Phase I Study of Adoptive Immunotherapy for Advanced ROR1+ Malignancies With Defined Subsets of Autologous T Cells Engineered to Express a ROR1-Specific Chimeric Antigen Receptor
Phase: PHASE1
Conditions: Hematopoietic and Lymphoid Cell Neoplasm, Malignant Solid Neoplasm, Metastatic Lung Non-Small Cell Carcinoma, Metastatic Triple-Negative Breast Carcinoma, Recurrent Acute Lymphoblastic Leukemia, Recurrent Mantle Cell Lymphoma, Refractory Chronic Lymphocytic Leukemia, Stage III Lung Non-Small Cell Cancer AJCC v7, Stage IIIA Lung Non-Small Cell Cancer AJCC v7, Stage IIIB Lung Non-Small Cell Cancer AJCC v7, Stage IV Breast Cancer AJCC v6 and v7, Stage IV Lung Non-Small Cell Cancer AJCC v7, Unresectable Lung Non-Small Cell Carcinoma
Sponsor: Fred Hutchinson Cancer Center

1 
FRED HUTCHINSON CANCER RESEARCH CENTER 
UNIVERSITY OF WASHINGTON SCHOOL OF MEDICINE 
PROTOCOL 9330 
Current Version: 08/19/2020 
Previous Version: 12/18/2018 
1.
TITLE OF PROTOCOL: Phase I study of adoptive immunotherapy for advanced
ROR1+ malignancies with defined subsets of autologous T cells engineered to express a ROR1-
specific chimeric antigen receptor
Investigators 
Professional Title 
Telephone Number 
PI - David Maloney, MD, PhD 
Professor, FHCRC 
 
206-667-5616
Jennifer Specht, MD 
Associate Professor, UW 
206-288-6889
Sylvia Lee, MD 
 
Assistant Professor, UW 
206-288-2274
Cameron Turtle, MBBS, PhD 
Associate Professor, FHCRC  
206-667-7073
Josh Veatch MD, PhD 
Post doctoral Fellow, FHCRC 
206-667-5108
Stanley Riddell, MD 
Professor, FHCRC 
 
206-667-5249
Immunotherapy MD on call 
Pager 206-314-3907 
Biostatistician 
Ted Gooley, PhD 
Professor, FHCRC 
206-667-6533
Research Staff 
Cari Morin  
  Study Coordinator 
206-667-5901
Susan Hildebrandt, RN 
Research Nurse 
206-667-7786
Emergency (24 hour) Phone:  206-598-8902 
IRO REC'D AUG 30 2021
FHCRC IRB Approval  
SEP 16 2021 
Document Released Date

2 
TABLE OF CONTENTS 
  PAGE 
1. TITLE OF PROTOCOL…………..………………………………............................1 
LIST OF FIGURES 
  4 
LIST OF TABLES 
  4 
2. INTRODUCTION……………………………………………………………….........5
3.  BACKGROUND……………………...…………………………………………….....5
3A. ROR1 expression in cancer and normal tissues 
  5 
3B. ROR1 Expressing Hematologic Malignancies 
  6 
3B.1. Chronic Lymphocytic Leukemia 
3B.2. Mantle Cell Lymphoma 
3B.3. Acute Lymphoblastic Leukemia 
3C. ROR1 Expressing Solid Tumors 
  7 
3C.1. Non-Small Cell Lung Cancer 
3C.2. Breast Cancer  
3D. Adoptive T cell therapy for human malignancies with unmodified T cells  
  10 
3E. Transfer of tumor targeting receptors into T cells to confer tumor-specificity 
  10 
3F. Clinical experience with the adoptive transfer of CAR-T cells 
  11 
3F.1. First generation CAR-T cells 
3F.2. CD19 CAR-T cells with 4-1BB/CD3ζ or CD28/CD3ζ signaling domains 
3G. Development and preclinical validation of ROR1-specific CARs 
  12 
3G.1. Development of ROR1 CAR 
  12 
3G.2. In vivo safety of ROR1 CAR-T cells in non-human primates  
  12 
3H. Safety considerations for adoptive therapy with genetically modified T cells 
  13 
3I.  Overview of the study  
  14 
4. OBJECTIVES…………………………………………………………………………14
4A. Primary  
  14 
4B. Secondary 
  14 
5. PATIENT SELECTION…………………………………………………………..….14 
5A. Inclusions/Exclusions for patients with CLL, MCL or ALL (Cohort A) 
 14 
5B. Inclusions/Exclusions for Patients with NSCLC or Breast Cancer (Cohort B) 
 15 
6. EVALUATION AND COUNSELING OF PATIENT……………………….……...19 
7. PROTOCOL REGISTRATION………………………………………………..….…19 
8. STUDY AGENT…………………………………………………………..……..….…19 
9. PLAN OF TREATMENT………………………………………………………….…20 
9A. Leukapheresis or blood-draw to obtain T cells for lentiviral transduction 
  20 
9B. Cytoreductive Chemotherapy  
  20 
9C. Clinical and/or Laboratory Exclusion Criteria for T Cell Infusions  
  20 
9D. Infusions of ROR1 CAR-T cells 
  21 

 
 
 
 
3 
 
10. MANAGEMENT OF POTENTIAL TOXICITIES AND COMPLICATIONS….23 
 
10A. Management of toxicity associated with the T cell infusion  
 
 
   23 
10B. Cytokine release                                                                                                           24 
10C. Management of tumor lysis syndrome 
24 
 
10D. Transgene Immunogenicity 
24 
 
10E. Management of other toxicities 
25 
 
 
11. SCHEDULE OF PATIENT EVALUATIONS……………………………………...26 
 
11A. Evaluation for Leukapheresis and Study Enrollment 
26 
 
11B. Disease Assessment 
26 
11C. Evaluations on the Day of Each T cell Infusion 
27 
11D. Patient Evaluations After Each T cell Infusion 
27 
 
12. PROTOCOL ENROLLMENT……………………………………………………….33 
 
 
  
13. RECORDS……………………………………………………………………………..33 
 
 
 
14. EVALUATION AND STATISTICAL CONSIDERATIONS………...………….....34  
14A. Type of study 
34 
14B. Definition of endpoints 
34  
14C. Endpoint evaluation 
34  
 
  15. GUIDELINES FOR REPORTING ADVERSE EVENTS AND DATA SAFETY   
  AND MONITORING PLAN…………………………………………………………......35 
 
15A. General issues and IRB reporting requirements 
 
 
 
 
    35  
15B. Data safety monitoring plan  
 
 
 
 
 
 
    37 
 
 
 
 
 
 
16. TERMINATION OF THE STUDY…………………………………………………..37  
 
17. REFERENCES…………………………………………………………………………38 
 
 
 
18. APPENDICES…………………………………………………………………………48  
18A. Suggested cytoreduction/lymphodepletion chemotherapy regimens 
 
    48 
18B. Karnofsky and ECOG Performance Score   
        
 
 
                49  
18C. Response Criteria for CLL, MCL, ALL, NSCLC and TNBC  
 
 
    50 
18D. Patient Evaluations  
 
 
 
 
 
 
                53 
18E.Research Sample Checklist 
 
 
 
 
 
 
 
    55 
18F. Long Term Follow-up 
 
 
 
 
 
 
 
    59 
18G. Grading Criteria for CRS 
 
 
 
 
 
 
 
    60 
18H. Management of CRS  
 
 
 
 
 
 
 
    61 
 
 
 
 
 
 
 

 
 
 
 
4 
 
LIST OF FIGURES           
   PAGE 
 
Figure 1. Study Treatment Plan ……………………………………………………….20 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
LIST OF TABLES           
 
 
 
 
 
 
       PAGE 
 
Table 1: Targeted / Planned Enrollment……………………………………………….33 
  
Table 2: FHCRC IRB Policies for Reportable Events……………………………..….37 
 
Table 3: FHCRC IRB Forms for Reporting……………………………………..…….37 

 
 
 
 
5 
 
2. 
INTRODUCTION 
 
The introduction of tumor-targeting receptors into T cells by gene transfer allows the rapid 
generation of tumor-specific T cells from any cancer patient for adoptive T cell therapy of their 
malignancy. A promising strategy involves engineering T cells with synthetic chimeric antigen 
receptors (CARs) that are comprised of a single chain antibody that is specific for a tumor cell-
surface molecule linked to one or more T cell signaling molecules.1-4 Recent trials, including 
FHCRC Protocol 2639 in which CAR-modified T cells (CAR-T cells) specific for the CD19 
molecule are being administered to patients with B-cell malignancies, have demonstrated marked 
tumor regression in a subset of patients with advanced acute lymphoblastic leukemia (ALL) and 
non-Hodgkin’s lymphoma (NHL).5 A subset of patients with B cell malignancies recur after CD19 
CAR-T cell therapy with tumor variants that lack the CD19 molecule6,7 and it may be necessary to 
target additional molecules to achieve maximal efficacy. In addition, it would be ideal to extend 
CAR-T cell therapy to common epithelial cancers. However this poses several challenges, 
including the identification of molecules expressed on tumor cells that can be targeted safely with 
CAR-T cells.  
 
We and others have investigated the tyrosine kinase orphan receptor ROR1 as a novel candidate 
target for CAR-T cells in patients with chronic lymphocytic leukemia (CLL), mantle cell 
lymphoma (MCL) and several epithelial cancers.8-15 We performed preclinical studies in vitro and 
in vivo in immunodeficient mice bearing human tumor xenografts that demonstrate ROR1 CAR-T 
cells have antitumor activity16. We have demonstrated that large doses of autologous ROR1 CAR-
T cells can be administered to non-human primates without toxicity,17 and have developed a 
clinical grade lentiviral vector that encodes a ROR1 CAR and can be used to transduce human T 
cells to re-direct their specificity to ROR1+ malignancies. This phase I clinical trial will evaluate 
the safety and antitumor activity of adoptively transferred autologous T cells genetically modified 
to express a CAR that targets ROR1. Two cohorts of patients will be eligible for treatment: Cohort 
A will enroll patients with advanced ROR1+ chronic lymphocytic leukemia (CLL), mantle cell 
lymphoma (MCL), or acute lymphoblastic leukemia (ALL); and Cohort B will enroll patients with 
ROR1+ non-small cell lung cancer (NSCLC) and triple negative breast cancer (TNBC). 
 
3. 
BACKGROUND  
 
3A. ROR1 expression in cancer and normal tissues 
 
The orphan tyrosine kinase receptor gene, ROR1, encodes two putative isoforms - a short 393 
amino acid (aa) intracellular protein and a long 937 aa type-1 transmembrane protein (isoform 1). 
ROR1 isoform 1 is expressed on many tissues during embryonic development, but absent in most 
adult tissues except a stage of immature B-cells in the bone marrow (BM) and low levels of 
expression in adipocytes, pancreas, and lung.16,18 We developed a monoclonal antibody that stains 
ROR1 in tissue sections and demonstrated that ROR1 was absent in vital tissues including brain, 
heart, lung and liver. However, we did observe some ROR1 expression in parathyroid, pancreatic 
islets cells, fat cells, and epithelial cells in the esophagus, stomach, duodenum, and colon. ROR1 is 
uniformly expressed on the surface of CLL, MCL, a subset of ALL, and on several common 
epithelial cancers including non-small cell lung cancer (NSCLC) and breast cancer.8-16,19-21 The 
role of ROR1 in tumor cells is not completely defined; however knockdown of ROR1 expression 
in leukemia, lung, and breast cancer cell lines causes growth arrest and/or apoptosis.9,13,20,22,23 
Analyses of outcomes in breast and ovarian cancer patients also suggest that high levels of ROR1 

 
 
 
 
6 
 
expression are associated with a poor prognosis.22,23 Together, these data suggest that targeting 
ROR1 with small molecules, monoclonal antibodies, or CAR-T cells may have therapeutic activity 
against ROR1+ malignancies. Our group has focused on the development of CAR-T cells to target 
ROR1 (see Section 3G). 
3B. ROR1 Expressing Hematologic Malignancies 
 
3B.1. Chronic Lymphocytic Leukemia  
CLL is the most prevalent adult leukemia and patients may have an indolent or aggressive clinical 
course. Asymptomatic patients without a large disease burden at diagnosis are usually observed 
until their disease progresses and requires therapy. Patients with high risk features including non-
mutated immunoglobulin genes, expression of ZAP-70, CD38, and/or adverse genetic changes 
(11q23 or 17p-) detected by cytogenetics or FISH often progress more rapidly to symptomatic 
disease and require treatment to control disease burden and reduce symptoms.24,25 Unfortunately, 
chemotherapy for CLL is not curative and patients remain at a continuous risk of relapse.  The 
prognosis for patients relapsing after first line therapy is variable but largely dependent on the 
duration and depth of first remission and on the interim development of adverse genetic risk 
features such as 17p deletion.  For patients with long remissions (> 2 years), the initial treatment 
can often be repeated.  However, the response rate, depth and duration of remission are usually 
shorter with subsequent therapy. 
 
For patients requiring antitumor therapy, the addition of monoclonal antibody therapy to 
combination chemotherapy has become standard of care.  For medically fit patients less than 65 
years of age, the CLL8 trial by the German CLL group demonstrated higher complete remission 
(CR) rates, longer progression free survival (PFS) and overall survival (OS) for patients 
randomized to fludarabine, cyclophosphamide, rituximab (FCR) chemotherapy compared to FC 
chemotherapy alone .26  In this trial, lower levels of minimal residual disease (MRD) as detected 
by flow cytometry 2 months after the completion of therapy were correlated with improved PFS 
and OS.27 Other approaches include combinations of fludarabine and rituximab or bendamustine 
and rituximab and are often used in older patients or those with comorbid medical conditions.  
 
Patients beyond first or second relapse are in need of novel treatments. New agents targeting the B 
cell Ig receptor signaling pathway including the PI3kinase-delta inhibitor (idelalisib) and the 
Brutons Tyrosine kinase (BTK) inhibitor (ibrutinib) are now approved for patients with CLL and 
improve survival, but are not curative.28,29 Allogeneic HCT using HLA matched related or 
unrelated donors following non-myeloablative conditioning may provide a curative option, 
however there is significant risk of non-relapse mortality of approximately 25% at 1 year due to 
GVHD and/or infections.30  
 
CLL cells nearly universally express both CD19 and ROR1.10,31 Adoptive T cell therapy with 
autologous CD19 CAR-T cells has induced complete remissions in up to 30% of patients with 
refractory CLL. Targeting CD19 can be associated with depletion of normal mature B cells, which 
express CD19, but do not express ROR1.32-35  Moreover, some patients treated with murine 
antibody-based CAR-T cells can make immune responses against antibody domains, as seen with 
CAIX-CAR-T cells in patients with metastatic renal cell carcinoma36 and in a subset of patients 
treated with our anti-CD19 CAR-T cell for patients with B-cell malignancies.5 The R12 ROR1 
scFv is from a rabbit antibody and is unlikely to be cross-reactive with the murine scFv used to 
construct the CD19 CAR. Thus, ROR1 CAR-T cells could provide a novel treatment option for 

 
 
 
 
7 
 
patients with CLL that is refractory to conventional therapy or CD19 CAR-T cells, and would not 
be expected to eliminate normal mature CD19+ B cells.   
 
3B.2. Mantle Cell Lymphoma  
MCL is a chronic relapsing NHL with an aggressive clinical course. Current standard of care 
includes induction with regimens such as R-CHOP or HyperCVAD/Mtx/Ara-C.37  Patients with 
responsive disease are considered for high-dose therapy and autologous HCT as consolidation in 
first remission.  A study by the German low-grade lymphoma group demonstrated improved PFS 
and OS with autologous HCT in first remission compared with interferon maintenance therapy.38 
Even with the use of autologous HCT, patients appear to remain at risk of relapse with no evidence 
of a plateau on the PFS curve. Patients who relapse following autologous HCT, and those unable 
to receive autologous HCT, have a poor outcome and are considered for experimental approaches 
and for allogeneic HCT. Similar to CLL, Bruton’s tyrosine kinase and PI3 kinase inhibitors have 
activity in MCL and are useful agents, however not all patients respond to treatment and relapses 
occur.39,40  
 
Allogeneic HCT appears to provide powerful graft-versus-tumor effect against MCL and several 
studies have demonstrated high CR rates and a low rate of relapse.41 This suggests MCL is 
susceptible to T cell recognition however as discussed above, the considerable risk of morbidity 
and mortality from GVHD and infection makes proceeding to an allogeneic HCT a difficult 
decision for many patients.  Patients unable to find a donor or unwilling to accept the risk of 
allogeneic HCT are usually treated with single agent or combination chemotherapy with palliative 
intent. MCL uniformly expresses ROR1, suggesting that as with CLL, treatment with ROR1 CAR-
T cells offers a novel approach to patients with disease that is refractory to conventional therapy.  
 
