Conditions: Metastatic Non-small Cell Lung Cancer
Keywords: lung cancer immune checkpoint PD-1 CRISPR autologous cell infusion
Allocation: NON_RANDOMIZED | Intervention model: PARALLEL
Enrollment: 12
Healthy volunteers: no

Brief summary:
This study will evaluate the safety of PD-1 knockout engineered T cells in treating metastatic non-small cell lung cancer. Blood samples will also be collected for research purposes.

Detailed description:
This is a dose-escalation study of ex-vivo knocked-out, expanded, and selected PD-1 knockout-T cells from autologous origin. Patients are assigned to 1 of 3 treatment groups to determine the maximal tolerant dose. After the lower number of cycles are considered tolerant, an arm of the next higher number of cycles will be open to next patients. Biomarkers and immunological markers are collected and analyzed as well.

Arms:
arm 1: Peripheral blood lymphocytes will be collected and Programmed cell death protein 1(PDCD1) gene will be knocked out by CRISPR Cas9 in the laboratory (PD-1 Knockout T cells). The lymphocytes will be selected and expanded ex vivo and infused back into patients. Cyclophosphamide at 20mg/kg single dose will be administered 3 days i.v. before cell infusion.

A total of 1 x 10\^7/kg PD-1 Knockout T cells will be infused in one cycle. Each cycle is divided into three administrations, with 20% infused in the first administration, 30% in the second, and the remaining 50% in the third. Patients will receive a total of two cycles of treatment. arm 2: Peripheral blood lymphocytes will be collected and Programmed cell death protein 1(PDCD1) gene will be knocked out by CRISPR Cas9 in the laboratory (PD-1 Knockout T cells). The lymphocytes will be selected and expanded ex vivo and infused back into patients. Cyclophosphamide at 20mg/kg single dose will be administered 3 days i.v. before cell infusion.

A total of 2 x 10\^7/kg PD-1 Knockout T cells will be infused in one cycle. Each cycle is divided into three administrations, with 20% infused in the first administration, 30% in the second, and the remaining 50% in the third. Patients will receive a total of two cycles of treatment. arm 3: Peripheral blood lymphocytes will be collected and Programmed cell death protein 1(PDCD1) gene will be knocked out by CRISPR Cas9 in the laboratory (PD-1 Knockout T cells). The lymphocytes will be selected and expanded ex vivo and infused back into patients. Cyclophosphamide at 20mg/kg single dose will be administered 3 days i.v. before cell infusion.

A total of 4 x 10\^7/kg PD-1 Knockout T cells will be infused in one cycle. Each cycle is divided into three administrations, with 20% infused in the first administration, 30% in the second, and the remaining 50% in the third. Patients will receive a total of two cycles of treatment.

Interventions: intervention 1: Cyclophosphamide intervention 2: PD-1 Knockout T Cells
Intervention descriptions: intervention 1: To deplete Tregs before collecting peripheral blood intervention 2: Autologous lymphocytes are collected and PDCD1 gene is knocked out in the laboratory. Cells are selected and expanded ex vivo. Cells are infused back to the patients for treatment

Protocol document:
Prot_SAP_000.pdf: 
 
 
Gene Edited T-Cell Therapy for Advanced Lung Cancer.  Version 1.9 
1 
 
 
 
 
A phase I clinical study of PD-1 knockout engineered T cells 
treating patients with advanced Non-Small Cell Lung Cancer 
Protocol No.: MHC-001 
 
 
 
 
Designers of this clinical study protocol:  
West China Hospital, Sichuan University  
Chengdu MedGenCell, Co., Ltd 
Sponsor:  
West China Hospital, Sichuan University  
Principal Investigator:  
Prof. You Lu 
Version No. of the study protocol:  
Version 1.9  
Amended version No.:  
Version 9 
Amended date:  
Dec 4, 2018  
 
 
 
 
 
All rights are reserved by the West China Hospital, Sichuan University 
Confidential 
Without the consent of the West China Hospital, Sichuan University, this protocol will 
not be allowed to be used, divulged, published or otherwise disclosed. This amended 
protocol will replace the protocol of previous version. Please provide copies of the amended 
protocol to clinical staff related and archive the previous version under your supervision. 


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Gene Edited T-Cell Therapy for Advanced Lung Cancer.  Version 1.9 
 
Contents 
List of abbreviations ......................................................................................................... 1 
Protocol synoposis ............................................................................................................ 4 
1 Study background ................................................................................................................. 11 
1.1 Overview of tumor immunotherapy background ................................................ 11 
1.2 Application prospect of gene editing in immunotherapy study ........................... 12 
1.3The study technology and work basis ................................................................... 13 
1.3.1 Study-related technology............................................................................. 13 
1.3.2 Pre-clinical data ........................................................................................... 14 
2 Study objectives ........................................................................................................... 18 
2.1 Primary objective ................................................................................................. 18 
2.2Secondary objectives ............................................................................................ 18 
2.3 Primary endpoint ................................................................................................. 18 
2.4Secondary endpoint .............................................................................................. 18 
3 Study design ........................................................................................................................... 18 
3.1 Description of the study design ........................................................................... 18 
3.1.1 Molecular screening .................................................................................... 20 
3.1.2 Screening period .......................................................................................... 20 
3.1.3 Treatment period ......................................................................................... 21 
3.1.4 End of treatment .......................................................................................... 21 
3.1.5 Follow-up period ......................................................................................... 21 
3.1.6 Survival follow-up period ........................................................................... 21 
3.2 Definition of the end of study .............................................................................. 21 
3.3 Early termination of the study ............................................................................. 21 
4 Population .................................................................................................................... 22 
4.1 Inclusion criteria .................................................................................................. 22 
4.2 Exclusion criteria ................................................................................................. 23 
5 Treatment ..................................................................................................................... 24 
5.1 Cell therapy protocol ........................................................................................... 25 
5.1.1 Pre-treatment ............................................................................................... 25 
5.1.2 Cell infusion ................................................................................................ 25 
5.1.2.1 Treatment assignment ............................................................................... 25 
5.1.2.2 Cell infusion procedure ............................................................................ 26 
5.2 Concomitant treatment ......................................................................................... 27 
5.2.1 Acceptable concomitant treatment .............................................................. 27 


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5.2.2 Local palliative treatment ............................................................................ 27 
5.2.3 Restricted treatment..................................................................................... 28 
5.2.4 Treatment duration ...................................................................................... 28 
5.3. Dose escalation guidelines .................................................................................. 28 
5.3.1 Starting dose rationale ................................................................................. 28 
5.3.2 Provisional dose levels ................................................................................ 29 
5.3.3 Guidelines for dose escalation and determination....................................... 29 
5.3.4 Intrapatient dose escalation ......................................................................... 29 
5.3.5 Other dosing regimens ................................................................................ 29 
5.3.6 Definition of Dose Limiting Toxicity .......................................................... 29 
5.4 Dose modification ................................................................................................ 30 
5.4.1 Criteria for cell therapy delay, reinitiation and discontinuation .................. 30 
5.4.1.1 Criteria for delayed cell infusion .............................................................. 30 
5.4.1.2 Criteria of reinitiation of cell therapy ....................................................... 31 
5.4.1.3 Criteria of discontinuation of cell therapy................................................ 31 
5.4.2 Follow-Up for DLT and other toxicities ...................................................... 31 
5.4.3 Anticipated risk and safety considerations of the study treatment .............. 32 
5.5 Supportive treatment ............................................................................................ 32 
5.5.1 Cytokine release syndrome (CRS) .............................................................. 32 
5.5.2 Treatment transfusion/allergic reaction related to cell therapy ................... 35 
6. Patient numbering, treatment assignment or randomization ................................ 37 
6.1 Patient numbering ................................................................................................ 37 
6.2 Treatment assignment or randomization .............................................................. 37 
6.3 Treatment blinding ............................................................................................... 37 
6.4 Preparations for cell therapy ................................................................................ 37 
6.4.1 Package and labeling of the study cells ....................................................... 37 
6.4.2 Cell supply and storage ............................................................................... 37 
6.4.3 Cell product compliance and records .......................................................... 38 
6.4.3.1 Compliance............................................................................................... 38 
6.4.3.2 Record of the study cell therapy ............................................................... 38 
6.4.3.3 Disposal and destruction .......................................................................... 38 
7 Visit schedule and assessments ................................................................................... 38 
7.1 Visit schedule ....................................................................................................... 38 
7.1.1 Molecular screening .................................................................................... 42 
7.1.2 Screening ..................................................................................................... 42 


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7.1.3 Screening failure ......................................................................................... 44 
7.1.4 Treatment phase........................................................................................... 44 
7.1.4.1 Major treatment period ............................................................................. 44 
7.1.4.1.1 Assessment within one week prior to each treatment cycle .................. 44 
7.1.4.1.2 Assessment on day eight of each cycle ................................................. 45 
7.1.4.1.3 End of cell therapy ................................................................................ 45 
7.1.5 Follow-Up after end of cell therapy ............................................................ 46 
7.1.6 End of study (EOT) ..................................................................................... 47 
7.1.7 Follow-ups ................................................................................................... 48 
7.1.7.1 The safety of follow-up period ................................................................. 48 
7.1.7.2 PFS follow-up period ............................................................................... 49 
7.1.7.3 Survival follow-up period ........................................................................ 49 
7.2 Assessment types ................................................................................................. 49 
7.2.1 Safety and tolerability assessments ............................................................. 49 
7.2.1.1 Physical examination ............................................................................... 49 
7.2.1.2 Vital signs ................................................................................................. 50 
7.2.1.3 Height and weight .................................................................................... 50 
7.2.1.4 Performance status ................................................................................... 50 
7.2.1.5 Laboratory assessments ............................................................................ 50 
7.2.1.6 Tolerability evaluation .............................................................................. 51 
7.2.2 Measurement of efficacy ............................................................................. 51 
7.2.3 Biomarkers .................................................................................................. 52 
7.2.3.1 ctDNA ...................................................................................................... 53 
7.2.3.2 Immune repertoire .................................................................................... 53 
7.2.3.3 Screening of the tumor biomarkers .......................................................... 54 
7.2.3.4 PDX model ............................................................................................... 54 
7.2.3.5 In vivo T cell tracking ............................................................................... 54 
7.2.3.6 Flow cytometery for regulatory T (Treg) cells, memory T cells, activated   
T cells and PD-1 ........................................................................................... 55 
7.2.4 Other descriptions ......................................................................................... 55 
7.2.5 Withdrawal of ICF ......................................................................................... 56 
7.2.6 Discontinuation of the study ......................................................................... 56 
8. Adverse events ............................................................................................................ 56 
8.1 Serious adverse events ......................................................................................... 58 
8.1.1 Definition of serious adverse events ........................................................... 58 


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8.1.2 Report .......................................................................................................... 58 
8.2 Pregnancy ............................................................................................................ 59 
9. Data management and analysis ................................................................................. 59 
9.1 Data confidentiality ............................................................................................. 59 
9.2 Site monitoring .................................................................................................... 60 
9.3 Data collection ..................................................................................................... 60 
9.4 Statistical methods and data analysis ................................................................... 60 
9.4.1 Analysis data sets ........................................................................................ 60 
9.4.2 Patient demographics / other baseline characteristics ................................. 61 
9.4.3 Treatments (study treatment, concomitant therapies, compliance) ............. 61 
9.4.4 Primary objective ........................................................................................ 61 
9.4.4.1 Variable ..................................................................................................... 61 
9.4.4.2 Safety analysis .......................................................................................... 61 
9.4.4.3 Tolerability ............................................................................................... 62 
9.4.5 Secondary objective analysis ...................................................................... 62 
9.4.5.1 Efficacy analysis ...................................................................................... 62 
9.4.5.2 In vivo activity status and duration of PD-1 edited T cells ...................... 63 
9.4.5.3 Immunological factors and biomarkers .................................................... 63 
9.4.6 Exploratory analysis .................................................................................... 63 
9.4.7 Interim analysis ........................................................................................... 63 
9.4.8 Sample size calculation ............................................................................... 64 
10 Ethical considerations ............................................................................................... 64 
10.1 Regulatory and ethical compliance .................................................................... 64 
10.2 Responsibilities of the investigators and IRB .................................................... 64 
10.3 Informed consent procedures ............................................................................. 64 
10.4 Discontinuation of the study .............................................................................. 65 
10.5 Publication of study protocol and results ........................................................... 65 
10.6 Study documentation, record keeping and retention of documents ................... 65 
10.7 Confidentiality of study documents and patient records ................................... 66 
10.8 Compensation and insurance ............................................................................. 66 
10.8.1 Payment for transportation expenses, etc. ................................................. 66 
10.8.2 Indemnity and insurance ........................................................................... 66 
11. Protocol compliance ................................................................................................. 66 
11.1 Study protocol amendment ................................................................................ 66 
12. Appendix ................................................................................................................... 68 


