Clinical trial NCT01454596
Conditions: Malignant Glioma Glioblastoma Brain Cancer Gliosarcoma
Keywords: Cell Therapy Gene Therapy Immunotherapy Brain Cancer Glioma Glioblastoma
Sponsor: National Cancer Institute (NCI)
Start date: 2012-05-16; Completion date: 2019-01-17
Enrollment: 18 participants; Allocation: NON_RANDOMIZED; Intervention model: SEQUENTIAL

Brief summary:
Background:

The National Cancer Institute (NCI) Surgery Branch has developed an experimental therapy for treating patients with gliomas that involves taking white blood cells from the patient, growing them in the laboratory in large numbers, genetically modifying these specific cells with a type of virus (retrovirus) to attack only the tumor cells, and then giving the cells back to the patient. This type of therapy is called gene transfer. In this protocol, we are modifying the patient's white blood cells with a retrovirus that has the gene for epidermal growth factor receptor (EGFR) vIII incorporated in the retrovirus.

Objective:

The purpose of this study is to determine a safe number of these cells to infuse and to see if these particular tumor-fighting cells (anti-EGFRvIII cells) are a safe and effective treatment for advanced gliomas.

Eligibility:

\- Adults age 18-70 with malignant glioma expressing the EGFRvIII molecule.

Design:

Work up stage: Patients will be seen as an outpatient at the National Institutes of Health (NIH) clinical Center and undergo a history and physical examination, scans, x-rays, lab tests, and other tests as needed

Leukapheresis: If the patients meet all of the requirements for the study they will undergo leukapheresis to obtain white blood cells to make the anti-EGFRvIII cells. {Leukapheresis is a common procedure, which removes only the white blood cells from the patient.}

Treatment: Once their cells have grown, the patients will be admitted to the hospital for the conditioning chemotherapy, the anti-EGFRvIII cells, and aldesleukin. They will stay in the hospital for about 4 weeks for the treatment.

Follow up: Patients will return to the clinic for a physical exam, review of side effects, lab tests, and scans every month for the first year, and then every 1-2 months as long as their tumors are shrinking. Follow up visits will take up to 2 days.

Detailed description:
BACKGROUND:

\- Patients with recurrent gliomas have very limited treatment options. Epidermal growth factor receptor (EGFR).

(EGFRvIII) is the most common mutant variant of EGFR and is present in 24-67% of patients with glioblastoma.

* EGFRvIII expression promotes oncogenesis and is associated with poor prognosis.
* EGFRvIII is not expressed in normal tissue and is an attractive target for immunotherapy.
* We have constructed a retroviral vector that contains a chimeric antigen receptor (CAR) that recognizes the EGFRvIII tumor antigen, which can be used to mediate genetic transfer of this CAR with high efficiency without the need to perform any selection.

OBJECTIVES:

Primary Objectives

* To evaluate the safety of the administration of anti-EGFRvIII CAR engineered peripheral blood lymphocytes in patients receiving the non-myeloablative, lymphodepleting preparative regimen and aldesleukin.
* Determine the six month progression free survival of patients receiving anti-EGFRvIII CAR-engineered peripheral blood lymphocytes and aldesleukin following a nonmyeloablative, lymphodepleting preparative regimen.

ELIGIBILITY:

* Histologically proven glioblastoma or gliosarcoma expressing EGFRvIII as determined by immunohistochemistry (IHC) or Reverse transcription polymerase chain reaction (RT-PCR)
* Failed prior standard treatment with radiotherapy with or without chemotherapy
* Karnofsky performance score (KPS) greater than or equal to 60
* Cardiac, pulmonary and laboratory parameters within acceptable limits

DESIGN:

* The study will be conducted using a Phase I/II design.
* Patients will receive a non-myeloablative, lymphodepleting preparative regimen consisting of cyclophosphamide and fludarabine followed by intravenous infusion of ex vivo tumorreactive, CAR gene-transduced peripheral blood mononuclear cells (PBMC), plus intravenous (IV) aldesleukin.
* Once the maximum tolerated cell dose (MTD) has been determined, the study will proceed to the phase II portion.
* In the phase II portion of the trial, patients will be accrued to two cohorts:

  * Patients with recurrent malignant glioma receiving steroids at the time of treatment.
  * Patients with recurrent malignant glioma not receiving steroids at the time of treatment.
* A total of 107 patients may be enrolled over a period of 7 years.

Arms: arm 1: Non-myeloablative lymphodepleting preparative regimen of cyclophosphamide and fludarabine + escalating doses of epidermal growth factor receptor (EGFRv)III Chimeric antigen receptor (CAR) transduced peripheral blood lymphocytes (PBL) + aldesleukin arm 2: Non-myeloablative lymphodepleting preparative regimen of cyclophosphamide and fludarabine + maximum tolerated dose (MTD) of anti-EGFRvIII CAR transduced PBL established in Phase I + aldesleukin
Interventions: intervention 1: Epidermal growth factor receptor(EGFRv)III Chimeric antigen receptor (CAR) transduced PBL intervention 2: Aldesleukin intervention 3: Fludarabine intervention 4: Cyclophosphamide
Intervention descriptions: intervention 1: Day 0: Cells will be infused intravenously over 20-30 minutes. Patients will receive two cell doses, 2 hours apart. intervention 2: Aldeskeukin 72,000 IU /kg intravenous (IV) or 720,000 IU /kg IV (based on total body weight) over 15 minutes every eight hours (+/- 1 hour) beginning within 24 hours of cell infusion and continuing for up to 5 days (maximum 15 doses). intervention 3: Days -7 to -3: Fludarabine 25 mg /m(2)/day intravenous piggyback (IVPB) daily over 30 minutes for 5 days. intervention 4: Days -7 and -6: Cyclophosphamide 60 mg/kg/day X 2 days IV in 250 ml dextrose 5% in water (D5W) with Mesna 15 mg/kg /day X 2 days over 1 hr.

Protocol documents:
Prot_SAP_000.pdf: 
Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
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Abbreviated Title: Anti-EGFRvIII CAR PBL  
CC Protocol Number: 11-C-0266 Q 
IBC Protocol Number: RD-11-III-06 
OSP Protocol Number: 1103-1095 
NCT Number: NCT01454596 
Version Date: June 28, 2018 
PROTOCOL TITLE 
A Phase I/II Study of the Safety and Feasibility of Administering T-Cells Expressing Anti-
EGFRvIII Chimeric Antigen Receptor to Patients with Malignant Gliomas Expressing EGFRvIII 
 
NIH Principal Investigator:  
Steven A. Rosenberg, M.D., Ph.D. 
   
 
 
 
 
 
Chief of Surgery, Surgery Branch, CCR, NCI 
 
 
 
 
 
 
 
Building 10, CRC, Room 3-3940 
  
9000 Rockville Pike, Bethesda, MD 20892 
 
 
 
 
 
Phone: 240-760-6218; Email: Steven.Rosenberg@nih.gov 
 
Investigational Agent: 
Drug Name: 
Anti-EGFRvIII CAR (PG13-139-CD8-
CD28BBZ (F10) transduced PBL  
IND Number: 
14767 
Sponsor: 
Center for Cancer Research 
Manufacturer: Surgery Branch Cell Production Facility 
 
Commercial Agents: Cyclophosphamide, Fludarabine, and Aldesleukin 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
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PRÉCIS 
Background: 
• Patients with recurrent gliomas have very limited treatment options. EGFR variant III 
(EGFRvIII) is the most common mutant variant of EGFR and is present in 24-67% of 
patients with glioblastoma.  
• EGFRvIII expression promotes oncogenesis and is associated with poor prognosis.  
• EGFRvIII is not expressed in normal tissue and is an attractive target for immunotherapy. 
• We have constructed a retroviral vector that contains a chimeric antigen receptor (CAR) 
that recognizes the EGFRvIII tumor antigen, which can be used to mediate genetic 
transfer of this CAR with high efficiency without the need to perform any selection. 
Objectives: 
• Primary objectives: 
o To evaluate the safety of the administration of anti-EGFRvIII CAR engineered 
peripheral blood lymphocytes in patients receiving the non-myeloablative, 
lymphodepleting preparative regimen and aldesleukin. 
o Determine the six-month progression free survival of patients receiving anti-
EGFRvIII CAR-engineered peripheral blood lymphocytes and aldesleukin following 
a non-myeloablative, lymphodepleting preparative regimen.  
Eligibility: 
• Histologically proven glioblastoma or gliosarcoma expressing EGFRvIII as determined 
by IHC or RT-PCR 
• Failed prior standard treatment with radiotherapy with or without chemotherapy 
• Karnofsky performance score (KPS) ≥ 60 
• Cardiac, pulmonary and laboratory parameters within acceptable limits  
Design: 
• The study will be conducted using a Phase I/II design.  
• Patients will receive a non-myeloablative, lymphodepleting preparative regimen 
consisting of cyclophosphamide and fludarabine followed by intravenous infusion of ex 
vivo tumor-reactive, CAR gene-transduced PBMC, plus IV aldesleukin. 
• Once the maximum tolerated cell dose (MTD) has been determined, the study will 
proceed to the phase II portion.  
• In the Phase II portion of the trial, patients will be accrued to two cohorts: 
o Patients with recurrent malignant glioma receiving steroids at the time of treatment. 
o Patients with recurrent malignant glioma not receiving steroids at the time of 
treatment. 
• A total of 107 patients may be enrolled over a period of 7 years. 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
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TABLE OF CONTENTS 
PRÉCIS .......................................................................................................................................... 2 
TABLE OF CONTENTS ............................................................................................................. 3 
1 
INTRODUCTION................................................................................................................. 8 
1.1 
Study Objectives ................................................................................................................. 8 
1.1.1 Primary Objectives.......................................................................................................... 8 
1.1.2 Secondary Objective ....................................................................................................... 8 
1.2 
Background and Rationale .................................................................................................. 8 
1.2.1 Gliomas ........................................................................................................................... 8 
1.2.2 EGFRvIII and Glioblastomas ......................................................................................... 9 
1.2.3 Preclinical Studies to Provide the Rationale for this Clinical Protocol ........................ 10 
1.2.4 Prior NCI-SB Trials of Cell Transfer Therapy Using Tumor-Infiltrating Lymphocytes 
(TIL) in Patients with Metastatic Melanoma ............................................................................ 15 
1.2.5 NCI-SB Trials of Cell Transfer Therapy Using Transduction of Anti-TAA TCR Genes 
into Non-reactive TIL or PBL .................................................................................................. 17 
1.2.6 Safety Considerations ................................................................................................... 19 
2 
ELIGIBILITY ASSESSMENT AND ENROLLMENT .................................................. 21 
2.1 
Eligibility Criteria ............................................................................................................. 21 
2.1.1 Inclusion Criteria .......................................................................................................... 21 
2.1.2 Exclusion Criteria ......................................................................................................... 22 
2.2 
Screening Evaluation ........................................................................................................ 23 
2.2.1 Within 3 Months Prior to Starting the Preparative Regimen ........................................ 23 
2.2.2 Within 8 Weeks Prior to Starting the Preparative Regimen ......................................... 23 
2.2.3 Within 4 Weeks Prior to Starting the Preparative Regimen ......................................... 23 
2.2.4 Within 14 Days Prior to Starting the Preparative Regimen .......................................... 23 
2.2.5 Within 7 Days Prior to Starting the Preparative Regimen ............................................ 23 
2.2.6 Within 1-4 Days Prior to Starting the Preparative Regimen ......................................... 24 
2.3 
Registration and Treatment Assignment Procedures ........................................................ 24 
2.3.1 Prior to Registration for this Protocol ........................................................................... 24 
2.3.2 Registration Procedure .................................................................................................. 24 
2.3.3 Treatment Assignment Procedures ............................................................................... 24 
3 
STUDY IMPLEMENTATION .......................................................................................... 25 
3.1 
Study Design ..................................................................................................................... 25 
3.1.1 Schema – Beginning with Amendment K .................................................................... 25 
3.1.2 Pre-Treatment Phase: Cell Preparation Performed on 03-C-0277 ................................ 26 
3.1.3 Treatment Phase ............................................................................................................ 26 
3.1.4 Dose-Limiting Toxicity ................................................................................................ 28 
3.1.5 Phase II Portion ............................................................................................................. 29 


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3.2 
Protocol Stopping Rules ................................................................................................... 29 
3.3 
Drug Administration ......................................................................................................... 30 
3.3.1 Preparative Regimen with Cyclophosphamide and Fludarabine .................................. 30 
3.3.2 Cell Infusion.................................................................................................................. 31 
3.3.3 Aldesleukin Administration .......................................................................................... 32 
3.3.4 Treatment Schedule ...................................................................................................... 32 
3.4 
Baseline Evaluation .......................................................................................................... 33 
3.4.1 Within 14 Days Prior to Starting the Preparative Regimen .......................................... 33 
3.5 
On-Study Evaluation ......................................................................................................... 33 
3.5.1 During the Preparative Regimen (Daily) ...................................................................... 33 
3.5.2 Post-Cell Infusion ......................................................................................................... 34 
3.5.3 During Aldesleukin Administration (Daily) ................................................................. 34 
3.5.4 During Hospitalization (Every 1-2 Days) ..................................................................... 34 
3.6 
Post-Treatment (Follow-up) Evaluation ........................................................................... 34 
3.6.1 Time-Period of Evaluations .......................................................................................... 35 
3.6.2 Scheduled Evaluations .................................................................................................. 35 
3.7 
Study Assessment Calendar .............................................................................................. 37 
3.8 
Criteria for Removal from Protocol Therapy and Off-Study Criteria .............................. 40 
3.8.1 Criteria for Removal from Protocol Therapy................................................................ 40 
3.8.2 Off-Study Criteria ......................................................................................................... 40 
3.8.3 Off Protocol Therapy and Off-Study Procedure ........................................................... 40 
4 
CONCOMITANT MEDICATIONS/MEASURES .......................................................... 40 
4.1 
Mannitol ............................................................................................................................ 41 
4.2 
Non-Enzyme Inducing Anti-Epileptic Drugs (Non-EIAED) ........................................... 41 
4.3 
Infection Prophylaxis ........................................................................................................ 41 
4.3.1 Pneumocystis Jirovecii Pneumonia............................................................................... 41 
4.3.2 Herpes Simplex or Varicella Zoster Virus Prophylaxis ................................................ 41 
4.3.3 Fungal Prophylaxis (Fluconazole) ................................................................................ 42 
4.3.4 Empiric Antibiotics ....................................................................................................... 42 
4.4 
Blood Product Support ...................................................................................................... 42 
5 
BIOSPECIMEN COLLECTION ...................................................................................... 42 
5.1 
Samples Sent to Figg Lab ................................................................................................. 42 
5.2 
Samples Sent to Surgery Branch Cell Production Facility ............................................... 43 
5.3 
Prior to Chemotherapy Administration ............................................................................. 43 
5.4 
Prior to Cell Infusion (Baseline Sample for Cytokine Analysis) ...................................... 43 
5.5 
Post-Cell Infusion Evaluations ......................................................................................... 43 
5.6 
Sample Collection Schedule ............................................................................................. 43 
5.7 
Immunological Testing ..................................................................................................... 44 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
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5.8 
Monitoring Gene Therapy Trials: Persistence and RCR .................................................. 44 
5.9 
Sample Storage, Tracking And Disposition For Surgery Branch Cell Production Facility
 
44 
5.10 
Sample Storage, Tracking and Disposition For Figg Lab ................................................. 45 
5.10.1 
Sample Data Collection ............................................................................................ 45 
5.10.2 
Sample Storage and Destruction ............................................................................... 45 
6 
DATA COLLECTION AND EVALUATION ................................................................. 46 
6.1 
Data Collection ................................................................................................................. 46 
6.1.1 Exclusions to Routine Adverse Event Recording ......................................................... 47 
6.2 
Data Sharing Plans ............................................................................................................ 47 
6.2.1 Human Data Sharing Plan ............................................................................................. 47 
6.2.2 Genomic Data Sharing Plan .......................................................................................... 47 
6.3 
Response Criteria .............................................................................................................. 47 
6.3.1 Definitions..................................................................................................................... 47 
6.3.2 Disease Parameters ....................................................................................................... 48 
6.3.3 Evaluation of Measurable Lesions ................................................................................ 48 
6.3.4 Definition of Response for Evaluable Lesions ............................................................. 49 
6.3.5 Best Response ............................................................................................................... 50 
6.3.6 Central Radiology Review ............................................................................................ 50 
6.3.7 Neurological Examination ............................................................................................ 50 
6.3.8 Progression-Free Survival ............................................................................................. 50 
6.3.9 Overall Survival ............................................................................................................ 50 
6.4 
Toxicity Criteria ................................................................................................................ 50 
7 
SAFETY REPORTING REQUIREMENTS/DATA AND SAFETY MONITORING 
PLAN ............................................................................................................................................ 51 
7.1 
Definitions......................................................................................................................... 51 
7.1.1 Adverse Event ............................................................................................................... 51 
7.1.2 Suspected Adverse Reaction ......................................................................................... 51 
7.1.3 Unexpected Adverse Reaction ...................................................................................... 51 
7.1.4 Serious........................................................................................................................... 51 
7.1.5 Serious Adverse Event .................................................................................................. 51 
7.1.6 Disability ....................................................................................................................... 52 
7.1.7 Life-Threatening Adverse Drug Experience ................................................................. 52 
7.1.8 Protocol Deviation (NIH Definition) ............................................................................ 52 
7.1.9 Non-Compliance (NIH Definition) ............................................................................... 52 
7.1.10 
Unanticipated Problem.............................................................................................. 52 
7.2 
NCI-IRB and Clinical Director (CD) Reporting ............................................................... 52 
7.2.1 NCI-IRB and NCI CD Expedited Reporting of Unanticipated Problems and Deaths . 52 


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7.2.2 NCI-IRB Requirements for PI Reporting at Continuing Review ................................. 53 
7.2.3 NCI-IRB Reporting of IND Safety Reports.................................................................. 53 
7.3 
IND Sponsor Reporting Criteria ....................................................................................... 53 
7.3.1 Waiver of Expedited Reporting to CCR ....................................................................... 53 
7.3.2 Reporting Pregnancy ..................................................................................................... 54 
7.4 
Institutional Biosafety Committee (IBC) Reporting Criteria ............................................ 55 
7.4.1 Serious Adverse Event Reports to IBC ......................................................................... 55 
7.4.2 Annual Reports to IBC.................................................................................................. 55 
7.5 
Data and Safety Monitoring Plan ...................................................................................... 56 
7.5.1 Principal Investigator/Research Team .......................................................................... 56 
7.5.2 Sponsor Monitoring Plan .............................................................................................. 56 
7.5.3 Safety Monitoring Committee (SMC) .......................................................................... 57 
8 
STATISTICAL CONSIDERATIONS .............................................................................. 57 
9 
COLLABORATIVE AGREEMENTS.............................................................................. 59 
10 HUMAN SUBJECTS PROTECTIONS ............................................................................ 59 
10.1 
Rationale for Subject Selection ......................................................................................... 59 
10.2 
Participation of Children ................................................................................................... 59 
10.3 
Participation of Subjects Unable to Give Consent ............................................................ 59 
10.4 
Evaluation of Benefits and Risks/Discomforts ................................................................. 60 
10.5 
Risk/Benefit Analysis ....................................................................................................... 60 
10.6 
Consent Process and Documentation ................................................................................ 60 
10.6.1 
Informed Consent of Non-English Speaking Subjects ............................................. 60 
11 PHARMACEUTICAL INFORMATION ......................................................................... 61 
11.1 
Investigational Regimen ................................................................................................... 61 
11.1.1 
Anti-EGFRvIII CAR Transduced PBL ..................................................................... 61 
11.1.2 
Retroviral Vector Containing the anti-EGFRvIII CAR Gene................................... 61 
11.1.3 
Interleukin-2 (Aldesleukin, Proleukin, Recombinant Human Interleukin 2) ........... 62 
11.1.4 
Fludarabine ............................................................................................................... 62 
11.1.5 
Cyclophosphamide .................................................................................................... 63 
11.2 
Support Medications ......................................................................................................... 64 
11.2.1 
Mesna (Sodium 2-mercaptoethanesulfonate, Mesnum, Mesnex, NSC-113891) ...... 64 
11.2.2 
Filgrastim (Granulocyte Colony-Stimulating Factor, G-CSF, Filgrastim, Neupogen)
 
64 
11.2.3 
Trimethoprim and Sulfamethoxazole Double Strength (TMP/SMX DS) ................ 65 
11.2.4 
Herpes Simplex and Varicella Zoster Virus Prophylaxis ......................................... 66 
11.2.5 
Fluconazole ............................................................................................................... 66 
11.2.6 
Ondansetron Hydrochloride ...................................................................................... 67 
11.2.7 
Furosemide ................................................................................................................ 67 


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12 REFERENCES .................................................................................................................... 68 
13 APPENDICES ..................................................................................................................... 72 
13.1 
Appendix A: Modification of Dose Calculations* in Patients whose BMI is Greater Than 
35 
72 
13.2 
Appendix B: Adverse Events Occurring in ≥10% of Patients Treated with Aldesleukin 
(n=525)1 ........................................................................................................................................ 73 
13.3 
Appendix C: Expected IL-2 Toxicities and their Management ........................................ 74 
13.4 
Appendix D: Interleukin-2 Toxicities Observed in Patients Treated at the NIH Clinical 
Center 77 
13.5 
Appendix E: Certificate of Analysis – anti-EGFRvIII CAR Transduced T-Cells ............ 78 
13.6 
Appendix F: Karnofsky Performance Score (KPS) .......................................................... 79 
13.7 
Appendix G: Anti-Epileptic Medications ......................................................................... 80 
 
 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
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1 
INTRODUCTION 
1.1 
STUDY OBJECTIVES 
1.1.1 Primary Objectives 
• To evaluate the safety of the administration of anti-EGFRvIII CAR engineered peripheral 
blood lymphocytes in patients receiving the non-myeloablative, lymphodepleting 
preparative regimen and aldesleukin 
• Determine the six-month progression free survival of patients receiving anti-EGFRvIII 
CAR-engineered peripheral blood lymphocytes and aldesleukin following a non-
myeloablative, lymphodepleting preparative regimen.  
1.1.2 Secondary Objective 
• Determine the in vivo survival of CAR gene-engineered cells. 
• Evaluate radiographic changes after treatment 
1.2 
BACKGROUND AND RATIONALE 
1.2.1 Gliomas 
The American Cancer Society estimates that approximately 20,500 new cases of primary brain 
and nervous system tumors will develop and approximately 12,740 patients will die in the U.S. 
each year(1). Brain tumors account for approximately 85 to 90% of all central nervous system 
malignancies. Glioblastoma is the most aggressive and most common glioma accounting for 
51% of all gliomas(2). Once the primary modalities of treatment, surgery, radiation, and 
chemotherapy have failed the treatment options are limited. With the current standard of care 
including resection, radiation and chemotherapy, the median overall survival is 14.6 months(3). 
There are no good options for patients with recurrent brain tumors and participation in clinical 
trials is warranted.  
Despite recent advances in neuro-imaging, neuro-anesthesia, and neuro-surgical techniques, the 
prognosis of patients with malignant gliomas treated by surgical resection alone remains dismal 
with a median survival of 4 to 6 months(4). The low survival rates reflect the unique infiltrative 
growth characteristics of malignant gliomas, which make true “total resection” impossible 
without causing unacceptable neurological damage to the patient. To date, radiotherapy has 
proven to be the most effective treatment for malignant gliomas extending median survival to 8 
to 12 months(5-9). Adjuvant chemotherapy with temozolomide prolongs median survival to 14 
months, however tumor progression is the rule and few patients survive beyond 18 months. 
Furthermore, once these tumors have progressed, conventional chemotherapy has not been 
shown to prolong survival(10). There are multiple reasons why gliomas are relatively resistant to 
standard chemotherapy including diminished drug delivery to the tumor as a consequence of the 
blood-brain barrier, tumor hypoxia, and their relatively low growth fraction. Most importantly is 
the fact that gliomas tend to have significant intrinsic resistance to most standard cytotoxic 
agents. Clearly, novel therapies are needed, and in recent years’ immunotherapeutic approaches 
for gliomas have emerged as a strategy of interest.  
Animal studies have demonstrated that the cellular rather than the humoral arm of the immune 
response plays the major role in the elimination of murine tumors(11). Much of this evidence was 
derived from studies in which the adoptive transfer of T lymphocytes from immune animals 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
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9
could transfer resistance to tumor challenge or in some experiments, the elimination of 
established cancer. Thus, most strategies for the immunotherapy of patients with cancer have 
been directed at stimulating strong T-cell immune reactions against tumor-associated antigens. In 
contrast to antibodies that recognize epitopes on intact proteins, T-cells recognize short peptide 
fragments (8-18 amino acids) that are presented on surface class I or II major histocompatibility 
(MHC) molecules, and it has been shown that tumor antigens are presented and recognized by T-
cells in this fashion. The molecule that recognizes these peptide fragments is the T-cell receptor 
(TCR). The TCR is analogous to the antibody immunoglobulin molecule in that two separate 
proteins (the TCR alpha and beta chains) are brought together to form the functional TCR 
molecule.  
An alternate approach to enable T-cells to recognize targets is to utilize a CAR which is 
constructed by using the variable regions of the heavy and light chains of an antibody connected 
by a linker sequence and fused to T-cell signaling protein domains from CD28, CD3 zeta, and 
41BB. When a CAR is introduced into a T-cell, the T-cell can recognize and signal based on the 
non-MHC restricted recognition of the target. The goal of this protocol is to transfer peripheral 
blood lymphocytes (PBL) transduced with genes encoding a CAR that recognizes the mutant 
protein EGFRvIII that is expressed on a subset of glioma patients into autologous PBL. Normal 
PBL derived from glioma patients will be genetically engineered with a viral vector encoding the 
CAR and infused into patients with the aim of mediating regression of their tumors. This trial is 
similar to previous National Cancer Institute Surgery Branch (NCI-SB) TCR gene transfer 
adoptive immunotherapy protocols except that we will use a receptor that recognizes targets 
based on the recognition of an antibody reactive with the EGFRvIII molecule expressed on the 
tumor surface. 
1.2.2 EGFRvIII and Glioblastomas 
The epidermal growth factor receptor (EGFR) is a 170-kDa transmembrane tyrosine kinase. 
Activation of EGFR launches a host of downstream signaling cascades leading to cell 
proliferation, invasion, motility and adhesion. EGFR overexpression leads to unregulated cell 
growth and malignant transformation, and is frequently implicated in a wide range of 
malignancies.  
Increased EGFR signaling via overexpression or mutation is the most frequent genetic alteration 
associated with GBMs, occurring in 40-50% of these tumors(12, 13). The most common EGFR 
mutant is the EGF receptor variant III (EGFRvIII) which harbors an in-frame deletion of exons 
2-7. This results in a truncated extracellular ligand binding domain, but renders the protein 
constitutively active in a ligand-independent fashion(14). EGFRvIII expression has been shown to 
enhance tumorigenicity(15, 16), promote cellular motility(17), and confer resistance to radiation and 
chemotherapy(18-20). EGFRvIII is present in 24-67% of GBMs but not in normal tissue(21, 22), 
making it an attractive target for therapy. 
Preclinical studies using monoclonal antibodies (mAbs), and small-molecule inhibitors against 
EGFRvIII have provided modest results, however only peptide vaccines against EGFRvIII are 
currently offered in experimental clinical trials.  
Preclinical experience with peptide vaccines involving mice that were pre-immunized with a 
peptide vaccine against EGFRvIII(23-25) led to significant inhibition of tumor formation by cells 
expressing EGFRvIII. Moreover, immunization beginning within 4 days of tumor injection of 
tumors expressing EGFRvIII led to modest prolongation of survival. Other strategies for 


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10
EGFRvIII antigen presentation have included the use of dendritic cells (DCs) (26, 27). Again, 
preimmunization with DCs pulsed with peptide vaccine increased median survival times in mice 
compared with controls.  
Based on these preclinical data, several early clinical trials have evaluated immunization with 
EGFRvIII peptide vaccines(28, 29). A phase I clinical trial in patients with newly diagnosed GBM 
demonstrated the safety of vaccination in this population using DCs loaded with an EGFRvIII 
peptide vaccine(28). A phase II trial evaluating EGFRvIII peptide vaccine administered with GM-
CSF again showed minimal toxicity, and a favorable median time to tumor progression of 64.5 
weeks compared with 28.5 weeks in historical controls. The safety of EGFRvIII peptide vaccines 
in conjunction with standard care using temozolomide chemotherapy has also been established. 
These various trials have demonstrated tolerability of peptide vaccines and documented their 
ability to elicit antigen specific cellular and humoral responses however the effectiveness of any 
of these approaches has yet to be proven.  
There has also been preclinical work in the arena of adoptive immunotherapy using genetically 
modified T-cells. Bullian, et al. created an EGFRvIII specific CAR, MR1-ζ. They went on to 
demonstrate that human T-cells genetically modified with MR1-ζ specifically recognized 
EGFRvIII expressing tumor cells and inhibited growth in vitro(30). Similarly, Ohno, et al. 
engineered T-cells expressing a CAR against EGFRvIII based on a mouse monoclonal antibody, 
3C10. They demonstrated in vivo inhibition of intracranial tumor growth in mice(31).  
1.2.3 Preclinical Studies to Provide the Rationale for this Clinical Protocol 
Based on a review of scientific literature and publicly available databases, the amino acid 
sequences for seven monoclonal antibodies (4 murine and 3 human) specific for EGFRvIII were 
obtained and used to assemble single-chain variable fragment (scFv) genes. These scFv genes 
were inserted into gamma-retroviral vector MSGV1 using T-cell signaling domains from CD28-
CD3zeta (28Z) or, CD8-CD28-41BB-CD3zeta (28BBZ) to produce CAR expression vectors 
(Figure 1). Gamma-retroviral vectors were used to transduce human PBL and tested for 
reactivity by co-culture assays using as targets a variety of cell lines engineered to express 
EGFRvIII (there are no established cell lines that express EGFRvIII). Initial characterization of 
the seven different anti-EGFRvIII CARs suggested that a CAR based on human mAb 139 
yielded high specific reactivity against EGFRvIII expressing targets, and was thus chosen for 
development as a clinical reagent. Human mAb-based CARs may also have an advantage that 
they may not elicit an anti-CAR immune response as has been seen in some patients being 
treated with murine-based CARs. Anti-EGFRvIII mAb 139-CAR vectors were used to transduce 
PBL and CAR expression determined by FACS (Figure 1) and biological activity tested by co-
culture (Table 1).  
 