3B.3. Acute Lymphoblastic Leukemia (ALL)  
ROR1 is expressed on some cases of B-cell ALL,9,11,42-44 although the uniformity of expression 
was not extensively characterized in these studies. Cell surface ROR1 expression was found in 
45% of pediatric B-ALL cases and was not limited to any particular genotype,11 The subset of 
adult and pediatric ALL cases with the t(1:19) chromosome translocation (encoding the oncogenic 
E2A-PBX1 fusion protein) exhibit uniform surface expression of ROR1 on leukemia blasts and 
silencing of ROR1 expression in t(1:19) leukemia cell lines increases their sensitivity to kinase 
inhibitors.9 Thus, ROR1 could serve as a target for CAR-T cells in ALL patients that are ROR1+, 
refractory to conventional therapies, and ineligible for or unresponsive to CD19 CAR-T cells or 
blinatumomab therapy.  
  
3C. ROR1 Expressing Solid Tumors  
 
Transcriptional profiling studies have demonstrated ROR1 expression by many different epithelial 
cancers, and linked expression of ROR1 to a poor prognosis.22,23  ROR1 expression at the cell 
surface has been confirmed in lung and breast cancer cell lines using flow cytometry, however 
protein expression in primary tumors in formalin fixed paraffin embedded (FFPE) sections has 
been difficult to demonstrate because of the inconsistent behavior of currently available anti-ROR1 
mAb that target the extracellular domain in immunohistochemistry (IHC). Our studies suggest that 
formalin fixation destroys the ROR1 epitope(s) targeted by these mAb and that routine antigen 
retrieval methods are insufficient for reproducible staining in IHC. We have developed new 
ROR1-specific mAb that target epitopes that are present in the intracellular domain of isoform 1 
and these mAb are effective for detecting ROR1 protein in FFPE specimens. We will use this 

 
 
 
 
8 
 
reagent to determine ROR1 positivity in solid tumors. The initial trial of ROR1 CAR-T cells will 
focus on subtypes of advanced NSCLC and breast cancer because of the data indicating these 
tumors are frequently positive for ROR1 expression, and the clear need for novel therapies for 
these malignancies.13,14,20,22  
 
3C.1 Non-small cell lung cancer (NSCLC)  
NSCLC is the leading cause of cancer deaths in the United States and worldwide and often 
diagnosed at an advanced stage. Histologically, NSCLC is divided into adenocarcinoma, squamous 
cell carcinoma (SCC), and large cell carcinoma. Platinum-based doublet chemotherapy regimens 
are standard for first-line treatment of advanced stage NSCLC, but have significant side effects and 
provide only a modest overall survival benefit (range 2 to 4 months) compared to best supportive 
care.45 Using conventional chemotherapy as the only treatment, survival beyond 2 years occurs in 
less than 20% of cases.46 Although many patients with Stage IIIA disease are candidates for 
curative-intent therapy, palliation is the common goal for patients who have extensive mediastinal 
tumor burdens.  
 
Driver mutations in the epidermal growth factor receptor (EGFR) and anaplastic lymphoma kinase 
(ALK) are identified in a subset of lung adenocarcinomas. In patients with EGFR abnormalities, 
response rates of up to 70% can be achieved with tyrosine-kinase inhibitors (TKIs), erlotinib or 
gefitinib.47-49 Other EGFR mutations are associated with lower response rates and acquired 
resistance to these TKIs.50 Even responding patients eventually develop resistance, resulting in 
short-lived responses measured in months.51,52 Additionally, although EGFR mutations are 
common in NSCLC cases in Asian populations (up to 40%), their incidence in Caucasian 
populations is only 5% to 15%.53,54 Furthermore, K-RAS mutations which are present in 10% and 
30% of Asian and Caucasian subjects respectively, act downstream of EGFR and tend to be 
resistant to erlotinib and gefitinib.55,56 
 
ALK mutations are observed in only about 4% of unselected NSCLC cases,57-59 and the therapeutic 
efficacy of ALK inhibitors such as crizotinib and ceritinib has been disappointing.60 Thus, despite 
targeted therapy advances, treatments that safely deliver durable responses are urgently needed.  
 
Early clinical trials using immune checkpoint blocking agents, including anti-PD-1 (nivolumab, 
BMS-936558) and anti-PD-L1 (BMS-936559) antibodies, have produced responses in a significant 
fraction of patients with NSCLC.61-64 Other anti-PD-1 and anti-PD-L1 agents (MK-3475, 
MPDL3280A, MEDI4736) have also produced responses63-65. Although most patients have not 
responded to monotherapy with these agents, durable responses have been observed, supporting 
approaches to achieve more robust (or less inhibited) T cell mediated anti-tumor immunity, 
including strategies that directly target tumor-associated antigens with T cells. ROR1 is expressed 
on a subset of lung adenocarcinomas, where its expression may be induced by the NKX2-1 
oncogene.13 ROR1 knockdown in lung adenocarcinoma cell lines inhibited cell proliferation 
irrespective of EGFR status,66 and cells that were resistant to the EGFR tyrosine kinase inhibitor, 
gefitinib. Thus, treatment with ROR1 CAR-T cells offers a novel therapeutic approach.   
 
3C.2. Breast Cancer  
Breast cancer is composed of different subtypes typically classified based on the expression of 
estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 
(HER2). Triple negative breast cancer (TNBC) is the clinical term used to describe breast 
carcinomas that do not express the estrogen receptor (ER), progesterone receptor (PR), or human 

 
 
 
 
9 
 
epidermal growth factor receptor type 2 (HER2 or c-ERB2).  TNBC comprises approximately 10-
17% of all breast cancers.67-69   Despite efforts of many investigators in recent years, there remain 
no well-defined therapeutic targets for TNBC and chemotherapy remains the standard of care in 
the early and advanced disease settings. As a group, TNBCs are associated with a poor prognosis, 
characterized by early relapse70,71 and patients with TNBC often experience a significantly shorter 
survival following recurrence compared with patients with other breast cancer subtypes.67 The 
triple negative phenotype is an independent predictor of distant metastasis and cause specific 
survival and remains an area of urgent, unmet need in breast cancer.   
 
Recent clinical studies suggest that immunotherapy has the potential to improve clinical outcomes 
for patients with breast cancer. Genomic profiling has identified an immuno-modularity subtype of 
TNBC characterized by elevated expression of genes involved in T cell function, immune 
transcription, and antigen processing.72 Gene expression profiling of breast cancer stroma has also 
revealed that cytotoxic CD8+T cell infiltration and the resulting immune gene signature are 
predictive of good prognosis.73-75  On a histologic level, tumor infiltration with CD8+ T cells are 
generally associated with better prognosis while CD4+ T cells, which include T-regulatory cells, 
have been associated with worse outcomes.76  Accumulating evidence suggests that basal TNBC 
may be the most regulated by intratumoral T cells and perhaps most response to 
immunotherapeutics.77  
 
The immune checkpoint molecule, PD-L1, is highly expressed in ~20% of TNBCs, in association 
with decreased T cell proliferation and increased apoptosis,78 supporting anti-PD-L1/anti-PD1 
therapy in this patient population. In the KEYNOTE-012 phase Ib trial, pembrolizumab (MK-
3475) resulted in a notable overall response rate of 18.5% in a heavily pretreated TNBC population 
with an acceptable safety profile (Nanda R et al., A phase Ib study of pembrolizumab (MK-3475) 
in patients with advanced triple negative breast cancer. SABCS 2014, S1-09) and is being 
evaluated in additional studies with chemotherapy in the metastatic and neoadjuvant settings.  
Vaccine strategies are being investigated79-81 as are adoptive T cell therapies, including a 
mesothelin-targeting T cells.82 CAR-T cells targeting cMet or carcinoembryonic antigen (CEA) 
have entered Phase 1 trials as treatments for patients with advanced breast cancers, including 
TNBC (NCT01837602; NCT00673829), however no data has been reported on these trials.  
 
ROR1 is expressed in a subset of breast cancers and high ROR1 expression is associated with 
higher grade and more aggressive breast cancer types, including TNBC.14,22 High levels of ROR1 
in patient samples and cell lines are associated with expression of genes involved in epithelial-
mesenchymal transition (EMT); silencing of ROR1 in TNBC-derived cell lines reduced EMT gene 
expression and inhibited in vitro cell migration and bone and lung foci size in xenografts22. High 
ROR1 levels in primary breast tumors predicted for shorter metastasis free survival, independent 
of ER and HER2 status.  Together, these findings suggest a role of ROR1 signaling in the 
progression of breast cancer and support ROR1 as a novel therapeutic target, especially for TNBC 
where therapeutic options are limited.  
 
3D. Adoptive T cell therapy for human malignancies with unmodified T cells 
The potential for adoptively transferred T cells to eradicate human malignancies was first 
suggested by the graft-versus-leukemia effect of allogeneic T cells administered as part of a 
hematopoietic stem cell transplant (HCT).83,84 The adoptive transfer of in vitro expanded, 
autologous T cells derived from melanoma tumor infiltrates (TIL) and administered after 
lymphodepleting chemotherapy has induced complete or partial remissions in up to 72% of 

 
 
 
 
10 
 
patients with metastatic melanoma.85 Adoptive transfer of T cells can be associated with toxicity, 
related to immune recognition of normal cells that express the antigen(s) targeted by the infused T 
cells. For example, patients that receive allogeneic HCT or donor lymphocyte infusions often 
develop graft-versus-host disease as a consequence of donor T cell recognition of epithelial cells 
that express minor histocompatibility antigens86 and melanoma patients treated with TIL may 
develop vitiligo due to recognition of normal melanocytes.85 Despite these complications, T cell 
therapy, including HCT, has cured a subset of patients with advanced malignancy that was 
resistant to conventional chemotherapy and/or radiotherapy.85,86  
 
The clinical trials at the National Cancer Institute demonstrated that administering lympho-
depleting chemotherapy comprised of fludarabine and cyclophosphamide, or fludarabine, 
cyclophosphamide and total body irradiation, prior to the transfer of polyclonal melanoma-specific 
T cells dramatically improved both the survival of the transferred T cells and therapeutic efficacy 
compared to previous studies where lymphodepleting chemotherapy was not administered before  
cell transfer.85,87,88 Studies in murine models subsequently confirmed the human data that lympho-
depletion improves the persistence and efficacy of transferred T cells.89 Lymphodepletion may 
improve the persistence of transferred T cells by several mechanisms including reducing 
competition for cytokines such as IL-15 and IL-7 that promote lymphocyte proliferation and 
survival, eliminating CD4+ CD25+ regulatory T cells, and activating antigen presenting cells that 
may promote the function of transferred T cells. The use of lymphodepletion prior to T cell 
transfer has now become a standard in adoptive T cell therapy studies, including those using CAR-
T cells. 
 
3E. Transfer of tumor targeting receptors into T cells to confer tumor-specificity 
An obstacle for applying T cell therapy more broadly is the difficulty isolating tumor-reactive T 
cells from patients with cancer. An approach to overcome the low frequency of tumor-reactive T 
cells in patients is to redirect the specificity of T cells by expressing a transgene that encodes a 
receptor specific for a tumor-associated antigen (TAA). Vector systems to deliver transgenes into 
primary human T cells have been developed and clinical trials in which autologous T cells are 
modified to express a tumor-reactive T cell receptor (TCR) have shown transient clinical 
responses.90,91 However, redirecting T cells to recognize tumor antigens through TCR gene transfer 
is inherently constrained because of the requirement for major histocompatibility complex (MHC) 
restricted peptide presentation by tumor cells, and because many tumors express low levels of 
MHC molecules to avoid T cell recognition.92  
 
An alternative method for re-directing T cell recognition to tumor cells is to express an artificial 
chimeric non-MHC restricted antigen receptor (CAR) that recognizes a tumor cell surface 
molecule.1-4 A CAR is typically comprised of a fusion gene that encodes a monoclonal antibody-
derived single chain variable fragment (scFv), consisting of heavy (VH) and light (VL) chains 
joined by a flexible linker, fused through a transmembrane domain to a cytoplasmic signaling 
moiety consisting of CD3ζ alone, or CD3ζ combined with activation domains from costimulatory 
molecules such as CD28, 4-1BB or OX40.93,94 CARs with specificity for tumor cell-surface 
epitopes are “universal” in that they bind antigen in an HLA independent fashion, and one receptor 
construct can be used to treat all patients with tumors that express the molecule targeted by the 
CAR. T cells obtained from the blood of cancer patients can be modified with CARs to generate 
anti-tumor effector cells (CAR-T cells) for adoptive therapy. CAR-T cells can recognize tumor 
cells that have down-regulated HLA molecules, thereby avoiding the need to isolate rare HLA-
restricted tumor-reactive T cells. CARs have been constructed for many tumor-associated cell 

 
 
 
 
11 
 
surface molecules, including CD19, CD20, EGFR, HER2neu, GD2, PSMA, mesothelin, CAIX and 
ROR1.16,93-97 
3F. Clinical experience with the adoptive transfer of CAR-T cells 
 
3F.1. First generation CAR-T cells  
The initial human trials of CAR-modified T cells employed first generation constructs that linked 
the scFv to the CD3ζ or FcR gamma epsilon chain as the only intracellular signaling domain. T 
cells engineered with first generation CARs specific for CAIX were administered to patients with 
renal cell cancer, and caused elevations in liver enzymes, possibly due to expression of CAIX on 
bile duct epithelium.98  T cells specific for CD171 (L1CAM) were administered to patients with 
neuroblastoma without toxicity.99 In each of these studies, T cell persistence was poor and 
sustained antitumor effects were not observed.  
 
Patients with advanced B-cell malignancies have also been treated with first generation CARs that 
were specific for CD20 or CD19. These patients experienced little or no toxicity, but the CAR-T 
cells persisted poorly and sustained antitumor efficacy was not observed.90,100 The suboptimal 
results in these initial trials may reflect several factors including the absence of signaling domains 
to provide costimulation necessary for sustained T cell proliferation and survival in first generation 
CARs; the methods used to engineer and expand T cells for adoptive transfer that required long 
term culture and favored infusing terminally differentiated effector cells; and the variability in 
phenotype of the starting population, which can influence the capacity of T cells to persist and 
function long term in vivo.100-104  
 
3F.2. CD19 CAR-T cells with 4-1BB/CD3ζ or CD28/CD3ζ signaling domains 
To overcome the limitations identified in the initial trials and enhance potency, costimulatory 
endodomains were added to CD3ζ to enhance signaling through the CD19 CAR,32,33,105-107 and 
improved methods for selecting and transducing T cells of defined phenotype have been developed 
that do not require long-term culture.32,33 Potent antitumor activity has been observed in patients 
with ALL, CLL and NHL treated with autologous CD19 CAR-T cells on FHCRC Protocol #2639 
and in studies at the National Institutes of Health, University of Pennsylvania, and Memorial Sloan 
Kettering Cancer Center.32-34,108 In all of these studies, lymphodepleting chemotherapy was 
administered to enhance T cell persistence. In some cases, IL-2 was administered after the T cell 
infusion although the most impressive antitumor responses were observed in patients that did not 
receive IL-2.32  
 
The major toxicities of CD19-targeting CAR-T cells include cytokine release syndrome (CRS), 
neurologic toxicity, and tumor lysis that can occur early (days) or later (up to 2 weeks) after 
infusion of CD19 CAR-T cells. Some patients have persistent depletion of normal CD19+ B cells, 
necessitating intravenous immunoglobulin treatment. A relationship between T cell dose, toxicity, 
and efficacy has not been clearly evident in most trials. The inconsistency in the behavior of the 
therapeutic products may be because polyclonal T cells from the patient were transduced to 
express the CAR, and the resulting cell products administered to each patient varied greatly in 
phenotypic composition (CD4, CD8) and subset derivation. 
 
In the trial at the FHCRC, we have formulated the CAR-T cell products in a defined ratio of CD8 
and CD4 T cells based on preclinical data demonstrating that cell products containing an equal 
ratio of CD8 and CD4 T cells had superior antitumor activity compared to either subset alone. 
Although the trial is not yet completed, the administration of cell products of defined composition 

 
 
 
 
12 
 
to the first 35 patients appears to provide a clearer relationship between CAR-T cell dose and 
toxicity in patients with different B cell histologies. In ALL, severe CRS has been observed in 
patients with a high tumor burden and in this setting, a cell dose of 2x105/kg is now uniformly used 
to achieve high complete response rates. Patients with low tumor burdens will tolerate higher cell 
doses, generally without severe CRS. In NHL, severe CRS was observed at 2 x 107 cells/kg and 
2x106/kg selected for study even in patients with high tumor burdens. These studies demonstrate 
that CAR-T cells can have potent efficacy in advanced B cell malignancies and support the 
investigation of novel targets and extending this therapy to solid tumors. 
 