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12.1 ECOG performance status ................................................................................. 68 
12.2 RECIST 1.1  Guideline ...................................................................................... 69 
12.3 irRC evaluation criteria ...................................................................................... 82 
References ....................................................................................................................... 86 


 
 
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1
List of abbreviations 
AE  
Adverse event  
A/G  
Albumin/ globulin  
AKP  
Alkaline phosphatase  
ALB  
Albumin  
ALK  
Anaplastic lymphoma kinase  
ALT  
Alanine transaminase / glutamic-pyruvic transaminase  
ANC  
Absolute neutrophil count  
APTT  
Activated partial thrombplastin time 
AST  
Aspartate aminotransferase / glutamic oxalacetic transaminase  
bTMB 
Blood tumor mutation burden 
BUN  
Blood urea nitrogen  
CABG  
Coronary artery bypass grafting  
CAR-T 
Chimeric antigen receptor T cell therapy 
CEA  
Carcinoma embryonic antigen  
CDR3  
Complementary determining region 3  
CHF  
Congestive heart failure  
CIK  
Cytokine-induced killer cells  
Cl  
Chlorine  
CK-MB  
Creatine kinase MB 
CNS  
Central nervous system  
CR  
Complete response  
Cr  
Creatinine  
CRF  
Case report form  
CRP  
C-reactive protein  
CRS  
Cytokine release syndrome  
CSR  
Clinical study report  
CT  
Computed tomography  
CTCAE4.03  
Common terminology criteria for adverse events, version 4.03  
CtDNA  
Circulating tumor DNA  
CTLA-4  
Cytotoxic T-lymphocyte antigen-4 
CYFRA21-1  
Cytokeratin fragment antigen 21-1  
DCR 
Disease control rate 
DFS  
Disease free survival  
DLT  
Dose-limiting toxicity  
DOR  
Duration of response  
EC  
Ethics committee  
ECG  
Electrocardiograph 


 
 
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ECOG  
Eastern Cooperative Oncology Group 
eCRF  
Electronic case report form  
EDC  
Electronic data capture  
LVEF  
Left ventricular ejection fraction  
EGFR  
Epidermal growth factor receptor  
EOT  
End of treatment 
FDG  
F-deoxyglucose  
GCP  
Good Clinical Practice  
G-CSF  
Granulocyte colony-stimulating factor  
GVHD  
Graft-versus-host disease  
HBV  
Hepatitis B virus  
HBV sAg  
Hepatitis B virus surface antigen  
HCV Ab  
Hepatitis C virus Antibody  
HCG  
Human chorionic gonadotropin  
HIV  
Human immunodeficiency virus  
ICF  
Informed consent form 
ICH  
International Council for Harmonization (drug registration)  
IFN-α 
Interferon-α 
IFN-γ  
Interferon-λ  
INR  
International normalized ratio  
IrRC  
Immune-related response criteria 
irCR  
Immune-related Complete Response  
irPR  
Immune-related Partial Response 
irSD  
Immune-related Stable Disease  
irPD  
Immue-related Progressive Disease 
IUD  
Intrauterine device  
IUS  
Intrauterine system  
Ka  
Potassium  
LAHB  
Left anterior hemi-block  
LDH  
Lactic dehydrogenase  
MRI  
Magnetic resonance imaging  
mTOR  
Mammalian target of rapamycin  
Na  
Sodium  
NCI  
National Cancer Institute  
NE  
Not evaluable  
NS  
Normal saline  
NSCLC  
Non-small cell lung cancer  
NSE  
Neuronspecific enolase  


 
 
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NY-ESO-1  
New York - esophageal squamous cell carcinoma-1 
OS  
Overall survival  
PD  
Progressive disease  
PFS  
Progression free survival  
PD-1  
Programmed cell death -1  
PD-L1  
Programmed death - ligand 1  
PET-CT  
Positron emission tomography Computed tomography 
PPF  
Production possibility frontier  
PR  
Partial response  
PTT  
Partial thromboplastin time  
QTc  
QT correction  
RECIST  
Response evaluation criteria in solid tumors  
RR  
Response rate 
SAE  
Serious adverse event  
SD  
Stable disease  
sgRNA  
Single-guide RNA  
TBL  
Total bilirubin  
TCR 
T cell receptor 
TKI  
Tyrosine kinase inhibitor  
TNF-α  
Tumor necrosis factor-α  
TSH  
Thyroid-stimulating hormone 
TTP  
Time to progress  
ULN  
Upper limit of the normal 
WBC  
White blood cell  
WOCBP  
Women of child-bearing period  
 
 
 
 
 
 
 
 
 
 
 
 


 
 
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Protocol synoposis 
 
Protocol No.  
MHC-001  
Title 
A phase I clinical study of PD-1 knockout engineered T cells treating 
patients with advanced Non-Small Cell Lung Cancer 
Sponsor  
West China Hospital, Sichuan University 
Clinical 
Phase 
Phase I  
Study type  
Interventional treatment  
Objective and 
principle  
To investigate the safety and tolerability of CRISPR-Cas9 edited 
autologous PD-1 T cells (PD-1 edited T cell) in patients with PD-L1 
postive advanced non-small cell lung cancer (NSCLC) after failure of 
multiple lines of prior standard treatments. 
Primary 
objective  
To investigate the safety and tolerability of CRISPR-Cas9 PD-1 edited 
T cell in patients with advanced NSCLC. 
Secondary 
objective  
1) To investigate the antitumor activity of CRISPR-Cas9 edited PD-1 T 
cells in patients with advanced NSCLC. 
2) To investigate the changes in the potential immunological indexes 
and biomarkers. 
3) To track the PD-1 edited T cells in patients and investigate the 
mechanism of the clinical antitumor activity. 
Primary 
endpoint  
The dose-limiting toxicity (DLT) rate of the cell therapy during the first 
treatment cycle.  


 
 
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Secondary 
endpoint  
1) To evaluate the objective response rate (ORR) and 8-week disease 
control rate (DCR) of the cell therapy.  
2) To evaluate time to first response, defined as the time from the first 
cell infusion to the first observed complete response (CR) or partial 
response (PR).  
3) To evaluate the duration of response (DOR), defined as the time from 
the first observed CR or PR to the first observed PD or death from any 
cause.  
4) To evaluate the progression free survival (PFS), defined as the time 
from the first cell infusion to the first observed PD or death from any 
cause. 
5) To evaluate the overall survival (OS), defined as the time from the 
first cell infusion to death.  
6) To evaluate the activation status and duration of PD-1 edited T cell 
after infusion.  
7) To evaluate the change of in vivo cytokine levels and their correlation 
to the tumor response and adverse events of after PD-1 edited T cell 
infusion. 
Study design  
This phase I clinical study aims to investigate the safety and 
tolerability of PD-1 edited T cell therapy in patients with advanced 
NSCLC.  
About 20 patients with PD-L1 positive advancedNSCLC who have 
progressed after the multiple lines of prior standard therapeutic 
regimens are planned to be enrolled in this study. The safety and 
tolerability of the therapy will be investigated by using intergroup dose 
escalation method.  
This clinical study was a first-in-human trial using CRISPR 
technology knockouting PD-1 gene in T cell to treat lung cancer 
patients. Although there were no exact dosage/regimen and safty data, 
starting dose was selected referring to cell therapy or CAR-T treatment 
for considering benefit-risk of cancer patients (see section 5.3.1 Starting 
dose rationale).  Pre-A: 2x107 PD-1 edited T cells per kilogram of body 
weight（percentage of PD-1 knockout T cell may vary depending on 


 
 
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technology efficiency. The study defines the Pre-A group as a 
preliminary exploratory dose group. The Pre-A group has a 28 days-
observation after the first treatment cycle. If no DLT was observed in 
Pre-A group during the defined observation period, the study will 
continue other treat groups according to provisional dosage; If DLT or 
other unexpected toxicity occurs in the Pre-A group, the investigator 
will adjust the cell dose and interval of sequential cell therapy based on 
data from Pre-A group. 
There was three treatment groups and 28 days/cycle. 
Group A: 1x107 PD-1 edited T cells per kilogram of body weight, 2 
treatment cycles;  
Group B: 2x107 PD-1 edited T cells per kilogram of body weight, 2 
treatment cycles;  
Group C: 4x107 PD-1 edited T cells per kilogram of body weight, 2 
treatment cycles. 
(Treatment may continue after 2 cycles if considered to be benefitial to 
patient) 
 
Inclusion 
criteria  
1.  Informed consent form (ICF) is signed:  
(1) The enrolled patients must sign and date the EC-approved written 
ICFs according to the guidelines of the competent authority and study 
institution. The ICF should be signed before implementing any 
protocol-related procedure (not including the routine medical care of the 
patients).  
(2) The enrolled patients must be willing and able to comply with the 
visits, therapeutic regimen, and laboratory examinations specified in the 


 
 
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schedule as well as the other requirements in the study.  
2.  Age: 18-70 years.  
3. ECOG performance status 0-2. 
4.  NSCLC, stage IIIB/IV confirmed by histology or cytology.   
All patients must be screened for EGFR gene mutation and ALK 
gene fusion; patients with positive EGFR mutation or ALK gene fusion 
treatment failure of EGFR-TKI or ALK-TKI must have been 
experienced. 
5.   Patients must fail from the standard third-line or above therapy 
scheme. 
6.  Patients must be positive for PD-L1staining, as tested through tumor 
biopsy (TPS ≥ 1%).  
7.  Patients must have a measurable lesion (tumor lesion with a long 
diameter of more than 10 mm through CT scan; lymph node with a short 
diameter of more than 15 mm through CT scan) in accordance with the 
RECIST 1.1. 
8.  Life expectency of ≥ 3 months.  
9.  Adequate bone marrow function:  
• WBC ≥ 3.0×109/L. 
• ANC ≥ 1.5 ×109/L.  
• PLT ≥ 90×109/L, without transfusions within 14 days before the first 
cell therapy. 
•  Hemoglobin ≥ 10.0 g/dL.  
10.  Adequate hepatic and renal function:  
Total bilirubin ≤ 1.5×ULN. 
• AST and ALT < 2.5×ULN; AST and ALT < 5×ULNif metastasis to 
liver.  
• Serum creatinine ≤1.5×ULN, or creatinine clearance rate ≥ 50 
ml/min (based on the Cockcroft/Gault formula). 
11. Coagulation functions:  
•  INR ≤ 1.5×ULN. 
•  PTT ≤ 1.5×ULN. 
12.Adequate cardiovascular function:  
•  Left ventricular ejection fraction ≥ 50%.  


 
 
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    •  QTcF interval ≤ 450 ms. 
13.  ALP ≤ 2.5×ULN.  
14. Recovery from all adverse events of previous systemic anti-
cancertherapies to baseline or Grade ≤ 1, except forAlopecia. 
15.  Washout period for the previous anti-tumor treatment is not less 
than four weeks and washout period for the molecular targeting drug is 
not less than five half-life periods. Palliative radiotherapy must have 
been completed for at least two weeks; thoracic radiation therapy must 
have been completed for at least three months, and major surgery must 
have been completed for at least four weeks with recovery.  
16.  The patients must take contraception measures strictly.  
• Within the 72-hour period prior to the therapy, the serum or urine 
pregnancy test (HCG sensitivity is 25 IU/L at least or equivalent) of the 
women of childbearing potential (WOCBP) must be negative.  
• WOCBP must agree to take contraceptive measures strictly during 
the treatment period and for at least 18 months after the final cell 
infusion until no knockout T cell is detected in the blood.  
•  The males who have sexual contact with WOCBP must agree to 
take contraceptive measures strictly during the treatment period and for 
at least 18 months after the final cell infusion until no knockout T cell 
is detected in the blood. 
Exclusion 
criteria 
1. Patients with active central nervous system (CNS) metastasis 
(including but not limited to carcinomatous meningitis and spinal cord 
compression) must be excluded. However,patients with metastatic CNS 
tumors may participate in this study if thepatients can be recoved to the 
baseline level at least two weeks before inclusion (without the residual 
signs or symptoms related to the CNS treatment).  In addition, the 
patients must have stopped using corticosteroids for four weeks before 
inclusion.  
2. Patients with weight loss in the recent six months ≥ 10%.  
3. Patients accompanied by emergent symptoms caused by tumor that 
need immediate intervention.  
4. Patients with medical history of interstitial lung disease (ILD) or 
immune related pneumonia.  