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Figure 1. Construction and analysis of anti-EGRFvIII CAR vectors. Shown on the top of the 
figure is a diagram of the two CAR vector designs showing the relative locations of the scFv and 
T-cell signaling domains. In the lower half of the figure, CAR vectors based on human mAb 139 
were used to engineer human T-cells. One week post-transduction, cells were stained with goat 
anti-human Fab and analyzed by FACS. The shift in the histogram versus untransduced cells 
(UnTd) demonstrated successful gene transfer and expression of the CD28-CD3zeta (28Z) and 
CD8-CD28-41BB-CD3zeta (28BBZ) based vectors. 
 
 
U251 
U251-EGFR 
U251-vIII 
 
Effector I 
 
 
 
 
UnTd 
0 
0 
0 
 
GFP 
0 
0 
0 
 
Anti-ERBB2 
1195 
2201 
2692 
 
139-28Z 
0 
0 
2743 
 
139-28BBZ 
0 
0 
1820 
 
 
 
 
 
 
 
U87 
U87-GFP 
U87-EGFR 
U87-vIII 
Effector II 
 
 
 
 
GFP 
203 
389 
236 
339 
Anti-ERBB2 
1959 
1061 
671 
932 
139-28Z 
759 
451 
561 
1797 
139-28BBZ 
605 
460 
499 
2217 
Table 1. Test of CARs targeting EGFR-vIII. Two donor PBL (Effector I and II) were 
transduced with anti-EGFRvIII CAR vectors containing CD28-CD3zeta (139-28Z) or CD8-


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
12
CD28-41BB-CD3zeta (139-28BBZ) signaling domains. 5-6 post transduction PBL were co-
cultured with glioma cell lines U251 and U87 that had been engineered to express wild type 
EGFR, EGFRvIII, or GFP. An anti-ERBB2 CAR served as a positive control in all co-cultures. 
Shown is resultant IFN-g production (pg/mL) following 18-hour incubation. 
 
While detection of the 28BBZ construction by FACS was less than the 28Z construct, the 
transduced T-cells were equally reactive against EGFRvIII expressing targets. We next 
determined the ability of EGFRvIII CAR engineered T-cells to lyse target cells in a standard 
51Cr-release assay (Figure 2 and Figure 3). As shown in Figure 2 and Figure 3, both vectors 
specifically lysed only cell lines engineered to express the mutant EGFRvIII and not control or 
wild-type EGFR engineered cell lines. Based on experience in the NCI-SB and in publications 
reported by others, the presence of signaling domains from the 4-1BB protein is associated with 
a better survival of CAR engineered cells in animal models. Thus, the construct containing the 4-
1BB signaling elements was chosen for clinical production.  
 
Figure 2. Specific lysis of U87 EGFRvIII by anti-EGFRvIII CAR transduced T-Cells. Anti-
EGFRvIII CAR transduced human PBL were co-cultured for 4 hours with indicated 51Cr labeled 
target tumor cell lines (parent U87, GFR, wild type EGFR, or EGFRvIII engineered). Specific 
lysis of tumor cells was measured at the given E:T ratio using the formula: [(specific release-
spontaneous release)/(total release-spontaneous release)]. Specific lysis of untransduced 
(UnTd), control GFR vector (GFP), 139-28Z CAR (vIII-28Z), and 139-28BBZ (vIII-BBZ) human 
PBL are plotted on the graph as indicated. 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
13
 
Figure 3. Specific lysis of U251 EGFRvIII by anti-EGFRvIII CAR trasduced T-cells. Anti-
EGFRvIII CAR transduced human PBL were co-cultured for 4 hours with indicated 51Cr labeled 
target tumor cell lines (parent U251, GFR, wild type EGFR, or EGFRvIII engineered). Specific 
lysis of tumor cells was measured at the given E:T ratio using the formula: [(specific release-
spontaneous release)/(total release-spontaneous release)]. Specific lysis of untransduced 
(UnTd), control GFR vector (GFP), anti-ERBB2 CAR (ERBB2), 139-28Z CAR (vIII-28Z), and 
139-28BBZ (vIII-BBZ) human PBL are plotted on the graph as indicated. 
 
By detailed molecular analysis of many different classes of cancer cell lines, it has now been 
demonstrated that established cancer cell lines often do not mirror the molecular characteristics 
of primary human cancers and this is the case for glioma lines. An alternative to the use of 
establish glioma cell lines is the analysis of tumor stem cell (TSC) lines. The TSC paradigm 
proposes that a subpopulation of cells exist in cancer that give rise to all the cells in a 
differentiated tumor. The Neuro-Oncology Branch NCI has demonstrated that in situ glioma 
cells share properties not found in glioma cell lines, and harbor features consistent with tumor 
stem cells. It was further demonstrated that marked phenotypic and genotypic differences exist 
between primary human tumor-derived TSCs and their matched glioma cell lines. TSCs derived 
directly from primary glioblastomas harbor extensive similarities to normal neural stem cells and 
recapitulate the genotype, gene expression patterns, and in vivo biology of human glioblastomas. 
These findings suggest that glioma-derived TSCs may be a more reliable model than many 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
14
commonly utilized glioma cell lines for understanding the biology of primary human tumors. We 
therefore analyzed three TSC lines for the presence of EGFRvIII and demonstrated by RT-PCR 
that EGFRvIII is expressed in these lines (Figure 4). PBL from two donors were then engineered 
with the EGFRvIII CAR (28BBZ) and co-culture with glioma TSC lines and control EGFRvIII 
expressing cell lines. Data shown in Table 2 demonstrate that EGFRvIII CAR engineered cells 
from both donors produced IFN-g following co-culture with the TSC lines. These results further 
support the use of EGFRvIII CAR engineered T-cells as a potential immunotherapy for glioma 
patients.  
 
Figure 4. Expression of EGFRvIII in glioma tumor stem cell lines. RNA from control U251 
cells (wild type EGFR or vIII engineered), TSC lines 308, 822, 1228, or these lines grown for 3 
days in serum (+symbol) were isolated and subject to RT-PCR using vIII specific primers. RT-
PCR products were subject to gel electrophoresis, and visualized with cyber green staining. The 
location of the appropriate size product bands is as indicated on the right. 
 
 
 
Media 
U251-
EGFR 
U251-
vIII 
TSC 
1228 
TSC 308 
308+serum 
TSC 822 
822+serum 
Effector I 
 
 
 
 
 
 
 
 
UnTd 
0 
0 
0 
47 
34 
0 
100 
0 
GFP 
0 
0 
0 
28 
34 
33 
30 
2 
ERBB2 
0 
2186 
2014 
1170 
1836 
2369 
187 
42 
EGFRvIII 
37 
16 
1821 
729 
473 
86 
918 
471 
 
 
 
 
 
 
 
 
 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
15
 
Media 
U251-
EGFR 
U251-
vIII 
TSC 
1228 
TSC 308 
308+serum 
TSC 822 
822+serum 
Effector II 
 
 
 
 
 
 
 
 
UnTd 
0 
0 
0 
0 
147 
0 
80 
80 
GFP 
0 
0 
0 
0 
0 
0 
180 
0 
ERBB2 
0 
4317 
4128 
2573 
3308 
4651 
1067 
259 
EGFRvIII 
384 
331 
4523 
3306 
3351 
680 
4406 
3153 
Table 2. Test of EGFR-vIII CAR recognition of glioma TSCs. Two donor PBL (Effector I and 
II) were transduced with anti-EGFRvIII CAR vector containing CD8-CD28-41BB-CD3zeta 
(EGFR-vIII) signaling domains. 5 post-transduction PBL were co-cultured with glioma TSC 
lines or cell line U251 that had been engineered to express wild type EGFR, or EGFRvIII. TSC 
lines 308 and 822 were also grown in serum for 3 days to induced differentiation (+ serum). 
GFP transduced cells served as negative control and an anti-ERBB2 CAR served as a positive 
control in all co-cultures (untransduced cells, UnTd). Shown is resultant IFN-g production 
(pg/mL) following 18-hour incubation. 
 
1.2.4 Prior NCI-SB Trials of Cell Transfer Therapy Using Tumor-Infiltrating Lymphocytes 
(TIL) in Patients with Metastatic Melanoma 
In the great majority of murine models demonstrating the therapeutic effectiveness of the 
adoptive transfer of lymphocytes mediating tumor regression, immunosuppression of the host 
prior to the adoptive transfer of lymphocytes was required. We thus incorporated a 
lymphodepleting chemotherapy into our human cell transfer studies. In the NCI-SB we 
developed a protocol to rapidly expand heterogeneous TIL for adoptive transfer. TIL were 
expanded using the rapid expansion protocol (REP) in the presence of OKT3, irradiated 
allogeneic feeder cells and IL-2. These REPed TIL retained highly specific in vitro anti-tumor 
activity, often contained reactivities against several antigenic epitopes and contained both CD8+ 
and CD4+ lymphocytes. These autologous bulk TIL were re-infused to patients following a 
nonmyeloablative chemotherapy with cyclophosphamide and fludarabine. These patients 
subsequently received high-dose IL-2 (protocol 99-C-0158). Forty-three patients received this 
treatment. This regimen resulted in objective cancer regressions in 49% of patients (21 of 43) 
with metastatic melanoma (Table 3)(32). Grade 3 and 4 toxicities are shown in Table 4. Murine 
models predicted that increasing the extent of lymphodepletion could increase the effectiveness 
of the cell transfer therapy. Thus, we performed two additional sequential trials of ACT with 
autologous TIL in patients with metastatic melanoma. Increasing intensity of host preparative 
lymphodepletion consisting of cyclophosphamide and fludarabine with either 200cGy (25 
patients) or 1200 cGy (25 patients) total body irradiation (TBI) was administered prior to cell 
transfer. While non-myeloablative chemotherapy alone showed an objective response rate of 
49%, when 200cGy or 1200cGy TBI was added the response rates were 52% and 72% 
respectively (Table 3). Complete response rates for the three trials were 13%, 20% and 40%. 
TBI appeared to result in increased patient survival. Nineteen of the 20 patients that experienced 
a complete response are ongoing from 3 to 5 years. Responses were seen in all visceral sites 
including brain.  


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
16
Host lymphodepletion was associated with increased serum levels of the lymphocyte 
homeostatic cytokines IL-7 and IL-15. Objective responses were correlated with the 
telomere length of the transferred cells. Patients exhibited the expected hematological toxicities 
associated with the cyclophosphamide, fludarabine and TBI preparative regimens. Patients 
recovered marrow function rapidly after cell infusion with absolute neutrophil counts greater 
than 500 per mm3 by day 12 and sustained platelet counts above 20,000 per mm3 by day 14 
(except 4 patients on the TBI 1200 protocol with platelet recovery on days 16, 17, 20, and 22). 
 
Table 3. Cell Transfer Therapy (8/1/10) 
 
 
 
 Treatment 
Total 
PR 
CR 
OR (%) 
 
  
 
number of patients (duration in months) 
 
 
 
 No TBI 
43 
16 
5 
21 (49%) 
 
  
 
(84, 36, 29, 28, 
14, 12, 11, 7, 
7, 7, 7, 4,  
4, 2, 2, 2) 
(82+, 81+, 79+, 
78+, 64+) 
 
 
  
 
 
 
 
 
 200 TBI 
25 
8 
5 
13 (52%) 
 
  
 
(14, 9, 6, 6, 
5, 4, 3, 3) 
(68+, 64+, 60+, 
57+, 54+) 
 
 
  
 
 
 
 
 
 1200 TBI 
25 
8 
10 
18 (72%) 
 
  
 
(21, 13, 7, 6, 
6, 5, 3, 2) 
(48+, 45+, 44+, 
44+, 39+, 38+, 
38+, 38+, 37+,  
19) 
 
 
  
 
 
 
 
 
 (52 responding patients: 42 had prior IL-2; 22 had prior IL-2 + chemotherapy) 
 
 (20 complete responses: 19 ongoing at 37 to 82 months) 
 
 
Table 4. Time in Hospital and Non-Hematological Grade 3 and 4 Toxicities Related to 
Lymphodepleting Chemotherapy and Cell Transfer 
Attribute Measured 
Duration, Number or Type 
Number of Patients (%) 
 
 
 
Days in Hospital1 
6-10 
6 (17%) 
 
11-15 
18 (51%) 
 
16-20 
4 (11%) 
 
21-25 
7 (20%) 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
17
 
 
 
pRBC Transfusions 
0 
2 (6%) 
 
1-5 
18 (51%) 
 
6-10 
13 (37%) 
 
11-15 
2 (6%) 
 
 
 
Platelet Transfusions 
0 
6 (17%) 
 
1-5 
21 (60%) 
 
6-10 
5 (14%) 
 
11-15 
2 (6%) 
 
16-20 
1 (3%) 
 
 
 
Autoimmunity 
Uveitis 
5 (14%) 
 
Vitiligo 
13 (37%) 
 
 
 
Opportunistic Infections 
Herpes zoster 
3 (9%) 
 
Pneumocystis pneumonia 
2 (6%) 
 
EBV-B cell lymphoma 
1 (3%) 
 
RSV pneumonia 
1 (3%) 
 
 
 
Other 
Febrile neutropenia 
13 (37%) 
 
Intubated for dyspnea  
3 (9%) 
 
Cortical blindness 
1 (3%) 
1Measured from the day of cell administration to discharge  
 
1.2.5 NCI-SB Trials of Cell Transfer Therapy Using Transduction of Anti-TAA TCR Genes 
into Non-reactive TIL or PBL 
It is often not possible to isolate sufficient tumor samples from melanoma patients and even 
when tumor is available, only about 60 to 70% generate melanoma reactive TIL cultures. As a 
potential alternative to the requirement to establish TIL cultures from melanoma patients, we 
sought methods that could be used to easily obtain a polyclonal population of T-cells with anti-
TAA properties. Transfer of antigen specific TCR genes to PBL is a potential method generating 
large numbers of reactive anti-cancer T-cells. In a murine model of this approach, TCR gene 
transfer into murine splenocyte T-cells was performed using a retroviral vector. The engineered 
T-cells were shown to expand in vivo upon viral challenge and efficiently homed to effector 
sites. In addition, small numbers of TCR-transduced T-cells promoted the inhibition of antigen-
expressing tumors in the mice. Retroviral vector mediated gene transfer can be used to engineer 
human T-cells with high efficiency. 
Studies in the NCI-SB identified genes that encode melanoma tumor associated antigens (TAA) 
recognized by TIL in the context of multiple MHC class I molecules (33-35). These TAA appeared 
to be clinically relevant antigens responsible for mediating tumor regression in patients with 
advanced melanoma since the TIL used to identify these antigens were often capable of 
mediating in vivo anti-tumor regression. Two antigens, which were present in virtually all fresh 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
18
and cultured melanomas, were called MART-1 (Melanoma Antigen Recognized by T-Cells - 1), 
and gp100. The genes encoding these two antigens have been cloned and sequenced. The 
MART-1 gene encodes a 118 amino acid protein of 13 kd. The gp100 gene encodes a protein 
identical to that recognized by monoclonal antibody HMB-45. These antigens were thus the 
original targets of our gene therapy cell transfer protocols in patients with metastatic melanoma. 
We thus isolated TCR genes from both gp100 and MART-1 reactive T-cells. These studies have 
recently been published and are presented briefly here(36, 37). To test the in vivo efficacy of these 
MART-1 TCR engineered T-cells, 31 HLA-A*0201 patients with progressive metastatic 
melanoma were treated. Results in the first 17 patients were published in(36). All patients were 
refractory to prior therapy with IL-2. T-cell cultures from all patients were biologically reactive, 
with specific secretion of interferon-γ following co-culture with either MART-1 peptide pulsed 
T2 cells and or melanoma cell lines expressing the MART-1 antigen. Gene transfer efficiencies 
measured by staining for V 12 expression in these lymphocytes ranged from 17% to 67% (mean 
value 42%). Four of the 31 patients demonstrated a sustained objective regression of their 
metastatic melanoma assessed by standard RECIST criteria. There were no toxicities in any 
patient attributed to the gene-marked cells. We thus demonstrated for the first time in humans, 
that normal autologous T lymphocytes, transduced ex vivo with anti-TAA TCR genes and 
reinfused in cancer patients can persist and express the transgene long-term in vivo and mediate 
the durable regression of large established tumors. A similar study was conducted using gp100 
TCR gene marked cells, however this retroviral vector had a low titer when produced under 
GMP conditions. Fourteen patients were treated on this study. No antitumor responses have been 
seen. There have been no grade 5 toxicities observed on this study, and all grade 3 and 4 
toxicities observed were expected toxicities associated with the non-myeloablative chemotherapy 
regimen or IL-2. 
The low response rate in our prior MART-1 TCR gene transfer protocol led us to identify 
MART-1 reactive TCR with higher avidity than the MART-1 F4 TCR used in the prior gene 
therapy clinical trial(37). We have now treated 24 patients with metastatic melanoma using 
autologous PBL transduced with an improved MART-1 F5 TCR following a non-myeloablative 
chemotherapy. Six patients (25%) have achieved an objective partial response. Toxicities were 
similar to those seen in the prior TCR gene therapy trial except that 15 patients developed a 
transient mild anterior uveitis easily reversed by steroid eye drops and ten patients developed 
decreased hearing reversed by middle ear steroid injections. Transient rashes have also been 
seen. There were no treatment related deaths. We have also conducted a clinical trial with a TCR 
that recognizes the gp100:154-162 melanoma peptide. This TCR was raised in an HLA-A2 
transgenic mouse immunized with this peptide(37). We have now treated 21 patients with 
metastatic melanoma using autologous PBL transduced with this improved gp100 TCR 
following a non-myeloablative chemotherapy. Four patients (19%), have achieved an objective 
partial response. Seven patients developed a transient mild anterior uveitis reversed by steroid 
eye drops and ten patients developed decreased hearing reversed by middle ear steroid injections. 
There were no treatment related deaths. 
In addition to the studies listed above in patients with metastatic melanoma, we have recently 
initiated several studies for patients with other metastatic cancers using PBL transduced with 
TCR genes or CAR genes. We have studied PBL transduced with TCR genes targeting p53, 
CEA, NY-ESO-1 and TRAIL bound to the DR4 Receptor, and PBL transduced with CAR 
targeting Her-2 and CD19. Patients on these studies also received a non-myeloablative 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
19
chemotherapy regimen consisting of cyclophosphamide and fludarabine, and high dose IL-2. In 
two studies, 08-C-0121 (anti-ESO-1 TCR) and 09-C-0082 (anti-CD19 CAR), we have seen 
impressive clinical responses. The clinical responses observed in seven of the fifteen evaluable 
patient receiving anti-ESO TCR-engineered PBL have been very encouraging. Of the six 
synovial cell sarcoma, four patients have confirmed PRs. Of the nine patients with highly ESO 
expressing melanoma, there have been 2 confirmed complete responses and 2 confirmed partial 
responses. In study 09-C-0082, an impressive ongoing partial remission at 7 months post 
treatment of the lymphoma occurred in the first patient treated on this study(38). This patient had 
heavily pre-treated, progressive follicular lymphoma that involved all major lymph node areas. 
Only three small lesions that were consistent with lymphoma persisted on positron emission 
tomography (PET) imaging post-treatment. Most toxicities observed in these studies were 
expected toxicities of the chemotherapy and aldesleukin administration. However, in 2 studies, 
we have observed serious adverse events related to the transduced cells. In 09-C-0051 (anti-Her2 
CAR transduced PBL), the first patient, with Her-2 expressing metastatic colorectal cancer, was 
treated with 1010 autologous T-cells transduced with the retrovirus encoding an anti-Her-2 CAR. 
This patient developed respiratory distress and died four days later(39). This toxicity was 
apparently due to a previously unrecognized ability of this CAR to recognize Her-2 expressed on 
lung epithelial cells although the exact explanation for the toxicity is not clear. In 09-C-0047 
(anti-CEA TCR transduced PBL), all three patients treated experienced a variety of 
gastrointestinal events which were attributed to the gene/cell therapy including diarrhea, and 
colitis. All gastrointestinal events have since resolved in these patients, and all of the patient’s 
colonic mucosa has returned to normal, and the patients have normal bowel function. Grade 3 
diarrhea lasting longer than 72 hours is considered a DLT per protocol and this event was 
observed in two of three patients enrolled in protocol 09-C-0047, meeting the criteria for 
stopping protocol accrual. One of the patients experienced an objective regression of liver 
metastases.  
1.2.6 Safety Considerations 
Several safety concerns regarding the infusion of large numbers of retrovirally modified tumor-
reactive T-cells have been addressed in our previous clinical studies. The non-myeloablative 
chemotherapy and the administration of high-dose IL-2 have expected toxicities discussed 
earlier. The non-myeloablative chemotherapy used in this protocol has been administered to over 
200 patients and all have reconstituted their hematopoietic systems. Since IL-2 can induce a 
capillary leak syndrome, patients with malignant gliomas may be at risk of increased 
intracerebral edema following IL-2 administration. Since the capillary leak syndrome occurs 
within hours of drug administration and all IL-2 administration will be done as an inpatient, any 
increases in cerebral edema will be followed closely and treated aggressively (i.e. mannitol) 
should it be clinically indicated. Due to the risks of thrombocytopenia in this patient population, 
once the platelet count drops below 30,000/mm3 platelets will be monitored and transfusions 
given as clinically indicated until the platelet count is greater than 30,000/mm3 for 24 hours 
without transfusion support. 
Although we recognize the dangers of giving a non-myeloablative chemotherapy and IL-2 to 
patients with recurrent brain tumors, and often after prior surgery, we are encouraged by our 
experience treating metastatic tumors to the brain in patients with melanoma using TIL along 
with IL-2 following the same preparative regimen used in the current protocol. Seven of 17 
patients (41%) achieved a complete response in the brain. Two of 9 patients treated using TCR 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
20
gene transduced cells (anti-MART-1 or anti-gp100) achieved a complete response. One of the 26 
patients experienced a tumor- associated subarachnoid hemorrhage during the thrombocytopenic 
phase of therapy and had an uneventful metastasectomy. 
CAR transduced human T-cells have been administered to humans in several clinical trials. Pule, 
et al. engineered human T-cells to express a CAR directed to the diasialoganglioside GD2, a 
tumor associated antigen expressed by human neuroblastoma cells(40). These CAR transduced 
cells were administered to eleven children between the ages of 3 and 10, all of whom had 
metastatic neuroblastoma. There were no adverse effects attributable to the genetically modified 
CAR T-cells in the eleven subjects that were followed for up to two years following cell 
infusion. Two patients exhibited an objective response. Persistence of the transduced cells was 
greater when the CAR were inserted into EBV specific cytolytic T lymphocytes than when the 
CAR were inserted into T-cells that were activated by an anti-CD3 monoclonal antibody.  
In the NCI-SB we have an approved clinical protocol to treat patients with B cell lymphomas 
utilizing lymphocytes transduced with a retrovirus encoding a CAR that recognizes the CD19 
molecule. Thus far six patients have been treated with between 2x108 to 4x109 cells. To date, 
three of the patients treated have exhibited partial responses and one complete response (ongoing 
at over 6 months) has been observed. As expected by the targeting of CD19, the protocol was 
associated with elimination of normal B cells. One patient required ICU support for toxicity 
similar to tumor lysis syndrome and a second patient treated on this study died prior to response 
evaluation from H1N1 influenza infection. This death is possibly related to the research but is 
not related to the anti-CD19 CAR transduced cells. 
In a second CAR at the NCI-SB a patient with metastatic colorectal cancer was treated with 1010 
autologous T-cells transduced with the retrovirus encoding an anti-Her-2 CAR. This patient 
developed respiratory distress and died four days later. This toxicity was apparently due to a 
previously unrecognized ability of this CAR to recognize Her-2 expressed on lung epithelial 
cells although the exact explanation for the toxicity is not clear. As the EGFRvIII protein is 
tumor specific, we do not expect any recognition of normal cells. In addition, as previously 
described, multiple EGFRvIII vaccine trials have been reported and no significant vaccine-
related toxicities have been observed. 
In other protocols we have administered over 3x1011 TIL with widely heterogeneous reactivity 
including CD4, CD8, and NK cells without difficulty. The expansion of tumor-reactive cells is a 
desirable outcome following the infusion of antigen reactive T-cells and this expansion could 
mimic the clonal expansion of a transformed lymphocyte. We do not believe the transfer of these 
gene modified cells has a significant risk for malignant transformation in this patient population. 
While the risk of insertional mutagenesis is a known possibility using retroviral vectors, this has 
only been observed in the setting of patients with primary immunodeficiencies using retroviral 
vector-mediated gene transfer into CD34+ bone marrow cells. In the case of retroviral vector-
mediated gene transfer into mature T-cells, there has been no evidence of long-term toxicities 
associated with these procedures since the first NCI sponsored gene transfer study in 1989. 
Although continued follow-up of all gene therapy patients will be required, data suggest that the 
introduction of retroviral vectors transduced into mature T-cells is a safe procedure. While we 
believe the risk of insertional mutagenesis is extremely low, the proposed protocol follows all 
current FDA guidelines regarding testing and follow up of patients receiving gene transduced 
cells.  