3G.  
Development and preclinical validation of ROR1-specific CARs 
 
3G.1. Development of ROR1 CAR 
We developed and optimized ROR1-specific CARs from the sequences of VH and VL chains of  
the 2A2 murine monoclonal antibody and from the R11 and R12 rabbit monoclonal 
antibodies.16,109 The 2A2 and R12 mAbs recognize an epitope in the Ig/Fz portion of the 
extracellular domain of ROR1, and R11 recognizes an epitope in the Kringle domain. T cells 
engineered by lentiviral transduction to express each of these ROR1 CARs optimized for spacer 
length specifically recognize ROR1+ tumor cells in vitro. A comparison of antitumor efficacy of 
2A2 and R12 CAR-T cells in immunodeficient mice engrafted with human tumor xenografts 
demonstrated that the R12 CAR was the most active.109 Lentiviral vectors encoding R12 CARs 
containing either 4-1BB/CD3ζ or CD28/CD3ζ were constructed and validated clinical lots of 
lentiviral supernatant have been prepared. The clinical lentiviral vector R12/S/4-1BBζ will be used 
in this clinical trial and encodes the truncated EGFR molecule (EGFRt) downstream of a T2A 
element in the vector such that T cells that express the ROR1 CAR will also express EGFRt. The 
clinical anti EGFR monoclonal antibody cetuximab binds to EGFRt and mediates antibody 
dependent cell mediated cytotoxicity (ADCC). Thus, cetuximab could be administered to patients 
to eliminate ROR1 CAR-T cells if toxicity occurred.   
 
3G.2. In vivo safety of ROR1 CAR-T cells in non-human primates 
The safety of targeting ROR1 with R12/4-1BB/CD3ζ CAR-T cells was evaluated in a non-human 
primate model to determine the potential for serious toxicity from recognition of rare normal tissue 
cells that might express ROR1. Human and macaca mulatta (M. mulatta) ROR1 are completely 
homologous in the Ig/Fz region that is recognized by the R12 scFv and we demonstrated that the 
tissue expression of ROR1 in macaques and humans is highly similar, including expression in 
parathyroid, pancreatic islets, esophagus, stomach, duodenum and colon 110. We adoptively 
transferred high doses (1-5x108/kg) of autologous macaque ROR1 CAR-T cells to 4 macaques 
without evidence of clinical toxicity or abnormalities in serum chemistry, or blood counts. ROR1 
CAR-T cells did not eliminate normal mature B cells in the peripheral blood. ROR1 CAR-T cells 
trafficked to bone marrow and lymph nodes, and were functional as shown both by the elimination 
of endogenous ROR1+ pre-B cells and by proliferation of CAR-T cells in response to challenging 
the animals with autologous T cells transfected to express ROR1. The T cell doses administered in 
this preclinical safety study are 2500-fold greater than the starting dose of CD19 CAR-T cells and 
25-fold greater than the highest dose of CD19 CAR-T cells administered to patients on Protocol 
2639.  
 
A limitation of the primate study was that long-term safety of ROR1 CAR-T cells could not be 
established because the animals were not immunosuppressed prior to CAR-T cell infusion and 
all animals developed a T cell immune response to the CAR-T cells after 3 to 4 weeks that 

 
 
 
 
13 
 
coincided with elimination of the CAR-T cells. We treated 2 additional macaques with ROR1 
CAR-T cells administered in cell doses of 1-5x108/kg following lymphodepleting chemotherapy 
to prolong CAR-T cell persistence. We did not observe any clinical toxicity and measurements of 
serum chemistry including calcium and glucose, and hepatic, pancreatic, and renal functions were 
normal. The CAR-T cells persisted for >40 days without any evidence of late toxicity.  
Transgene product-specific immune responses may occur in humans as well because the R12 scFv 
is derived from a rabbit antibody, and later immune rejection might serve as a safety mechanism to 
avoid long-term toxicity. The duration that CAR-T cells must persist to eliminate tumors is not yet 
established, and if targeting ROR1 shows short term safety in humans, future studies could use a 
human or humanized scFv specific for ROR1 in the design of the CAR combined with EGFRt to 
provide a safety mechanism. These preclinical findings support the clinical evaluation of ROR1 
CAR-T cells for the treatment of ROR1+ hematologic malignancies and solid tumors in carefully 
designed clinical trials 
 
3H.  
Safety considerations for adoptive therapy ROR1 CAR-T cells 
 
There are several potential toxicities of adoptive CAR-T cell therapy. The first relates to normal 
cell expression of the targeted molecule. In the case of targeting cells that express the CD19 
molecule, re-directed T cells do recognize and eliminate non-malignant CD19+ B-cells, potentially 
resulting in long-term B-cell immunodeficiency.32,34,106,111,112 In the case of ROR1 targeting in our 
non-human primate model, we observed no overt clinical toxicity even after transferring ROR1 
CAR-T cells doses equivalent to or exceeding those used in most human cancer immunotherapy 
trials, and observed no depletion of normal mature peripheral B cells (these cells do not express 
ROR1). We had previously shown low level of ROR1 on the surface of adipocytes differentiated 
from preadipocytes in vitro,16 and plasma adiponectin levels were transiently elevated in our 
macaque model of ROR1 CAR-T cell therapy. This could suggest a low-level of recognition of 
adipocytes, however analysis of necropsy specimens did not indicate accumulation of ROR1 CAR-
T cells in fat tissues and animals did not exhibit weight loss. It is possible that patients that receive 
ROR1 CAR-T cells may develop serious toxicity due to recognition of normal cells that express 
ROR1, including parathyroid, pancreatic islets, gastrointestinal tract epithelium, or other normal 
tissues.  
 
 
A second potential toxicity relates to the possibility of transformation of the gene-modified T cells 
as a consequence of insertional mutagenesis. The development of T cell leukemia was reported in 
a subset of patients on two gene therapy trials for X-linked severe combined immunodeficiency 
syndrome (SCID), in which bone marrow derived CD34 cells were transduced with a retroviral 
vector encoding the common cytokine receptor gamma chain 113-117. Animal studies have shown 
that mature T cells are resistant to transformation after retroviral integration118, and leukemia has 
never been observed in clinical trials involving gene transfer into mature T cells, despite more than 
10 years of follow-up in some studies.  Our study also differs from the X-linked SCID trial in 
several important aspects: 1) we will use a lentiviral vector rather than a retrovirus to genetically 
modify cells, which reduces the risk of integrating into a transcriptionally active site; 2) the target 
of the genetic modification will be mature T cells and not hematopoietic progenitor cells; and 3) 
the genetically modified ROR1-specific T cells will not constitutively express a functional growth 
factor receptor as was the case with transduced cells in the X-linked SCID trial.  
 

 
 
 
 
14 
 
A third potential toxicity is that ROR1 CAR-T cells might cause cytokine release after engagement 
of target cells that can result in fever, hypotension, and neurologic toxicity, or induce rapid tumor 
cell death resulting in a tumor lysis syndrome. Both cytokine release and tumor lysis syndromes 
have been observed in clinical trials of CD19 CAR-T cell therapy in patients, often those with high 
tumor burdens, and represent the most serious common complication of therapy.32,34  In the 
majority of patients the CRS is transient and subsides with tumor elimination, or if severe can be 
mitigated by administering corticosteroids or tocilizumab.119-122 
 
3I. Overview of the Study  
 
This is a Phase I/II, open-label, nonrandomized study that will evaluate the safety and potential 
antitumor activity of adoptively transferring autologous ROR1 CAR-T cells for patients with 
advanced ROR1+ B-cell malignancies (Cohort A) or advanced ROR1+ NSCLC and TNBC (Cohort 
B).  Patients with ROR1+ CLL, MCL or ALL that are refractory to conventional therapy will be 
eligible for Cohort A. Patients with ROR1+ NSCLC or TNBC who have failed conventional 
chemotherapy or targeted therapy, or for whom these therapies are not effective will be eligible for 
Cohort B.  
 
To provide for greater reproducibility of T cell products and to facilitate safety and efficacy 
evaluation, we will enrich CD8+ and CD4+ T cells separately from the leukapheresis product or 
blood of each patient, transduce each cell subset with the R12/S/4-1BBζ CAR lentivirus 
independently, expand and pool T cells in a 1:1 ratio to achieve the specified cell dose for each 
cohort of patients. The primary endpoint of the trial will be to determine the safety of ROR1 CAR-
T cells using a dose escalation/de-escalation design to define a cell dose that has acceptable 
toxicity, and might be used for a future larger phase II trial. Because toxicity may depend on the 
tumor type (hematologic/solid tumor), each cohort will be evaluated separately for toxicity.  
 
 
4. 
OBJECTIVES 
4A.  
Primary objectives 
To evaluate the safety of adoptive T cell therapy using ex vivo expanded autologous CD8+ and 
CD4+ ROR1 CAR-T cells for patients with advanced ROR1+ hematologic (Cohort A) and 
epithelial (Cohort B) malignancies. 
 
4B.  
Secondary objectives 
To determine:  
1. Duration of in vivo persistence of adoptively transferred T cells, and the phenotype of persisting 
T cells, 
2. Trafficking of adoptively transferred T cells traffic to the bone marrow or other tumor site and 
function in vivo, and  
3. Preliminary antitumor activity of the adoptive transfer of ROR1 CAR-T cells in patients with 
measurable tumor burden prior to T cell transfer  
 
 
 
 
 

 
 
 
 
15 
 
5. 
PATIENT SELECTION  
5A. 
Inclusion/Exclusion Criteria for patients with CLL, MCL or ALL (Cohort A)  
 
Inclusions 
1. Patients with  
a. CLL who are beyond first remission and who have failed combination 
chemoimmunotherapy with regimens containing a purine analogue and anti-CD20 
antibody, or who have failed tyrosine kinase or PI3 kinase inhibitors, or who were not 
eligible for or declined such therapy. Patients with fludarabine refractory disease are 
eligible.  
b. Mantle cell lymphoma patients who are beyond first remission and previously treated with 
chemoimmunotherapy. Patients who have relapsed following autologous HCT are eligible. 
c. ALL patients who have relapsed or have residual disease following treatment with curative 
intent. ALL patients must have ROR1 expressed on > 90% of the leukemia blasts to be 
eligible.  
 
2. Confirmation of diagnosis by internal pathology review of initial or subsequent biopsy or other 
pathologic material at the FHCRC/SCCA.  
3. Evidence of ROR1 expression by immunohistochemistry or flow cytometry on any prior or 
current tumor specimen. 
4. Male or female subject, greater than or equal to18 years of age. 
5. Karnofsky performance status ≥70% 
6. Negative pregnancy test for women of childbearing potential. Subjects of childbearing potential 
are those who have not been surgically sterilized or have not been free from menses for > 1 year.  
7. Fertile male and female patients must be willing to use a contraceptive method before, during, 
and for at least two months after the T cell infusion. 
8. Ability to understand and provide informed consent. 
 
Exclusions 
1. Treatment with other investigational agent(s) within 30 days of planned lymphodepletion. 
2. Patients requiring ongoing daily corticosteroid therapy at a dose of >15 mg of prednisone per 
day (or equivalent). Pulsed corticosteroid use for disease control is acceptable. 
3. Active autoimmune disease requiring immunosuppressive therapy. 
4. Major organ dysfunction defined as: 
a. Serum creatinine >2.5 mg/dL 
b. Significant hepatic dysfunction (SGOT >5x upper limit of normal; bilirubin >3.0 mg/dL) 
c. Patients with clinically significant pulmonary dysfunction, as determined by medical history 
and physical exam should undergo pulmonary function testing. Those with an FEV1 of < 50% 
or DLCO (corrected) <40% will be excluded. 
d. Significant cardiovascular abnormalities as defined by any one of the following: congestive 
heart failure, clinically significant hypotension, symptomatic coronary artery disease, or a 
documented ejection fraction of <45%. Any patient with an EF of 45-49% must receive 
clearance by a cardiologist to be eligible for the trial.  

 
 
 
 
16 
 
5. Patients who are HIV seropositive.  
6. Uncontrolled active infection (bacterial, viral, fungal, mycobacterial) not responding to 
treatment with intravenous antibiotics, antiviral or antifungal agents, or long-term treatment with 
oral agents.   
7. Women who are breast-feeding.  
8. 
Patients who have contraindication to cyclophosphamide chemotherapy. 
9. Known additional malignancy that is progressing or requires active treatment.  Exceptions 
include basal cell carcinoma of the skin, squamous cell carcinoma of the skin, or in situ cervical 
cancer that has undergone potentially curative therapy.   
10. Untreated central nervous system (CNS) metastases and/or carcinomatous meningitis.  Subjects 
with previously treated brain metastases may participate provided they are stable (without 
evidence of progression by imaging for at least four weeks prior to enrollment and any neurologic 
symptoms have returned to baseline), have no evidence of new or enlarging brain metastases. 
 
5.B Inclusion/Exclusion Criteria for Patients with NSCLC or TNBC (Cohort B) 
 
5B.1. NSCLC 
 
Inclusions 
1.  Patients with non-small cell lung cancer that is metastatic or inoperable and who have been 
treated with at least one line of prior therapy or declined conventional therapy.  
2.  Patients with known EGFR or ALK mutations must have been treated on at least one line of 
molecularly targeted therapy (e.g., erlotinib, crizotinib). 
3.  Patients must have measurable disease by at least one of the criteria below: (See Appendix C) 
a. Extra skeletal disease that can be accurately measured in at least one dimension as ≥10 mm 
with conventional CT techniques as defined by RESIST 1.1.   
b. Skeletal or bone-only disease measurable by FDG PET imaging.  
4.  ROR1 expression in >20% of the primary tumor or metastasis by IHC.  
5.  Karnofsky performance status of  ≥ 70%.  
6.  Patients must be off chemotherapy for a minimum of 3 weeks prior to start of treatment.   
Targeted therapies must be stopped at least 3 days prior to start of lymphodepletion. 
7.   Male or female subject, greater than or equal to18 years of age. 
8. Negative pregnancy test for women of childbearing potential. Subjects of childbearing 
potential are those who have not been surgically sterilized or have not been free from menses 
for > 1 year. 
9. Fertile male and female patients must be willing to use a contraceptive method before, during 
and for at least two months after the T cell infusion. 
 
10. Ability to understand and provide informed consent. 
 
 

 
 
 
 
17 
 
Exclusions 
1.   ANC < 1000/mm3 
2.   Hgb < 9 mg/dl (transfusion permitted to achieve this) 
3.   Platelet count < 75,000/mm3 
4.  Treatment with other investigational agent(s) within 30 days of planned lymphodepletion. 
5. Patients requiring ongoing daily corticosteroid therapy at a dose of > 15 mg of prednisone per 
day (or equivalent). Pulsed corticosteroid use for disease control is acceptable. 
6. Active autoimmune disease requiring immunosuppressive therapy. 
7. Major organ dysfunction defined as: 
a. Serum creatinine > 2.5 mg/dL 
b. Significant hepatic dysfunction (SGOT >5x upper limit of normal;      
bilirubin > 3.0 mg/dL) 
c. Patients with clinically significant pulmonary dysfunction, as determined by medical history 
and physical exam should undergo pulmonary function testing. Those with an FEV1 of < 50% 
or DLCO (corrected) < 40% will be excluded. 
d.  Significant cardiovascular abnormalities as defined by any one of the following: 
congestive heart failure, clinically significant hypotension, symptomatic coronary artery 
disease, or a documented ejection fraction of < 45%. Any patient with an EF of 45-49% 
must receive clearance by a cardiologist to be eligible for the trial.  
8. Patients who are HIV seropositive.  
9. Uncontrolled active infection (bacterial, viral, fungal, mycobacterial)  not responding to 
treatment with intravenous antibiotics, antiviral or antifungal agents, or long-term treatment 
with oral agents.   
10. Women who are breastfeeding. 
11. Patients who have contraindication to cyclophosphamide chemotherapy. 
12. Known additional malignancy that is progressing or requires active treatment.  Exceptions 
include basal cell carcinoma of the skin, squamous cell carcinoma of the skin, or in situ 
cervical cancer that has undergone potentially curative therapy.   
13.  Untreated central nervous system (CNS) metastases and/or carcinomatous meningitis.  
Subjects with previously treated brain metastases may participate provided they are stable 
(without evidence of progression by imaging for at least four weeks prior to enrollment and 
any neurologic symptoms have returned to baseline), have no evidence of new or enlarging 
brain metastases. 
 
5B.2. TNBC 
 
Inclusions 
1.  Histologically confirmed diagnosis of metastatic TNBC; i.e. breast cancer that is ER negative 
(≤ 10%), PR negative (≤ 10%), and HER2 negative (0 or 1+ by immunohistochemistry or negative 
for gene amplification by FISH).     

