotherapy  
b. CD19 or CD20 positive B cell NHL limited to the following 
histologies: Advanced Stage III or IV Follicular Lymphoma, 
Diffuse Large B cell Lymphoma and associated subtypes (e.g. 
aggressive 
B-cell 
lymphoma, 
T-cell/histocyte 
rich 
B-cell 

 
 
 
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lymphoma, primary mediastinal B-cell lymphoma, EBV+ diffuse 
large B-cell lymphoma, transformed lymphoma such as 
transformed follicular or marginal zone lymphoma, and Richter’s 
transformation) and Mantle cell lymphoma. Specific criteria 
include: 
- 
Patients must have active, measurable disease after two lines 
of cytotoxic chemotherapy of which one must be anthracycline 
containing.  
- 
Must have received Rituximab or another CD20 antibody and 
at minimum two chemotherapy regimens appropriate for their 
disease.  
- 
Either failed autologous transplant or ineligible to receive 
autologous transplant 
7. Karnofsky performance score ≥70. See Appendix A for scales. 
8. Normal Baseline Neurological Evaluation: Mini-Mental Status 
Exam Score 24-30 (Appendix B) 
9. Adequate hepatic function, defined as AST and ALT <5 x upper 
limit of normal (ULN); serum bilirubin and alkaline phosphatase 
<5 x ULN, or considered not clinically significant as per the clinical 
PIs discretion (e.g. Gilbert’s or indirect hyperbilirubinemia) or felt 
to be due to underlying disease.  
10. Adequate renal function, defined as creatinine clearance>60 
ml/min 
11. Able to provide written informed consent  
12. Agree to practice birth control during the study 
13. Adequate cardiac function as indicated by New York Heart 
Association (NYHA) classification I or II AND left ventricular 
ejection fraction of ≥35% (by cardiac ECHO or MUGA) and 
adequate pulmonary function as indicated by room air oxygen 
saturation of ≥92%. 
14. Expected survival >12 weeks 
15. Negative urine or serum pregnancy test in females of child bearing 
potential at study entry and again within 48 hours’ prior 
lymphodepleting chemotherapy. 
16. Patients with prior blinatumomab treatment require repeat biopsy 
post-blinatumomab treatment that demonstrates CD19 or CD20 
positive disease.  
17. Meet criteria for regarding fertility and contraception detailed in 
section 4.4 
18. Central line access will be required for CAR-20/19-T cell infusion. 
 
Exclusion Criteria 
 
1. Positive beta-HCG in female of child-bearing potential defined as 
per table 1. 
2. Patients with known systemic allergy to bovine or murine 
products. 
3. Known prior positive serology for human anti-mouse antibody 
(HAMA).  
4. Confirmed active human immunodeficiency virus (HIV), Hepatitis 
B or C infection. 
5. History of significant autoimmune disease OR active, uncontrolled 
autoimmune phenomenon: such as systemic lupus erythematous, 

 
 
 
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Wegner’s glomerulonephritis, autoimmune hemolytic anemia, 
idiopathic thrombocytopenic purpura (AIHA, ITP) requiring steroid 
therapy defined as >20 mg of prednisone or equivalent daily. 
6. Presence of ≥grade 3 non-hematologic toxicities as per CTCAE 
version 5.0 from any previous treatment unless it is felt to be due 
to underlying disease.  
7. Concurrent use of investigational therapeutic agents or enrollment 
on another therapeutic clinical trial at any institution. Minimum of 
≥4 weeks required from administration of any other investigational 
agents on other clinical trials prior to enrollment on this CAR-T 
protocol. 
8. Refusal to participate in the long-term follow-up protocol 
9. Patients with active CNS involvement by malignancy on MRI or 
by lumbar puncture. 
a. 
Patients with prior CNS disease that has been effectively 
treated will be eligible providing treatment was >4 weeks 
before enrollment and a remission documented within 8 
weeks of planned CAR-T cell infusion by MRI brain and CSF 
analysis. 
10. Previous recipients of allogeneic hematopoietic stem cell 
transplantation (AHCT) are excluded if they are <100 days’ post-
transplant, have evidence of active graft-versus-host-disease 
(GVHD) of any grade, or are currently on immunosuppression.  
11. Previous CAR-T cell therapy directed at either CD19 or CD20 
within 100 days of planned CAR-20/19-T cell infusion (does not 
include re-enrollment) 
a. 
Patients with prior CAR-T treatment against CD19 or CD20 
must have repeat biopsy confirming a minimum of 5% CD19 
or CD20 positivity by immunohistochemistry or flow 
cytometry 
12. Anti-CD20 antibody treatment within 4 weeks of cell infusion 
13. Anti-CD19 antibody treatment within 4 weeks of cell infusion 
14. Cytotoxic chemotherapy other than lymphodepletion within 14 
days of CAR-T cell infusion  
15. Cytotoxic chemotherapy treatment within 14 days or steroid 
treatment (other than replacement dose steroids) within 7 days 
prior to apheresis collection for CAR-T cells 
16. Patients post solid organ transplant who develop high grade 
lymphomas or leukemias 
17. Concurrent active malignancy other than basal or squamous cell 
carcinomas of the skin 
 
Study Rationale 
The study is divided into two phases. Phase 1 and Phase 1b.   
 
Phase 1:  To evaluate a novel closed processing method for the 
production of the first dual CD20 and CD19 CAR-T cell and 
determine the safety of administration of these cells as part of a first 
in human study consisting of 2 cohorts; dose escalation and dose 
expansion. The CAR-T cells were given over 2 days.  
 
 

 
 
 
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Phase 1b: To determine the safety of single dose (unfractionated 
CAR-T cell infusion at 2.5 x 106 cells/kg  
Primary Objectives 
1. Determine the safety (dose level) of CAR-20/19-T cell 
administration in relapsed refractory CLL/SLL and B-cell NHL 
2. Determine the feasibility to manufacture CAR-20/19-T cells 
locally from patient apheresis products using the CliniMACS 
Prodigy Cell processing device 
Secondary 
Objectives 
1. Determine the anti-tumor responses as measured by response 
rates.  
2. Describe the duration of response for responding patients 
3. Determine persistence of CAR-20/19-T cells 
4. Determine the effects of CAR-20/19-T infusion on non-neoplastic 
CD19 & CD20 B cells in vivo 
5. Determine if cellular or humoral host immunity develops against 
CAR-20/19-T cells  
6. Evaluate MRD using molecular technologies 
Endpoints 
1. Ability to manufacture CAR-20/19-T cells from patient apheresis 
products using the CliniMACS Prodigy processing device (Miltenyi 
Biotec). The number T cell lines that do not meet criteria for T cell 
purity, total transduced T cell content, viability, and sterility will be 
determined.  
2. Occurrence of adverse events, defined as either CRS related 
Grade 3/4 toxicity or other NCI CTCAE version 5.0 non-hematologic 
≥ grade 3 signs/symptoms, laboratory toxicities and clinical events 
that are possibly, probably or definitely related to study treatment at 
any time from the infusion until day +28 post CAR-T infusion 
Study Design 
This is a phase 1/1b, interventional, single arm, open label, treatment 
study designed to evaluate the safety and feasibility of infusion of 
CAR-20/19-T in patients with B cell malignancies that have relapsed 
after prior therapies and are not candidates for curative intent 
standard therapy.  
 
The study is divided into two phases. Phase 1 and Phase 1b.   
 
Phase 1:  consists of 2 cohorts; dose escalation and dose 
expansion. The CAR-T cells were given over 2 days.  
In the phase 1 portion, there will be a dose escalation cohort to 
determine the safe CAR-20/19-T cell dose in patients with CLL/SLL 
and NHL. Once the desired dose has been identified there will be a 
6 patient dose expansion phase at the specified dose level. 
Phase 1b: CAR-T infusion given as a single dose.  
In the Phase 1b portion of the study, we will test the safety of 
unfractionated CAR-T cells utilizing the safe dose identified in the 
phase 1 portion (2.5 x 106 cells/kg).  
Study Agent 
Description 
CAR-20/19-T cells will be administered either fresh or thawed after 
cryopreservation by IV injection. Patients will receive one of four 
dose levels of CAR-20/19-T cells based on our dose escalation 
design (Section 6). Cells will be given over 2 days, 30% on Day 0 

 
 
 
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and 70% on Day 1 in the Phase 1 portion and in Phase 1b as a 
single infusion on Day 0. Cell dose will be maxed at 80 kg.  
 
Phase 1 
Dose Level -1: 1 x 105 CAR-20/19-T cells/kg  
Dose Level 0: 2.5 x105 CAR-20/19-T cells/kg (starting dose level) 
Dose Level 1: 7.5 x105 CAR-20/19-T cells/kg 
Dose Level 2: 2.5 x106 CAR-20/19-T cells/kg (goal cell dose) 
 
Phase 1b 
Expansion Dose Level: 2.5 x 106 cells/kg (single infusion) 
 
Number of Subjects 
A maximum of 24 patients will be enrolled on this Phase 1/1b study. 
Subject Participation 
Duration 
Subjects will be followed for up to 2 years post infusion of the CAR-
20/19-T cells  
Duration of Follow up 
Patients will be followed for up to 2 years post infusion of CAR-20/19-
T cells infusion as part of this protocol and then placed on a long-
term follow-up protocol that will follow patients for up to a total of 15 
years post CAR-T cell infusion  
Estimated Time to 
Complete Enrollment: 
18-24 months 
Statistical 
Methodology: 
 
This is a phase 1/1b, interventional single arm, open label, 
treatment study designed to evaluate the safety and feasibility of 
infusion of CAR-20/19-T cells. In the phase 1 portion the study will 
utilize a 3+3 dose escalation design in NHL & CLL/SLL followed by 
a 6 patient expansion cohort. 
In the Phase 1b portion, 9 patients will be treated at the safe dose 
from the Phase 1 portion (2.5 x 106 cells/kg).and will receive a 
single infusion rather than fractionated dose.  
Safety Assessments 
Safety will be determined by assessments of cytokine release 
syndrome (CRS) using the Lee et al. grading scale [1] and adverse 
events via NCI CTCAE version 5.0. Dose limiting toxicities (DLT) will 
be monitored for the first 28 days after CAR-20/19-T cell infusion as 
per Table 1. DLT will be defined a grade 3-4 non-hematologic toxicity 
possibly, probably, or definitely related to the infusion of CAR-20/19-
T cells. All deaths (grade 5 toxicity) in the first 28 days felt to be either 
possibly, probably, or definitely related to CAR-20/19-T cell therapy 
will be considered a DLT. CRS related DLTs and hematologic DLT 
are defined in the protocol (section 3.4) 
 
Efficacy 
Assessments 
CLL/SLL & B-cell NHL at Day +28 (+/- 3 days) 
PET/CT or CT neck/chest/abd/pelvis + BM Aspirate and Biopsy 

 
 
 
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Unique Aspects of 
this Study 
This is the first phase 1/1b study to evaluate the safety and efficacy 
of a CAR-T cell therapy directed against two B cell antigens and 
produced under GMP conditions using the closed system 
CliniMACS Prodigy device in B cell malignancies. 

 
 
 
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FIGURE 1: STUDY SCHEMA 
  
 
 
Phase 1: Cells administered over 2 days (30% on Day 0, 70% on Day 1) 
Phase 1b: Cells administered as a single infusion on Day 0 (100%) 

 
 
 
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TABLE 1: SCHEDULE OF EVENTS 
 
Week -8      
to  
Day -15 
Day 
-15 
 
ᶲ2 to 4 
days 
prior to 
day -4 
 
♦Day    
-4 
LDP 
§Day 0 
+6 
weeks 
Day 
+1 
#Day 
+2  
to Day 
+7 
Day 
+10 
Day 
+14 
Day 
+21 
Day 
+28 
Day 
+60 
Day 
+90 
Day 
+120 
Day 
+150 
Day 
+180  
Day 
+270 
Day 
+365 
$Follow 
up 
(Year 
1-2) 
Visit Window 
Screening/
Enrollment 
Aphere
sis 
 +/- 1 
day 
 +/- 5 
days 
CAR-20/19-T 
cells infusion 
  
+/- 1 
day 
+/- 2 
days 
+/- 2 
days 
+/- 3 
days 
+/- 7 
days 
+/- 7 
days 
+/- 7 
days 
+/- 7 
days 
+/- 
14 
days 
+/- 
14 
days 
+/- 1 
month 
+/- 1 
month 
Signed Inform Consent  
x 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Apheresis  
  
x 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
CAR-20/19-T cells infusion1 
  
  
  
  
x 
 x1 
  
  
  
  
  
  
  
  
  
  
  
  
  
Confirmation of Diagnosis of CD19 
or CD20 positive B-cell NHL or 
CLL/SLL2 
x 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Recent History and Physical 
Examination 3 
x 
x  
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
Karnofsky performance status 
x 
 
 
 
x 
 
 
 
 
 
x 
 
 
 
 
 
 
x 
 
Vitals4 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
Concomitant Medication & 
Evaluation of Adverse Events 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
Bone Marrow Transplant Markers 5 
x 
  
  
  
x 
  
  
  
  
  
x 
  
x 
  
  
x 
  
x 
 x  
Tumor Measurements by PET/CT, 
CT Scans6 
x 
  
x6  
 
  
  
  
  
  
  
x 
  
 
  
  
 x 
  
x 
x 
ECG7 
x 
  
  
 
x 
 
  
  
  
  
x 
  
  
  
  
  
  
 
  
ECHO/MUGA 
x 
  
  
  
  
  
  
  
  
  
 
  
  
  
  
  
  
  
  
Lumber Puncture 8 
x 
  
x8  
 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
MRI Brain9 
x 
 
x9 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Bone Marrow Biopsy & Aspirate 10-11 
x 
  
  
  
  
  
  
  
  
  
x 
  
  
  
  
x 
  
x 
 x 
CBC and Differential  
x 
x 
  
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
Comprehensive Metabolic Panel12  
x 
x 
  
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
LDH/Uric Acid 
x 
x 
  
x 
x 
x 
x 
x 
x 
x 
x 
 
x 
 
 
x 
 
x 
x 
Quantitative Immunoglobulins 
x 
  
  
  
x 
 
  
 
 
 
x 
x 
x 
x 
x 
x 
x 
x 
x 

 
 
 
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Week -8      
to  
Day -15 
Day 
-15 
 
ᶲ2 to 4 
days 
prior to 
day -4 
 
♦Day    
-4 
LDP 
§Day 0 
+6 
weeks 
Day 
+1 
#Day 
+2  
to Day 
+7 
Day 
+10 
Day 
+14 
Day 
+21 
Day 
+28 
Day 
+60 
Day 
+90 
Day 
+120 
Day 
+150 
Day 
+180  
Day 
+270 
Day 
+365 
$Follow 
up 
(Year 
1-2) 
Visit Window 
Screening/
Enrollment 
Aphere
sis 
 +/- 1 
day 
 +/- 5 
days 
CAR-20/19-T 
cells infusion 
  
+/- 1 
day 
+/- 2 
days 
+/- 2 
days 
+/- 3 
days 
+/- 7 
days 
+/- 7 
days 
+/- 7 
days 
+/- 7 
days 
+/- 
14 
days 
+/- 
14 
days 
+/- 1 
month 
+/- 1 
month 
Ferritin/CRP/ESR 
x 
  
  
  
x 
x 
x 
x 
x 
x 
x 
  
  
  
  
  
  
  
  
PT/INR/PTT/D-Dimer/Fibrinogen 
x 
  
  
 
x 
x 
x 
x 
x 
x 
x 
  
  
  
  
  
  
  
  
TSH/Free T4 
x 
  
  
  
  
  
  
  
 
 
x 
 
 
 
  
 
  
x 
  
Lipid Panel 
x 
  
  
  
  
  
  
  
  
  
x 
  
  
  
  
  
  
 
  
Urinalysis 
x 
  
 x 
 
  
  
  
  
  
  
x 
  
  
  
  
  
  
 
  
Beta-2-Microglobulin 
x 
 
 
 
 
 
 
 
 
 
x 
 
 
 
 
 
 
 
 
Autoimmune Screen (ANA) 
x 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
 
Pregnancy Test (serum or urine) 13 
x 
  
 x 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Infectious Disease Markers 14 
x 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Respiratory Virus Panel 15 
 
  
x 
 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Creatinine Clearance 16 
x 
  
  
  
 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
Central Line17 
 
 
x 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Research Studies 
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
  
T Cell Persistent Studies 18 
  
  
  
  
x 
x 
x 
x 
x 
x 
x 
 
x 
 
 
x 
 
x 
x 
Q-PCR for integrated vector DNA19 
  
  
  
  
x 
 
 
 
 
 
x 
 
x 
 
 
x 
 
x 
x 
Presence of replication competent 
virus by Q-PCR for VSV-g20 
 
 
 
 
x 
 
 
 
 
 
x 
 
x 
 
 
x 
 
x 
x 
Cytokine Studies) 21 
  
  
  
  
x 
x 
x 
x 
x 
x 
x 
  
  
  
  
  
  
  
  
Neurological evaluations22 
x 
 
 
 
 
x 
x 
 
 
 
x 
 
 
 
 
 
 
 
 
T-cell repertoire Diversity Studies23 
 
x 
 
x 
 
 
x 
 
 
 
x 
 
 
 
 
 
 
 
 
PaxGene RNA Tubes25 
 
x 
 
 
 
 
 
 
x 
 
x 
 
x 
 
 
 
 
 
 
Tryptophan Metabolites25 
 
x 
 
 
 
 
 
 
x 
 
x 
 
x 
 
 
 
 
 
 
Quality of Life Studies26 
 
x 
 
 
 
 
 
 
x 
 
x 
 
x 
 
 
x 
 
x 
x 
 
Footnotes: 
        ♦  Start of Lymphodepleting Chemotherapy (LDP) Theses assessments should be completed daily during each day of LDP unless otherwise specified. Start date  
of LDP chemotherapy will be determined on the date of CAR-T cell infusion. 
        ᶲ 
Pre-lymphodepletion visit will be based on date of CAR-T cell administration and hence can move forward by up to 6 weeks 

 
 
 
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§ 
CAR-20/19-T cells may be given fresh after manufacturing without cryopreservation or can be cryopreserved and given up to 6 weeks after production 
# 
Daily from Days +1 through Day +7 unless otherwise specified in Section 5.4.6 
$ 
Clinical Visits every 6 months (+/- 1 month) from Year 1 to Year 2 
1- 
Split-dosing of CAR-T cells through Phase 1 escalation/expansion. Single dosing of CAR-T cells in Phase 1b 
2- 
Patients will need documentation of relapsed or refractory B cell malignancies. CD19 or CD20 positivity ≥5% by IHC or Flow cytometry required on one biopsy. 
Please see eligibility criteria for specific requirements for each patient.  
3- 
History and height only required at screening. Physical exam, weight at all other time points 
4- 
Vitals should include Blood Pressure, Temperature, Respiratory rate, Pulse & Pulse Ox. On the days of CAR-20/19-T cell infusion, vitals should be done prior 
to infusion, at the end of infusion, and every 15 minutes thereafter for a minimum of one-hour post infusion. 
5- 
Absolute numbers of CD3+ T cells, CD4+ T cells and CD8+ T cells should be assessed before scheduling for apheresis collection to determine protocol 
eligibility. Minimum of ≥50 CD3+ T cells/mm3 required, with a mixture of both CD4+ and CD8+ T cells.   
6- 
Diagnostic imaging is required at baseline (within 8 weeks of CAR-20/19-T cell infusion) and at 1 month post-cell infusion (PET/CT preferred). Diagnostic CT 
scans of the neck/chest/abdomen/pelvis must be performed if PET/CT scans cannot be obtained at baseline due to timing constraints, insurance or 
reimbursement issues, or other reasons. Either PET/CT or diagnostic CT scans are required to document response. If >8 weeks’ elapse from date of imaging 
and planned T-cell infusion, repeat imaging will be required.  For follow-up patients with get either PET/CT or diagnostic CT scans at Day +28, 6 months, and 1 
year. For year 1-2 scans will be done yearly. 
7- 
ECG to be performed prior to CAR-20/19-T cell infusion on Day 0 
8- 
Lumbar Puncture with CSF analysis by cytology and flow cytometry is required at screening/eligibility for all patients with prior history of CNS disease.  If >8 
weeks elapse from last LP and planned CAR-20/19-T cell infusion, a repeat LP will be required in patients with a history of CNS disease and who exhibit CNS 
symptoms. 
9- 
MRI Brain will be performed only for patients with history of prior CNS disease or signs/symptoms of active CNS disease. MRI will be repeated if >8 weeks 
elapse between previous MRI and scheduled CAR-20/19-T cell infusion.  
10- Patients should have a staging bone marrow biopsy and aspirate ≤ 8 weeks prior to CAR-20/19-T cell infusion. If >8 weeks’ elapse, a repeat biopsy will be 
indicated. 
11- All patients will have a repeat marrow at Day +28 post-CAR-T cell infusion. Beyond Day +28, for patients with B-cell NHL ongoing marrow biopsy core and 
aspirate at 6 months, 1, year, and 2 year is indicated only if positive at baseline. Patients with CLL/SLL are required to have BM biopsy at all specified 
timepoints (6 months, 1 year, 2 year). Bone marrow biopsy and aspirate should also be obtained at time of suspected relapse. If baseline marrow is positive 
only by flow cytometry, the aspirate and flow cytometry should be repeated.  
12- Comprehensive Metabolic Panel: Albumin, Alkaline Phosphatase, AST, Bicarb, Bili T, Calcium (total), Chloride, Creatinine, Glucose, Potassium (K), Sodium 
(NA), Protein Total, BUN, ALT.  
13- Within 48 hours prior to the start of lymphodepleting chemotherapy. Definition of female of childbearing potential(FCBP): A FCBP is a sexually mature female 
regardless of sexual orientation or whether she has undergone a tubal ligation who:  1) has not undergone a hysterectomy or bilateral oophorectomy or 2) has 
not been naturally postmenopausal (amenorrhea following cancer therapy does not rule out childbearing potential) for at least 24 consecutive months (i.e., has 
had menses at any time in the preceding 24 consecutive months). 
14- Patients must be tested for HIV, Hepatitis B surface antigen/antibody and core antibody, and HCV within 8 weeks prior to CAR T cell infusion.   If the HCV 
antibody is positive, a screening HCV RNA by any RT-PCR or bDNA assay must be performed. Repeat testing is NOT required.  
15- Viral Respiratory NAAT will be done 2-4 days prior to initiation of lymphodepleting (LDP) chemotherapy if patients exhibit symptoms concerning for viral 
infection (cough, cold, fever). In the setting of a positive result, initiation of LPD chemotherapy and CAR-T treatment will be delayed until resolution of 
symptoms.  
16- Calculate Creatinine (by Cockcroft-Gault) at screening.  
17- Central Line is required prior to initiation of LDP chemotherapy.  
18- T-cell persistent studies: baseline sample to be drawn prior to CAR-20/19-T cell infusion on Day 0 followed by Day +1, +7, +10, +14, +21, +28 and then per 
calendar above. If there are two consecutive negative tests—this will not need to be repeated.  Additional samples (blood, CSF, ascites, tissue, etc.)  for T-
cell persistence can be performed per the investigators discretion as felt to be clinically indicated.  
19- Q-PCR for Integrated Vector DNA: Will be done prior to infusion then at Day +28, 3 months, 6 months, 1 year, 1.5 years, 2 years. If there are two consecutive 
negative tests—this will not need to be repeated.  

 
 
 
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20- Replication competent lentivirus (RCL) will be monitor by Q-PCR for VSV-G (Vesicular stomatitis virus-G) which is a surrogate for RCL. Will be done prior to 
infusion then at Day +28, 3 months, 6 months, 1 year, 1.5 years, 2 years.  Testing will continue until Q-PCR for integrated vector DNA is negative for two 
consecutive timepoints.  
21- At minimum, will include assays for the following cytokines: IFN-ϒ, IL10, IL6 and TNFα. Additional cytokines will be monitored at discretion of investigators 
Cytokines will be checked on Day 0, +1, Day +7, Day+10, Day+14, Day+21, Day+28. Additional samples may be taken if indicated based on clinical 
presentation.  
22- Given concern for neurotoxicity with CAR-T cells, neurological evaluation will be done at baseline and on Day+1, Day+7 and every third day beyond if the 
patient develops neurological toxicity. For patients without neurological toxicity after CAR-T therapy, re-evaluation will occur on Day+28. Neurological 
Assessment Tool is found in Appendix B. 
23- T-cell receptor diversity studies. Sample 1 at Week -4 will be from apheresis product. Sample 2 will be from the CAR-T cell product. Sample 3 and 4 will be a 
peripheral blood sample (10 ml) on Day +7 and Day +28 
24- Pax Gene tubes will be collected and stored until sent for analysis as described in Section 11.2 until disease progression 
25- Red top tube will be collected at the noted time points and delivered to Dr. Hillard’s MCW lab for analysis until disease progression 
26- Quality of Life studies until disease progression 
 
 
VISITS REQUIRED AT MCW: All visits through Day+28 will be required to occur at MCW. For responding patients, beyond Day +28, 
patients will have required visits at MCW on Day +90 (+/- 7 days), Day+180 (+/- 14 days), Day 365 (+/- 1 month), +1.5 years (+/- 1 month) 
and 2 years (+/- 1 month). For timepoints not falling on those dates, local oncology evaluation will suffice, and records will be obtained 
accordingly.  
For patients who progress, follow-up will be as per Section 5.4.7  

 
 
 
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TABLE OF CONTENTS 
PROTOCOL SUMMARY ............................................................................................................ 3 
FIGURE 1: STUDY SCHEMA ..................................................................................................... 9 
TABLE 1: SCHEDULE OF EVENTS ........................................................................................ 10 
TABLE OF CONTENTS ........................................................................................................... 14 
LIST OF ABBREVIATIONS ...................................................................................................... 18 
1 INTRODUCTION ................................................................................................................... 22 
1.1 BACKGROUND ...................................................................................................................22 
1.2 INVESTIGATIONAL AGENT(S) ...............................................................................................23 
1.2.1 FIGURE 3—GENE MAP AND ANIMATION OF THE CD20/CD19 VECTOR ...............................24 
1.3 PRECLINICAL DATA ............................................................................................................25 
1.4 MCW PRECLINICAL DATA ..................................................................................................25 
1.5 CLINICAL DATA ..................................................................................................................31 
1.6 RATIONALE FOR LYMPHODEPLETION AND CAR-T CELL DOSING ............................................32 
1.7 KNOWN AND POTENTIAL RISKS AND BENEFITS ....................................................................33 
2 HYPOTHESIS AND OBJECTIVES ........................................................................................ 36 
2.1 HYPOTHESIS .....................................................................................................................36 
2.2 PRIMARY OBJECTIVES .......................................................................................................36 
2.3 SECONDARY OBJECTIVES ..................................................................................................36 
2.4 RATIONALE FOR THE OUTCOME MEASURES SELECTION .......................................................36 
3 STUDY DESIGN .................................................................................................................... 37 
3.1 GENERAL DESCRIPTION .....................................................................................................37 
3.2 NUMBER OF SUBJECTS ......................................................................................................37 
3.3 PRIMARY ENDPOINT(S) ......................................................................................................37 
3.4 DOSE LIMITING TOXICITY ...................................................................................................38 
3.5 SECONDARY ENDPOINT(S) .................................................................................................39 
3.6 STUDY TIMELINE ...............................................................................................................39 
3.7 PRIMARY COMPLETION ......................................................................................................39 
3.8 STUDY COMPLETION ..........................................................................................................40 
4 PATIENT SELECTION .......................................................................................................... 40 
4.1 ELIGIBILITY CRITERIA .........................................................................................................40 
4.2 INCLUSION CRITERIA ..........................................................................................................40 
4.3 EXCLUSION CRITERIA ........................................................................................................41 
4.4 SPECIAL CRITERIA FOR REGARDING FERTILITY AND CONTRACEPTION ...................................42 
4.5 SUBJECT RECRUITMENT AND SCREENING ...........................................................................42 
5  STUDY ENTRY AND WITHDRAWAL; STUDY PROCEDURES .......................................... 43 
5.1 STUDY ENTRY PROCEDURES .............................................................................................43 
5.2 PRETREATMENT PERIOD ....................................................................................................43 
5.3 ENROLLMENT ....................................................................................................................44 
5.4 STUDY PROCEDURES RELATED TO TREATMENT ..................................................................45 
5.4.1 STUDY PROCEDURES – DAY -15 (APHERESIS DAY) ...........................................................45 
5.4.2 MONONUCLEAR CELL APHERESIS ....................................................................................45 
5.4.3 STUDY PROCEDURES WEEK -1 + 6 WEEKS (~2 TO 4 DAYS PRIOR TO LYMPHODEPLETION) ...45 

 
 