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
21
2 
ELIGIBILITY ASSESSMENT AND ENROLLMENT 
2.1 
ELIGIBILITY CRITERIA 
2.1.1 Inclusion Criteria 
2.1.1.1 Patients with histologically proven glioblastomas or gliosarcomas that express 
EGFRvIII as assessed by IHC or PCR confirmed by the NCI Laboratory of Pathology.  
2.1.1.2 Patients must have progression of disease after radiotherapy (including patients that 
undergo surgery for recurrent disease and are rendered NED). This includes recurrent 
GBM after receiving all standard first-line treatment, including surgery (if feasible due 
to neurosurgical and neuro-anatomical considerations) and adjuvant radiotherapy ± 
chemotherapy. 
2.1.1.3 Patients must either not be receiving steroids, or be on a stable dose of steroids for at 
least five days prior to registration. 
2.1.1.4 Age ≥ 18 years and ≤ 70 years. 
2.1.1.5 Ability of subject to understand and the willingness to sign a written informed consent 
document. 
2.1.1.6 Willing to sign a durable power of attorney. 
2.1.1.7 KPS ≥ 60 (see Appendix F). 
2.1.1.8 Patients of both genders must be willing to practice birth control from the time of 
enrollment on this study and for four months after treatment. 
2.1.1.9 Women of child-bearing potential must have a negative pregnancy test because of the 
potentially dangerous effects of the treatment on the fetus. 
2.1.1.10 Serology  
• Seronegative for HIV antibody. (The experimental treatment being evaluated in this 
protocol depends on an intact immune system. Patients who are HIV seropositive may 
have decreased immune-competence and thus be less responsive to the experimental 
treatment and more susceptible to its toxicities.)  
• Seronegative for hepatitis B antigen, and seronegative for hepatitis C antibody. If 
hepatitis C antibody test is positive, then patients must be tested for the presence of 
antigen by RT-PCR and be HCV RNA negative.  
2.1.1.11 Hematology 
• WBC ≥ 3000/mm3 
• ANC ≥ 1000/mm3 without the support of filgrastim 
• Platelet count ≥ 100,000/mm3 
• Hemoglobin ≥ 8.0 g/dL. Subjects may be transfused to reach this cut-off. 
2.1.1.12 Chemistry 
• Serum ALT/AST ≤ 2.5 x ULN 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
22
• Serum creatinine ≤ 1.6 mg/dL 
• Total bilirubin ≤ 1.5 mg/dL, except in patients with Gilbert’s Syndrome, who must 
have a total bilirubin < 3.0 mg/dL. 
2.1.1.13 Patients must be at least 4 weeks from radiation therapy. Additionally, patients must be 
at least 6 weeks from nitrosoureas, 4 weeks from temozolomide, 3 weeks from 
procarbazine, 2 weeks from vincristine and 4 weeks from last bevacizumab 
administration. Patients must be at least 4 weeks from other cytotoxic therapies not 
listed above and 2 weeks for non-cytotoxic agents (e.g., interferon, tamoxifen) 
including investigative agents. All toxicities from prior therapies should be resolved to 
CTCAE ≤ grade 1 (except for toxicities such as alopecia, or vitiligo). 
2.1.1.14 Subjects must be co-enrolled on protocol 03-C-0277. 
2.1.2 Exclusion Criteria 
2.1.2.1 A prior history of gliadel implantation in the past six months. 
2.1.2.2 Women of child-bearing potential who are pregnant or breastfeeding because of the 
potentially dangerous effects of the treatment on the fetus or infant. 
2.1.2.3 Active systemic infections requiring anti-infective treatment, coagulation disorders, or 
any other active or uncompensated major medical illnesses. 
2.1.2.4 Any form of primary immunodeficiency (such as Severe Combined Immunodeficiency 
Disease).  
2.1.2.5 Concurrent opportunistic infections (The experimental treatment being evaluated in this 
protocol depends on an intact immune system. Patients who have decreased immune-
competence may be less responsive to the experimental treatment and more susceptible 
to its toxicities.) 
2.1.2.6 History of severe immediate hypersensitivity reaction to cyclophosphamide, 
fludarabine, or aldesleukin.  
2.1.2.7 History of coronary revascularization or ischemic symptoms. 
2.1.2.8 Clinically significant hemorrhagic or ischemic stroke, including transient ischemic 
attacks and other central nervous system bleeding in the preceding 6 months that were 
not related to glioma surgery. History of prior intratumoral bleeding is not an exclusion 
criterion; patients who with history of prior intratumoral bleeding, however, need to 
undergo a non-contrast head CT to exclude acute bleeding.  
2.1.2.9 Other concomitant anti-cancer therapy except corticosteroids. 
2.1.2.10 Any patient known to have LVEF ≤ 45%. 
2.1.2.11 Documented FEV1 ≤ 60% predicted tested in patients with: 
• A prolonged history of cigarette smoking (≥ 20 pack-year smoking history, with 
cessation within the past two years). 
• Symptoms of respiratory dysfunction. 
2.1.2.12 Patients who are receiving any other investigational agents. 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
23
2.1.2.13 Documented LVEF ≤ 45% tested in patients: 
• Age ≥ 65 years 
• With clinically significant atrial and/or ventricular arrhythmias, including but not 
limited to: atrial fibrillation, ventricular tachycardia, second- or third-degree heart block 
or have a history of ischemic heart disease and/or chest pain. 
2.2 
SCREENING EVALUATION 
Note: Testing for screening evaluation is conducted under the NCI-SB companion protocol, 99-
C-0128 (Evaluation for NCI Surgery Branch Clinical Research Protocols). 
2.2.1 Within 3 Months Prior to Starting the Preparative Regimen 
• HIV antibody titer, HBsAg determination, and anti HCV 
• Immunohistochemistry or PCR confirmation of tumor tissue for expression of EGFRvIII 
for histologically proven glioblastomas or gliosarcomas confirmed by the NCI 
Laboratory of Pathology. (Note: Testing is permitted to be conducted at any time prior to 
starting the preparative regimen.) 
2.2.2 Within 8 Weeks Prior to Starting the Preparative Regimen 
• Pulmonary function testing for patients with a prolonged history of cigarette smoking (≥ 
20 pack-year smoking history, with cessation within the past two years), symptoms of 
respiratory dysfunction, or other clinical indications which may include thoracic 
surgeries.  
• Cardiac evaluation commensurate to patients’ history and clinical presentation (e.g., 
stress thallium, echocardiogram, MUGA) for patients who are ≥ 65 years of age, or who 
have a history of ischemic heart disease, chest pain, or clinically significant atrial and/or 
ventricular arrhythmias, including but not limited to: atrial fibrillation, ventricular 
tachycardia, heart block. Patients with a LVEF of ≤ 45% will not be eligible. Patients < 
65 years of age with cardiac risk factors (e.g., diabetes, hypertension, obesity) may 
undergo cardiac evaluations as noted above.  
2.2.3 Within 4 Weeks Prior to Starting the Preparative Regimen 
• Complete history and physical examination, including weight and vital signs, noting any 
organ system involvement and any allergies/sensitivities to antibiotics. (Note: Patient 
history may be obtained within 8 weeks prior to starting the preparative regimen). 
2.2.4 Within 14 Days Prior to Starting the Preparative Regimen 
• Screening blood tests: 
o CBC w/differential 
o Chemistries: Creatinine, ALT/GPT, AST/GOT, Total bilirubin 
• Urinalysis, with culture if indicated  
2.2.5 Within 7 Days Prior to Starting the Preparative Regimen 
• β-HCG pregnancy test (serum or urine) on all women of child-bearing potential. 


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• Physical examination, including neurological examination 
• KPS ≥ 60 (see Appendix F). 
2.2.6 Within 1-4 Days Prior to Starting the Preparative Regimen 
• Dynamic contrast-enhanced MRI scan with perfusion of the head. For patients on 
steroids, the dose must be fixed for at least 5 days prior to imaging. If the steroid dose is 
increased between the date of imaging and registration, a new baseline MRI is required. 
2.3 
REGISTRATION AND TREATMENT ASSIGNMENT PROCEDURES 
2.3.1 Prior to Registration for this Protocol 
Patients will sign the consent for and enroll on the NCI-SB companion protocol 03-C-0277 (Cell 
Harvest and Preparation for Surgery Branch Adoptive Cell Therapy Protocols), prior to 
transduction of PBL cells (either fresh or cryopreserved samples). Once cells exceed the potency 
requirement and are projected to exceed the minimum number specified in the Certificate of 
Analysis (CoA), patients will sign the consent document for this protocol.  
2.3.2 Registration Procedure 
Authorized staff must register an eligible candidate with NCI Central Registration Office (CRO) 
within 24 hours of signing consent. A registration Eligibility Checklist from the website 
(http://home.ccr.cancer.gov/intra/eligibility/welcome.htm) must be completed and sent via 
encrypted email to the NCI-CRO at ncicentralregistration-l@mail.nih.gov. After confirmation of 
eligibility at the NCI-CRO, CRO staff will call pharmacy to advise them of the acceptance of the 
patient on the protocol. Verification of registration will be forwarded electronically via email to 
the research team. A recorder is available during non-working hours.  
2.3.3 Treatment Assignment Procedures 
2.3.3.1 Cohorts 
Cohort 
Name 
Description 
1A 
Steroids (Phase I) 
Patients receiving steroids at the time of treatment. 
1B 
No Steroids (Phase I)  
Patients not receiving steroids at the time of treatment. 
2A 
Steroids (Phase II) 
Patients receiving steroids at the time of treatment. 
2B 
No Steroids (Phase II) 
Patients not receiving steroids at the time of treatment. 
2.3.3.2 Arms 
Number 
Name 
Description 
1 
Phase I Arm 
Non-myeloablative, lymphodepleting preparative regimen of 
cyclophosphamide and fludarabine + escalating doses of anti-
EGFRvIII CAR transduced PBL + aldesleukin 
2 
Phase II Arm 
Non-myeloablative, lymphodepleting preparative regimen of 
cyclophosphamide and fludarabine + MTD of anti-EGFRvIII 
CAR transduced PBL established in Phase I + aldesleukin 


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2.3.3.3 Randomization and Arm Assignment 
This is a non-randomized study. All patients will be directly assigned based on cohort as follows: 
• Patients in Cohorts 1A and 1B will be directly assigned to Arm 1. 
• Patients in Cohorts 2A and 2B will be directly assigned to Arm 2. 
3 
STUDY IMPLEMENTATION 
3.1 
STUDY DESIGN 
The study will be conducted using a Phase I/II design. With the approval of Amendment K, 
additional dose levels were added: dose levels 9-10. Prior to receiving the engineered PBL cells, 
patients will receive a non-myeloablative, lymphodepleting preparative regimen consisting of 
cyclophosphamide and fludarabine followed in 2-4 days by intravenous infusion of ex vivo 
tumor-reactive, EGFRvIII CAR gene-transduced PBMC, plus IV high- or low-dose aldesleukin 
(720,000 or 72,000 IU/kg every 8 hours for a maximum of 15 doses). 
Patients will undergo complete evaluation of tumor with physical and neurological examination, 
MRI of the brain with and without gadolinium, and clinical laboratory evaluation four weeks (± 
7 days) after completion of treatment. If the patient has stable disease or tumor shrinkage, repeat 
complete evaluations will be performed every 1 month (± 7 days). After the first year, patients 
continuing to respond will continue to be followed with this evaluation every 2 months (± 7 
days) until off-study criteria are met. All patients will receive one course of treatment. The start 
date of the course will be the start date of the chemotherapy; the end date will be the day of the 
first post-treatment evaluation. 
3.1.1 Schema – Beginning with Amendment K 
 


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3.1.2 Pre-Treatment Phase: Cell Preparation Performed on 03-C-0277 
PBMC will be obtained by leukapheresis (approximately 1x1010 cells). PBMC will be obtained 
while enrolled on protocol 03-C-0277. PBMC will be grown, transduced and expanded for this 
trial according to standard operating procedures submitted in the IND. Whole PBMC will be 
cultured in the presence of anti-CD3 (OKT3) and aldesleukin in order to stimulate T-cell growth. 
Transduction is initiated by exposure of approximately 1x107 to 5x108 cells to supernatant 
containing the anti-EGFRvIII CAR retroviral vector. These transduced cells will be expanded 
and tested for their anti-tumor activity. Successful CAR gene transfer will be determined by 
FACS analysis for the CAR protein and anti-tumor reactivity will be tested by cytokine release 
as measured on EGFRvIII expressing cells. Successful CAR gene transfer for each transduced 
PBL population will be defined as > 10% CAR positive cells and for biological activity, gamma-
interferon secretion must be ≥ 200 pg/mL and twice the background level as tested against the 
positive control and < 400 pg/mL against the negative control including wt-EGFRvIII. If the 
gamma-interferon secretion against the negative control is greater than 400 pg/mL the cells will 
remain in culture and tested daily until the target level is reached.  
3.1.3 Treatment Phase 
Note: Paragraphs highlighted in grey below are no longer applicable after the approval of 
amendment K. 
3.1.3.1 Phase I – Dose Escalation: Closed with Amendment H 
The protocol will begin with a Phase I dose escalation design, with eight dose levels and with 
two different cohorts; one for patients receiving steroids at the time of treatment (Cohort 1A) and 
one for patients not receiving steroids at the time of treatment (Cohort 1B). Each cohort in Phase 
I will be escalated independently of the other, and different MTDs may be identified.  
Initially, the protocol will enroll one patient in each of the first 4 dose levels unless that patient 
experiences a dose-limiting toxicity (DLT). Following Dose Level 4, all subsequent dose levels 
will proceed in a Phase I dose escalation design, with 4 dose levels of n = 3. 
The total number of EGFRvIII engineered cells transferred for each dose level will be: 
Dose Escalation Schedule 
Dose Level 
Dose of Anti-EGFRvIII CAR T-Cells  
Level 1 (Cohorts 1A & 1B) 
107 
Level 2 (Cohorts 1A & 1B) 
3x107 
Level 3 (Cohorts 1A & 1B) 
108  
Level 4 (Cohorts 1A & 1B) 
3x108 
Level 5 (Cohorts 1A & 1B) 
109 
Level 6 (Cohorts 1A & 1B) 
3x109 
Level 7 (Cohorts 1A & 1B) 
1010 
Level 8 (Cohorts 1A & 1B) 
3 – 6x1010 (closed following patient 17) 


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Patients will be enrolled sequentially, therefore enrollment will not proceed to a higher dose 
level until patients have been treated in the prior dose level. Patients, however, will be dose-
escalated to the next dose level within a given cohort independent on what is occurring in the 
other strata. If sufficient cells cannot be grown to meet the criteria for the assigned dose level, 
the patient will be enrolled in the appropriate dose level for the number of cells infused.  
In dose levels 1-4, if the patient experiences a DLT, five more patients would be treated at that 
dose level to confirm that no greater than 1/6 patients have a DLT prior to proceeding to the next 
higher level. If a level with 2 or more DLTs in 3-6 patients has been identified, five 
additional patients will be accrued at the next-lowest dose level, for a total of 6, in order to 
further characterize the safety of the MTD prior to starting the phase II portion. If there are 1 or 
fewer DLTs in the first dose level, the study will proceed to the second dose level. If a DLT 
occurs in the first dose level, that dose level will be expanded to n = 6 patients. If two DLTs 
occur in the first dose level, the study will be terminated.  
In dose levels 5-8, should a single patient experience a DLT due to the cell infusion at a 
particular dose level, three more patients would be treated at that dose level to confirm that no 
greater than 1/6 patients have a DLT prior to proceeding to the next higher level. If a level with 2 
or more DLTs in 3-6 patients has been identified, three additional patients will be accrued at the 
next-lowest dose level, for a total of 6, in order to further characterize the safety of the MTD 
prior to starting the phase II portion.  
The MTD is the highest dose at which < 1 of 6 patients experienced a DLT or the highest dose 
level studied if DLTs are not observed at any of the dose levels.  
Prior to receiving the engineered PBL cells, all patients will receive a non-myeloablative, 
lymphodepleting preparative regimen consisting of cyclophosphamide and fludarabine followed 
in 1-4 days by intravenous infusion of in vitro tumor-reactive, EGFRvIII CAR gene-transduced 
PBL plus IV aldesleukin (72,000 IU/kg every 8 hours for a maximum of 15 doses).  
A two-week safety assessment period following regimen completion (defined as the last dose of 
IL-2) will be conducted at the following times for each cohort: 
• For the first patient in each dose level before a second patient will be accrued in the dose 
level. 
• After the last patient is treated in each dose level prior to enrollment into the next dose 
level. 
• For the second and third patients in dose levels 1-4 (if the first patient treated in dose 
levels 1-4 experiences a DLT and the dose level is expanded).  
3.1.3.2 Phase I – Dose Escalation: Beginning with Amendment K 
Beginning with Amendment K, patients enrolled in dose levels 9-10 will receive a non-
myeloablative, lymphodepleting preparative regimen followed by intravenous infusion of 
EGFRvIII CAR gene-transduced PBMC, plus high- or low-dose aldesleukin.  
The total number of EGFRvIII engineered cells transferred for each dose level will be: 
 
 


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Dose Escalation Schedule 
Dose Level 
Dose of Anti-EGFRvIII CAR T-Cells  
Aldesleukin (IL-2) 
Level 6 (Completed) 3x109 
Low-Dose 
Level 7 (Completed) 1010 
Low-Dose 
Level 8 (Closed) 
3 – 6x1010 
Low-Dose 
Level 9  
3x1010 given as a split dose, 2 hours apart Low-Dose 
Level 9A 
3x1010 given as a single dose 
Low-Dose 
Level 9D-1 
1010 
High-Dose 
Level 9D-2 
3x109 
High-Dose 
Level 10 
3x1010 
High-Dose 
If one DLT is observed in Dose Level 9 or 9A, the dose will be de-escalated (Dose Level 9D-1) 
and high-dose IL-2 will be added. If no more than one DLT is seen, a total of 6 patients will be 
enrolled in Dose Level 9D-1 to further characterize the safety of the MTD. 
If more than one DLT is observed in Dose Level 9D-1, then the dose will be de-escalated (Dose 
Level 9D-2) and high-dose IL-2 will be added. If no more than one DLT is seen, a total of 6 
patients will be enrolled in Dose Level 9D-2 to further characterize the safety of the MTD. If 
more than one DLT is observed in Dose Level 9D-2, we will re-evaluate our strategy.  
If no DLTs are observed in Dose Level 9 or 9A, the dose will be escalated to include high-dose 
IL-2 (Dose Level 10). If no more than one DLT is seen in Dose Level 10, a total of 6 patients 
will be enrolled in Dose Level 10 to further characterize the safety of the MTD. If more than one 
DLT is observed in Dose Level 10, a total of 6 patients will be enrolled in Dose Level 9D-1. 
If sufficient cells cannot be grown to meet the criteria for the assigned dose level, the patient will 
be enrolled in the appropriate dose level for the number of cells infused.  
The MTD is the highest dose at which < 1 of 6 patients experienced a DLT or the highest dose 
level studied if DLTs are not observed at any of the dose levels.  
A two-week safety assessment period following regimen completion (defined as the last dose of 
IL-2) will be conducted at the following times for each dose level: 
• After the first patient in each dose level before a second patient will be accrued in the 
dose level. 
• After any patient experiences a DLT. 
3.1.4 Dose-Limiting Toxicity 
A DLT is defined as follows:  
• Grade 2 or greater allergic reaction or reaction that involves bronchospasm or generalized 
urticaria. 
• All grade 3 and greater toxicities with the exception of: 


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o Myelosuppression, defined as lymphopenia, neutropenia, decreased hemoglobin, and 
thrombocytopenia, due to the chemotherapy preparative regimen. 
o IL-2 expected toxicities as defined in Appendix B and Appendix C. 
o Expected chemotherapy toxicities as defined in Section 11. 
o Immediate hypersensitivity reactions occurring within 2 hours of cell infusion (related 
to cell infusion) that are reversible to a grade 2 or less within 24 hours of cell 
administration with standard therapy.  
o Grade 3 fever 
o Grade 3 metabolic laboratory abnormalities without significant clinical sequela that 
resolve to grade 2 within 7 days. 
o Grade 3 autoimmunity that resolves to less than or equal to a grade 2 autoimmune 
toxicity within 10 days. 
3.1.5 Phase II Portion 
Once the MTD has been determined, the study will then proceed to the Phase II single stage 
portion utilizing the MTD of anti-EGFRvIII engineered cells as determined in the Phase I 
portion. Patients will be accrued into two cohorts: those patients with recurrent malignant glioma 
receiving steroids at the time of treatment (Cohort 2A), and those patients with recurrent 
malignant glioma not receiving steroids at the time of treatment (Cohort 2B).  
Similar to the Phase I portion, prior to receiving the engineered PBL cells, patients in the Phase 
II portion will receive a non-myeloablative, lymphodepleting preparative regimen consisting of 
cyclophosphamide and fludarabine followed in 1-4 days by intravenous infusion of ex vivo 
tumor-reactive, anti-EGFRvIII CAR gene-transduced PBL plus IV high-dose aldesleukin.  
Note: If the MTD is not established in Dose Level 9D, we will re-evaluate our strategy and 
amend the protocol. 
3.2 
PROTOCOL STOPPING RULES 
New subject enrollment to the protocol will be temporarily halted if any of the following 
conditions are met in each cohort separately, and discussions with the FDA or NIH IRB 
regarding safety and the need for protocol revisions if applicable: 
• During the Phase I portion of the study – If two or more patients develop a grade 3 or 
greater toxicity related to the cell product, with the exception of: 
o Grade 3 metabolic laboratory abnormalities without significant clinical sequela that 
resolve to grade 2 or less within 7 days 
o Grade 3 fever  
• If 1 of the first 3 patients (OR 2 of the first 6 patients, OR 3 of the first 9 patients, OR 4 
of the first 12 patients) develop grade 3 autoimmunity, that cannot be resolved to less 
than or equal to a grade 2 autoimmune toxicity within 10 days, or any grade 4 or greater 
autoimmune toxicity.  
• Two or more patients remain neutropenic (grade 3 or 4 ANC) 4 weeks following 
treatment.  


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• Two or more patients remain thrombocytopenic (grade 3 or 4 platelets) 8 weeks 
following treatment. 
• Two or more patients develop a major neurological event defined as: 
o Significant grade 3 toxicity that does not resolve to ≤ grade 1 with best medical 
management and supportive care within 3 weeks 
o Significant grade 4 toxicity  
• During the Phase II portion of the study – Once five or more patients have been enrolled, 
if grade 4 non-reversible non-hematological toxicities exceed 20% or more in patients 
cumulatively enrolled in this phase. Grade 4 non-reversible toxicities are defined as those 
grade 4 toxicities that are not reversible over a 7-day period.  
• During the Phase I and II portions of the study – Any death that is at least possibly 
attributed to the investigational agent and which occurs within 30 days of receiving the 
investigational agent. 
The rationale for these neurological guidelines is based on this patient population’s inherent 
increased risk of cerebral edema. In particular, aldesleukin is known to cause capillary leak 
syndrome which can lead to cerebral edema and its ensuing symptoms including the following: 
seizures, depressed level of consciousness, encephalopathy, headache, clinically evident 
herniation syndromes, and worsening of baseline focal neurological symptoms. 
These symptoms are generally transient, directly related to cerebral edema, and reversible with 
supportive care. Therefore, any grade 3 neurological toxicity should first be treated with best 
medical management and supportive care (see table below) until resolution of the toxicity to ≤ 
grade 1. If the neurological toxicity does not resolve to grade 1 with best medical management 
and supportive care, it will qualify as a major neurological event as defined above.  
3.2.1.1 Management of Expected Significant Neurological Toxicities ≥ Grade 3 
Expected Toxicity 
Medical Management/Supportive Care 
Cerebral Edema 
IV Steroids 
Mannitol, per protocol 
Seizures 
Institute AEDs, up to a maximum of 3 agents 
Encephalopathy 
Treat metabolic abnormalities 
IV Steroids 
Headache 
IV Steroids 
Analgesics, including at least 1 narcotic agent 
3.3 
DRUG ADMINISTRATION 
3.3.1 Preparative Regimen with Cyclophosphamide and Fludarabine 
Treatment will be according to the schedule described below and in Section 3.3.4. Starting on 
Day -6, study medication start times for drugs given once daily should be given within 2 hours of 
the scheduled time. Chemotherapy infusions may be slowed or delayed as medically indicated. 


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Administration of diuretics, electrolyte replacement, and hydration and monitoring of 
electrolytes should all be performed as clinically indicated. 
Days -7 and -6 
Approximately 6 Hours Prior to Cyclophosphamide 
Hydrate: Begin hydration with 0.9% Sodium Chloride Injection containing 10 meq/L of 
potassium chloride at 1.5-2.6 mL/kg/hour (starting approximately 6 hours pre-cyclophosphamide 
and continuing until 24 hours after last cyclophosphamide infusion). The hydration rate will be 
capped at 250 mL/hour. At any time during the preparative regimen, if urine output is <1.0 
mL/kg/hour or if body weight is >2 kg over pre-cyclophosphamide value, furosemide 10-20 mg 
IV may be administered.  
Approximately 1 Hour Prior to Cyclophosphamide  
Ondansetron (0.15 mg/kg/dose [rounded to the nearest even mg dose between 8 mg and 16 mg 
based on patient weight] IV every 8 hours x 3 days) will be given for nausea.  
Cyclophosphamide 60 mg/kg/day x 2 days IV in 250 mL D5W infused simultaneously with 
mesna 15 mg/kg/day over 1 hour x 2 days. If patient is obese (BMI > 35), drug dosage will be 
calculated using practical weight as described in Appendix A.  
A decreased dose of cyclophosphamide at 30 mg/kg/day (x 2 days) will be considered for 
patients who have a history of prolonged hematologic recovery from prior chemotherapy 
treatments. 
Immediately following the end of Cyclophosphamide 
Begin mesna infusion at 3 mg/kg/hour intravenously diluted in a suitable diluent (see Section 
11.2.1) over 23 hours after each cyclophosphamide dose. If patient is obese (BMI > 35), drug 
dosage will be calculated using practical weight as described in Appendix A.  
Days -7 to -3 
Fludarabine 25 mg/m2/day IVPB daily over 30 minutes for 5 days. If patient is obese (BMI > 
35), drug dosage will be calculated using practical weight as described in Appendix A. 
(Fludarabine will be started approximately 1-2 hours after the cyclophosphamide and mesna on 
Days -7 and -6). 
3.3.2 Cell Infusion 
Day 0 (2-4 Days After the Last Dose of Fludarabine) 
Cells are delivered to the Patient Care Unit by a staff member from the Surgery Branch Cell 
Production Facility (SB-CPF). Prior to infusion, the cell product identity label is double-checked 
by two authorized staff (MD or RN), and an identification of the product and documentation of 
administration are entered in the patient’s chart. The cells are to be infused intravenously on the 
Patient Care Unit over 20-30 minutes or as clinically determined by an investigator for patient 
safety via non-filtered tubing, gently agitating the bag during infusion to prevent cell clumping.  
Note: For patients enrolled in Dose Level 9, the cell dose will be given in two doses, 2 hours 
apart. If the patient experiences a grade 1 toxicity that does not resolve within one hour or 
a grade 2 or greater toxicity, the second half of the dose will be withheld. 