 
 
 
 
18 
 
2.  Patients must have measurable disease by at least one of the criteria below: (See Appendix C) 
a. Extra skeletal disease that can be accurately measured in at least one dimension as ≥ 10 mm 
with conventional CT techniques as defined by RESIST 1.1.   
b. Skeletal or bone-only disease measurable by FDG PET imaging. 
3. Patients must have received standard adjuvant, neoadjuvant, and/or metastatic chemotherapy 
per NCCN or institutional practice.  No maximum on number of prior systemic treatment 
regimens.  
4.  Patients may receive agents to protect against skeletal related complications such as zolendronic 
acid or denosumab.  
5.   ROR1 expression in > 20% of the primary tumor or metastasis by IHC.  
6.  Karnofsky performance status of ≥ 70%. 
7.  Patients must be off chemotherapy for a minimum of 3 weeks prior to planned leukapheresis. 
8.  Subjects must be ≥ 18 years old. 
9.  Negative pregnancy test for women of childbearing potential. Subjects of childbearing potential 
are those who have not been surgically sterilized or have not been free from menses for > 1 year.   
10.  Fertile male and female patients must be willing to use a contraceptive method before, during 
and for at least two months after the T cell infusion. 
 
11. Ability to understand and provide informed consent. 
Exclusions 
1.  ANC < 1000/mm3 
2.  Hgb < 9 mg/dl (transfusion permitted to achieve this) 
3.  Platelet count < 75,000/mm3 
4.  Treatment with other investigational agent(s) within 30 days of planned lymphodepletion. 
5.  Patients requiring ongoing daily corticosteroid therapy at a dose of > 15 mg of prednisone per 
day (or equivalent). Pulsed corticosteroid use for disease control is acceptable. 
6.  Active autoimmune disease requiring immunosuppressive therapy. 
7.  Major organ dysfunction defined as: 
a. Serum creatinine > 2.5 mg/dL 
b. Significant hepatic dysfunction (SGOT > 5X upper limit of normal; bilirubin > 3.0 mg/dL) 
c. Patients with clinically significant pulmonary dysfunction, as determined by medical history 
and physical exam should undergo pulmonary function testing. Those with an FEV1 of < 50% 
or DLCO (corrected) < 40% will be excluded. 
d. Significant cardiovascular abnormalities as defined by any one of the following: congestive 
heart failure, clinically significant hypotension, symptomatic coronary artery disease or a 
documented ejection fraction of < 45%. Any patient with an EF of 45-49% must receive 
clearance by a cardiologist to be eligible for the trial.   
8.  Patients who are HIV seropositive.  

 
 
 
 
19 
 
9. Uncontrolled active infection (bacterial, viral, fungal, mycobacterial) not responding to 
treatment with intravenous antibiotics, antiviral or antifungal agents, or long-term treatment 
with oral agents.   
10. Breast-feeding women. 
11. Patients who have contraindication to cyclophosphamide chemotherapy. 
12. Has a known additional malignancy that is progressing or requires active treatment.  
Exceptions include basal cell carcinoma of the skin, squamous cell carcinoma of the skin, or in situ 
cervical cancer that has undergone potentially curative therapy.   
13. Untreated central nervous system (CNS) metastases and/or carcinomatous meningitis.  Subjects 
with previously treated brain metastases may participate provided they are stable (without 
evidence of progression by imaging for at least four weeks prior to enrollment and any neurologic 
symptoms have returned to baseline) and have no evidence of new or enlarging brain metastases.   
 
6. 
EVALUATION AND COUNSELING OF PATIENT  
 
Patients will be seen at the Seattle Cancer Care Alliance or the Fred Hutchinson Cancer Research 
Center (FHCRC) for consideration of treatment options for their disease. The protocol will be 
discussed thoroughly with the patient and other family members if appropriate, and all known and 
potential risks to the patient will be described. The procedure and alternative forms of therapy will 
be presented as objectively as possible, and the risks and hazards of the procedure explained to the 
patient. Consent from the patient will be obtained using forms approved by the Institutional 
Review Board (IRB) of the FHCRC. A summary of the clinic visit detailing what was covered will 
be dictated for the medical record. 
 
7. 
PROTOCOL REGISTRATION  
 
Eligible subjects will be identified and registered into the system by the Clinical Coordinators 
Office (CCO) (Intake Office) and assigned a UPN (Unique Patient Number). The CCO will 
register the subject on to the protocol through the Data Management Office. A 
screening/leukapheresis consent must be signed prior to the performance of study related 
assessments or procedures. The T cell therapy treatment consent will be signed after successful 
collection of T cells and prior to chemotherapy and treatment with ROR1 CAR-T cells.  
 
8. 
STUDY AGENT 
 
8A. 
ROR1 CAR-T cells 
1. The methods employed to derive ROR1 CAR-T cells from the patient’s CD8+ and CD4+ T cells 
enriched from PBMC, and release tests of the cell products prior to infusion are outlined in the 
Chemistry, Manufacturing and Controls (CMC) section of the Investigational New Drug (IND) 
application to the FDA. Modifications to the CMC section during the course of the study must be 
submitted for FDA review. 

 
 
 
 
20 
 
2. The autologous T cell product will be prepared and administered to the patient by intravenous 
infusion. Patients will be pretreated with cytoreductive/lymphodepleting chemotherapy and receive 
the T cell infusions 36-96 hours after completing chemotherapy (Section 9.B).  
3. A schematic of the treatment plan is shown in Figure 1. 
  
 
 
 
 
 
 
 
9. 
PLAN OF TREATMENT  
 
9A. 
Leukapheresis or blood-draw to obtain T cells for ROR1 CAR-T cell manufacturing 
1. A leukapheresis will be performed on each patient in the Apheresis Unit at the SCCA using 
standard operating procedures for obtaining peripheral blood mononuclear cells. Should a 
technical issue arise during the procedure or in the processing of the product, or insufficient ROR1 
CAR-T cells be manufactured for the prescribed ROR1 CAR-T cell dose, a second procedure may 
be performed.  
Patients ineligible for a vein to vein apheresis may elect to have a percutaneous central venous 
access catheter inserted to support this collection. Patients ineligible for leukapheresis and who 
have a hematocrit of >38 and a total non-malignant (normal) lymphocyte count of >2000 may 
undergo phlebotomy of 400 ml of blood to obtain PBMCs necessary to establish the T cell 
cultures. This approach will only be taken in patients that would be enrolled at dose levels 1 and 2 
and if the absolute CD8 and CD4 T lymphocyte counts are deemed to be sufficient by the Clinical 
PI to obtain enough T cells for successful manufacturing. 
2. The leukapheresis or phlebotomy product will be delivered to the Cell Processing Facility 
(CPF) at the FHCRC or the Cell Therapy Laboratory (CTL) at the SCCA. CD8+ and CD4+ cell 
selections may be performed in the FHCRC CPF or SCCA CTL. PBMC not required for cell 
selections may be archived for research.  
If lymphocyte subset counts are considered adequate, the product will be divided into two aliquots. 
One aliquot will be enriched for CD8+ T cells and the second aliquot will be enriched for CD4+ T 
cells using clinical grade reagents and SOPs developed at the FHCRC/SCCA. Subsequent 
processing, after selection of CD4+ and CD8+ T cells and cryopreservation (if required), is 
performed in the FHCRC CPF. 
If processing of CD8+ and CD4+ T cells is not considered suitable due to lymphopenia, low CD8+ 
T and CD4+ cell counts or other reasons, we may use other strategies that are approved for use in 
this protocol by the FDA to manufacture ROR1 CAR-T cells without selection of subsets. 
3. Quality control and release testing will be performed on the ROR1 CAR-T cell product prior to 
its release for patient infusion.  
  

 
 
 
 
21 
 
4. Patients may receive chemotherapy after screening and leukapheresis to control disease during 
production of ROR1 CAR-T cells. Decisions regarding chemotherapy in this interval should be 
discussed with protocol Principal Investigator (PI). 
5. Patients will be scheduled for cytoreductive and/or lymphodepleting chemotherapy to be 
completed 36-96 hours prior to the first T cell infusion. 
 
9B. 
Cytoreductive/Lymphodepleting Chemotherapy 
1. Prior to the first infusion of ROR1 CAR-T cells, patients will receive chemotherapy as 
determined by the referring physician in consultation with the protocol Principal Investigator (PI). 
The objectives of administering chemotherapy are to provide lymphodepletion to facilitate T cell 
survival and to reduce the tumor burden prior to infusion of ROR1 CAR-T cells.  
2. Refer to Appendix A for suggested chemotherapy regimens based on disease cohort. 
3. There must be at least a 36-hour interval between the last dose of chemotherapy and the T cell 
infusion. 
9C.  
T Cell Infusions 
1. ROR1 CAR-T cells should be administered between 36 and 96 hours after the completion of 
lymphodepleting chemotherapy.  
2. On the day of scheduled T cell infusion the patients should undergo a clinical evaluation and a 
clinical determination for appropriateness to proceed with T cell administration.: 
3. Patients deemed not appropriate for T cell administration may receive a T cell infusion outside 
the 36-96 hour window if they subsequently resolve the clinical and/or laboratory concerns which 
deemed them inappropriate. 
4.  Patients who receive lymphodepleting chemotherapy, but who are unable to undergo CAR-T 
cell infusion within the intended 36-96 hour window due to CAR-T cell manufacturing or release 
issues may receive the cell infusion outside of this window if the issues can be resolved.  
Discussion with the protocol PI and IND holder is required. 
 
9D. 
ROR1 CAR-T cells dosing  
 
1. Cohort A (CLL, MCL and ALL) - Cell dose escalation/de-escalation   
The primary objective of this study is to estimate the maximum tolerated dose (MTD) of CAR-T 
cells. EGFRt is co-expressed in the CAR vector and serves as a transduction marker to determine 
the number of CAR-T cells in the cell product to define cell dose. The MTD for these purposes 
will be defined as a true dose limiting toxicity rate of 25%, where DLT is defined in Section 
14C.1. A modification of the continual reassessment method (CRM) will be used to estimate the 
MTD. The modifications include treating patients in groups of at least two (rather than one), and 
allowing a maximum increase of one dose level between groups. Patients will receive a single 
intravenous infusion of ROR1 CAR-T cells at one of four escalating dose levels beginning with 
dose level 1 for the first group of two patients. Dose escalation or de-escalation is determined by 
the CRM algorithm, taking into account the number of patients experiencing a serious 
toxicity at each dose level, as described in Section 14C.1.  
Dose level 0: 
 
up to 1x105 EGFRt+ cells/kg 
Dose level 1: 
 
up to 3.3x105 EGFRt+ cells/kg (Starting dose level)  
Dose level 2: 
 
up to 1x106 EGFRt+ cells/kg  

 
 
 
 
22 
 
Dose level 3: 
 
up to 3.3x106 EGFRt+ cells/kg 
Dose level 4:  
 
up to 1.0x107 EGFRt+ cells/kg  
Treatment of patients in each of the dose-escalation/de-escalation groups will be staggered with a 
minimum of a 21-day interval following infusion between each patient and 35 days before 
escalation to the next dose level. The CRM algorithm will be followed until up to 30 patients have 
been enrolled and treated, and the MTD will be estimated after the 30th patient is evaluable for 
DLT. Preliminary estimates of anti-tumor efficacy will be obtained among all patients and among 
those who were treated at the estimated MTD. 
 
2. Cohort B (NSCLC and TNBC) - Cell dose escalation/de-escalation 
The toxicity of ROR1 CAR-T cells resulting from on-target recognition of tumor cells could differ 
in epithelial malignancies due to the sites of primary and metastatic disease, differences in ROR1 
expression levels, and differences in the tumor microenviroment that could suppress transferred T 
cell immunity. Thus, we will perform the dose escalation/de-escalation of ROR1 CAR-T cells 
separately in patients in Cohort B. Our goal will be to have relatively balanced enrollment of 
TNBC and NSCLC in this cohort depending on patients meeting eligibility criteria. The primary 
objective for this cohort is to estimate the maximum tolerated dose (MTD) of ROR1 CAR-T cells, 
as above, with the MTD defined as the dose of T cells associated with a true DLT rate of 25%, 
where DLT is defined in Section 14C.1. Dose escalation or de-escalation is determined by the 
CRM algorithm, taking into account the number of patients experiencing a serious toxicity at 
each dose level, as described in Section 14C.1.  
Dose level 0: 
 
up to 1x105 EGFRt+ cells/kg 
Dose level 1: 
 
up to 3.3x105 EGFRt+ cells/kg (Starting dose level)  
Dose level 2: 
 
up to 1x106 EGFRt+ cells/kg  
Dose level 3: 
 
up to 3.3x106 EGFRt+ cells/kg 
Dose level 4:  
 
up to 1.0x107 EGFRt+ cells/kg  
Treatment of patients in the dose-escalation/de-escalation groups will be staggered with a 
minimum of a 21-day interval following infusion between each patient and 35 days before 
escalation to the next dose level.. The CRM algorithm will be followed until up to 30 patients have 
been enrolled and treated, and the MTD will be estimated after the 30th patient is evaluable for 
DLT. Preliminary estimates of anti-tumor efficacy will be obtained among all patients and among 
those who were treated at the estimated MTD. 
 
3. Cell administration. Individual aliquots of CD4+ and CD8+ ROR1 CAR-T cells will be prepared 
for each T cell infusion according to protocols established in the GMP Cell Processing Facility and 
after passing release criteria will be formulated to provide a single cell product at the specified cell 
dose. The specified T cell dose refers to CAR+ T cells determined by the expression of the 
truncated EGFR transduction marker, which is expressed coordinately with the CAR in the vector. 
Each T cell infusion should be administered intravenously over approximately 20 – 30 minutes at 
the specified T cell dose. At least the first patient treated at each dose level will receive the T cell 
infusion at the University of Washington Medical Center.  
 
4. Patient monitoring during T cell infusions. All patients will be monitored during each T cell 
infusion. Patients should receive the T cells under physician guidance and nursing supervision, 
with vital signs being monitored and recorded before, q 15 mins during and at the conclusion of 

 
 
 
 
23 
 
the infusion and then approximately hourly for 2 hours after the infusion. O2 saturation should be 
monitored continuously during the infusion and results documented before, q 15 mins during and 
at the conclusion of the infusion. O2 saturation should then be assessed and documented 
approximately hourly for 2 hours after the infusion 
 
5. Products that cannot be formulated to meet cell dose specification.  The intent for each infusion 
is to provide a cell product that reaches at least 85% of the targeted  cell dose and contains 
approximately 50% (+/- 15%) of ROR1 CAR-modified CD4+ T cells and 50% (+/- 15%) of ROR1 
CAR-modified CD8+ T cells (i.e. CAR-modified T cells in 1:1 CD4+/CD8+ ratio)  If a T cell 
product cannot be formulated to meet this cell dose specification because of low transduction 
efficiency, suboptimal growth of one of the subsets, or failure of either the CD4+ or CD8+ T cell 
product to meet release criteria, the cell product should be infused at or as close as possible to the 
specified phenotype allocation and total T cell dose providing: 
i. The total EGFRt+ CAR-T cell dose exceeds 5x104 cells/kg, 
ii. The cell product passes other release criteria including endotoxin, gram stain, sterility, 
mycoplasma, RCL and viability. 
iii.Toxicity and safety data will be collected and reported for subjects receiving such 
products. 
Such patients will not be considered evaluable for safety analysis in the assigned dose cohort and a 
replacement subject will be added to the cohort.  Toxicity and safety data will be collected and 
reported on such patients, but will not be incorporated into the CRM algorithm for the cell dose 
cohort under study. 
 
6. Retreatment of patients on the study. Patients enrolled in the study may be eligible to receive a 
second ROR1 CAR-T cell infusion with or without additional cytoreductive chemotherapy at the 
same (for those that received the highest cell dose) or up to the next highest dose level if adequate 
ROR1 CAR-T cells can be produced and the following criteria are met: 
a. There is evidence of persistent disease after the first T cell infusion.  
b. There were no toxicities attributed to the first infusion that were dose-limiting or 
required dose de-escalation (Section 14.C.1).  
c. The patient is ≥ 21 days from the first T cell infusion.  
d. There are no clinical and/or laboratory exclusion criteria (Section 9C.1). 
 
10. 
MANAGEMENT OF TOXICITIES AND COMPLICATIONS 
 
Acute infusional toxicity may occur during or shortly after T cell infusion. In addition, cytokine 
release syndrome, tumor lysis syndrome, and neurologic toxicity have been reported specifically 
after CD19 CAR-T cells and may occur after the infusion of ROR1 CAR-T cells. 
Recommendations for the management of these complications is addressed in 10A-10E.Clinical 
interventions should be provided in accordance with the treating clinician’s judgment and patient’s 
clinical conditions. A table of proposed grading criteria for cytokine release syndrome is given in 
Appendix G 
 
10A. Management of acute toxicity associated with T cell infusion 
 

 
 
 
 
24 
 
The results of our prior studies of adoptive immunotherapy for CMV, HIV, leukemia and 
lymphoma, and melanoma suggest that serious acute toxicities resulting simply from infusing the 
numbers of T cells proposed in this study are unlikely to occur.  However, fevers and other 
transient constitutional symptoms are often observed with T cell infusions, and can occur very 
shortly after the T cell infusion in patients with a high antigen (tumor) burden.  
 