 
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5.4.4 START OF LYMPHODEPLETION CHEMOTHERAPY (DAY -4 + 6 WEEKS) .................................46 
5.4.5 CAR-20/19-T CELL INFUSION (+ 6 WEEKS) ....................................................................46 
5.4.6 DAY +1 THROUGH DAY +7 AFTER CELL INFUSION ..............................................................47 
5.4.7 FOLLOW-UP VISITS (DAY+10 TO 2 YEARS) .......................................................................47 
5.5 STUDY WITHDRAWAL PROCEDURES ...................................................................................49 
6 TREATMENT PLAN .............................................................................................................. 50 
6.1 LYMPHODEPLETION (WITHIN 1 WEEK PRIOR TO CAR-20/19-T CELL INFUSION) .......................50 
6.2 PREPARATION OF CAR-20/19-T CELL INFUSION .................................................................50 
6.3 ADMINISTRATION OF CAR-20/19-T CELL INFUSION ON DAY 0 AND DAY +1 (PHASE 1) OR DAY 0 
(PHASE 1B) ..................................................................................................................52 
6.4 DOSE ESCALATION SCHEDULE ...........................................................................................54 
6.5 DOSE EXPANSION PHASE ..................................................................................................57 
6.6 PHASE 1B (SINGLE DOSE OF CAR-T CELL) .........................................................................57 
6.7 TREATMENT/DOSE LIMITING TOXICITY ................................................................................57 
6.8 USAGE OF CONCURRENT/CONCOMITANT MEDICATIONS .......................................................58 
6.9 DIETARY RESTRICTIONS.....................................................................................................58 
6.10 PROHIBITED MEDICATIONS ...............................................................................................58 
6.11 MONITORING SUBJECT COMPLIANCE ................................................................................58 
6.12 RE-TREATMENT ...............................................................................................................58 
7 DOSING DELAYS/DOSE MODIFICATIONS ......................................................................... 59 
7.1 DOSING ADMINISTRATION/DELAYS ......................................................................................59 
7.2 MONITORING AND TOXICITY MANAGEMENT..........................................................................60 
7.3 OTHER TOXICITIES ............................................................................................................61 
8ADVERSE EVENTS: DEFINITIONS AND REPORTING REQUIREMENTS ........................... 62 
8.1 DEFINITIONS .....................................................................................................................62 
8.2 UNANTICIPATED PROBLEM INVOLVING RISK TO SUBJECT OR OTHER (UPIRSO) ....................63 
8.3 AE ATTRIBUTION AND GRADING .........................................................................................63 
8.4 MONITORING AND RECORDING AN ADVERSE EVENT ............................................................64 
8.5 PROCEDURE FOR REPORTING DRUG EXPOSURE DURING PREGNANCY AND BIRTH EVENTS ....64 
8.6 SUBJECT COMPLAINTS .......................................................................................................65 
8.7 ROUTINE REPORTING PROCEDURES FOR AES ....................................................................65 
9 PHARMACEUTICAL INFORMATION ................................................................................... 67 
9.1.1 CAR-20/19-T CELLS .......................................................................................................67 
9.1.2 SOLUTION PREPARATION ................................................................................................68 
9.1.3 INVESTIGATIONAL AGENT ADMINISTRATION ......................................................................68 
9.1.4 STORAGE REQUIREMENTS ..............................................................................................68 
9.1.5 STABILITY ......................................................................................................................68 
9.1.6 ROUTE OF ADMINISTRATION ............................................................................................69 
9.1.7 NURSING IMPLICATIONS ..................................................................................................70 
9.1.8 HANDLING ......................................................................................................................70 
9.1.9 AVAILABILITY ..................................................................................................................70 
9.1.10 AGENT ORDERING ........................................................................................................70 
9.1.11 AGENT ACCOUNTABILITY ...............................................................................................70 
9.1.12 AGENT DESTRUCTION AND RETURN ...............................................................................70 
9.2.1 TOCILIZUMAB .................................................................................................................70 
9.2.2 PREPARATION ................................................................................................................71 
9.2.3 TOCILIZUMAB ADMINISTRATION ........................................................................................71 

 
 
 
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9.2.4 STORAGE .......................................................................................................................71 
9.2.5 STABILITY ......................................................................................................................71 
9.2.6 HANDLING ......................................................................................................................72 
9.2.7 AVAILABILITY ..................................................................................................................72 
9.2.8 AGENT ORDERING ..........................................................................................................72 
9.2.9 AGENT DESTRUCTION AND RETURN .................................................................................72 
10 REPORTING AND DOCUMENTING RESPONSE (MEASUREMENT OF EFFECT) ........... 72 
10.1 EVALUATION OF EFFICACY (OR ACTIVITY) ..........................................................................72 
10.2 RESPONSE CRITERIA .......................................................................................................72 
10.3 CHRONIC LYMPHOCYTIC LEUKEMIA RESPONSE CRITERIA ...................................................72 
10.4 NON-HODGKIN’S LYMPHOMA RESPONSE CRITERIA ............................................................74 
10.5 RESPONSE DEFINITIONS ..................................................................................................75 
10.6 EVALUATION OF SAFETY ..................................................................................................76 
11 CORRELATIVE STUDIES/SPECIAL STUDIES................................................................... 76 
11.1 T CELL DIVERSITY ANALYSIS DURING CAR-T CELL THERAPY ............................................76 
11.2: CIRCULATING TRYPTOPHAN METABOLITES AND GENE EXPRESSION PATTERNS AND RISK FOR 
CNS TOXICITY DURING CAR-T CELL THERAPY ...............................................................77 
11.3: PATIENT-REPORTED OUTCOME (PRO) MEASURES FOLLOWING CAR-T CELL THERAPY ........78 
12 STATISTICAL CONSIDERATIONS ..................................................................................... 80 
12.1 STUDY ENDPOINTS ..........................................................................................................80 
12.2 ACCRUAL ESTIMATES ......................................................................................................80 
12.3 INTERIM ANALYSES AND STOPPING RULES ........................................................................80 
12.4 STATISTICAL ANALYSES PLANS ........................................................................................80 
12.5 ANALYSIS POPULATION ....................................................................................................82 
13 DATA AND SAFETY MONITORING PLAN (DSMP) ........................................................... 82 
13.1 STUDY TEAM ...................................................................................................................82 
13.2 QUALITY ASSURANCE ......................................................................................................82 
13.3 CLINICAL TRIALS OFFICE ..................................................................................................82 
13.4 DSMC ............................................................................................................................83 
14 REGULATORY COMPLIANCE, ETHICS AND STUDY MANAGEMENT ............................ 83 
14.1 ETHICAL STANDARD.........................................................................................................83 
14.2 REGULATORY COMPLIANCE ..............................................................................................83 
14.3 INSTITUTIONAL REVIEW BOARD ........................................................................................84 
14.4 SUBJECT CONFIDENTIALITY AND ACCESS TO SOURCE DOCUMENTS/DATA ..........................84 
14.5 QUALITY ASSURANCE ......................................................................................................85 
14.6 CHANGES IN THE PROTOCOL ............................................................................................85 
15 DATA HANDLING AND RECORD KEEPING ..................................................................... 86 
15.1 OVERVIEW ......................................................................................................................86 
15.2 STUDY RECORD RETENTION ............................................................................................86 
15.3 PUBLISHING DATA ...........................................................................................................86 
16 REFERENCES .................................................................................................................... 87 
APPENDIX A: PERFORMANCE STATUS CRITERIA ............................................................. 94 
APPENDIX B:  NEUROLOGICAL ASSESSMENT ................................................................... 95 

 
 
 
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APPENDIX C: 2004 MAS/HLH DIAGNOSTIC CRITIERA ........................................................ 96 
APPENDIX D: SCHEMA FOR TESTING OF CART CELLS FOR FRESH INFUSION OR 
CRYOPRESERVATION (FOR LATER INFUSION) IN PHASE 1 DOSE 
ESCALATION/EXPANSION ..................................................................................................... 97 
APPENDIX E: CAR-T CELL TREATMENT GUIDELINES........................................................ 98 
APPENDIX F: ASSESSMENT/MANAGEMENT OF CYTOKINE RELEASE SYNDROME AND 
NEUROTOXICITY ASSOCIATED WITH CAR-T CELLS [1, 56] ............................................... 99 
APPENDIX G: SUPPORTIVE CARE MEASURES FOR CAR-T CELL TREATMENT ............ 103 
APPENDIX H: OPERATING PROCEDURES FOR PATIENTS TREATED WITH CAR-20/19-T 
CELLS AT A NON-SPECIFIED DOSE ................................................................................... 104 
APPENDIX I: QUALITY OF LIFE (QOL) MEASURES ........................................................... 105 
APPENDIX J: SPECIMEN COLLECTION AND HANDLING .................................................. 115 

 
 
 
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LIST OF ABBREVIATIONS 
AAHRPP 
Association for the Accreditation of Human Research Protection Programs 
AE 
 
Adverse Event 
ALC 
 
Absolute Lymphocyte Count 
ALL 
 
Acute Lymphoblastic Leukemia 
ALP 
 
Alkaline Phosphatase 
ANA   
Anti-Nuclear Antibody 
ALT 
 
Alanine Aminotransferase 
ANC 
 
Absolute Neutrophil Count 
AST 
 
Aspartate Aminotransferase 
ATC 
 
Anatomical Therapeutic Chemical (Classification System) 
BCW  
Blood Center of Wisconsin 
BUN 
 
Blood Urea Nitrogen 
CAR 
 
Chimeric Antigen Receptor 
CBC 
 
Complete Blood Cell (count) 
Cl 
 
Chloride 
CLL 
 
Chronic Lymphocytic Leukemia 
Cr 
 
Creatinine 
CR 
 
Complete Response 
CRC 
 
Clinical Research Coordinator 
CRF 
 
Case Report Form 
CRP 
 
C-Reactive Protein 
CRS 
 
Cytokine Release Syndrome 
CSF 
 
Cerebral Spinal Fluid 
CT 
 
Computerized Tomography  
CTCAE  
Common Terminology Criteria for Adverse Events 

 
 
 
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CTEP  
Cancer Therapy Evaluation Program 
CTMS  
Clinical Trial Management System 
DFS 
 
Disease-Free Survival 
DIC 
 
Disseminated Intravascular Coagulation 
DLT 
 
Dose Limiting Toxicity 
DLBCL  
Diffuse Large B cell Lymphoma 
DSMB  
Data and Safety Monitoring Board 
DSMC  
Data and Safety Monitoring Committee 
DSMP  
Data and Safety Monitoring Plan 
EBV 
 
Epstein-Barr Virus 
ECG 
 
Electrocardiography 
ECHO  
Echocardiogram 
ECOG  
Eastern Cooperative Oncology Group 
 
ESR 
 
Erythrocyte Sedimentation Rate 
FACT  
Foundation for the Accreditation of Cellular Therapy 
FCBP  
Female of Childbearing Potential 
FDA 
 
Food and Drug Administration 
FH 
 
Froedtert Hospital 
FT4 
 
Free Thyroxine 
GCP 
 
Good Clinical Practice 
HAMA  
Human Anti-Mouse Antibody 
HBeAg  
Hepatitis B “e” Antigen 
HBV 
 
Hepatitis B Virus 
HCT 
 
Hematocrit 
HCV 
 
Hepatitis C Virus 
HGB 
 
Hemoglobin 

 
 
 
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HIV 
 
Human Immunodeficiency Virus 
HLH 
 
Hemophagocytic Lymphohistiocytosis 
ICH 
 
International Conference on Harmonization 
IHC 
 
Immunohistochemistry 
IND 
 
Investigational New Drug Application 
IP 
 
Investigational Product 
IRB 
 
Institutional Review Board 
iwCLL  
International Workshop on Chronic Lymphocytic Leukemia 
IV 
 
Intravenous 
IVIG 
 
Intravenous Immunoglobulin 
K 
 
Potassium 
LDH 
 
Lactate Dehydrogenase 
LDP 
 
Lymphodepleting 
LP 
 
Lumbar Puncture 
LPL 
 
Lymphocyte Propagation Laboratory 
LFT 
 
Liver Function Test 
MAS 
 
Macrophage Activation Syndrome 
MCWCC  
Medical College of Wisconsin Cancer Center 
MedDRA  
Medical Dictionary for Regulatory Activities 
MMSE  
Mini-mental Status Exam 
MNC  
Mononuclear cells 
MRI 
 
Magnetic Resonance Imaging 
MUGA     
Multigated Acquisition Scan 
Na 
 
Sodium 
NAAT  
Nucleic Acid Amplification Test 
NCI 
 
National Cancer Institute 

 
 
 
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NHL 
 
Non-Hodgkin’s Lymphoma 
ORR  
Overall Response Rate 
PET/CT  
Positron Emission Tomography-Computed Tomography 
PD 
 
Progressive Disease 
PO 
 
Per os (by mouth, orally) 
PR 
 
Partial Response 
QOL 
 
Quality of Life 
Q-PCR   
Quantitative Polymerase Chain Reaction 
RBC 
 
Red Blood Cell (count) 
RCL 
 
Replication Competent Lentivirus 
SAE 
 
Serious Adverse Event 
SD 
 
Stable Disease 
SLL 
 
Small Lymphocytic Lymphoma 
SRC 
 
Scientific Review Committee 
SGOT  
Serum Glutamic Oxaloacetic Transaminase 
SGPT  
Serum Glutamic Pyruvic Transaminase 
TSH 
 
Thyroid Stimulating Hormone 
ULN 
 
Upper Limit of Normal 
UP 
 
Unanticipated Problem 
UPIRSO   
Unanticipated Problems Involving Risks to Subjects or Others 
VSV-G  
Vesicular Stomatitis Virus-G 
WBC  
White Blood Cell (count) 
WM 
 
Waldenstroms Macroglobulinemia 

 
 
 
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1 INTRODUCTION 
1.1 Background 
CD19 and CD20 are antigens expressed on both healthy B cells and a multitude of B cell- 
derived hematological malignancies including non-Hodgkin's lymphomas (NHL), acute 
lymphoblastic leukemia (ALL), and chronic lymphocytic leukemia (CLL). An estimated 162,000 
new cases of hematological malignancies were expected to be diagnosed in 2015.[2]  
 
CD19 and CD20 are both highly attractive targets for B cell neoplasm immunotherapy. CD19 is 
expressed by most B cell malignancies including NHL, ALL, CLL, hairy cell leukemia, and a 
subset of acute myelogenous leukemia.[3] CD19 is a 95kDa glycoprotein present on the B cell 
surface from early development until differentiation into plasma cells. It is a member of the 
immunoglobulin superfamily and a component of a cell surface signal transduction complex that 
regulates signal transduction through the B cell receptor. CD19 expression is restricted to B 
lineage cells and is not expressed by pluripotent blood stem cells or on most other normal 
tissues.[4, 5] Several clinical trials demonstrated effectiveness of CD19 chimeric antigen 
receptor (CAR) modified T cells in patients with B-cell ALL, CLL/SLL, or B-cell NHL. [6-10]  
 
CD20 is a non-glycosylated 33 to 37 kDa phosphoprotein that is expressed on both normal and 
malignant B cells. It normally functions as a component of signal transduction in growth 
regulation of B-lymphocytes. CD20, like CD19, is a cell surface receptor restricted to B cell 
precursors and mature B cells but is lost following plasma cell differentiation.[11, 12] Monoclonal 
antibodies against CD20 are effective in treating B cell malignancies. Unlike CD19 CAR, CD20 
CAR-T therapies are in early stages of development but have shown significant promise in 
some clinical trials.[13, 14]  
 
The primary advantage of adoptive immunotherapy using CAR-T cells compared to alternative 
modalities lies in the ability of T cells to expand and persist upon antigen binding of the target. 
[15] Persistence of CAR-T cells in patients with refractory NHL and ALL has correlated with 
sustained progression free survival likely due to the long-term surveillance these cells provide 
against recurrent malignancy.[6] However, despite encouraging results, not all patients respond 
to this treatment and even those who have achieved remission may relapse. An important 
mechanism of relapse to both targeted monoclonal antibody treatment and CAR-T therapy was 
shown to be downregulation of the targeted receptor allowing tumor cells to escape 
destruction.[7, 16] Given the overlap of CD19 and CD20 expression on B cells (Figure 2) we 
speculate that simultaneous targeting of these two separate B cell receptors may result in a 
more complete B cell ablation and a reduced risk of tumor cell escape.  
 
In this Phase 1/1b study we first hope to demonstrate the feasibility of manufacturing CAR-T 
cells expressing scFv specific for CD20-CD19 (CAR-20/19-T) in a completely closed system 
using the CliniMACS Prodigy device and then determine the safety of this dual targeted CAR in 
a first-in-human study of patients with relapsed and refractory B cell malignancies. Secondary 
outcomes will include response rates, and observed toxicities of the treatment, specifically the 
development of cytokine release syndrome (CRS)[7], an inflammatory storm that has been seen 
with previous CAR-T therapies.  
 
 
 

 
 
 
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1.2 Investigational Agent(s)  
The investigational agent in this protocol is the CAR-20/19-T cells. The CAR-T cells will be 
produced in the Lymphocyte Propagation Laboratory (LPL) under an IND to be submitted to 
FDA prior to trial initiation. The Cell lines will be produced as follows: A non-mobilized 
mononuclear cell (MNC) apheresis product (MNC, Apheresis) will be obtained from the patient 
approximately 4 weeks before infusion.  The MNC, Apheresis product will be washed with buffer 
to remove platelets and plasma, treated with a combination of antibodies to CD4 and CD8 that 
have been conjugated to paramagnetic particles and enriched for CD4+ and CD8+ T cells.  
Minimally 1.0 x 108 enriched cells will be activated using Miltenyi TransACT reagent, which 
consists of nanoparticles conjugated to CD3 and CD28. The cells will be incubated for 24 hours 
at 37oC and 5% CO2 prior to transduction.  The activated cells will then be incubated with a 
lenti-virus vector that will introduce a chimeric antigen receptor that will externally express 
antibody binding regions to CD19 and CD20, with transmembrane signaling domains from 4-
1BB and CD3ζ (Figure 3).  The cells will be further cultured in medium containing interleukin-2, 
fed, and expanded within the Prodigy culture chamber to numbers suitable for clinical use. After 
13-14 days of expansion in culture the cells will be washed and tested for expression of the 
CD20/CD19 antibody and for function using cytotoxicity and IFN-ϒ production assays in 
response to B cells prior to infusion. The lines will undergo further testing for sterility, viability, 
average copy number/cell, VSVg qPCR, and for relative subsets include central memory, 
effector memory, and regulatory T cells. Processing prior to the final preparation for either 
immediateadministration or cryopreservation will be performed in a closed and sterile system 
using GMP grade reagents and materials obtained from Miltenyi Biotec within the ISO 7 
environment of the LPL. Copy number and VSVg qPCR will be tested by Lentigen Technology. 
 
Closed Processing Using the CliniMACS Prodigy Device 
The ability to produce CAR-T cell lines suitable for clinical use requires manufacturing 
processes that are reproducible, scalable and that maintain sterility. Most CAR-T cell trials have 
involved complex manual processing procedures that include: enrichment and/or depletion of 
cellular subsets from starting cells, activation and exposure to viral vectors to introduce the 
chimeric antigen receptors, and cell expansion steps that may require weeks in culture.  There 
have been attempts to automate individual steps, or to close the system.[17] However, the 
CliniMACS Prodigy device (Miltenyi Biotec) is the first computer-controlled unit that automates 
each of the required processing steps while in a completely closed system.[17, 18] The Prodigy 
device has been approved for use in cell separation procedures and for the selection and 
culture of antigen reactive T cells.[19, 20]  We will use the CliniMACS Prodigy for the 
automated, closed system processing of the CAR-20/19-T cell line to be used in this study. 
Figure 2: B-cell 
maturation and 
markers of 
differentiation 

 
 
 
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1.2.1 Figure 3—Gene Map and Animation of the CD20/CD19 Vector 
 
 
 

 
 
 
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1.3 Preclinical Data 
A wealth of preclinical studies demonstrates the effectiveness of engineered T cells against 
CD19 and CD20 positive malignancies.  While the construction and the vector (gammaretroviral 
vectors vs lentiviral vectors) used to create CAR T cells has varied, the incorporation of a co-
stimulatory signaling domain in 2nd generation CARs with either CD28 or 4-1BB (CD137) 
increases the potency of these cells.[21, 22] Lentiviral vectors have been favored due to their 
ability to transduce non-dividing and dividing cells, insert a large genetic segment and allow for 
long-term transgene expression.[23-25] The pre-clinical data supporting the efficacy of CAR 
cells in xenograft models is published [26-28] and more recently the feasibility of the production 
of a dual antigen CAR-20/19-T cell was presented.[29] This preclinical model established the in 
vitro effectiveness of cytotoxic tumor cell killing by dual expressing CARs and also 
demonstrated a decreased level of antigen escape by down-regulation when compared to single 
antigen targeted CAR-T cells.[30] 
1.4 MCW Preclinical Data 
Clinical Experience at MCW with CliniMACS Devices 
The CliniMACS Prodigy device was recently introduced to the North American market as a 
follow-up to the CliniMACS Cell Separation System.  The MCW Cell Processing laboratory has 
an extensive experience with the CliniMACS Cell Separation systems, having employed them 
since 2005 for clinical protocols involving graft engineering to deplete and/or enrich specific 
cellular subsets as detailed below: 
 
 
MCW’s experience with the CliniMACS Cell Separation system indicates that the device 
produces reproducible and highly purified target populations, rigorous depletions of unwanted 
populations, and consistent recoveries of >50% of desired cell subsets. The MCW Cell 
Therapy Lab has additional experience with use of this device in a multi-center trial 
conducted by the Blood and Marrow Transplant Clinical Trial Network (CTN), CTN0303 that 
resulted in the FDA approval of the device for clinical use. Dr. Taylor (previous Lab Dierctor) 
had primary responsibility for preparing the Standard Operating Procedures for the device’s 
use and analyzing the device’s performance in that trial [31] as well as her current 
participation on the steering committee of the ongoing CTN1301 multi-center trial. The 
Prodigy device incorporates the same technology as the Cell Separation System but with 
additional functionalities that include pre-separation steps and post-separation culture steps. 
 
Preclinical Experience using CliniMACS Prodigy Device to produce CAR-T cells 
In September 2015, Dr. Fenlu Zhu, Assistant Lab Director, underwent 2 ½ days of intensive 
training to learn the steps of CAR-T cell production via the Prodigy device at Lentigen 
Technology, Inc.  Eight preclinical experiments using the CliniMACS Prodigy were conducted. 
Three experiments were performed using a CD19 vector with a design similar to the 
CD20/CD19 vector to be used clinically, 4 were using a research grade production of the vector 

 
 
 
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to be used clinically, and the 5th experiment used the GMP grade vector that will be used for the 
clinical protocol. For all 8 experiments cells were processed through the CD4/CD8 positive cell 
enrichment phase. There were two failed experiments of these first 8 studies. One failure was 
due to contamination of the starting cell product caused by a failure of aseptic technique during 
an open processing step; a step that would not be a factor for the clinical study. The second 
failure was device-related secondary to a software update that was not properly verified by the 
service technician from Miltenyi. The error prevented us from moving forward after the cell 
enrichment phase. The first line that was generated was compromised by the decision to start 
the experiment even though the source material was 72 hours old due to delays in receiving it 
into the laboratory. Going forward product processing will begin no longer than 48 hours after 
collection. The results obtained are summarized below. 
 
CD4/CD8 Enrichment:  Data for enrichment were obtained for all 8 lines and are summarized in 
the table below. Purity and Recovery post CD4/CD8 enrichment (N=8) 
 
Cell Population 
% of cells % recovery  
CD4+ plus CD8+ 
98±4 
54±7 
CD3+ 
90±10 
54±9 
CD3+CD4+ 
65±8 
60±9 
CD3+CD8+ 
26±11 
52±16 
CD3-CD56+ 
6±9 
27±15 
 
 
The enriched cells were highly purified for CD4+ and CD8+ cells.  While CD4 is limited to T 
cells, CD8 is also expressed on a subset of NK cells that may or may not be CD3+.  An average 
of 90±10% of the enriched cells were CD3+ and 6±9% were CD3-CD56+ NK cells that express 
a form of CD8 that has a lower antigen density than CD3+CD8+ T cells.  >50% of the starting 
target population of CD4+ plus CD8+ cells were recovered.  Importantly monocytes, which are 
known to suppress T cell proliferation of patient-derived material, were absent.[32, 33]  
 
Cell Expansion:  Cell lines were started using 1.0 x 108 starting CD4/CD8 enriched cells and 
were harvested on day 13.  The first experiment using a buffy coat product as the starting 
material expanded only 5.4-fold from the start, likely due to the age of the cells at the start of 
processing.  One line using the CD19 vector expanded 25.7-fold and the 4 lines using the 
CD20/CD19 vector expanded 414.5, 33.8, 24.5 and 24.8-fold.  Therefore, expansion exceeded 
the expected results from the Lentigen training experience of approximately 20-fold 
 
Transduction efficiency:  The percentage of viable transduced cells was measured by Protein L 
expression, a protein unique to immunoglobulin kappa light chains.[34]  Expression is shown in 
the table below and flow data from one representative line is shown in Figure 4. 
 
 
 
 
 
 
 
 
 
 
 

 
 
 
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Transduction efficiency as measured by Protein L expression 
 
Vector (MOI) 
% of CD3 % of CD4 % of CD8 Copy#/All Cells 
#1  CD19 (5) 
41.0 
42.3 
26.4 
1.01 
#2  CD19 (5) 
45.7 
52.3 
43.7 
0.48 
#3  CD20/CD19 (5) 
21.0 
22.7 
16.5 
0.23 
#4  CD20/CD19 (10) 
34.0 
28.7 
35.4 
0.53 
#5  CD20/CD19 (10) 
56.6 
57.6 
60.3 
0.67 
#6  CD20/CD19 (10) 
25.8 
28.7 
23.9 
nd 
 
 
Figure 4: Transduction efficiency representative experiment 
 
 
 
Figure 4: The cells were gated to remove debris, then to include only 7-AAD negative (viable), 
cells. The data are shown gated on all CD3+ cells (which were >99% of the CAR-T line) in the 
far left panel, gated on CD4+ T cells in the center panel, or gated on CD8+ T cells in the far right 
panel. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

 
 
 
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Post Expansion Phenotype:  The phenotype of the expanded population changed from the 
starting CD4/CD8 enriched cells in that fewer CD3-CD56+ NK cells expanded, as shown most 
clearly for line #3. The percentage of CD8+ cells increased from the starting cells expect for 
Line #1 where the line was greatly enriched for CD4+ cells, possibly due to the age of the cells 
at the start of the culture. The goal is to have both CD4+ and CD8+ cells in the cultures, though 
cytotoxicity is expected to be mediated predominately by the CD8+ T cells.  CD4+ CAR cells are 
expected to provide a helper function to the CD8+ cells secondarily to activation after exposure 
to the ligand bearing B cell. CAR-T cell line viability at the time of harvest was 90.1± 6.5%. 
 
 
Line Designation 
%CD3 
%CD3+ 
CD4+ 
%CD3+ 
CD8+ 
%CD3+ 
CD56+ 
%CD3-
CD56+ 
Line#1 Pre 
99.5% 
74.8% 
23.9% 
0.9% 
0.2% 
Post 
99.3% 
93.8% 
8.5% 
ND 
0.6% 
Line #2 Pre 
97.9% 
51.7% 
49.5% 
4.2% 
2.2% 
Post 
98.8% 
29.3% 
71.8% 
4.3% 
1.1% 
Line #3 Pre 
78.7% 
63.2% 
16.2% 
1.7% 
18.4% 
Post 
99.5% 
50.2% 
47.1% 
1.6% 
0.5% 
Line #4 Pre 
91.7% 
64.9% 
27.9% 
8.9% 
1.1% 
Post 
97.3% 
46.5% 
50.5% 
9.7% 
0.5% 
 
 
 
 
 
 
Line #5 Pre 
90.1% 
75.0% 
16.3% 
2.4% 
0.6% 
Post 
98.5% 
42.7% 
59.1% 
3.4% 
0.9% 
 
 
 
 
 
 
Line #6 Pre 
90.3% 
69.4% 
22.5% 
2.3% 
1.7% 
Post 
99.4% 
58.3% 
43.7% 
1.4% 
0.1% 
 
It is important that any method of CAR-T cell production should preserve a population of central 
memory T cells to ensure longevity in circulation.  An advantage of immunotherapy using CAR-
T cells is that during transduction the cells are not being expanded through antigen stimulation 
and thus should not become enriched for T cells with an exhausted phenotype (Terminal 
Memory T cells).  As shown in figure 5 below for Line#5, a population of Central Memory CD4+ 
and CD8+ T cells are maintained using the CAR-20/19-T cell vector manufactured both in the 
CliniMACS Prodigy and using plates followed by expansion in the G-Rex Device (Wilson-Wolf, 
Inc).  
 