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3.3.3 Aldesleukin Administration 
Days 0-5 (Day 0 = Day of Cell Infusion)  
• Beginning on Day 1 or 2, filgrastim will be administered subcutaneously at a dose of 300 
mcg/day (dose may be adjusted as clinically indicated). Filgrastim administration will 
continue daily until neutrophil count > 1x109/L x 3 days or > 5x109/L. 
• Aldesleukin 72,000 IU/kg IV (Dose Levels 9 & 9A) or 720,000 IU/kg IV (Dose Levels 
9D & 10) (based on total body weight) over 15 minutes approximately every eight hours 
beginning within 24 hours of cell infusion and continuing for up to 5 days (maximum of 
15 doses). 
• For Phase II portion: Aldesleukin 720,000 IU/kg IV (based on total body weight) over 15 
minutes approximately every eight hours beginning within 24 hours of cell infusion and 
continuing for up to 5 days (maximum of 15 doses). If high-dose aldesleukin is not 
tolerated in Dose Levels 9D or 10, then we will re-evaluate our strategy and amend the 
protocol. 
Aldesleukin (based on total body weight) will be administered as an intravenous bolus over a 15-
minute period approximately every 8 hours beginning within 24 hours of cell infusion and 
continuing for up to 5 days (maximum 15 doses). Doses will be preferentially administered every 
eight hours; however, up to 24 hours may elapse between doses depending on patient tolerance. 
Aldesleukin dosing will be stopped if toxicities are not sufficiently recovered by supportive 
measures within 24 hours of the last dose of aldesleukin. Dosing will be delayed or stopped if 
patients reach grade 3 or 4 toxicity due to aldesleukin, except for the reversible grade 3 toxicities 
common to aldesleukin such as diarrhea, nausea, vomiting, hypotension, skin changes, anorexia, 
mucositis, dysphagia, or constitutional symptoms and laboratory changes as detailed in 
Appendix B. Toxicities will be managed as outlined in Appendix C. Dosing may be held or 
stopped at the discretion of the treating investigator. (Appendix D lists the toxicities seen in 
patients treated with aldesleukin at the NIH Clinical Center).  
Because confusion is a possible side effect of aldesleukin administration, a Durable Power of 
Attorney will be signed by the patient to identify a surrogate to make decisions about their 
medical care if the patient becomes incapacitated or cognitively impaired. 
3.3.4 Treatment Schedule 
Day 
-7 
-6 
-5 
-4 
-3 
-2 
-1 
0 
1 
2 
3 
4 
5 
Therapy 
 
 
 
 
 
 
 
 
 
 
 
 
 
Cyclophosphamide (60 mg/kg) 
X 
X 
 
 
 
 
 
 
 
 
 
 
 
Fludarabine (25 mg/m2) 
X 
X 
X 
X 
X 
 
 
 
 
 
 
 
 
Anti-EGFRvIII PBL 
 
 
 
 
 
 
 
X1 
 
 
 
 
 
Aldesleukin 
 
 
 
 
 
 
 
X2 
X 
X 
X 
X 
X 
Filgrastim3 (5 mcg/kg) 
 
 
 
 
 
 
 
 
X 
X 
X 
X 
X 
TMP/SMX4 
160 mg/800 mg (example) 
 
 
 
 
 
 
 
X 
 
X 
 
X 
 


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1Two to four days after the last dose of fludarabine. The cells will be split into two doses, two hours apart 
for patients in Dose Level 9. 
2Initiate within 24 hours after cell infusion. 
3Continue until neutrophil count > 1x109/L for 3 consecutive days or > 5x109/L. 
4The TMP/SMX schedule should be adjusted to QD three times per week (Monday, Wednesday, Friday) 
and continue for at least six months and until CD4 > 200 x2, starting Day 0 or within one week of 
anticipated lymphopenia. 
5Continue until ANC > 1000/mm3 
6In patients positive for HSV or VZV, continue for at least 6 months and until CD4 > 200 x2. 
3.4 
BASELINE EVALUATION 
3.4.1 Within 14 Days Prior to Starting the Preparative Regimen 
• Apheresis, as indicated 
• Baseline blood tests: 
o CBC w/differential 
o Acute Care Panel (sodium, potassium, chloride, bicarbonate, creatinine, glucose, 
BUN), Hepatic Panel (alkaline phosphatase, AST, ALT, total bilirubin, direct 
bilirubin), Mineral Panel (albumin, calcium, magnesium, phosphorus), Uric acid, 
Creatinine kinase, Lactate dehydrogenase, Total protein  
o PT-INR/PTT 
o TBNK 
o Thyroid Panel 
o Anti-CMV antibody titer, HSV and VZV serology, and EBV panel (may be 
performed within 3 months prior to starting the preparative regimen) 
• Urinalysis, with culture if indicated 
• Chest x-ray 
• EKG 
3.5 
ON-STUDY EVALUATION 
3.5.1 During the Preparative Regimen (Daily) 
• CBC w/differential 
o Note: Once the platelet count drops below 80,000/mm3, platelets will be monitored 
and transfusions given as clinically indicated until the platelet count is greater than 
30,000/mm3 for 24 hours without transfusion support. 
Fluconazole5 (400 mg PO) 
 
 
 
 
 
 
 
X 
X 
X 
X 
X 
X 
Valacyclovir PO or Acyclovir IV6 
 
 
 
 
 
 
 
X 
X 
X 
X 
X 
X 


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• Acute Care Panel (sodium, potassium, chloride, bicarbonate, creatinine, glucose, BUN), 
Hepatic Panel (alkaline phosphatase, AST, ALT, total bilirubin, direct bilirubin), Mineral 
Panel (albumin, calcium, magnesium, phosphorus), Uric acid, Creatinine kinase, Lactate 
dehydrogenase, Total protein  
• PT-INR/PTT (every 3 days) 
• Urinalysis, as needed 
• Weight, as indicated 
3.5.2 Post-Cell Infusion 
• Vital signs, including neurological examinations, will be monitored hourly (± 15 
minutes) for four hours and then routinely (every 4-6 hours) unless otherwise clinically 
indicated. 
3.5.3 During Aldesleukin Administration (Daily)  
Note: Until values return to normal limits/baseline. 
• CBC (except platelets as noted in Section 3.5.1) 
• Acute Care Panel (sodium, potassium, chloride, bicarbonate, creatinine, glucose, BUN), 
Hepatic Panel (alkaline phosphatase, AST, ALT, total bilirubin, direct bilirubin), Mineral 
Panel (albumin, calcium, magnesium, phosphorus), Uric acid, Creatinine kinase, Lactate 
dehydrogenase, Total protein   
3.5.4 During Hospitalization (Every 1-2 Days) 
Note: Once electrolytes are stable and platelet count is above 30,000/mm3 for 24 hours without 
support. 
• Physical examination, including weight and vital signs, as clinically indicated 
• Toxicity assessment, including a review of systems, as clinically indicated 
• CBC w/differential 
• Acute Care Panel (sodium, potassium, chloride, bicarbonate, creatinine, glucose, BUN), 
Hepatic Panel (alkaline phosphatase, AST, ALT, total bilirubin, direct bilirubin), Mineral 
Panel (albumin, calcium, magnesium, phosphorus), Uric acid, Creatinine kinase, Lactate 
dehydrogenase, Total protein 
• Once total lymphocyte count is > 200/mm3, TBNK for peripheral blood CD4 count will 
be drawn weekly (while the patient is hospitalized). See Section 5.5 for post-cell infusion 
evaluations. 
• Other tests will be performed as clinically indicated. 
3.6 
POST-TREATMENT (FOLLOW-UP) EVALUATION  
• All patients will return to the NIH Clinical Center for their first follow-up evaluation for 
response 4 weeks (± 7 days) following the administration of the cell product.  
• Patients who have received multiple transfusions during the treatment phase or have been 
discharged with grade 3 or greater significant adverse events should be evaluated by the 


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referring physician within two weeks of discharge and repeat labs as appropriate to be 
faxed to the research team. Patients will receive appropriate treatment as determined by 
their treating physician 
• Patients who are unable or unwilling to return for follow-up evaluations may be followed 
via phone or email contact. A request will be made to send laboratory, imaging, and 
physician exam reports performed by their treating physician. Any outstanding toxicities 
will be reviewed with the patient. 
3.6.1 Time-Period of Evaluations 
Patients who experience stable disease, a partial response, or a complete response or have 
unresolved toxicities after their first follow-up evaluation will return to the NIH Clinical Center 
as noted below: 
• Monthly (± 7 days) for 12 months  
• Every 1-2 months (± 7 days) as feasible until off-study criterion is met 
Note: Patients may be seen more frequently as clinically indicated. 
3.6.2 Scheduled Evaluations 
At each scheduled evaluation for response, patients will undergo: 
• Physical examination, including neurological examination 
• KPS  
• Weight and vital signs 
• Toxicity assessment, including a review of systems 
• CBC w/differential 
• Acute Care Panel (sodium, potassium, chloride, bicarbonate, creatinine, glucose, BUN), 
Hepatic Panel (alkaline phosphatase, AST, ALT, total bilirubin, direct bilirubin), Mineral 
Panel (albumin, calcium, magnesium, phosphorus), Uric acid, Creatinine kinase, Lactate 
dehydrogenase, Total protein  
• PT-INR/PTT 
• Urinalysis, as needed 
• Thyroid Panel, as clinically indicated 
• TBNK 
• MRI of the brain with and without gadolinium 
• A 5-liter apheresis may be performed at the first follow-up visit. If the patient is unable to 
undergo apheresis, approximately 96 mL of blood may be obtained. Subsequently, 
approximately 60 mL of blood will be obtained at follow-up visits for at least 3 months. 
Peripheral blood mononuclear cells will be cryopreserved so that immunologic testing 
may be performed. This will be performed on protocol 03-C-0277. 


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• Detection of RCR and persistence of CAR gene transduced cells (see Section 5.8). This 
will be performed on the NCI-SB long-term follow-up protocol 09-C-0161 (Follow up 
Protocol for Subjects Previously Enrolled in NCI Surgery Branch Studies). 
• Long-term follow-up of patients receiving gene transfer: Physical examinations will be 
performed and documented annually for 5 years following cell infusion to evaluate long-
term safety. After 5 years, health status data will be obtained from surviving patients via 
telephone contact or mailed questionnaires. The long-term follow-up period for retroviral 
vectors is 15 years. This will be performed on protocol 09-C-0161.


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3.7 
STUDY ASSESSMENT CALENDAR 
Assessments 
Prior to Starting Preparative Regimen 
Prior to 
Preparative 
Regimen 
During 
Preparative 
Regimen 
(Daily) 
Prior to 
Cell 
Infusion 
Post-Cell 
Infusion 
During 
IL-2 
Admin.1 
During 
Hospitalization 
(Every 1-2 
Days)2 
Post-
Treatment 
Follow-up3 
Within 3 
Months 
Within 8 
Weeks 
Within 4 
Weeks 
Within 
14 Days 
Within 
7 Days 
Within  
1-4 Days 
IHC or PCR 
confirmation of 
tumor tissue4 
X 
 
 
 
 
 
 
 
 
 
 
 
 
Medical history5 
 
X 
X 
 
 
 
 
 
 
 
 
 
 
Physical exam 
 
 
X 
 
X 
 
 
 
 
 
 
X 
X 
Neurological exam 
 
 
 
 
X 
 
 
 
 
X6 
 
 
X 
Performance score 
(KPS)7 
 
 
 
 
X 
 
 
 
 
 
 
 
X 
Weight 
 
 
X 
 
 
 
 
X 
 
 
 
X 
X 
Vital signs 
 
 
X 
 
 
 
 
 
 
X8 
 
X 
X 
β-HCG pregnancy 
test9 
 
 
 
 
X 
 
 
 
 
 
 
 
 
                                                 
1 Until values return to normal limits/baseline. 
2 Once electrolytes are stable and platelet count is above 30,000/mm3 for 24 hours without support. 
3 All patients will return to the NIH Clinical Center for their first follow-up evaluation for response 4 weeks (± 7 days) following the 
administration of the cell product. Patients who experience stable disease, a partial response, or a complete response or have unresolved toxicities 
after their first follow-up evaluation will return to the NIH Clinical Center monthly (± 7 days) for 12 months and then every 1-2 months (± 7 days) 
as feasible until off-study criterion is met. See Section 3.6. 
4 Immunohistochemistry or PCR confirmation of tumor tissue for expression of EGFRvIII for histologically proven glioblastomas or gliosarcomas 
confirmed by the NCI Laboratory of Pathology. Testing is permitted to be conducted at any time prior to starting the preparative regimen. 
5 Note organ system involvement and any allergies/sensitivities to antibiotics. Patient history may be obtained within 8 weeks prior to starting the 
preparative regimen. 
6 Neurological examinations will be monitored hourly (± 15 minutes) for four hours and then routinely (every 4-6 hours) unless otherwise 
clinically indicated. 
7 KPS ≥ 60 (see Appendix F). 
8 Vital signs will be monitored hourly (± 15 minutes) for four hours and then routinely (every 4-6 hours) unless otherwise clinically indicated. 
9 Serum or urine; on all women of child-bearing potential. 


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Assessments 
Prior to Starting Preparative Regimen 
Prior to 
Preparative 
Regimen 
During 
Preparative 
Regimen 
(Daily) 
Prior to 
Cell 
Infusion 
Post-Cell 
Infusion 
During 
IL-2 
Admin.1 
During 
Hospitalization 
(Every 1-2 
Days)2 
Post-
Treatment 
Follow-up3 
Within 3 
Months 
Within 8 
Weeks 
Within 4 
Weeks 
Within 
14 Days 
Within 
7 Days 
Within  
1-4 Days 
Urinalysis10 
 
 
 
X 
 
 
 
X 
 
 
 
 
X 
Pulmonary 
function test11 
 
X 
 
 
 
 
 
 
 
 
 
 
 
Cardiac 
evaluation12 
 
X 
 
 
 
 
 
 
 
 
 
 
 
EKG 
 
 
 
X 
 
 
 
 
 
 
 
 
 
Toxicity 
assessment13 
 
 
 
 
 
 
 
 
 
 
 
X 
X 
Serology 
HIV antibody, 
HBsAg, anti-HCV 
X 
 
 
 
 
 
 
 
 
 
 
 
 
Anti-CMV, HSV 
and VZV serology, 
EBV panel 
X 
 
 
X14 
 
 
 
 
 
 
 
 
 
Laboratory Procedures 
CBC w/differential 
 
 
 
X 
 
 
 
X15 
 
 
X16 
X 
X 
                                                 
10 With culture if indicated. 
11 For patients with a prolonged history of cigarette smoking (≥ 20 pack-year smoking history, with cessation within the past two years), 
symptoms of respiratory dysfunction, or other clinical indications which may include thoracic surgeries. 
12 Commensurate to patients’ history and clinical presentation (e.g., stress thallium, echocardiogram, MUGA) for patients who are ≥ 65 years of 
age, or who have a history of ischemic heart disease, chest pain, or clinically significant atrial and/or ventricular arrhythmias, including but not 
limited to: atrial fibrillation, ventricular tachycardia, heart block. Patients with a LVEF ≤ 45% will not be eligible. Patients < 65 years of age with 
cardiac risk factors (e.g., diabetes, hypertension, obesity) may undergo cardiac evaluation as noted above. 
13 Including a review of systems. 
14 Anti-CMV antibody titer, HSV and VZV serology, and EBV panel may be performed within 3 months prior to starting the preparative regimen. 
15 Once the platelet count drops below 80,000/mm3, platelets will be monitored and transfusions given as clinically indicated until the platelet 
count is greater than 30,000/mm3 for 24 hours without transfusion support. 
16 Except platelets as noted in Section 3.5.1. 


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Assessments 
Prior to Starting Preparative Regimen 
Prior to 
Preparative 
Regimen 
During 
Preparative 
Regimen 
(Daily) 
Prior to 
Cell 
Infusion 
Post-Cell 
Infusion 
During 
IL-2 
Admin.1 
During 
Hospitalization 
(Every 1-2 
Days)2 
Post-
Treatment 
Follow-up3 
Within 3 
Months 
Within 8 
Weeks 
Within 4 
Weeks 
Within 
14 Days 
Within 
7 Days 
Within  
1-4 Days 
Blood 
chemistries17 
 
 
 
X 
 
 
 
X 
 
 
X 
X 
X 
PT-INR/PTT 
 
 
 
X 
 
 
 
X18 
 
 
 
 
X 
TBNK 
 
 
 
X 
 
 
 
 
 
 
 
X19 
X 
Thyroid Panel 
 
 
 
X 
 
 
 
 
 
 
 
 
X 
Additional 
apheresis20 
 
 
 
X 
 
 
 
 
 
 
 
 
X 
Correlatives21 
CPT tubes (SB-
CPF) 
 
 
 
 
 
 
X 
 
 
X 
 
 
 
SST tube (Figg 
Lab) 
 
 
 
 
 
 
X 
 
X 
X 
 
 
 
Imaging 
MRI scan 
 
 
 
 
 
X22 
 
 
 
 
 
 
X23 
Chest x-ray 
 
 
 
X 
 
 
 
 
 
 
 
 
 
                                                 
17 Screening: Creatinine, ALT/GPT, AST/GOT, Total bilirubin. All other times: Acute Care Panel (sodium, potassium, chloride, bicarbonate, 
creatinine, glucose, BUN), Hepatic Panel (alkaline phosphatase, AST, ALT, total bilirubin, direct bilirubin), Mineral Panel (albumin, calcium, 
magnesium, phosphorus), Uric acid, Creatinine kinase, Lactate dehydrogenase, Total protein. 
18 Every 3 days. 
19 Once total lymphocyte count is > 200/mm3, TBNK for peripheral blood CD4 count will be drawn weekly (while the patient is hospitalized). 
20 Apheresis may be performed prior to and 6 weeks (± 2 weeks) following the administration of the cell product. If the patient is unable to 
undergo apheresis, approximately 96 mL of blood may be obtained. Subsequently, approximately 60 mL of blood will be obtained at follow-up 
visits for at least 3 months. Peripheral blood mononuclear cells will be cryopreserved so that immunologic testing may be performed. This will be 
performed on protocol 03-C-0277. 
21 Research samples, as described in Section 5. 
22 Dynamic contrast-enhanced MRI scan with perfusion of the head. For patients on steroids, the dose must be fixed for at least 5 days prior to 
imaging. If the steroid dose is increased between the date of imaging and registration, a new baseline MRI is required. 
23 MRI of the brain with and without gadolinium. 


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3.8 
CRITERIA FOR REMOVAL FROM PROTOCOL THERAPY AND OFF-STUDY CRITERIA 
Prior to removal from study, effort must be made to have all subjects compete an evaluation 
safety visit approximately 4 weeks (± 7 days) following administration of the cell product (at the 
first follow-up evaluation). 
3.8.1 Criteria for Removal from Protocol Therapy  
Patients will be taken off treatment for the following: 
• Completion of first follow-up evaluation 
• Progression of disease 
• Investigator discretion 
• Patient requests to be withdrawn from protocol therapy 
• Positive pregnancy test 
3.8.2 Off-Study Criteria 
Patients will be taken off-study for the following: 
• Completion of study follow-up period 
• Progression of disease 
• Patient begins a new therapy for their cancer 
• Investigator discretion 
• Patient requests to be withdrawn from the study 
• Significant noncompliance 
• Patient lost to follow-up 
• Death 
All patients will be co-enrolled on protocol 09-C-0161. Patients who are taken off-study for 
progressive disease or study closure on this treatment protocol may be followed on protocol 09-
C-0161. 
Once a subject is taken off-study, no further data can be collected on this treatment protocol. 
3.8.3 Off Protocol Therapy and Off-Study Procedure 
Authorized staff must notify the NCI-CRO when a subject is taken off protocol therapy and 
when a subject is taken off-study. A Participant Status Update Form from the website 
(http://home.ccr.cancer.gov/intra/eligibility/welcome.htm) main page must be completed and 
sent via encrypted email to the NCI-CRO at ncicentralregistration-l@mail.nih.gov.  
4 
CONCOMITANT MEDICATIONS/MEASURES 
Concomitant medications to control side effects of therapy will be given. Meperidine (25-50 mg) 
will be given intravenously if severe chilling develops. Other supportive therapy will be given as 
required and may include acetaminophen (650 mg every 4 hours), indomethacin (50-75 mg every 
6 hours) and ranitidine (150 mg every 12 hours). Ondansetron 0.15 mg/kg/dose IV every 8 hours 


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will be administered for nausea and vomiting. Additional antiemetics will be administered as 
needed for nausea and vomiting uncontrolled by ondansetron. 
4.1 
MANNITOL  
Mannitol will be preferentially used in the acute hospital setting to control the signs and 
symptoms of increased intracranial pressure. If necessary, corticosteroids may then be used in 
the subacute and outpatient setting for the control of the signs and symptoms of increased 
intracranial pressure. However, all attempts will be made to withhold steroids in the non-steroid 
strata unless clearly medically indicated. Increases in steroid dose should be discussed with the 
Principal Investigator (PI). Changes in the steroid dose should be carefully documented on the 
flow sheets. 
4.2 
NON-ENZYME INDUCING ANTI-EPILEPTIC DRUGS (NON-EIAED) 
• Non-EIAEDs are permitted but should be recorded in the patient’s chart. If anti-seizure 
medications are required during study, every effort should be made to choose non-
EIAEDs (Appendix G). 
• Changes in the doses of dexamethasone and anti-epileptics will be allowed, but must be 
clearly documented in the patient’s chart. It is strongly recommended that if a patient 
needs to be taken off a particular anti-epileptic agent (i.e. allergic reaction, ineffective), 
that it be replaced with another agent that is a non-inducer of the cytochrome P450 
system. 
4.3 
INFECTION PROPHYLAXIS 
Note: Other anti-infective agents may be substituted at the discretion of the treating investigator. 
4.3.1 Pneumocystis Jirovecii Pneumonia 
All patients will receive the fixed combination of trimethoprim and sulfamethoxazole 
(TMP/SMX) as double strength (DS) tab (DS tabs = TMP 160 mg/tab, and SMX 800 mg/tab) PO 
daily three times a week on non-consecutive days, beginning Day 0 or within one week of 
anticipated lymphopenia.  
Dapsone (in G6PD sufficient patient), atovaquone, or pentamidine may be substituted for 
TMP/SMX-DS in patients with sulfa allergies.  
4.3.2 Herpes Simplex or Varicella Zoster Virus Prophylaxis 
Patients with positive HSV or VZV serology will be given valacyclovir orally at a dose of 500 
mg daily starting on the day of cell infusion, or acyclovir, 250 mg/m2 IV every 12 hours if the 
patient is not able to take medication by mouth. Reversible renal insufficiency has been reported 
with IV but not oral acyclovir. Neurologic toxicity including delirium, tremors, coma, acute 
psychiatric disturbances, and abnormal EEGs have been reported with higher doses of acyclovir. 
Should this occur, a dosage adjustment will be made or the drug will be discontinued. Acyclovir 
will not be used concomitantly with other nucleoside analogs which interfere with DNA 
synthesis, e.g. ganciclovir. In renal disease, the dose is adjusted as per product labeling. 
Prophylaxis for pneumocystis, HSV, and VZV will continue for 6 months post-chemotherapy. If 
the CD4 count is < 200 at 6 months post-chemotherapy, prophylaxis will continue for at least 6 
months and until CD4 count is > 200 for two consecutive measures. 


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Note: A missed prophylactic dose will not be considered a protocol deviation, and thus a 
deviation will not be reported to the NIH-IRB (see sections 7.1.8 and 7.2), if the patient is 
compliant with taking at least 75 percent of their required dose. 
4.3.3 Fungal Prophylaxis (Fluconazole) 
Patients will start fluconazole 400 mg PO starting on the day of cell infusion and continue until 
the absolute neutrophil count is > 1000/mm3. The drug may be given IV at a dose of 400 mg in 
0.9% sodium chloride USP daily in patients unable to take it orally. 
4.3.4 Empiric Antibiotics 
Patients will start on broad-spectrum antibiotics, in accordance with the current institutional 
guidelines, for fever of 38.3°C once or two temperatures ≥ 38.0°C at least one hour apart, AND 
an ANC < 500/mm3. Infectious disease consultation will be obtained for all patients with 
unexplained fever or any infectious complications. 
4.4 
BLOOD PRODUCT SUPPORT 
Using daily CBCs as a guide, the patient will receive platelets and packed red blood cells 
(PRBCs) as needed. As a general guideline, patients may be transfused for: 
• Hemoglobin < 8 gm/dL 
• Platelets < 10,000/mm3 
All blood products will be irradiated. Leukocyte filters will be utilized for all blood and platelet 
transfusions to decrease sensitization to transfused WBCs and decrease the risk of CMV 
infection. 
5 
BIOSPECIMEN COLLECTION 
Blood and tissue are tracked at the patient level and can be linked to all protocols on which the 
patient has been enrolled. Samples will be used to support the specific objectives listed in the 
treatment protocol(s), e.g., immunologic monitoring, cytokine levels, persistence, as well as to 
support long-term research efforts within the NCI-SB and with collaborators as specified in 
protocol 03-C-0277. 
The amount of blood that may be drawn from adult patients for research purposes shall not 
exceed 10.5 mL/kg or 550 mL, whichever is smaller, over any eight-week period. 
Samples will be ordered in CRIS and tracked through a Clinical Trial Data Management system. 
Should a CRIS screen not be available, the CRIS downtime procedures will be followed. 
Samples will not be sent outside NIH without IRB notification and an executed MTA. 
5.1 
SAMPLES SENT TO FIGG LAB 
• Venous blood samples will be collected in either a 4-mL or an 8-mL SST tube to be 
processed for serum and stored for future research. Record the date and exact time of 
draw on the tube. Blood tubes may be kept in the refrigerator until pick-up.  
• For sample pick-up, page 102-11964. 
• For immediate help, call 240-760-6180 (main blood processing core number) or, if no 
answer, 240-760-6190 (main clinical pharmacology lab number). 


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• For questions regarding sample processing, contact Julie Barnes by e-mail or at 240-760-
6044. 
• The samples will be processed, barcoded, and stored in the Figg lab until requested by the 
investigator. 
5.2 
SAMPLES SENT TO SURGERY BRANCH CELL PRODUCTION FACILITY 
• Venous blood samples will be collected in 8-mL CPT tubes to be processed and stored 
for future research. Record the date and exact time of draw on the tube. Blood tubes are 
kept at room temperature until pick-up. 
• Samples will be picked up by the research nurse or designee and transported to the SB-
CPF within 24 hours of blood draw. 
• The samples will be processed, barcoded, and stored in SB-CPF. 
5.3 
PRIOR TO CHEMOTHERAPY ADMINISTRATION 
• 5 CPT tubes (8 mL each): SB-CPF 
• 1 SST tube (8 mL): Figg Lab 
• 1 SST tube (4 mL): Daily, starting day of chemotherapy through day of discharge. Send 
to Figg Lab. 
5.4 
PRIOR TO CELL INFUSION (BASELINE SAMPLE FOR CYTOKINE ANALYSIS) 
• 1 SST tube (8 mL): Figg Lab 
5.5 
POST-CELL INFUSION EVALUATIONS 
Once total lymphocyte count is > 200/mm3, the following samples will be drawn and sent to the 
SB-CPF on Monday, Wednesday, and Friday x5 days, then weekly (while the patient is 
hospitalized): 
• 5 CPT tubes (8 mL each): SB-CPF 
• 1 SST tube (8 mL): Figg Lab 
5.6 
SAMPLE COLLECTION SCHEDULE 
Test 
Volume Blood 
Type of tube 
Collection Point 
Disposition 
Research blood 
52 mL 
CPT and SST 
Prior to chemo 
Deliver SST tubes to Figg Lab. 
Deliver CPT tubes to SB-CPF. 
Research blood 
8 mL 
SST 
Prior to cell 
infusion 
Deliver to Figg Lab. 
Research blood 
48 mL 
CPT and SST 
Post-cell infusion 
and at follow-up 
visits1 
Deliver SST tubes to Figg Lab. 
Deliver CPT tubes to SB-CPF. 
1 Research blood will be obtained at follow-up visits for at least 3 months. 


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5.7 
IMMUNOLOGICAL TESTING 
• Apheresis may be performed prior to and 6 weeks (± 2 weeks) following the 
administration of the cell product. At other time points, patient PBL will be obtained 
from whole blood by purification using centrifugation on a Ficoll cushion. Aliquots of 
these PBMC will be cryopreserved for immunological monitoring of cell function. 
• Lymphocytes will be tested directly and following in vitro culture using some or all of 
the following tests. Direct immunological monitoring will consist of quantifying T-cells 
reactive with EGFRvIII by FACS analysis using CAR-specific staining. Ex vivo 
immunological assays will consist of cytokine release by bulk PBL (± antigen 
stimulation) and by other experimental studies such as cytolysis if sufficient cells are 
available. If cell numbers are limiting, preference will be given to the direct analysis of 
immunological activity. Immunological assays will be standardized by the inclusion of 1) 
pre-infusion PBMC and 2) an aliquot of the engineered PBL cryopreserved at the time of 
infusion. In general, differences of 2- to 3-fold in these assays are indicative of true 
biologic differences.  
5.8 
MONITORING GENE THERAPY TRIALS: PERSISTENCE AND RCR 
• Engineered cell survival. CAR and vector presence will be quantitated in PBMC samples 
using established PCR techniques. Immunological monitoring using CAR-specific 
staining will be used to augment PCR-based analysis. This will provide data to estimate 
the in vivo survival of lymphocytes derived from the infused cells. In addition, 
measurement of CD4 and CD8 T-cells will be conducted and studies of these T-cell 
subsets in the circulation will be determined by using specific PCR assays capable of 
detecting the unique DNA sequence for each retroviral vector engineered T-cell. Note: 
samples will be batched and assayed at the conclusion of the study. 
• Patients will be co-enrolled on protocol 09-C-0161 and will adhere to the follow-up 
schedule described in that protocol. Patients’ blood samples will be obtained and undergo 
analysis for detection of RCR by PCR prior to cell infusion and RCR PCR will be 
performed at 3 and 6 months, and at one-year post cell administration. Blood samples 
will be archived annually thereafter if all previous testing has been negative with a brief 
clinical history. If a patient dies or develops neoplasms during this trial, efforts will be 
made to assay a biopsy sample for RCR. If any post-treatment samples are positive, 
further analysis of the RCR and more extensive patient follow-up will be undertaken, in 
consultation with the FDA. RCR PCR assays detect the GaLV envelop gene and are 
performed under contract by the Indiana University Vector Production Facility. The 
results of these tests are maintained by the contractor performing the RCR tests and by 
the NCI-SB research team. 
5.9 
SAMPLE STORAGE, TRACKING AND DISPOSITION FOR SURGERY BRANCH CELL 
PRODUCTION FACILITY 
Blood and tissue collected during the course of this study will follow the Cell Tracking and 
Labeling System established by the SB-CPF. The Cell Tracking and Labeling System is designed 
to unambiguously ensure that patient/data verification is consistent. The patients’ cell samples 
(blood or tissue) are tracked by distinct identification labels that include a unique patient 
identifier and date of specimen collection. Cryopreserved blood and tissue samples also bear the 


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date the sample was frozen. All cryopreserved samples are tracked for freezer location and 
storage criteria. All samples are stored in monitored freezers/refrigerators in 3NW NCI-SB 
laboratories at specified temperatures with alarm systems in place. Serum samples will be sent to 
the Blood Processing Core (BPC) for storage. Samples will be barcoded and stored on site or 
offsite at NCI Frederick Central Repository Services in Frederick, MD. All samples collected 
(blood or tissue) are entered into a central computer database with identification and storage 
location, and this database is backed up every night. 
If, at any time, a patient withdraws from the study and does not wish for their existing samples to 
be utilized, the individual must provide a written request. Following receipt of this request, the 
samples will be destroyed and reported as such to the IRB. Any samples lost (in transit or by a 
researcher) or destroyed due to unknown sample integrity (i.e. broken freezer allows for 
extensive sample thawing, etc.) will be reported as such to the IRB. 
Blood and tissue collected during the course of this study will be stored, tracked, and disposed of 
as specified in protocol 03-C-0277. 
5.10 SAMPLE STORAGE, TRACKING AND DISPOSITION FOR FIGG LAB 
5.10.1 Sample Data Collection 
All samples sent to the BPC will be barcoded, with data entered and stored in the LABrador (aka 
LabSamples) utilized by the BPC, and data will be updated to the NCI-SB central computer 
database weekly. This is a secure program, with access to LABrador limited to defined Figg Lab 
personnel, who are issued individual user accounts. Installation of LABrador is limited to 
computers specified by Dr. Figg. These computers all have a password-restricted login screen. 
All Figg Lab personnel with access to patient information annually complete the NIH online 
Protection of Human Subjects course.  
LABrador creates a unique barcode ID for every sample and sample box, which cannot be traced 
back to patients without LABrador access. The data recorded for each sample includes the 
patient ID, name, trial name/protocol number, time drawn, cycle time point, dose, material type, 
as well as box and freezer location. Patient demographics associated with the clinical center 
patient number are provided in the system. For each sample, there are notes associated with the 
processing method (delay in sample processing, storage conditions on the ward, etc.). 
5.10.2 Sample Storage and Destruction 
Barcoded samples are stored in barcoded boxes in a locked freezer at either -20 or -80°C 
according to stability requirements. These freezers are located onsite in the BPC and offsite at 
NCI Frederick Central Repository Services in Frederick, MD. Visitors to the laboratory are 
required to be accompanied by laboratory staff at all times.  
Access to stored clinical samples is restricted. Samples will be stored until requested by a 
researcher named on the protocol. All requests are monitored and tracked in LABrador. All 
researchers are required to sign a form stating that the samples are only to be used for research 
purposes associated with this trial (as per the IRB approved protocol) and that any unused 
samples must be returned to the BPC. It is the responsibility of the NCI PI to ensure that the 
samples requested are being used in a manner consistent with IRB approval. 