 
 
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5.  Patients with peripheral neuropathy. 
6. Patients with clinically significant coagulation abnormalities, or 
patients who are receiving thrombolytic or anticoagulant therapy.  
7. Patients with a history of human immunodeficiency virus (HIV) 
positivity or with the acquired immunodeficiency syndrome (AIDS), or 
with active hepatitis B or hepatitis C. 
8. Patients that have other malignancy.  
9. Patients with active, known or suspected autoimmune diseases.  
10. Patients who have received systemic treatment by corticosteroids 
(the 
equivalent 
dose 
>10 
mg 
prednisone/day) 
or 
other 
immunomodulators (interleukin-2, IFN-α, IFN-γ, cyclosporine, G-CSF, 
mTOR inhibitor) within the 28 days prior to the study. Patients who 
receive inhaled or topical corticosteroids and adrenal corticosteroids 
(the equivalent treating dose <10 mg prednisone/ day) can participate in 
the study if they are lack of active autoimmune diseases.  
11.  Clinically significant cardiac disease or impaired cardiac function, 
such as: 
•  Clinically significant heart disease such as CHF requiring 
treatment, or uncontrolled hypertension defined by blood pressure > 
140/100 mmHg at rest (taking mean of 3 consecutive readings). 
•  History (< 6 months prior to screening) or current evidence 
ofclinically 
significant 
cardiac 
arrhythmias, 
atrial 
fibrillation 
and/orconduction abnormality, e.g. congenital long QT syndrome, 
Grade ≥2/complete AV-blockage or hypokaliemia CTCAE Grade ≥ 3. 
• History/evidence of acute coronary syndromes (including 
myocardialinfarction, unstable angina, coronary artery bypass graft 
(CABG),coronary angioplasty, or stenting), < 6 months prior to 
screening. 
• Complete left bundle branch block (CLBBB).  
• Right bundle branch block (RBBB) + left anterior hemiblock 
(LAHB - bifascicular block).   
12. Patients with a history of allergies to cyclophosphamide and other 
agents involved in this study. 
13. Patients that do not have enough venous access.  


 
 
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14. Pregnant or lactating women, where pregnancy is defined as the 
state of a female from conception until the termination of gestation.  
15. Patients with mental disorders are mandatorily detained for 
treatment or are judged as inappropriate for inclusion by the 
investigator.  
16. Patients who are difficult to communicate with or difficult to be 
followed up for a long time.  
17. Other inappropriate conditions in discretion of the investigator’s 
judgment. 
Safety 
evaluation  
The incidence rate and severity of AE (CTCAE4.03) and SAE, 
including laboratory tests, vital signs, ECG changes, etc.  
Effectiveness 
evaluation  
Tumors are assessed at baseline, the 8th week, the 12th week and once 
every 8 weeks during the treatment and follow-up period per RECIST 
1.1.  
Schedule  
2016/08-2018/05: Recruitment and treatment periord.  
2018/06-2018/12: Follow-up, data analysis and reporting. 


 
 
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1. Study background 
1.1 Overview of tumor immunotherapy background 
Immunotherapy has been considered as an effective treatment for cancer, following surgery, 
chemoradiotherapy and targeted therapy. In the battle against tumor cells, the activity, survival, 
and migration of T cells are being inhibited by the evolved tumor cells through different ways. 
Therefore, the anti-tumor effect of T cells decreases or dysfunctions significantly, resulting in 
immune tolerance and immune escape. The immune checkpoints, such as PD-1/PD-L1 axis, are 
vital factors for immune escape.  
Antibodies targeting the immune checkpoints of PD-1/PD-L1 and cytotoxic T cell antigen-
antibody 4 (CTLA-4) are the two most attractive immunotherapy targets, which have achieved a 
breakthrough in caner therapeutic field [1-3]. The clinical application of a CTLA-4 inhibitor for 
melanoma was approved by the U.S. Food and Drug Administration (FDA) in 2011, and a PD-1 
inhibitor was approved for NSCLC in 2015. The CheckMate 017 study [4] indicated that Nivolumab 
(a PD-1 inhibitor), as a second line treatment for patients with squamous cell cancer, could prolong 
the overall survival (OS) by 3.2 months and acheived a higher one-year OS rate than the standard 
chemotherapy therapy (42% vs 24%; NCT01642004). The CheckMate 057 study also revealed 
that Nivolumab could extend the OS by 2.8 months as a second-line treatment for non-squamous 
cell lung cancer compared with standard chemotherapy (NCT01673867) [5]. The KEYNOTE 001 
reported that Pembrolizumab (a PD-1 inhibitor) treatment extended the OS to 12 months in 
NSCLC patients (NCT01295827) [6]. These inspiring results make immunotherapy to target T cells 
by blocking immune checkpoints a research hotspot.  
However, monoclone antibody to blocking immune checkpoint has inherent disadvantages: 
(1) there are different immune checkpoints functioning on specific sites, and it is difficult to apply 
antibodies in combination; (2) antibodies only act on membrane proteins; (3) the action of 
antibodies is temporary and their effect will disappear rapidly due to degradation; (4) resistance to 
immunotherapy is common with neutralized antibody.  
Transfusion of specific PD-1 knockout T cells back into the body might not only overcome 
the above disadvantages, but also avoid susceptibility to tuberculosis and viral hepatitis, and reduce 
the incidence of dilated cardiomyopathy and lupus-like autoimmune disease after complete-PD-1 
knock therapy [7-10]. It has been reported that the transfusion of PD-1 knockout CD8+T cells from 
PD-1 knockout mouse spleen and peripheral blood into wild-type mice decreases the functional 
cytokine secretion of individual cells for a short period of time. However, accelerated proliferation 
of the PD-1 knockout CD8+T cells produced more cells and finally increased the overall functional 
cytokine secretion.  It has also been reported that apoptosis of the PD-1 knockout CD8+T cells 


 
 
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increases late after 42 days of transfusion, which indirectly suggests that PD-1 knockout CD8+T 
cell transfusion is safe [11].  
1.2 Application prospect of gene editing in immunotherapy study 
Conventional gene knockout technology is very complex, expensive, inefficient, easily off-
target and time-consuming, limiting its application in cell immunotherapy. The CRISPR-Cas9-
OPT gene editing system that will be applied in this study is the third generation of gene editing 
system, following ZFN system and TALEN system. CRISPR (Clustered Regularly Interspaced 
Short Palindromic Repeat), in this system, consists of the exogenous DNA that is induced by 
approximately 20 spacers and repeated sequences at both ends, which is transcribed into guide 
RNAs and then forms a complex with CAS9 protein. The complex will recognize and bind to the 
specific sequences of DNA through guide RNAs, and then CAS9 will target and remove the DNA 
in the binding region [12]. Comparing with other gene editing technologies, the CRISPR-Cas9 is 
simple to manipulate and highly efficient in knocking out target genes, and can edit multiple 
genomic regions in the same cell [13]. These advantages have promoted the application of CRISPR-
Cas9 in clinical translational studies in recent years.  
In a cancer gene therapy study, MCL-1, which is a crucial gene for Burkitt lymphoma, can be 
specifically edited by the CRISPR-Cas9 system, leading to the inhibition of tumor growth [14]. 
Similar studies have also demonstrated that using CRISPR-Cas9 to edit E-cadherin in a bladder 
carcinoma cell line can inhibit the growth of the cancer cells [15].  
As for treating virus-related tumors, many in vitro studies have reported that the CRISPR-
Cas9 system can successfully and specifically edit the genome of human hepatitis B virus (HBV) 
and reduce the generation of the HBV antigens [16].  Human papilloma virus (HPV) can integrate 
its own DNA into the host genome, highly express of the virus proto-oncogenes E6 and E7, and 
then induce cervical cancer. Several studies have shown that an editing system that targets E6 and 
E7 genes can be established using CRISPR-Cas9 and that this can be used to inhibit the 
proliferation and tumorigenic ability of HPV16-infected SiHa cells [17].  
Basic research in cancer immunotherapy has demonstrated that using CRISPR-Cas9 to knock 
out the PDCD1 gene (encoding the PD-1 protein) can decrease the PD-1 expression on T cells and 
increase the secretion of IFN-γ [18]. The anti-tumor ability of chimeric antigen receptor T cell 
immunotherapy (CAR-T) cells is significantly improved after using CRISPR-Cas9 technology, 
which combines immune checkpoint blockade therapy with CAR-T, to further knock out the 
expression of PD-1 in CD19 CAR-T cells in in vitro experiments. Additionally, this has been 
demonstrated in experimental animals with PD-1 positive expression [19]. In June 2016, the 
recombinant DNA advisory committee (RAC) of the national institutes of health (NIH) first 


 
 
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approved a phase I clinical trial led by Professor Carl June from the University of Pennsylvania. 
Initially, the investigators plan to transfect NY-ESO-1 into T cells that have been isolated from 
patients with myeloma, sarcoma, and melanoma. Secondly, CRISPR will be used to knock out the 
PDCD1 gene in these T cells. They hypothesize that it will bring a beneficial effect by combining 
these two gene-editing approaches.  
Cell therapy in cancer hasalso evoluted rapidly in recent years. Clinical studies using CAR-T 
for treatment of malignant hematological tumors, such as acute lymphoblatic leukemia, and 
lymphoma such as diffuse large B cell lymphoma, indolent B cell lymphoma and acute 
lymphoblastic lymphoma, have demonstrated the safety and efficacy of this type of therapy [20, 21]. 
For solid tumors, clinical trials for the application of CAR-T in neuroblastoma, metastatic renal 
cell carcinoma, and ovarian cancer have been reported and demonstrated its safety and feasibility 
in clinical application [22-25].  
Since September 2013, we have used the CRISPR-Cas9-OPT optimized platform to 
specifically knock out the PD-1 gene and the immune checkpoint gene CTLA-4 in T cells, to 
improve the function of T cells. Our preliminary data from in vivo and in vitro experiments suggest 
that: (1) the sgRNAs can efficiently knock out PD-1 and CTLA-4; (2) it is feasible to knock-out 
PD-1 in T cells; (3) there is a significant improvement in the secretion of tumor killing cytokine 
from T cells after the knock-out of PD-1. These results suggest applying this optimized system to 
target PD-1 on T cells in cancer immunotherapy. Due to the ease of manipulation and development, 
and the low cost, this therapy shows the potential for broad application.  
1.3 The study technology and work basis 
1.3.1 Study-related technology 
This novel project has been co-developed by the Thoracic Oncology Department of the West 
China Hospital at the Sichuan University and MedGenCell Biotech Co., Ltd. MedGenCell Biotech 
Co., Ltd. is a biotech company located in the Tianfu Life Science Park (Chengdu, Sichuan) and 
provides biotech development, technology consultation, technology promotion, clinical trials, and 
technical support.  
The CRISP-Case9 vector has been established, which knocks out PD-1 in healthy human T 
cells with a high efficiency (ranging from 40%-90%). The cytokine secretion was increased in the 
PD-1 knock-out T cells in vitro. The enyme-linked immunoSpot (ELISPOT) test showed that the 
function of the specifically activated T cells was enhanced following PD-1 knock-out.  
There were two major challenges of genome editing technology: (1) the low efficiency of 
gene knockout; (2) the potential off-target toxicity caused by CRISPR-Cas. The team collaborating 
on this project has been working on the optimization of the Cas9 system since July 2012. The 


 
 
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works including not limiting are as followed:  
1. Optimization of the CRISPR-Cas9 system for the knockout of mammal genes. This system 
that uses CRISPR-Cas9 to establish a gene knockout mouse model through pronuclear injection. 
An application for the patent has been filed (‘A eukaryotic gene targeting method and helical 
structural DNA sequence without species limit or bio-safety problem’, Application 
No.201310028668.2), and the related article was published in Cell Research[26]. This optimization 
laid the foundation for all genome manipulation in different animals through the use of the Cas9 
system via pronuclear injection.  
2. Multiple gene knock-outs have been accomplished in mice and rats through pronuclear 
injection. We performed gene manipulation in mice at 10 points of five target genes by injecting 
10 sgRNAs at the same time. These 10 gene regions were all manipulated and detected in the 
founders. This operation enables the knock-out of 10 genes in rats or mice simultaneously. This 
has several advantages: first, it reduces the working time and cost of experiments; second, it 
simplifies the process of knocking out multiple genes, which has previously been hard to perform. 
As human diseases are usually caused by multiple gene alterations, the simultaneous manipulation 
of multiple genes will be more attractive. Based on the above-mentioned research, we performed 
the conditional gene knock-out in rat by using the CRISPR-Cas9 system. After the conditional 
knock-outloxP template was introduced, the homologous recombination was mediated by 
CRISPR-Cas9. The articles reporting this research were published inCell Research[27].  
3. Due to its high efficiency, CRISPR-Cas9 causes an off-target effect, which limits its 
application. Through collaboration with the Sanger institute, we successfully reduced the off-target 
effect to an undetectable level by combinating mutant Cas9 with double sgRNA technology. This 
was published inNature Methods[28].  
4. By cooperating with domestic investigators, we performed monkey gene knock-out 
through pronuclear injection. This research work was published in Cell[29].  
1.3.2 Pre-clinical data18:  
(1) The identification of sgRNAs that efficiently knockout PD-1. 