Examples of potential symptoms/signs due to T cell infusions and their initial management are: 
• Fever, chills, and temperature elevations >101°F may be managed with acetaminophen 650 
mg p.o. q 4-6 hrs.  All subjects who develop fever or chills should have a blood culture 
drawn. 
• Headache may be managed with acetaminophen. 
• Nausea, vomiting may be managed with diphenhydramine 25-50 mg IV or other 
antiemetics (excluding corticosteroids). 
• Hypotension should be managed initially by fluid administration. 
• Hypoxemia should be managed initially with supplemental oxygen. 
 
If the following signs appear during T cell infusion, the infusion should be paused and the patient 
assessed:  
• Systolic BP < 80 mmHg or > 40 mmHg fall from baseline 
• Heart rate > 140/min or increase from baseline > 40/min (confirmed by palpation or EKG) 
• Respiratory rate > 35/min or increase from baseline of > 10/min 
 
 
 
• Arterial O2 saturation < 88% on room air  or fall from baseline > 5% 
 
If after assessment by the PI/designee the patient’s condition is stable then the infusion may be 
resumed.  
 
 
If a T cell infusion is terminated due to acute toxicity, the residual T cells should be returned to the 
Cell Processing Lab for analysis. Investigation of possible causes of observed signs should proceed 
and, if necessary, additional medical treatment will be instituted. 
 
Patients requiring discontinuation of T cell infusion may be eligible for re-treatment if the cause is 
deemed not related to the T cell infusion.  
 
10B.  Management of cytokine release syndrome  
 
1. If patients become febrile or develop symptoms of cytokine release, we may measure cytokine 
levels, serum ferritin, C-reactive protein and markers of tumor lysis syndrome (e.g. chemistry, uric 
acid, LDH), and evaluate persistence and phenotype of the transgene-expressing cells, as clinically 
indicated. 
 
2. Any patient who develops clinical evidence of symptoms related to cytokine release will have a 
work-up to exclude infection or other causes. Initial treatment should consist of supportive 
measures as dictated by the clinical and laboratory findings, and may include fluid replacement, 
medications to support blood pressure, antipyretics, oxygen supplementation, and broad-spectrum 

 
 
 
 
25 
 
antibiotics if infection cannot be excluded as a potential etiology for the signs and symptoms. 
Patients with Grade > 3 CRS (severe CRS; sCRS) and or Grade 2 CRS with progressive symptoms 
and signs should be treated with tocilizumab (4-8 mg/kg IV) and corticosteroids (dexamethasone 
10 mg IV every 12 hours). Higher doses of steroids may be given after discussion with the 
Principal Investigator or designee and repeated doses of tocilizumab may be given if necessary. 
See Appendix G for cytokine release syndrome grading and Appendix H for recommended 
management guidelines. 
 
10C. Management of tumor lysis syndrome 
 
1. All patients will be considered at risk for tumor lysis and should receive allopurinol prophylaxis 
before chemotherapy begins, unless contraindicated. Allopurinol should be continued for as long 
as the medical team determines appropriate after the T cell infusion. They may receive additional 
hydration and urine alkalinization for the first 2 weeks after the T cell infusion.  
 
2. If tumor lysis syndrome develops, as defined by the Cairo Bishop criteria 123, the Attending 
Physician will direct patient management with guidance from the study staff 124.  Conservative 
therapy, including allopurinol, urinary alkalinization, and IV fluid hydration may be instituted 
immediately for suspected tumor lysis syndrome. Hyperkalemia may be treated with potassium 
binding resins, diuresis, or insulin/dextrose therapy. Hyperphosphatemia may be treated with 
phosphate binding resins. In severe cases, rasburicase (in non-G6PD-deficient individuals) or renal 
dialysis may be necessary. 
 
 
10D. Management of neurotoxicity 
 
Neurotoxicity, manifest as delirium, seizures and/or focal neurologic deficits, has been 
reported after CD19 CAR-T cell therapy. Neurotoxicity that is attributed to the T cell infusion/s 
should be treated with corticosteroids (e.g. dexamethasone 10 mg IV q 4-12 hours). Tocilizumab 
may also be given, preferably after discussion with the Principal Investigator or designee. 
Patients with a history of seizures or history of CNS involvement or high tumor burden ALL 
should receive prophylactic treatment with anti convulsants (Keppra or similar) following CAR-
T cell infusion for approximately 4 weeks.  Patients who develop significant neurotoxicity 
should be also treated with anti-convulsants if not already started.  See Appendix H for 
recommended management guidelines. 
 
10E. Management of other toxicities 
 
1. If a new onset CTCAE v4.0 grade ≥ 3 toxicity is observed following any T cell infusion, the 
patients will receive investigation and medical treatment appropriate for the physiological 
abnormalities. 
 
2. Grade ≥ 3 toxicity that is attributed to the T cell infusion/s and is unresponsive to supportive 
measures or persists for > 7 days may be treated with corticosteroids (e.g. dexamethasone 10 mg 
IV q 4-12 hours) after discussion with the Principal Investigator or designee. 
 
3. Uncontrolled proliferation of CAR-T cells has not been observed in clinical trials to date. 
However, in the unlikely event uncontrolled proliferation of ROR1 CAR-T cells occurred in a 
study subject, initial therapy may involve treatment with corticosteroids (e.g. methylprednisolone 1 

 
 
 
 
26 
 
g IV). Anti-lymphocyte globulin or cytotoxic drugs would also be considered in serious cases. If 
we observe an increase in CAR positive cells to greater than 10% of T cells at more than 3 months 
after last infusion we will analyze for clonal expansion by deep sequencing of the TCR beta gene 
(ImmunoSeq Adaptive Biotechnology). 
 
4. The vector encodes EGFRt and our preclinical data in animal models demonstrates that 
administering two doses of Erbitux (cetuximab) can eliminate CAR-T cells that express EGFRt 
and reverse on-target toxicity not attributed to cytokine storm, for which there is no suitable 
animal model. Thus, in patients without cytokine storm who have an ongoing organ toxicity that 
is attributed to ROR1 CAR-T cells, we could consider administering up to two doses one week 
apart of intravenous cetuximab in standard dosing at 400 mg/m2  (dose 1) and 250 mg/m2 (dose 
2). Patients would be pre-medicated 30-60 minutes prior to each dose of cetuximab with 50 mg 
of diphenhydramine intravenously. Blood samples (30 ml in sodium heparin tubes) should be 
collected prior to cetuximab and on approximately days 1, 4, 7, 14, and 21 to determine if 
cetuximab resulted in a decline or elimination of ROR1 CAR-T cells.  
 
 
11. SCHEDULE OF PATIENT EVALUATIONS (Appendix D - Table of Evaluations). 
Please note that results of tests and/or procedures conducted as per standard of care purposes 
may be used for research purposes if conducted within the protocol-defined window prior to 
screening/leukapheresis (11.A.) and/or T-Cell Therapy (11.B.).  
 
 
11A. Evaluation for Leukapheresis (Completed within 15 days of leukapheresis procedure) 
1. Informed consent and HIPPA signing 
2. Laboratory evaluation, including: 
a. CBC, differential and platelet count 
b. Renal function panel and hepatic function panel with LDH 
c. Blood Works NW Recipient Donor Battery Panel 
d. Serum pregnancy test for females of child-bearing potential 
e. ABO blood typing and antibody screen 
f. Patient/donor CD3 chimerism, at any time point post transplant, if prior allogeneic 
transplant 
3. Medical history, including:  
a. Hematologic, cytogenetic, flow cytometric, and histologic findings at diagnosis and at the 
time of enrollment on the study 
 
b. Prior therapies and response to therapy if known 
4.  Confirmation of diagnosis by internal pathology review of initial or subsequent biopsy or other 
pathologic material at the FHCRC/SCCA. 
5.  Physical Exam and Karnofsky Performance Status 
 
6.  Research testing:  

 
 
 
 
27 
 
  a. Confirmation of ROR1 expression on tumor cells  
 
i Cohort A: Flow cytometry of tumor cells from blood, lymph node, or bone marrow 
specimen or IHC on a prior diagnostic lymph node or other site. 
 
ii. Cohort B: IHC on a prior diagnostic specimen or fresh biopsy specimen. 
  b. 10 ml blood sample, sodium heparin (green top tube) to Berger Lab for evaluation of CD3, 
CD4, and CD8 T cells subsets 
 
 
11.B  Evaluation prior to T cell therapy (to be performed if studies in 11A. 1-7 deem patient to be 
eligible and T cell collected) Should be completed within 30 days of enrollment unless otherwise 
specified 
1. Cohort A (CLL, MCL, and ALL) 
1. Laboratory evaluations 
a. CBC, differential and platelet count 
b. Renal function panel and hepatic function panel with LDH 
c. Uric acid 
d. Serum ferritin 
e. CRP  
f. DIC panel without platelets 
g. Parathyroid hormone level (PTH) 
h. G6PD screen 
i.  Serum pregnancy test for females of child bearing potential  
j. Quantitative immunoglobulins (IgG, IgA and IgM) 
k. Immunophenotyping of normal and/or malignant B cells. If clinically indicated, 5 
ml blood in sodium heparin should be sent to SCCA Hematopathology Laboratory 
for analysis of circulating normal and/or malignant B cells. This may be omitted if 
previously performed within 30 days of the planned T cell infusion AND the patient 
has not received anti-tumor therapy in the interim.  
2. A bone marrow aspirate/biopsy should be performed with pathology, flow cytometry, 
karyotyping, FISH and other molecular studies as indicated by the disease. This may be 
omitted if done within 30 days of planned T cell infusion AND the patient has not received 
anti-tumor therapy in the interim. 
3. A CT scan (preferably diagnostic quality) and, if possible, a PET scan should be performed 
to evaluate disease status in patients with lymphoma and CLL. Patients with B-ALL should 
undergo CT +/- PET imaging if clinically indicated. Imaging studies may be omitted in 
patients who have had recent imaging within 30 days before scheduled T cell infusion AND 
have not received anti-tumor therapy in the interim.  
4. Lumbar puncture with CSF evaluation is required for any patient with a history of CNS 
disease or signs and symptoms of CNS or epidural disease. This may be omitted if a lumbar 

 
 
 
 
28 
 
puncture performed within 30 days of scheduled T cell infusion did not show evidence of 
CNS disease.  
5. Baseline CXR 
6. Baseline EKG 
7. MUGA scan or echocardiogram 
8. Physical Exam and Karnofsky Performance Status 
9. Research samples:  
a. A blood sample (30 ml, heparin/green top tubes) should be obtained for baselineT 
cell persistence and sent to the Berger Lab at the FHCRC. 
 
b. A blood sample (up to10 ml, serum separator tube) should be obtained for 
measurement of serum inflammatory cytokine levels and baseline transgene 
immunogenicity. Samples should be sent as soon as possible to the Berger Lab at 
the FHCRC. 
 
c.  If a bone marrow is being done clinically then an additional 5 - 10 ml of marrow 
aspirate should be obtained and sent in heparin/green top tubes to the Berger Lab 
at the FHCRC.  
 
d.  If biopsy or sampling of tissues other than bone marrow; ie CSF, pleural fluid, 
lymph nodes etc. can be performed and the patient has provided consent for the collection 
of research tissue,  than these samples may be obtained and sent to the Berger lab for 
research studies. Please discuss the planned procedure with the PI. 
 
2. Cohort B (NSCLC and TNBC) 
1. Laboratory evaluations 
a. CBC, differential and platelet count 
b. Renal function panel and hepatic function panel with LDH 
c. Uric acid 
d. Serum ferritin 
e. CRP 
f. DIC panel without platelets 
g. Parathyroid hormone levels (PTH) 
h. G6PD screen 
i. Serum pregnancy test for females of child bearing potential  
j. Quantitative immunoglobulins (IgG, IgA and IgM) 
2. A CT scan (preferably diagnostic quality) and if indicated a PET scan should be performed 
to evaluate sites and status of disease. Imaging studies may be omitted in patients who have 
had recent imaging within 30 days before scheduled T cell infusion AND have not received 
anti-tumor therapy in the interim. 

 
 
 
 
29 
 
3. Baseline brain CT or MRI 
4. Baseline CXR 
5. Baseline EKG 
6. MUGA scan or echocardiogram 
7. Physical Exam and Karnofsky Performance Status          
8. Research samples. 
a. A blood sample (30 ml, heparin/green top tubes) should be obtained for baseline 
T cell persistence and sent to the Berger Lab at the FHCRC. 
b. A blood sample (up to10 ml, serum separator tube) should be obtained for 
measurement of serum inflammatory cytokine levels and baseline transgene 
immunogenicity. Samples should be sent as soon as possible to the Berger Lab at 
the FHCRC. 
 
c.  If biopsy or sampling of tissues; ie CSF, pleural fluid, bone marrow, lymph node, 
etc.  can be performed and the patient has provided consent for the collection of 
research tissue then these samples may be obtained and sent to the Berger lab for 
research studies. Please discuss the planned procedure with the PI. 
 
11C.  Evaluation Prior to Each T cell Infusion: Day 0 
 
1. Clinical: 
a. CBC, differential and platelets 
b. Renal panel and hepatic panel with LDH 
c. Uric acid 
d.  Serum ferritin  
e. CRP 
f. DIC panel without platelets 
g. Vital signs at the approximate times:  before starting, every 15 minutes during the T cell 
infusion, and hourly for 2 hours following the T cell infusion 
h. O2 saturation should be monitored continuously by pulse oximetry during the T cell 
infusion. Values should be recorded prior to initiating the infusion, approximately 15 
minutes during the T cell infusion and approximately hourly for 2 hours post infusion. 
2. Research:  
a. 
Blood samples (up to10 ml, serum separator tube) should be obtained prior to the T cell 
infusion for measurement of serum inflammatory cytokine levels and human anti-chimeric 
anti-ROR1 antibodies (HACA). Samples should be sent as soon as possible to the Berger 
Lab at the FHCRC. 
b. 
Blood samples (30 ml, heparin/green top tubes) should be obtained prior to the T cell 
infusion for baseline analysis of the presence of transferred T cells by q-PCR for vector 

 
 
 
 
30 
 
sequences and/or for expression of the EGFR and ROR1 CAR transgenes on CD8+ and 
CD4+ T cells by flow cytometry, if sufficient PBMC can be obtained.  
 
11D.  Patient Evaluations After Each T Cell Infusion  
 
The following evaluations should be performed after each T cell infusion. If a patient receives a 
second T cell infusion, the day of the second infusion will be designated ‘day 0’ for evaluations 
thereafter.  
1. Clinical 
a. Record new findings on history and physical exam 1 day after the T cell infusion and at 
least weekly for a minimum of five weeks. 
b. Laboratory studies at least twice weekly for two weeks then weekly for three weeks after 
each T cell infusion. 
i.  CBC, differential and platelet count 
ii. Renal function and hepatic function panel with LDH 
iii. Uric acid, CRP and serum ferritin, DIC panel without platelets 
c. If patients become febrile or develop symptoms of cytokine release or tumor lysis between 
the indicated time points, we may measure serum ferritin, C-reactive protein, DIC panel 
and tumor lysis markers at additional times, as clinically indicated. 
d. PTH levels should be measured 1 and 2 months after CAR-T cells. If patients develop 
hypocalcemia, additional serum PTH levels may be obtained. 
2. Immunoglobulin levels 
a. Patients receiving CAR T cells who develop B cell depletion may develop an IgG 
deficiency. Recommendations will be made for monitoring of IgG levels monthly and 
administering intravenous immunoglobulin (IVIG) as clinically indicated.  
b. Serum IgG, IgA and IgM levels may be measured at approximately 3 and 12 months after 
the final T cell infusion in patients who develop B cell aplasia. 
3. Immunophenotyping 
ROR1 is not expressed on mature normal B cells or plasma cells in the peripheral blood, but is 
expressed at a stage of B cell development in the bone marrow. The absolute numbers of normal B 
cells in the peripheral blood (and abnormal ROR1+ B cells in patients with MRD or relapse) 
should be determined on approximately days 14, 28, 60, and day 120 after the last T cell infusion, 
unless the patient initiates other non-CAR T cell systemic therapy. Blood, 5 ml in sodium heparin 
should be sent to SCCA Hematopathology Laboratory for analysis.  If a deficiency of B cells 
develops by day 120 or there are persisting ROR1 CAR-T cells, we may monitor B cell numbers 
again on D180 and then every 6 months until normal B cells recover.  
4. Serum Cytokines  
a. A blood sample (up to 10 ml, serum separator tube) should be obtained approximately on 
days 1, , 4, 7, 10, 14, 21 ,28 and 35 after each T cell infusion for measurement of serum 
inflammatory cytokine levels. Any patient that stays on the Immunotherapy service beyond 

 
 
 
 
31 
 
D35 may have weekly serum cytokine samples obtained as clinically indicated. Samples 
should be sent to the Berger Lab at the FHCRC. 
b. If patients become febrile, develop symptoms of cytokine release, or assessment of 
cytokines is clinically appropriate at times other than those indicated, we may measure 
cytokine levels at additional times. 
5. Evaluation of persistence and phenotype of ROR1 CAR-T Cells 
a. Blood samples (30 ml, heparin/green top tubes) should be obtained on approximately days 
1, 4, 7, 10, 14, 21, 28, 35, 60, 120, 180, and 365 after the T cell infusion for analysis of the 
persistence of transferred T cells. Additional samples may be collected at other times than 
those indicated if required for evaluation of persistence of CAR-T cells. Persistence 
monitoring may be discontinued beyond day 28 in patients who proceed to allogeneic 
hematopoietic stem cell transplantation and/or do not have detectable transgene-expressing 
T cells on two consecutive occasions. Samples should be sent to the Berger Lab at the 
FHCRC. A subset of blood samples obtained after the infusion should also be analyzed by 
multiparameter flow cytometry for the phenotype of persisting CD8+ and CD4+ ROR1 
CAR-T cells. Markers that may be analyzed include CD62L, CCR7, CD28, CD27, CD127, 
PD1, Lag3, Tim3 and PDL1. 
b. If patients become febrile, develop possible signs of toxicity, or assessment of CAR-T cell 
persistence is clinically appropriate at times other than those indicated, we may measure 
the persistence of transferred T cells at additional times.  
 