 
 
 
 
 
 
 
 
 
 
 

 
 
 
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Figure 5: T cell Subset before and after CAR-T cell production in Plate vs. Prodigy 
 
 
 
Figure 5: T cell subsets were identified by gating on CD4 or CD8 positive cells and then assessing the percentage of 
cells expressing CD45RA and CD62Ligand.  Naïve T cells are CD45RA+/CD62L+. Central Memory (CM) T cells are 
CD45RA-/CD62L+, Effector Memory (EM) T cells are CD45RA-/CD62L-, and Terminal (exhausted) CD8+ T cell 
effectors are CD45RA+/CD62L-.  Data shown are from Line #5. 
 
Lines #1, #2, and #3 were tested for the presence of regulatory T cells (Tregs) before and after 
line generation.  Low numbers of Tregs were present in the starting fractions, but were 
undetectable in the lines post expansion, indicating that our approach does not enrich for 
populations that might suppress tumor killing.   
 
Effector cell function:  Effector cell function was measured both in 51Cr release cytotoxicity 
assays and by assessment of intracellular gamma interferon production after target cell co-
culture.  All the lines, except the Line #1 experiment, mediated lytic activity to B cell lines 
expressing both CD19 and CD2 (Raji or EBV transformed B cells). This was likely due to the 
failure of the CD8+ transduced cells to expand in this line.  However, both the few CD8+ cells 
and the CD4+ cells produced IFN-γ after co-culture with B cell targets  We obtained K562 cells 
that were separately transduced with CD19 and CD20 after production of Line #4 the second 
CD20/CD19 transduced line and could demonstrate preferable killing of both transduced targets 
in addition to Raji cell killing, though this line did contain a higher percentage of CD3+CD56+ 
cells than the other lines after expansion that resulted in detectable lysis of the parental K562 
line.  However, an IFN-ϒ intracellular cytokine assay showed cell activation only after co-culture 
with the transduced lines, Raji cells, but not parental K562 as shown in Figure 6. 
 
 

 
 
 
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Similar results were obtained with Line #5 where there was lysis of the parental K562 line that 
was greatly reduced by depletion of CD56+ cells, but failed to stimulate IFN-ϒ production even 
without CD56+ cell reduction.  For both Line #4 and Line #5 depletion of CD56+ cells had little 
effect on the killing of CD19 or CD20 expressing target cells.  
 
Figure 6: Effector function of CAR-20/19-T cell line as demonstrated by cytotoxicity and 
IFN-ϒ production 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Lysis of the K562 parental line was markedly reduced after depletion of CD56+ cells in the 
cultures, demonstrating that this was NK-mediated lysis, as shown in Figure 7. 
 
 
 
 
 
 
 
 

 
 
 
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For all six lines that were generated, we recovered sufficient viable transduced cells to be able 
to meet the doses required for this phase I study. 
 
1.5 Clinical Data 
B-cell ALL 
ALL is a fatal lymphoid malignancy that can develop in patients of all ages. Although children 
with ALL have impressive outcomes with combination chemotherapy, those with high-risk 
cytogenetics or a Philadelphia (Ph) chromosome do not fare as well despite intensive therapy 
and allogeneic transplantation. Adults have significantly worse prognosis than children.[35] 
Therapies for relapsed or refractory ALL are limited, and many patients are unable to tolerate 
the successive lines of cytotoxic chemotherapy necessary to achieve disease control.[36]  
 
CD19 positivity is almost ubiquitous in ALL and is a marker used in diagnosis of the disease. 
Consequently, many of the first CD19 CAR-T cell studies involved patients with either relapsed 
or refractory ALL with remarkable outcomes. In a combined pediatric/adult ALL study, a 
complete remission (CR) was achieved in 90% of patients with relapsed ALL with sustained 
remissions in two-thirds of those patients.[6]  
 
B-cell Lymphomas 
There are several subtypes of B cell NHL, including Burkitt lymphoma, diffuse large B cell 
lymphoma (DLBCL), follicular lymphoma (FL), CLL/SLL (small lymphocytic lymphoma), mantle 
cell lymphoma (MCL), marginal zone lymphoma (MZL), and lymphoplasmacytic lymphoma 
(LPL)/Waldenstroms Macroglobulinemia (WM). As these are mature B cell malignancies, CD20 
expression is almost universal, and treatment with rituximab is standard of care in all 
subtypes.[37] Standard first line therapy includes the CD20 antibody rituximab in combination 
with cytotoxic chemotherapy. After relapse, salvage chemotherapy +/- autologous transplant are 
considerations and can cure a proportion of patients. However, treatment options for those 
failing second line treatment or refractory to first or second line treatment are quite limited.[38]  
 
Similar to CD20, CD19 expression is also seen in most B cell lymphomas although the level of 
expression varies with histologic subtype.[39]  CD19 CAR-T therapy resulted in dramatic 
responses in a heavily pretreated population of patients with multiple histologic subtypes of 
lymphoma. The initial publication for CAR-T cell therapy against CD19 demonstrated significant 
efficacy in 3 patients with relapsed, refractory CLL[40]. In a single arm study utilizing a CD28, 
CD3ζ co-stimulatory domain CAR-T cells, 8/15 patients treated with anti-CD19 CARs achieved 
a CR. Four of seven patients with chemo refractory DLBCL achieved CR; these were durable in 
3 patients.[9] In another study, presented at ASH 2015, patients with relapsed, refractory 
DLBCL, MCL, and FL treated with  CD19 CAR-T cells had an overall response rate of 68% with 
minimal toxicity.[10] More recent data from ASH 2016 from KITE, a large pharmaceutical 
 
Figure 7: Effect of 
CD56+ cell depletion 
on lysis of parental 
K562 targets 

 
 
 
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company evaluating CD19 CAR-T cells, demonstrated an overall response rate of 76% in 
patients with relapsed, refractory NHL with 39% of patients having an ongoing response at 3 
months.[41] Additionally, from ASH 2016, ibrutinib refractory patients receiving CAR-T cells 
against CD19 were found to have an overall response rate of 76% with CRs achieved in 5 
patients.[42] These above data support the efficacy of CAR-T cells in both NHL and CLL.  
CAR-T Therapy 
The CAR approach uses genetically programmed, patient-derived lymphocytes transfected with 
chimeric receptor genes to combine the effector functions of T lymphocytes with the ability of 
antibodies to recognize predefined surface antigens with high specificity in a non-major 
histocompatibility complex (MHC) restricted manner.[43] Consequently, CAR-T cells have the 
ability to recognize intact membrane proteins independent of antigen processing or presentation 
by the host’s immune cells. CARs typically encode a single chain variable fragment (scFv) 
extracellular domain to bind tumor or virus linked to an intracellular signaling domain that 
mediates T cell activation. In principle, universal targeting vectors can be constructed because 
the scFv binds to native cell surface epitopes and bypass the requirement for MHC restriction 
(interaction with recipient HLA molecules on host immune cells). The scFv antibody, contains 
the VH and VL chains joined by a peptide linker of about 15 residues in length.[44]  We intend to 
use a second generation CAR approach which provides costimulatory signaling domains 4-1BB 
and CD3ζ together with two scFv chains recognizing both CD19 and CD20.[15, 45] (Figure 3) 
There are several potential limitations to activity of CAR-T cells: 1) the tumor must express the 
target antigen on the cell surface; 2) large amounts of shed or soluble antigen could inhibit the 
CAR-T cells; 3) the chimeric receptor may be immunogenic, resulting in the elimination of the 
redirected T cells by the host immune system.  
 
1.6 Rationale for Lymphodepletion and CAR-T cell dosing 
Lymphodepletion 
It is increasingly clear that adoptive immunotherapy strategies are more effective in the context 
of homeostatic T cell proliferation – the proliferation of naive T cells and their differentiation into 
memory-like T cells when total numbers of naive T cells are reduced by lymphodepleting 
chemotherapy. Lymphodepletion also eliminates regulatory T cells and other competing 
elements of the immune system that act as “cytokine sinks”, enhancing the availability of 
cytokines such as IL-7 and IL-15.[46, 47] In this protocol we propose to transfer CAR-20/19-T 
cells into subjects that are rendered lymphopenic as a result of cytotoxic chemotherapy. 
 
The optimal regimen for lymphodepletion remains unknown. Various CAR trials have utilized a 
combination of fludarabine, cyclophosphamide, pentostatin, or disease specific 
chemotherapeutic agents.[7, 9, 10, 40, 48, 49] Given the heterogeneity of previous CAR trials 
and using the previously published literature, we will, like most other institutions, utilize a 
combination of a purine analog (fludarabine) with cyclophosphamide, which is the most common 
lymphodepletion regimen used.[48, 49] The regimens for this trial is listed in section 6.1. 
 
Dose Rationale 
Previous literature has demonstrated a response to CAR-T cells at a variety of dose levels from 
1.0-5.0 x106 cells/kg to 1.0-5.0x109 CAR-T cells as a flat, weight independent dose.[9, 10, 40] 
Although an optimal dose has not been determined, based on prior studies, our goal dose level 
is 2.5 x 106 cells/kg. However, due to concern for increased toxicity with a dual targeted CAR-T 
cell, we will start at a dose level of 2.5 x 105 cells/kg and dose escalate per our study design 
(Section 6.4) to our target dose. Based on FDA recommendations we will dose per kg for 
uniformity and the dose will be maxed at a weight of 80 kilograms. Recent data regarding cell 

 
 
 
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dose optimization presented at the American Society of Clinical Oncology meeting in 2016 
demonstrated that administration in split-dose fashion may improve safety without altering 
efficacy and that a flat cell dose of 108 was the optimal dose level. Higher doses led to more 
CRS related deaths. [50, 51] Based on that data we will split our infusion during the phase 1 
portion over 2 days, 30% on Day 0 and 70% on Day 1. If there is no significant toxicity during 
the phase 1 portion, we will transition to single day dosing in our Phase 1b portion of the study. 
Phase 1b will be open to enrollment once a subject has consented for the final slot on the 
Phase 1 portion. Our target dose was chosen on the basis of efficacy and safety data from 
CD19 directed CAR-T cells with similar 4-1BB and CD3ζ co-stimulatory domains.[48, 49] Given 
that CAR-T cell expansion occurs in vivo post infusion, there has not been clear evidence of 
dose response related to efficacy or toxicity.  
 
1.7 Known and Potential Risks and Benefits 
Potential Risks 
Patients enrolled on this study will be exposed to a genetically engineered autologous T cell. 
Potential risks of the CAR-20/19-T cells include the following: 
 
• 
B cell ablation and consequent increased risk of infection: As both healthy and malignant 
B cells express CD19 and CD20 antigens, B cell ablation with consequent elimination of 
immunoglobulin production is an expected side effect of administration of the CAR-
20/19-T cells. This may be transient or permanent depending on the longevity of the 
CAR-T cells; it generally resolves when the CAR-20/19-T cells are cleared. Patients with 
persistent low levels of immunoglobulins can be given intravenous immune globulin if felt 
to be indicated by their treating physician.  
 
• 
Immune Reaction: Although we are utilizing autologous T cells, it is possible that the 
processing will make these cells immunogenic and that patients will have an immune 
response directed against the scFv. If such a response occurs, it is possible that the 
cells will be rejected or a serious allergic reaction will occur. Thus far, such allergic 
reactions have not been reported in clinical studies of CAR-T cells. [6, 8, 9, 40] 
Additionally patients can get exacerbation of pre-existing auto-immune conditions or 
development of new auto-immune or neurologic conditions.  
 
• 
Transformation: Gene transfer may result in new tumors derived from genetically 
modified cells, due to incorporation of viral DNA, which may dysregulate genes 
controlling cell apoptosis and proliferation leading to an uncontrolled malignant clone or 
new hematologic malignancy. Again, this is not yet reported in clinical studies of CAR-T 
cells in human patients, which now number greater than 100. [6, 8, 9, 40] 
 
• 
Graft-Versus-Host Disease (GVHD): In patients who have underwent previous 
allogeneic transplant and then relapse with disease and receive CAR-T therapy with 
allogeneic cells, one potential risk is re-activation of GVHD. This was not seen in one 
recent prospective study of patients receiving allogeneic CAR-T cells from their donor. 
[52] 
 
• 
Risk of tumor lysis syndrome (TLS) related to lymphodepleting chemotherapy or CAR-T 
cells: The risk of TLS is dependent on the burden of disease and rapidity of response. 
There are reports of patients treated with CD19 CAR-T cells developing tumor lysis 
syndrome. Therefore, all patients will be closely monitored both before and after 

 
 
 
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chemotherapy and CAR-20/19-T cell infusions including blood tests for potassium, uric 
acid, and calcium. All subjects will receive allopurinol prophylactically. TLS resulting in 
renal insufficiency, or rapidly rising uric acid, or evidence of organ dysfunction will be 
managed with intravenous fluids and rasburicase as needed and determined by the 
treating physicians. Appropriate clinical therapy will be administered should any 
significant TLS occur. 
 
• 
Cytokine Release Syndrome (CRS): CRS is an inflammatory syndrome that occurs as a 
result of T cell activation and proliferation associated with engagement of the targeted 
antigen. It manifests when large number of lymphocytes become activated and release 
inflammatory cytokines. Its symptoms (Figure 8) can vary from mild flu-like complaints 
with fevers, anorexia, nausea/vomiting, headache, and rash to a severe inflammatory 
storm that can result in hypotension, multi-system organ failure, shock, and even death 
despite treatment with anti-cytokine therapy and steroids. [1, 53, 54]  It is associated with 
markedly increased production of acute phase reactants such as C-reactive protein 
(CRP) and ferritin as well as increases in the erythrocyte sedimentation rate (ESR).  The 
presentation of CRS is most similar with MAS (macrophage activation syndrome) or HLH 
(hemophagocytic lymphohistiocytosis) and is felt to be a reaction to the immune 
activation that occurs with antigen binding by the CAR-T cells. Diagnosis of MAS or HLH 
is based on the 2004 HLH criteria (Appendix C). [55]                      
 
It is unclear if the development of CRS predicts clinical response.  Initial reports of CRS 
in CAR-T cell clinical trials revealed dramatically increased levels of IL-6 that peak 
during maximal T cell proliferation. Inhibition of IL-6 binding with the IL-6 receptor 
antibody, tocilizumab results in a rapid and dramatic improvement in the life-threatening 
symptoms associated with CAR-T therapies. It is unclear if this inhibition dampens the 
efficacy of the modified T cells.[53, 56]  Given the unique presentation of CRS 
associated with CAR-T cell therapy, a revised grading system has been recommended 
for appropriate classification of degree of toxicity. We will employ this revised grading 
system when evaluating patients with CRS in this study.[1]  
 
 
 
Figure 8  
Clinical Symptoms 
associated with CRS [1] 
 
 
 
 
 
 
 
 
 
 
• 
CRS related Neurological Toxicity: Neurological toxicity is a specific risk that has been 
observed with anti-CD19 CAR-T cell therapy. This has manifested with symptoms such 
as hallucinations, delirium, aphasia, and confusion. These toxicities range from mild to 
severe (requiring ventilator support for airway protection).[1, 53, 57, 58] Neurological 
toxicity generally emerges within the first week after anti-CD19 CAR-T cell infusion, and 

 
 
 
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resolves within a few days. Similar neurological changes are observed with 
blinatumomab administration and CD28 mAb ligation, for which it has been speculated 
that neurological toxicity arises from generalized T cell mediated inflammation rather 
than direct toxicity mediated by T cells on CNS tissue.[56] The neurological toxicities 
reported with anti-CD19 CAR-T cell therapy to date have generally been medically 
manageable and reversible. However, recently Juno Therapeutics reported 3 deaths due 
to CNS toxicity as a result of their anti-CD19 CAR-T cell trial.[59] Their CAR utilizes a 
different signaling domain than the one proposed in this study. In addition, we will 
require prophylactic levetiracetam in all patients at the time of lymphodepletion 
chemotherapy to prevent potential neurotoxicity.  
 
• 
Infection: Patients who receive fresh CAR-T cells are at risk of developing an infection 
as results of complete sterility testing will not be available until after infusion. To 
decrease this risk, in process testing (Appendix D) will be performed prior to start of 
lymphodepletion chemotherapy. Should the cultures return positive post-infusion 
appropriate antibiotics and infectious disease work-up will be performed.  
 
Potential risks associated with lymphodepleting chemotherapy 
A combination of fludarabine with cyclophosphamide will be utilized for lymphodepletion in all 
patients undergoing treatment with CAR-20/19-T cells. Potential risks associated with this 
combination chemotherapy include the following 
 
• 
Cytopenias: Cytotoxic chemotherapy can result in drug-induced pancytopenia placing 
the patient at risks of several complications including bleeding related to 
thrombocytopenia, infection related to neutropenia, or fatigue/hypotension associated 
with anemia. Patients will be allowed to receive prophylactic transfusions as indicated in 
Appendix E. In the event of neutropenic fever, patients will be appropriately managed 
with broad spectrum antibiotics.  
 
• 
Tumor Lysis Syndrome: As defined above, patients will be monitored for TLS post 
treatment with both lymphodepleting and CAR-20/19-T cell treatment.  
 
• 
Organ Toxicity: Any cytotoxic chemotherapy regimen can result in organ damage as 
defined as hypoxia (requiring O2), an elevated creatinine, neurological toxicity, or 
elevations in liver function tests. These parameters will be monitored per our follow-up 
schedule in Table 1.  
 
Unknown Risks 
In the setting of genetic therapy and a first-in-human study there are potential unknown risks. 
The degree of CRS with dual antigen binding has not been previously tested; it is possible that 
this will lead to a more robust immune response, and therefore more severe CRS, than 
previously seen. Additionally, as there is limited long-term data in patients treated with CAR-T 
cell therapies, it is unknown if there will be treatment-related secondary malignancies or other 
unexpected toxicities in the future.  
 
Potential Benefits 
Patients with relapsed leukemia or lymphoma as based on the inclusion criteria have very poor 
outcomes with standard treatments options. Allogeneic transplant can be offered to those 
patients in remission but this comes with high treatment related mortality and many patients are 
not eligible due to active or progressive disease.  Recent data published and presented at 

 
 
 
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national conferences establishes the efficacy of CAR-T cells in the treatment of relapsed, 
refractory leukemia and lymphoma [6, 8-10, 40] Although clinical responses are plausible, given 
this is a phase 1/1b study the clinical benefit of this specific CAR-20/19-T cell infusion is 
unknown.   
 
2 HYPOTHESIS AND OBJECTIVES 
 
2.1 Hypothesis 
Single antigen targeted monoclonal antibodies and CAR-T therapies have significant efficacy in 
the treatment of patients with lymphoma and leukemia. However, not all patients respond, and 
those that do respond can still experience progressive disease. In the setting of monoclonal 
antibodies such as rituximab (CD20 targeted antibody), one method of resistance has been the 
downregulation of the CD20 receptor leading to tumor cell escape.[16] Similarly, the proposed 
mechanism of relapse in CD19 positive ALL treated with CD19 CAR-T cells has been 
downregulation of the CD19 receptor and development of a clonal CD19 negative population.[7] 
Preclinical studies have demonstrated the decreased downregulation of tumor cell antigens 
when treated with combined CD20/CD19 CAR-T cells.[29]  We hypothesize that a dual targeted 
CD20/CD19 CAR-T cell treatment will be more efficacious than single antigen targeting and will 
decreased the risk of progression or relapse after treatment. 
 
2.2 Primary Objectives 
1. Determine the safety (dose level) of CAR-20/19-T cell administration in relapsed 
refractory CLL/SLL and B-cell NHL 
2. Determine the feasibility of manufacturing CAR-20/19-T cells locally from patient 
apheresis products using the CliniMACS Prodigy Cell processing device 
 
2.3 Secondary Objectives 
1. Determine the anti-tumor responses as measured by response rates.  
2. Describe the duration of response for responding patients 
3. Determine persistence of CAR-20/19-T cells 
4. Determine the effects of CAR-20/19-T infusion on non-neoplastic CD19 & CD20 B cells 
in vivo 
5. Determine if cellular or humoral host immunity develops against CAR-20/19-T cells  
6. Evaluate MRD using molecular technologies   
 
2.4 Rationale for the Outcome Measures Selection 
As a first-in-human and Phase 1 study, our primary outcome is to demonstrate both the safety 
and feasibility of CAR-20/19-T cell production and administration to patients. Production 
feasibility will be measured by our ability to perform apheresis and produce gene modified CAR-
20/19-T cells that can be administered to enrolled patients. Safety will be determined by 
assessments of CRS using the Lee et al. grading scale [1] and adverse events via NCI CTCAE 
version 5.0. Response assessment will vary by specific disease subtype and will occur initially 
at Day +28 post-treatment (+/- 3 days) with CAR-20/19-T cells. Ongoing response assessment 
will continue until progression and/or death up to 2 years (see Table 1), after which patients will 
be transitioned to a long-term follow-up protocol. 
 
At a minimum, the Day +28 (+/- 3 days) response assessment will include the following:  
 
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3 STUDY DESIGN 
3.1 General Description 
This is a phase 1/1b, interventional single arm, open label, treatment study designed to evaluate 
the safety and feasibility of infusion of CAR-20/19-T cells in adult patients with B cell 
malignancies that have failed prior therapies. The primary objective is safety of infusion and 
feasibility of manufacturing CAR-20/19-T cells at MCW-FH. We will monitor and describe safety, 
tolerability and engraftment potential of CAR-20/19-T cells in these patients. The general 
protocol schema is shown in Figure 1. 
Subjects who have adequate T cells as per eligibility criteria will be leukopheresed to obtain a 
MNC apheresis product for CAR-20/19-T manufacturing. The apheresis product will be washed 
free of platelets and plasma and the CD4+ and CD8+ T cells will be purified from the MNC, 
activated through the CD3 and CD28 antigens, then transduced with the CD19 and CD20 scFv 
regions coupled to CD3ζ/4-1BB signaling domains using a lentiviral vector. The cells will be 
expanded in vitro washed from culture, removed from the device, and either administered to the 
patient within 48 hours of production or cryopreserved for later administration. In process and 
end of production release testing will be performed (Appendix D for schema) The entire 
manufacturing process up until removal from culture will be performed using a computer 
controlled program within the closed system of the CliniMACS Prodigy device. 
  
Patients will receive CAR-20/19-T cells through a central venous catheter and then will be 
monitored closely with physical examinations and blood tests to assess safety, engraftment, and 
persistence of the CAR-20/19-T cells at regular intervals as outlined (Table 1). Response 
assessments will also be performed for secondary endpoint analysis as described in Section 
3.5. Patients will be followed daily from Day 0 to Day +7. Then seen on Day +10 (+/- 1 day), 
Day +14 (+/- 2 days), Day +21 (+/- 2 days) and then on Day 28 (+/- 3 days). After Day +28 
patients will be seen monthly up to 6 months, every 3 months up to year 1, and every 6 months 
up to year 2. Details of further follow-up are in listed in the study calendar. Following completion 
of this study, subjects will be enrolled onto a roll-over long-term follow-up protocol for up to an 
additional thirteen years to assess the diagnosis of long-term health problems, such as 
development of new malignancies and persistent of CAR-T cells. 
3.2 Number of Subjects  
A maximum of 24 patients will be treated with CAR-20/19-T cells. In the phase 1 portion an 
initial dose escalation cohort using 3+3 patients at the first dose level in this study. Once a safe 
dose level has been identified, there will be a 6 patient dose expansion cohort at the selected 
dose level. Once the final subject has enrolled on the Phase 1 portion of the study, the Phase 
1b portion of the study will be open to accrual of 9 patients).  
3.3 Primary Endpoint(s) 
Primary feasibility endpoints assessed include: 
Ability to manufacture CAR-20/19-T cells from patient apheresis products using the CliniMACS 
Prodigy processing device (Miltenyi Biotec). The number of T cell lines that do not meet criteria 
for T cell purity, total transduced T cell content, viability, and sterility will be determined.  
 
 

 
 
 
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Primary safety endpoints assessed include:  
 
Occurrence of adverse events, defined as either CRS related Grade 3/4 toxicity or other NCI 
CTCAE version 5.0 non-hematologic ≥ grade 3 signs/symptoms, laboratory toxicities and clinical 
events that are possibly, probably or definitely related to study treatment at any time from the 
infusion until week 4. The revised CRS grading system will be used for patients who develop 
CRS (Appendix G).[1] This will include infusion toxicity and any toxicity possibly related to the 
CAR-20/19-T cells including but not limited to:  
 
a. Fevers  
b. Rash  
c. Hepatic dysfunction  
d. Pulmonary infiltrates or other pulmonary toxicity (unrelated to concomitant infections) 
e. CRS/MAS 
f. Neurologic toxicity  
 
3.4 Dose Limiting Toxicity 
Dose-limiting toxicity (DLT) will be defined as a grade 3-4 non-hematologic toxicity possibly, 
probably, or definitely related to the infusion of CAR-20/19-T cells. All deaths (grade 5 toxicity) 
in the first 28 days felt to be either possibly, probably, or definitely related to CAR-20/19-T cell 
therapy will be considered a DLT. All Grade 4 non-hematologic toxicity felt to be either possibly, 
probably, or definitely related to CAR-20/19-T cell infusion will be considered a DLT. If multiple 
toxicities are seen, the presence of a DLT will be based on the most severe toxicity 
experienced. The DLT will be based on the tolerability observed during the first 28 days of 
treatment/observation. 
 
CRS and neurotoxicity are possible DLTs that can occur as a result of administration of CAR-T 
cells. For this protocol, any grade 4 CRS related toxicity per revised CRS grading system 
(Appendix G) will be considered a DLT. In addition, patients with Grade 3 CRS toxicity that 
does not improve to grade 1 or 2 despite treatment with cytokine inhibition and/or 
immunosuppression within 7 days will also be considered a DLT. All Grade 4 neurotoxicity will 
also be considered a DLT and similarly any Grade 3 neurotoxicity which does not improve to 
grade 1 or 2 despite treatment within 7 days will also be considered a DLT. All infusional 
reactions ≥grade 2 lasting more than 24 hours will be considered a DLT. Lastly, while 
hematologic toxicity is an expected side effect of lymphodepletion, persistent Grade≥3 
neutropenia that does not resolve to Grade≤2 neutropenia by Day +28 will be considered a 
DLT. Refractory thrombocytopenia and anemia are known potential complications among 
heavily pre-treated patients with hematological malignancies and lymphodepletion 
chemotherapy and will be managed per supportive care guidelines. While grade and toxicity will 
be recorded for this clinical trial the degree of thrombocytopenia/anemia will not count towards 
our DLT evaluation. 
 
The first 28 days were chosen for the DLT monitoring period given the variability in the 
presentation of CRS, which can occur within hours to weeks after infusion [1]. Additionally, as 
more severe CRS tends to occur earlier after infusion compared to non-severe CRS, 28 days 
was felt to be an appropriate monitoring period.[6] The attribution of adverse events with respect 
to their relationship to the infusion of CAR-20/19-T cells will be the responsibility of the site PI. 
 

 
 
 
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3.5 Secondary Endpoint(s)  
Secondary endpoints include assessment of the following: 
 
1. The number of patients who have CAR-20/19-T cells manufactured but not infused due 
to failure to meet release criteria.  
2. Disease assessment will include the following:  
o Number of patients with anti-tumor responses after CAR-20/19-T cell infusion. 
Since patients with a variety of histologies will be treated – response will be 
assessed using disease appropriate imaging/marrow and international 
consensus criteria most applicable to the disease (Section 10). 
o Repeat bone marrow biopsy and aspirate with evaluation of minimal residual 
disease when indicated (i.e., patients with baseline disease on bone marrow 
biopsy). These will be analyzed by multiparameter flow cytometry (MFC) and/or 
molecular analysis of immunoglobulin rearrangements, as available. 
o Duration of response in responders 
o Relapse rates and time to relapse  
o Cause of death in treated patients  
o Overall survival and progression free survival of individual patients 
 
   Exploratory Endpoints 
1. Characterize CAR-20/19-T cells with respect to their expansion, persistence, phenotype, 
and function. This will be assessed by flow cytometry, Q-PCR, RNA sequence, and in 
vitro functional assays.  
2. Determine if host immunity develops against the CAR-20/19-T cell construct or other 
elements of the transgene or vector and assess correlation with loss of detectable CAR-
20/19-T cells (loss of engraftment).  
3. Evaluate the effect of CAR-20/19-T cells on B cell and plasma cell compartments 
assessed by MFC to measure both benign and malignant B cell populations and plasma 
cell populations. 
4. Describe the incidence of new-onset secondary malignancies if any.  
5. Determine the utility of a T-cell diversity assay in predicting outcomes with CAR-T cell 
therapy 
 
3.6 Study Timeline 
Preclinical lines of CD19 and CD20/CD19 CAR T-cells have been generated in support of the 
IND application.  These studies have demonstrated that the MCW lymphocyte propagation 
laboratory can produce CAR-T cell lines using the Miltenyi CliniMACS Prodigy device.    
 
1) Submission of IND (1-month review time for FDA) 
2) Submission of clinical protocol through IRB, DSMC at local institution 
3) Clinical accrual after FDA and IRB approved protocol along with regulatory approval from 
local institutions (MCW and FH). 
 
3.7 Primary Completion 
The study is expected to reach primary completion 18-24 months from the time the study opens 
to accrual. 