 
 
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Figure 1. The knockout efficiency of different sgRNAs targeting PD-1 was tested in HeLa cell line.  
(2) Electro-transfer efficiency of T cells. 
 
Figure 2. Representative images of the transfected cell by light microscope and fluorescence microscope. The 
positive infected cells are observed under fluorescence microscope. Results: Qualitative detection of GFP by 
fluorescence microscope72 h after electrotransfection was performed to evaluatetransfection efficiency.The 
outcome from flow cytometry revealed that the transfection efficiency was 35-40%. 


 
 
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(3) Electrotransfection of the T cells from three unscreened donors.  
 
Figure 3. Electrotransfection of the T cells from three unscreened donors. There was discrepancy in the 
electrotransfection efficiency among different individuals. Without screening, the electrotransfection efficiency 
was 30-50%; under external screening, the efficiency was 70-95%; the QC standard for the transfusion product 
is above 10%.  
(4) The knock-out of PD-1 has been performed in T cells, and the capability of cytokine secretion 
of T cells has been significantly improved. 
 
Figure 4. The knock-out of PD-1 in T cells, and the capability of cytokine secretion of T cells has been 
significantly improved. The changes in the cytokine secretion of the T cells that were isolated from patients' 
ascites and electrotransfected with Cas9/sgRNA were detected by Ellispot.  
(5) The cytotoxicity test proved that PD-1 KO T cells had higher efficiency in targeted tumor cell 
killing than the common CIK cells.  


 
 
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Figure 5. The cytotoxicity tests. PD-1 KO T cells showed higher efficiency in the targeting of tumor cells than 
the common CIK cells and the NT cells.  
(6) The in vivo toxicity of PD-1 KO T cells in NSG mice.  
Group  
N  
Accidental deaths  Pathologic change a GVHD b 
WT T cells  
5  
0/5  
0/5  
0/5  
PD-1 knockout T cells  5  
0/5  
0/5  
0/5  
a: On the 30th day after cell transfusion, the mice were sacrificed. H&E staining of the heart, liver, kidney, 
and lung tissues were performed. The Giemsa-Wright staining of bone marrow smear was also performed to 
check the infiltration of the T cells. 
b: The wrinkled skin, body weight changes, and activity changes of the mice were observed.  
 
Figure 7. The in vivo toxicity test results. Images were captured using a 10X magnification. 
Since September 2013, we have utilized the CRISPR-Cas9 system and completed the 
following research: 1. sgRNAs that can highly efficiently knock out PD-1 and CTLA-4 have been 


 
 
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identified; 2. The PD-1 knock-out has been performed on T cells, leading to a significant 
improvement in the cytokine secretion of the engineered T cells. These results demonstrate the 
feasibility of the use of the CRISPR-Cas9 system to target T cell PD-1 in cancer immunotherapy 
for patients. Considering the simple operation and reasonable development expenses, this 
therapeutic method shows the potential for broad application.  
2 Study objectives 
2.1 Primary objective 
To investigate the safety and tolerability of CRISPR-Cas9 edited PD-1 T cell in patients with 
advanced NSCLC. 
2.2Secondary objectives 
1) To investigate the antitumor activity of CRISPR-Cas9 edited PD-1 T cells in patients with 
advanced NSCLC. 
2) To investigate the changes in the potential immunological indexes and biomarkers. 
3) To track the PD-1 edited T cells in patients and investigate the mechanism of the clinical 
antitumor activity. 
2.3 Primary endpoint 
The dose-limiting toxicity (DLT) rate of the cell therapy during the first treatment cycle. 
2.4 Secondary endpoint 
1) To evaluate the response rate (RR) and 8-week disease control rate (DCR) of the cell therapy.  
2) To evaluate time to first response, defined as the time from the first cell infusion to the first 
observed complete response (CR) or partial response (PR).  
3) To evaluate the duration of response (DOR), defined as the time from the first observed CR 
or PR to the first observed PD or death from any cause.  
4) To evaluate the progression free survival (PFS), defined as the time from the first cell infusion 
to the first observed PD or death from any cause. 
5) To evaluate the overall survival (OS), defined as the time from the first cell infusion to death.  
6) To evaluate the activation status and duration of PD-1 edited T cell after infusion.  
7) To evaluate the change of in vivo cytokine levels and their relation to the tumor response and 
adverse events of after PD-1 edited T cell infusion. 
3Study design 
3.1 Description of the study design 
This phase I clinical study aims to investigate the safety and tolerability of PD-1 edited T cell 
therapy in patients with advanced NSCLC.  


 
 
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About 20 patients with PD-L1 positive advancedNSCLC who have progressed after the 
multiple lines of prior standard therapeutic regimens are planned to be enrolled in this study. The 
safety and tolerability of the therapy will be investigated by using intergroup dose escalation 
method.  
This clinical study was a first-in-human trial using CRISPR technology knockouting PD-1 
gene in T cell to treat lung cancer patients. Although there were no exact dosage/regimen and safty 
data, starting dose was selected referring to cell therapy or CAR-T treatment for considering 
benefit-risk of cancer patients (see section 5.3.1 Starting dose rationale).  Pre-A: 2x107 PD-1 edited 
T cells per kilogram of body weight (percentage of PD-1 knockout T cell may vary depending on 
technology efficiency). The study defines the Pre-A group as a preliminary exploratory dose group. 
The Pre-A group has a 28 days-observation after the first treatment cycle. If no DLT was observed 
in Pre-A group during the defined observation period, the study will continue other treat groups 
according to provisional dosage; If DLT or other unexpected toxicity occurs in the Pre-A group, 
the investigator will adjust the cell dose and interval of sequential cell therapy based on data from 
Pre-A group. 
There was three treatment groups and 28 days/cycle. 
Group A: A dose of 1×107 PD-1 edited T cells per kilogram of body weight splited into three 
doses of 20%, 30%, and 50% will be conducted on the 1st, 3rd, and 5th day of each cycle, 2 
treatment cycles;  
Group B: A dose of 2×107 PD-1 edited T cells per kilogram of body weight splited into three 
doses of 20%, 30%, and 50% will be conducted on the 1st, 3rd, and 5th day of each cycle, 2 
treatment cycles;  
Group C: A dose of 4×107 PD-1 edited T cells per kilogram of body weight splited into three 
doses of 20%, 30%, and 50% will be conducted on the 1st, 3rd, and 5th day of each cycle, 2 
treatment cycles. 
(Treatment may continue after 2 cycles if considered to be benefitial) 
Summary schema of the study design:  


 
 
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3.1.1 Molecular screening 
All of the patients must sign the ICFs for molecular screening. Please find more information 
in section 7.1.1.  
All of the patients enrolled in this study should be confirmed as PD-L1 positive (TPS ≥ 1%) 
by immunohistochemical staining.  
The main standard procedure for detecting PD-L1 is as following: 
 
After the tumor being confirmed as PD-L1positive, the patient can sign the study main ICF, 
and start the screening/baseline visit. 
 Remarks: If the patients have already been confirmed to bePD-L1positive through staining 
with Roche Ventana PD-L1 (SP-142, SP-263) and Dako PD-L1 (Dako28-8, Dako22c3) regent 
prior to this clinic study, they can directly sign the main study ICFs, and the screening/baseline 
visit can be initiated [30].  
3.1.2 Screening period 
The screening period is composed of the screening phase one and phase two. Molecular pre-
screening should be completed in the screening phase one, the manufactured T-cell expansion and 


 
 
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baseline evaluation will be performed in the screening phase two. The patients must sign the ICFs 
before blood collection and screening evaluation. The baseline evaluation must be performed 
within the four weeks before the first treatment cycle. Refer to 7.1.2 for additional information.  
3.1.3 Treatment period 
The enrolled patients will be divided into three treatment groups. The treatment will be 
discontinued if the treatment is finished or if any of the following conditions occur: progressive 
disease (PD) confirmed by the investigators (according to RECIST 1.1), unacceptable toxicities, 
pregnancy, end of the treatment bythe investigator or the patients. 
The safety will be monitored according to the scheduled follow-up plan (Table 7-1).  
For the imaging assessments of the tumor (such as CT and MRI scan), all of the image scans 
will be scheduled according to the study scheme from the beginning of this study. The scan after 
the first treatment cycle will be performed at weeks 8 (+1) and 12 (±1). Afterward, the tumor 
should be assessed every 8 (±1) weeks until the disease progresses, the patient dies, the patient is 
lost to follow-up, or the patient withdraws the ICF.  
The evaluation of tumor responseshould follow the RECIST 1.1guideline in Appendix 12.2; 
refer to the irRC in Section 12.3.  
3.1.4 End of treatment 
The end of treatment (EOT) visit will be arranged for patients within the seven days following 
the confirmation of PD or the discontinuation of treatment due to any other reasons (table 7-1).  
3.1.5Follow-upperiod 
All patients will be followed up every two months within the 24-month period following the 
last treatment cycle, to assess the safety (AE or SAE).  
3.1.6 Survival follow-upperiod 
Once a patient completes the treatment and/or the following tumor assessment, he/she will be 
followed up every two months to check the survival until the patient is dead, lost to follow-up, or 
withdraws the consent for survival follow-ups, or the study is completed (refer to the definition of 
EOT in 3.2). The survival follow-up will be conducted through phone calls.  
3.2 Definition of the end of study 
The end of study (EOS) was defined as the sixth month after the last enrolled patient finishes 
the last treatment.  
3.3 Early termination of the study 
The investigators can terminate the study at any time. If the study has to be terminated early, 
a patient visit should be arranged as soon as possible, and the assessment for early study 
termination that is listed in Section 7.1.6 should be performed. To protect the benefit of the patients, 


 
 
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the investigators should inform the patients of other procedures which they should comply with. 
The investigators are responsible for reporting to the ethics committee and the relevant 
departments regarding the early termination of this study.  
4Population 
Advanced NSCLC patients with positive PD-L1 expression (TPS ≥ 1%) who previously failed 
at least three-lines of treatments were enrolled. 
4.1 Inclusion criteria 
1. Informed consent form (ICF) is signed:  
(1) The enrolled patients must sign and date the EC-approved written ICFs. 
 according to the guidelines of the competent authority and study institution. The ICFs should be 
signed before implementing any protocol-related procedure (not including the routine medical care 
of the patients).  
(2) The enrolled patients must be willing and able to comply with the visits, therapeutic regimen, 
and laboratory examinations specified in the schedule as well as the other requirements in the study.  
2. Age: 18-70 years.   
3. ECOG performance status 0-2. 
4. NSCLC, stage IIIB/IV confirmed by histology or cytology.   
All patients must be screened for EGFR gene mutation and ALK gene fusion; patients with positive 
EGFR mutation or ALK gene fusion treatment failure of EGFR-TKI or ALK-TKI must have been 
experienced. 
5. Patients must fail from the standard third-line or above therapy scheme. 
6. Patients must be positive for PD-L1staining, as tested through tumor biopsy (TPS ≥ 1%).  
7. Patients must have a measurable lesion (tumor lesion with a long diameter of more than 10 mm 
through CT scan; lymph node with a short diameter of more than 15 mm through CT scan) in 
accordance with the RECIST 1.1. 
8. Life expectency of ≥ 3 months. 
9. Adequate bone marrow function:  
• WBC ≥ 3.0×109/L. 
• ANC ≥ 1.5 ×109/L.  
• PLT ≥ 90×109/L, without transfusions within 14 days before the first cell therapy. 
•  Hemoglobin ≥ 10.0 g/dL. 
10.  Adequate hepatic and renal function:  
Total bilirubin ≤ 1.5×ULN.  
• AST and ALT < 2.5×ULN; AST and ALT < 5×ULN if metastasis to liver.  