6. Evaluation of human anti-chimeric anti-ROR1 antibodies (HACA) 
a. A blood sample (up to 10 ml, serum separator tube) should be obtained on approximately 
days 7, 14, 21, 28, 35, 60, 90 and 180 after each T cell infusion for measurement of human 
anti-chimeric anti-ROR1 antibodies (HACA). Samples should be sent to the Berger Lab at 
the FHCRC. 
7. Evaluation of transgene immunogenicity 
Loss of persistence of transferred T cells by flow cytometry for the EGFRt marker (<0.01% of 
CD3+ cells) is possible.  Patients will be evaluated for antibody or T cell-mediated transgene 
immune responses as follows: 
a. Blood (10ml, serum separator tube) should be collected around days 28, 60, 120, 180, 365 
after the final T cell infusion and serum extracted for evaluation of antibody-mediated 
immune responses. Samples should be sent to the Berger Lab at the FHCRC.  
b. Blood (20 ml, sodium heparin/green top tubes) should be collected around days 28, 60, 120, 
180, 365 after the final T cell infusion) and PBMC isolated for evaluation of cellular 
immune responses. Samples should be sent to the Berger Lab at the FHCRC.  
8.   Archival samples for future studies of T cell function  
a. Blood (60 ml, sodium heparin/green top tubes) OPTIONAL may be obtained from patients 
once between days 0 and 21 after ROR1 CAR-T cell infusion and once between days 21 and 
90 for archival purposes. 
b.  Blood (20 ml, sodium heparin/green top tubes) should be obtained from patients around 
days 120, 180, and 365 after the final T cell infusion for archival purposes. Samples should 
be sent to the Berger Lab at the FHCRC. 

 
 
 
 
32 
 
c. Serum (up to 10 ml, serum separator tube) should be obtained from patients at around days 
120, 180, and 365 after the final T cell infusion for archival purposes. Samples should be 
sent to the Berger Lab at the FHCRC. 
9. Evaluation of migration of adoptively transferred ROR1 CAR-T cells 
a. If aspirations and/or biopsies of bone marrow are performed for evaluation of tumor 
response or other clinical indications then additional aspirates (5 - 10 ml in heparin/green 
top tubes) should be obtained and sent to the Berger Lab, FHCRC.  
b. If biopsy or sampling of tissues other than bone marrow is performed for clinical indications 
then additional tissue may be obtained during the same procedure for research studies. 
Please discuss the planned procedure with the PI. 
10.  Evaluation of tumor response  
 
Cohort A (CLL, MCL, and ALL) 
a. Bone marrow aspiration and biopsy.  
i. Because the timing of disease response to ROR1 CAR-T cells is unknown, a bone 
marrow aspirate and biopsy should be performed between 14 days and 3 months after each 
T cell infusion unless the patient did not have bone marrow involvement prior to therapy. 
Bone marrow evaluation should be performed before the patient receives additional 
conventional anti-tumor therapy. Evaluation should include pathology analysis, flow 
cytometry, karyotyping, FISH studies and other molecular studies, according to disease-
specific guidelines.  
ii. A bone marrow aspirate and biopsy should also be performed at approximately 6 and at 
approximately 12 months after the first T cell infusion, as clinically indicated. 
b.   Fresh tumor biopsy: If biopsy or sampling of tissues other than bone marrow; ie CSF, 
pleural fluid, lymph nodes etc. can be performed and the patient has provided consent for the 
collection of research tissue,  then these samples may be obtained and sent to the Berger lab for 
research studies.. Please discuss the planned procedure with the PI. 
c. Imaging studies 
 
i. A CT scan (preferably diagnostic quality) and, if possible, a PET scan of the neck, chest, 
abdomen and pelvis should be obtained between 1 and 3 months after each T cell infusion, 
and should be performed before the patient receives additional conventional anti-tumor 
therapy. Patients with B-ALL do not require CT/PET imaging, unless it is clinically 
indicated.  
ii. Patients with a response to treatment should have CT or PET/CT scans performed at 6 
and 12 months after the first T cell infusion, as clinically indicated. 
d. CBC with diff, serum chemistry panel and LDH should be performed concurrently with 
restaging by biopsy or imaging.  
e. Peripheral blood flow cytometry for patients in Cohort A should be performed around the 
time of restaging by biopsy or imaging.  
f. Standard criteria (as detailed in Appendix C) will be used to define tumor response. 

 
 
 
 
33 
 
g. Evaluation of tumor response may be discontinued in patients who proceed to other 
systemic non-CAR T cell therapies 
Cohort B (NSCLC and TNBC) 
a.  Imaging studies 
i. A CT scan (preferably diagnostic quality) and if indicated a PET scan of the chest, 
abdomen and pelvis should be obtained between 1 and 3 months after each T cell infusion, 
and should be performed before the patient receives additional conventional anti-tumor 
therapy.  
ii.  A MRI of the brain should be obtained between 1 and 3 months after each T cell 
infusion for any patient with a history of brain metastases. 
iii. Patients with a response to treatment should have follow-up imaging studies at 
approximately 6 and 12 months after the T cell infusion, and every 6 months thereafter, as 
clinically indicated. 
 
    b. Fresh tumor biopsy: If biopsy or sampling of tissues other than bone marrow; ie CSF, pleural 
fluid, lymph nodes etc. can be performed and the patient has provided consent for the collection of 
research tissue,  than these samples may be obtained and sent to the Berger lab for research studies. 
Please discuss the planned procedure with the PI. 
 
c. CBC with diff, serum chemistry panel and LDH should be performed around the time of 
restaging by biopsy or imaging.  
d.  Standard criteria (as detailed in Appendix C) will be used to define tumor response. 
Evaluation of tumor response may be discontinued in patients who proceed to other non-CAR-T 
cell therapies. 
11. Long Term Follow-Up (Appendix F 
Enrolled patients who receive ROR1 CAR-T cells will be asked to participate in long-term follow-
up (LTFU) according to the guidelines set forth by the FDA’s Biologic Response Modifiers 
Advisory Committee that apply to gene transfer studies. Current recommendations from the FDA 
suggest a minimum of 15 years of follow-up. Recommendations will be made for an autopsy to be 
conducted if the research participant dies. 
 
12. 
PROTOCOL ENROLLMENT 
 
12A. Projected Target Accrual 
 
The number of patients required to complete dose escalation/de-escalation studies is variable, and 
dependent on observed toxicity. We estimate that defining a dose level in Cohorts A and B with 
acceptable toxicity will take up to 30 patients in each cohort.  Thus, up to a total of 60 patients 
could be enrolled on the trial.   
 
Table 1: Targeted / Planned Enrollment 

 
 
 
 
34 
 
 
Ethnic Category 
            Gender                               
Number of  
                                                          
Subjects 
Females 
Males 
Total 
Hispanic or Latino 
4 
2 
6 
Not Hispanic or Latino 
32 
22 
54 
Ethnic Category Total of All 
36 
24 
60 
 
Racial Category 
 
American Indian / Alaska 
0 
1 
1 
Asian 
3 
3 
6 
Native Hawaiian or Other 
Pacific Islander
0 
0 
0 
Black or African American 
3 
1 
4 
White 
30 
19 
49 
Racial Categories:  Total of All 
Subjects* 
36 
24 
60 
 
 
13.  RECORDS 
 
The medical record containing information regarding treatment of the patient will be maintained as 
a confidential document, within the guidelines of the Fred Hutchinson Cancer Research Center, the 
University of Washington Medical Center and the Seattle Cancer Care Alliance. The investigators 
will ensure that data collected conform to all established guidelines for coding collection, key entry 
and verification. Each patient is assigned a unique patient number to assure patient confidentiality.  
Patients will not be referred to by name or by any other personal identifier in any publication or 
external presentation.  The Clinical Statistics Departments at FHCRC maintain a patient database 
to allow storage and retrieval of patient data collected from a wide variety of sources. The licensed 
medical records departments, affiliated with the institution where the patient receives medical care, 
maintains all original inpatient and outpatient chart documents.  
The primary research records are kept in access controlled office spaces or password protected 
computer based applications. Information gathered from this study regarding patient outcomes and 
adverse events may be made available to the Federal Drug Administration, NIH and Juno 
Therapeutics. All precautions to maintain confidentiality of medical records will be taken.  
 
14. EVALUATION AND STATISTICAL CONSIDERATIONS 
 
14A.  Type of study 
 
This is a phase 1/2 study to assess the safety and antitumor activity of adoptive T cell therapy with 
autologous CD4+ T cells and CD8+ T cells transduced to express a ROR1-specific CAR for 
patients with advanced ROR1+ B cell malignancies.  
 
14B. Definition of endpoints 
 

 
 
 
 
35 
 
1. Safety assessment (relevant data being obtained) 
a. General Toxicity Assessment 
• History and physical exam before and at intervals after T cell infusions. 
• Pulse oximetry before and during the infusion 
• CBC and chemistry battery before and at intervals after the T cell infusion 
• Toxicity grading according to NCI CTCAE Version 4.0 
• Serum cytokine levels 
•  Immunophenotyping  
• Serum immunoglobulin levels 
• Replication competent lentivirus  
• Adverse event reporting 
 
2. Efficacy Assessment 
a. Evaluation of the duration of persistence of adoptively transferred ROR1 CAR-T cells 
b. Evaluation of the migration of adoptively transferred ROR1 CAR-T cells 
c. Evaluation of antitumor activity of adoptively transferred ROR1 CAR-T cells by evaluating 
the objective response rate of complete remission and partial remission, and determining 
progression free survival, and overall survival. 
 
14C.  Endpoint Evaluation 
 
1.  Safety Assessment 
Patients will be treated in cohorts of two patients at one of five dose levels of ROR1 CAR-T cells 
starting at dose level 1 (Section 9D.1 and 9D.2) to determine cell doses associated with an 
estimated toxicity rate of less than 25%.  
 
a. Dose limiting toxicities 
DLTs will be assessed for 28 days following the infusion of the CAR T cell product. The 
following events will be considered DLTs if they are attributed as at least possibly related 
to T cell administration. 
Grading will be done in accordance with the NCI Common Terminology Criteria for 
Adverse Events (CTCAE) Version 4.0 unless otherwise specified. 
 
1. Grade ≥ 3 allergic reaction related to the CAR T cell infusion 
2. Grade ≥ 3 autoimmune reactions  
3. Any Grade 3 or 4 non-hematologic event that has not resolved to < grade 3 by day 28 
post T cell infusion 
4. Grade ≥ 3 neurotoxicity of greater than 7 days duration 
5. Grade ≥ 3 neurotoxicity that does not revert to Grade 1 or baseline within 28 days 
6. Grade ≥ 3 seizures that do not resolve to < grade 3 within 3 days 
7. Grade ≥ 4 CRS (using criteria modified from Lee 2014) 
8. Grade 3 CRS that does not resolve to < grade 3 within 7 days 
9. Any other toxicity not meeting the above criteria that is deemed by the PI to represent 
a DLT 

 
 
 
 
36 
 
 
b. Stopping and Suspension Rules  
If there ever exists sufficient evidence to suggest that the true probability of treatment-
related death by day 100 exceeds 20% (regardless of dose), enrollment of patients 
suspended pending a detailed review by the PI, study monitor and statistician and 
DSMB.  Sufficient evidence for this purpose will be defined as any observed outcome 
whose lower 80% confidence limit exceeds 20%. Operationally, any of the following ratios 
of treatment-related deaths to patients treated would trigger such a rule: 2/2-4, 3/5-7, 4/8-
11, 5/12-15, 6/16-20, 7/21-24, 8/25-28, or 9/29-30. If the true probability of treatment-
related death is 0.10, then the probability of stopping after 15 or 30 patients is 
approximately 0.07.  If the true probability of treatment-related death is 0.40, then the 
probability of stopping after 15 or 30 patients is approximately 0.85 and 0.91, respectively 
(probabilities estimated from 5,000 simulations). 
 
2.  Assessment of efficacy of transferred T cells 
Data should be collected for persistence, migration and efficacy of transferred T cells and 
descriptive statistics will be used to summarize the changes from baseline where possible. For 
those patients with measurable disease at the time T cell therapy commences, responses will be 
evaluated using standard response criteria based on CT or PET imaging and histologic analysis of 
bone marrow or other tissue samples. 
 
15.   
GUIDELINES FOR REPORTING ADVERSE EVENTS AND DATA SAFETY AND 
MONITORING PLAN 
 
15A. General issues and IRB reporting requirements  
1. Definitions associated with reportable events and reporting requirements can be found on the 
FHCRCs Institutional Review Office (IRO) extranet website (Table 1).  
2. The review and reporting of adverse events will be in accordance with the Cellular 
Immunotherapy AE reporting for separate FH Sponsor and Investigator SOP. 
3. The NCI Common Terminology Criteria for Adverse Events (CTCAE) Version 4 
(http://ctep.cancer.gov/protocolDevelopment/electronic_applications/ctc.htm#ctc_40) will be used 
for grading and analysis of adverse events. All grade 3 or greater adverse events will be collected 
during and for 48 hrs post leukapheresis and again from the start of lymphodepletion through day 
35 after each T cell infusion. The collection of adverse events will stop at the time of 
commencement of new anti-tumor therapy. 
To ensure that investigative treatment-related conditions are distinguished from disease-related 
conditions, attribution of causality will be established in grading adverse events. For each event, 
the Principal Investigator or designee, in conjunction with the physician or research nurse who 
examined and evaluated the research participant, will assign the attribution.  Data managers who 
are removed from the clinical assessment of the research participant should not perform this.   
Attribution of adverse events attributed to the infused genetically modified T cells should be 
determined using the following criteria: 
• Definite - the adverse event is clearly related to the infused T cells 
• Probable - the adverse event is likely related to the infused T cells 

 
 
 
 
37 
 
• Possible - the adverse event may be related to the infused T cells 
• Unlikely - the adverse event is doubtfully related to the infused T cells 
• Unrelated - the adverse event is clearly not related to the infused T cells 
Deaths occurring 35 days or later from a T cell infusion and death due to progression of known 
malignancy will be scored as a grade 5 toxicity, but with a low level of attribution, unlikely or not 
related, unless a serious toxicity that was attributed to the T cell infusion occurred prior to day 35, 
did not resolve and was considered a significant factor in the patient’s demise.   
4. The FHCRC IRB will be notified of reportable events by the FHCRC Principal Investigator 
(PI) or study nurse according to current reporting obligations as found on the FHCRC Institutional 
Review Office extranet website. 
5. Reporting of unanticipated adverse effects to the FDA will be the responsibility of the IND 
sponsor. 
6. The FHCRC PI and Research Nurse and study personnel should meet regularly (in person or 
via teleconference) to review all reported events.  
 