 
 
 
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3.8 Study Completion 
Initial data on the safety, feasibility and short-term toxicity of CAR-20/19-T cells will be complete 
and available 4 months after completion of accrual. Thus, preliminary toxicity and efficacy data 
will reach completion 24 months from the time the study opens to accrual. 
 
4 PATIENT SELECTION 
 
4.1 Eligibility Criteria 
 
Patients must meet all the below inclusion and exclusion criteria prior to apheresis. After 
consent patients can receive additional bridging chemotherapy except during listed time periods 
in inclusion/exclusion criteria prior to apheresis and prior to CAR-20/19-T cell infusion.  
 
4.2 Inclusion Criteria  
1. Diagnosis of B-cell NHL or CLL/SLL: Patients must be aged≥18 years with relapsed, 
refractory disease and no available curative options that meet clinical criteria to initiate 
treatment. 
2. Patients with B-cell NHL or CLL/SLL must have either CD19 or CD20 positive disease 
on most recent biopsy performed (a repeat biopsy is not mandatory for this study except 
as noted below). A minimum of 5% CD19 or CD20 positivity by immunohistochemistry or 
flow cytometry on prior or repeat biopsy is required.  
3. Absolute CD3+ T cell count ≥50/mm3  
4. MRI brain and Lumbar Puncture with CSF analysis by cytology and flow cytometry 
without evidence of CNS involvement ONLY in patients with history of CNS involvement 
5. Measurable disease must have been documented within 4 weeks of the time of consent 
defined as the following by disease specific subtype: 
c. B-cell NHL: Active disease defined as nodal lesions greater than 20 mm in the 
long axis or extranodal lesions >10 mm in long and short axis or bone marrow 
involvement that is biopsy proven 
d. CLL/SLL: Active disease by either bone marrow, peripheral flow cytometry, or CT 
and/or PET imaging with nodal disease 
6. Patients should have failed at least two lines of a standard treatment and meet disease 
specific criteria detailed below: 
a. CLL/SLL: measurable disease as defined above that has relapsed after at least 
one line of chemo-immunotherapy and progressed or intolerant to ibrutinib 
monotherapy  
b. CD19 or CD20 positive B cell NHL limited to the following histologies: Advanced 
Stage III or IV Follicular Lymphoma, Diffuse Large B cell Lymphoma and 
associated subtypes (e.g. aggressive B-cell lymphoma, T-cell/histocyte rich B-
cell lymphoma, primary mediastinal B-cell lymphoma, EBV+ diffuse large B-cell 
lymphoma, transformed lymphoma such as transformed follicular or marginal 
zone lymphoma, and Richter’s transformation) and Mantle cell lymphoma. 
Specific criteria include: 
- 
Patients must have active, measurable disease after two lines of cytotoxic 
chemotherapy of which one must be anthracycline containing.  
- 
Must have received Rituximab or another CD20 antibody and at minimum two 
chemotherapy regimens appropriate for their disease.  
- 
Either failed autologous transplant or ineligible to receive autologous 
transplant 

 
 
 
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7. Karnofsky performance score ≥70. See Appendix A for scales. 
8. Normal Baseline Neurological Evaluation: Mini-Mental Status Exam Score 24-30 
(Appendix B) 
9. Adequate hepatic function, defined as AST and ALT <5 x upper limit of normal (ULN); 
serum bilirubin and alkaline phosphatase <5 x ULN, or considered not clinically 
significant as per the clinical PIs discretion (e.g. Gilbert’s or indirect hyperbilirubinemia) 
or felt to be due to underlying disease.  
10. Adequate renal function, defined as creatinine clearance>60 ml/min 
11. Able to provide written informed consent  
12. Agree to practice birth control during the study 
13. Adequate cardiac function as indicated by New York Heart Association (NYHA) 
classification I or II AND left ventricular ejection fraction of ≥35% (by cardiac ECHO or 
MUGA) and adequate pulmonary function as indicated by room air oxygen saturation of 
≥92%. 
14. Expected survival >12 weeks 
15. Negative urine or serum pregnancy test in females of child bearing potential at study 
entry and again within 48 hours’ prior lymphodepleting chemotherapy. 
16. Patients with prior blinatumomab treatment require repeat biopsy post-blinatumomab 
treatment that demonstrates CD19 or CD20 positive disease.  
17. Meet criteria for regarding fertility and contraception detailed in section 4.4 
18. Central line access will be required for CAR-20/19-T cell infusion. 
 
4.3 Exclusion Criteria 
A potential subject who meets any of the following exclusion criteria is ineligible to participate in 
the study. 
 
1. Positive beta-HCG in female of child-bearing potential defined as per table 1. 
2. Patients with known systemic allergy to bovine or murine products. 
3. Known prior positive serology for human anti-mouse antibody (HAMA).  
4. Confirmed active human immunodeficiency virus (HIV), Hepatitis B or C infection. 
5. History of significant autoimmune disease OR active, uncontrolled autoimmune 
phenomenon: such as systemic lupus erythematous, Wegner’s glomerulonephritis, 
autoimmune hemolytic anemia, idiopathic thrombocytopenic purpura (AIHA, ITP) 
requiring steroid therapy defined as >20 mg of prednisone or equivalent daily. 
6. Presence of ≥grade 3 non-hematologic toxicities as per CTCAE version 5.0 from any 
previous treatment unless it is felt to be due to underlying disease.  
7. Concurrent use of investigational therapeutic agents or enrollment on another 
therapeutic clinical trial at any institution. Minimum of ≥4 weeks required from 
administration of any other investigational agents on other clinical trials prior to 
enrollment on this CAR-T protocol. 
8. Refusal to participate in the long-term follow-up protocol 
9. Patients with active CNS involvement by malignancy on MRI or by lumbar puncture. 
a. Patients with prior CNS disease that has been effectively treated will be eligible 
providing treatment was >4 weeks before enrollment and a remission 
documented within 8 weeks of planned CAR-T cell infusion by MRI brain and 
CSF analysis. 
10. Previous recipients of allogeneic hematopoietic stem cell transplantation (AHCT) are 
excluded if they are <100 days’ post-transplant, have evidence of active graft-versus-
host-disease (GVHD) of any grade, or are currently on immunosuppression.  

 
 
 
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11. Previous CAR-T cell therapy directed at either CD19 or CD20 within 100 days of 
planned CAR-20/19-T cell infusion (does not include re-enrollment) 
a. Patients with prior CAR-T treatment against CD19 or CD20 must have repeat 
biopsy confirming a minimum of 5% CD19 or CD20 positivity by 
immunohistochemistry or flow cytometry 
12. Anti-CD20 antibody treatment within 4 weeks of cell infusion 
13. Anti-CD19 antibody treatment within 4 weeks of cell infusion 
14. Cytotoxic chemotherapy other than lymphodepletion within 14 days of CAR-T cell 
infusion  
15. Cytotoxic chemotherapy treatment within 14 days or steroid treatment (other than 
replacement dose steroids) within 7 days prior to apheresis collection for CAR-T cells 
16. Patients post solid organ transplant who develop high grade lymphomas or leukemias 
17. Concurrent active malignancy other than basal or squamous cell carcinomas of the skin 
 
 
4.4 Special Criteria for regarding Fertility and Contraception 
Female subjects of reproductive potential (women who have reached menarche or women who 
have not been post-menopausal for at least 24 consecutive months, i.e., who have had menses 
within the preceding 24 months, or have not undergone a sterilization procedure [hysterectomy 
or bilateral oophorectomy]) must have a negative serum or urine pregnancy test performed as 
part of eligibility criteria and again within 48 hours of initiation of lymphodepleting chemotherapy. 
 
Due to the high-risk level of this study, while enrolled, all subjects must agree not to participate 
in a conception process (e.g., active attempt to become pregnant or to impregnate, sperm 
donation, in vitro fertilization). Additionally, if participating in sexual activity that could lead to 
pregnancy, the study subject must agree to use reliable and double barrier methods of 
contraception during the follow-up period of the protocol. 
 
Acceptable birth control includes a combination of two of the following methods: 
• 
Condoms* (male or female) with or without a spermicidal agent. 
• 
Diaphragm or cervical cap with spermicide 
• 
Intrauterine device (IUD) 
• 
Hormonal-based contraception 
 
Subjects who are not of reproductive potential (women who are premenarche or have been 
post-menopausal for at least 24 consecutive months or have undergone hysterectomy tubal 
ligation, salpingectomy, and/or bilateral oophorectomy or men who have documented 
azoospermia) are eligible without requiring the use of contraception.   
 
4.5 Subject Recruitment and Screening 
Subjects will be identified through the clinical practices of the investigator or sub-investigators 
and through referrals from outside hospitals and physicians.  No direct-to-patient advertising will 
be performed.   
To be eligible, the subjects must have an adequate number of T cells (CD3+ ≥50/mm3) for CAR-
20/19-T cell line production, as determined from a peripheral blood sample before being 
considerable as eligible to undergo apheresis collection. The purpose of this screening 
procedure is to exclude subjects from participation who would otherwise undergo a futile 

 
 
 
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apheresis and restaging. We will additionally determine the % of CD4 and CD8 in circulation to 
ensure the CAR-20/19-T line contains both helper and effector T cells. 
5  STUDY ENTRY AND WITHDRAWAL; STUDY PROCEDURES 
5.1 Study Entry Procedures 
Required Preregistration Screening Tests and Procedures: 
 
The study-specific assessments are detailed in this section and outlined in Table 1. Screening 
assessments must be performed within 8 weeks prior to enrollment. Any results falling outside 
of the reference ranges may be repeated at the investigator’s discretion. All on-study visit 
procedures are allowed a window as noted in the study calendar. Treatment or visit delays for 
public holidays or weather conditions do not constitute a protocol violation. 
A written, signed informed consent form (ICF) and a Health Insurance Portability and 
Accountability Act (HIPAA) authorization must be obtained before any study-specific 
assessments are initiated. A signed ICF copy will be given to the subject and a copy will be filed 
in the medical record. The original will be kept on file with the study records.  
All patients who are consented will be registered in OnCore®, the MCW Cancer Center Clinical 
Trial Management System. The system is password protected and meets HIPAA requirements. 
Registration Process: 
Subjects will be identified through the clinical practices of the investigator or sub-investigators 
and through referrals from outside hospitals and physicians.   
 
To be eligible, patients need a diagnosis of CLL/SLL or B-cell NHL as specified in Section 4. 
CD19 or CD20 expression should be confirmed by IHC or flow cytometry (>5% expression of 
either CD19 or CD20) or MFC on malignant tissue, if it has not previously been documented.  In 
the Phase 1 portion, at each dose level, the first two patients will be monitored for a full 28 days 
after CAR-T infusion prior to subsequent patient’s treatment with CAR-T cells to allow 
monitoring of acute and sub-acute toxicities. After that patients will be infused no sooner than at 
14-day intervals among the specified dose level or in-between two dose levels. (e.g. 28 days 
between patient 1 and patient 2, and 28 days between patient 2 and patient 3). 
 
For the Phase 1b portion, please see section 6.6 for details on dosing. The PI on the study, Dr. 
Nirav Shah, should be contacted regarding any questions regarding subject eligibility. Contact 
information for the PI is listed on the front page of the protocol. 
 
5.2 Pretreatment Period 
Screening Assessments 
The screening procedures and assessments must be completed within ≤8 weeks prior to CAR-
20/19-T cells infusion (except as noted otherwise). For patients who fail to get their CAR-T cells 
within that time frame the studies below will have to be reported unless noted otherwise.  
 
Eligible subjects who have signed an informed consent and have adequate pre-screening 
evaluation will undergo a routine staging workup including: 
• 
History and Physical Examination including height and weight, Performance Status 

 
 
 
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Assessment (Karnofsky score), concomitant medications, and review of adverse events.  
• 
Vitals: Blood Pressure, Temperature, Respiratory rate, Pulse & Pulse Ox. 
• 
Radiologic imaging:  
o Diagnostic PET-CT scans or CT of the Neck, Chest, Abdomen, and Pelvis; If >8 
weeks’ elapses between screening assessment and planned CAR-20/19-T cells 
infusion, scans will be required to be repeated. 
• 
Complete Blood Count and differential. 
• 
Comprehensive Metabolic Panel: Albumin, Alkaline Phosphatase, AST, Bicarb, Bili T, 
Calcium (total), Chloride, Creatinine, Glucose, Potassium (K), Sodium (NA), Protein 
Total, BUN, and ALT. 
• 
Uric Acid 
• 
LDH 
• 
Ferritin, ESR, CRP 
• 
Lipid Panel 
• 
Beta-2-Microgloublin 
• 
Creatinine Clearance 
• 
Coagulation Assessments: PT/INR, PTT, Fibrinogen, and D-Dimer. 
• 
Thyroid Function Tests:  TSH, Free T4 
• 
Bone Marrow Transplant Markers/Apheresis Screening (Absolute numbers of CD3+ T 
cells, CD4+ T cells and CD8+ T cells should be assessed before scheduling for 
apheresis collection to determine protocol eligibility. Minimum of ≥50 CD3+ T cells/mm3 
required, with a mixture of both CD4+ and CD8+ T cells).  
• 
Quantitative Immunoglobulins (IgM, IgG, IgA) 
• 
Autoimmune Screen: ANA  
• 
Urinalysis 
• 
Serum or urine pregnancy test for females of child bearing potential 
• 
Infectious Disease Markers: HIV, Hepatitis B surface antigen, and HCV antibody within 8 
weeks prior to CAR-20/19-T cells infusion. If the HCV antibody is positive, a screening 
HCV RNA by any RT-PCR or bDNA assay must be performed. Patients with HIV or Hep 
B positivity will be excluded from the study. (Does not need to be repeated) 
• 
Bone marrow and aspirate evaluation within 8 weeks of CAR-20/19-T cells infusion 
• 
Baseline cardiac ECHO/MUGA (Does not need to be repeated if no new cardiac 
symptoms) 
• 
Electrocardiogram (ECG)  
• 
Lumbar puncture as specified in Table 1 and inclusion criteria (Does not need to be 
repeated if no active CNS symptoms) 
• 
MRI Brain (Only in patients with history of CNS disease or signs/symptoms of active 
CNS disease) 
• 
Baseline neurological assessment 
 
5.3 Enrollment 
Documents required: 
 Signed informed consent forms and eligibility documentation  
 Assignment of subject sequence number. 
 

 
 
 
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5.4 Study Procedures Related to Treatment 
5.4.1 Study Procedures – Day -15 (Apheresis day) 
 
Physical Examination including weight, concomitant medications, and review of adverse 
events.  
 
Vital signs: Blood Pressure, Temperature, Respiratory rate, Pulse & Pulse Ox. 
 
CBC with differential 
 
Comprehensive Metabolic Panel: Albumin, Alkaline Phosphatase, AST, Bicarb, Bili T, 
Calcium (total), Chloride, Creatinine, Glucose, Potassium (K), Sodium (NA), Protein 
Total, BUN, and ALT. 
 
Uric Acid 
 
LDH 
 
Apheresis 
 
T-cell repertoire diversity testing from apheresis sample 
 
Quality of Life Studies 
 
Tryptophan Metabolites 
 
PaxGene RNA gene expression 
 
5.4.2 Mononuclear Cell Apheresis  
A 2-5 blood volume apheresis will be targeted if tolerated by the patient, to be collected at 
Froedtert Hospital, as appropriate for CAR-20/19-T cell production. From a single apheresis, 
the intention is to harvest 1.0-5.0 x 109 MNC cells total to manufacture CAR-T cells. 
Baseline blood leukocytes for FDA requirements and for research are also obtained and 
cryopreserved.  
 
5.4.3 Study Procedures Week -1 + 6 weeks (~2 to 4 days prior to lymphodepletion) 
Patients will be seen 2-4 days prior to lymphodepleting chemotherapy. 
• 
Physical Examination including weight, concomitant medications, and review of adverse 
events.  
• 
Vital signs: Blood Pressure, Temperature, Respiratory rate, Pulse & Pulse Ox. 
• 
Urine or serum pregnancy test in females of childbearing potential  
• 
Viral Respiratory NAAT will be performed in patients with signs/symptoms concerning for 
viral infection (cough, cold). In the setting of a positive result, initiation of LDP 
chemotherapy will be delayed until resolution of symptoms 
• 
Urinalysis 
• 
Lumbar Puncture: For high risk patients with concern for CNS involvement, at the 
discretion of the PI, a repeat LP may be done prior to lymphodepleting chemotherapy 
• 
If >8 weeks’ elapse from staging bone marrow biopsy and aspirate, a repeat biopsy will 
be indicated 
• 
Confirm patient has central line access 
• 
Radiologic imaging:  
o Diagnostic PET-CT scans or CT of the Neck, Chest, Abdomen, and Pelvis; If >8 
weeks’ elapses between screening assessment and planned CAR-20/19-T cells 
infusion, scans will be required to be repeated. 
 
 

 
 
 
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5.4.4 Start of Lymphodepletion Chemotherapy (Day -4 + 6 weeks) 
Patients will then be seen again the day of initiation of Lymphodepletion chemotherapy. The 
following procedures must be completed prior to initiation of lymphodepleting chemotherapy 
 
 
Physical Examination including weight, concomitant medications, and review of adverse 
events.  
 
Vital signs: Blood Pressure, Temperature, Respiratory rate, Pulse & Pulse Ox. 
 
CBC with differential 
 
Comprehensive Metabolic Panel: Albumin, Alkaline Phosphatase, AST, Bicarb, Bili T, 
Calcium (total), Chloride, Creatinine, Glucose, Potassium (K), Sodium (NA), Protein 
Total, BUN, and ALT. 
 
Uric Acid 
 
LDH 
 
T-cell repertoire diversity testing from CAR-T sample 
 
A chemotherapy cycle (3 days of chemotherapy with a day of rest) is planned for all patients 
to start within approximately one week of CAR-20/19-T cell infusion. Patients will receive a 
combination of fludarabine/cyclophosphamide lymphodepletion. These chemotherapy 
agents are all commercially available and will be infused per institutional standards. 
Antimicrobial prophylaxis and TLS prophylaxis is outlined in Appendix E. 
 
The chemotherapy will be planned so that it completes BEFORE the infusion of the first 
dose of CAR-20/19-T cells. The purpose of the chemotherapy is to induce lymphopenia to 
facilitate expansion of infused T cells. Although the chemotherapy is not investigational, 
patients will be required to receive it at Froedtert Hospital (FH) and/or the Medical College of 
Wisconsin Cancer Center. Patients will be started on allopurinol for tumor lysis prevention.  
5.4.5 CAR-20/19-T Cell Infusion (+ 6 weeks) 
Infusion begins 1-7 days after the completion of chemotherapy and will be administered over 
two days (Day 0 and Day +1) in the Phase 1 portion. In the phase 1b portion cells will be 
administered as a single dose on Day 0.  Patients can receive cells fresh, if appropriate or 
after cryopreservation for up to 6 weeks post-harvest.  Patients will be either admitted to FH 
under the care of the hematology/oncology service for the cell infusion or seen daily in the 
FH Day Hospital for infusion from Days 0-7. Details of the infusion and hospitalization plan 
can be found in the treatment plan section 6. On these days, the following evaluations will 
occur prior to CAR-20/19-T cells infusion unless stated otherwise:  
 
• 
Physical Examination including weight, Performance Status Assessment (Karnofsky 
score), concomitant medications, and review of adverse events.  
• 
Vital signs: Blood Pressure, Temperature, Respiratory rate, Pulse & Pulse Ox 
• 
CBC with differential 
• 
Comprehensive Metabolic Panel: Albumin, Alkaline Phosphatase, AST, Bicarb, Bili T, 
Calcium (total), Chloride, Creatinine, Glucose, Potassium (K), Sodium (NA), Protein 
Total, BUN, and ALT.  
• 
Uric Acid  
• 
LDH  
• 
Ferritin/ESR/CRP 
• 
Bone Marrow Transplant Markers (Day 0 only) 
• 
Quantitative Immunoglobulins (IgM, IgG, IgA) only on Day 0 prior to infusion 
• 
Electrocardiogram (ECG)  

 
 
 
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• 
Coagulation Assessments: PT/INR, PTT, Fibrinogen, and D-Dimer. 
• 
Research Studies  
o T-Cell Persistent Studies: Baseline sample to be drawn prior to CAR-20/19-T 
cells infusion on Day 0 only. 
o Cytokine Studies: At minimum, will include IFN-ϒ, IL10, IL6 and TNFα 
o Q-PCR for Integrated Vector DNA baseline sample on Day 0 prior to infusion 
o Detectable RCL by Q-PCR for VSV-G baseline sample on Day 0 prior to infusion 
 
5.4.6 Day +1 through day +7 after cell infusion 
Patients will be seen daily through day+7 of infusion. This will occur either in the inpatient 
setting or with daily visits to the Day Hospital through day+7. At these evaluations, the 
following will occur:  
 
• 
Physical Examination including weight, concomitant medications, and review of adverse 
events.  
• 
Vital signs: Blood Pressure, Temperature, Respiratory rate, Pulse & Pulse Ox. 
• 
CBC with differential 
• 
Comprehensive Metabolic Panel: Albumin, Alkaline Phosphatase, AST, Bicarb, Bili T, 
Calcium (total), Chloride, Creatinine, Glucose, Potassium (K), Sodium (NA), Protein 
Total, BUN, and ALT. 
• 
Uric Acid 
• 
LDH 
• 
Coagulation Assessments: PT/INR, PTT, Fibrinogen, and D-Dimer. 
• 
Ferritin/CRP/ESR 
• 
Research Studies as per schedule of events (Table 1) 
o T-Cell Persistent Studies—Will not be repeated once there are two consecutive 
negative results. 
o Cytokine Studies 
o Neurological assessments 
o T-cell repertoire diversity testing from peripheral blood 
 
 
5.4.7 Follow-Up Visits (Day+10 to 2 years) 
ONGONG RESPONSE 
Patients with an ongoing response will be followed for up to 2 years after CAR-20/19-T cells 
infusion as per this protocol. After Day 28, testing can be performed at the local oncologist 
office and results sent to MCW/FH other than dates mentioned in the study calendar. The 
following procedures will be performed at specified intervals, see Table 1 per schedule of 
events. 
• 
Physical Examination, including weight, Performance Status Assessment (Karnofsky 
score), concomitant medications, and review of adverse events.  
• 
Vital signs: Blood Pressure, Temperature, Respiratory rate, Pulse & Pulse Ox 
• 
CBC with differential 
• 
Comprehensive Metabolic Panel: Albumin, Alkaline Phosphatase, AST, Bicarb, Bili T, 
Calcium (total), Chloride, Creatinine, Glucose, Potassium (K), Sodium (NA), Protein 
Total, BUN, and ALT. 

 
 
 
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• 
Uric Acid 
• 
LDH 
• 
Coagulation Assessments: PT/INR, PTT, Fibrinogen, and D-Dimer 
• 
Lipid Panel 
• 
Thyroid Function Tests:  TSH, Free T4 
• 
Quantitative Immunoglobulins (IgM, IgG, IgA) 
• 
Urinalysis 
• 
Bone Marrow Transplant markers  
• 
Beta-2-Microglobulin 
• 
Ferritin/ESR/CRP 
• 
Radiologic imaging:  
o CT neck/chest/abd/pelvis or PET/CT skull base to mid-thigh 
o At 1 month, 6 months, and 1 year. From 1 year to 2 years, scans will be done 
yearly.  
a) Scans would only be completed in follow-up only for patients who remain 
in remission since entering the study. Patients with progressive disease 
will not require any further imaging.  
• 
Bone Marrow Biopsy and aspirate as per Table 1 
• 
Electrocardiogram (ECG) 
• 
Research Studies: 
o T-Cell Persistent Studies—Will not be repeated once there are two consecutive 
negative results. 
o Q-PCR for Integrated Vector DNA—Will not be repeated once there are two 
consecutive negative results. 
o Detectable RCL by Q-PCR for VSV-G—Will not be repeated once Q-PCR for 
integrated vector DNA is negative at two consecutive timepoints.  
o Cytokine Studies  
o Neurological assessments 
o T-cell repertoire diversity testing from peripheral blood 
o Quality of Life Studies 
o Tryptophan Metabolites 
o PaxGene RNA gene expression 
 
VISITS REQUIRED AT MCW/FH: Beyond Day +28, patients will have required visits at 
MCW/FH on Day +90 (+/- 7 days), Day+180 (+/- 14 days), Day 365 (+/- 1 month), +1.5 
years (+/- 1 month) and 2 years (+/- 1 month). For timepoints not falling on those dates, local 
oncology evaluation will suffice and records will be obtained accordingly.  
 
PROGRESSIVE DISEASE 
For patients who develop progressive disease will not follow the above study calendar and will 
placed in a follow-up plan as detailed below with visit with their local oncologist every 6 months 
starting from CAR-T infusion date for up to 2 years and if alive will subsequently roll into our 
long-term follow-up protocol. At these visits the following should be performed 
 
• 
Physical Examination, including vitals 
• 
Survival/Progression status 
• 
Subsequent Treatments Post-CAR-T infusion 
• 
Targeted concomitant medications 
o Including IVIG, immunosuppressive medications, prophylactic anti-infective 
medications 

 
 
 
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• 
Targeted adverse events recording 
o Including New malignancies, new incidence or exacerbation of pre-existing 
neurologic disorder, new incidence or exacerbation of prior rheumatologic 
disorder, or other autoimmune condition, and new incidence of hematologic 
disorder 
• 
CBC with differential 
• 
Comprehensive Metabolic Panel: Albumin, Alkaline Phosphatase, AST, Bicarb, Bili T, 
Calcium (total), Chloride, Creatinine, Glucose, Potassium (K), Sodium (NA), Protein 
Total, BUN, and ALT. 
• 
Quantitative Immunoglobulins (IgM, IgG, IgA) 
• 
Absolute B and T cell lymphocyte counts   
• 
Research Studies: 
o Patients will be required to have blood sent to MCW (See Appendix K for 
instructions on sending samples to MCW) or return to MCW for testing at the 
following timepoints until testing is negative for persistence of CAR-T cells: +6 
months, +1 year, +1.5 years, and +2 years post-CAR-T infusion and will include 
the following listed below. Once there are TWO negative results, no further visits 
to MCW or blood samples to MCW will be mandated:  
a) T-Cell Persistent Studies—Will not be repeated once there are two 
consecutive negative results. Additional samples (blood, CSF, ascites, 
tissue, etc.)  for T-cell persistence can be performed per the investigators 
discretion as felt to be clinically indicated.  
b) Q-PCR for Integrated Vector DNA—Will not be repeated once there are 
two consecutive negative results. 
c) Detectable RCL by Q-PCR for VSV-G—Will not be repeated once Q-PCR 
for integrated vector DNA is negative at two consecutive timepoints.  
 
  
5.4.8 Long-term/Survival Follow-Up Procedures 
 
Patients will be followed for up to 2 years after CAR-20/19-T cells infusion or until progression. 
Patients who progress will be followed for survival, progression, persistence of CAR-T cells, and 
future treatments. Patients who progress post treatment will not have any further required 
disease assessment on study, and management of their disease will return to their treating 
physician. They will be required to return for other study laboratory testing and evaluations as 
mandated per the protocol as outlined above. Patients who are alive beyond 2 years will be 
placed on a long-term follow-up protocol for a total of up to 15 years for monitoring of post-
genetically modified therapies. 
 
5.5 Study Withdrawal Procedures 
Patient-Initiated Withdrawal:  A patient may decide to withdraw from the study at any time. 
 
Investigator-Initiated Withdrawal:  The Investigator will withdraw a patient whenever 
continued participation is no longer in the patient’s best interests. Reasons for withdrawing a 
patient include, but are not limited to, disease progression, the occurrence of an adverse event 
or a concurrent illness that prevents further administration of treatment, a patient’s request to 
end participation, a patient’s noncompliance or simply significant uncertainty on the part of the 
Investigator that continued participation is prudent. There may also be administrative reasons to 

 
 
 
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terminate participation, such as concern about a patient’s compliance with the prescribed 
treatment regimen.  
 
Withdrawal Documentation Procedure:  The reason for study withdrawal and the date the 
patient was removed from the study must be documented in the case report form. 
6 
TREATMENT PLAN 
6.1 Lymphodepletion (within 1 week prior to CAR-20/19-T cell infusion) 
Patients will receive lymphodepleting conditioning regimen prior to CAR-20/19-T cells starting 
on Day -4. Lymphodepletion will start based on the planned date of infusion of CAR-20/19-T 
cells. Tumor lysis prevention with allopurinol daily (dose per institutional standards) will be 
initiated the day of chemotherapy. Levetiracetam (Keppra) prophylaxis dosed per institutional 
standards will be initiated in all patients to prevent possible neurologic toxicity. Anti-emetics and 
steroids can be provided pre-treatment as per standard of care protocols for these treatment 
regimens. Please see Appendix E for guidelines for CAR-T cell infusion. The lymphodepletion 
regimens is as follows:  
 
Fludarabine 30 mg/m2 on Day -4, -3, -2 and Cyclophosphamide 500 mg/m2 on Day -2[41] 
 
This regimen is based on published lymphodepletion protocols for adult patients receiving CD19 
CAR-T cells. Fludarabine and Cyclophosphamide combination is the most utilized 
lymphodepletion regimen prior to CAR-T cell therapy and the above regimen was presented at 
ASH 2016 and used in a large cohort of >100 patients with published safety and toxicity data. 
[41, 48] Fludarabine and Cyclophosphamide dosing will be based on actual body weight.  For 
patients who are neutropenic (ANC<1000 k/µL) at the start of lymphodepletion due to underlying 
hematologic disease or as a result of prior chemotherapy related toxicity, cyclophosphamide will 
be omitted from the lymphodepletion regimen.  
 