 
 
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• Serum creatinine ≤1.5×ULN, or creatinine clearance rate ≥ 50 ml/min (based on the 
Cockcroft/Gault formula) . 
11. Coagulation functions:  
•  INR ≤ 1.5×ULN.  
•  PTT ≤ 1.5×ULN. 
12. Adequate cardiovascular function:  
•  Left ventricular ejection fraction ≥ 50%.  
    •  QTcF interval ≤ 450 ms.  
13. ALP ≤ 2.5×ULN.  
14.Recovery from all adverse events of previous systemic anti-cancer therapies to baseline or 
Grade ≤ 1, except for Alopecia.  
15. Washout period for the previous anti-tumor treatment is not less than four weeks and washout 
period for the molecular targeting drug is not less than five half-life periods. Palliative radiotherapy 
must have been completed for at least two weeks; thoracic radiation therapy must have been 
completed for at least three months, and major surgery must have been completed for at least four 
weeks with recovery.  
16. The patients must take contraception measures strictly.  
• Within the 72-hour period prior to the therapy, the serum or urine pregnancy test (HCG sensitivity 
is 25 IU/L at least or equivalent) of the women of childbearing potential (WOCBP) must be 
negative.  
• WOCBP must agree to take contraceptive measures strictly during the treatment period and for 
at least 18 months after the final cell infusion until no knockout T cell is detected in the blood.  
•  The males who have sexual contact with WOCBP must agree to take contraceptive measures 
strictly during the treatment period and for at least 18 months after the final cell infusion until no 
knockout T cell is detected in the blood. 
4.2 Exclusion criteria 
1. Patients with active central nervous system (CNS) metastasis (including but not limited to 
carcinomatous meningitis and spinal cord compression) must be excluded. However,patients with 
metastatic CNS tumors may participate in this study if thepatients can be recoved to the baseline 
level at least two weeks before inclusion (without the residual signs or symptoms related to the 
CNS treatment).  In addition, the patients must have stopped using corticosteroids for four weeks 
before inclusion.  
2. Patients with weight loss in the recent six months ≥ 10%.  
3. Patients accompanied by emergent symptoms caused by tumor that need immediate intervention.  


 
 
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4. Patients with medical history of interstitial lung disease (ILD) or immune related pneumonia.  
5.  Patients with peripheral neuropathy. 
6. Patients with clinically significant coagulation abnormalities, or patients who are receiving 
thrombolytic or anticoagulant therapy.  
7. Patients with a history of human immunodeficiency virus (HIV) positivity or with the acquired 
immunodeficiency syndrome (AIDS), or with active hepatitis B or hepatitis C. 
8. Patients that have other malignancy.  
9. Patients with active, known or suspected autoimmune diseases.  
10. Patients who have received systemic treatment by corticosteroids (the equivalent dose >10 mg 
prednisone/day) or other immunomodulators (interleukin-2, IFN-α, IFN-γ, cyclosporine, G-CSF, 
mTOR inhibitor) within the 28 days prior to the study. Patients who receive inhaled or topical 
corticosteroids and adrenal corticosteroids (the equivalent treating dose <10 mg prednisone/ day) 
can participate in the study if they are lack of active autoimmune diseases.  
11. Clinically significant cardiac disease or impaired cardiac function, such as: 
•  Clinically significant heart disease such as CHF requiring treatment, or uncontrolled 
hypertension defined by blood pressure > 140/100 mmHg at rest (taking mean of 3 consecutive 
readings). 
•  History (< 6 months prior to screening) or current evidence ofclinically significant cardiac 
arrhythmias, atrial fibrillation and/orconduction abnormality, e.g. congenital long QT syndrome, 
Grade ≥2/complete AV-blockage or hypokaliemia CTCAE Grade ≥ 3. 
• History/evidence of acute coronary syndromes (including myocardialinfarction, unstable 
angina, coronary artery bypass graft (CABG),coronary angioplasty, or stenting), < 6 months prior 
to screening. 
• Complete left bundle branch block (CLBBB).  
• Right bundle branch block (RBBB) + left anterior hemiblock (LAHB - bifascicular block).   
12. Patients with a history of allergies to cyclophosphamide and other agents involved in this study. 
13. Patients that do not have enough venous access.  
14. Pregnant or lactating women, where pregnancy is defined as the state of a female from 
conception until the termination of gestation.  
15. Patients with mental disorders are mandatorily detained for treatment or are judged as 
inappropriate for inclusion by the investigator.  
16. Patients who are difficult to communicate with or difficult to be followed up for a long time.  
17. Other inappropriate conditions in discretion of the investigator’s judgment. 
5. Treatment 


 
 
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5.1 Cell therapy protocol 
5.1.1 Pre-treatment 
A single dose (20 mg/kg) of cyclophosphamide will be administrated as a pre-treatment for 
lymphocytes depletion on the third day before the cell transfusion of the first treatment cycle. 
Cyclophosphamide will be diluted with 20-30 ml of normal saline (NS) and intravenously injected 
for more than 10 min.  
5.1.2 Cell infusion 
5.1.2.1 Treatment assignment 
Pre-A: 2x107 PD-1 edited T cells per kilogram of body weight (percentage of PD-1 knockout 
T cell may vary depending on technology efficiency). The study defines the Pre-A group as a 
preliminary exploratory dose group. The Pre-A group has a 28 days-observation after the first 
treatment cycle. If no DLT was observed in Pre-A group during the defined observation period, the 
study will continue other treat groups according to provisional dosage; If DLT or other unexpected 
toxicity occurs in the Pre-A group, the investigator will adjust the cell dose and interval of 
sequential cell therapy based on data from Pre-A group; 
Group A: A dose of 1×107 PD-1 edited T cells per kilogram of body weight splited into three 
doses of 20%, 30%, and 50% will be conducted on the 1st, 3rd, and 5th day of each cycle, 2 
treatment cycles, 28 days/cycle;  
Group B: A dose of 1×107 PD-1 edited T cells per kilogram of body weight splited into three 
doses of 20%, 30%, and 50% will be conducted on the 1st, 3rd, and 5th day of each cycle, 2 
treatment cycles, 28 days/cycle;  
Group C: A dose of 1×107 PD-1 edited T cells per kilogram of body weight splited into three 
doses of 20%, 30%, and 50% will be conducted on the 1st, 3rd, and 5th day of each cycle, 2 
treatment cycles, 28 days/cycle. 
     In this study, each treatment group has two treatment cycles. The safety and tumor 
progressionwill be evaluated when the patients have received two treatment cycles. If no DLT or 
PD is observed, the investigators willdiscuss with the patientthe further therapeutic regimens based 
on the potential benefitial to the patient who is willing to continueto receive treatment: 
1) If patient decides not to continue treatment, the patient will be followed upaccording to the 
study workflow (7 visits and evaluation) until end of study. Additionally, the patient has the right 
to choose other cancer therapeutic strategies. 
2) If the patient agrees to continuecell therapy, she/he can receive treatment until end of study. 
According to the irRC criteria, if PD is identified through tumor imaging, another evaluation 
should be conducted after at least four weeks to confirm the PD status. During the period to 


 
 
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confirmation of the PD, if the patient’s performance status does not deviate from the baseline 
(ECOG：0-2 scores), or no intolerable therapeutic side effects/other conditions that are 
inappropraite for further cell infusion therapy occur, and the patient agrees to continue treatment, 
the patient may continue to receive the cell infusion treatment. If the next imaging test is scheduled 
within four weeks, it is not necessary for the patient to take the next appointed imaging test.  If the 
clinical conditions of the patient are stable, the tumor imaging test can be re-initiated at an 
appointed time point and continued until PD is confirmed through imaging assessment. Given that 
tumors may pseudoprogress within the first several months after the initial cell infusion therapy 
and then shrink in some patients, patients are allowed to continue the cell infusion therapy after 
the first presence of PD until confirmed. For the patients with unstable clinical conditions, it is not 
necessary to repeat the imaging examination to confirm PD. Tumor pseudoprogression includes: 
• 
Worsening of the original target lesion 
• 
Worsening of the original non-target lesion  
• 
Identification of new lesions 
Cells for each infusion treatment cycle should be derived from the newly collected blood. The 
total number of cells for three treatment groups is: 1x107, 2x107, and 4x107 PD-1 edited T cells per 
kilogram of body weight, respectively, and the total volume are 100 ml/time. 
5.1.2.2 Cell infusion procedure 
1. The vital signs, temperature and blood pressure should be tested and recorded within 30 min 
before the cell infusion. If any fever occurs (> 38.5°C), the cell infusion shall not be performed.  
2. Diphenhydramine will be intramuscularly injected within approximately 30 min before the 
cell infusionto prevent the cell infusion reaction.  
3. Use 0.9% normal saline (NS) to establish the intravenous line.  
4. Gently shake the cell bag 3-4 times to suspend the cells.  
5. Start infusion, shake the cell bag gently every 20 min during the infusion and slightly pinch 
the bottom of the cell bag several times to prevent cell aggregation.  
6. Cell infusion speed: during the first cell infusion to a patient, the initial speed should be 
adjusted to about 15-20 drops/min. The patient’s tolerability will be evaluated 10-15 min after the 
infusion. If no adverse event is observed in the patient, the infusion speedwill be adjusted to be 
reasonably faster, around 60 drops/min. For elderly patients and patients with cardio-pulmonary 
insufficiency, the infusion speed should be slowed downand close attention should be paidto the 
patient’s tolerability to the infusion speed. The speed of infusion can be adjusted according to the 
condition of the individual.  


 
 
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7. The NS will be administratedafter the cell transfusion. 
Remarks: The vital signs and symptoms of patient should be monitored within 30 min before 
and after the cell infusion and at the time when the cell infusion is finished.  
5.2 Concomitant treatment 
5.2.1 Acceptable concomitant treatment 
Recordall of the medicine and non-medicine treatment (includingphysical therapy, natural 
drugs, and blood transfusion) that is administrated to the patients within the four weeks before the 
treatment and during the treatment period.  
During the study, the patients will be allowed to use external, ophthalmic, intraarticular, intranasal 
and inhaled corticosteroids (at extremely low systemic inhaled doses).  
Patientes will be allowed to take corticosteroids to prevent (for instance, the allergy to contrast 
agent) or cure (for instance, delayed allergic reaction caused by contact allergen) non-autoimmune 
diseases in a short term (< 1 week).  
Combined palliative and supportive treatments are allowed to be used to treat the disease-
related symptoms (e.g. diphosphonate). During the cell infusion treatment, the use of RANK-L 
inhibitors (including, but not limited to, Xgeva) will not be allowed. The previous palliative 
radiotherapy should be finished at least two weeks before the cell therapy is commenced.  
5.2.2 Local palliative treatment 
Patients who have no evidence of clinical PD or imaging PD (according to RECIST 1.1) are 
allowed to receive local palliative treatments (including palliative radiotherapy or palliative 
surgical resection to the symptomatic non-target bone lesions, skin lesions or CNS lesions) before 
the treatment ends. Patients who have no evidence of clinical or imaging progression (according 
to RECIST 1.1), and for whom the lesion that causes hemoptysis is not the only measurable target 
lesion, will be allowed to receive the local palliative treatment to that target lesion after discussing 
with the principle investigator and getting permission.  
Before starting palliative therapies, the objective evidences of the PD of the patients should be 
evaluated, especially if the latest tumor response evaluation is conducted more than four 
weeksbefore the local palliativetreatment. If the second tumor progression is confirmedaccording 
to irRECIST before the local palliative treatment, the treatment of the patients will be stopped.  
It is currently unclear whether the toxicity that is induced by radiotherapy overlaps with the 
toxicity that is induced by the cell therapy. Because no data regarding the effects of combined 
radiotherapy and cell therapy has been reported, if the patient requires palliative 
radiotherapy(radiotherapy is not allowed in thoracic lesions) then the cell therapy should be 
stopped for at least one week before the radiotherapy. Cell therapy could be resumed at least one 