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Following completion of this study, samples will remain in storage as detailed above. Access to 
these samples will only be granted following IRB approval of an additional protocol, granting 
the rights to use the material.  
If, at any time, a patient withdraws from the study and does not wish for their existing samples to 
be utilized, the individual must provide a written request. Following receipt of this request, the 
samples will be destroyed, and reported as such to the IRB. Any samples lost (in transit or by a 
researcher) or destroyed due to unknown sample integrity (i.e. broken freezer allows for 
extensive sample thawing, etc.) will be reported as such to the IRB. 
Sample barcodes are linked to patient demographics and limited clinical information. This 
information will only be provided to investigators listed on this protocol, via registered use of the 
LABrador. It is critical that the sample remains linked to patient information such as race, age, 
dates of diagnosis and death, and histological information about the tumor, in order to correlate 
genotype with these variables. 
6 
DATA COLLECTION AND EVALUATION 
6.1 
DATA COLLECTION  
The PI will be responsible for overseeing entry of data into an in-house password-protected 
electronic system (C3D) and ensuring data accuracy, consistency and timeliness. The PI, 
associate investigators (AI), research nurses, and/or a contracted data manager will assist with 
the data management efforts. All data obtained during the conduct of the protocol will be kept in 
secure network drives or in approved alternative sites that comply with NIH security standards. 
Primary and final analyzed data will have identifiers so that research data can be attributed to an 
individual human subject participant. Data will be entered into the NCI CCR C3D database.  
All adverse events (AEs), including clinically significant abnormal findings on laboratory 
evaluations, regardless of severity, will be followed until return to baseline or stabilization of 
event. Patients will be followed for AEs until their first follow-up evaluation (4 weeks (± 7 days) 
following administration of the cell product) or until off-study, whichever comes first.  
An abnormal laboratory value will be recorded in the database as an AE only if the laboratory 
abnormality is characterized by any of the following: 
• Results in discontinuation from the study 
• Is associated with clinical signs or symptoms  
• Requires treatment or any other therapeutic intervention 
• Is associated with death or another serious adverse event, including hospitalization  
• Is judged by the Investigator to be of significant clinical impact 
• If any abnormal laboratory result is considered clinically significant, the investigator 
will provide details about the action taken with respect to the test drug and about the 
patient’s outcome. 
All AEs must be recorded on the AE case report form unless otherwise noted below in Section 
6.1.1. 


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End of study procedures: Data will be stored according to HHS and FDA regulations, and NIH 
Intramural Records Retention Schedule as applicable.  
Loss or destruction of data: Should we become aware that a major breach in our plan to protect 
subject confidentiality and trial data has occurred, the IRB will be notified. 
6.1.1 Exclusions to Routine Adverse Event Recording 
Patients will be receiving multiple agents, which include commercially available agents 
(fludarabine, cyclophosphamide, aldesleukin, and supportive medications) in combination with 
the investigational agents; therefore, grade 1 events not related to the cell product will not be 
reported/recorded. 
6.2 
DATA SHARING PLANS 
6.2.1 Human Data Sharing Plan 
De-identified human data generated for use in future and ongoing research will be shared 
through a NIH-funded or approved repository (ClinicalTrials.gov) and BTRIS. At the completion 
of data analysis, data will be submitted to ClinicalTrials.gov either before publication or at the 
time of publication or shortly thereafter. Data may also be used to support long-term research 
efforts within the NCI-SB, and de-identified data may also be shared with collaborators as 
specified in protocol 03-C-0277. 
6.2.2 Genomic Data Sharing Plan 
The NIH Genomic Data Sharing Policy does not apply to this study. 
6.3 
RESPONSE CRITERIA 
For the purpose of this study, patients will be evaluated for response 4 weeks (± 7 days) after cell 
infusion. If the patient has stable disease or tumor shrinkage, repeat complete evaluations will be 
performed monthly (± 7 days) as feasible for 12 months, and then every 1-2 months (± 7 days). 
In addition to a baseline scan, confirmatory scans should also be obtained at least 4 weeks (not 
less than 4) following initial documentation of objective response. 
Response and progression will be evaluated in this study using the international criteria proposed 
by the revised Response Evaluation Criteria in Solid Tumors (RECIST) guideline (version 1.0).  
As part of this trial, as well as to assist in the determination of tumor progression, all efforts will 
be made to observe radiographic changes in the patient’s tumors over time.  
6.3.1 Definitions 
Evaluable for toxicity: All patients will be evaluable for toxicity from the time of their first 
treatment with Cyclophosphamide. 
Evaluable for objective response: Only those patients who have measurable disease present at 
baseline, have received at least one cycle of therapy, and have had their disease re-evaluated will 
be considered evaluable for response. These patients will have their response classified 
according to the definitions stated below. (Note: Patients who exhibit objective disease 
progression prior to the end of treatment will also be considered evaluable.) 
Evaluable Non-Target Disease Response: Patients who have lesions present at baseline that are 
evaluable but do not meet the definitions of measurable disease, have received at least one cycle 


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of therapy, and have had their disease re-evaluated will be considered evaluable for non-target 
disease. The response assessment is based on the presence, absence, or unequivocal progression 
of the lesions. 
6.3.1.1 Objective Criteria for Measurable Lesions(41, 42):  
If there are too many measurable lesions to measure at each evaluation, choose the largest two to 
be followed before a patient is entered on study. The remaining lesions will be considered 
evaluable for the purpose of objective status determination. 
6.3.2 Disease Parameters 
6.3.2.1 Measurable Criteria 
Measurable disease: Bi-dimensionally contrast-enhancing lesions with clearly defined margins 
by MRI scan, with two perpendicular diameters of at least 10 mm, visible on two or more axial 
slices. Measurement of tumor around a cyst or surgical cavity represents a particularly difficult 
challenge. In general, such lesions should be considered nonmeasurable unless there is a nodular 
component measuring ≥ 10 mm in diameter. The cystic or surgical cavity should not be 
measured in determining response. 
Note: Measurable criteria always take precedence over evaluable criteria. For enhancing tumors, 
contrast-enhancement measurable or evaluable disease will be recorded. For non-enhancing 
tumors, which are a small percentage of high-grade gliomas, the T2/FLAIR measurable or 
evaluable disease will be recorded.  
Non-measurable but evaluable disease: Unidimensionally measurable lesions, masses with 
margins not clearly defined, or lesions with a multiple cystic component. 
Non-evaluable disease: No definitive, measurable or evaluable tumor. 
6.3.3 Evaluation of Measurable Lesions 
6.3.3.1 Definition of Response 
Complete Response (CR): Complete response requires all of the following: complete 
disappearance of all enhancing measurable and nonmeasurable disease sustained for at least 4 
weeks; no new lesions; stable or improved nonenhancing (T2/FLAIR) lesions; and patient must 
be off corticosteroids or on physiologic replacement doses only, and stable or improved 
clinically. In the absence of a confirming scan 4 weeks later, this response will be considered 
only stable disease. 
Partial Response (PR): Partial response requires all of the following: ≥50% decrease, compared 
with baseline, in the sum of products of perpendicular diameters of all measurable enhancing 
lesions sustained for at least 4 weeks; no progression of nonmeasurable disease; no new lesions; 
stable or improved nonenhancing (T2/FLAIR) lesions on same or lower dose of corticosteroids 
compared with baseline scan; and patient must be on a corticosteroid dose not greater than the 
dose at time of baseline scan and is stable or improved clinically. In the absence of a confirming 
scan 4 weeks later, this response will be considered only stable disease. 
Stable: Stable disease occurs if the patient does not qualify for complete response, partial 
response, or progression and requires the following: stable nonenhancing (T2/FLAIR) lesions on 
same or lower dose of corticosteroids compared with baseline scan and clinically stable status. In 
the event that the corticosteroid dose was increased for new symptoms and signs without 


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confirmation of disease progression on neuroimaging, and subsequent follow-up imaging shows 
that this increase in corticosteroids was required because of disease progression, the last scan 
considered to show stable disease will be the scan obtained when the corticosteroid dose was 
equivalent to the baseline dose. 
Progression: Progression is defined by any of the following: ≥ 25% increase in sum of the 
products of perpendicular diameters of enhancing lesions (compared to best response or with 
baseline if no decrease) on stable or increasing doses of corticosteroids; a significant increase in 
T2/FLAIR nonenhancing lesions on stable or increasing doses of corticosteroids compared with 
baseline scan or best response after initiation of therapy, not due to comorbid events; the 
appearance of any new lesions; clear progression of nonmeasurable lesions; or definite clinical 
deterioration not attributable to other causes apart from the tumor, or to decrease in 
corticosteroid dose. Failure to return for evaluation as a result of death or deteriorating condition 
should also be considered as progression. Patients with nonmeasurable enhancing disease whose 
lesions have significantly increased in size and become measurable (minimal bidirectional 
diameter of ≥10 mm and visible on at least two axial slices) will also be considered to have 
experienced progression. The transition from a nonmeasurable lesion to a measurable lesion 
resulting in progression can theoretically occur with relatively small increases in tumor size (e.g., 
a 9 x 9 mm lesion [nonmeasurable] increasing to a 10 x 11 mm lesion [measurable]). Ideally, the 
change should be significant (> 5 mm increase in maximal diameter or ≥ 25% increase in sum of 
the products of perpendicular diameters of enhancing lesions). In general, if there is doubt about 
whether the lesion has progressed, continued treatment and close follow-up evaluation will help 
clarify whether there is true progression.   
6.3.3.2 Evaluable Lesions 
Contrast enhancing evaluable lesions will be recorded at each evaluation. FLAIR or T2- 
weighted images should also be assessed as evaluable disease if appropriate. 
The following scale will be used to designate relative changes in MRI scans: 
+3 
disappearance of tumor (CR) 
+2 
definitely better (PR) 
+1 
possibly better 
0 
unchanged 
-1 
possibly worse 
-2 
definitely worse (PD) 
-3 
development of a new lesion (PD) 
6.3.4 Definition of Response for Evaluable Lesions 
Complete Response (CR): Defined as the circumstance when the MRI scan is ranked +3 and the 
tumor is no longer seen by neuroimaging, and the patient no longer requires steroids for control 
of tumor-induced cerebral edema.  
Partial Response (PR): Defined as a MRI scan ranked +2 provided that the patient has not had 
his/her dose of steroids increased since the last evaluation period.  


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Progression (PD): Defined as the circumstance when the MRI scan is ranked -2 or -3, or the 
presence of a new lesion. 
Stable disease (SD): Defined as the circumstance when the MRI scan shows no change or 
possible (-1 or +1) changes. Patients should be receiving stable or decreasing doses of steroids. 
6.3.5 Best Response  
Two assessments at least 4 weeks apart are required for a description of complete response and 
partial response. 
6.3.6 Central Radiology Review  
We will centrally (NCI) review all MRI scans that have been designated as a partial or complete 
response. Dr. John Butman of NIH Neuro-Radiology or his designee will assist in the review of 
all MRI scans. 
6.3.7 Neurological Examination 
Since it is not used for determining response, an objective assessment of the neurological exam 
will not be required data. Nevertheless, the neurological exam could be useful as a corollary 
piece of data to support or refute the validity of subtle MRI changes. Thus, we will request (but 
not require) that the following information will be recorded at each follow-up visit: 
• Normal versus abnormal neurologic exam 
• Status of neurologic exam compared to last exam 
o Definitely better 
o Possibly better 
o Stable 
o Possibly worse 
o Definitely worse 
• BARS assessment and score 
6.3.8 Progression-Free Survival  
Defined as the time from the date of registration to the date of first observation of progressive 
disease. If a patient was removed from a study for a reason other than progression, the patient 
will be censored for further evaluation of progression in that study as of the date of starting other 
therapy, if that was known. Otherwise, the date the patient was removed from the study will be 
used. If the patient was followed routinely for progression after being removed from the study 
and had progression without further therapy, that progression date will be used.  
6.3.9 Overall Survival 
Defined as the time from date of registration to date of death due to any cause. Patients not 
known to have died will be censored for survival as of the last date known alive. 
6.4 
TOXICITY CRITERIA 
The following adverse event management guidelines are intended to ensure the safety of each 
patient while on the study. The descriptions and grading scales found in the revised NCI 


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Common Terminology Criteria for Adverse Events (CTCAE) version 4.0 will be utilized for AE 
reporting. All appropriate treatment areas should have access to a copy of the CTCAE version 
4.0. A copy of the CTCAE version 4.0 can be downloaded from the CTEP web site 
(http://ctep.cancer.gov/protocolDevelopment/electronic_applications/ctc.htm#ctc_40). 
7 
SAFETY REPORTING REQUIREMENTS/DATA AND SAFETY MONITORING 
PLAN 
7.1 
DEFINITIONS 
7.1.1 Adverse Event 
Any untoward medical occurrence in a human subject, including any abnormal sign (for 
example, abnormal physical exam or laboratory finding), symptom, or disease, temporally 
associated with the subject’s participation in research, whether or not considered related to the 
subject’s participation in the research. 
7.1.2 Suspected Adverse Reaction 
Suspected adverse reaction means any adverse event for which there is a reasonable possibility 
that the drug caused the adverse event. For the purposes of IND safety reporting, ‘reasonable 
possibility’ means there is evidence to suggest a causal relationship between the drug and the 
adverse event. A suspected adverse reaction implies a lesser degree of certainty about causality 
than adverse reaction, which means any adverse event caused by a drug. 
7.1.3 Unexpected Adverse Reaction 
An adverse event or suspected adverse reaction is considered “unexpected” if it is not listed in 
the Investigator’s Brochure or is not listed at the specificity or severity that has been observed; 
or, if an Investigator’s Brochure is not required or available, is not consistent with the risk 
information described in the general investigational plan or elsewhere in the current application. 
"Unexpected”, also refers to adverse events or suspected adverse reactions that are mentioned in 
the Investigator’s Brochure as occurring with a class of drugs or as anticipated from the 
pharmacological properties of the drug, but are not specifically mentioned as occurring with the 
particular drug under investigation. 
7.1.4 Serious 
An Unanticipated Problem or Protocol Deviation is serious if it meets the definition of a Serious 
Adverse Event or if it compromises the safety, welfare or rights of subjects or others. 
7.1.5 Serious Adverse Event 
An adverse event or suspected adverse reaction is considered serious if in the view of the 
investigator or the sponsor, it results in any of the following: 
• Death 
• A life-threatening adverse drug experience 
• Inpatient hospitalization or prolongation of existing hospitalization 
• A persistent or significant disability/incapacity or substantial disruption of the ability to 
conduct normal life functions 
• A congenital anomaly/birth defect 


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• Important medical events that may not result in death, be life-threatening, or require 
hospitalization may be considered a serious adverse drug experience when, based upon 
appropriate medical judgment, they may jeopardize the patient or subject and may require 
medical or surgical intervention to prevent one of the outcomes listed in this definition 
7.1.6 Disability 
A substantial disruption of a person’s ability to conduct normal life functions. 
7.1.7 Life-Threatening Adverse Drug Experience 
Any adverse event or suspected adverse reaction that places the patient or subject, in the view of 
the investigator or sponsor, at immediate risk of death from the reaction as it occurred, i.e., it 
does not include a reaction that had it occurred in a more severe form, might have caused death. 
7.1.8 Protocol Deviation (NIH Definition) 
Any change, divergence, or departure from the IRB approved research protocol. 
7.1.9 Non-Compliance (NIH Definition) 
The failure to comply with applicable NIH Human Research Protections Program (HRPP) 
policies, IRB requirements, or regulatory requirements for the protection of human research 
subjects.  
7.1.10 Unanticipated Problem 
Any incident, experience, or outcome that: 
1. Is unexpected in terms of nature, severity, or frequency in relation to: 
a. the research risks that are described in the IRB-approved research protocol and 
informed consent document; Investigator’s Brochure or other study documents, 
and 
b. the characteristics of the subject population being studied; AND 
2. Is related or possibly related to participation in the research; AND 
3. Suggests that the research places subjects or others at a greater risk of harm (including 
physical, psychological, economic, or social harm) than was previously known or 
recognized. 
7.2 
NCI-IRB AND CLINICAL DIRECTOR (CD) REPORTING 
7.2.1 NCI-IRB and NCI CD Expedited Reporting of Unanticipated Problems and Deaths 
The Protocol PI will report in the NIH Problem Form to the NCI-IRB and NCI Clinical Director: 
• All deaths, except deaths due to progressive disease 
• All protocol deviations  
• All unanticipated problems 
• All non-compliance  
Reports must be received within 7 days of PI awareness via iRIS. 


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7.2.2 NCI-IRB Requirements for PI Reporting at Continuing Review 
The protocol PI will report to the NCI-IRB:  
1. A summary of all protocol deviations in a tabular format to include the date the deviation 
occurred, a brief description of the deviation and any corrective action. 
2. A summary of any instances of non-compliance. 
3. A tabular summary of the following adverse events: 
• 
All Grade 2 unexpected events that are possibly, probably or definitely related to 
the research  
• 
All Grade 3 and 4 events that are possibly, probably or definitely related to the 
research  
• 
All Grade 5 events regardless of attribution  
• 
All serious events regardless of attribution.  
Note: Grade 1 events are not required to be reported. 
7.2.3 NCI-IRB Reporting of IND Safety Reports 
Only IND Safety Reports that meet the definition of an unanticipated problem will need to be 
reported to the NCI-IRB. 
7.3 
IND SPONSOR REPORTING CRITERIA 
From the time the subject receives the investigational agent/intervention to the time of the first 
follow-up evaluation (4 weeks (± 7 days) following the administration of the cell product), the 
investigator must immediately report to the sponsor, using the mandatory MedWatch Form FDA 
3500A or equivalent, any serious adverse event, whether or not considered drug-related, 
including those listed in the protocol or Investigator’s Brochure and must include an assessment 
of whether there is a reasonable possibility that the drug caused the event. For serious adverse 
events that occur after the first follow-up evaluation, only those events that have an attribution of 
at least possibly related to the agent/intervention will be reported. 
Required timing for reporting per the above guidelines: 
• Deaths (except death due to progressive disease) must be reported via email within 24 
hours. A complete report must be submitted within one business day.  
• Other serious adverse events as well as deaths due to progressive disease must be 
reported within one business day 
Events will be submitted to the Center for Cancer Research (CCR) at: CCRsafety@mail.nih.gov 
and to the CCR PI and study coordinator. 
7.3.1 Waiver of Expedited Reporting to CCR 
The investigators are requesting a waiver from reporting specific events in an expedited manner 
to the CCR. Patients will be receiving commercially available agents, such as fludarabine, 
cyclophosphamide, and aldesleukin. The majority of toxicities observed on NCI-SB adoptive cell 
therapy (ACT) protocols are expected toxicities of the non-myeloablative, lymphodepleting 
preparative regimen or IL-2 and occur in approximately 95% of the patients enrolled, therefore, 


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we are requesting a waiver from reporting the following events in an expedited manner to the 
CCR.  
• Grade 3 or greater myelosuppression, defined as lymphopenia, neutropenia, decreased 
hemoglobin, and thrombocytopenia. 
• Grade 3 or greater nausea, vomiting, mucositis - oral, anorexia, diarrhea, fever, chills, 
fatigue, and rash maculo-papular. 
• Grade 3 hypoxia, dyspnea, hematuria, hypotension, sinus tachycardia, urine output 
decreased, confusion, infections, and febrile neutropenia. 
The PI will submit a summary table of all grade 3-5 events, whether or not considered related to 
the product, every 6 months. The report shall include the number of patients treated in the 
timeframe, the number of events per AE term per grade which occurred in the 6-month 
timeframe and in total since the start of the study, attribution, and type/category of serious. 
Reports will be submitted to the Center for Cancer Research (CCR) at: 
CCRsafety@mail.nih.gov. 
7.3.2 Reporting Pregnancy  
7.3.2.1 Maternal Exposure 
If a patient becomes pregnant during treatment and for the first four months following treatment, 
the study treatment should be discontinued immediately and the pregnancy should be reported to 
the Sponsor. The potential risk of exposure of the fetus to the investigational agent(s) or 
chemotherapy agents(s) should be documented in box B5 of the Medwatch form “Describe 
Event or Problem”. 
Pregnancy itself is not regarded as an SAE. However, as patients who become pregnant on study 
risk intrauterine exposure of the fetus to agents which may be teratogenic, the CCR is requesting 
that pregnancy should be reported in an expedited manner as grade 3 “Pregnancy, puerperium 
and perinatal conditions - Other (pregnancy)” under the Pregnancy, puerperium and perinatal 
conditions SOC.  
Congenital abnormalities or birth defects and spontaneous miscarriages should be reported and 
handled as SAEs. Elective abortions without complications should not be handled as AEs. The 
outcome of all pregnancies (spontaneous miscarriage, elective termination, ectopic pregnancy, 
normal birth, or congenital abnormality) should be followed up and documented. 
If any pregnancy occurs in the course of the study, then the investigator should inform the 
Sponsor within one day, i.e., immediately, but no later than 24 hours of when he or she 
becomes aware of it. 
The designated Sponsor representative will work with the investigator to ensure that all relevant 
information is provided to the Sponsor within 1 to 5 calendar days for SAEs and within 30 days 
for all other pregnancies. 
The same timelines apply when outcome information is available. 
7.3.2.2 Paternal Exposure 
Male patients should refrain from fathering a child or donating sperm during the study and for 
120 days after the last dose of aldesleukin. 


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Pregnancy of the patient’s partner is not considered to be an AE. However, the outcome of all 
pregnancies (spontaneous miscarriage, elective termination, ectopic pregnancy, normal birth, or 
congenital abnormality) occurring from the date of the first dose until 120 days after the last dose 
should, if possible, be followed up and documented. 
7.4 
INSTITUTIONAL BIOSAFETY COMMITTEE (IBC) REPORTING CRITERIA 
7.4.1 Serious Adverse Event Reports to IBC 
The PI (or delegate) will notify IBC of any unexpected fatal or life-threatening experience 
associated with the use of the cell product as soon as possible but in no event later than 7 
calendar days of initial receipt of the information. Serious adverse events that are unexpected and 
associated with the use of the cell product, but are not fatal or life-threatening, must be reported 
to the NIH IBC as soon as possible, but not later than 15 calendar days after the investigator’s 
initial receipt of the information. Adverse events may be reported by using the FDA Form FDA 
3500A or equivalent. 
7.4.2 Annual Reports to IBC 
Within 60 days after the one-year anniversary of the date on which the IBC approved the initial 
protocol, and after each subsequent anniversary until the trial is completed, the PI (or delegate) 
shall submit the information described below. Alternatively, the IRB continuing review report 
can be sent to the IBC in lieu of a separate report. Please include the IBC protocol number on the 
report. 
7.4.2.1 Clinical Trial Information  
A brief summary of the status of the trial in progress or completed during the previous year. The 
summary is required to include the following information:  
• Title and purpose of the trial  
• Clinical site 
• Principal Investigator 
• Clinical protocol identifiers  
• Participant population (such as disease indication and general age group, e.g., adult or 
pediatric);  
• Total number of participants planned for inclusion in the trial; the number entered into 
the trial to date whose participation in the trial was completed; and the number who 
dropped out of the trial with a brief description of the reasons  
• Status of the trial, e.g., open to accrual of subjects, closed but data collection ongoing, or 
fully completed  
• If the trial has been completed, a brief description of any study results.  
7.4.2.2 Progress Report and Data Analysis  
Information obtained during the previous year's clinical and non-clinical investigations, 
including:  


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• Narrative or tabular summary showing the most frequent and most serious adverse 
experiences by body system 
• Summary of all serious adverse events submitted during the past year 
• Summary of serious adverse events that were expected or considered to have causes not 
associated with the use of the gene transfer product such as disease progression or 
concurrent medications 
• If any deaths have occurred, the number of participants who died during participation in 
the investigation and causes of death 
• Brief description of any information obtained that is pertinent to an understanding of the 
gene transfer product’s actions, including, for example, information about dose-response, 
information from controlled trials, and information about bioavailability. 
7.5 
DATA AND SAFETY MONITORING PLAN 
Careful evaluation to ascertain the toxicity, immunologic effects and anti-tumor efficacy of cell 
infusions will be performed. Due to the nature of these studies, it is possible that expansion of 
specific T-cell clones will be observed as tumor-reactive T-cell proliferate in response to tumor 
antigens. Therefore, care will be taken to track T-cell persistence both immunologically and 
molecularly according to plan specified in sections 5.7 and 5.8.  
7.5.1 Principal Investigator/Research Team 
The clinical research team will meet on a regular basis when patients are being actively treated 
on the trial to discuss each patient. Decisions about enrollment will be made based on the 
toxicity data from prior patients.  
All data will be collected in a timely manner and reviewed by PI or AI. Adverse events will be 
reported as required above. Any safety concerns, new information that might affect either the 
ethical and or scientific conduct of the trial, or protocol deviations will be reported to the IRB 
using iRIS.  
The PI will review adverse event and response data on each patient to ensure safety and data 
accuracy. The PI will personally conduct or supervise the investigation and provide appropriate 
delegation of responsibilities to other members of the research staff. 
7.5.2 Sponsor Monitoring Plan 
As a sponsor for clinical trials, FDA regulations require the CCR to maintain a monitoring 
program. The CCR’s program allows for confirmation of: study data, specifically data that could 
affect the interpretation of primary study endpoints; adherence to the protocol, regulations, and 
SOPs; and human subject’s protection. This is done through independent verification of study 
data with source documentation focusing on: 
• Informed consent process 
• Eligibility confirmation 
• Drug administration and accountability 
• Adverse events monitoring 


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• Response assessment. 
The monitoring program also extends to multi-site research when the CCR is the coordinating 
center. 
This trial will be monitored by personnel employed by a CCR contractor. Monitors are qualified 
by training and experience to monitor the progress of clinical trials. Personnel monitoring this 
study will not be affiliated in any way with the trial conduct. 
7.5.3 Safety Monitoring Committee (SMC) 
This protocol will require oversight from the Safety Monitoring Committee (SMC). Initial 
review will occur as soon as possible after the annual NCI-IRB continuing review date. 
Subsequently, each protocol will be reviewed as close to annually as the quarterly meeting 
schedule permits or more frequently as may be required by the SMC. For initial and subsequent 
reviews, protocols will not be reviewed if there is no accrual within the review period. Written 
outcome letters will be generated in response to the monitoring activities and submitted to the PI 
and Clinical Director or Deputy Clinical Director, CCR, NCI. 
8 
STATISTICAL CONSIDERATIONS 
The primary objectives of this study are to determine the safety and six-month progression free 
survival of patients receiving anti-EGFRvIII CAR-engineered PBL and aldesleukin following a 
non-myeloablative, lymphodepleting preparative regimen. The secondary objectives are to 
determine the in vivo survival of CAR gene-engineered cells, and to evaluate radiographic 
changes after treatment.  
The protocol will begin with a Phase I dose escalation design, with eight dose levels and with 
two different cohorts; one for patients receiving steroids at the time of treatment (Cohort 1A) and 
one for patients not receiving steroids at the time of treatment (Cohort 1B). Each cohort in Phase 
I will be escalated independently of the other, and different MTDs may be identified. With 
Amendment H, patients were no longer divided among two separate cohorts for the Phase I dose 
escalation portion of the trial. Thus, dose levels 6-8 included all patients (those who required 
steroids and those who did not). In addition, the safety assessment period was revised as 
described in Section 3.1.3.2. With Amendment K, three dose levels (9, 9A, 10) and two de-
escalation levels (9D-1 and 9D-2) were added to the Phase I dose escalation portion. Prior to 
receiving the engineered PBL cells, patients will receive a non-myeloablative, lymphodepleting 
preparative regimen consisting of cyclophosphamide and fludarabine followed in 2-4 days by 
intravenous infusion of ex vivo tumor-reactive, EGFRvIII CAR gene-transduced PBMC, plus IV 
high- or low-dose aldesleukin (720,000 or 72,000 IU/kg every 8 hours for a maximum of 15 
doses). 
Refer to Section 3.1 for additional details.  
In the Phase II portion of the trial, patients treated with (Cohort 2A) or without (Cohort 2B) 
steroids will be evaluated separately based on the dose found appropriate in Phase I, using the 
same single-stage Phase II trial design. The objective of this part of the trial is to determine if the 
protocol treatment is able to result in a 6-month progression-free survival of 30% rather than the 
historical value for recurrent glioblastoma of 10% based on a recent study(43). In this portion of 
the trial, we will plan to accrue 32 patients in each cohort. With 32 patients, an exact binomial 