Phase 1 Dose Escalation/Expansion: CAR-20/19-T cells will be administered per actual body 
weight in the lymphopenic phase on Day 0 & Day 1. Dose will be maxed at a weight of 80 
kilograms.  
 
Phase 1b Cohort : CAR-20/19-T cells will be administered as a single dose on Day 0. Dose will 
be maxed at a weight of 80 kilograms 
 
6.2 Preparation of CAR-20/19-T Cell Infusion 
Cell Manufacturing 
The MNC, Apheresis product obtained from the patient on Day -15 will be washed free of 
platelets and plasma and the CD4+ and CD8+ T cells will be purified from the MNC, activated 
through the CD3 and CD28 antigens, then transduced with the CD20/CD19 CD3ζ/41BB 
lentiviral vector. The cells will be expanded in vitro, harvested from culture, and then 
administered within 48 hours of production in an isotonic electrolyte solution (Plasma-Lyte® A or 
Normosol® R) containing 2.5% HSA or cryopreserved for administration in a freezing solution of 
Plasmalyte A or Normosol R containing 10% DMSO and 4% HSA. 
 

 
 
 
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Testing will be performed at two mandatory time points (in process day-6 and at harvest) and 
will include the following (Please see Appendix D for details on testing, including those tests for 
which results are required for product release):  
 
In-Process Testing 
• 
Transduction efficiency 
• 
Viability 
• 
Gram Stain 
• 
In-house Mycoplasma 
 
Final Product Testing (Harvest) 
• 
Sterility  
o endotoxin 
o mycoplasma 
o bacterial and fungal cultures  
• 
Phenotype 
o CD3, CD4, CD8, central memory and effector memory T cell subsets, 
o NK cells at harvest 
o NK-T cells at harvest 
o B cells at harvest 
o T regs at harvest 
• 
Viability and viable cell numbers 
• 
Effector activity (lysis and cytokine product in response to B cell targets) 
• 
Copy number (qPCR) at harvest 
• 
VSVg (qPCR) at harvest 
• 
% transduced cells (light chain expressing T cells) 
 
Cells will be harvested on Day 14 of production for fresh administration or cryopreserved for 
later infusion (Appendix D). The day cells are ultimately administered will be deemed Day 0 for 
study calendar and schedule of activities. All testing will be performed in accordance with FACT 
and FDA regulations. The entire manufacturing process up until removal from culture will be 
performed by a computer- controlled program using the CliniMACS Prodigy device. 
  
Cell thawing (if indicated)  
For patients who require cryopreservation the following will be performed: The frozen cells will 
be thawed in the cell processing lab and transported by laboratory staff to the subject’s bedside 
either at the inpatient oncology unit of FH or the outpatient day hospital under the care of a 
Hematology/Oncology attending. If the CAR-20/19-T cell product appears to have a damaged or 
leaking bag, or otherwise appears to be compromised, it should not be infused, and should be 
returned to production laboratory.  
 
Positive Sterility Test (after infusion) 
In the scenario that CAR-T cells are administered prior to completion of the 14 days of sterility 
testing, it is possible that a sterility test returns positive after administration. In that scenario, we 
will follow our standard operating procedure for positive cultures after infusion of cell therapy 
products. In general, this involves the following: 
 
1. Cell processing lab will contact the PI on the study at the time of this finding.  
2. The PI will discuss these findings with the patient 

 
 
 
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3. The PI will order appropriate infectious work-up and provide antimicrobial coverage as 
indicated based on the results of the sterility tests.  
4. Reporting—the PI on the clinical trial will report this to the DSMC, IRB, and the FDA 
 
6.3 Administration of CAR-20/19-T cell infusion on Day 0 and Day +1 (Phase 1) or Day 0 
(Phase 1b) 
Patients will receive the infusion of the CAR-20/19-T cells as an inpatient on the oncology unit 
or in the outpatient Day Hospital under the clinical care of hematology/oncology attending. 
Patients will receive institutional standard for infusion reaction prophylaxis (e.g., Acetaminophen 
and diphenhydramine hydrochloride, or other acceptable alternatives, as pre-treatment). 
Patients will be on allopurinol for tumor lysis prevention and levetiracetam (Keppra) with 
initiation of lymphodepleting chemotherapy (See Appendix E for details). Pretreatment with 
steroids is contraindicated.  Patients will receive CAR-20/19-T cells by IV infusion via central 
line for a total viable, transduced, cell dose ranging from 1.0 x 105 cells/kg to 2.5 x 106 cells/kg 
based on phase of study. The dose of cells may be limited if calculations performed at the time 
of administration or cryopreservation indicates that the patient would receive > 1.8 x 108 non-
viable cells/kg. Patients who fail to meet the target cell dose will be allowed infusion and will be 
followed for toxicity and adverse events as all other patients. The infusions will occur 
approximately 1-7 days following chemotherapy.  
Premedication 
Potential side effects following T cell infusions include transient fever, chills, and/or nausea. The 
subject will be pre-medicated with acetaminophen and/or diphenhydramine hydrochloride as per 
institutional standards prior to the infusion of CAR-20/19-T cells. These medications may be 
repeated every six hours as needed. Epinephrine will be readily available in the rare scenario of 
an allergic reaction.  A course of non-steroidal anti-inflammatory medication may be prescribed 
if the patient continues to have fever not relieved by acetaminophen. It is recommended that 
patients not receive systemic corticosteroids such as hydrocortisone, prednisone, prednisolone 
(Solu-Medrol) or dexamethasone (Decadron) at any time, except in the case of a life-threatening 
emergency, since this may have an adverse effect on T cells. If corticosteroids are required for 
an acute infusional reaction, an initial dose of hydrocortisone 100 mg is recommended. 
 
Prophylaxis Therapies 
Patients will receive appropriate prophylaxis with CAR-T cell therapy as recommended in 
Appendix E. Briefly levetiracetam will be given for seizure prophylaxis, institutional standards 
will be applied for pneumocystis pneumonia (PCP), viral, and neutropenic prophylaxis.  
 
Infusion 
On the day of the first infusion (Day 0), patients will have a CBC with differential, and 
assessment of T-cell subsets prior to administration of CAR-20/19-T cells since chemotherapy 
is given in part to induce lymphopenia.   
 
The CAR-20/19-T cell dose will be administered by IV infusion. Dosing will be split during the 
phase 1 portion of the trial, with 30% given on day 0, and 70% given on day 1. If there is no 
significant infusional toxicity, in the phase 1b portion of the trial cells will be administered as a 
single dose on Day 0. As detailed above, the cells will be administered fresh or thawed if 
indicated in the LPL and placed in a validated transport container at 1-10oC and brought directly 
to the patient’s bedside. If the CAR-20/19-T cell product appears damaged or leaking, or 
otherwise appears to be compromised, it should not be infused, and should be returned to 
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units or in the outpatient day hospital BMT rooms, using precautions for immunosuppressed 
patients. 
 
The transduced T cells will be administered by intravenous infusion. The infusion nurse spikes 
HPC product bag and administers through blood tubing accordingly. The duration of the infusion 
will be approximately 5-15 minutes. Each infusion bag will have affixed to it a label containing 
the following: “FOR AUTOLOGOUS USE ONLY”. In addition, the label will have at least two 
unique patient identifiers such as the subject’s initials, birth date, and study number. Prior to the 
infusion, two individuals will independently verify all this information in the presence of the 
subject and so confirm that the information is correctly matched to the participant. Subjects will 
be infused and premedicated as described.  
 
Subjects’ vital signs (temperature, respiration rate, pulse, and blood pressure, pulse oximetry) will 
be done prior to dosing, at the end of the infusion and every 15 minutes thereafter for a minimum 
of 1 hour. A blood sample for determination of a baseline CAR-20/19-T cell level is obtained any 
time prior to the infusion and Day+1 (and sent to the LPL Labs) and then will be completed as per 
the study calendar. Emergency medical equipment (i.e., emergency trolley) will be available 
during the infusion in case the subject has an allergic response, or severe hypotensive crisis, or 
any other reaction to the infusion.  
 
Patients experiencing toxicities from their preceding cytoreductive chemotherapy will have their 
infusion schedule delayed until these toxicities have resolved. The specific toxicities warranting 
delay of T cell infusions include:  
 
1) Pulmonary: Requirement for supplemental oxygen >2L by nasal cannula to keep saturation 
greater than 95% or presence of radiographic abnormalities on chest x-ray that are progressive; 
2) Cardiac: New cardiac arrhythmia not controlled with medical management 
3) Hypotension: requiring pressor support 
4) Active Infection: Fever>101 F and/or positive cultures for bacteria, fungus, or virus. Patients 
who are unable to receive the CAR-20/19-T cells within 7 days of completion of chemotherapy 
will be removed from the protocol.  
 
Febrile reaction 
In the unlikely event that the subject develops sepsis or systemic bacteremia following CAR-T- 
cell infusion, appropriate cultures and medical management should be initiated.  
 

 
 
 
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Table 2: Regimen Description 
Study Drug 
Premedication3 
   Dose 
    Route 
Schedule 
Cycle 
Length 
CAR-20/19-T cells1       Tylenol PO            1.0 x 105-2.5 x 106cells/kg2      IV           30% on Day 0   
 (Phase 1)                   Benadryl IV/PO                                                                      70% on Day 1  
 
CAR-20/19-T cells          As above                  2.5 x 106 cells/kg2                IV        100% on Day 0 
 (Phase 1b) 
                                                                        
 28 days   
 
 
    28 days 
 
    
1Patients with ≥ grade 2 non-hematologic CTCAE toxicity will have the 2nd dose held up to 7 days until 
symptoms resolve or will not be administered at the discretion of the PI 
2Cell number include only viable T cells expressing the CAR as determined by flow cytometry. Cell dose maxed 
at 80 kg or if the infusion contains >1.8 x 108/kg non-viable cells. 
3Recommended premedication, dosed per institutional standards 
 
6.4 Dose Escalation Schedule 
For this phase 1/1b study we plan to include multiple histologies including CLL/SLL and B-cell 
NHL.  Based on previous CAR-T studies (as outlined in Section 1.6) our goal treatment dose is 
2.5 x 106 cells/kg. Due to concern that a dual CAR-T cell may increase risk of CRS our starting 
dose for this study will be 2.5 x 105 cells/kg (1 log reduction) and we will escalate based on 
DLTs to our goal dose. We will utilize a 3+3 escalation design for this study (Figure 9). At each 
dose level, the first two patients will be monitored for a full 28 days after CAR-T infusion prior to 
subsequent patient’s treatment with CAR-T cells to allow monitoring of acute and sub-acute 
toxicities. After that patients will be infused no sooner than at 14-day intervals among the 
specified dose level or in-between two dose levels. The first three patients with NHL or CLL/SLL 
will be treated with 2.5 x 105 cells/kg. If 0/3 patients experience a DLT, the dose will increase to 
7.5 x 105 cells/kg. If 1/3 patients at the 2.5 x 105 cells/kg experiences a toxicity, then an 
additional three patients will be treated at this dose level. If ≤1/6 patients have a DLT at this 
level, the dose will increase to 7.5 x 105 cells/kg. If ≥2/6 patients have a DLT at 2.5 x 105 
cells/kg, the dose will de-escalate to 1.0 x 105 cells/kg. If 2/3 patients in our first cohort have 
DLTs, the dose will also de-escalate to 1.0 x 105 cells/kg. If at the 1.0 x 105 cells/kg there are ≥2 
DLTs out of a maximum of 6 patients, the study will close due to excessive toxicity. If there is 0-
1 DLTs in 3-6 patients, the 1.0 x 105 cells/kg dose will be selected for expansion cohort (Figure 
10).  
We will utilize a similar dosing schema should we escalate to the 7.5 x 105 cells/kg dose. If there 
are 0-1 toxicities out of 3-6 patients, the dose will escalate to our goal dose of 2.5 x 106 cells/kg. 
If there are ≥2 toxicities in 3-6 patients at 7.5 x 105 dose, then the 2.5 x 105 cells/kg dose will be 
selected for our expansion cohort. At our goal dose level (2.5 x 106 cells/kg) if there is 0-1 DLTs 
out of 3-6 patients, this dose will be selected for expansion. If there are ≥2 toxicities in 3-6 
patients at the 2.5 x 106 cells/kg dose, then the 7.5 x 105 cells/kg dose will be selected.  
Once a dose has been selected within the NHL and CLL/SLL cohort, this dose will be utilized for 
the Dose Expansion phase of the study (Section 6.5). The dose expansion cohort will proceed 
only after approval from the local IRB/DSMC and the FDA. 

 
 
 
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Table 3: Dose Escalation CAR-T cells 
 
Dose Level 
Dose of Study Drug* 
Minimum # of Patients/Dose 
Level 
-1 
1.0 x 105 cells/kg 
2 
0 
2.5 x 105 cells/kg 
2 
1 
7.5 x 105 cells/kg 
2 
2 
2.5 x 106 cells/kg (goal cell dose)  
2 
 
Patients who Fail to Produce Target Cell Dose 
 
It is possible that some patients fail to make the targeted cell dose as required per our dose 
escalation/expansion schema (Figures 9 & 10). For those patients, we will allow administration 
of CAR-20/19-T cells but these patients must be replaced on the study for the purposes of our 
Phase 1/1b study. Despite not achieving the desired dose level there are still potential toxicities 
associated with CAR-20/19-T cell administration and so these patients will undergo identical 
study procedures and will be followed for DLTs, AEs, and survival as all other patients. For this 
cohort of patients who receive a non-specified dose, if there is ≥33% rate of DLT as defined in 
section 3.4, then future patients who fail to make the desired dose level will no longer be 
allowed to be treated in this manner. Standard operating procedures for patients treated at a 
non-specified cell dose are listed in Appendix H.  
 
 
 
 

 
 
 
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Figure 9: Dose Escalation Phase 

 
 
 
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6.5 Dose Expansion Phase  
 
 
 
 
 
 
Once a dose has been identified in our Dose Escalation phase we will perform a Dose 
Expansion at that dose level (termed Dose Level X). We will evaluate an additional 6 patients in 
NHL/CLL at this Dose Level X (Figure 10). In this dose expansion phase is there are ≥2 
patients with DLTs, no further patients will be allowed enrollment.  
The dose expansion phase will commence after identification of a dose level in the dose 
escalation cohort and after approval from both internal and external monitoring agencies 
(DSMB/IRB/FDA). 
6.6 Phase 1b (Single Dose of CAR-T cell) 
Once the final patient on the Phase 1 protocol has enrolled on the trial, we will initiate 
enrollment on the Phase 1b portion to test the safety of single dose administration compared to 
fractionated dosing. This phase will include a 9 patient expansion at 2.5 x 106 cells/kg CAR-T 
cells administered in a single dose (rather than split dose). The first two patients at this dose 
level will be admitted to the hospital and monitored for 14-days prior to subsequent infusion to 
ensure there are not unexpected toxicities associated with single-dosing. If there are no DLTs in 
these first two patients, then additional patients can receive cells either outpatient or inpatient 
and be treated as indicated.  This cohort will close if there is ≥3 DLTs among the nine treated 
patients. 
6.7 Treatment/Dose Limiting Toxicity 
Management and dose modifications associated with the AEs are outlined in Section 7: Dosing 
Delays/Dose Modifications.  
Dose-limiting toxicity (DLT) will be defined as a grade 3 or 4 non-hematologic toxicity possibly, 
probably, or definitely related to the infusion of CAR-20/19-T cells. See section 3.4 for a 
detailed definition. The dose limiting toxicity will be based on the tolerability observed during 
the first 28 days of treatment/observation. If multiple toxicities are seen, the presence of dose 
limiting toxicity should be based on the most severe toxicity experienced.  
 
Grade 3 or 4 CRS will be treated initially with IL-6 inhibition with tocilizumab and/or 
corticosteroids (Appendix F). Additional agents that can be used for refractory CRS include 
siltuximab or other immunosuppression at the discretion of the PI. A detailed algorithm for the 
management of CRS and neurotoxicity related to CAR-T cells for this study is provided in 
Appendix F. Supportive care measures to consider for patients receiving CAR-T cells are 
provided in Appendix G. 
 
As cytokine release syndrome is a new entity observed with novel cellular therapies, a modified 
grading system has been proposed for more appropriate assessment of dose limiting toxicities. 
Figure 10: Dose Expansion Phase 
Dose Level X—Selected from 
Dose Escalation Phase 
6 patient expansion cohort 
in NHL and CLL patients at 
Dose Level X 

 
 
 
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[1, 60] We will employ this grading system to accurately grade toxicity of our CAR-20/19-T cells 
and a copy of the grading system can be found in Appendix F.  
 
6.8 Usage of Concurrent/Concomitant Medications 
The investigational agent in this protocol is autologous CAR-20/19-T cells which as a living 
product will not undergo metabolism as would a pharmaceutical therapy. As such, 
concurrent/concomitant medications can be prescribed at the discretion of the treating physician 
for treatment of symptomatic conditions such as fever, nausea/vomiting, diarrhea, anxiety, etc. 
Antimicrobial prophylaxis can be started as per treating physician. This can include ciprofloxacin 
or cefepime (or other appropriate antibiotic if allergic) for antibacterial prophylaxis, acyclovir or 
alternative agent for antiviral prophylaxis, fluconazole (or alternative agent as indicated) for 
fungal prophylaxis, bactrim (or other PCP prophylaxis if sulfa allergic). Prohibited and limited 
therapies are detailed in the following section 6.9.   
 
6.9 Dietary Restrictions 
On the inpatient service, patients will follow the FDA’s Food Safety for Cancer Patient 
Guidelines. Otherwise no specific dietary recommendations are indicated for this study.  
 
6.10 Prohibited Medications 
Patients should be off all systemic steroids, oral or IV, (not including inhaled or intranasal 
steroids) at the time of apheresis (for at least 7 days prior) and again starting Day 0 after 
administration of the CAR-20/19-T cells. Exception to this will be steroids for replacement 
dosing for adrenally insufficient patients or steroids for the purposes of the treatment of CRS at 
the discretion of the investigators listed on this protocol.  
With regards to growth factors, long-acting products such as Neulasta (pegfilgrastrim) will not be 
permitted starting Day -7 to Day +28. Given the potential risk of exacerbation of CRS or 
triggering of CRS, short-acting growth factors such as filgrastrim (neupogen) or sargramostim 
(leukine) will be limited to patients with persistent neutropenia  starting Day+21 provided that 
there are no active signs or symptoms consistent with CAR-T cell associated CRS or 
neurotoxicity.  
6.11 Monitoring Subject Compliance  
CAR-20/19-T cells will be administered only to eligible patients under the supervision of the 
investigator or identified sub-investigator(s). The appropriate study personnel will maintain 
records of study drug receipt and dispensing.  The study agent will be given on the inpatient 
oncology units. There will be no self-administered agents as part of the investigational portion of 
this protocol.  
6.12 Re-treatment 
Patients who have received treatment on the CAR-20/19-T cell protocol may be retreated if they 
meet the same inclusion/exclusion criteria as listed in the clinical protocol and meet the 
following requirements:  
1. Patient must be ≥100 days from first dose of CAR-20/19-T cells but ≤24 months from 
initial treatment.  

 
 
 
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2. Have <1% detectable CAR-T cells by flow cytometry in the peripheral blood 
3. Had demonstrated clinical benefit from the first dose (either PR or CR at Day 28 
evaluation).  
4. Have a repeat biopsy that confirms either CD19 or CD20 positive disease post-CAR-T 
cell relapse.  
5. Have not received another genetically modified CAR-T cell product either commercially 
or as part of a separate clinical trial.  
 
Patients who are re-treated will not count towards our primary safety endpoint but AEs and 
testing will be monitored/performed in an identical fashion as their first infusion (see schedule of 
events). Research studies including t-cell repertoire studies, Paxgene RNA analysis, tryptophan 
metabolites, and QOL studies will not be repeated. Cells will be given as a single infusion on 
Day 0 and the lymphodepletion regimen outlined in this protocol will be utilized. Patients will not 
undergo repeat apheresis and CAR-T cell production, rather, they will receive remainder CAR-T 
cells that are cryopreserved from their initial production (see section 9.1.5 on stability data of 
CAR-T cells post cryopreservation). Patients will receive no more than the goal dose of 2.5 x 
10^6 cells/kg (max 80 kg). If the patient does not have that dose, they will receive the remainder 
of the CAR-T cells left.  
To ensure safety of re-treatment, DLTs through Day 28 will be monitored specific to this cohort. 
At first, a maximum of 3 patients will be allowed to be treated in this manner with concurrent 
enrollment allowed. If there is ≤1 DLT within the first three patients, further patients will be 
allowed re-treatment with CAR-20/19-T cells as clinical indicated and if they have met above 
criteria. After enrollment of the 4th patient or beyond in this cohort, if the DLT rate exceeds 
≥33%, no further patients will be allowed retreatment.  
All patients will be followed for AEs, survival, response rates as per Table 1.  
7 
DOSING DELAYS/DOSE MODIFICATIONS 
7.1 Dosing Administration/Delays 
Patients will receive the modified CAR-20/19-T cells in one fixed dose on Day 0 or two doses 
given 24 hours apart on Day 0 and Day +1 of this protocol. Patients will be either admitted to the 
oncology unit for this infusion or seen daily in the outpatient BMT Day hospital suite and will be 
under the care of a hematology/oncology attending. Criteria for infusion of cellular therapy 
include the following: 
1) No new medical complication that would unduly increase risk of infusion 
2) No requirement of supplemental oxygen >2L by nasal cannula to keep saturation >95% 
3) No active cardiac arrhythmias not controlled with medical management 
4) No hypotension requiring pressor support 
5) Study treatment administration will not be delayed as a result of neutropenia, 
thrombocytopenia, or anemia, which are all potential risks of lymphodepleting 
chemotherapy 
6) No uncontrolled active infectious complications 
7) No fever>101o F in last 24 hours prior to infusion 
 

 
 
 
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Patients who do not meet these criteria will have the first dose of their cells delayed for up to a 
maximum of 7 days beyond their goal treatment date (Day 0). If there is not resolution of those 
issues, the patients will be taken off protocol.  
 
For patients who received split dose cells, the second dose of cells given on Day 1, patients 
must meet the above same criteria. In addition, patients who develop clinically significant 
toxicities within 24 hours after their first infusion will not have their subsequent infusion given. 
These include grade 2 or higher infusion reactions, allergic responses, cytokine release 
syndrome, cardiac or pulmonary toxicities, or others deemed clinically significant by the study 
investigators. Transient grade 1 chills, nausea, vomiting and other mild expected infusion-
related toxicities that respond to usual supportive care measures will not require holding 
subsequent doses. Patients with fever≥101o F after the first cell dose felt to be related to CRS 
will have the second dose held for up to 7 days until there is resolution. Any questions about 
subsequent dosing should be discussed with the Principal Investigator. The second dose can be 
delayed up to maximum of 7 days after the initial dose as per the discretion of the PI. 
Dose Modifications and Dosing Delay Table for Specific Adverse Events 
Table 4: Potential Dose Modifications for Dose #2 in setting of Cytokine Release Syndrome 
Grade of Event 
Management 
 
≤ Grade 1 
No change in planned 2nd infusion 
 
   Grade 2* 
Hold 2nd infusion until ≤ Grade 1 
Resume at same dose level 
 
 Grade 3 
Hold 2nd infusion 
 
 Grade 4 
Hold 2nd infusion 
 
Management:  CRS management (Appendix F) 
*Patients requiring a delay of > 7 days will not receive the 2nd dose of modified CAR-20/19-T cells 
 
7.2 Monitoring and Toxicity Management 
Each patient receiving CAR-20/19-T cells will be evaluable for safety. The safety parameters 
include all laboratory tests and hematological abnormalities, physical findings, and spontaneous 
reports of adverse events reported to the investigator by patients. Each patient will be assessed 
periodically for any toxicity development as per the schedule of procedures (Table 1). Toxicity 
will be assessed per the NCI CTCAE v5.0 except for CRS which will be graded as previously 
defined.  
For expected cytopenias patients will be transfused platelets and red blood cells as per 
Appendix G.  
We will monitor specifically for the development of CRS, a known complication that can occur 
with the treatment of CAR modified T cells. Signs/symptoms related to CRS were listed in 
section 1.7 (Figure 8). Grading of CRS will be by the revised recommended grading system for 
CRS found in Appendix F. Any patient who develops fever as an outpatient will be admitted for 
work-up and evaluation of CRS. Detailed algorithm for CAR-T cell associated CRS or 
neurotoxicity based on current recommendations can be found in Appendix F. The current 
recommendations for CRS management include Tocilizumab and/or steroids as first-line 
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for the treatment of several rheumatological conditions. Tocilizumab prevents IL-6 binding to 
both cell associated and soluble IL-6Rs inhibiting signaling and has led to clinical improvement 
in patients with life-threatening CRS within 24 hours of administration.[1, 53] For patients failing 
to respond after two doses of tocilizumab, steroids will be second line for CRS. Further 
management beyond second line therapy will depend upon the judgment of the clinician and 
may include additional immunosuppression such as Siltuximab or other agents that can inhibit T 
cell proliferation/function.  
7.3 Other Toxicities 
Other toxicities include those associated with lymphodepleting chemotherapy regimen which 
includes fludarabine with cyclophosphamide. 
 
Hematologic Toxicities 
Grade 3 or 4 anemia, neutropenia, and thrombocytopenia are 
potential toxicities associated with lymphodepleting chemotherapy. 
Infection 
Neutropenic fever is a potential toxicity associated cytotoxic 
chemotherapy.  
Tumor Lysis Syndrome 
Can occur as a result of tumor cell death with treatment with 
cytotoxic chemotherapy.  
Organ toxicity 
Can be pulmonary, renal, liver, or cardiac toxicity related to 
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8 
ADVERSE EVENTS: DEFINITIONS AND REPORTING 
REQUIREMENTS 
8.1 Definitions 
Adverse Event (AE) and Serious Adverse Events (SAE) 
The investigator and his or her team will follow the Medical College of Wisconsin policies related 
to adverse event reporting. This information may be found on the Human Research Protection 
Program website. (http://www.mcw.edu/hrpp/InvestigatorsandStudyStaff.htm) 
Serious AE (SAE) means any untoward medical occurrence that at any dose: 
• 
Death. Results in death. 
• 
Life threatening. Is life threatening (refers to an AE 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). 
• 
Hospitalization. Requires inpatient hospitalization or prolongation of an existing 
hospitalization (see clarification in the paragraph below on planned hospitalizations). 
• 
Disability/incapacity. Results in persistent or significant disability or incapacity. 
(Disability is defined as a substantial disruption of a person’s ability to conduct normal 
life functions). 
• 
Medically important event. This refers to an AE that may not result in death, be 
immediately life threatening, or require hospitalization, but may be considered serious 
when, based on appropriate medical judgment, may jeopardize the patient, require 
medical or surgical intervention to prevent 1 of the outcomes listed above, or involves 
suspected transmission via a medicinal product of an infectious agent.  Examples of 
such medical events include allergic bronchospasm requiring intensive treatment in an 
emergency room or at home, blood dyscrasias or convulsions that do not result in 
inpatient hospitalization; any organism, virus, or infectious particle (e.g., prion protein 
transmitting Transmissible Spongiform Encephalopathy), pathogenic or nonpathogenic, 
is considered an infectious agent.  
Clarification should be made between a serious AE (SAE) and an AE that is considered severe 
in intensity (Grade 3 or 4), because the terms serious and severe are NOT synonymous. The 
general term severe is often used to describe the intensity (severity) of a specific event; the 
event itself, however, may be of relatively minor medical significance (such as a Grade 3 
headache). This is NOT the same as serious, which is based on patient/event outcome or action 
criteria described above, and is usually associated with events that pose a threat to a patient’s 
life or ability to function. A severe AE (Grade 3 or 4) does not necessarily need to be considered 
serious. Seriousness (not intensity) serves as a guide for defining regulatory reporting 
obligations. Planned hospitalizations (e.g. admission for CAR-T infusion) will not be considered 
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8.2 Unanticipated Problem Involving Risk to Subject or Other (UPIRSO) 
The investigator and his or her team will follow the Medical College of Wisconsin policies related 
to unanticipated problems involving risks to subjects or others. This information may be found 
on the Human Research Protection Program website 
(http://www.mcw.edu/hrpp/InvestigatorsandStudyStaff.htm) 
8.3 AE Attribution and Grading 
Adverse Event Grading  
Will be performed using the NCI CTCAE Version 5.0 except for CRS which will be graded as 
described in Appendix F per clinical guidelines.  
Adverse Event Attribution 
Attribution is an assessment of the relationship between the AE and the medical intervention. 
Relationship Assessment:  In-Depth Definitions 
For all collected AEs, the clinician who examines and evaluates the subject will determine the 
adverse event’s causality based on temporal relationship and his/her clinical judgment. The 
degree of certainty about causality will be graded using the categories below: 
Definitely Related: There is clear evidence to suggest a causal relationship, and other possible 
contributing factors can be ruled out. The clinical event, including an abnormal laboratory test 
result, occurs in a plausible time relationship to CAR-20/19-T cell administration and cannot be 
explained by concurrent disease or other drugs or chemicals. The response to withdrawal of the 
drug (dechallenge) should be clinically plausible. The event must be pharmacologically or 
phenomenologically definitive, with use of a satisfactory rechallenge procedure if necessary.  
Probably Related: There is evidence to suggest a causal relationship, and the influence of 
other factors is unlikely. The clinical event, including an abnormal laboratory test result, occurs 
within a reasonable time sequence to CAR-20/19-T cell administration, is unlikely to be 
attributed to concurrent disease or other drugs or chemicals, and follows a clinically reasonable 
response on withdrawal (dechallenge). Rechallenge information is not required to fulfill this 
definition.  
Possibly Related: There is some evidence to suggest a causal relationship (e.g., the event 
occurred within a reasonable time after administration of the trial medication). However, the 
influence of other factors may have contributed to the event (e.g., the subject’s clinical condition, 
other concomitant events). Although an adverse event from CAR-20/19-T cell infusion may rate 
only as “possibly related” soon after discovery, it can be flagged as requiring more information 
and later be upgraded to “probably related” or “definitely related”, as appropriate. 
Unlikely: A clinical event, including an abnormal laboratory test result, whose temporal 
relationship to CAR-20/19-T cell administration makes a causal relationship improbable (e.g., 
the event did not occur within a reasonable time after administration of the trial medication) and 
in which other drugs or chemicals or underlying disease provides plausible explanations (e.g., 
the subject’s clinical condition, other concomitant treatments).  