 
 
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week after the radiotherapy is completed. Close monitoring should be paid to the patients during 
and after the radiotherapy. Once the toxicities occur, the cell therapy should be continued only after 
AEs recover to Grade 1 and below, or to the baseline level or stable.  
5.2.3 Restricted treatment 
During the study, the investigators will judge whether the patients will benefit from the 
concomitant therapy and they will decide whether to apply it or not.The investigators will record 
this in the corresponding sections of the case report form.  
During the study, the following drugs are contraindicated (unless they are used to treat the 
drug-related AEs):  
1. Immunosuppressors. 
2. During the study, the prophylactic usage of recombinant human growth factors (such as G-
CSF and its analogues and hemopoietin-α and its analogues) is contraindicated. The use of 
prophylactic usage of recombinant human growth factors will be acceptable only if it really 
medically benefits the patients and the patient’s eligbility for further treatment will not be affected, 
as judged by the investigators. GM-CSF is contraindicated throughout the study process.  
3. High doses of systemic corticosteroid that will lead to immunosuppression (except for those 
specified in Section 5.1.5.1). 
4. Any antitumor medicine is contraindicated, including, but not limited to, chemotherapy, 
targeted drugs (Gefitinib, Tarceva, AZD9291 active ingredient, antitumor Chinese medicine, etc), 
hormonotherapy, immunotherapy (including but not limited to treatment of thymus gland), 
extensive nonpalliative radiotherapy, and investigated treatment for NSCLC).  
Except for the local palliative treatment specified in Section 5.2.2, any other radiotherapies or 
surgerical treatment of any tumor lesions are not allowed during this study. Patients who receive 
other non-palliative therapy will have to discontinue the study treatment.  
5.2.4 Treatment duration 
For proper planning and evaluation in this study, a treatment cycle is defined as 28 days. Patients 
will receive treatment following the protocol until PD (RECIST 1.1) or intolerable toxicity occurs, 
the termination decision is made by the investigator, the consent forms are withdrawed by the 
patient, or the patient dies.  
5.3. Dose escalation guidelines 
5.3.1 Starting dose rationale 
In 2010, the FDA approved the first therapeutic vaccine PROVENGE which is infusedat least 
5×107 cellsinto the patients per two weeks for three times. In addition, some clinical studies 
indicated that no treatment related adverse events above Grade 3 were observed and no related 


 
 
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lethal effect was observed after multiple myeloma patients received the receptor-engineered T cell 
therapy at an average dose of 2.4×109 (0.45-3.9×109) 31. Single infusion is mainly used in the 
chimeric antigen receptor engineered T cell (CAR-T) therapy with a commonly used dose of 
1×106/kg. A phase I clinical study indicated that no treatment related adverse events of Grade 3 or 
above were observed after the patients received the escalated CAR-T dosing (1×108, 1×109, 1×1010 
cells) 32. Anotherrecent study in NSCLC patients indicated that no serious adverse events were 
observed after the patients received the CAR-T infusion at an average dose of 0.97×107/kg (0.45 
to 1.09×107/kg) 33. Therefore, accordingly to thedose levels of DC-T cell therapy approved by the 
FDA in 2010 and the CAR-T cell therapy in the existing clinical studies, and considering the 
balance of potential treatment safety and benefits to decide the initial dose of this study.  
5.3.2 Provisional dose levels 
The initial dose of the exploratory group Pre-A was 2x107 PD-1 edited T cells per kilogram of 
body weight. This dose will be split into three applications (20%, 30%, and 50%, respectively).A 
cycle is 28-days, and one treatment cycles. Besides, the cell dose of the treatment group A/B/C 
will sequentially increase 1x107 to 2x107 and 4x107 PD-1 edited T cells per kilogram of body 
weight.  
5.3.3 Guidelines for dose escalation and determination 
If DLT occurs in more than 1/3 of cases in the treatment group, another three patients will be 
enrolled into this group. Further enrollement for this group will be stopped if DLT occurs in one 
more case, and the data will be updated and re-evaluated.  
To apply the rule of dose escalation, the investigators and related staff (including study 
physicians and statisticians) will evaluate the toxicity (including the AEs unrelated to DLT and 
laboratory test abnormality) and safety profile of the tested group. Only when the safety evulation 
of the tested group has been completed, another treatment group will be started.  
5.3.4 Intrapatient dose escalation 
The escalation dose is from 1x107 to 2x107 and 4x107 PD-1 edited T cells per kilogram of body 
weight in the three treatment groups A, B, and C. Engineered T-cells spliting into three applications 
(20%, 30% and 50%) will be conducted on the 1st, 3rd, and 5th day of each cycle. The patient must 
be able to tolerate the lowest dose, and the effect of the prior dose has beenassessed, the patient 
can receive the sequential treatments.  
5.3.5 Other dosing regimens 
N/A  
5.3.6 Definition of Dose Limiting Toxicity 
Unless otherwise specified, NCI CTCAE 4.03 will be applied to assess the toxicity. DLT is 


 
 
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defined as an AE or a laboratory abnormal value that is probabaly attributable to the T cell infusion 
within first 28 days after cell transfusion. An abnormal laboratory value can be attributed to the T 
cell infusion if it happens within one cycle after the first cell infusion (≤ 28 days) or occurs in 
thefirst cycle but is later confirmed in the second cycle, and meets any criterion of the following 
conditions:  
• Any adverse event that is ≥ Grade 3 and is confirmed to be or probably attributable to the 
cell therapy.  
• Any toxicity related to the cell therapy that requires systemic glucocorticoid treatment.  
• Autoimmune toxicity that is ≥ Grade 2.  
Treatment will be discontinued when a patient experiences DLT, and the follow-up will be 
initiated according to the descriptions in Table 7-1.  
Before patients enroll in a higher dose group, all of the AEs ≥ Grade 2 that have occurred in 
the the current dose level must be analyzed.  
5.4 Dose modification 
5.4.1 Criteria for cell therapy delay, reinitiation and discontinuation 
5.4.1.1 Criteria for delayed cell infusion 
1.  Any AEs are ≥Grade 2 that are related to the cell therapy, except the following events:  
• For any Grade 2 fatigue or non-clinically significant laboratory abnormality that is related 
to the cell therapy, it is not neccessary to delay the treatment;  
• If the baseline level of AST, ALT or total bilirubin is within Grade 1, and the patient 
presents Grade 2 after the treatment while does not receive the hormonotherapy. In this case, 
it is not neccessary to delay the treatment; 
• Any Grade 3 laboratory abnormality that is related to the cell therapy, except the following 
conditions:  
1)  Recovery from Grade 3 lymphopenia or leukopenia.  
2) Any amylase or lipase abnormality ≥ Grade 3, which is related to the cell therapy, without 
pancreatitis symptoms or clinical manifestations, do not require the therapy to be delayed 
but to require consultion of the major investigators. 
2. Any AE or laboratory abnormality that investigators consider it necessary to delay the therapy.  
3. If there are any clinical manifestations, the patient should be re-evaluated every week or more 
frequently, and therapy should be reinitiated when the patient conforms to resuming treatment 
criteria.  
4. For other conditions that may deteriorate the safety of the patient, the cell infusion can be 
delayed or stopped according to the evaluation of safety by investigators based on the actual 


 
 
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conditions.   
If the cell fusion therapy is postponed for more than 14 days, the patients will not reinitiate the 
treatment, unless otherwise specificied. If the reason for a delay of more than two weeks of therapy 
is not related to the cell therapy, continuation is acceptable upon the approval of the principle 
investigator. Before the treatment is resumed in the patients with more than two weeks of delay, it 
is necessary to consult the principle investigator and perform tumor evaluation according to the 
study protocol.  
5.4.1.2 Criteria of reinitiation of cell therapy 
After the cell therapy, related AEs are controlled to below Grade 1 level or baseline level, the 
patient can reinitiate cell therapy. The following conditions are listed:  
• Patients with Grade 2 fatigue can resume the therapy.  
• Patients with Grade 2 but not Grade 3 cell therapy related skin AE can undergo the therapy 
again. 
• Patients, whom with an abnormal baseline level of AST/ALT or total bilirubin ≤ Grade 1 
toxicity, has Grade 2 AST/ALT or total bilirubin right now, while the previous treatment delay 
was not caused by the Grade 2 AST/ALT or total bilirubin. In this case, the patients can resume 
the therapy. 
• Patients with continuous Grade 1 pneumonia can undergo further therapy after being 
approved through discussion.  
5.4.1.3 Criteria of discontinuation of cell therapy 
The cell therapy should be discontinued permanently in cases with the following conditions:  
• ≥ Grade 3 Cytokine release syndrome;  
• Any cell therapy-related condition ≥ Grade 2 that requires systemic glucocorticoid therapy, 
such as pneumonia, nervous system damage, diarrhea/colitis, liver damage, nervous system 
damage, impaired kidney function, symptomatic endocrinopathy, uveitis, ophthalmodynia or 
blurred vision;  
• Patients with cell treatment-related Grade 3-4 lung toxicity, diarrhea or colitis and ≥ Grade 
2 cell therapy related endocrinopathy will stop the treatment; 
• Any AE, laboratory abnormality or intercurrent illness that, the investigators judge, would 
lead to great clinical risk in the patient if the cell therapy was continued.  
5.4.2 Follow-Up for DLT and other toxicities 
If the treatment is stopped temporarily or permanently due to AE or laboratory abnormality, 
the patient should be followed up once a week for at least four weeks. After that, the patient should 
be followed up once every four weeks until the event disappears or becomes stable. If the cell 


 
 
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infusion is to be delayed more than 14 days beyond the next scheduled infusion day, patients should 
stop the treatment permanently and be followed up as described above. All AEs and SAEs related 
to the therapy should be followed up to 24 months after the end of study.  
Follow-up for other toxicity: if discontinued treatment due to AE or laboratory abnormality, 
the patient must be followed up at least once a week for four weeks, and then once every four 
weeks until the event disappears or becomes stable. 
5.4.3 Anticipated risk and safety considerations of the study treatment 
This study protocol provides dose modification criteria, DLT definition and detailed guidance 
for delay, reinitiation and discontinuation of the study treatment. The recommended supportive 
treatment and cancer immunotherapy guidelines for anticipated toxicities are provided in Appendix 
5.5, including the treatment for AEs (CRS, etc).  
5.5 Supportive treatment 
The cell therapy treated patients may, as recommended by the investigators, be treated with 
appropriate supportive therapies, based on the guidelines for immune-related advert events (irAE). 
5.5.1 Cytokine release syndrome (CRS) 
Cell therapy may cause the release of inflammatory cytokines into the blood as a cytokine 
release syndrome. The cytokines usually reach the peak level at 5-7 days after cell infusion. A 
serious CRS may result in acute respiratory distress syndrome, multiple organ failure, and even 
death.  
In this study, the biomarkers of CRS such as IL-6, IL-10, CRP, TNF-α, etc will be routinely 
monitored in each cycle at the time before cell infusion, 24 hours after cell infusion, and on the 7th 
day after the cell infusion. In case of clinical suspicion or consideration of the CRS, the patient 
will be intensively clinically monitored for body temperature, blood pressure, oxygen saturation 
and neurological disorders. The blood sampling for cytokine detection is also recommended.  
The patients with CRS will not present specific symptoms initially, but some non-specific 
symptoms of the respiratory, gastrointestinal, cardiovascular, renal, and neurological systems may 
be observed 34-36.  
Main clinical signs and symptoms of CRS: 
Organ system  
Signs and Symptoms 
Constitutional 
Fever ± rigors, malaise, fatigue, anorexia, myalgia, arthralgia, nausea, emesis, 
headache  
Skin 
Rash 
Gastrointestinal  Nausea, vomiting, diarrhea 


 
 
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Respiratory  
Tachypnea, hypoxia  
Cardiovascular  Tachycardia, widened pulse pressure, hypotension, increased cardiac output 
(early), possible diminished cardiac output (late)  
Coagulation 
Elevated D-dimer, hypofibrinogenemia ± bleeding  
Renal 
Azotemia  
Hepatic 
Transaminitis, hyperbilirubinernia 
Neurologic 
Headache, mental status changes, confusion, delirium, word 
finding difficulty or frank aphasia, hallucinations, tremor, 
dymetria, altered gait, seizures 
 
Supportive care and close monitoring without special treatment are recommened for Grade 1 
CRS and Grade 2 CRS without systemic complications. 
The main clinical symptoms of severe CRS are persistent fever, hypotension, hypoxia and 
neurological symptoms. Fever may start 24 hours after the cell infusion and last for about 10 days. 
CRP will change during the CRS, which usually significantly increases at 4-9 days following the 
cell infusion. CRP > 20mg/dl may indicate severe CRS. Severe CRS is life-threatening and must 
be identified and intervened as early as possible. The diagnostic criteria are as follows: 
• Continuous fever lasting at least 3 days;  
• The maximum fold increase of two cytokines is 75-fold or more, or the maximum fold 
increase of one cytokine is 250-fold or more;  
At least one symptom from clinical toxicity, such as hypotension (requiring treatment with at 
least one kind of pressor) or hypoxia (PO2 < 90%) or neurological symptoms (such as mental 
status changes, unresponsiveness, seizures). 
 