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test with a 0.10 one-sided significance level will have 94% power to detect the difference 
between 10% as the 6 month PFS probability and 30% as the 6 month PFS probability. 
In addition to determining the 6-month PFS probability, a Kaplan-Meier curve of PFS will be 
constructed for each cohort. 
In order to ensure that patients are not receiving a treatment which may be associated with worse 
results than those normally considered to be available, an early stopping rule will be imposed 
independently for each cohort. If after 16 patients have been enrolled and potentially followed 
for 6 months in either cohort, fewer than 10% of patients have been progression-free for 6 
months, then no further patients will be enrolled in that particular cohort as soon as this has been 
determined. 
As of Amendment Q, 18 patients have been accrued. The last patient treated on study at Dose 
Level 9 (the first patient treated at this dose level) experienced a DLT. Thus, the next patient will 
be treated at Dose Level 9D-1.  
Enrollment Summary per Dose Level 
Dose Level 
Dose of Anti-EGFRvIII CAR T-Cells  
Total Patients Enrolled 
Level 1 (Cohort 1A) – Completed  
107 
1 
Level 1 (Cohort 1B) – Completed  
107 
1 
Level 2 (Cohort 1A) – Completed  
3x107 
1 
Level 2 (Cohort 1B) – Completed  
3x107 
1 
Level 3 (Cohort 1A) – Completed  
108 
1 
Level 3 (Cohort 1B) – Completed  
108 
1 
Level 4 (Cohort 1A) – Eliminated 
3x108 
0 
Level 4 (Cohort 1B) – Completed  
3x108 
1 
Level 5 (Cohort 1A) – Eliminated 
109 
0 
Level 5 (Cohort 1B) – Completed 
109 
3 
Level 6 (All patients) – Completed  3x109 
3 
Level 7 (All patients) – Completed  1010 
3 
Level 8 (All patients) – Closed  
3 – 6x1010  
1 
Level 9 (All patients) 
3x1010 given as a split dose, 2 hours apart 
1 
Level 9A (All patients) 
3x1010 given as a single dose 
0 
Level 9D-1 (All patients)* 
1010 
0 
Level 9D-2 (All patients) 
3x109 
0 
Level 10 (All patients) 
3x1010 
0 
 
*Next enrolled patient will be treated at this dose level 
 


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To complete the Phase I dose-escalation portion, and both cohorts in the Phase II portion of the 
study, a total of up to 107 patients may be required (including up to 17 patients already 
accrued)—approximately 9 to 12 patients for dose levels 9 and 10 (6 patients will be the 
maximum for dose level 9D or 10), 2 additional patients in case patients need to be replaced for 
any reason, and two Phase II cohorts with a maximum of 32 each. Provided that about 1-2 
patients per month will be able to be enrolled onto this trial, approximately 7 years may be 
required to enroll up to 107 patients onto this trial.  
9 
COLLABORATIVE AGREEMENTS  
We have established a Cooperative Research and Development Agreement (CRADAs #02716 
and #03168) with Kite Pharma, Inc., and will be sharing data with them. 
10 HUMAN SUBJECTS PROTECTIONS 
10.1 RATIONALE FOR SUBJECT SELECTION 
The patients to be entered in this protocol have malignant gliomas which are refractory to 
standard therapy, and have limited life expectancies. 
Subjects from both genders and all racial/ethnic groups are eligible for this study if they meet the 
eligibility criteria. To date, there is no information that suggests that differences in drug 
metabolism or disease response would be expected in one group compared to another. Efforts 
will be made to extend accrual to a representative population, but in this preliminary study, a 
balance must be struck between patient safety considerations and limitations on the number of 
individuals exposed to potentially toxic and/or ineffective treatments on the one hand and the 
need to explore gender and ethnic aspects of clinical research on the other hand. If differences in 
outcome that correlate to gender or to ethnic identity are noted, accrual may be expanded or a 
follow-up study may be written to investigate those differences more fully. 
10.2 PARTICIPATION OF CHILDREN 
The use of the non-myeloablative, lymphodepleting preparative regimen in this protocol is a 
major procedure which entails serious discomforts and hazards for the patient, such that fatal 
complications are possible. It is therefore only appropriate to carry out this experimental 
procedure in the context of life threatening malignant gliomas. Since the efficacy of this 
experimental procedure is unknown, it does not seem reasonable to expose children to this risk 
without further evidence of benefit.  
Should results of this study indicate efficacy in treating malignant gliomas, which is not 
responsive to other standard forms of therapy, future research can be conducted in the pediatric 
population to evaluate potential benefit in that patient population.  
10.3 PARTICIPATION OF SUBJECTS UNABLE TO GIVE CONSENT 
Adults unable to give consent are excluded from enrolling in the protocol. However, re-consent 
may be necessary and there is a possibility, though unlikely, that subjects could become 
decisionally impaired. For this reason and because there is a prospect of direct benefit from 
research participation (Section 10.5), all subjects ≥ age 18 will be offered the opportunity to fill 
in their wishes for research and care, and assign a substitute decision maker on the “NIH 
Advance Directive for Health Care and Medical Research Participation” form so that another 
person can make decisions about their medical care in the event that they become incapacitated 
or cognitively impaired during the course of the study. The PI or AI will contact the NIH Ability 


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to Consent Assessment Team (ACAT) for evaluation as needed for the following: an 
independent assessment of whether an individual has the capacity to provide consent; assistance 
in identifying and assessing an appropriate surrogate when indicated; and/or an assessment of the 
capacity to appoint a surrogate. For those subjects that become incapacitated and do not have 
pre-determined substitute decision maker, the procedures described in MAS Policy 87-4 and 
NIH HRPP SOP 14E for appointing a surrogate decision maker for adult subjects who are (a) 
decisionally impaired, and (b) who do not have a legal guardian or durable power of attorney, 
will be followed. 
10.4 EVALUATION OF BENEFITS AND RISKS/DISCOMFORTS 
The experimental treatment has a chance to provide clinical benefit though this is unknown. The 
NCI-SB has extensive experience with ACT following treatment with high-dose aldesleukin, 
however, this experimental treatment is only available at a very few centers throughout the 
country. Although we have seen responses to this treatment, we do not know if this change in our 
process will improve patient outcome. The risks associated with ACT are substantial, including a 
delay in treatment due to the need to harvest and grow the cells; a surgical procedure (possibly 
major) to obtain tumor for the cell product; the possibility that a cell product cannot be 
generated; infection and sepsis due to the non-myeloablative, lymphodepleting preparative 
regimen; intubation; renal toxicities due to aldesleukin; and death. The risks in this treatment are 
detailed in Section 11.  
10.5 RISK/BENEFIT ANALYSIS 
Because all patients in this protocol have malignant glioma and limited life expectancies, the 
potential benefit is thought to outweigh the potential risks. The risk/benefit analysis for adults 
with the capacity to consent, as well as for adults who may become unable to provide consent, is 
greater than minimal risk but presenting the prospect of direct benefit to individual subjects 
based on the risks and potential benefits described in Section 10.4.   
10.6 CONSENT PROCESS AND DOCUMENTATION 
Patients are initially consented on protocols 99-C-0128 and 03-C-0277. If the lymphocytes can 
be generated for infusion and the patient meets the thorough screening for eligibility, the patient, 
with family members or friends at the request of the patient, will be presented with a detailed 
description of the protocol treatment. The procedures and tests involved in this study and the 
associated risks, discomforts and benefits of these processes, will be carefully explained to the 
patient and a signed informed consent document will be obtained prior to entry onto the study. 
The informed consent document is given to the patient, who is requested to review it and to ask 
questions prior to agreeing to participate. The patient is reassured that participation on trial is 
entirely voluntary and that he/she can withdraw or decide against treatment at any time without 
adverse consequences. The PI, AI, or clinical fellow is responsible for obtaining written consent 
from the patient.  
10.6.1 Informed Consent of Non-English Speaking Subjects 
If there is an unexpected enrollment of a research participant for whom there is no translated 
extant IRB-approved consent document, the PI and/or those authorized to obtain informed 
consent will use the Short Form Oral Consent Process as described in MAS Policy M77-2, 
OHSRP SOP 12, 45 CFR 46.117 (b) (2), and 21 CFR 50.27 (b) (2). The summary that will be 
used is the English version of the extant IRB approved consent document. Signed copies of both 


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the English version of the consent and the translated short form will be given to the subject or 
their legally authorized representative and the signed original will be filed in the medical record. 
Unless the PI is fluent in the prospective subject’s language, an interpreter will be present to 
facilitate the conversation (using either the long-translated form or the short form). Preferably 
someone who is independent of the subject (i.e., not a family member) will assist in presenting 
information and obtaining consent. Whenever possible, interpreters will be provided copies of 
the relevant consent documents well before the consent conversation with the subject (24 to 48 
hours if possible).  
We request prospective IRB approval of the use of the short form process for non-English 
speaking subjects and will notify the IRB at the time of continuing review of the frequency of the 
use of the Short Form. 
11 PHARMACEUTICAL INFORMATION 
Cyclophosphamide, fludarabine, and aldesleukin, the commercial drugs used in this study, will 
not alter labeling of the FDA-approved drugs. The investigation is not intended to support a new 
indication for use or any other significant changes to labeling or advertising in 
cyclophosphamide, fludarabine, or aldesleukin. The investigation does not involve a route of 
administration or dosage level in use in a patient population or other factor that significantly 
increases the risks (or decreases the acceptability of the risks) associated with the use of the drug 
products. 
11.1 INVESTIGATIONAL REGIMEN 
11.1.1 Anti-EGFRvIII CAR Transduced PBL 
The procedure for expanding the human PBL and the CoA are similar to those approved by the 
Food and Drug Administration, and used at the NCI in ongoing protocols evaluating cell therapy 
in the NCI-SB. The CoA is included in Appendix E and in the IND submission for these cells. 
The PBL will be transduced with retroviral supernatant containing the chimeric anti-EGFRvIII 
CAR. 
11.1.2 Retroviral Vector Containing the anti-EGFRvIII CAR Gene 
The retroviral vector supernatant (PG13-139-CD8-CD28BBZ (F10) encoding a CAR directed 
against the antigen, EGFRvIII, was prepared and preserved following cGMP conditions in the 
SB-CPF. The retroviral vector utilizes the MSGV1 retroviral vector backbone and consists of 
4,032 bps including the 5' LTR from the murine stem cell virus (promoter), packaging signal 
including the splicing donor (SD) and splicing acceptor sites, human anti-EGFRvIII scFv-based 
(mAb 139) CAR protein containing a signal peptide signal (human GM-CSFR), 139 light chain 
variable region, linker peptide, 139 heavy chain variable region, CD8 (hinge, transmembrane), 
CD28 ( cytoplasmic region), 4-1BB (cytoplasmic region) and TCR zeta (cytoplasmic region), 
followed by the murine stem cell virus 3'LTR. The physical titer will be determined by RNA dot 
blot according to sponsor certificate. The supernate will be stored at SBVPF upon the completion 
of production at –80° C with around-the-clock temperature monitoring. Upon request, 
supernatant will be delivered on dry ice to be used in in vitro transduction. There will be no re-
use of the same unit of supernate for different patients. Retroviral titer has been shown to be 
stable after immediate thawing and immediate administration (coating the tissue culture wells 
previously coated with Retronectin). Handling of the vector should follow the guidelines of 


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Biosafety Level-2 (BSL-2). The specific guidelines for Biosafety Level-2 (BSL-2) can be viewed 
at http://bmbl.od.nih.gov/sect3bsl2.htm.  
Note: Penicillin, streptomycin, and gentamycin will not be used in the manufacture of products 
for patients with documented allergies to these drugs. 
11.1.3 Interleukin-2 (Aldesleukin, Proleukin, Recombinant Human Interleukin 2) 
How Supplied: Interleukin-2 (aldesleukin) will be provided by the NIH Clinical Pharmacy 
Department from commercial sources. 
Formulation/Reconstitution: Aldesleukin, NSC #373364, is provided as single-use vials 
containing 22 million IU (−1.3 mg) IL-2 as a sterile, white to off-white lyophilized cake plus 50 
mg mannitol and 0.18 mg sodium dodecyl sulfate, buttered with approximately 0.17 mg 
monobasic and 0.89 mg dibasic sodium phosphate to a pH of 7.5 (range 7.2 to 7.8). The vial is 
reconstituted with 1.2 mL of Sterile Water for Injection, USP, and the resultant concentration is 
18 million IU/mL or 1.1 mg/mL. Diluent should be directed against the side of the vial to avoid 
excess foaming. Swirl contents gently until completely dissolved. Do not shake. Since vials 
contain no preservative, reconstituted solution should be used with 24 hours. 
Storage: Intact vials are stored in the refrigerator (2-8°C) protected from light. Each vial bears an 
expiration date. 
Dilution/Stability: Reconstituted aldesleukin should be further diluted with 50 mL of 5% Human 
Serum Albumin (HSA). The HSA should be added to the diluent prior to the addition of RIL-2. 
Dilutions of the reconstituted solution over a 1000-fold range (i.e., 1 mg/mL to 1 mcg/mL) are 
acceptable in either glass bottles or polyvinyl chloride bags. Aldesleukin is chemically stable for 
48 hours at refrigerated and room temperatures, 2-30°C. 
Administration: The dosage will be calculated based on total body weight. The final dilution of 
aldesleukin will be infused over 15 minutes. Aldesleukin will be administered as an inpatient.  
Toxicities: Expected toxicities of aldesleukin are listed in the product label and in Appendix B 
and Appendix C. Grade 3 toxicities common to aldesleukin include diarrhea, nausea, vomiting, 
hypotension, skin changes, anorexia, mucositis, dysphagia, or constitutional symptoms and 
laboratory changes as detailed in Appendix B. Additional grade 3 and 4 toxicities seen with 
aldesleukin are detailed in Appendix C.  
11.1.4 Fludarabine 
 (Please refer to the FDA-approved package insert for complete product information)  
Description: Fludarabine phosphate is a synthetic purine nucleoside that differs from physiologic 
nucleosides in that the sugar moiety is arabinose instead of ribose or deoxyribose. Fludarabine is 
a purine antagonist antimetabolite.  
How Supplied: It will be purchased by the NIH Clinical Pharmacy Department from commercial 
sources. Fludarabine is supplied in a 50 mg vial as a fludarabine phosphate powder in the form of 
a white, lyophilized solid cake.  
Stability: Following reconstitution with 2 mL of sterile water for injection to a concentration of 
25 mg/mL, the solution has a pH of 7.7. The fludarabine powder is stable for at least 18 months 
at 2-8°C; when reconstituted, fludarabine is stable for at least 16 days at room temperature. 
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within 8 hours. Specialized references should be consulted for specific compatibility 
information. Fludarabine is dephosphorylated in serum, transported intracellularly and converted 
to the nucleotide fludarabine triphosphate; this 2-fluoro-ara-ATP molecule is thought to be 
required for the drug’s cytotoxic effects. Fludarabine inhibits DNA polymerase, ribnucleotide 
reductase, DNA primase, and may interfere with chain elongation, and RNA and protein 
synthesis. 
Storage: Intact vials should be stored refrigerated (2-8°C). 
Administration: Fludarabine is administered as an IV infusion in 100 mL 0.9% sodium chloride, 
USP over 15-30 minutes. The doses will be based on body surface area (BSA). If patient is obese 
(BMI > 35), drug dosage will be calculated using practical weight as described in Appendix A.  
Toxicities: At doses of 25 mg/m2/day for 5 days, the primary side effect is myelosuppression; 
however, thrombocytopenia is responsible for most cases of severe and life-threatening 
hematologic toxicity. Serious opportunistic infections have occurred in CLL patients treated with 
fludarabine. Hemolytic anemia has been reported after one or more courses of fludarabine with 
or without a prior history of a positive Coomb’s test; fatal hemolytic anemia has been reported. 
In addition, bone marrow fibrosis has been observed after fludarabine therapy. Other common 
adverse effects include malaise, fever, chills, fatigue, anorexia, nausea and vomiting, and 
weakness. Irreversible and potentially fatal central nervous system toxicity in the form of 
progressive encephalopathy, blindness, and coma is only rarely observed at the currently 
administered doses of fludarabine. More common neurologic side effects at the current doses of 
fludarabine include weakness, pain, malaise, fatigue, paresthesia, visual or hearing disturbances, 
and sleep disorders. Adverse respiratory effects of fludarabine include cough, dyspnea, allergic 
or idiopathic interstitial pneumonitis. Tumor lysis syndrome has been rarely observed in 
fludarabine treatment of CLL. Treatment on previous ACT protocols in the NCI-SB have caused 
persistently low (below 200) CD4 counts, and one patient developed polyneuropathy manifested 
by vision blindness, and motor and sensory defects. 
11.1.5 Cyclophosphamide 
(Please refer to FDA-approved package insert for complete product information) 
Description: Cyclophosphamide is a nitrogen mustard-derivative alkylating agent. Following 
conversion to active metabolites in the liver, cyclophosphamide functions as an alkyating agent; 
the drug also possesses potent immunosuppressive activity. The serum half-life after IV 
administration ranges from 3-12 hours; the drug and/or its metabolites can be detected in the 
serum for up to 72 hours after administration.  
How Supplied: Cyclophosphamide will be obtained from commercially available sources by the 
Clinical Center Pharmacy Department.  
Stability: Following reconstitution as directed with sterile water for injection, cyclophosphamide 
is stable for 24 hours at room temperature or 6 days when kept at 2-8°C.  
Administration: It will be diluted in 250 mL D5W and infused over one hour. The dose will be 
based on the patient’s body weight. If patient is obese (BMI > 35), drug dosage will be 
calculated using practical weight as described in Appendix A. 
Toxicities: Hematologic toxicity occurring with cyclophosphamide usually includes leukopenia 
and thrombocytopenia. Anorexia, nausea and vomiting, rash and alopecia occur, especially after 


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high-dose cyclophosphamide; diarrhea, hemorrhagic colitis, infertility, and mucosal and oral 
ulceration have been reported. Sterile hemorrhagic cystitis occurs in about 20% of patients; 
severity can range from microscopic hematuria to extensive cystitis with bladder fibrosis. 
Although the incidence of hemorrhagic cystitis associated with cyclophosphamide appears to be 
lower than that associated with ifosfamide, mesna (sodium 2-mercaptoethanesulfonate) has been 
used prophylactically as a uroprotective agent in patients receiving cyclophosphamide. 
Prophylactic mesna is not effective in preventing hemorrhagic cystitis in all patients. Patients 
who receive high dose cyclophosphamide may develop interstitial pulmonary fibrosis, which can 
be fatal. Hyperuricemia due to rapid cellular destruction may occur, particularly in patients with 
hematologic malignancy. Hyperuricemia may be minimized by adequate hydration, 
alkalinization of the urine, and/or administration of allopurinol. If allopurinol is administered, 
patients should be watched closely for cyclophosphamide toxicity (due to allopurinol induction 
of hepatic microsomal enzymes). At high doses, cyclophosphamide can result in a syndrome of 
inappropriate antidiuretic hormone secretion; hyponatremia with progressive weight gain without 
edema occurs. At high doses, cyclophosphamide can result in cardiotoxicity. Deaths have 
occurred from diffuse hemorrhagic myocardial necrosis and from a syndrome of acute 
myopericarditis; in such cases, congestive heart failure may occur within a few days of the first 
dose. Other consequences of cyclophosphamide cardiotoxicity include arrhythmias, potentially 
irreversible cardiomyopathy, and pericarditis. Other reported adverse effects of 
cyclophosphamide include headache, dizziness, and myxedema; faintness, facial flushing, and 
diaphoresis have occurred following IV administration. Mesna (sodium 2-
mercaptoethanesulphonate; given by IV injection) is a synthetic sulfhydryl compound that can 
chemically interact with urotoxic metabolites of cyclophosphamide (acrolein and 4-
hydroxycyclophosphamide) to decrease the incidence and severity of hemorrhagic cystitis.  
11.2 SUPPORT MEDICATIONS 
11.2.1 Mesna (Sodium 2-mercaptoethanesulfonate, Mesnum, Mesnex, NSC-113891) 
(Please refer to the FDA-approved package insert for complete product information) 
Description: Mesna will be obtained commercially by the Clinical Center Pharmacy Department 
and is supplied as a 100 mg/mL solution.  
Storage: Intact ampoules are stored at room temperature.  
Stability: Diluted solutions (1-20 mg/mL) are physically and chemically stable for at least 24 
hours under refrigeration. Mesna is chemically stable at room temperature for 48-72 hours in 
D5W, 48-72 hour in D5W/0.45% NaCl, or 24 hours in 0.9% NaCl. 
Administration: Dilute to concentrations ≤ 20 mg mesna/mL fluid in D5W or 0.9% NaCl and to 
be administered intravenously as a continuous infusion. If patient is obese (BMI > 35), drug 
dosage will be calculated using practical weight as described in Appendix A. Toxicities include 
nausea, vomiting and diarrhea.  
11.2.2 Filgrastim (Granulocyte Colony-Stimulating Factor, G-CSF, Filgrastim, Neupogen) 
Filgrastim will be obtained commercially by the Clinical Center Pharmacy Department and is 
supplied in 300 µg/mL and 480 µg/1.6 mL vials. Filgrastim should be refrigerated and not 
allowed to freeze. The product bears the expiration date. The product should not be shaken. It is 


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generally stable for at least 10 months when refrigerated. The appropriate dose is drawn up into a 
syringe.  
Filgrastim will be given as a daily subcutaneous injection. The side effects of filgrastim are skin 
rash, myalgia and bone pain, an increase of preexisting inflammatory conditions, enlarged spleen 
with occasional associated low platelet counts, alopecia (with prolonged use) elevated blood 
chemistry levels. 
11.2.3 Trimethoprim and Sulfamethoxazole Double Strength (TMP/SMX DS) 
TMP/SMX DS will be obtained by the Clinical Center Pharmacy Department from commercial 
sources. It will be used for the prevention of PCP pneumonia. The oral dose is 1 tablet PO daily 
three times a week (MUST be on non-consecutive days) beginning on Day 0 or within one week 
of anticipated lymphopenia and continuing for at least 6 months and until the CD4 count is > 200 
on two consecutive lab studies. Like other sulfa drugs, TMP/SMX DS can cause allergies, fever, 
photosensitivity, nausea, and vomiting. Allergies typically develop as a widespread itchy red 
rash with fever 8-14 days after beginning the standard dose. Neutropenia, a reduction in the 
number of neutrophils, can also occur.  
Dapsone (in G6PD sufficient patient), atovaquone, or pentamidine will may be substituted for 
TMP/SMX-DS in patients with sulfa allergies. 
11.2.3.1 Dapsone 
Dapsone will be obtained by the Clinical Center Pharmacy Department from commercial 
sources. It will be used for the prevention of Pneumocystis pneumonia. The dose is 100 mg by 
mouth daily, starting on Day 0 (± 7 days) and continuing at least 6 months and until the CD4+ 
count is > 200 on two consecutive lab studies. It is supplied as 25 mg and 100 mg tablets. 
Dapsone contains a sulfa group, although the cross reactivity in patients with sulfa allergies is 
quite low. Dapsone may be considered in patients with mild to moderate sulfa allergies. Dapsone 
should be avoided in patients with severe (i.e., a history of anaphylaxis or other equally serious 
reaction) reactions to sulfa drugs. Additionally, dapsone has been reported to cause hemolytic 
anemia is patients with G6PD deficiency. It is recommended that patients be tested for G6PD 
deficiency prior to the initiation of dapsone therapy. Dapsone is generally well tolerated, but may 
cause a number of hematologic adverse reactions, including increased reticulocyte counts, 
hemolysis, decreased hemoglobin, methemoglobinemia, agranulocytosis, anemia, and 
leukopenia. Other rare but serious adverse reactions include bullous exfoliative dermatitis, 
Stevens-Johnson syndrome, toxic epidermal necrolysis, pancreatitis, interstitial pneumonitis, and 
pulmonary eosinophilia. For more detailed information about adverse reactions, consult the 
package insert. 
11.2.3.2 Atovaquone 
Atovaquone will be obtained by the Clinical Center Pharmacy Department from commercial 
sources. It will be used for the prevention of Pneumocystis pneumonia in patients who cannot 
tolerate or are allergic to sulfamethoxazole/trimethoprim, dapsone, or pentamidine. Atovaquone 
may be given as a single daily dose of 1500 mg orally or the dose may be split into 750 mg given 
orally twice daily. Atovaquone will be started on Day 0 (± 7 days), and will continue for at least 
6 months and until the CD4+ count is > 200 on two consecutive lab studies. Atovaquone is 
supplied as an oral suspension containing 150 mg/mL. Common adverse reactions to atovaquone 
include: headache, rash, diarrhea, nausea, vomiting, abdominal pain, cough, and fever. Rare but 


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serious adverse reactions include acute renal failure, hepatitis and hepatic failure, angioedema, 
pancreatitis, and Stevens-Johnson syndrome. For more detailed information about adverse 
reactions, consult the package insert. 
11.2.3.3 Aerosolized Pentamidine 
Patients with sulfa allergies will receive aerosolized pentamidine 300 mg per nebulizer within 
one week prior to admission and continued monthly until the CD4 count is above 200 on two 
consecutive follow up lab studies and for at least 6-months post chemotherapy. Pentamidine 
Isethionate will be obtained by the Clinical Center Pharmacy Department from commercial 
sources. It will be used to prevent the occurrence of PCP infections. It is supplied in 300 mg vials 
of lyophilized powder and will be administered via nebulizer. Toxicities reported with the use of 
Pentamidine include metallic taste, coughing, bronchospasm in heavy smokers and asthmatics; 
increased incidence of spontaneous pneumothorax in patients with previous PCP infection or 
pneumatoceles, or hypoglycemia. 
11.2.4  Herpes Simplex and Varicella Zoster Virus Prophylaxis 
11.2.4.1 Valacyclovir (Valtrex) 
Valacyclovir will be obtained by the Clinical Center Pharmacy Department from commercial 
sources. It will be used orally to prevent the occurrence of herpes virus infections in patients with 
positive HSV serology. It is supplied in 500 mg tablets. Valacyclovir will be started at a dose of 
500 mg orally daily if the patient is able to tolerate oral intake. See package insert for dosing 
adjustments in patients with renal impairment. Common side effects include headache, upset 
stomach, nausea, vomiting, diarrhea or constipation. Rare serious side effects include hemolytic 
uremic syndrome and thrombotic thrombocytopenic purpura.  
11.2.4.2 Acyclovir 
Acyclovir will be obtained by the Clinical Center Pharmacy Department from commercial 
sources. It will be used to prevent the occurrence of herpes virus infections in patients who 
cannot take oral medications. It is supplied as powder for injection in 500 mg/vials. Reconstitute 
in 10 mL of sterile water for injection to a concentration of 50 mg/mL. Reconstituted solutions 
should be used within 12 hours. IV solutions should be diluted to a concentration of 7 mg/mL or 
less and infused over 1 hour to avoid renal damage. Reversible renal insufficiency has been 
reported with IV but not oral acyclovir. Neurologic toxicity including delirium, tremors, coma, 
acute psychiatric disturbances, and abnormal EEGs have been reported with higher doses of 
acyclovir. Should this occur, a dosage adjustment will be made or the drug will be discontinued. 
Stomach upset, headache or nausea, rash or hives; peripheral edema; pain, elevated liver function 
tests; and leukopenia, diarrhea, lymphadenopathy, myalgias, visual abnormalities and elevated 
creatinine have been reported. Hair loss from prolonged use has been reported. Acyclovir will 
not be used concomitantly with other nucleoside analogs which interfere with DNA synthesis, 
e.g. ganciclovir. In renal disease, the dose is adjusted as per product labeling.  
11.2.5 Fluconazole 
Fluconazole will be obtained by the Clinical Center Pharmacy Department from commercial 
sources. It will be used to prophylax against fungal infections. It is available in 200 mg tablets. It 
can cause headache, nausea, vomiting, diarrhea or abdominal pain, and liver damage which may 
be irreversible. It can cause rashes and itching, which in rare cases has caused Stevens Johnson 


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Syndrome. It has several significant drug interactions. The package insert should be consulted 
prior to prescribing. For IV administration in patients who cannot tolerate the oral preparation, 
Fluconazole comes in 2 mg/mL solution for injection, and prepared according to Clinical Center 
Pharmacy standard procedures. It should be administered at a maximum IV rate of 200 mg/hour. 
11.2.6 Ondansetron Hydrochloride 
Ondansetron hydrochloride will be obtained by the Clinical Center Pharmacy Department from 
commercial sources. It will be used to control nausea and vomiting during the chemotherapy 
preparative regimen. It can cause headache, dizziness, myalgias, drowsiness, malaise, and 
weakness. Less common side effects include chest pain, hypotension, pruritis, constipation and 
urinary retention. Consult the package insert for specific dosing instructions. 
11.2.7 Furosemide 
Furosemide will be obtained by the Clinical Center Pharmacy Department from commercial 
sources. It will be used to enhance urine output during the chemotherapy preparative regimen 
with cyclophosphamide. Adverse effects include dizziness, vertigo, paresthesias, weakness, 
orthostatic hypotension, photosensitivity, rash and pruritis. Consult the package insert for a 
complete list of all side effects. 
 