 
 
 
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Unrelated: The AE is completely independent of CAR-20/19-T cell administration, and/or 
evidence exists that the event is definitely related to another etiology. There must be an 
alternative, definitive etiology documented by the clinician.  
Known AEs List 
Potential risks associated with this treatment include symptoms/complications associated with 
CRS and neurotoxicity. Section 1.7 lists the known symptoms/complications associated with 
CRS and other known risks of CAR-T cell administration.  
Time Period for AE Capture 
AE collection schedule detailed in Table 1: Schedule of Events 
8.4 Monitoring and Recording an Adverse Event  
Definition. Any clinically relevant deterioration in laboratory assessments or other clinical 
finding is considered an AE.  
Reporting source. AEs may be spontaneously reported by the patient and/or in response to an 
open question from study personnel or revealed by observation, physical examination or other 
diagnostic procedures.  
Prior to the trial. Planned hospital admissions or surgical procedures for an illness or disease 
that existed before the patient was enrolled in the trial are not to be considered AEs unless the 
condition deteriorated in an unexpected manner during the trial (e.g., surgery was performed 
earlier or later than planned).  
Pretreatment events following signed informed consent. For serious pretreatment events, 
the investigator must determine both the intensity of the event and the relationship of the event 
to study procedures.  
Treatment events. For serious AEs, the investigator must determine both the intensity of the 
event and the relationship of the event to study drug administration.  
Not serious AEs. For non-serious AEs, the investigator must determine both the intensity of the 
event and the relationship of the event to study drug administration. 
Follow-up of Adverse Events 
All adverse events will be followed with appropriate medical management 30 days following the 
last dose of the study drug or treatment or until they are resolved, if they are related to the study 
treatment. 
8.5 Procedure for Reporting Drug Exposure during Pregnancy and Birth Events 
If a woman becomes pregnant, or suspects that she is pregnant, while participating in this study, 
she must inform the investigator immediately. The sponsor-investigator must notify the DSMC 
and IRB. The pregnancy must be followed for the final pregnancy outcome. As this study utilizes 
an investigational agent under IND with the FDA, this will be reported to the FDA.  

 
 
 
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If a female partner of a male patient becomes pregnant during the male patient’s participation in 
this study, the sponsor-investigator must also immediately notify the DSMC by email. Every 
effort should be made to follow the pregnancy for the final pregnancy outcome. 
8.6 Subject Complaints 
If a complaint is received by anyone on the study staff, it will be discussed with the study staff 
and will be addressed on a case-by-case basis. The PI will be notified of any complaints. 
Complaints will be reported to the IRB if indicated.  
If the subject has questions about his or her rights as a study subject, wants to report any 
problems or complaints, obtain information about the study or offer input, the subject can call 
the Medical College of Wisconsin/Froedtert Hospital research subject advocate at 414-955-
8844. This information is provided to the subject in their consent.  
A product complaint is a verbal, written or electronic expression that implies dissatisfaction 
regarding the identity, strength, purity, quality or stability of a drug product. Individuals who 
identify a potential product complaint situation should immediately contact the sponsor and 
report the event. Whenever possible, the associated product should be maintained in 
accordance with the label instructions pending further guidance from a sponsor representative. 
Product complaints in and of themselves are not Reportable Events. If a product complaint 
results in an SAE, an SAE form should be completed.  
8.7 Routine Reporting Procedures for AEs 
Expedited Reporting Procedures for SAEs, UPIRSOs and DLTs. 
 
Since this is an investigator-initiated study, the principal investigator, also referred to as the 
sponsor-investigator, is responsible for reporting serious adverse events (SAEs) to any 
regulatory agency and to the sponsor- investigator’s IRB. Regardless of expectedness or 
causality, all SAEs (including serious pretreatment events) must also be reported to the DSMC 
as soon as possible, but no later than five calendar days of the sponsor-investigator’s 
observation or awareness of the event.  
 
Signs or symptoms reported as adverse events will be graded and recorded by the investigator, 
according to the CTCAE. When possible, signs and symptoms indicating a common underlying 
pathology should be noted as one comprehensive event.  
 
The investigator will assess all adverse events and determine reporting requirements to the 
MCWCC Data and Safety Monitoring Committee (DSMC) and MCW’s Institutional Review 
Board, and, when the study is conducted under an Investigational New Drug Application (IND), 
to the Food and Drug Administration (FDA), if it meets the FDA reporting criteria. The 
investigator will report SAEs to any regulatory agency and to the sponsor- investigator’s IRB. 
 
All adverse events, whether or not unexpected, and whether or not considered to be associated 
with the use of the study drug, will be entered into OnCore®. 
 
Reporting to the Data and Safety Monitoring Committee 
Regardless of expectedness or causality, all SAEs (including serious pretreatment events) must 
also be reported to the DSMC as soon as possible, but no later than five calendar days of the 
sponsor-investigator’s observation or awareness of the event.  

 
 
 
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Report Method:  The investigator will use email to report SAEs to the DSMC. The SAE report 
must include event term(s), serious criteria, and the sponsor-investigator’s or sub-investigator’s 
determination of both the intensity of the event(s) and the relationship of the event(s) to study 
drug administration. Intensity for each SAE, including any lab abnormalities, will be determined 
by using the NCI CTCAE as a guideline whenever possible.  
The criteria are available online at http://ctep.cancer.gov/reporting/ctc.html. 
Reporting to MCW Committee Institutional Review Board 
The principal investigator must report unanticipated problems involving risks to subjects or 
others (UPIRSOs) to the MCW IRB within five business days of his/her awareness of the event.  
[Guidance on Adverse Event Reporting to the IRB is available online at MCW IRB Policies and 
Procedures.] (http://www.mcw.edu/hrpp/policiesprocedures.htm) 
Expedited Reporting to the Food and Drug Administration 
As this study is being conducted under an IND, the sponsor-investigator is responsible for 
determining whether the suspected adverse reaction meets the criteria for expedited reporting in 
accordance with Federal Regulations (21 CFR §312.32).  
The investigator will report in an IND safety report any suspected adverse reaction that is both 
serious and unexpected. In addition, per FDA recommendations, the sponsor-investigator will 
report all CRS ≥Grade 3 and Neurotoxicity ≥Grade 3 that occurs as a result of CAR-20/19-T cell 
infusion. For other events, the sponsor-investigator needs to ensure that the event meets all 
three definitions: suspected adverse reaction, unexpected, and serious  
 
If the adverse event does not meet one of the above definitions, it should not be submitted as 
an expedited IND safety report. The timeline for submitting an IND safety report to FDA is no 
later than 15 calendar days after the investigator determines that the suspected adverse 
reaction qualifies for reporting (21 CFR 312.32(c)(1)).  
 
Any unexpected fatal or life-threatening suspected adverse reaction will be reported to FDA no 
later than seven calendar days after the Investigator’s initial receipt of the information (21 CFR 
312.32(c)(2)).  
Any relevant additional information that pertains to a previously submitted IND safety report will 
be submitted to FDA as a Follow-up IND Safety Report without delay, as soon as the 
information is available (21 CFR 312.32(d)(2)).  
Suggested Reporting Form: 
 
US FDA MedWatch 3500A:  
http://www.fda.gov/Safety/MedWatch/HowToReport/DownloadForms/default.htm 
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9 PHARMACEUTICAL INFORMATION 
 
9.1.1 CAR-20/19-T cells 
 
Product Description 
Each CAR-20/19-T cell line is a unique product prepared from patient peripheral blood CD4+ 
and CD8+ T cells. The cells have been engineered to express two different extracellular single 
chain antibodies (scFv) recognizing CD20 and CD19. The antibodies are linked to tandem 
signaling domains comprised of the CD3ζ signaling module linked to the 4-1BB costimulatory 
domain (Figure 3). The CAR-20/19-T cells if given fresh without cryopreservation will be in an 
infusion medium of an isotonic electrolyte solution (Plasma-Lyte® A or Normosol® R) containing 
2.5% HSA. If needed, the CAR-T cells will be cryopreserved in infusible grade cryomedium at 
two different cell numbers.  For cryopreserved cells the infusion medium will consist of 10% di-
methylsulfoxide, 4% human serum albumin, in 85% Normosol R. 
Mechanism of Action: 
When the antibody chains on the CAR-20/19-T cells bind to their ligands on the surface of B 
cells the T cells are triggered to kill the cells to which they bind.  A given T cell can bind and kill 
multiple target cells. 
Metabolism: 
The CAR-20/19-T cells will be activated by ligand engagement and are expected to expand in 
vivo.  Transduced cells are expected to persist for a period of months to years. 
Contraindications: 
Please see section 7.1 for contraindications to infusion  
Side Effects:   
Side effects are thoroughly detailed in section 1.7. Briefly major toxicities include: 
• 
B cell ablation- Since CD19 and CD20 are also expressed on healthy B cells, the CAR-
20/19-T cells are expected to eliminate these cells from the circulation as long as the 
cells persist.  The loss of B cells could result in an increased risk of infection. 
• 
Allergic Reaction- The scFv on the T cell surface may invoke an immune reaction that 
might eliminate the CAR-20/19-T cells and secondarily cause an autoimmune response, 
though such responses have not been reported in clinical trials to date. 
• 
Transformation- The genetic modification of the CAR-20/19-T cells could result in 
transformation. To date transformations of CAR-T cells have not been reported 
• 
Tumor lysis syndrome (TLS) - Patients with a high tumor burden may experience a 
reaction due to lysis of the tumor cells either from the lymphodepleting preparative 
regimen or from the CAR-20/19-T cells. Such reactions have been reported and can be 
severe. With careful monitoring, appropriate therapies are available that can mitigate this 
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• 
Cytokine Release Syndrome (CRS)- In vivo expansion of the infused CAR-20/19-T cells 
may be associated with a release of inflammatory cytokines. Such responses have been 
reported, some have been severe. Close monitoring and appropriate interventions are 
available that can mitigate this reaction. 
• 
Neurotoxicity- CAR-T therapy has led to some patients having unpredictable 
neurotoxicity in the form of seizures, aphasias, and in rare cases life threatening cerebral 
edema. Clinical vigilance and early intervention can help prevent and/or treat this 
complication. 
• 
Unknown- Given this first-in-human use of CAR-20/19-T cells unknown side effects may 
occur either associated with the infusion or long term.  Monitoring for such effects is 
described in this protocol 
9.1.2 Solution Preparation  
The CAR-20/19-T cells will be administered either fresh following harvest or within 6 weeks of 
cryopreservation in the appropriate doses for infusion. Cryopreserved cells will be thawed within 
the LPL prior to administration. The CAR-T cells will be transported at 1-10oC to the infusion 
site. If the dose level changes from the time of product cryopreservation to time of infusion, the 
volume of product infused will be adjusted accordingly. 
 
9.1.3 Investigational Agent Administration  
The cells will be administered from 1-7 days after completion of chemotherapy to deplete 
lymphocytes.  The patient will be premedicated.  Vital signs will be monitored 15 minutes prior to 
infusion and every 15 minutes after infusion for the first hour. Monitoring will be continued until 
all signs are stable.  The cells will be administered over a period of 5-15 minutes by IV injection.  
See Section 6 for further details. 
9.1.4 Storage Requirements 
For patients requiring cryopreservation, the cells will be stored cryopreserved in the LPL until 
the time of infusion.  Cryopreservation storage is in vapor phase liquid nitrogen at temperatures 
<-150oC in devices that are continuously monitored for temperature according to standard 
laboratory practices.  
Fresh cells will be stored at 1-10oC in a monitored refrigerator and will be given within 48 hours 
of harvest or cryopreserved accordingly.   
9.1.5 Stability 
Fresh cells have been demonstrated to be stable in infusion medium up to 48 hours after 
harvest when stored at a concentration <20 x 106 per mL at 1-10oC. Longer term phenotypic 
and functional stability data up to 1 year is shown in the below table and Figure 11. Based on 
the 1-year data, the product is likely stable indefinitely while in the cryopreserved state. Once 
thawed, the cell infusion must start within 45 minutes. 
 
 

 
 
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Phenotypic Stability of CAR-20/19-T Cell Products 
 
 
Figure 11: Phenotypic and Functional Stability of CAR-20/19-T Cell Products. CAR-20/19-
T cell products generated from Subjects 002-004 in the Phase I trial were cryopreserved, 
thawed at the indicated time points to assess (A) Protein L positivity to assess surface CAR 
expression, (B) cytotoxicity versus CD19+ target cells, and (C) cytotoxicity versus CD20+ target 
cells. Cell cytotoxicity was conducted at an effector:target (E:T) cell ratio of 50:1 (note: the E:T 
ratio was not adjusted based on the percentage of CAR-T cell present in the product; it was 
based on total T cells). 
9.1.6 Route of Administration  
The CAR-20/19-T cells will be administered intravenously. 

 
 
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9.1.7 Nursing Implications 
Premedication and patient monitoring before, during, and after each split dose infusion is 
required. See Section 6 for details 
9.1.8 Handling 
The required dose of CAR-20/19-T cells are transported at 1-10oC. The cells are infused at 
room temperature over a 5-15-minute period by IV injection. For cryopreserved cells, infusion 
should start no longer than 45 minutes after thawing. 
 
9.1.9 Availability 
Each lot of CAR-20/19-T cells is patient-specific. The cell lines are produced on site by the 
MCW cell processing laboratories.   
9.1.10 Agent Ordering 
CAR-20/19-T cells are produced at MCW and are a patient-specific material. A physician order 
for infusion specifying the dose of transduced cells per kg to be infused is required. 
9.1.11 Agent Accountability 
CAR-20/19-T cells are identified by a donor identification number (DIN) that is used to track the 
product at collection to the product at disposition (infused or discarded) and to trace the product 
at infusion to the donation.  CAR-20/19-T cells are for autologous use only and are labeled 
accordingly. 
9.1.12 Agent Destruction and Return 
Once issued from the LPL, the CAR-20/19-T cell line cannot be returned for reissue.  Aliquots of 
the lines must be maintained for potential further analysis; however, the cells are for autologous 
use only and may not be released to another patient. 
9.2.1 Tocilizumab 
Product Description 
Tocilizumab is a monoclonal antibody and immunosuppressant; specifically, tocilizumab is an 
interleukin-6 (IL-6) receptor antagonist. IL-6 is a pleiotropic pro-inflammatory cytokine and is 
produced by various cells including T- and B cells, lymphocytes, monocytes and fibroblasts. 
This agent will be part of the first line treatment for patients with CRS requiring more than 
supportive measures for medical management (Appendix F for CRS management 
guidelines). 
 
Mechanism of Action 
 
Tocilizumab binds IL-6 receptors (sIL-6R and mIL-6R) and inhibits IL-6-mediated signaling 
through these receptors. IL-6 has been shown to be involved in T cell activation, induction of 
immunoglobulin secretion, initiation of hepatic acute phase protein synthesis, and stimulation of 
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Contraindications 
 
Administration of live vaccines is contraindicated during treatment with tocilizumab.  Use is 
contraindicated in patients with known hypersensitivity to tocilizumab.  
 
Drug Interactions 
 
Use of tocilizumab may cause increased metabolism of medications that are CYP450 
substrates including CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6, and CYP3A4. Caution 
should be used in medications that are CYP450 substrates including simvastatin, 
dextromethorphan and omeprazole, and medications with a narrow therapeutic index.  
 
Side Effects 
• 
Myelosuppression: Tocilizumab may lead to neutropenia and thrombocytopenia.   
• 
Hepatic enzyme and lipid parameter alterations:  Tocilizumab may lead to an elevation in 
liver enzymes and an increase in lipid parameters including: total cholesterol, 
triglycerides, LDL cholesterol, and HDL cholesterol.  
• 
Immunosuppression: Tocilizumab may lower patient’s resistance to infections.  
• 
Gastrointestinal perforation: Events have occurred in clinical trials, caution in patients at 
increased risk of developing gastrointestinal perforation 
9.2.2 Preparation 
Tocilizumab should be diluted in 100 mL of 0.9% sodium chloride. Prior to addition of 
tocilizumab, withdraw a volume of 0.9% sodium chloride from the infusion bag equal to the 
volume of tocilizumab required (total volume of final preparation should be 100 mL). Gently mix 
the infusion to avoid foaming 
 
9.2.3 Tocilizumab Administration 
Tocilizumab should be administered over 60 minutes as an intravenous infusion. The infusion 
should have a dedicated line and not be infused concomitantly with other drugs. 
 
9.2.4 Storage 
Vials must be refrigerated at 2ºC to 8ºC (36°F to 46°F).  Do not freeze.  Vials should be 
protected from light and stored in the original package until time of use. Prepared parenteral 
drug products should be inspected visually for particulate matter and discoloration prior to 
administration. Do not use the product if visibly opaque particles, discoloration, or other foreign 
particles are observed in the solution. 
 
9.2.5 Stability 
Prepared solutions may be stored at 2ºC to 8ºC (36°F to 46°F) or room temperature for up to 24 
hours. 
 

 
 
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9.2.6 Handling 
Tocilizumab will be handled following the institutional hazardous medication policy.  
 
9.2.7 Availability 
Tocilizumab is available in single dose vials (20 mg per mL) of 80 mg per 4 mL, 200 mg per 10 
mL, and 400 mg per 20 mL.  
 
9.2.8 Agent Ordering 
Tocilizumab is commercially available  
 
9.2.9 Agent Destruction and Return 
Tocilizumab will be destroyed based on institutional standards 
10 REPORTING AND DOCUMENTING RESPONSE (MEASUREMENT 
OF EFFECT) 
10.1 Evaluation of Efficacy (or Activity) 
Definitions 
Evaluable for toxicity 
- All patients will be evaluable for toxicity from the time of their first study drug treatment. 
Evaluable for objective response 
- Only those patients who have received CAR-20/19-T cells will be re-evaluated and considered 
evaluable for response. These patients will have their response classified according to the 
definitions stated below. (Note: Patients who exhibit objective disease progression prior to the 
first disease assessment at Day +28 will also be considered evaluable.) 
10.2 Response Criteria 
Response assessment will vary by disease subtype and specific criteria are listed below for 
CLL/SLL and NHL. 
10.3 Chronic Lymphocytic Leukemia Response Criteria 
Response criteria as per 2008 guidelines for diagnosis and treatment of CLL/SLL [61] 
Complete Response (CR) 
1. Peripheral blood lymphocytes (evaluated by blood and differential count) below 4.0 x 
109/L (4000/µL) 
2. Absence of significant lymphadenopathy (e.g., lymph nodes >1.5 cm in diameter) by 
physical examination and CT scan 
3. No hepatomegaly or splenomegaly by physical examination or CT imaging 
4. Absence of constitutional symptoms 
5. ANC >1500/µL, Hemoglobin >11.0 g/dL, Platelets >100,000/ µl 
6. Bone Marrow Biopsy confirmation 

 
 
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Complete response with incomplete marrow recovery (CRi) 
1. Fulfill all criteria for a CR but have persistent anemia, thrombocytopenia, or neutropenia 
felt to be unrelated to CLL but related to drug toxicity.  
 
Partial Remission (PR) 
1. A decrease in the number of blood lymphocytes by 50% or more from value before 
therapy 
2. Reduction in lymphadenopathy by CT scans as defined by the following 
a. A decrease in lymph node (LN) size by 50% or more either in the sum products 
of up to 6 lymph nodes, or in the largest diameter of the enlarged node(s) 
detected prior to therapy 
b. No increase in any LN and no new enlarged LN. In small LN (<2 cm), an increase 
of less than 25% is not considered to be significant.  
3. A reduction in the pretreatment enlargement of the spleen or liver by 50% or more as 
detected by CT scan 
4. Blood count should show one of the following results 
a. Neutrophils >1500/µl without exogenous growth factors 
b. Platelet count >100,000/µl or 50% improvement over baseline without need for 
exogenous growth factors 
c. Hemoglobin >11.0g/dL or 50% improvement from baseline without requiring RBC 
transfusions or exogenous erythropoietin.  
 
Progressive Disease 
1. Lymphadenopathy 
a. Appearance of any new lesion, such as enlarged lymph nodes (>1.5 cm), 
splenomegaly, hepatomegaly, or other organ infiltrates. 
b. An increase by 50% or more in greatest determined diameter of any previous 
site. 
2. An increase in the previously noted enlargement of the liver or spleen by 50% or more or 
the de novo appearance of hepatomegaly or splenomegaly. 
3. An increase in the number of blood lymphocytes by 50%or more with at least 5000 B 
lymphocytes per microliter. 
4. Transformation to a more aggressive histology (e.g., Richter syndrome). Whenever 
possible, this diagnosis should be established by lymph node biopsy. 
5. Occurrence of cytopenia (neutropenia, anemia, or thrombocytopenia) attributable to CLL 
 
Stable Disease 
1. Patients who have not achieved a CR or a PR, and who have not exhibited progressive 
disease, will be considered to have stable disease (which is equivalent to a 
nonresponse). 
 
 
 
 

 
 
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10.4 Non-Hodgkin’s Lymphoma Response Criteria 
2014 Lugano Classification will be utilized for response 
assessment in patients with NHL. [62] 
 
Complete Response (CR) 
PET/CT Based Response 
o Complete metabolic response: Score 1, 2, 3 with or 
without a residual mass on the 5 point Deauville Scale 
(Figure 12) 
o No evidence of FDG-avid disease in marrow 
 
CT-Based Response 
o Target nodes/nodal masses must regress to ≤1.5 cm in longest transverse diameter 
(LDi) 
o No extralymphatic sites of disease 
o Negative BM biopsy if positive at baseline 
o Regression of organ enlargement to normal 
 
Partial Response (PR) 
PET/CT Based Response 
o Score 4 or 5 on Deauville scale with reduced uptake compared with baseline and 
residual masses of any size 
CT-Based Response 
o ≥50% decrease sum of the product of the perpendicular diameters (SPD) of up to 6 
dominant measurable nodes and extranodal sites; no criteria for progressive disease are 
met 
o When a lesion is too small to measure on CT, assign 5 mm x 5 mm as the default 
value. When no longer visible, 0 x 0 mm 
o For a node >5 mm x 5 mm, but smaller than normal, use actual measurement for 
calculation 
o If enlarged, spleen must have regressed by >50% in length  
 
Stable Disease (SD) 
PET/CT Based Response 
o Score 4 or 5 on Deauville scale with no significant change in FDG uptake from the 
baseline scan 
 
CT-Based Response 
o <50% decrease from baseline SPD of up to 6 dominant measurable nodes and 
extranodal sites, no criteria for progressive disease are met.  
o No increase in non-measured lesions consistent with progression 
o No increased in organ enlargement consistent with progression.  
 
Progressive Disease (PD) 
PET/CT Based Response 
o Score 4 or 5 on Deauville scale with increase in intensity of uptake from baseline 
o New FDG-avid foci consistent with lymphoma  
o New or recurrent FDG-avid foci in bone marrow 
 
CT-Based Response 
Figure 12: 5 point Deauville Scale 

 
 
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o An individual node/lesion must be abnormal with longest transverse diameter >1.5 cm 
and have an increase by ≥50% from cross product of the LDi and perpendicular 
diameter and have an increase in the LDi or shortest axis perpendicular (SDi) from nadir 
as follows: 
o 0.5 cm for lesions ≤2 cm 
o 1.0 cm for Lesions >2 cm 
o In the setting of splenomegaly, the splenic length must increase by >50% of the extent of 
its prior increase beyond baseline (eg: a 15-cm spleen must increase to >16 cm). If no 
prior splenomegaly, must increase by at least 2 cm from baseline 
o New or recurrent splenomegaly 
o New or clear progression of pre-existing non-measured lesions 
o Re-growth of previously resolved lesions 
o A new node >1.5 cm in any axis 
o A new extra nodal site > 1.0 cm in any axis; if <1.0 cm in any axis is presence must be 
unequivocal and must be attributable to lymphoma 
o Assessable disease of any size unequivocally attributable to lymphoma 
 
10.5 Response Definitions 
Evaluation of Best Overall Response 
The best overall response is the best response recorded from the start of the treatment until 
disease progression/recurrence (taking as reference for progressive disease the smallest 
measurements recorded since the treatment started). The patient's best response assignment 
will depend on the achievement of both measurement and confirmation criteria. The first 
disease assessment will occur at 4 weeks’ post treatment (Day+28). Response assessment will 
continue up to 2 years or until progression/death.  
 
Duration of Response 
 
Duration of overall response 
The overall response duration is measured from the time measurement criteria are met for CR 
or PR (whichever is first recorded) until the first date that recurrent or progressive disease is 
objectively documented (taking as reference for progressive disease the smallest 
measurements recorded since the treatment started). 
 
The overall CR duration is measured from the time measurement criteria are first met for CR 
until the first date that recurrent disease is objectively documented. 
 
Duration of stable disease 
Stable disease is measured from the start of the treatment until the criteria progression are met, 
taking as reference the smallest measurements recorded since the treatment started. 
 
Progression-Free Survival 
Progression-free survival (PFS) is defined as the time duration from treatment start to 
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10.6 Evaluation of Safety 
Analyses will be performed for all patients having received at least one dose of study drug. The 
study will use the CTCAE v5.0 for reporting of non-hematologic adverse events and modified 
criteria for hematologic adverse events.  
11 CORRELATIVE STUDIES/SPECIAL STUDIES 
11.1 T Cell Diversity Analysis During CAR-T Cell Therapy 
Diversity of the T cell repertoire (as measured by T cell receptor sequencing) has been 
associated with effective T cell immunity. Conversely, low levels of T cell diversity have been 
correlated with aging, infection, and the development of cancer. The relation between the 
diversity of the T cells used for CAR-T therapy and outcomes are not understood. Neither is the 
effect of generating CAR-T on repertoire diversity understood. Both could be important 
parameters in therapy outcomes. 
 
We will measure the diversity of the circulating T cell repertoire at three time points during the 
study to examine: 1) the effect of generating the CAR-T cells and 2) any possible effect of 
infusion of the CAR-T product on the overall circulating repertoire of subject. The initial sample 
of 107 mononuclear cells will be obtained at time of apheresis and the T cell repertoire diversity 
will act as the benchmark for the two comparative analyses. The second sample will consist of 
the same number of cells taken from the final CAR-T product prior to infusion into the subject. 
The final sample will consist of 10ml of blood and will be obtained as part of the 28-day disease 
assessment.  
 