Table 2: FHCRC IRB Policies for Reportable Events.  
(Relevant FHCRC policies include, but are not limited to, the following documents. Please also 
refer to the FHCRC IRO website) 
IRB Policy 
2.6 
Adverse Events and Other 
Unanticipated Problems 
Involving Risks to Subjects or
Others 
http://extranet.fhcrc.org/EN/sections/iro/irb/ae.html 
IRB Policy 
1.9 
Noncompliance with the 
Office of the Director’s 
Human Research Protection 
Program Policy 
http://extranet.fhcrc.org/EN/sections/iro/irb/ae.html 
IRB Policy 
1.1 
Reporting Obligations for 
Principal Investigators 
http://extranet.fhcrc.org/EN/sections/iro/irb/policy/i
ndex.html 
IRB Policy 
2.2 
Continuing Review 
http://extranet.fhcrc.org/EN/sections/iro/irb/policy/i
ndex.html 
IRB Policy 
1.13 
Investigational New Drugs 
(IND), Biologics and 
Investigational Device 
Exemptions (IDE) 
http://extranet.fhcrc.org/EN/sections/iro/irb/policy/i
ndex.html 
 
Table 3: FHCRC IRB Forms for Reporting 
Adverse Event Reporting Form 
http://extranet.fhcrc.org/EN/sections/iro/irb/forms/in
dex.html  
Unanticipated Problem Reporting Form 
http://extranet.fhcrc.org/EN/sections/iro/irb/forms/in
dex.html  
Noncompliance Reporting Form 
http://extranet.fhcrc.org/EN/sections/iro/irb/forms/in
dex.html  
 
15B.  Reporting to the FDA 

 
 
 
 
38 
 
As a study conducted under IND (Investigational New Drug) regulations we will comply 
with the FDA regulations regarding safety reporting 21CFR312.32 including the 
following requirements: 
1. 
A sponsor must promptly review all information relevant to the safety of the drug 
21CFR312.32 (b). 
2. 
A sponsor must notify FDA in an IND safety report of potential serious risks, as 
soon as possible but in no case later than 15 calendar days after the sponsor determines 
that the information qualifies for reporting under 21 CFR312.32 (c)(1). Information that 
is required to be reported includes, but is not limited to, a. Serious and unexpected 
adverse reactions and b. An increased rate of occurrence of serious suspected adverse 
reactions.  
3. 
The IND safety report must be completed and sent to the FDA in a narrative format, 
on FDA Form 3500A, or an electronic format. 
4. 
A sponsor must also notify FDA of any unexpected fatal or life-threatening 
suspected adverse reaction as soon as possible but in no case later than 7 calendar days 
after the sponsor’s initial receipt of the information 21CFR312.32 (c)(2). 
15C.   Data and Safety Monitoring Plan 
 
1.  Definition of Risk Level 
This phase I/II trial involves genetic modification of somatic cells and requires an IND. At the 
FHCRC, this type of trial has independent monitoring through the Clinical Research Support 
Office at the FHCRC, and has a Data and Safety Monitoring Board (Section 15C. 3).  
2. Monitoring and Personnel Responsible for Monitoring 
a. The Principal Investigator (P.I.) is responsible for every aspect of the design, conduct and 
final analysis of the protocol.  Regulations defining the responsibilities for assessment and 
reporting of adverse events (AE), serious AE and unexpected AE are defined by the Code 
of Federal Regulations:  21 CFR 312.32 and Common Terminology Criteria for Adverse 
Events (CTCAE) Version 4.0 published by the Cancer Therapy Evaluation Program 
(CTEP), a division of the NCI/NIH. Sponsor responsibilities include providing the PI with 
the necessary information to conduct the investigation, ensuring proper monitoring, 
ensuring that the investigation is conducted in accordance with the protocol, conducting 
ongoing review of the safety and efficacy of the investigational product, and reporting to 
the FDA consistent with 21 CFR 312 Subpart D. 
 
b. This clinical study will rely upon the monitoring of the trial by the P.I. in conjunction with 
a Study Physician(s), Physician Assistant(s) (PA) or Nurse Practitioner(s), Research 
Nurse(s), Research Coordinator(s), statistician, and an independent Study Monitor assigned 
by the FHCRC Clinical Research Support Office (CRSO). Institutional support of trial 
monitoring will be in accordance with the FHCRC/University of Washington Cancer 
Consortium Institutional Data and Safety Monitoring Plan.  Under the provisions of this 
plan, FHCRC Clinical Research Support coordinates data and compliance monitoring 
conducted by consultants, contract research organizations, or FHCRC employees 
unaffiliated with the conduct of the study.  Independent monitoring visits occur at specified 

 
 
 
 
39 
 
intervals determined by the assessed risk level of the study and the findings of previous 
visits per the institutional DSMP.  
 
In addition, protocols are reviewed at least annually and as needed by the Consortium Data 
and Safety Monitoring Committee (DSMC), FHCRC Scientific Review Committee (SRC) 
and the FHCRC/University of Washington Cancer Consortium Institutional Review Board 
(IRB).  The review committees evaluate accrual, adverse events, stopping rules, and 
adherence to the applicable data and safety monitoring plan for studies actively enrolling or 
treating patients.  The IRB reviews the study progress and safety information to assess 
continued acceptability of the risk-benefit ratio for human subjects.  Approval of 
committees as applicable is necessary to continue the study. 
   
 
The trial will comply with the standard guidelines set forth by these regulatory committees 
and other institutional, state and federal guidelines.  
c. Continuous monitoring of the data and safety of this study should be performed by the 
Protocol Management Team (PMT), which consists of the Principal Investigators, Research 
Nurse, and Statistician. Monitoring by the PMT should be performed at least monthly or 
more often if necessitated by the development of adverse events, and should include review 
of the data on all enrolled patients with a summary of grade 3+ adverse events and follow up 
information for each patient. This list of patients should be included with the PMT report 
form. The PMT will be responsible for implementation of the stopping rules for safety if 
necessary. 
d. A Case Report Form (CRF) should be completed for every patient that was registered for 
participation in the study. The Principal Investigator or a Co-Investigator will sign and date 
the indicated places of the CRF.  This signature will indicate that thorough inspection of the 
data therein has been made, and will certify the contents of the form. 
 
3. Data and Safety Monitoring Board 
The study will be monitored by the Immunotherapy Integrated Research Center (IIRC) DSMB. 
The DSMB will be responsible for safeguarding the interests of trial participants and assessing 
the safety and efficacy of the interventions during the trial.  This responsibility will be exercised 
by providing recommendations about stopping or continuing the trial.  To contribute to 
enhancing the integrity of the trial, the DSMB may also formulate recommendations relating to 
the selection, recruitment and retention of participants and their management; adherence to 
protocol-specified regimens; and the procedures for data management and quality control.  
 
The DSMB will be advisory to the study Sponsor and the PI, who will be responsible for prompt 
review of the DSMB recommendations to guide decisions regarding continuation or termination 
of the trial and whether amendments to the protocol or changes in study conduct are required. 
 
The DSMB is an independent, multidisciplinary group consisting of clinical experts and a 
statistician who collectively have experience in leukemia, lymphoma, hematology, biostatistics, 
and the conduct and monitoring of clinical trials. The DSMB will meet approximately every 6 
months to review data. The current members are listed in the IIRC DSMB charter. 
 

 
 
 
 
40 
 
16. TERMINATION OF THE STUDY 
The study may be terminated at any time by the Protocol PI, the IND Sponsor, the FHCRC IRB, or 
the FDA. 

 
 
 
 
41 
 
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49 
 
18. APPENDICES 
 
APPENDIX A - Suggested Cytoreduction/Lymphodepletion Chemotherapy Regimens 
 
The following is a guide to chemotherapy regimens that may be used immediately prior to 
adoptive T cell therapy.  The selection of appropriate chemotherapy regimen to provide tumor 
cytoreduction and lymphodepletion should be discussed with the study PI’s.  The preferred 
chemotherapy regimen for patients on the trial is Regimen C however other regimens may be 
used depending on PI discretion. Chemotherapy regimens that employ prolonged administration 
of corticosteroids that would continue after the T cell infusion cannot be used for prior to T cell 
infusion. Patients should be scheduled for T cell infusions starting 36-96 hours after the last dose 
of chemotherapy.  
 
 
Regimen A  
Cyclophosphamide 30-60 mg/kg on day 1; fludarabine 25 mg/m2 on days 2-4 
 
Regimen B 
Cyclophosphamide 30-60 mg/kg on day 1; fludarabine 25 mg/m2 on days 2-6 
 
Regimen C  (Preferred) 
Cyclophosphamide 300 mg/m2 on day 1-3; fludarabine 30 mg/m2 on days 1-3 
 
Regimen D 
Cyclophosphamide 2-4 gm/m2 on day 1 

 
 
 
 
50 
 
APPENDIX B - PERFORMANCE STATUS SCALES 
These indices allow patients to be classified as to their functional impairment and can be used to 
compare effectiveness of different therapies and to assess the prognosis in individual patients.  
 
KARNOFSKY PERFORMANCE STATUS SCALE 
Karnofsky DA Burchenal JH. (1949). "The Clinical Evaluation of Chemotherapeutic Agents in 
Cancer." In: MacLeod CM (Ed), Evaluation of Chemotherapeutic Agents. Page 196. 
DEFINITIONS 
RATING (%) 
CRITERIA 
Able to carry on normal 
activity and to work; no 
special care needed. 
  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.  
Unable to work; able to live a
home and care for most 
personal needs; varying 
amount of assistance needed.
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 personal needs. 
50 
Requires considerable assistance and frequent 
medical care.  
Unable to care for self; 
requires equivalent of 
institutional or hospital care; 
disease may be progressing 
rapidly. 
40 
Disabled; requires special care and assistance. 
30 
Severely disabled; hospital admission is indicated
although death not imminent. 
20 
Very sick; hospital admission necessary; active 
supportive treatment necessary. 
10 
Moribund; fatal processes progressing rapidly. 
0 
Dead 
 
ECOG PERFORMANCE STATUS SCALE 
Oken, M.M., Creech, R.H., Tormey, D.C., Horton, J., Davis, T.E., McFadden, E.T., Carbone P.P. 
(1982) Am J Clin Oncol 5:649. 
GRADE 
CRITERIA 
0 
Fully active, able to carry on all pre-disease performance without restriction 
1 
Restricted in physically strenuous activity but ambulatory and able to carry out work 
of a light or sedentary nature, e.g., light house work, office work 
2 
Ambulatory and capable of all selfcare but unable to carry out any work activities. Up
and about more than 50% of waking hours 
3 
Capable of only limited selfcare, confined to bed or chair more than 50% of waking 
hours 
4 
Completely disabled. Cannot carry on any selfcare. Totally confined to bed or chair 
5 
Dead 

 
 
 
 
51 
 
APPENDIX C – RESPONSE CRITERIA 
 
For Patients with Chronic lymphocytic leukemia (CLL)  * 
 
Complete remission (CR): Normal imaging studies (X-ray, CT, MRI) (nodes, liver, and spleen), 
peripheral blood by flow cytometry has no clonal lymphocytes, bone marrow by flow cytometry 
has no clonal lymphocytes, bone marrow by morphology has no nodules (or if present, nodules are 
free from CLL cells by immunohistochemistry), and the duration is at least 2 months. 
CR with minimal residual disease (CR-MRD): CR by above criteria except peripheral blood or 
bone marrow by flow cytometry with >0 - <1 CLL cells/1000 leukocytes (0.1%) 
Partial remission (PR): Absolute lymphocyte count in peripheral blood at least 50% decreased 
and physical exam/Imaging studies (nodes, liver, and/or spleen) at least 50% decreased.  Duration 
is at least 2 months. 
Progressive disease (PD): at least one of: Physical exam/imaging studies (nodes, liver, and/or 
spleen) >50% increase or new, circulating lymphocytes by morphology and/or flow cytometry 
>50%increase, and lymph node biopsy with Richter’s transformation 
Stable disease (SD): Did not meet any of the above criteria for complete or partial remission or 
progression. 
* Hallek et al. (2008) Guidelines for the diagnosis and treatment of chronic lymphocytic leukemia: 
a report from the International Workshop on Chronic Lymphocytic Leukemia updating the 
National Cancer Institute-Working Group 1996 guidelines. Blood. 111(12): 5446-56.  
 
For Patients with Mantle Cell Lymphoma (Cheson Criteria) ^ 
 
Response 
Definition 
Nodal Masses 
Spleen, Liver 
Bone Marrow 
CR 
Disappearance of 
all evidence of 
disease 
(a) FDG-avid or PET positive prior to 
therapy; mass of any size permitted if 
PET negative (b) Variably FDG-avid   
or PET negative; regression to normal 
size on CT 
Not palpable, nodules 
disappeared 
Infiltrate cleared on repeat 
biopsy; if indeterminate by 
morphology, immuno-
histochemistry should be 
negative 
PR 
Regression of 
measurable   
disease and no new
sites 
≥ 50% decrease in SPD of up to 6 
largest dominant masses; no increase   
in size of other nodes (a) FDG-avid or 
PET positive prior to therapy; one or 
more PET positive at previously 
involved site (b) Variably FDG-avid    
or PET negative; regression on CT 
≥ 50% decrease in SPD of 
nodules (for single nodule in
greatest transverse diameter)
no increase in size of liver or
spleen 
Irrelevant if positive prior   
to therapy; cell type should 
be specified 
SD 
Failure to attain 
CR/PR or PD 
(a) FDG-avid or PET positive prior to 
therapy; PET positive at prior sites of 
disease and no new sites on CT or 
PET; (b) Variably FDG-avid or PET 
negative; no change in size of    
previous lesions on CT 
  
  

 
 
 
 
52 
 
Relapsed 
disease or 
PD 
Any new lesion or 
increase by ≥ 50% 
of previously 
involved sites from
nadir 
Appearance of a new lesion(s) >  
1.5 cm in any axis, ≥ 50% increase in 
SPD of more than one node, or ≥ 50% 
increase in longest diameter of a 
previously identifed node > 1 cm in 
short axis 
> 50% increase from nadir in
the SPD of any previous 
lesions 
New or recurrent  
involvement 
Abbreviations: CR, complete remission; FDG, [18F]fluorodeoxyglucose; PET, positron emission 
tomography; CT, computed tomography; PR, partial remission; SPD, sum of the product of the 
diameters; SD, stable disease; PD, progressive disease.   
 
 
 
 
 
 
^ Cheson BD et al. (2007) JCO 25(5): 579. 
 
For Patients with Acute lymphoblast leukemia (ALL)  
 
Remission status will be determined by restaging of bone marrow and other involved sites by 
morphology, flow cytometry and molecular studies, as appropriate. 
 
For Patients with NSCLC and TNBC ~ 
 
Tumor response and progression will be evaluated in this study using the international criteria 
proposed by the Response Evaluation Criteria in Solid Tumors (RECIST) Committee (Eisenhauer 
EA, Therasse P, Bogaerts J, et al. New response evaluation criteria in solid tumours: revised 
RECIST guideline (version 1.1). European journal of cancer (Oxford, England : 1990). Jan 
2009;45(2):228-247).  Tumor flare has not been observed in our experience with CD19 CAR-T 
cells; however, patients with an increase in tumor burden at initial restaging will have confirmation 
by a repeat assessment at ≥ 4 weeks from restaging (Wolchok JD, Hoos A, O'Day S, et al. 
Guidelines for the evaluation of immune therapy activity in solid tumors: immune-related response 
criteria. Clinical cancer research : an official journal of the American Association for Cancer 
Research. Dec 1 2009;15(23):7412-7420) Measurable lesions are defined as those that can be 
measured in at least one dimension as > 20 mm with conventional CT techniques (cuts of 10 mm 
or less in slice thickness) or as > 10 mm with spiral CT (5 mm contiguous algorithm).  All tumors 
must be recorded in millimeters or decimal fractions of centimeters.  All other lesions are 
considered non-measurable.  Measurable lesions will be identified as target lesions.  A sum of the 
longest diameter for all the target lesions will be calculated and reported as the baseline sum.  The 
baseline sum will be used as a reference by which to characterize the objective tumor response. 
Given potential for differential trafficking of CAR T cells to nodal sites, site of target lesions will 
be carefully captured to assess for possible differential radiographic response in visceral versus 
nodal disease sites.  
 
Target lesion responses will be described as (1) Complete response (CR): disappearance of all 
target lesions; (2) Partial Response (PR): at least 30% decrease in the sum of the longest diameter 
(LD) of target lesions; (3) Progressive disease (PD): at least a 20% increase in the sum of the LD 
of the target lesions, taking as a reference the smallest LD recorded prior to receiving the first T 
cell infusion; and, (4) Stable Disease: neither sufficient shrinkage to qualify for PR nor sufficient 
growth to qualify for PD.  Non-target lesions will be described as CR: disappearance of all non-
target lesions and normalization of tumor-marker; Incomplete response/SD: persistence of one or 
more non-target lesion(s) and or persistent abnormal tumor marker; PD: Appearance of  ≥1 new 
lesions and/or unequivocal progression of existing non-target lesion. 
 

 
 
 
 
53 
 
We anticipate some patients may have bone-only disease.  The RECIST system incorporates CT 
imaging and considers bone disease to be “non-measurable”. FDG PET imaging may be useful to 
monitor response to therapy in bone-only lesions.  PET scans are interpreted quantitatively by 
using the maximum SUV of the most conspicuous bone lesion at baseline PET scan and 
monitoring change in that value. Based on recommendations proposed by the European 
Organization for Research and Treatment of Cancer (EORTC) regarding the use of FDG-PET for 
disease assessment, we will use FDG PET imaging for monitoring response to T cell infusions in 
patients with bone-only metastasis as outlined in the following Table. 
 