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(c-erbB) in human brain tumors. Mol Cell Biol. 1988;8(4):1816-20. 
14. 
Chu CT, Everiss KD, Wikstrand CJ, Batra SK, Kung HJ, Bigner DD. Receptor 
dimerization is not a factor in the signalling activity of a transforming variant epidermal growth 
factor receptor (EGFRvIII). Biochem J. 1997;324 ( Pt 3):855-61. 
15. 
Batra SK, Castelino-Prabhu S, Wikstrand CJ, Zhu X, Humphrey PA, Friedman HS, et al. 
Epidermal growth factor ligand-independent, unregulated, cell-transforming potential of a 
naturally occurring human mutant EGFRvIII gene. Cell Growth Differ. 1995;6(10):1251-9. 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
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16. 
Lal A, Glazer CA, Martinson HM, Friedman HS, Archer GE, Sampson JH, et al. Mutant 
epidermal growth factor receptor up-regulates molecular effectors of tumor invasion. Cancer 
Res. 2002;62(12):3335-9. 
17. 
Boockvar JA, Kapitonov D, Kapoor G, Schouten J, Counelis GJ, Bogler O, et al. 
Constitutive EGFR signaling confers a motile phenotype to neural stem cells. Mol Cell Neurosci. 
2003;24(4):1116-30. 
18. 
Lammering G, Valerie K, Lin PS, Hewit TH, Schmidt-Ullrich RK. Radiation-induced 
activation of a common variant of EGFR confers enhanced radioresistance. Radiother Oncol. 
2004;72(3):267-73. 
19. 
Montgomery RB, Guzman J, O'Rourke DM, Stahl WL. Expression of oncogenic 
epidermal growth factor receptor family kinases induces paclitaxel resistance and alters beta-
tubulin isotype expression. J Biol Chem. 2000;275(23):17358-63. 
20. 
Nagane M, Coufal F, Lin H, Bogler O, Cavenee WK, Huang HJ. A common mutant 
epidermal growth factor receptor confers enhanced tumorigenicity on human glioblastoma cells 
by increasing proliferation and reducing apoptosis. Cancer Res. 1996;56(21):5079-86. 
21. 
Humphrey PA, Wong AJ, Vogelstein B, Zalutsky MR, Fuller GN, Archer GE, et al. Anti-
synthetic peptide antibody reacting at the fusion junction of deletion-mutant epidermal growth 
factor receptors in human glioblastoma. Proc Natl Acad Sci U S A. 1990;87(11):4207-11. 
22. 
Wong AJ, Ruppert JM, Bigner SH, Grzeschik CH, Humphrey PA, Bigner DS, et al. 
Structural alterations of the epidermal growth factor receptor gene in human gliomas. Proc Natl 
Acad Sci U S A. 1992;89(7):2965-9. 
23. 
Moscatello DK, Ramirez G, Wong AJ. A naturally occurring mutant human epidermal 
growth factor receptor as a target for peptide vaccine immunotherapy of tumors. Cancer Res. 
1997;57(8):1419-24. 
24. 
Heimberger AB, Crotty LE, Archer GE, Hess KR, Wikstrand CJ, Friedman AH, et al. 
Epidermal growth factor receptor VIII peptide vaccination is efficacious against established 
intracerebral tumors. Clin Cancer Res. 2003;9(11):4247-54. 
25. 
Ciesielski MJ, Kazim AL, Barth RF, Fenstermaker RA. Cellular antitumor immune 
response to a branched lysine multiple antigenic peptide containing epitopes of a common 
tumor-specific antigen in a rat glioma model. Cancer Immunol Immunother. 2005;54(2):107-19. 
26. 
Heimberger AB, Archer GE, Crotty LE, McLendon RE, Friedman AH, Friedman HS, et 
al. Dendritic cells pulsed with a tumor-specific peptide induce long-lasting immunity and are 
effective against murine intracerebral melanoma. Neurosurgery. 2002;50(1):158-64; discussion 
64-6. 
27. 
Wu AH, Xiao J, Anker L, Hall WA, Gregerson DS, Cavenee WK, et al. Identification of 
EGFRvIII-derived CTL epitopes restricted by HLA A0201 for dendritic cell based 
immunotherapy of gliomas. J Neurooncol. 2006;76(1):23-30. 
28. 
Sampson JH, Archer GE, Mitchell DA, Heimberger AB, Bigner DD. Tumor-specific 
immunotherapy targeting the EGFRvIII mutation in patients with malignant glioma. Semin 
Immunol. 2008;20(5):267-75. 


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29. 
Heimberger AB, Hussain SF, Aldape K, Sawaya R, Archer GA, Friedman H, et al. 
Tumor-specific peptide vaccination in newly-diagnosed patients with GBM. Journal of Clinical 
Oncology. 2006;24(18):107s-s. 
30. 
Bullain SS, Sahin A, Szentirmai O, Sanchez C, Lin N, Baratta E, et al. Genetically 
engineered T cells to target EGFRvIII expressing glioblastoma. J Neurooncol. 2009;94(3):373-
82. 
31. 
Ohno M, Natsume A, Ichiro Iwami K, Iwamizu H, Noritake K, Ito D, et al. Retrovirally 
engineered T-cell-based immunotherapy targeting type III variant epidermal growth factor 
receptor, a glioma-associated antigen. Cancer Sci. 2010;101(12):2518-24. 
32. 
Dudley ME, Yang JC, Sherry R, Hughes MS, Royal R, Kammula U, et al. Adoptive cell 
therapy for patients with metastatic melanoma: Evaluation of intensive myeloablative 
chemoradiation preparative regimens. J Clin Oncol. 2008;26:5233-9. 
33. 
Kawakami Y, Eliyahu S, Delgado CH, Robbins PF, Rivoltini L, Topalian SL, et al. 
Cloning of the gene coding for a shared human melanoma antigen recognized by autologous T 
cells infiltrating into tumor. Proc Natl Acad Sci USA. 1994;91:3515-9. 
34. 
Kawakami Y, Eliyahu S, Delgado CH, Robbins PF, Sakaguchi K, Appella E, et al. 
Identification of a human melanoma antigen recognized by tumor infiltrating lymphocytes 
associated with in vivo tumor rejection. Proc Natl Acad Sci USA. 1994;91:6458-62. 
35. 
Kawakami Y, Eliyahu S, Sakaguchi K, Robbins PF, Rivoltini L, Yannelli JR, et al. 
Identification of the immunodominant peptides of the MART-1 human melanoma antigen 
recognized by the majority of HLA-A2-restricted tumor infiltrating lymphocytes. J Exp Med. 
1994;180(1):347-52. 
36. 
Morgan RA, Dudley ME, Wunderlich JR, Hughes MS, Yang JC, Sherry RM, et al. 
Cancer regression in patients after transfer of genetically engineered lymphocytes. Science. 
2006;314(5796):126-9. 
37. 
Johnson LA, Morgan RA, Dudley ME, Cassard L, Yang JC, Hughes MS, et al. Gene 
therapy with human and mouse T-cell receptors mediates cancer regression and targets normal 
tissues expressing cognate antigen. Blood. 2009;114:535-46. 
38. 
Kochenderfer JN, Wilson WH, Janik JE, Dudley ME, Stetler-Stevenson M, Feldman SA, 
et al. Eradication of B-lineage cells and regression of lymphoma in a patient treated with 
autologous T cells genetically engineered to recognize CD19. Blood. 2010;116:4099-102. 
39. 
Morgan RA, Yang JC, Kitano M, Dudley ME, Laurencot CM, Rosenberg SA. Case 
report of a serious adverse event following the administration of T cells transduced with a 
chimeric antigen receptor recognizing ERBB2. Mol Ther. 2010;18(4):843-51. 
40. 
Pule MA, Savoldo B, Myers GD, Rossig C, Russell HV, Dotti G, et al. Virus-specific T 
cells engineered to coexpress tumor-specific receptors: persistence and antitumor activity in 
individuals with neuroblastoma. Nat Med. 2008;14(11):1264-70. 
41. 
Wen PY, Macdonald DR, Reardon DA, Cloughesy TF, Sorensen AG, Galanis E, et al. 
Updated response assessment criteria for high-grade gliomas: response assessment in neuro-
oncology working group. J Clin Oncol. 2010;28(11):1963-72. 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
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71
42. 
Weitzner MA, Meyers CA, Gelke CK, Byrne KS, Cella DF, Levin VA. The Functional 
Assessment of Cancer Therapy (FACT) scale. Development of a brain subscale and revalidation 
of the general version (FACT-G) in patients with primary brain tumors. Cancer. 
1995;75(5):1151-61. 
43. 
Ballman KV, Buckner JC, Brown PD, Giannini C, Flynn PJ, LaPlant BR, et al. The 
relationship between six-month progression-free survival and 12-month overall survival end 
points for phase II trials in patients with glioblastoma multiforme. Neuro-oncology. 
2007;9(1):29-38. 
 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
72
13 APPENDICES  
13.1 APPENDIX A: MODIFICATION OF DOSE CALCULATIONS* IN PATIENTS WHOSE BMI IS 
GREATER THAN 35 
 
Unless otherwise specified in this protocol, actual body weight is used for dose calculations of 
treatment agents. In patients who are determined to be obese (BMI > 35), the practical weight 
(see 3 below) will be used. 
 
1. BMI Determination: 
BMI = weight (kg) / [height (m)]2 
 
2. Calculation of ideal body weight: 
 
Male = 50 kg + 2.3 (number of inches over 60 inches) 
 
 
Example: Ideal body weight of 5’10” male 
 
 
 
50 + 2.3 (10) = 73 kg 
 
 
Female = 45.5 kg + 2.3 (number of inches over 60 inches) 
 
 
Example: Ideal body weight of 5’3” female 
 
 
 
45.5 + 2.3 (3) = 57 kg 
3. Calculation of “practical weight”: 
 
 
Calculate the average of the actual and the ideal body weights. This is the practical 
weight to be used in calculating the doses of chemotherapy and associated agents 
designated in the protocol. 
 
*Practical weight will NOT be used in the calculation of dose for aldesleukin. 
 
 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
73
13.2 APPENDIX B: ADVERSE EVENTS OCCURRING IN ≥10% OF PATIENTS TREATED WITH 
ALDESLEUKIN (N=525)1 
 
Body System  
% Patients  
 
Body System  
 % Patients 
Body as a Whole  
 
 
 
Metabolic and Nutritional Disorders 
Chills   
 
 
52  
 
Bilirubinemia   
 
40 
Fever   
 
 
29  
 
Creatinine increase  
 
33 
Malaise  
 
 
27  
 
Peripheral edema  
 
28 
Asthenia  
 
 
23 
 
SGOT increase  
 
23 
Infection  
 
 
13  
 
Weight gain   
 
16 
Pain   
 
 
12  
 
Edema  
 
 
15 
Abdominal pain  
 
11  
 
Acidosis  
 
 
12 
Abdomen enlarged  
 
10  
 
Hypomagnesemia  
 
12 
Cardiovascular  
 
 
 
Hypocalcemia  
 
11 
Hypotension   
 
71  
 
Alkaline phosphatase incr 
10 
Tachycardia   
 
23 
 
Nervous 
Vasodilation   
 
13  
 
Confusion  
 
 
34 
Supraventricular tachycardia  12  
 
Somnolence   
 
22 
Cardiovascular disordera  
11  
 
Anxiety  
 
 
12 
Arrhythmia  
 
 
10  
 
Dizziness  
 
 
11 
Digestive  
 
 
 
 
Respiratory 
Diarrhea  
 
 
67  
 
Dyspnea  
 
 
43 
Vomiting  
 
 
50  
 
Lung disorderb  
 
24 
Nausea  
 
 
35  
 
Respiratory disorderc   
11 
Stomatitis  
 
 
22  
 
Cough increase  
 
11 
Anorexia  
 
 
20  
 
Rhinitis  
 
 
10 
Nausea and vomiting   
19  
 
Skin and Appendages 
Hemic and Lymphatic  
 
 
Rash   
 
 
42 
Thrombocytopenia  
 
37 
 
Pruritus  
 
 
24 
Anemia  
 
 
29  
 
Exfoliative dermatitis  
18 
Leukopenia 
 
 
16  
 
Urogenital 
Oliguria  
 
 
63 
Legend: 
a. Cardiovascular disorder: fluctuations in blood pressure, asymptomatic ECG changes, CHF. 
b. Lung disorder: physical findings associated with pulmonary congestion, rales, rhonchi. 
c. Respiratory disorder: ARDS, CXR infiltrates, unspecified pulmonary changes. 
1Source: Proleukin® Prescribing Information – June 2007 
 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
74
13.3 APPENDIX C: EXPECTED IL-2 TOXICITIES AND THEIR MANAGEMENT 
Expected Toxicity 
Expected 
Grade 
Supportive 
Measures 
Stop Cycle* 
Stop Treatment 
** 
Chills 
3 
IV Meperidine 25-
50 mg, IV q1h, prn 
No 
No 
Fever 
3 
Acetaminophen 650 
mg, po, q4h; 
Indomethicin 50-75 
mg, po, q8h 
No 
No 
Pruritis 
3 
Hydroxyzine HCL 
10-20 mg po q6h, 
prn; 
Diphenhydramine 
HCL25-50 mg, po, 
q4h, prn 
No 
No 
Nausea/Vomiting/ 
Anorexia 
3 
Ondansetron 10 mg, 
IV, q8h, prn; 
Granisetron 0.01 
mg/kg IV daily prn; 
Droperidol 1 mg, IV 
q4-6h, prn; 
Prochlorperazine 25 
mg q4h PO, prn or 
10 mg IV q6h prn 
No 
No 
Diarrhea 
3 
Loperamide 2mg, 
po, q3h, prn; 
Diphenoxylate HCl 
2.5 mg and atropine 
sulfate 25 mcg, po, 
q3h, prn; codeine 
sulfate 30-60 mg, 
po, q4h, prn 
If uncontrolled 
after 24 hours 
despite all 
supportive 
measures 
No 
Malaise 
3 or 4 
Bedrest interspersed 
with activity 
If other 
toxicities occur 
simultaneously 
No 
Hyperbilirubinemia 
3 or 4 
Observation 
If other 
toxicities occur 
simultaneously 
No 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
75
Anemia 
3 or 4 
Transfusion with 
PRBCs 
If uncontrolled 
despite all 
supportive 
measures 
No 
Thrombocytopenia 
3 or 4 
Transfusion with 
platelets 
If uncontrolled 
despite all 
supportive 
measures 
No 
Edema/Weight gain 
3 
Diuretics prn 
No 
No 
Hypotension 
3 
Fluid resuscitation 
Vasopressor support 
If uncontrolled 
despite all 
supportive 
measures 
No 
Dyspnea 
3 or 4 
Oxygen or 
ventilatory support 
If requires 
ventilatory 
support 
No 
Oliguria 
3 or 4 
Fluid boluses or 
dopamine at renal 
doses 
If uncontrolled 
despite all 
supportive 
measures 
No 
Increased creatinine 
3 or 4 
Observation 
Yes (grade 4) 
No 
Renal failure 
3 or 4 
Dialysis 
Yes 
Yes 
Pleural effusion 
3 
Thoracentesis 
If uncontrolled 
despite all 
supportive 
measures 
No 
Bowel perforation 
3 
Surgical intervention Yes 
Yes 
Confusion 
3 
Observation 
Yes 
No 
Somnolence 
3 or 4 
Intubation for 
airway protection 
Yes 
Yes 
Arrhythmia 
3 
Correction of fluid 
and electrolyte 
imbalances; 
chemical conversion 
If uncontrolled 
despite all 
supportive 
measures 
No 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
76
or electrical 
conversion therapy 
Elevated troponin 
levels 
3 or 4 
Observation 
Yes 
If changes in LV 
function have not 
improved to 
baseline by next 
dose 
Myocardial 
infarction 
4 
Supportive care 
Yes 
Yes 
Elevated 
transaminases 
3 or 4 
Observation 
For grade 4 
without liver 
metastases 
If changes have 
not improved to 
baseline by next 
dose 
Hyperbilirubinemia 
3 or 4 
Observation 
For grade 4 
without liver 
metastases 
If changes have 
not improved to 
baseline by next 
dose 
Electrolyte 
imbalances 
3 or 4 
Electrolyte 
replacement 
If uncontrolled 
despite all 
supportive 
measures 
No 
Neutropenia 
4 
Observation 
No 
No 
*Unless the toxicity is not reversed within 12 hours. 
** Unless the toxicity is not reversed to grade 2 or less by next treatment. 
 
 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
77
13.4 APPENDIX D: INTERLEUKIN-2 TOXICITIES OBSERVED IN PATIENTS TREATED AT THE 
NIH CLINICAL CENTER  


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
78
13.5 APPENDIX E: CERTIFICATE OF ANALYSIS – ANTI-EGFRVIII CAR TRANSDUCED T-
CELLS 
Date of preparation of final product: 
Patient: 
Tests performed on final product: 
Test 
Method 
Limits 
Result 
Tests 
Performed by 
Initials
/Date 
Cell viability1 
trypan blue exclusion 
>70% 
 
 
 
Total viable cell 
number1 
visual microscopic count 
>1 x107 
 
 
 
Tumor reactivity2 
γ-IFN release vs. positive 
control 
 
γ-IFN release vs. negative 
control including wt-
EGFRvIII 
>200 pg/mL, and > 2 
times background 
 
<400 pg/mL 
 
 
 
CAR expression2 
FACS analysis of the 
transduced cells 
PBL, >10% 
 
 
 
Microbiological 
studies  
gram stain1,3,  
no micro-organisms 
seen 
 
 
 
 
aerobic culture3,4 
no growth 
 
 
 
 
fungal culture3,4 
no growth 
 
 
 
 
anaerobic culture3,4 
no growth 
 
 
 
 
mycoplasma test5 
negative 
 
 
 
Endotoxin  
limulus assay1 
≤ 5 E.U./kg 
 
 
 
RCR 
S+L- Assay4 
RCR-PCR6 
 
negative 
 
 
 
1 Performed on sample of the final product immediately prior to infusion. Results are available at the time of infusion. 
2 Performed 2-10 post transduction. Results are available at the time of infusion.  
3 Performed 2-4 days prior to infusion. Results are available at the time of infusion but may not be definitive. 
4 Sample collected from the final product prior to infusion. Results will not be available before cells are infused into the patient. 
5 Performed 2-10 days prior to infusion. Results are available at the time of infusion. 
6 Performed on sample approximately 1-4 days prior to infusion. Results are available at the time of infusion.  
Prepared by: _______________________________________Date: ____________________ 
QC sign-off: _______________________________________Date: ____________________ 
Qualified Laboratory or Clinical Supervisor 
 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
79
13.6 APPENDIX F: KARNOFSKY PERFORMANCE SCORE (KPS) 
 
KPS  
Description  
100 
Normal, no complaints, no signs of disease 
90 
Capable of normal activity, few symptoms or signs of disease 
80 
Normal activity with some difficulty, some symptoms or signs 
70 
Caring for self, not capable of normal activity or work 
60 
Requiring some help, can take care of most personal requirements 
50 
Requires help often, requires frequent medical care 
40 
Disabled, requires special care and help 
30 
Severely disabled, hospital admission indicated but no risk of death 
20 
Very ill, urgently requiring admission, requires supportive measures or treatment 
10 
Moribund, rapidly progressive fatal disease processes 
0 
Death 
 
 


Abbreviated Title: Anti-EGFRvIII CAR PBL 
Version Date: June 28, 2018 
 
80
13.7 APPENDIX G: ANTI-EPILEPTIC MEDICATIONS 
 
Non-EIAEDs 
 
EIAEDs 
Valproic acid 
 
Carbamazapine 
Gabapentin 
 
Oxcarbazepine* 
Lamotrigine 
 
Phenytoin 
Topriamate 
 
Fosphenytoin 
Levetiracetam 
 
Phenobarbital 
Tiagabine 
 
Primidone 
Zonisamide 
 
 
Clonazepam 
 
 
 
* enzyme induction limited and less potent than carbamazapine 
 
 
 
 

ICF_001.pdf: 
MEDICAL RECORD 
CONSENT TO PARTICIPATE IN A CLINICAL RESEARCH STUDY 
• 
Adult Patient or 
• Parent, for Minor Patient 
INSTITUTE: 
National Cancer Institute 
STUDY NUMBER: 
11-C-0266 
PRINCIPAL INVESTIGATOR: 
Steven A. Rosenberg, M.D., Ph.D 
 
STUDY TITLE:  
 
A Phase I/II Study of the Safety and Feasibility of Administering T-Cells Expressing Anti-
EGFRvIII Chimeric Antigen Receptor to Patients with Malignant Gliomas Expressing 
EGFRvIII 
Continuing Review Approved by the IRB on 07/09/18 
Amendment Approved by the IRB on 07/10/18 (Q) 
Date posted to web:  08/03/18 
Standard 
PATIENT IDENTIFICATION 
CONSENT TO PARTICIPATE IN A CLINICAL RESEARCH STUDY 
• 
Adult Patient or 
• Parent, for Minor Patient 
NIH-2514-1 (07-09) 
P.A.: 09-25-0099 
File in Section 4: Protocol Consent (1) 
 
 
INTRODUCTION 
 
We invite you to take part in a research study at the National Institutes of Health (NIH). 
First, we want you to know that: 
 
Taking part in NIH research is entirely voluntary. 
You may choose not to take part, or you may withdraw from the study at any time. In either 
case, you will not lose any benefits to which you are otherwise entitled. However, to receive 
care at the NIH, you must be taking part in a study or be under evaluation for study 
participation. 
You may receive no benefit from taking part. The research may give us knowledge that 
may help people in the future. 
Second, some people have personal, religious or ethical beliefs that may limit the kinds of medical 
or research treatments they would want to receive (such as blood transfusions). If you have such 
beliefs, please discuss them with your NIH doctors or research team before you agree to the study. 
Now we will describe this research study. Before you decide to take part, please take as much time 
as you need to ask any questions and discuss this study with anyone at NIH, or with family, friends 
or your personal physician or other health professional. 
 
 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 11-C-0266 
CONTINUATION: page 2 of 17 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.: 09-25-0099 
File in Section 4: Protocol Consent 
 
Why is this study being done? 
You have been diagnosed with malignant glioma and the standard treatments available have not 
been effective. We have developed an experimental procedure for treating patients with cancer 
that uses cells found in their blood. We genetically modify these cells and grow them in the 
laboratory. We will be using the anti-EGFRvIII gene and a type of virus (retrovirus) in making 
these special cells (anti- EGFRvIII cells). The anti-EGFRvIII cells target a form of the Epidermal 
Growth Factor Receptor (EGFR) which is only present in gliomas, not in normal tissue in your 
body. By blocking EGFRvIII, we hope that these cells when infused will decrease the size of your 
tumors. However, it is possible that these cells will not have this effect. 
The anti-EGFRvIII cells will be given to you as an intravenous (IV) infusion. This type of 
experimental regimen is called “gene transfer” and is very closely monitored by the Food and Drug 
Administration (FDA) and other regulatory agencies. The risks of gene transfer will be described 
later in this document. 
Why are you being asked to take part in this study? 
You are being asked to participate in this study because you have been diagnosed with cancer that 
has the EGFRvIII molecule on the surface of the tumor. 
How many people will take part in this study? 
Up to 107 patients may participate in this study. 
Description of Research Study  
This study has several stages outlined below: 
• Stage 1 is performed under the screening protocol, 99-C-0128 (Evaluation for NCI Surgery 
Branch Clinical Research Protocols), to which you have already enrolled. 
• Stage 2 is performed under the cell harvest protocol, 03-C-0277 (Cell Harvest and 
Preparation for Surgery Branch Adoptive Cell Therapy Protocols), to which you have 
already enrolled. 
• Stages 3-6 are performed under this protocol. 
Stage 
Timeframe 
Location 
Comments & Instructions 
1. Work-up 
1-2 weeks 
Inpatient and/or 
outpatient  
Scans, x-rays, labs and other tests 
as needed 
2. Cell 
Manufacturing 4-6 weeks 
Inpatient and/or 
outpatient 
Leukapheresis 
3. Chemotherapy 
 
1 week 
Inpatient 
Receive IV chemotherapy to 
prepare your immune system for 
the cells. 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 11-C-0266 
CONTINUATION: page 3 of 17 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.: 09-25-0099 
File in Section 4: Protocol Consent 
 
4. Cells and 
aldesleukin 
(IL-2) 
1-5 days 
Inpatient and 
possibly ICU 
Receive anti-EGFRvIII cells IV, 
and then aldesleukin (IL-2) about 
every 8 hours for up to 15 doses. 
5. Recovery 
1-3 weeks 
Inpatient unit 
Recover from the effects of 
treatment. 
6. Follow-up 
Ongoing 
until disease 
progression 
Outpatient 
Return to clinic for physical exam, 
review of side effects, labs, scans 
approximately 4 weeks following 
treatment, and then monthly for 12 
months and then every 1-2 months. 
The first few patients enrolled will participate in the Phase I portion of the study, called the “dose 
escalation” phase. The purpose of dose escalation is to determine the most effective yet safe dose 
of anti-EGFRvIII cells. There will be 13 dose levels of anti-EGFRvIII cells. The first patients 
enrolled get the smallest dose and the dose is increased when a level has been determined to be 
safe.  
Your study doctor will discuss with you which dose of anti-EGFRvIII cells you will be receiving.  
What will happen if you take part in this research study?  
Before you Begin the Study 
The following procedures are conducted under protocols 99-C-0128 or 03-C-0277. 
Cell Harvest and Growth 
You underwent a process called “apheresis”, while enrolled on our companion protocol 03-C-
0277. This process obtained certain types of blood cells from you. Some of these cells will be 
grown in the lab and genetically modified to recognize a protein on your tumor cells. If your cells 
do not grow, unfortunately you will not be able to receive the cell infusion. If that happens, we 
will look at alternative experimental treatments at the NIH Clinical Center or refer you to the care 
of your referring physician. We usually know after about 4 weeks whether the cells will grow well 
enough to be used as an experimental treatment on this protocol. At the time we determine that 
your cells are not growing, we will inform you and discuss your options with you. Several 
medications are used during the preparation of your cell product, be sure to tell your doctor 
if you are allergic to any antibiotics. 
Work-up 
Prior to receiving the experimental regimen, you will undergo many tests. We will evaluate you 
for eligibility for participation on this trial with a physical examination, CT and/or MRI scans, x-
rays, EKG, heart and lung function tests, and blood tests. Patients who have a positive HIV blood 
test will not be eligible for this protocol because it may put them at higher risk of developing 
infections. If you are a woman, you will undergo a pregnancy test. You may be admitted to the 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 11-C-0266 
CONTINUATION: page 4 of 17 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.: 09-25-0099 
File in Section 4: Protocol Consent 
 
hospital for these tests. However, you will be allowed to leave on pass on the days that you are not 
having tests performed. 
Catheter Insertion 
Prior to beginning the experimental regimen, you will have an intravenous (IV) catheter placed in 
your upper chest. The area will be numbed with an anesthetic before the catheter is put in. Although 
rare, putting these catheters in can sometimes cause collapse of a lung or cause bleeding. Lung 
collapse is treated by putting a tube into your chest for a few days to allow your lung to expand. 
Pressure is placed on any area that might bleed. Other IVs may be needed in one or both of your 
arms if we to give you extra fluids, medicines, or nutrition. 
Leukapheresis 
Leukapheresis, a specific type of “apheresis”, is a procedure that allows us to remove certain types 
of blood cells from you and return the rest of your blood. It is a very common procedure that is 
done routinely here at the NIH with very few risks. During leukapheresis, blood is removed from 
you through a needle in your arm, circulated through a machine that divides whole blood into red 
cells, plasma (the liquid component of blood), and lymphocytes (or white cells), and then the 
plasma and red cells are returned to you through a second needle in your other arm. The white 
blood cells collected before treatment may be used to help grow the cells. In addition to the 
leukapheresis you will undergo as part of your work-up, we will also ask you to undergo one 
additional apheresis procedure between 4 and 6 weeks after you receive the cell infusion to see the 
impact of this therapy on the immune system and see if cells we gave you are still active. 
The leukapheresis procedure takes between 4-5 hours to complete. Rarely, people may experience 
lightheadedness or dizziness. We ask that you eat prior to the procedure to prevent this. 
During the Study 
Chemotherapy Regimen (Day -7 through Day -3) 
After we have grown the anti-EGFRvIII cells to large numbers in the laboratory, you will be 
admitted to the hospital to begin your experimental regimen. You will be given two chemotherapy 
medicines, cyclophosphamide and fludarabine, to suppress your immune system so the anti-
EGFRvIII cells can work without any interference from the cells in your immune system. (These 
medicines will not treat your cancer. They may cause your tumor to shrink some, but this shrinkage 
is anticipated to be only partial and only for a short time.) The main purpose of the chemotherapy 
is to see if we can make the cells more effective in fighting cancer tumors. Animal experiments 
have indicated that this can make the cells more effective in fighting cancer tumors, and we think 
this is true in humans. You will receive the cyclophosphamide through your catheter over 1 hour 
for two days (Day -7 and Day -6) and the fludarabine will be given through your catheter for 30 
minutes every day for five days (Day -7 through Day -3). The side effects of these medicines are 
described on the following pages. 