 
 
 
 
CTCAEclassi
fication  
Clinical manifestation  
Treatment  


 
 
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Vasopressors dose conversion (≥ 3 h infusion time is required for all doses)  
Norepinephrine  
Monotherapy  
≥ 20 ug / min  
Dopamine  
Monotherapy  
≥ 10 ug / kg / min  
Phenylephrine  
Monotherapy  
≥ 200 ug /min  
Epinephrine  
Monotherapy  
≥ 10 ug / min  
Use 
of 
antidiuretic 
hormone  
 
Antidiuretic 
hormone 
+ 
norepinephrine 
corresponds to ≥ 10 ug / min  
Grade 1  
Non-specific 
clinical 
symptoms. 
 
Fever, 
nausea, 
fatigue, 
headache, myalgias, malaise.  
 
Supportive treatment: Close monitoring: 
Cytokine, CRP, ESR, TNF-α: blood 
routine, biochemical test.  Pathogenic 
bacteria culture to evaluate infection. 
Treatment is necessary if a fever or 
granulocytopenia occurs. 
Grade 2  
Symptoms 
require and 
respond to 
moderate 
intervention  
Oxygen requirement of <40% 
orhypotension responsive to 
fluids or a low dose of one 
vasopressor (such as dopamine, 
norepinephrine) or Grade 2 
organ toxicity. 
Evaluate whether there are extensive co-
morbities or older age. 
If not: supporting treatment (pay close 
attention to the cardiac functions and 
function of other organsand the cytokine 
level)  
If yes: refer to grade 3-4 CRS  
Grade 3  
Symptoms 
require and 
respond to 
aggressive 
intervention 
Oxygen requirement ≥ 40% or 
hypotension requiring a high 
dose or multiple vasopressors 
or Grade 3 organ toxicity or 
grade 4 transaminitis. 
Viligant supporting treatment.  
Administration 
of 
tocilizumab* 
(4 
mg/kg). If no improvement in the 
patients’ condition is observed within 
24-48 hours, the above dose can be 
administrated again ± a glucocorticoid 
(the initial dose of methylprednisolone is 
2 mg/kg/day and can be regulated 
correspondingly). For patients with 
serious nervous system symptoms, 
dexamethasone can be considered (0.5 
mg/kg/day, single dose ≤ 10 mg).  
Grade 4  
Life-
threatening 
symptoms  
Requirement 
for 
ventilator 
support 
orGrade 
4 
organ 
toxicity 
(excluding 
transaminitis). 


 
 
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Use 
of 
combined 
vasopressors  
Non-antidiuretic 
hormone  
Norepinephrine equals to ≥ 20 ug / min  
Norepinephrine equivalent dose = [norepinephrine equivalent dose (ug / min)] + [dopamine (ug / kg / min) ÷ 2] 
+ [phenylephrine (ug / min)] + [epinephrine (Ug / min) ÷ 10]  
 
5.5.2 Treatment transfusion/allergic reaction related to cell therapy 
Transfusion reaction and allergic reaction may occur in the process of cell infusion, including 
chills and fever (usually about 38C or over 40C in serious cases), accompanied with headache, 
nausea, emesis, pruritus, urticaria, fast pulse, general malaise, arthralgia, hypotension, 
hypertension, bronchospasm and other allergic reactions. In serious cases, it may cause 
angioneurotic edema, laryngismus, asthma, anaphylactic shock, heart failure and pulmonary 
edema.  
The transfusion/allergic reaction occur quickly in general and is potentially life threatening for 
critical ill patients.  These patients shall be closely monitored clinically and treated promptly and 
properly.  
Some patients may suffer from delayed allergic reactions, which occur within one week of the 
cell infusion. An allergic reaction is characterized by local or systemic pruritus, rash and other 
signs. The treatment for these symptomes includes oral antihistamine and/or glucocorticoid. 
Measurements for treating transfusion/allergic reactions 
Transfusion/allergic 
reaction classification  
Treatment  
Follow-up  
Grade 1 symptoms:  
Slight, temporary reaction; 
discontinuation of infusion 
is not needed; no need for 
treatment.  
Stay at the bedside.  
Slow down the infusion rate.  
Closely monitor the patient's 
symptoms, consciousness, 
blood pressure, breathing, 
pulse, urine output, etc., until 
the symptoms are remitted.  
Medication prevention: It is 
recommended to use appropriate 
drugs 30 minutes prior to treatment 
for the prevention of the recurrence 
of the transfusion reaction when 
further cell infusions are conducted, 
such as: Diphenhydramine at 50 
mg, intramuscular injection (or 
other similar drugs), acetaminophen 
at 325-1000 mg (or other similar 
drugs), etc.  
 
Grade 2 symptoms:  
treatment or discontinuation 
of infusion is needed, but 
Stop the cell infusion 
immediately  
Replace infusion fluid and 
If the patient's symptoms remit 
within a short time: restart cell 
infusion at 50% of the original 


 
 
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symptomatic treatment 
(antihistamine NSAIDAS, 
narcotics, infusion 
treatment) has a rapid effect; 
prevention of medication ≤ 
24 hr.  
 
conduct intravenous infusion 
with normal saline. Use 
diphenhydramine 50 mg (or an 
equivalent dose of a similar 
drug) for intravenous infusion; 
acetaminophen 325-1000 mg; 
proper use of glucocorticoid 
and/or bronchodilator; stay at 
the bedside and closely 
observe the patient's 
symptoms, consciousness, 
blood pressure, breathing, 
pulse, urine output, etc. until 
symptom remission. If 
necessary, conduct on-site 
physical storage, blood 
sampling (blood routine test, 
blood culture, etc.).  
infusion speed and if there is no 
notable discomfort after 30 
minutes, speed up the infusion rate, 
and closely monitor, or resume the 
original speed  
Medication prevention: when 
another cell infusion is conducted, 
it is recommended to reventively 
use the following drugs 30 min in 
advance of treatment: 
diphenhydramine, acetaminophen, 
etc. Use glucocorticoids for 
prevention when necessary  
If ≥ Grade 2 transfusion/allergic 
reaction reoccur, terminate cell 
therapy. 
Grade 3-4 symptoms  
Grade 3: delay in symptom 
remission (such as failure to 
respond quickly to 
symptomatic treatment 
and/or discontinuation of 
infusion); relapse of 
improved symptoms; 
hospitalization needed for 
sequelae.  
Grade 4: life-threatening; 
emergency treatment needed  
 
Stop the cell infusion 
immediately  
Change infusion liquid and 
tubes; conduct intravenous 
infusion of normal saline; 
proper use of bronchodilators 
is recommended: including but 
not limited to: epinephrine 0.2-
1 mg (diluted to 1:1000) 
through subcutaneous 
injection or epinephrine 0.1-
0.25 mg (diluted to 1:10,000) 
through slow intravenous 
injection; diphenhydramine 50 
mg + methylprednisolone 100 
mg or an equivalent dose of 
other glucocorticoids through 
intravenous infusion. 
Cell therapy is permanently 
terminated  
Stay at the bedside and closely 
monitor the patient until the 
symptoms completely remit and 
condition is stable.  
If necessary, conduct on-site 
physical therapy, check blood 
routine and biochemical indexes, 
etiological culturing (such as: the 
liquid being infused, blood, etc.).  
Continue follow-up: Some patients 
may suffer from a delayed allergic 
reaction within one week of the cell 
infusion, including expression local 
or systemic pruritus, rash and 
others symptoms. For these 
symptoms, such symptomatic 
treatment as oral antihistamine drug 
and/or glucocorticoid will be 
provided.  


 
 
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6. Patient numbering, treatment assignment or randomization 
6.1 Patient numbering 
In this study, the subject ID is used to identify each patient when the patient is first enrolled for 
molecular screening, and is used as the major identification of the patient throughout the entire 
study. The format of the subject ID is as follows: 00101, 00102, and 00110. Each patient has a 
unique subject ID in the database. Once a subject ID is assigned to one patient, it can not be used 
again. 
6.2 Treatment assignment or randomization 
According to the study design, the patients will be assigned into a specified treatment group 
without randomization.  
6.3 Treatment blinding 
N/A. This is an open-label, phase I study.  
6.4 Preparations for cell therapy 
The investigators or the responsible staff members at the study center must inform the patients 
that the study treatment needs to be performed according to the study protocol. Only the authorized 
staffs in the study center can perform the cell transfusion for the patients. The conditions of the 
patients must be recoded in detail throughout the whole study.  
Table 6-4.  Preparation for cell therapy  
Therapy  
Application  
Preparations  
Cell 
therapy  
In this study, cell infusion must be performed by 
the authorized staff.  
See Section 5.1.2.1 for detailed 
cell transfusion procedure. 
6.4.1 Package and labeling of the study cells 
The package label of the study cells is displayed as below. The label is produced in duplicate 
and one label should be taken from the cell storage bag and filed in the study document.  
Patient No.:                          Cell type:  
Cell quantity:                       Capacity:  
Storage temperature:           Storage condition:  
Date of manufacture:   MM/DD/YY   00:00  
Expiration date:  
Manufacturer:  
Study ID: MHC-001 
6.4.2 Cell supply and storage 
The cells should be maintained between 2C to 8C during transportation and storage in the 


 
 
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laboratory and the study center. The study center must appoint an authorized staff member to 
receive, handle and store the cells for the study treatment. The cells must be stored safely and 
properly in a location that only the investigators and the appointed staff members at the study 
center have access to. On the reception of the cells, they should be transfused into the patients 
according to the cell therapy requirements (Section 5.1.2.1).  
6.4.3 Cell product compliance and records 
6.4.3.1 Compliance 
During every patient visit, the investigators and/or the study staff members will evaluate the 
compliance of the patient, and record the information on the original document.  
6.4.3.2 Record of the study cell therapy 
In the study treatment, the investigators or the authorized representative will record the 
distribution and usage of the cells accurately after the cells arrive at the study center. The monitors 
will review all of the record forms of the cell therapy during the visit to the center and at the 
completion time of the cell therapy.  
At the end of the study, or at any designated time during the study, the investigators will submit 
all unused records, used study cell therapy supplies, labels and copies of the completed cell therapy 
record forms to the monitoring party for review.  
6.4.3.3 Disposal and destruction 
The used cell packages should be destroyed as medical waste in the study center. If the cells 
are not completely infused for some special reason, the residual cells should be sealed, packed and 
frozen/refrigerated for storage strictly according to the operation standards, and further disposal 
conditions should be recorded clearly.   
7 Visit schedule and assessments 
7.1 Visit schedule 
All of the visits are listed in the following Table (Table 7-1). If the corresponding evaluation is 
performed during the visit, it should be marked with an X. All data that are generated from these 
evaluations must be supported by the original documents of the patients. At any time during the 
study, additional assessment can be performed according to the clinical symptoms. Especially for 
the clinically significant changes, additional laboratory test should be performed during follow-
ups, based on the investigators' judgement. The patients in each dose group should be hospitalized 
for observation during days 1 to 14. Before discharge, the patient should be evaluated and the 
safety of the patient should be confirmed.  
If it is an unscheduled evaluation, these results must be recorded as “an unscheduled evaluation” 
in the eCRF.  