 
 
EORTC Criteria for Assessment of Response by FDG-PET ~ 
Progressive Metabolic Disease 
• Increase of SUV >25% 
• Visible increase of FDG uptake (>20% of longest dimension) 
• Appearance of new focus 
Stable Metabolic Disease 
• Increase of SUV <25% or decrease <15% 
• No visible increase of the extent of FDG uptake 
 
Partial Metabolic Response 
• Reduction of a minimum of 15-25% of SUV after 1 treatment cycle; >25% after more than 
1 treatment cycle 
Complete Metabolic Response 
• Complete resolution of FDG uptake 

 
 
 
 
54 
 
APPENDIX D - Patient evaluations over the course of the study 
 
 
 
 
 
Approximate days post T cell infusion 
Study Assessments/ Testing 
Screen 
Enrolled 
0 
During and following 
each T cell infusion 
1 
 
2 
4 
7 
10 
14 
21 
28 
 
35 
 
60 
 
90 
120 
180 
 
365 
History and Physical1 
X 
X 
 
Day 1 and weekly x 5 
X 
 
 
X 
 
X 
X 
X 
X 
 
 
 
 
 
Karnofsky  performance status 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Vitals including O2 sats 
 
 
X 
q 15 min during then 
hourly x 2 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Automated CBC with 
differential, platelet count 
 
X 
 
X 
 
X 
 
2 x in first two weeks 
then weekly x3 
X 
 
 
X 
X 
X 
X 
X 
 
X 
 
 
 
 
 
Renal panel and Hepatic panel 
with LDH 
X 
X 
X 
2x in first two weeks 
then weekly x3 
X 
 
 
X 
X 
X 
X 
 
X 
 
X 
 
 
 
 
 
Uric acid, ferritin, CRP 
 
X 
X 
2 x in first two weeks 
then weekly x3 
X 
 
 
X 
X 
X 
X 
 
X 
 
X 
 
 
 
 
 
DIC panel without platelets 
 
X 
X 
2 x in first two weeks 
then weekly x 3 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
Pregnancy Test2 
X 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Bloodworks NW Recipient 
Donor Battery Panel 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
G6PD screening 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Patient/Donor CD3 
chimerism3 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
ABO blood typing and 
antibody screen 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Muga scan or echocardiogram 
 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
ROR1 expression confirmed 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Quantitative IgG, IgM, IgA4  
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
 
 
X 
EKG 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
CXR 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Lumbar Puncture with CSF 
evaluation5 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

 
 
 
 
55 
 
Study Assessments/ Testing 
Screen 
Enrolled 
0 
During and following 
each T cell infusion 
1 
 
2 
4 
7 
10 
1
4 
2
1 
2
8 
 
3
5 
 
60 
 
90 
120 
180 
 
365 
Tumor/tissue  biopsy6 
 
X 
 
 
 
 
 
 
 
 
 
X 
 
 
 
 
 
 
Bone marrow aspirate7 
 
 
X 
 
 
 
 
 
 
 
 
X 
 
X 
 
X 
CT/PET scan8 
 
X 
 
 
 
 
 
 
 
 
 
X 
 
X 
 
Brain CT or MRI 
 
X 
 
 
 
 
 
 
 
 
 
 
X 
 
 
 
PB review of lymphocyte 
subsets 
 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
PBMC collection 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
PTH levels 
 
 
 
 
 
 
 
 
 
 
 
X 
 
X 
 
 
 
 
 Cytokine levels 
 
X 
X  
X 
 
 
X 
X 
X 
X 
X 
X 
X 
 
 
 
 
 
HACA 
 
 
X  
 
 
 
X 
 
X 
X 
X 
X 
X 
X 
 
X 
 
Immunophenotyping9  
 
 
X 
 
 
 
 
 
 
 
X 
 
X 
 
X 
 
X 
 
 
T cell persistence9 
 
X 
X 
 
X 
 
X 
X 
X 
X 
X 
X 
X 
X 
 
X 
X 
X 
Transgene 
immunogenicity_serum 
 
X 
 
 
 
 
 
 
 
 
 
X 
 
X 
 
X 
X 
 
X 
Transgene 
immunogenicity_PBMC 
 
 
 
 
 
 
 
 
 
 
 
X 
 
X 
 
X 
X 
 
X 
Serum archive10 
 
 
 
 
 
 
X 
X 
X 
PBMC archive10 
 
 
 
0-21 and 21-90 
X 
X 
X 
X 
  X 
 
 
 
1. 
History to include hematologic, cytogenetic, flow cytometric and histologic findings at diagnosis and time of enrollment as well as prior therapies and response to therapy. 
2. 
If clinically indicated.  
3. 
In patients who have had an allo transplant only. 
4. 
Timepoints after enrollment may occur in patients who develop B cell aplasia. 
5. 
Required for any patient with a history of CNS disease or signs and symptoms of CNS or epidural disease, unless a negative lumbar puncture was performed within 30 days prior to 
scheduled T cell infusion. 

 
 
 
 
56 
 
6. 
As clinically indicated. or as consented by patient for research only 
7. 
Bone marrow aspirates/biopsies should be sent for pathology analysis as clinically indicated and according to the protocol. A 5-10 ml aliquot of the bone marrow aspirate in sodium heparin 
should be sent to the Berger Lab for research if done in conjunction with clinical procedure or if patient consented to reseach only 
8. 
Diagnostic CT/PET scan to include neck, chest, abdomen and pelvis, as clinically indicated by disease and status and according to the protocol. Staging evaluations may be ceased if the 
patient proceeds to other anti-tumor therapy. 
9. 
Immunophenotyping (Cohort A) may be omitted after day 28 in patients who proceed to other systemic therapies, with a recommendation to be made to the treating physician to follow Ig 
levels and replace if necessary. 
10. Blood for archival purposes may be collected as follows: day 0-21 and 21-90, days120, 180 and 365. 
 
T cell infusion time points represent guidelines for performance of required evaluations. Due to numerous factors influencing scheduling (pt and provider availability, testing 
services limitations etc.), variation in evaluation performance dates is anticipated and acceptable to the protocol (e.g., within +/- 7 days of time points < day 35; +/- 30 days for 
time points > day 35).

 
 
 
 
57 
 
APPENDIX E - Research sample checklist  
 
 RECIPIENT RESEARCH EVALUATIONS BEFORE T CELL INFUSION  
SAMPLE 
 
TIME 
TEST 
TUBE 
VOL. 
LAB 
Blood, 
lymph 
node, 
tumor 
specimen or 
bone 
marrow 
Time of 
initial 
Screening 
Confirmation of ROR1 expression 
by flow cytometry or IHC 
Sodium 
heparin 
(green) 
5 ml 
UW 
Hematopath 
lab 
Blood 
Screening 
Review of lymphocyte subsets 
Sodium 
heparin 
(green) 
10 ml 
Berger, 
D3-276 
Blood 
At 
Enrollment 
Baseline T cell persistence 
Sodium 
heparin 
(green) 
30 ml 
Berger, 
D3-276 
Blood 
At 
Enrollment 
Serum cytokine levels 
Serum 
separator 
5ml 
Berger, 
D3-276 
Blood 
At 
Enrollment 
Baseline transgene 
immunogenicity 
Serum 
separator 
5 ml 
Berger, 
D3-276 
Bone 
Marrow 
(if 
collected) 
At 
Enrollment 
Research studies 
Sodium 
heparin 
(green) 
5-10 
ml 
Berger, 
D3-276 
Any tissue  
(if 
collected) 
(requires 
specific 
consent if 
for 
research 
only) 
At 
Enrollment 
Research studies 
 
Formalin 
varies 
Berger, 
D3-276 
Blood 
Day 0 
Serum cytokine levels 
Serum 
separator 
5 ml 
Berger, 
D3-276 
Blood 
Day 0 
Baseline HACA 
Serum 
separator 
5 ml 
Berger, 
D3-276 
Blood 
Day 0 
T cell baseline persistence 
Sodium 
heparin 
(green) 
30 ml 
Berger, 
D3-276 
 
RECIPIENT RESEARCH EVALUATIONS AFTER T CELL INFUSION.  
 
SAMPLE 
 
TIME 
AFTER 
FINAL 
INFUSION 
TEST 
TUBE 
VOL. 
LAB 
 
 
 
 
 
 

 
 
 
 
58 
 
 
 
 
 
 
 
Blood 
Day 1 
Serum cytokine levels 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 1 
T cell persistence 
Sodium 
heparin 
(green) 
30 ml 
Berger, 
D3-276 
 
 
 
 
 
 
Blood 
Day  4 
Serum cytokine levels 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 4  
T cell persistence 
Sodium 
heparin 
(green) 
 
30 ml 
Berger, 
D3-276 
Blood 
Day 7 
Serum cytokine levels 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 7 
T cell persistence 
Sodium 
heparin 
(green) 
30 ml 
Berger, 
D3-276 
Blood 
Day 7 
HACA 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 10 
Serum cytokine levels 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 10 
T cell persistence 
Sodium 
heparin 
(green) 
 
30 ml 
Berger, 
D3-276 
Blood 
Day 14 
Serum cytokine levels 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 14 
T cell persistence 
Sodium 
heparin 
(green) 
30 ml 
Berger, 
D3-276 
Blood 
Day 14 
HACA 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 21 
Serum cytokine levels 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 21 
T cell persistence 
Sodium 
heparin 
(green) 
30 ml 
Berger, 
D3-276 
Blood 
Day 21 
HACA 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 0-21 
(optional) 
PBMC archive  
Sodium 
heparin 
(green) 
60 ml 
Berger, 
D3-276 
Blood 
Day 28 
Serum cytokine levels 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 28 
T cell persistence 
Sodium 
heparin 
(green) 
30 ml 
Berger, 
D3-276 

 
 
 
 
59 
 
Blood 
Day 28 
HACA 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 28 
PBMC transgene immunogenicity 
Sodium 
heparin 
(green) 
20 ml  
Berger, 
D3-276 
Blood 
Day 28 
Serum transgene immunogenicity 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Any tissue 
(requires 
specific 
consent if 
for 
research 
only) 
At restaging 
Research studies 
 
Formalin 
varies 
Berger, 
D3-276 
 
 
 
 
 
 
Blood 
Day 35 
Serum cytokine levels 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 35 
HACA 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 35 
         T cell persistence 
Sodium 
heparin 
(green) 
30 ml 
Berger, 
D3-276 
Blood 
 Weekly 
after day 35 
Serum cytokine levels 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 60 
T cell persistence 
Sodium 
heparin 
(green) 
30 ml 
Berger, 
D3-276 
Blood 
Day 60 
HACA 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 60 
PBMC transgene immunogenicity 
Sodium 
heparin 
(green) 
20 ml 
Berger, 
D3-276 
Blood  
Day 60 
Serum transgene immunogenicity 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 21-90 
(optional) 
PBMC archive 
Sodium 
heparin 
(green) 
60 ml 
Berger, 
D3-276 
Blood 
Day 90 
HACA 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 120 
PBMC archive 
Sodium 
heparin 
(green) 
20 ml 
Berger, 
D3-276 
Blood 
Day 120 
Serum archive 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 120 
T cell persistence 
Sodium 
heparin 
(green) 
30 ml 
Berger, 
D3-276 
Blood 
Day 120 
PBMC transgene immunogenicity 
Sodium 
heparin 
(green) 
20 ml 
Berger, 
D3-276 
Blood 
Day 120 
Serum transgene immunogenicity  
Serum 
10ml  
Berger, 

 
 
 
 
60 
 
separator  
D3-276 
Blood 
Day 180 
T cell persistence 
Sodium 
heparin 
(green) 
30 ml 
Berger, 
D3-276 
Blood 
Day 180 
HACA 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 180 
PBMC transgene immunogenicity 
Sodium 
heparin 
(green) 
20 ml 
Berger, 
D3-276 
Blood 
Day 180 
Serum transgene immunogenicity 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 180 
PBMC archive 
Sodium 
heparin 
(green) 
20 ml 
Berger, 
D3-276 
Blood 
Day 180 
Serum archive 
Serum 
separator 
10 ml 
Berger, 
D3-276 
 
 
 
 
 
 
Blood 
Day 365 
T cell persistence 
Sodium 
heparin 
(green) 
30 ml 
Berger, 
D3-276 
Blood 
Day 365 
PBMC transgene immunogenicity 
Sodium 
heparin 
(green) 
20 ml 
Berger, 
D3-276 
Blood 
Day 365 
Serum transgene immunogenicity 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Blood 
Day 365 
PBMC archive 
Sodium 
heparin 
(green) 
20 ml 
Berger, 
D3-276 
Blood 
Day 365 
Serum archive 
Serum 
separator 
10 ml 
Berger, 
D3-276 
 
 
 
 
 
 
Blood 
Clinical 
events 
T cell persistence 
Sodium 
heparin 
(green) 
30 ml 
Berger, 
D3-276 
Blood 
Clinical 
events 
Serum cytokine levels 
Serum 
separator 
10 ml 
Berger, 
D3-276 
Bone 
marrow 
Any clinical 
time point 
Migration 
Sodium 
heparin 
(green) 
5-10 ml 
Berger, 
D3-276 
Tissue 
samples 
Any clinical 
time point 
Migration 
RPMI 
varies 
Berger, 
D3-276 
 
 
 
 
 
 
 
 
 

 
 
 
 
61 
 
 
 
 
 
APPENDIX F - Long Term Follow-Up 
 
Study participants should be asked to participate in long term follow-up, as directed by the FDA 
Guidance for Industry – Gene Therapy Clinical Trials: Observing Subjects for Delayed Adverse 
Events. 
(http://www.fda.gov/BiologicsBloodVaccines/GuidanceComplianceRegulatoryInformation/Guida
nces/CellularandGeneTherapy/ucm072957.htm#5). 
 
Long term follow-up should commence one year after the final T cell infusion. The planned 
recommendations for follow-up are as follows:  
 
Years 1 - 15: 
1. Recommendation that patients undergo at least annual history and physical examination with 
their primary physician: 
• Adverse event screening guidance for the primary physician in the form of a gene 
therapy LTFU-directed screening survey may be available. 
• A request for the study team to be notified of all new malignancies and unexpected 
illnesses. 
• The primary physician may be provided with a blood draw courier kit to enable 
samples to be returned to the Berger Lab for archival purposes, and for analysis for 
transgene and vector persistence, and RCL, as dictated by studies of transferred T cell 
persistence. 
2. Annual phone call survey or questionnaire to the participant to screen for adverse events. 
3. Offer the opportunity to return to FHCRC for an annual LTFU clinic visit. 
4. Compliance with 21 CFR 312.32 in adverse event reporting. 
5. Research studies 
5.i.  Evaluation for transgene vector sequence by PCR of PBMC every 6 months for years 
1 to 5 and every year for years 6-15 until the transgene becomes undetectable.  
5.ii. If > 1% of cells express the transgene or if clonality is suggested, vector integration 
sites or TCRB sequence utilization may be analyzed in PBMC, CAR-T cells or other 
tissue. If clonality is suggested, repeat testing may be performed 3 months later. 
Persistent monoclonality, clonal expansion or vector integration near a known 
oncogenic locus should precipitate careful attention to the possibility of malignancy. 
However, the need for additional intervention should be guided by the clinical 
circumstances and not solely by the presence of these factors. 
5.iii. Annual testing of PBMC for RCL by VSVG QPCR. If there is no evidence of 
transgene persistence, RCL assays may be suspended after one year and samples may 
be archived.  
 
 
 
 
 

 
 
 
 
62 
 
 
 
 
 
APPENDIX G– Grading Criteria for CRS 
 
Grade 
Description of Symptoms 
1: Mild 
Not life-threatening, require only symptomatic treatment such as antipyretics and anti-
emetics (e.g., fever, nausea, fatigue, headache, myalgia, malaise) 
2: Moderate 
Require and respond to moderate intervention: 
• 
Oxygen requirement < 40%, or 
• 
Hypotension responsive to fluids or low dose of a single vasopressor, or 
• 
Grade 2 organ toxicity (by CTCAE v4.03) 
3: Severe 
Require and respond to aggressive intervention: 
• 
Oxygen requirement ≥ 40%, or 
• 
Hypotension requiring high dose of a single vasopressor (e.g., norepinephrine 
≥ 20 µg/min, dopamine ≥ 10 µg/kg/min, phenylephrine ≥ 200 µg/min, or 
epinephrine ≥ 10 µg/min), or 
• 
Hypotension requiring multiple vasopressors (e.g., vasopressin + one of the above 
agents, or combination vasopressors equivalent to ≥ 20 µg/min norepinephrine), or 
• 
Grade 3 organ toxicity or Grade 4 transaminitis (by CTCAE v4.03) 
4: Life-threatening 
Life-threatening: 
• 
Requirement for ventilator support, or 
• 
Grade 4 organ toxicity (excluding transaminitis) 
5: Fatal 
Death 
Adapted from Lee et al., 2014 (Lee 2014) 
 

 
 
 
 
63 
 
 
APPENDIX H – Recommended management guidelines for CRS and neurotoxicity