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STUDY NUMBER: 11-C-0266 
CONTINUATION: page 5 of 17 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
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P.A.: 09-25-0099 
File in Section 4: Protocol Consent 
 
Cell Infusion and Aldesleukin (IL-2) Regimen (Day 0 through Day 5) 
Two to four days after the last dose of chemotherapy, you will be given the anti-EGFRvIII cells. 
The anti-EGFRvIII cells will be given through your catheter over 20-30 minutes. Within 24 hours 
after the anti-EGFRvIII cell infusion, you will be given low-dose or high-dose aldesleukin through 
your catheter. Patients enrolled in Dose Level 9D-1, 9D-2, or 10 will receive high-dose 
aldesleukin. Prior dose levels have identified this dose of cells is safe to administer with low-dose 
aldesleukin and we will now be testing whether high-dose IL-2 can be administered after the gene 
engineered cells (CAR T-cells). Aldesleukin is approved by the FDA for treatment of metastatic 
melanoma and metastatic renal cell cancer. The purpose of giving the aldesleukin with this therapy 
is to keep the cells we give you active for as long as possible so they will fight your tumor. 
Aldesleukin will be given as a 15-minute infusion about every 8 hours for up to five days after the 
cell infusion (maximum number of doses is 15). Doses may be skipped or delayed depending on 
how well you tolerate the doses. The risks of the cells and aldesleukin are described on the 
following pages. 
The day after you receive the anti-EGFRvIII cells, we may give you G-CSF (filgrastim) as a shot 
or injection under the skin every day to stimulate your blood cells until they increase to a sufficient 
number. This will continue until your white blood cell counts begin to return to normal. 
We will watch you closely during this entire time for any side effects of this experimental regimen. 
We will discuss the side effects below and we will include in your care all the medicines and 
treatments to prevent as many of these side effects and to make you as comfortable as we can.  
When you are Finished with Treatment 
Recovery 
You will recover in the hospital until you are well enough to go home. This usually takes 7 to 21 
days after you have received the cells or your last dose of aldesleukin; however, you may need to 
stay in the hospital for longer than this before you are well enough to go home. We will continue 
to give you support medications, do laboratory tests, and watch you closely for any side effects 
until we feel your condition is stable.  
In addition to the laboratory tests to monitor your condition, we will remove approximately 1-9 
teaspoons of blood three times per week to study the effects of this regimen on your immune 
system. The maximum amount of blood for research is approximately 2.3 cups in 8 weeks. 
Follow-up and Evaluation of Experimental Regimen  
You will need to continue to take Bactrim, an antibiotic, for at least 6 months following your 
treatment to prevent you from catching a certain type of pneumonia seen in patients who have low 
white blood cell counts. You may also need to take Valtrex, an anti-viral, for at least 6 months 
following your treatment to prevent any type of herpes simplex virus, like shingles.  


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STUDY NUMBER: 11-C-0266 
CONTINUATION: page 6 of 17 pages 
 
PATIENT IDENTIFICATION 
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P.A.: 09-25-0099 
File in Section 4: Protocol Consent 
 
After you are discharged, we will ask you to return to the NIH Clinical Center approximately 4 
weeks following treatment, and then if you are responding to the treatment, every month for one 
year and then every 1-2 months after one year. 
The first follow-up visit will probably take 2 days; the following visits may only take one day. At 
each visit you will have lab tests, imaging studies, and a physical examination. At some of your 
follow-up visits, you may undergo leukapheresis or have about 8 tubes of blood drawn (4 
tablespoons) so that we can see the effect this therapy has had on your immune system and if the 
cells we gave you are still alive. If you are unwilling or unable to travel to the NIH Clinical Center, 
we will contact you by phone or email and we may ask you to send us lab, imaging, and physical 
exam reports. If your tumor appears to be growing, we will look for other investigational therapies 
you may be eligible for, or refer you back to the care of your local physician.  
Birth Control 
If you are a woman who is breast feeding or pregnant, you may not participate in the study because 
we do not know how this medicine would affect your unborn child or your baby. If you are a 
woman who can become pregnant, or are the partner of a woman who can become pregnant, you 
will need to practice an effective form of birth control before starting study treatment, during study 
treatment, and for four months after you finish study treatment. Males should also refrain from 
sperm donation during this time-period. If you think that you or your partner is pregnant, you 
should tell your study doctor or nurse as soon as possible.  
Effective forms of birth control include:  
• Abstinence 
• Intrauterine device (IUD) 
• Hormonal [birth control pills, injections, or implants] 
• Tubal ligation 
• Vasectomy 
Gene Therapy Long-Term Follow-up (Retroviral Vectors)  
Because we do not know the long-term side effects of gene therapy, we will collect your blood 
over the next several years, frequently at first and then less frequently. If you return to your 
referring physician after treatment here, we will ask you to have your physician send your blood 
specimens here for this testing. This testing will determine if the cells have grown or changed in 
your body. We will test your blood immediately after you receive the cells, and then at 3, 6, and 
12 months (2 teaspoons each time) post-cell administration. If all of the tests are normal and show 
no change, we will collect blood from you every year after that to store in case you develop 
symptoms later.  
According to FDA requirements, we need you to return annually to the NIH for a physical 
examination for five years after you receive the cells. After that time, we will be sending you a 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
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STUDY NUMBER: 11-C-0266 
CONTINUATION: page 7 of 17 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
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NIH-2514-2 (10-84) 
P.A.: 09-25-0099 
File in Section 4: Protocol Consent 
 
questionnaire to get information regarding your health for the next ten years, for a total follow-up 
time-period of 15 years. For this reason, we ask that you continue to provide us with a current 
address and telephone number, even after you complete this research study. At the time of your 
death, no matter the cause, we may request permission for an autopsy in order to obtain vital 
information concerning the safety of this experimental treatment approach. Please discuss this with 
your family to inform them of this request. These long-term follow-up evaluations will be 
conducted under our companion protocol 09-C-0161, “Follow up Protocol for Subjects Previously 
Enrolled in NCI Surgery Branch Studies”, to which you will be enrolled following treatment. 
Risks or Discomforts of Participation 
What side effects or risks can I expect from being in this study?  
The risks and discomforts of this research study can be significant. This experimental treatment 
can lead to long-term decrease in your immune function. It is also possible that you may lose your 
fertility following this experimental treatment. It is possible, although unlikely, that this 
experimental treatment may cause your death.  
As of December 2014, we have treated approximately 18 patients since the study opened. Out of 
the 18 patients enrolled on this study, we have encountered one treatment-related death. This 
patient’s cells were much more “active” than other patients’ cells (producing a large amount of a 
substance that probably helps fight the tumor cells but also causes toxicities). We have revised the 
process so that future cells will not be as “active” when they are given back to the patients. 
The major side effects of this experimental regimen (described in detail on the following 
pages) that are most severe include:  
• Infection and low blood counts caused by the chemotherapy  
• Confusion and changes in mental status caused by the IL-2 
• Fluid retention, low blood pressure, and high heart rate caused by the IL-2 
We will discuss the side effects of this experimental regimen with you. You will be given 
medicines, transfusions, and treatments to prevent or treat the side effects including drugs to 
prevent and/or treat different types of infections. We will try to make you as comfortable as 
possible. You should talk to your study doctor about any symptoms that you experience while 
taking part in the study. 
As this is a new experimental therapy, you may experience side effects that we do not expect that 
may cause your condition to worsen. Any new information that becomes available during the 
course of this study will be shared with you. 
Anti-EGFRvIII Cell Infusion (Gene Transfer)  
The cells we will be giving you have a type of virus (retrovirus) put into them along with the anti- 
EGFRvIII protein. Although this retrovirus is not active, there is the rare possibility that it may 
cause infection. The cells could also cause you to develop another type of cancer, such as leukemia 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
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NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 11-C-0266 
CONTINUATION: page 8 of 17 pages 
 
PATIENT IDENTIFICATION 
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File in Section 4: Protocol Consent 
 
or lymphoma. We do not have much information about the specific side effects of the anti- 
EGFRvIII cells since this is our first study using these cells. We have studied similar types of cells 
in patients with cancer in other studies, and have seen the following side effects: 
• Fever, chills and shortness of breath, which may last for a few hours (common) 
• Lung congestion  
• Immune-type reaction  
o In similar clinical trials with cells targeting a melanoma protein, we have observed 
the following immune-mediated toxicities: loss of skin pigment (known as vitiligo), 
inflammation of the eye (uveitis), hearing loss, and dizziness. The skin, the eye, and 
the ear are all sites where that targeted melanoma protein is known to exist. 
o In another clinical trial with a similar type of cell, a patient died due to an immune 
type reaction from the cells.  
o The anti-EGFRvIII cells may cause an immune type of reaction in your brain which 
may result in seizures, swelling, and bleeding. 
You will be treated on this gene transfer protocol with a viral vector that was manufactured at the 
NCI Surgery Branch Vector Production Facility before May 2016. An internal review of the 
facility that made the vector for this protocol determined that the facility needed to be closed due 
to manufacturing issues. We know of no additional risks related to the previously produced vector 
for patients who have received cells with vectors made in this facility as the vectors were 
extensively tested by outside experts. Therefore, the IRB has determined that the potential benefit 
to you outweighs the potential risks. 
Aldesleukin (IL-2)  
When IL-2 is given through an intravenous catheter, it can make you feel like you have the flu. It 
can also cause confusion and mental status changes making you unable to make sound decisions. 
Prior to beginning treatment, we will ask you to complete a Durable Power of Attorney so that a 
person of your choosing can make health care decisions for you in case you develop these side 
effects. In our experience giving IL-2 to over 2,000 patients, we have found that these side effects 
go away within a few days of stopping the IL-2. 
Medications 
The side effects of cyclophosphamide, fludarabine, IL-2 and some of the other medications you 
will receive are listed below.  
Cyclophosphamide and Fludarabine Side Effects 
Common 
Less Common 
Rare 
 
Changes in blood counts 
including: low red cell 
 
Bleeding 
 
Heart damage 


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CONTINUATION: page 9 of 17 pages 
 
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count (causing fatigue and 
shortness of breath), low 
platelet count (increasing 
the risk of bleeding and 
bruising), decrease in 
white blood cells 
(increasing the risk of 
infection and the need for 
treatment with antibiotics 
or other treatment)  
 
Loss of appetite, nausea, 
vomiting  
 
Diarrhea, stomach pain 
 
Mouth sores 
 
Hair loss 
 
Fatigue 
 
Muscle or joint aches  
 
Infection 
 
Bladder irritation with 
bloody urine 
 
Severe allergic reaction 
(difficulty 
breathing/swelling) 
 
Headache or dizziness 
 
Sweating 
 
Swelling of arms or legs 
 
Skin changes, rash, 
blisters  
 
Weakness 
 
Hearing loss 
 
Lung damage 
 
Kidney damage 
 
Inflammation of the eye 
resulting in blindness 
 
Inflammation of nervous 
system resulting in death 
 
Epstein Barr Virus 
Lymphoma. This can be 
fatal. (Two patients on 
other studies in the 
Surgery Branch 
developed EBV 
lymphoma, and both died 
as a result of this disease.) 
 
Loss of fertility 
 
Complications resulting 
from suppression of 
immune function, which 
can result in a severe 
infection and can be fatal. 
Low-Dose Aldesleukin (IL-2) Side Effects 
Common 
Unlikely 
 
Local swelling at site of injection 
 
Low blood pressure 
 
Tiredness 
 
Swelling in hands and feet 
 
Rash 
 
Nausea 
 
Vomiting 
 
Diarrhea 
 
Infection 
 
Laboratory changes including kidney 
blood tests 
 
Decreased urine 
 
Changes in consciousness 
 
Infections 
High-Dose Aldesleukin (IL-2) Side Effects 
Common 
Less Common  
Rare 


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CONTINUATION SHEET for either: 
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PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
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P.A.: 09-25-0099 
File in Section 4: Protocol Consent 
 
 
Fever, chills, and fatigue  
 
Lowered platelet and red 
blood cell levels that may 
require transfusions  
 
Significant fluid retention 
causing weight gain (as 
much as 20 pounds)  
 
Low blood pressure 
 
Increased heart rate 
 
Shortness of breath 
 
Low urine output 
 
Swelling in your 
extremities  
 
Fluid in your lungs that 
can require oxygen  
 
Dry mouth, nausea, 
vomiting, diarrhea 
 
Rash, itching 
 
Changes in skin or hair 
pigmentation, called 
vitiligo  
 
Changes in mental status, 
including confusion, 
difficulty sleeping or 
vivid dreams; this can be 
severe and require 
sedation and monitoring 
in the ICU. 
 
Decrease in thyroid 
function that may require 
daily thyroid hormone 
replacement 
 
Abnormal kidney and 
liver function that can be 
severe  
 
Abnormal heartbeats or 
low blood pressure that 
may require treatment in 
the ICU  
 
Breathing problems 
which may need 
monitoring in ICU and 
insertion of a breathing 
tube 
 
 
Bowel perforation (a 
hole) requiring longer 
hospitalization or surgery. 
 
Autoimmune disease, 
where your immune 
system attacks cells in 
organs of your body. 
Should this occur, you 
will be treated with 
steroids to stop the 
immune response. 
 
Damage to the heart 
muscle or heart attack 
 
Loss of blood flow to the 
extremities due to 
medicines used to treat 
very low blood pressure 
and shock. In one 
instance, a patient had to 
have her lower arm 
amputated after treatment 
with these medicines. 
 
IL-2 is mixed with human 
albumin which could 
cause an allergic reaction 
or potentially transmit 
viral infections, although 
we have not had this 
occur. 
Support Medications – Side Effects 
Common 
Less Common 
Rare 
Filgrastim (To increase production of white blood cells) 
 
Bone pain 
 
Severe headache 
 
Severe breathing 
problems 
 
Rupture of your spleen 
Bactrim (To prevent a specific type of pneumonia) 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
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STUDY NUMBER: 11-C-0266 
CONTINUATION: page 11 of 17 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
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P.A.: 09-25-0099 
File in Section 4: Protocol Consent 
 
 
 
Fever 
 
Nausea, vomiting  
 
Skin rash with itching 
 
Reduced number of white 
blood cells  
 
Allergic reaction 
 
Fluconazole (To prevent fungal infections) 
 
Headache 
 
Nausea, vomiting, 
diarrhea, abdominal pain 
 
Itching 
 
 
A skin disorder called 
Stevens Johnson 
Syndrome, which can be 
fatal 
 
Liver damage, which may 
be permanent 
Acyclovir and Valacyclovir 
 
 
Temporary decrease in 
kidney function, which 
may not cause any 
symptoms 
 
Nausea, vomiting, 
diarrhea, constipation  
 
Pain and irritation at place 
of injection 
 
Skin rash, hives, itching  
 
Tremors, dizziness, 
confusion, seizures 
 
Fatigue  
 
Blood in the urine 
Gene Therapy Risk of Cancer and Other Disease 
When retroviral vectors enter a normal cell in the body, the DNA of the vector inserts itself into 
the normal DNA in that cell; this process is called integration. Most integration is expected to 
cause no harm to the cell or to the patient. However, there is a chance that there may be some 
regions of the normal human DNA where integration of the viral vector’s DNA may result in 
activation of neighboring genes. For example, if one of these genes were a growth factor, this may 
cause uncontrolled division of the cell, resulting in a cancer. This type of event has occurred in 
one animal study in mice where the vector integration site correlates with the occurrence of cancer 
in these mice. Five instances of a similar event have been reported in five children out of 22 who 
received a retroviral vector in two experimental gene transfer studies for X-linked Severe 
Combined Immunodeficiency (SCID) conducted in Europe, not under the jurisdiction of the U.S. 
Food and Drug Administration (FDA). While most of the children who participated in this clinical 
trial appear to have been cured of their disease, five children developed leukemia (a form of cancer 
of the blood) approximately 2-6 years after receiving the gene transfer regimen. The first patient 
had extensive testing done to determine the cause of the leukemia. A group of experts in this field 


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PATIENT IDENTIFICATION 
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have looked at all the test results, and concluded that the gene transfer caused the leukemia in the 
first child. One of the children died as a result of their leukemia. The risk of another cancer 
developing in you, including leukemia, is unknown, but you need to be aware of this possible risk. 
To monitor you for this risk, we will be testing your blood 3 months after cell infusion, then at 6 
and 12 months, and then annually thereafter. If we find that the cells we have given you grow out 
of control, chemotherapy will be given to you to kill the cells, given their risk of causing leukemia 
or a second cancer. This testing will be conducted on protocol 09-C-0161. 
Potential Benefits of Participation 
Are there benefits to taking part in this study? 
We do not know if you will receive personal, medical benefit from taking part in this study. These 
potential benefits could include shrinking of your tumor or lessening of your symptoms, such as 
pain, that are caused by the cancer. Because there is not much information about the effect of this 
treatment on cancer, we do not know if you will benefit from taking part in this study, although 
the knowledge gained from this study may benefit others in the future who have cancer. 
Alternative Approaches or Treatments 
What other choices do I have if I do not take part in this study?  
Instead of being in this study, you have the following options: 
• Taking part in another study 
• Getting treatment or care for your cancer without being in a study. 
• Getting no treatment or getting comfort care, which is also called palliative care. This type 
of care helps reduce pain, tiredness, appetite problems, and other problems caused by 
cancer. It does not treat the cancer directly, but instead tries to improve how you feel. 
Comfort care tries to keep you as active and comfortable as possible. 
Please talk to your doctor about these and other options. 
Stopping Therapy  
Your doctor may decide to take you off this study if: 
• He/she believes that it is in your best interest 
• Your disease comes back during treatment 
• You become pregnant 
• You have side effects from the treatment that your doctor thinks are too severe 
• New information becomes available that shows another treatment would be better for you 
In this case, you will be informed of the reason for the decision to take you off the study. 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
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STUDY NUMBER: 11-C-0266 
CONTINUATION: page 13 of 17 pages 
 
PATIENT IDENTIFICATION 
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P.A.: 09-25-0099 
File in Section 4: Protocol Consent 
 
You can stop participating in the study at any time. However, if you decide to stop participating in 
the study, we encourage you to talk to the study doctor and your regular doctor first. If you refuse 
to participate or withdraw from the protocol or at the completion of the protocol, we will attempt 
to offer you participation in other NIH protocols if these are available, or will refer you to your 
home physician for further management. 
If you decide at any time to withdraw your consent to participate in the trial, we will not collect 
any additional medical information about you. However, according to FDA guidelines, 
information collected on you up to that point may still be provided to the Sponsor. If you withdraw 
your consent and leave the trial, any samples of yours that have been obtained for the study and 
stored at the NCI can be destroyed upon request. However, any samples and data generated from 
the samples that have already been distributed to other researchers or placed in the research 
databases cannot be recalled and destroyed. 
Research Subject's Rights 
What are the costs of taking part in this study?  
If you choose to take part in the study, the following will apply, in keeping with the NIH policy: 
• You will receive study treatment at no charge to you. This may include surgery, medicines, 
laboratory testing, x-rays or scans done at the Clinical Center, National Institutes of Health 
(NIH), or arranged for you by the research team to be done outside the Clinical Center, 
NIH if the study related treatment is not available at the NIH.  
• There are limited funds available to cover the cost of some tests and procedures performed 
outside the Clinical Center, NIH. You may have to pay for these costs if they are not 
covered by your insurance company. 
• Medicines that are not part of the study treatment will not be provided or paid for by the 
Clinical Center, NIH. 
• Once you have completed taking part in the study, medical care will no longer be provided 
by the Clinical Center, NIH. 
Will your medical information be kept private?   
We will do our best to make sure that the personal information in your medical record will be kept 
private. However, we cannot guarantee total privacy. Organizations that may look at and/or copy 
your medical records for research, quality assurance, and data analysis include: 
• The National Cancer Institute (NCI) and other government agencies, like the Food and 
Drug Administration (FDA), which are involved in keeping research safe for people. 
• NCI Institutional Review Board 
• The study Sponsor, Center for Cancer Research, or their agents 
• Our research collaborator, Kite Pharma, Inc. 


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PATIENT IDENTIFICATION 
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A description of this clinical trial will be available on http://www.Clinicaltrials.gov, as required 
by U.S. Law. This website will not include information that can identify you. At most, the website 
will include a summary of the results. You can search this website at any time.  
Certificate of Confidentiality  
To help us protect your privacy, we have obtained a Certificate of Confidentiality. The researchers 
can use this Certificate to legally refuse to disclose information that may identify you in any 
federal, state, or local civil, criminal, administrative, legislative, or other proceedings, for example, 
if there is a court subpoena. The researchers will use the Certificate to resist any demands for 
information that would identify you, except as explained below. 
You should also know that there are several circumstances in which the Certificate does not 
provide coverage. These include when information:  
• will be used for auditing or program evaluation internally by the NIH; or  
• must be disclosed to meet the legal requirements of the federal Food and Drug 
Administration (FDA).  
• is necessary for your medical treatment and you have consented to this disclosure;  
• is for other research.  
In addition, identifiable, sensitive information protected by this Certificate cannot be admissible 
as evidence or used for any purpose in any action, suit, or proceeding without your consent.  
You should understand that a Certificate of Confidentiality does not prevent you or a member of 
your family from voluntarily releasing information about yourself or your involvement in this 
research. If an insurer, employer, or other person obtains your written consent to receive research 
information, then the researchers will not use the Certificate to withhold that information. 
The Certificate of Confidentiality will not protect against the required reporting by hospital staff 
of information on suspected child abuse, reportable communicable diseases, and/or possible threat 
of harm to self or others. 
Conflict of Interest 
The National Institutes of Health (NIH) reviews NIH staff researchers at least yearly for conflicts 
of interest. This process is detailed in a Protocol Review Guide. You may ask your research team 
for additional information or a copy of the Protocol Review Guide. Members of the research team 
who do not work for NIH are expected to follow these guidelines but they do not need to report 
their personal finances to the NIH.  
Members of the research team working on this study may have up to $15,000 of stock in the 
companies that make products used in this study. This is allowed under federal rules and is not a 
conflict of interest.  


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 11-C-0266 
CONTINUATION: page 15 of 17 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.: 09-25-0099 
File in Section 4: Protocol Consent 
 
The NIH and the research team for this study are working with Kite Pharma, Inc. to see if this type 
of study could be done at institutions other than the NIH Clinical Center. Kite Pharma, Inc. also 
provides financial support for this study. 
Use of Specimens and Data for Future Research 
Blood and tissue collected during the course of this study will be used for future research and will 
be stored, tracked and disposed of under our companion protocol 03-C-0277.  
In addition, to advance science, it is helpful for researchers to share information they get from 
studying human samples. They do this by putting it into one or more scientific databases, where it 
is stored along with information from other studies. A researcher who wants to study the 
information must apply to the database and be approved. Researchers use specimens and data 
stored in scientific databases to advance science and learn about health and disease.  
We plan to keep some of your specimens and data that we collect and use them for future research 
and share them with other researchers. We will not contact you to ask about each of these future 
uses. These specimens and data will be stripped of identifiers such as name, address or account 
number, so that they may be used for future research on any topic and shared broadly for research 
purposes. Your specimens and data will be used for research purposes only and will not benefit 
you. It is also possible that the stored specimens and data may never be used. Results of research 
done on your specimens and data will not be available to you or your doctor. It might help people 
who have cancer and other diseases in the future.  
If you do not want your stored specimens and data used for future research, please contact us in 
writing and let us know that you do not want us to use your specimens and/or data. Then any 
specimens that have not already been used or shared will be destroyed and your data will not be 
used for future research. However, it may not be possible to withdraw or delete materials or data 
once they have been shared with other researchers. 
 


MEDICAL RECORD 
CONSENT TO PARTICIPATE IN A CLINICAL RESEARCH STUDY 
• 
Adult Patient or 
• Parent, for Minor Patient 
STUDY NUMBER: 11-C-0266 
CONTINUATION: page 16 of 17 pages 
 
PATIENT IDENTIFICATION 
CONSENT TO PARTICIPATE IN A CLINICAL RESEARCH 
STUDY (Continuation Sheet) 
• 
Adult Patient or 
• Parent, for Minor Patient 
NIH-2514-1 (07-09) 
P.A.: 09-25-0099 
 
File in Section 4: Protocol Consent 
 
 
 
OTHER PERTINENT INFORMATION 
1. 
Confidentiality. When results of an NIH research study are reported in medical journals 
or at scientific meetings, the people who take part are not named and identified. In most cases, the 
NIH will not release any information about your research involvement without your written 
permission. However, if you sign a release of information form, for example, for an insurance 
company, the NIH will give the insurance company information from your medical record. This 
information might affect (either favorably or unfavorably) the willingness of the insurance 
company to sell you insurance. 
The Federal Privacy Act protects the confidentiality of your NIH medical records. However, you 
should know that the Act allows release of some information from your medical record without 
your permission, for example, if it is required by the Food and Drug Administration (FDA), 
members of Congress, law enforcement officials, or authorized hospital accreditation 
organizations. 
2. 
Policy Regarding Research-Related Injuries. The Clinical Center will provide short-
term medical care for any injury resulting from your participation in research here. In general, no 
long-term medical care or financial compensation for research-related injuries will be provided by 
the National Institutes of Health, the Clinical Center, or the Federal Government. However, you 
have the right to pursue legal remedy if you believe that your injury justifies such action. 
3. 
Payments. The amount of payment to research volunteers is guided by the National 
Institutes of Health policies. In general, patients are not paid for taking part in research studies at 
the National Institutes of Health. Reimbursement of travel and subsistence will be offered 
consistent with NIH guidelines. 
4. 
Problems or Questions. If you have any problems or questions about this study, or about 
your rights as a research participant, or about any research-related injury, contact the Principal 
Investigator, Steven A. Rosenberg, M.D., Ph.D., Building 10 CRC, Room 3-3940, Telephone: 
240-760-6218. If you have any questions about the use of your specimens or data for future 
research studies, you may also contact the Office of the Clinical Director, Telephone: 240-760-
6070. You may also call the Clinical Center Patient Representative at 301-496-2626.  
5. 
Consent Document. Please keep a copy of this document in case you want to read it again. 
 
 


MEDICAL RECORD 
CONSENT TO PARTICIPATE IN A CLINICAL RESEARCH STUDY 
• 
Adult Patient or 
• Parent, for Minor Patient 
STUDY NUMBER: 11-C-0266 
CONTINUATION: page 17 of 17 pages 
 
PATIENT IDENTIFICATION 
CONSENT TO PARTICIPATE IN A CLINICAL RESEARCH 
STUDY (Continuation Sheet) 
• 
Adult Patient or 
• Parent, for Minor Patient 
NIH-2514-1 (07-09) 
P.A.: 09-25-0099 
 
File in Section 4: Protocol Consent 
 
 
 
 
COMPLETE APPROPRIATE ITEM(S) BELOW: 
A. Adult Patient’s Consent 
B. Parent’s Permission for Minor Patient 
I have read the explanation about this study 
and have been given the opportunity to 
discuss it and to ask questions. I hereby 
consent to take part in this study. 
I have read the explanation about this study 
and have been given the opportunity to 
discuss it and to ask questions. I hereby give 
permission for my child to take part in this 
study.  
(Attach NIH 2514-2, Minor’s Assent, if 
applicable.) 
 
  
  
 
 
 
Signature of Adult Patient/ 
Legal Representative 
Date 
Signature of Parent(s)/ 
Guardian 
Date 
 
 
Print Name 
 
 
Print Name 
C. Child’s Verbal Assent (If Applicable) 
The information in the above consent was described to my child and my child agrees to 
participate in the study. 
 
 
 
 
 
 
 
Signature of Parent(s)/Guardian 
Date 
Print Name 
THIS CONSENT DOCUMENT HAS BEEN APPROVED FOR USE  
FROM JULY 09, 2018 THROUGH FEBRUARY 04, 2019. 
 
  
 
 
  
 
Signature of Investigator 
Date 
Signature of Witness 
Date 
 
 
Print Name 
 
 
Print Name