• 
Sample 1: Will be obtained from the apheresis product collected at Day -15 
• 
Sample 2: From final CAR-T cell product prior to patient infusion 
• 
Sample 3: Peripheral blood sample (10 ml) on Day +7  
• 
Sample 4: Peripheral blood sample (10 ml) on Day +28 (+/- 3 days) 
 
The samples will be labeled with a unique patient identifier and sent to BloodCenter of 
Wisconsin (BCW) for T cell repertoire analysis. At BCW the de-identified samples will be subject 
to cell separation into CD4 and CD8 T cells, cDNA tagged with unique barcodes will be 
prepared and subject to BV gene-specific amplification and single cell RNA sequencing 
(scRNA-Seq). The amplicons will be pooled and subject to NGS sequencing on the Illumina 
platform. The sequence data will be clustered on the basis of unique CDR3 sequence 
(clonotype), whole transcriptome and the number of unique cDNA-defining barcodes counted to 
quantify the clonotype. Diversity and complexity measures will be derived, the latter involving 
not just the number of unique species but their frequency distribution. A similar analysis will be 
performed on the basis of the amino acids encoded by the CDR3 sequence (motif analysis). As 
needed the depth of sequencing can be modulated to bring up the resolution level. 
 
The repertoire diversity analyses and scRNA-Seq will describe the initial diversity of the 
repertoire and show whether the CliniMACS Prodigy-derived CAR-T cells have undergone 
major shifts in the T cell population being infused into the patient. Both results will be most 
informative when compared to other clinical parameters and outcomes. Comparison of the initial 
circulating repertoire diversity with that of the circulating repertoire after infusion of the CAR-T 
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states due to the functioning of the CAR-T cells, and comparison of the data with clinical 
outcomes should be informative. The risk associated with these analyses is minimal. The lab at 
BCW will be working with de-identified samples. The investigators at BCW are experts in this 
area of analysis. The information gained will be useful for the overall interpretation of outcomes.      
 
11.2: Circulating Tryptophan Metabolites and Gene Expression Patterns and risk for CNS 
toxicity during CAR-T cell Therapy 
Cytokine release syndrome (CRS) is a systemic inflammatory reaction that is observed in a high 
proportion of patients receiving CAR-T-based therapies.  CRS is driven by an increase in the 
release of proinflammatory cytokines that are released after the activation of monocytes, 
macrophages and some lymphocyte populations.  In addition to CRS, and perhaps as a 
consequence, patients receiving CAR-T therapies also experience CNS toxicity, including 
tremor, seizures and life-threatening cerebral edema.  Although the pathophysiology of these 
neurological effects is not clear, inflammatory cytokines are thought to be involved. 
 
Human studies, primarily carried out in patients with major depressive illness, indicate that the 
kynurenine pathway is a critical link between inflammation and damage to the brain.  The 
kynurenine pathway is activated by inflammatory cytokines (predominantly interferon gamma) 
released in the brain by activated T cells and leads to the production of kynurenine from 
tryptophan as a result of up-regulation of the enzyme indoleamine 2,3-dioxygenase (IDO) by 
microglia or brain-resident macrophages.  Kynurenine is further metabolized to neurotoxic 
metabolites 3-hydroxykynurenine (3-HK) and quinolinic acid (QA) and to the neuroprotective 
metabolite, kynurenic acid (KA).  The ratios of serum KA/3-HK and/or KA/QA have been used 
previously as indices of the role of this inflammatory pathway on brain function.  Low ratios are 
seen in patients with mood disorders, for example, compared to controls. Further, we have 
identified a gene expression profile of 53-genes termed the “conserved transcriptional response 
to adversity” (CTRA), a gene expression pattern associated with exposure to chronic stressors 
and whose largest gene subset is comprised of proinflammatory genes. This CTRA gene 
expression has been associated with worse outcomes following HCT. 
 
We hypothesize that neuroinflammation is responsible for the adverse effect of CAR-T cell 
therapy on the brain and that the serum KA/3-HK and KA/QA ratios as well as CTRA profiles will 
serve as biomarkers for neuroinflammation, and thus, as biomarkers for the risk of CNS toxicity. 
 
We will measure the serum concentrations of tryptophan, kynurenine, 3-HK, QA, KA, and gene 
expression patterns (CTRA) at 4 time points during the study to examine: (1) the trajectory in 
the concentration of each metabolite and gene expression in the month following CAR-T cell 
therapy; and (2) the correlation between the KA/3-HK and KA/QA ratios as well as gene 
expression and the occurrence of CNS toxicity. A baseline sample will be assessed on the day 
of apheresis and will serve as a benchmark for the trajectory study.  Two additional blood 
samples will be analyzed during the first month after treatment (14 and 28 days) and one 
sample will be analyzed after three months (day 90) to assess recovery.  The samples will be 
collected along with other blood samples at these time points in the study. 
 
The samples will be collected via standard venipuncture in serum separating tubes 
(metabolites) and PaxGene RNA tubes (gene expression) and will be collected using standard 
protocols and stored at -80 degrees in cryotubes labeled with unique patient identifiers until 
analysis.  The samples will be sent to the laboratory of Dr. Cecilia Hillard in the Neuroscience 
Research Center. The serum metabolite tubes will be processed for the concentrations of 5 
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chromatography with tandem mass spectrometry.  The extraction and derivatization of the 
samples will be carried out in the Hillard laboratory and the mass spectroscopy will be 
performed in the Department of Pharmacology at the Medical College of Wisconsin.  The 
PaxGene RNA tubes will be stored until they can be shipped in batch to the UCLA Social 
Genomics Core (Directed by Dr. Steve Cole, Professor, Hematology/Oncology, UCLA), who will 
conduct all gene expression profiling on obtained blood samples by using Illumina HT-12 human 
gene expression bead arrays. Total RNA will be extracted from the whole blood samples stored 
at MCW, subjected to quality assurance assays to test suitable mass (by spectroscopy) and 
integrity (by Agilent Bioanalyzer RNA Integrity Score) for analysis, and subjected to microarray 
target synthesis and hybridization in collaboration with the UCLA Neuroscience Genomics Core 
Laboratory using standard Illumina assay equipment and protocols. The output of these 
analyses is quantification of whole genome RNA production. The laboratory staff will work with 
de-identified samples and the data will be provided to the study PI for correlation with CNS 
toxicity in the same patients. 
 
The information gathered from this study will provide important mechanistic information 
regarding the relationships among the tryptophan-kynurenine pathway, proinflammatory gene 
expression, and the occurrence of CNS toxicity, and could elucidate these metabolites and gene 
expression clusters as potential biomarkers for the occurrence of CNS toxicity in patients 
undergoing CAR-T therapy. 
 
11.3: Patient-reported outcome (PRO) measures following CAR-T cell therapy 
Related to or as a consequence of CNS and its neurotoxicities, including neuroinflammation, 
patients receiving CAR-T therapies may be at risk for diminished quality of life and patient-
reported outcomes (PROs), including the development of depression, anxiety, fatigue, sleep 
disturbances, and pain. There are several mechanistic modalities by which engineered 
autologous CAR-T cells may influence and/or be affected by PRO measures. 
 
First, immune activation, inflammation, and cytokine activation - as occurs with CRS - are 
known to affect cognitive and/or emotional functioning,[63] including among cancer patients. 
[64] Second, while patient-reported symptomatology may be less salient during the initial acute 
period of potentially serious illness, as can occur with CRS [1, 53, 54], the acute and likely 
subsequent inflammation may play a key role in the development of depression and other 
psychopathologic symptoms; it is unknown what longer-term QOL sequelae may be present 
after an initial severe (and potentially ongoing) inflammatory insult. Prior research indicates that 
inflammation itself is involved in both the genesis and foreshadowing of future depressive or 
psychotic episodes.[65] 
 
Third, altered central nervous system (CNS) functioning and subsequent emotional/cognitive 
responses influence immunity and T cell function among cancer patients.[66]Further, these 
adverse responses are predictive of compromised immune reconstitution[67] and adverse 
outcomes following HCT.[68-70]Therefore, it is critical to assess whether or how CNS and 
psychological functioning will impact the efficacy of this new and likely increasingly used cancer 
treatment modality. As infection remains a significant risk of this therapy, it will be important to 
understand and optimize any factors that could contribute to increased infectious susceptibility, 
including neuroimmunologic influence. 
 
Finally, it is unknown what longer-term (1-2 years and beyond) effects such enhanced immune 
activation will have. Given the aforementioned immune/CNS effects and increasing emphasis 

 
 
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on QOL and PROs as mainstay parameters in assessing cancer treatment efficacy,[71, 72] it is 
important to ascertain late effect PROs.    
 
We will measure the following PROs (see Appendix I for questionnaires) during the ancillary 
study at the same time points as the biomarkers described in section 11.2 (at apheresis, D+14, 
D+28, D+90), as well as at the additional time points of 6 months, 1 year, and 2 years: 
 
Depression: Depression will be assessed through the General Depression subscale of 
the Inventory of Depression and Anxiety Symptoms (IDAS) and includes 20 questions 
with a scoring range of 20-100 (mean in community dwelling adult of 44.99 and standard 
deviation of 14.75).[73] 
 
Anxiety: Anxiety will be assessed using two subscale items of the IDAS including panic 
(health population mean = 12.58, SD = 5.26) and traumatic intrusions (healthy 
population mean = 7.60, SD = 4.20)[73] 
 
Fatigue: The Fatigue Severity Index (FSI) will be utilized to assess fatigue; a score of 3 
or greater on items assessing fatigue in the past week (average of items 1-3; FSI 
Composite) indicates clinically meaningful fatigue.[74] The FSI can also be evaluated 
using the average rating of the degree to which fatigue interfered with some general 
activities (0-10; FSI Interference); participants’ ratings of the number of days in the past 
week they felt fatigued (0-7; FSI Days); and participants’ rating of what percent of each 
day in the past week, on average, they felt fatigued (0-100; FSI Percent).[75] Individuals 
scoring at or above the cutoff also report significantly greater scores on these other 
subscales. 
 
Sleep: Sleep will be assessed using the Pittsburgh Sleep Quality Index (PSQI), with a 
score of >5 considered disturbed sleep as adjusted for cancer populations.[76, 77] 
 
Pain: The Brief Pain Inventory (BPI) assesses pain intensity as well as pain-related 
interference in function.[78] BPI Pain Severity score ranges from 0-40 (first four items), 
and the BPI Pain Interference score is a mean of the last 7 items (5a-5g) with a range of 
0-10. 
 
The information gathered from this study will provide important data regarding the impact of 
CAR-T therapy on patient QOL outcomes. 
Guidelines for reporting of Depression and/or Suicidal Ideation: 
Should the patient endorse any thoughts of suicidality or self harm per the IDAS, the study co-
investigator for this portion of the ancillary testing (Dr. Jennifer Knight) will contact them by 
phone and standard of care referrals will be made. Should completion of the study surveys 
prompt participants to want treatment for any of the other symptoms, participants will be offered 
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12 STATISTICAL CONSIDERATIONS 
12.1 Study Endpoints 
There are two primary endpoints in this study—feasibility and safety.  
 
Feasibility of CAR-20/19-T cell production by the CliniMACS Prodigy processing device (Miltenyi 
Biotec) will be measured. At minimum, we expect to reach our target CAR-20/19-T cell dose in 
at least 75% of patients with an adequate T cell starting dose. Feasibility will be measured at the 
end of the study after completion of accrual. If >25% of patients with an adequate T cell starting 
dose were unable to achieve goal CAR-20/19-T cell dose, we would deem that this technique is 
not a feasible method for production. 
Safety of administration of CAR-20/19-T cells in patients will be assessed as outlined in Section 
6 Treatment plan. 
12.2 Accrual Estimates 
 
This is a Phase 1/1b study evaluating the safety and efficacy CAR-20/19-T cells as part of a 
first-in-human study. We will utilize a 3+3 escalation design to identify the target dose level 
followed by a dose expansion at the selected dose level. The minimum number of patients 
enrolled will be 4 patients (2 at dose level 0 and 2 at dose level -1). The maximum number of 
patients enrolled would be 24, which is dependent on the number of DLTs experienced on trial.  
 
Should this treatment be safe and feasible, we would use the identified dose to open a larger 
phase II study to demonstrate the efficacy of our treatments.  
 
12.3 Interim Analyses and Stopping Rules 
 
There are no planned interim analyses as part of this Phase 1/1b Study.  
 
Toxicity will be considered unacceptable and lead to clinical hold of the study if any of the 
following occur in the first 5 patients: one treatment related mortality (TRM) or three grade 4 
non-hematological toxicity events as assessed by PI or DSMB. This ensures that there is only a 
5% probability of stopping if the actual TRM is <1%. For Grade 4 non-hematological events not 
resulting in death, this rule ensures a 5% probability of stopping if the true incidence is <20% 
and an 68% probability of stopping if the incidence is 60% or more. Additionally, if there are ≥ 2 
DLTs at any dose level, no further patients will be allowed administration at that dose level.  
 
In terms of production, if within the first 8 patients with an adequate starting T-cell dose ≥3 
patients fail to meet the targeted CAR-20/19-T cell dose, the study will be placed on a clinical 
hold to re-evaluate production methods and optimize manufacturing.  
 
12.4 Statistical Analyses Plans 
 
Phase 1 
 
The first 3 patients with NHL or CLL/SLL will be 
treated consecutively at Dose Level 0 and will 
be monitored for DLTs for a full 28 days. At 
each dose level, the first two patients treated 

 
 
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will be treated no sooner than 28 days apart to allow monitoring of acute and sub-acute toxicities. 
After that patients will be infused no sooner than at 14 day intervals among the specified dose 
level or in-between two dose levels.  
 
Dose escalation will proceed within each cohort per the following scheme: A cohort of three 
patients with NHL or CLL/SLL will receive 2.5 x 105 cells/kg (Dose Level 0). If there are no DLTs 
in the first three patients, the dose will be escalated to 7.5 x 105 cells/kg (Dose Level 1). If 1/3 
patients have a DLT at Dose Level 0, an additional three patients will be enrolled at this dose 
level. If there is ≤1/6 patients with a DLT, the dose will increase to Dose Level 1. A similar dose 
escalation schema will be used to escalate from Dose Level 1 to Dose Level 2 (2.5 x 106 
cells/kg) which is our goal dose. Dose escalations will NOT occur until all patients at the current 
level are monitored for 28 days after CAR-20/19-T cell infusion. 
 
If two of the first three or six patients with NHL or CLL/SLL develop DLT at Dose Level 0, the 
dose will de-escalate to 1.0 x 105 cells/kg and accrual will proceed per the same schema as 
above.  If two of the first three or six patients with NHL or CLL have a DLT at the lower dose, the 
study will be stopped for failure to achieve the desired safety endpoint. The minimum number of 
patients enrolled will be 4 patients (2 at dose level 0 and 2 at dose level -1) if the first four 
patients all experience a DLT. The maximum number of patients enrolled in our Dose Escalation 
phase would be 18.  
Once a safe dose has been identified, the Dose Expansion phase of the study will be open to 
accrual once approval is obtained from local and federal regulatory bodies. In this Phase, there 
will be a 6-12 patient expansion cohort in NHL and CLL/SLL. DLT monitoring will continue in the 
Dose Expansion phase. If ≥2 patients experience a DLT within this expansion cohort, the 
expansion phase will be placed on hold and will be re-opened only after discussion with the 
local DSMC and IRB.  
Phase 1b 
The Phase 1b cohort will be open to accrual once the final subject has enrolled on the Phase 1 
portion of the study. This will test the safety of single day CAR-T infusion.  
Expansion Single Infusion 
9 patients will be enrolled at the goal dose of 2.5 x 106 cells/kg. The first two patients at this 
dose level will be observed for 14 days prior to additional enrollment. Among this cohort if there 
are ≥3 DLTs, this arm will be closed early.  
Dose limiting Toxicity 
The dose limiting toxicity will be based on the tolerability observed during the first 28 days of 
treatment/observation. If multiple toxicities are seen, the presence of dose limiting toxicity will be 
based on the most severe toxicity experienced. If a patient dies from a cause felt to be unrelated 
to CAR-T cells (e.g., progressive disease) before completion of the 28 day DLT monitoring 
period, that patient will be replaced. Patients who experience toxicity from lymphodepletion 
chemotherapy that prevents administration of CAR-20/19-T cells will also be replaced. All 
patients who fail to produce the desired dose level will be followed for DLTs, AEs, and survival 
but will be replaced for this dose-finding and dose-expansion study.  
The definition of DLT for this study is detailed in Section 3.4 

 
 
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The statistical analysis will be primarily descriptive in keeping with the exploratory nature of the 
study. We will present summary tables of baseline demographic and clinical characteristics of 
the patients. Adverse events will be tabulated for both overall and within major categories and 
by grade. Analysis of other secondary endpoints such as anti-tumor activity will be presented as 
response rates by disease specific sub-type.  
 
Statistical support will be provided by Mei-Jie Zhang, PhD the senior statistician on this study. 
Dr. Zhang will participate in the study design, data analyses, and the scientific manuscript 
writing associated with this project.  
 
12.5 Analysis Population 
All patients enrolled and consented for this protocol who underwent CAR-20/19-T cell 
production will be evaluable for our feasibility endpoint. Only patients who receive CAR-20/19-T 
cells will be evaluable for our safety endpoint.  
12.6 Lost to Follow-up 
Should a subject fail to return to the clinic for a scheduled protocol specific visit, personnel at the 
investigational site will need to make 2 attempts by a combination of telephone/e-mail and mail 
to contact the subject. Sites must document both attempts to contact the subject. If a subject 
does not respond within 1 month after the second contact the subject will be considered lost to 
follow-up and no additional contact will be required. 
13 DATA AND SAFETY MONITORING PLAN (DSMP)  
Data and Safety Management Overview 
The Medical College of Wisconsin (MCW) Data and Safety Monitoring Committee (DSMC) and 
the MCW Institutional Review Board (IRB) will approve protocol-specific DSM plans. A local, 
investigator-initiated trial will be required to be continuously monitored by the principal 
investigator of the study with monthly (or as each new patient is dosed if no patients enrolled in 
the month) safety and progress reports submitted to the DSMC. For the purpose of this study, a 
medical monitor will be appointed to evaluate serious AEs (Grade 3-5) on a real-time basis. This 
medical monitor is responsible for evaluating serious AEs and can convene a conference call of 
DSMC chair, DSMC reviewers, or external reviewers when necessary.   
13.1 Study Team 
The study team consists of the investigators, clinical research coordinators/assistants, 
regulatory specialist, pharmacists, laboratory team, and the study biostatistician.  
13.2 Quality Assurance  
The MCWCC Clinical Trials Office provides ongoing quality assurance audits 
13.3 Clinical Trials Office 
The MCWCC Clinical Trials Office [CTO] provides administrative assistance and support to the 
DSMC.  

 
 
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13.4 DSMC 
The Medical College of Wisconsin Cancer Center places the highest priority on ensuring the 
safety of patients participating in clinical trials. Every cancer interventional trial conducted at 
MCW includes a plan for safety and data monitoring. 
More information can be found related to the MCWCC Data and Safety Monitoring Plan at the 
MCWCC website (Data and Safety Monitoring Plan).  
This study will be reviewed by the Medical College of Wisconsin Cancer Center Data Safety 
Monitoring Committee (MCW-CC DSMC). External reviewers with CAR-T experience will serve 
on this committee to provide their expertise. A summary of the MCW CC DSMC activities are as 
follows:  
 
Review the clinical trial for data integrity and safety  
 
Review all unexpected grade 3, and all grade 4, and 5 adverse events, as well as any 
others requiring expedited reporting as defined in this protocol. (Grades 4 & 5 events 
must be reported to the DSMC within 5 calendar days of study staff’s knowledge.) 
Review all DSM reports  
 
Submit a summary of any recommendations related to study conduct  
 
Terminate the study if deemed unsafe for patients  
A copy of the MCW-CC Data and Safety Monitoring Plan and membership roster will be 
maintained in the study research file and updated as membership changes. The committee will 
review reports from the study PI twice annually (or more frequently if needed) and provide 
recommendations on trial continuation, suspension or termination as necessary.  
Any available DSMC letters will be submitted to the IRB of record as required. 
14 REGULATORY COMPLIANCE, ETHICS AND STUDY MANAGEMENT 
14.1 Ethical Standard 
This study will be conducted in accordance with the ethical principles that have their origin in the 
Declaration of Helsinki as stated in 21 CFR §312.120(c)(4); consistent with GCP and all 
applicable regulatory requirements. 
14.2 Regulatory Compliance 
This study will be conducted in compliance with: 
 
The protocol 
 
Federal regulations, as applicable, including: 21 CFR 50 (Protection of Human 
Subjects/Informed Consent); 21 CFR 56 (Institutional Review Boards) and §312 
(Investigational New Drug Application; and 45 CFR 46 Subparts A (Common Rule), B 
(Pregnant Women, Human Fetuses and Neonates), C (Prisoners), and D (Children), 

 
 
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GCP/ICH guidelines, and all applicable regulatory requirements. The IRB must comply 
with the regulations in 21 CFR §56 and applicable regulatory requirements. 
14.3 Institutional Review Board 
The protocol, the proposed informed consent form and all forms of participant information 
related to the study (e.g., advertisements used to recruit participants) will be reviewed and 
approved by the MCW Institutional Review Board. Prior to obtaining MCW approval, the 
protocol must be approved by the Medical College of Wisconsin Cancer Center Scientific 
Review Committee. The initial protocol and all protocol amendments must be approved by the 
IRB prior to implementation. The MCW IRB is AAHARPP accredited.  
Informed Consent Process 
Informed consent is a process that is initiated prior to the individual’s agreeing to participate in 
the study and continues throughout the individual’s study participation. Extensive discussion of 
risks and possible benefits of this therapy will be provided to the subjects and their families. 
Consent forms describing in detail the study interventions/products, study procedures and risks 
are given to the subject and written documentation of informed consent is required prior to 
starting intervention/administering study product. 
Informed consent from the patient will be obtained using forms approved by the local 
institution’s IRB. Informed consent will be obtained by the Principal Investigator or Co-
Investigators. Consent forms will be IRB-approved and the subject will be asked to read and 
review the document. Upon reviewing the document, the investigator will explain the research 
study to the subject and answer any questions that may arise. In accordance with 46 CR 
46.111, the subject or representative legal guardian will sign and date the informed consent 
document prior to any procedures being done specifically for the study. The subjects may 
withdraw consent at any time throughout the course of the trial. 
A copy of the informed consent document will be given to the subjects for their records. The 
MCWCC CTO will follow the MCW/FH IRB’s policy for subjects who demonstrate limited English 
proficiency or limited literacy. The original consent is kept with the subject’s study file, and a 
copy of the consent is sent to the OCRICC office, which will then submit to HIM a copy of the 
signed consent to be scanned into EPIC, the legal medical record. 
14.4 Subject Confidentiality and Access to Source Documents/Data 
Subject confidentiality is strictly held in trust by the sponsor-investigator, participating 
investigators, and any staff. This confidentiality includes the clinical information relating to 
participating subjects, as well as any genetic or biological testing. 
The study protocol, documentation, data, and all other information generated will be held in 
strict confidence. No information concerning the study or the data will be released to any 
unauthorized third party without prior written approval of the sponsor-investigator. 
The conditions for maintaining confidentiality of the subjects’ records are required for the life of 
the data. These rules apply equally to any and all MCWCC projects.  
While data are being collected and after all data have been collected but are still in the process 
of being analyzed, the subject’s data/PHI are stored in the locked Clinical Research office in the 

 
 
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Clinical Trials Office. Databases in which the study subject information is stored and accessed 
are password protected, allowing for limited access by authorized personnel only. Data/PHI kept 
in the Case Report Forms in Oncore contain the study identifiers, subject initials, date of birth 
and date of service.  
The principal investigator will allow access to all source data and documents for the purposes of 
monitoring, audits, IRB review, and regulatory inspections.  
The Clinical Trials Office quality assurance monitor may inspect all documents and records 
required to be maintained by the investigator, including but not limited to, medical records 
(office, clinic, or hospital) and pharmacy records for the subjects in this study. The clinical study 
site will permit access to such records. 
Onsite Audits 
Auditing is essential to ensure that research conducted at the Medical College of Wisconsin 
(MCW) Cancer Center is of the highest quality and meets MCW and regulatory agency 
standards.  
Regulatory authorities, the IRB and/or sponsor may request access to all source documents, 
data capture records and other study documentation for on-site audit or inspection. Direct 
access to these documents must be guaranteed by the investigator, who must provide support 
at all times for these activities. 
14.5 Quality Assurance 
This protocol will be reviewed internally every six months by the Cancer Center Clinical Trials 
Office Quality Assurance Staff. Approximately 30% of subject files will be selected randomly for 
review (max 10 subjects at each monitoring timepoint). Consent, eligibility and objective based 
data will be reviewed for those files selected.  One file will be selected randomly for a 
comprehensive review at each quality assurance review timepoint. Regulatory will be reviewed 
at the time of each review.  
 
14.6 Changes in the Protocol 
Once the protocol has been approved by the MCW IRB, any changes to the protocol must be 
documented in the form of an amendment. The amendment must be signed by the investigator 
and approved by IRB prior to implementation.  
If it becomes necessary to alter the protocol to eliminate an immediate hazard to patients, an 
amendment may be implemented prior to IRB approval. In this circumstance, however, the 
Investigator must then notify the IRB in writing within five working days after implementation.  
The IRB may provide, if applicable regulatory authority(ies) permit, expedited review and 
approval/favorable opinion for minor change(s) in ongoing studies that have the approval 
/favorable opinion of the IRB. The investigator will submit all protocol modifications to the 
sponsor and the regulatory authority(ies) in accordance with the governing regulations. 
Any departures from the protocol must be fully documented in the source documents. 

 
 
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15 DATA HANDLING AND RECORD KEEPING 
15.1 Overview 
Every effort is made to uphold the integrity of the project, the research, the institution, and the 
researchers involved. Data collection guidelines and methodologies are carefully developed 
before the research begins. Investigators focus on the following to ensure data integrity:  well-
trained data collectors/recorders to ensure consistency and quality, well-designed data 
collection protocols and ongoing monitoring. In this way, study rigor and validity are maintained. 
Data is protected from physical damage as well as from tampering, loss or theft. This project’s 
data management is a multidisciplinary activity that includes investigators, research 
coordinators and nurses, data mangers, support personnel, biostatisticians and database 
programmers. Quality control will be applied to each stage of data handling to ensure that all 
data are reliable and have been processed correctly. MCW Cancer Center database will be 
utilized for data collection and case report forms through OnCore.  
15.2 Study Record Retention 
The principal investigator will maintain adequate records of the disposition of the CAR-20/19-T 
cells, including dates, quantity and use by subjects and the disposition of cells manufactured but 
not infused. 
The principal investigator, along with assistance from the research coordinators, is required to 
prepare and maintain adequate and accurate case histories that record all observations and 
other data pertinent to the investigation on each individual administered the CAR-20/19-T cells. 
Case histories include the case report forms and supporting data including, for example, signed 
and dated consent forms and medical records including, for example, progress notes of the 
physician, the individual's hospital chart(s), and the nurses' notes. The case history for each 
individual shall document that informed consent was obtained prior to participation in the study. 
Study documentation includes all CRFs, data correction forms or queries, source documents, 
sponsor-investigator correspondence, monitoring logs/letters, and regulatory documents (e.g., 
protocol and amendments, IRB correspondence and approval, signed patient consent forms). 
Source documents include all recordings of observations or notations of clinical activities and all 
reports and records necessary for the evaluation and reconstruction of the clinical research 
study. 
In accordance with FDA regulations, the investigator shall retain records for a period of two 
years following the date a marketing application is approved for the drug for the indication for 
which it is being investigated; or, if no application is to be filed or if the application is not 
approved for such indication, until two years after the investigation is discontinued and FDA is 
notified. 
15.3 Publishing Data 
Data from this study will be managed by the clinical trials office at MCW and the investigators 
listed on this study. Upon completion of the study, the data will be submitted to national meeting 
followed by a plan to publish in peer-reviewed journal.  
 
 

 
 
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APPENDIX A: PERFORMANCE STATUS CRITERIA 
 
Karnofsky Performance Scale       
Percent 
Description 
100 
Normal, no complaints, no evidence 
of disease 
90 
Able to carry on normal activity; minor 
signs or symptoms of disease 
80 
Normal activity with effort; some signs 
or symptoms of disease 
70 
Cares for self, unable to carry on 
normal activity or to do active work 
60 
Requires occasional assistance, but 
is able to care for most of his/her 
needs 
50 
Requires considerable assistance and 
frequent medical care 
40 
Disabled, requires special care and 
assistance 
30 
Severely disabled, hospitalization 
indicated 
Death not imminent 
20 
Very sick, hospitalization indicated 
Death not imminent 
10 
Moribund, fatal processes 
progressing rapidly 
0 
Dead 
 
 
 

 
 
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APPENDIX B:  NEUROLOGICAL ASSESSMENT 
The Mini-Mental State Examination will be performed as per Table 1: Schedule of Events.  

 
 
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APPENDIX C: 2004 MAS/HLH DIAGNOSTIC CRITIERA 
Diagnosis of MAS/HLH requires meeting 5/8 following criteria 
 
• 
Fever 
• 
Splenomegaly 
• 
Cytopenias (affecting at least 2 of 3 lineages in the peripheral blood) 
o Hemoglobin <9 g/dL  
o Platelets <100,000/µL 
o Absolute neutrophil count<1000/µL 
• 
Hypertriglyceridemia and/or hypofibrinogenemia: 
o Fasting triglycerides >265 mg/dl 
o Fibrinogen <1.5 g/L 
• 
Hemophagocytosis in bone marrow or spleen or lymph nodes 
• 
Low or absent NK-cell activity (according to local laboratory reference) 
• 
Ferritin >500 ng/mL 
• 
Soluble CD25 (i.e., soluble IL-2 receptor) >2,400 U/ml 
 
 

 
 
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APPENDIX D: SCHEMA FOR TESTING OF CART CELLS FOR FRESH 
INFUSION OR CRYOPRESERVATION (FOR LATER INFUSION) IN 
PHASE 1 DOSE ESCALATION/EXPANSION 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Day -6 In-Process Testing (minimal testing requirements) 
(Rationale:  For those receiving CAR-T cells as a fresh infusion, lymphodepleting 
chemotherapy would be initiated on Day -4 This allows 2 days to verify CAR-T potency before 
lymphodepletion is initiated.) 
1. Sterility cultures 
2. Transduction efficiency (Protein L staining for CD20_19 expression) 
3. Basic phenotyping (CD3, CD4, CD8 from Protein L panel) 
4. Viability (from Protein L panel) and viable cell numbers 
5. Chromium release assays versus CD19+/ CD20+ Raji target cells (potency) 
6. Mycoplasma testing (in-house) 
7. Endotoxin testing (sendout) 
 
Day 0 Testing of Final Product 
(Important Note:  For those receiving CAR-T cells as a fresh infusion, results of gram stains, 
flow cytometry (to assess transduction efficiency) and 7-AAD viability will be required for product 
release.) 
1. Sterility cultures 
2. Gram stain 
3. Transduction efficiency (by Protein L staining) 
4. Comprehensive phenotyping 
5. Chromium release assays versus CD19+/ CD20+ Raji target cell and transfected CD19+ 
and CD20+ target cells (potency) 
6. Intracellular cytokine staining assays (potency) 
7. VCN & VSVg testing (sendout) 
8. Sendout for final mycoplasma and endotoxin testing 
Day 0 + 6 weeks 
Harvest for Fresh Infusion or 
Cryopreservation for later infusion 
Final Testing 
Day -15 
Apheresis 
Day -14 
Cell 
Separation 
Day -13 
Transduction 
Day -8 (per Lab PI 
discretion) and 
Day  -6 
Sampling & 
Testing 
Day -4  
LDP 

 
 
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APPENDIX E: CAR-T CELL TREATMENT GUIDELINES 
□ Tumor Lysis Prevention 
o Allopurinol 300 mg daily for tumor lysis prevention starting Day -4 until 
Day+141 
□ Seizure prophylaxis  
o Keppra (Levetiracetam) 500 mg twice daily from Day -4 until Day +28  
□ Prophylactic Antibiotics 
o PCP prophylaxis to be started Day -4 through Day+28 (agent and dosing 
per institutional standards) 
o Anti-viral prophylaxis: Acyclovir or valacyclovir starting Day -4 through 
Day+28. Valganciclovir or IV ganciclovir can be given as a substitute as 
indicated by treating provider. 
o Neutropenic prophylaxis: As per institutional standards, not mandatory 
(e.g. ciprofloxacin or cefepime) 
o Fungal prophylaxis as per treating physician, not mandatory 
□ Pre-medications for CAR-20/19-T cells (1 hour prior to CAR-T cell infusion) 
o Acetaminophen 650 mg x 1 orally  
o Diphenhydramine hydrochloride 50 mg x 1 oral or IV2 
Transfusion Parameters 
o Maintain Platelets>20 K/µL and Hemoglobin>8.0 mg/dL 
o Filgrastrim can be given at the discretion of the treating provider starting 
Day +21, provided the patient does not have signs/symptoms consistent 
with CAR-T cell neurotoxicity or cytokine release syndrome.  
 
 
 
 
 
 
 
 
 
 
 
 
 
1- Can be stopped at discretion of treating physician 
2- Alternatives anti-histamines in place of diphenhydramine hydrochloride can be given per discretion of 
treating physician (e.g. cetirizine, ranitidine) Dosed per institutional standards. 
 

 
 
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APPENDIX F: ASSESSMENT/MANAGEMENT OF CYTOKINE RELEASE SYNDROME AND 
NEUROTOXICITY ASSOCIATED WITH CAR-T CELLS [1, 56] 
 
Lee et al. CRS Grading System1  
 
 
 
  
 
 
 
1. Lee, D.W., et al., Current concepts in the diagnosis and management of cytokine release syndrome. Blood, 2014. 124(2): p. 188-
95. 

 
 
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Table 5: CRS Symptoms, Evaluation, Management, Response 
CRS assessed 
via Lee et al. 
Revised CRS 
grading system.  
Grade 1 
toxicities 
Grade 2 
toxicities1 
Grade 3 or Grade 4 toxicities 
Symptoms 
Fevers, myalgias, 
nausea/fatigue, malaise, 
headache 
Hypotension that 
responds to fluid or one 
low dose pressor, oxygen 
requirement<40%, Grade 
2 organ toxicity per 
CTCAE 5.0 
Hypotension requiring high dose or multiple 
pressors, Grade 3 or 4 organ toxicity, life 
threatening symptoms, mechanical ventilation 
Evaluation & 
Management 
Infectious work-up, 
initiation/escalation of 
antibiotics per institutional 
standards. IV fluids, anti-
pyretic therapy as 
indicated with cooling 
blankets and 
acetaminophen (avoid 
NSAIDs). Supportive care 
measure2 
Admission to hospital (if 
not admitted). IV fluids, 
infectious work-up, 
supportive care measures 
Admission to hospital, intensive care unit 
transfer as indicated. IV fluids, infectious 
work-up, respiratory support, and renal 
replacement therapy as indicated 
Response & 
Escalation 
Persistent Grade 1 
symptoms>24-48 hours 
or escalation to Grade 3-4 
toxicities initiate CRS 
directed treatment3 
Persistent symptoms 
>12-24 hours despite 
interventions or 
escalation to Grade 3-4 
toxicity initiate CRS 
directed treatment3 
Immediate initiation of CRS directed 
treatments3 
 
1-At direction of treating physician, CRS treatment can be initiated in any patient with Grade≥2 toxicity  
2-Recommended Supportive care measure listed in Appendix G 
3-CRS Directed Treatment Algorithm is on Page 91 
 
 

 
 
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Table 6: Neurotoxicity Evaluation and Management 
 
Neurotoxicity 
assessed via 
CTCAE 5.0 
Grade 1 
Grade 2 
Grade 3 
Grade 4 
Examples 
Mild memory 
impairment, reduced 
awareness or alertness, 
mild paresthesia, etc.  
Moderate memory 
impairment or moderate 
lethargy limiting ADL. 
Moderate weakness or 
sensory loss moderate 
neuropathy limiting 
instrumental ADL, etc. 
Severe memory 
impairment limiting self-
care, severe weakness 
or sensory loss, multiple 
seizures, severe 
neurological deficit, etc.  
Life threatening 
consequence, cerebral 
edema1 urgent 
intervention indicated 
Evaluation 
Neurological Exam, 
additional work-up as 
indicated 
Neurological Examination, MRI Brain +/- lumbar puncture and EEG as 
indicated.  Neurological consultation.  
Management 
Clinical vigilance, 
monitoring, supportive 
care 
Admission to hospital (if 
not admitted), neuro-
checks Q4 hours, 
supportive care, work-up 
as above. 
Dexamethasone can be 
given at discretion of 
treating physician 
Admission to Intensive Care Unit. Dexamethasone 
10 mg IV/PO every 6-12 hours (max dose 10 mg 
Q6 hours). If not improving consider tocilizumab 8 
mg/kg IV x 1 (max dose 800 mg). Can give up to 2 
doses of Tocilizumab in 24 hours.  
1-For patients with evidence of cerebral edema (Grade 4 toxicity) the following specific interventions will be performed 
• 
Intensive Care Unit care 
• 
Neurosurgical consultation 
• 
Mechanical ventilation 
• 
Pharmacological support per intensivist and neurological and neurosurgical recommendations 

 
 
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Cytokine Release Syndrome Directed Treatment[56] 
 
Without Neurological Toxicities 
 
 
 
 
 
 
 
 
 
 
 
 
 
With Neurological Toxicities1 
 
 
 
 
 
 
 
 
 
 
1-Patients with neurological toxicities (Grade≥3 by CTCAE 5.0) will be treated with dexamethasone with or without the presence of systemic CRS. 
Tocilizumab 8 mg/kg 
(Max dose 800 mg) 
Response 
within 12-24 
hours? 
NO 
YES 
Grade 3 or Grade 4 Neurotoxicity 
as listed in Table 6 
Repeat Tocilizumab 8 
mg/kg (Max Dose 800 mg)  
Supportive Care per 
Appendix G 
Dexamethasone 10 mg 
IV/PO every 6-12 hours 
(max dose 10 mg Q6 hours)  
Dexamethasone 10 mg IV/PO 
every 6-12 hours (max dose 10 
mg Q6 hours). If not improving 
consider or with concurrent 
systemic CRS, can administer 
tocilizumab 8 mg/kg IV x 1 (max 
dose 800 mg). Can give up to 
two doses of Tocilizumab in 24 
hours.
Persistent 
symptoms 
Continue Dexamethasone until 
significant improvement + supportive 
care as per Appendix G. Patients with 
progressive systemic CRS symptoms 
see above algorithm. 

 
 
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APPENDIX G: SUPPORTIVE CARE MEASURES FOR CAR-T CELL 
TREATMENT 
Guidelines developed and adapted from Brudno, J. N. and J. N. Kochenderfer (2016). 
"Toxicities of chimeric antigen receptor T cells: recognition and management." Blood 127(26): 
3321-3330. 
 
 
Table 7: Supportive Care Guidelines for Patients Receiving CAR-T cells 
 
Constitutional 
• 
Administer acetaminophen for symptomatic management of fevers in patients 
with normal hepatic function.  
• 
Provide cooling blankets for fevers >40oC 
• 
Avoid corticosteroids and NSAIDs 
• 
Avoid meperidine  
Cardiovascular 
• 
Minimize anti-hypertensives prior to cell infusion  
• 
Monitor vital signs at least every 4 hours for up to 48 hours’ post CAR-20/19-T 
cell infusion 
• 
Monitor vital signs Q2H in patients with fevers and tachycardia 
• 
Initiate replacement of IV fluids for patients with poor oral intake or high 
insensible losses to maintain net even fluid balance 
• 
IV fluid boluses for patients with SBP less than their pre-infusion baseline or 
those with hypotension SBP<90 
• 
Persistent hypotension not responsive to fluids should be initiated on 
vasopressors and patients transferred to intensive care unit 
• 
For patient on vasopressors evaluate cardiac function with troponin and ECG. 
Echocardiogram can be performed as indicated.  
Infectious disease 
• 
Prophylactic antibiotics as per Appendix E 
• 
Neutropenic Fever management as per institutional guidelines with broad 
spectrum antibiotics. Appropriate fever work-up with chest x-ray, urinalysis, 
and blood cultures 
Hematologic 
 
• 
Initiate Allopurinol for TLS prevention as per Appendix E 
• 
Transfuse RBCs for Hgb<8 g/dL 
• 
Transfuse platelets for Platelets<20 k/µL 
• 
Monitor ANC as per schedule of events (Table 1). Neupogen can be started at 
Day+21 in patients with persistent neutropenia as per guidelines in Section 6.9 
• 
Transfuse fresh frozen plasma in patients with PTT prolongation >2.0 fold 
upper limit of normal 
• 
Transfuse cryoprecipitate to maintain fibrinogen ≥100 mg/dL 
Neurologic 
• 
Neuro checks Q4H by nursing staff in patients with ≥Grade 2 neurotoxicity 
• 
MRI Brain +/- lumbar puncture and EEG in patients with ≥Grade 2 
neurotoxicity 
• 
Neurological consultation for patients with ≥Grade 2 neurotoxicity 
• 
MMSE evaluation as per Table 1 
 
 

 
 
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APPENDIX H: OPERATING PROCEDURES FOR PATIENTS TREATED 
WITH CAR-20/19-T CELLS AT A NON-SPECIFIED DOSE 
Some patients will have successful production of CAR-20/19-T cells but fail to make the 
adequate cell dose as required per our protocol. Those patients will be allowed treatment but 
required to be replaced for the purposes of this Phase 1/1b study. Patients will be followed as 
per this clinical protocol with monitoring for DLT, safety, adverse events, and long-term survival.  
 
Specific Criteria for Infusion of Non-Specified CAR-20/19-T cell dose 
 
1) Meet all eligibility criteria a listed in Section 4 
2) Meet all release criteria for CAR-T cell product 
3) Meet same infusion criteria as listed in Section 6.3 and 7.1 
4) Patients will be followed per schedule of events listed in Section 5 and Table 1 
 
Analysis of Patients Treated with Non-Specified CAR-20/19-T cell dose 
 
At first, a maximum of 3 patients will be allowed to be treated in this manner. If there is ≤1 DLT 
within the first 28 days for the first three patients, further patients will be allowed study drug with 
CAR-20/19-T cells even if they don’t make the required treatment dose. After enrollment of the 
4th patient or beyond in this cohort, if the DLT rate exceeds ≥33%, no further patients will be 
allowed treatment outside of a specified cell dose per the clinical protocol.  
All patients will be followed for AEs, survival, response rates as per Table 1.  
 
 

 
 
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APPENDIX I: QUALITY OF LIFE (QOL) MEASURES 
QOL MEASURES 
 
Participant # _____ 
IDAS 
Instructions: Below is a list of feelings, sensations, problems, and experiences that people sometimes have. Read each item 
to determine how well it describes your recent feelings and experiences.  Then select the option that best describes how 
much you have felt or experienced things this way during the past two weeks, including today.   
 
 
Not at all 
A little bit 
Moderately 
Quite a bit 
Extremely 
1. I was proud of myself 
1 
2 
3 
4 
5 
2. I felt exhausted 
1 
2 
3 
4 
5 
3. I felt depressed 
1 
2 
3 
4 
5 
4. I felt inadequate 
1 
2 
3 
4 
5 
5. I slept less than usual 
1 
2 
3 
4 
5 
6. I felt fidgety, restless 
1 
2 
3 
4 
5 
7. I had thoughts of suicide
 
 
1 
2 
3 
4 
5 
8. I slept more than usual 
1 
2 
3 
4 
5 
9. I hurt myself purposely   
1 
2 
3 
4 
5 
10. I slept very poorly 
1 
2 
3 
4 
5 
11. I blamed myself for things
 
 
1 
2 
3 
4 
5 
12. I had trouble falling asleep
 
 
1 
2 
3 
4 
5 
13. I felt discouraged about things 
1 
2 
3 
4 
5 
14. I thought about my own death
 
 
1 
2 
3 
4 
5 

 
 
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15. I thought about hurting myself
 
 
1 
2 
3 
4 
5 
16. I did not have much of an appetite
 
 
1 
2 
3 
4 
5 
17. I felt like eating less than usual
 
 
1 
2 
3 
4 
5 
19. I did not feel much like eating 
1 
2 
3 
4 
5 
21. I felt optimistic 
1 
2 
3 
4 
5 
23. I felt that I had accomplished a lot
 
 
1 
2 
3 
4 
5 
24. I looked forward to things with enjoyment
 
 
1 
2 
3 
4 
5 
25. I was furious 
1 
2 
3 
4 
5 
26. I felt hopeful about the future
 
 
1 
2 
3 
4 
5 
27. I felt that I had a lot to look forward to
 
 
1 
2 
3 
4 
5 
28. I felt like breaking things
 
  
1 
2 
3 
4 
5 
29. I had disturbing thoughts of something bad that happened to me
 
 
1 
2 
3 
4 
5 
30. Little things made me mad
 
 
1 
2 
3 
4 
5 
31. I felt enraged 
 
1 
2 
3 
4 
5 
32. I had nightmares that reminded me of something bad that happened
 
 
1 
2 
3 
4 
5 
33. I lost my temper and yelled at people 
1 
2 
3 
4 
5 
34. I felt like I had a lot of interesting things to do.
 
 
1 
2 
3 
4 
5 
35. I felt like I had a lot of energy
 
 
1 
2 
3 
4 
5 
36. I had memories of something scary that happened
 
 
1 
2 
3 
4 
5 
37. I felt self-conscious knowing that others were watching me
 
 
1 
2 
3 
4 
5 

 
 
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38. I felt a pain in my chest  
1 
2 
3 
4 
5 
39. I was worried about embarrassing myself socially
 
 
1 
2 
3 
4 
5 
40. I felt dizzy or light headed
 
 
1 
2 
3 
4 
5 
41. I cut or burned myself on purpose
 
 
1 
2 
3 
4 
5 
42. I had little interest in my usual hobbies or activities
 
 
1 
2 
3 
4 
5 
43. I thought that the world would be better off without me 
1 
2 
3 
4 
5 
 
 
44. I felt much worse in the morning than later in the day
 
 
1 
2 
3 
4 
5 
45. I felt drowsy, sleepy 
1 
2 
3 
4 
5 
46. I woke up early and could not get back to sleep 
1 
2 
3 
4 
5 
47. I had trouble concentrating
 
 
1 
2 
3 
4 
5 
48. I had trouble making up my mind
 
 
1 
2 
3 
4 
5 
49. I talked more slowly than usual
 
 
1 
2 
3 
4 
5 
50. I had trouble waking up in the morning 
1 
2 
3 
4 
5 
51. I found myself worrying all the time
 
 
1 
2 
3 
4 
5 
52. I woke up frequently during the night
 
 
1 
2 
3 
4 
5 
53. It took a lot of effort for me to get going
 
 
1 
2 
3 
4 
5 
54. I woke up much earlier than usual
 
 
1 
2 
3 
4 
5 
55. I was trembling or shaking 
1 
2 
3 
4 
5 
56. I became anxious in a crowded public setting
 
 
1 
2 
3 
4 
5 
57. I felt faint 
1 
2 
3 
4 
5 

 
 
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58. I found it difficult to make eye contact with people
 
 
1 
2 
3 
4 
5 
59. My heart was racing or pounding
 
 
1 
2 
3 
4 
5 
60. I got upset thinking about something bad that happened
 
 
1 
2 
3 
4 
5 
61. I found it difficult to talk with people I did not know well
 
 
1 
2 
3 
4 
5 
62. I had a very dry mouth 
1 
2 
3 
4 
5 
63. I was short of breath 
1 
2 
3 
4 
5 
64. I felt like I was choking 
1 
2 
3 
4 
5 
 
 
 

 
 
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FSI 
 
Instructions: The following questions ask about fatigue during the past week. For each of the following, circle the 
one number that best indicates how that item applies to you. 
 
1. Rate your level of fatigue on the day you felt most fatigued during the past week. 
0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
Not at all 
fatigued 
 
 
 
 
 
 
 
 
 
As fatigued as 
I could be 
 
2. Rate your level of fatigue on the day you felt least fatigued during the past week. 
0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
Not at all 
fatigued 
 
 
 
 
 
 
 
 
 
As fatigued as I 
could be 
 
3. Rate your level of fatigue on the average in the last week. 
0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
Not at all 
fatigued 
 
 
 
 
 
 
 
 
 
As fatigued as I 
could be 
 
4. Rate your level of fatigue right now. 
0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
Not at all 
fatigued 
 
 
 
 
 
 
 
 
 
As fatigued as I 
could be 
 
5. Rate how much, in the past week, fatigue interfered with: 
 
No 
interference 
 
Extreme 
interference 
a. Your general level of activity ..........................  0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
b. Your ability to bathe and dress yourself .........  0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
c.  Your normal work activity (includes both 
work outside the home and housework) .........  
 
0 
 
1 
 
2 
 
3 
 
4 
 
5 
 
6 
 
7 
 
8 
 
9 
 
10 
d. Your ability to concentrate ..............................  0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
e. Your relations with other people .....................  0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
f. Your enjoyment of life .....................................  0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
g. Your mood ......................................................  0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
 
6. Indicate how many days, in the past week, you felt fatigued for any part of the day: 
0 
1 
2 
3 
4 
5 
6 
7 
 
7. Rate how much of the day, on average, you felt fatigued in the past week 
0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
None of the 
day 
 
 
 
 
 
 
 
 
 
The entire day 
 
 

 
 
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PSQI 
 
Instructions: The following questions relate to your usual sleep habits during the past month only. Your answers 
should indicate the most accurate reply for the majority of days and nights in the past month. Please answer all 
questions. 
 
1. During the past month, what time have you usually gone to bed at night? 
 
BED TIME ___________ 
 
2. During the past month, how long (in minutes) has it usually taken you to fall asleep each night? 
 
NUMBER OF MINUTES ___________ 
 
3. During the past month, what time have you usually gotten up in the morning? 
 
GETTING UP TIME ___________ 
 
4. During the past month, how many hours of actual sleep did you get at night? (This may be 
different than the number of hours you spent in bed.) 
 
HOURS OF SLEEP PER NIGHT ___________ 
 
For each of the remaining questions, check the one best response. Please answer all questions. 
 
5. During the past month, how often have you had trouble sleeping because you . . . 
 
a) Cannot get to sleep within 30 minutes 
 
Not during the 
past month _____  
Less than 
once a week _____  
Once or twice 
a week _____  
Three or more 
times a week _____  
b) Wake up in the middle of the night or early morning 
 
Not during the 
past month _____  
Less than 
once a week _____  
Once or twice 
a week _____  
Three or more 
times a week _____  
c) Have to get up to use the bathroom 
 
Not during the 
past month _____  
Less than 
once a week _____  
Once or twice 
a week _____  
Three or more 
times a week _____  
d) Cannot breathe comfortably 
 
Not during the 
past month _____  
Less than 
once a week _____  
Once or twice 
a week _____  
Three or more 
times a week _____  
 

 
 
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e) Cough or snore loudly 
 
Not during the 
past month _____  
Less than 
once a week _____  
Once or twice 
a week _____  
Three or more 
times a week _____  
f) Feel too cold 
 
Not during the 
past month _____  
Less than 
once a week _____  
Once or twice 
a week _____  
Three or more 
times a week _____  
g) Feel too hot 
 
Not during the 
past month _____  
Less than 
once a week _____  
Once or twice 
a week _____  
Three or more 
times a week _____  
h) Had bad dreams 
 
Not during the 
past month _____  
Less than 
once a week _____  
Once or twice 
a week _____  
Three or more 
times a week _____  
i) Have pain 
 
Not during the 
past month _____  
Less than 
once a week _____  
Once or twice 
a week _____  
Three or more 
times a week _____  
j) Other reason(s), please describe_________________________________________________ 
__________________________________________________________________________ 
 
How often during the past month have you had trouble sleeping because of this? 
 
Not during the 
past month _____  
Less than 
once a week _____  
Once or twice 
a week _____  
Three or more 
times a week _____  
 
 
 
 
6. During the past month, how would you rate your sleep quality overall? 
 
Very good 
__________ 
Fairly good 
__________ 
Fairly bad 
__________ 
Very bad 
__________ 
 
 
 
 
 

 
 
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7. During the past month, how often have you taken medicine to help you sleep (prescribed or 
"over the counter")? 
 
Not during the 
past month _____  
Less than 
once a week _____  
Once or twice 
a week _____  
Three or more 
times a week _____  
 
8. During the past month, how often have you had trouble staying awake while driving, eating 
meals, or engaging in social activity? 
 
Not during the 
past month _____  
Less than 
once a week _____  
Once or twice 
a week _____  
Three or more 
times a week _____  
9. During the past month, how much of a problem has it been for you to keep up enough enthusiasm to get things 
done? 
No problem at all 
__________ 
Only a very slight problem 
__________ 
Somewhat of a problem 
__________ 
A very big problem 
__________ 
 
 
 

 
 
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BPI 
 
Instructions: The following questions ask about pain during the past week. For each of the following, circle the one 
number that best indicates how that item applies to you. 
 
1. Please rate your pain by circling the number that best describes your pain at its worst in the last week. 
0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
No 
pain 
 
 
 
 
 
 
 
 
 
Pain as bad as you 
can imagine 
 
2. Please rate your pain by circling the number that best describes your pain at its least in the last week. 
0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
No 
pain 
 
 
 
 
 
 
 
 
 
Pain as bad as you 
can imagine 
 
3. Please rate your pain by circling the number that best describes your pain on the average in the last week. 
0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
No 
pain 
 
 
 
 
 
 
 
 
 
Pain as bad as you 
can imagine 
 
4. Please rate your pain by circling the number that best describes your pain right now. 
0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
No 
pain 
 
 
 
 
 
 
 
 
 
Pain as bad as you 
can imagine 
 
5. Circle the number that describes how, during the past week, pain has interfered with your: 
 
Does not 
interfere 
 
Completely 
interferes 
a. General activity ...............................................  0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
b. Mood ...............................................................  0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
c. Walking ability ................................................  0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
d. Normal work (includes both work outside the 
home and housework) ....................................  
 
0 
 
1 
 
2 
 
3 
 
4 
 
5 
 
6 
 
7 
 
8 
 
9 
 
10 
e. Relations with other people .............................  0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
f. Sleep ................................................................  0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
g. Enjoyment of life ............................................  0 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
 
 
 
 

 
 
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THESE QUESTIONS NEED ONLY BE ANSWERED ONCE; NO NEED TO ANSWER AT SUBSEQUENT 
STUDY FOLLOW-UP VISITS: 
Please indicate below what best represents your household income: 
_____ <$10,000 
_____ $10,001-$25,000 
_____ $25,001-$40,000 
_____ $40,001-$55,000 
_____ $55,001-$70,000 
_____ $70,001-$85,000 
_____ $85,001-$100,000 
_____ > $100,000 
   
Please indicate which education level best describes you: 
_____ Less than 12 years 
_____ High School 
_____ Trade School 
_____ Some College 
_____ College Graduate 
_____ Post-Graduate Degree  
 
 
 
  
 
 

 
 
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APPENDIX J: SPECIMEN COLLECTION AND HANDLING 
For patients who have progressed, mandatory blood samples for monitoring for CAR-T cell persistence 
which include T-cell persistence by flow and by quantitative PCR and presence of replication competent 
virus by Q-PCR for VSV-g can be drawn locally and sent to Medical College of Wisconsin for analysis.  
Peripheral blood sample:   
- 
T-cell Persistent Studies: A minimum of 20mL of peripheral blood, separated into 2-10mL EDTA 
(purple top) tubes.  
- 
Q-PCR Samples: A minimum of 20mL of peripheral blood, separated into 2- 10mL EDTA (purple 
top) tubes. 
- 
VSV-g Samples: A minimum of 20mL of peripheral blood, separated into 2- 10mL EDTA (purple 
top) tubes. 
 
Specimen Labeling  
All submitted specimens must be labeled with the patient’s initials, patient study ID#, patient’s date 
of birth and date/time of specimen collection. (All supplies and airbills will be provided in research 
sample kit).  
 
Shipping Requirements 
All specimens must be accompanied by a Specimen Shipping Requisition Form. Samples must be 
received by the Medical College of Wisconsin within 24 hours of draw. The Froedtert-Cell 
Processing Lab is open M-F, 8:00am – 4:30 pm.  Please do not draw samples on Fridays for 
delivery on Saturday.   
 
The sample must be placed in a leak proof primary receptacle (ex: vacutainer tube). Multiple 
primary receptacles must be individually wrapped or separated to prevent contact.  
 
The primary receptacle must be placed into a leak proof secondary container (ex: resealable 
biohazard bag) in such a way that under normal conditions of transport, they cannot break or leak.  
Absorbent material, such as paper towels or absorbent pads, must be placed in the secondary 
container with sufficient capacity to absorb the entire contents of the primary receptacle(s).  
 
The secondary container must be placed into an outer shipping container with suitable cushioning 
material. (ie: Gel pack) Ice packs to be added to the bottom of shipping container, and on top of the 
samples. The shipping container must be labeled with the universal biohazard symbol.  
 
The outer packaging must be marked with the name, address, and phone number of both the 
sender and recipient. All packages shipped via aircraft must display a 2-inch diamond with 
"UN3373" inside of the diamond.  
 
Send the container via next day delivery at refrigerated temperature to: 
 
 
Froedtert- Cell Processing Lab   
 
Attn: Fenlu Zhu 
 
9200 W. Wisconsin Ave  
 
 
Milwaukee, WI 53226 
 
Please call/email the Cell Processing Lab and Coordinator as soon as the tracking number is available: 
(414)805-6144, Fenlu Zhu:FZhu@mcw.edu, and (414)805-8378, Sharon Yim: syim@mcw.edu, The Cell 

 
 
IIT-CAR-T-Shah 
116 
Version 5  
 
 
Version Date:06/18/2019 
Processing Lab is open M-F, 8:00am – 4:30 pm.  Please do not draw samples on Fridays for delivery on 
Saturday.