39 
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Table 7-1 Flowchart of Study Visits:  
Study procedure 
Screening 1 
Screening 2 
Treatment  
Cell therapy completion 
Follow-up for safety 
(every 2 month) j 
End 
of 
treatment 
Follow-up 
Molecular 
screening 
28 days before drug 
administration 
C1D1 C1D8 
≥ C2D1 ≥ C2D8 Cell 
therapy 
completion 
28 
days 
after 
completion 
Sign the ICF 
X 
X 
 
 
 
 
 
 
 
 
 
PD-L1 status test l 
X 
 
 
 
 
 
 
 
 
 
 
Collection of fresh tumor 
tissue samplesm 
 
X 
 
 
 
 
 
 
 
 
 
Blood culture 
 
X 
 
 
 
 
 
 
 
 
 
Inclusion/ exclusion criteria  
X 
 
 
 
 
 
 
 
 
 
 Medical 
history/smoking 
history 
 
X 
 
 
 
 
 
 
 
 
 
Demographic data 
 
X 
 
 
 
 
 
 
 
 
 
Vital signs 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
Physical examination, body 
weight 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
Height 
 
X 
X b 
ECOG score 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
12 leads ECG 
 
X 
X 
 
X 
 
X 
X 
X 
X 
 
Heart Doppler ultrasound 
 
X 
X c 
 
Hematology/blood 
biochemistry/ coagulation 
profile/urine test 
 
X 
X a 
 
X 
X 
X 
X 
X 
X 
 
Cardiac markers 
 
X 
X c 
 
Thyroid function test 
 
X 
X a 
 
X 
 
X 
X 
X 
 
 


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Gene Edited T-Cell Therapy for Advanced Lung Cancer. Version 1.9  
 
 
 
 
Tumor markers 
 
X 
X a 
 
X 
 
X 
X 
X 
 
 
HIV/HBV/HCV 
 
X 
 
 
 
 
 
 
 
 
 
Stool routine test 
 
X 
X a 
 
X 
 
X 
X 
X 
X 
 
Cytokine level, erythrocyte 
sedimentation rate (ESR) 
 
X 
X a 
X e 
 
 
 
 
 
 
 
Lymphocyte subset 
distribution 
 
X 
X a 
X 
X 
X 
X 
X 
X 
X 
 
Pregnancy test f 
 
X 
X 
 
X 
 
X 
 
 
X 
 
Collection of blood samples  
X 
X d The details are seen in 7.2.3 Biomarkers. 
 
Tumor evaluation i 
 
X 
 
 
X 
 
X 
 
 
X 
 
blood collectedh  
 
X 
 
X 
 
X 
 
 
 
 
 
Cell therapy g 
 
 
X 
 
X 
 
 
 
 
 
 
AEs 
 
Continuously monitored during the study 
 
Combination medication 
 
Continuously monitored during the study 
 
Follow-up survival k 
 
 
 
 
 
 
 
 
 
 
X 


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Amendment of the clinical research proposal: Vision 1.9 
Confidential 
Protocol No.: MHC-001 
 
 
Note: Each evaluation should be completed within seven days before the treatment. In the major treatment period, one treatment course (cycle) is defined as 28 days. All safety evaluation should be 
completed before the treatment. D: day. C: cycle. 
a. If no PD or clinical indication, which prompts further assessment, is found in a patient in the screening assessment within seven days before the study treatment, there is no need to perform 
further blood biochemistry testing, hematology testing, coagulation profiling, thyroid function tesing, tumor marker testing, cytokine level testing, urine routine testing, stool routine testing or ECG 
examinations.  
b. Body weight measurement. The patient’s body weight should be measured on the screening/baseline visit day, the first day of the first cycle and the first day of the following cycles. If the baseline 
evaluation is performed within 72 hours before the first cell therapy, the body weight will not be measured again on the first day of the first cycle.  
c. The baseline visit evaluation should be performed. In case of any abnormality, additional examinations should be performed.  
d. The detailed blood sampling is displayed in Section 7.2.3. Biomarkers, including:ctDNA andblood samplingfor diverse T lymphocyte receptors diversity.  
e. Cytokine level detection at: 24h after each transfusion, the seventh day after the final infusion in each cycle; if cytokine storm is suspected in clinic, the number of tests can be increased according 
to the clinical guidelines. Refer to Section 7.2.2.5 for details.  
f. For all WOCBPs, the result of serum pregnancy test obtained within 72 hours before the study treatment should be negative.  
g. Cell therapy: the cell therapy is performed on the day 1, 3 and 5 of each cycle.   
h. Blood collected: The blood for amplifying the cells for further treatment cycles should be collected within three weeks before the treatment and on the 8th day after the prior treatment cycle. 
i. The assessment results that are determined by imaging during the screening period are valid within 28 days before the treatment. The RECIST 1.1 criteria will be used for assessmentduring the 
screening (by using chest + abdomen CT or MRI scans); meanwhile, bone scintigraphy/bone scan and brain scan/head CT/MRI are required.  Further scans should be performed according to the 
schedule, calculated from the completion day of the treatment (± 7 days).  If any brain metastasis occurs in the patient, a brain imaging examination must be performed in the following tumor 
evaluations. If any bone metastasis occurs at the time of baseline assessment, a bone scintigraphy/bone scan is further required in these patients every six months (± 7 days), to monitor new lesions. 
If the radiologic assessment can evaluate the bone metastasis lesion, then a further bone scintigraphy/bone scan is not required. For other conditions, corresponding examinations should be performed 
according to the clinical indications. All of the evaluations must be completed before the next treatment. If the imaging evaluation has been performed within four weeks of the treatment completion, 
there is no need for re-examination. The tumor evaluation should be performed in the eighth (or ninth), twelfth week and every eight weeks during the following period (refer to Section 13.2 for 
details).  
j. After completion of the cell therapy, patients should be followed up for safety every two months.  
k. After PD or withdrawal from the treatment, patients should be followed up every two months to collect the survival information, including anti-tumor therapy.  
l.  PD-L1 status test was used by fresh tissue or paraffin-embedded tissue. 
m. The investigators could collect unscheduled tissue sample based on clinical situations.


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7.1.1 Molecular screening 
The study center is required to perform a molecular screen to the patients for the PD-
L1 expression positivity. The molecular screening will be performed in the Pathology 
Department of the study center using the Leica bondmax automatic immunohistochemistry 
machine. PD-L1 status detection was used by fresh tissue or paraffin-embedded tissue. 
Before the molecular screening, the candidate patients are asked to sign the ICFs for 
molecular screening.  
The details of the PD-L1 status are displayed in Section 7.2.3.3.  
After confirmation that the tumor is PD-L1 positive (TPS ≥ 1%), the patient should sign 
the main ICF for the study before the screening/baseline visit.  
Note: If the patient’s tumor has previously been identified to be PD-L1 positive by 
Roche Ventana PD-L1 (SP-142, SP-263) and Dako PD-L1 (Dako28-8, Dako22c3), it is 
unnecessary to test the PD-L1 status again.  The patient should sign the main ICF for the 
study before the screening/baseline visit.  
7.1.2 Screening 
All patients should sign the ICFs that has been approved by the ethics committee before 
any evaluation and operation of the clinical study is to be performed. The patient or the 
person signing the ICF must acquire one copy of the signed ICF (the clinical routine tests 
can be performed before signing the ICF). The investigator must record this process in the 
original medical record.  
The baseline evaluation during the screening phase must be performed within the four 
weeks before the first treatment cycle. The screening examination should be performed 
within the 28 days before the first cell infusion. All the results should be obtained before 
the patients are confirmed to be eligible for the study. If the patient conforms to all inclusion 
criteria without conformation to any of the exclusion criteria, then he/she can be enrolled 
in the study.  
The following procedures and evaluations should be conducted within the 28 days 
before the first cell infusion:  
• Medical record collection: the records, such as past medical history, operation 
history, demographic data and baseline information.  
• Evaluation based on the inclusion and exclusion criteria.  
• Complete physical examination: examine all of the systems in the body, including 
body weight, height, body surface area and vital signs (blood pressure, heart rate, 
respiratory rate and pulse rate).  
• ECOG performance score. 


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• Collection of fresh tumor tissue samples. 
• HIV, HBVsAg and HCV Ab must be tested within the 28 days before the study 
treatment, and if the HBVeAb is positive, the HBV- DNA must be tested and 
confirmed to be negative.  
• Blood culture. 
• Hematology test (including hemoglobin, red blood cell, white blood cell count and 
classification, and platelet count).  
• Blood biochemistry: liver function test (TBL, ALT, AST, AKP, ALB, A/G and LDH), 
kidney function test (BUN and Cr), and amylase, lipase, blood glucose, and blood 
electrolyte (K+, Na+ and Cl-) measurement.  
• Thyroid function test: TSH, free T3, and free T4. 
• Myocardial enzymes: troponin, myoglobin and CK-MB. 
• Tumor markers (CEA, CYFRA21-1 and NSE). 
• Lymphocyte subpopulation: CD3/CD4/CD8/NK. 
• Urine routine test. 
• Stool routine test. 
• Coagulation test (aPTT and INR). 
• Twelve lead ECG. 
• Ultrasonic cardiogram. 
• Pregnancy test: For all WOCBPs, the serum pregnancy test result obtained within 
72 hours before the study treatmentmust be negative.  
• Imaging study: mainly use chest plus abdomen enhanced CT, bone scan (or systemic 
PET-CT) and head enhanced MRI.  
• Collection of blood for T cells expansion: patients in group A: 40 ml; groups B and 
C: 60-80 ml. The actual blood volume is confirmed according to the routine blood 
test result of the patient at that time (please see Section 7.2.3 for details).  
• Cytokine level monitoring and erythrocyte sedimentation rate (ESR): IL-6, IL-10, 
CRP, TNF-α, ESR, IL-2 receptor, IL-2, IL-8, Fractalkine, IFN-r, MCP-1, IL-5, GM-
CSF, and FLT3L. Please refer to Section 7.2.1.5 for details.   
Remarks: if the peripheral blood of the patient that is collected during the screening 
period for editing and amplification does not generate enough edited T cells for the 
treatment transfusion within 28 days, the patient is disqualified from the screening. The 
patients who fail the screening can be screened again and may participate in the study.


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7.1.3 Screening failure 
If a patient signs the main ICF but does not meet the eligibility criteria, it will be 
considered a screening failure. The investigators should record the reason for the failure in 
the original document and in the eCRF during the screening phase. For the patients that failed 
in the screening, the evaluation date including the demographic data, ICF,and 
inclusion/exclusion criteria must be collected. Other data from these patients will not be 
recorded in the clinical database except for the SAEs occurred during the screening phase 
(refer to Section 8.1 for the SAE reporting process details).  
7.1.4 Treatment phase 
Each assessment should be completed within the seven days before the treatment. During 
the major treatment period, one treatment cycle is defined as 28 days. All of the safety 
assessments should be completed before the treatment. A three-day delay is allowed in each 
cycle, due to holidays, weekends, bad weather or other unpredictable conditions, and is not 
considered to be a protocol deviation.  
7.1.4.1 Main treatment period 
One cell therapy cycle is defined as 28 days. The following assessment procedures are 
required during the study.  
7.1.4.1.1 Assessment within one week prior to each treatment cycle 
•    General physical examination: examine all of the systems/organs in the body, 
including body weight, height, body surface area and vital signs (blood pressure, heart 
rate, respiratory rate and pulse rate).  
• ECOG performance status. 
• Hematology test (including hemoglobin, red blood cell, white blood cell count and 
classification, and platelet count). 
• Blood biochemistry: liver function test (TBL, ALT, AST, AKP, ALB, A/G and LDH), 
kidney function test (BUN and Cr), and amylase, lipase, blood glucose, blood 
electrolyte (K+, Na+ and Cl-) measurements.  
• Thyroid function test: TSH, free T3, free T4. 
• Tumor marker (CEA, CYFRA21-1 and NSE). 
• Lymphocyte subpopulation: CD3/CD4/CD8/NK. 
• Urine routine test, stool routine test, coagulation profile (aPTT and INR).  
•    Twelve lead ECG. 
• Pregnancy test: for all WOCBPs, a negative result in serum or urine pregnancy test 
must be obtained within the 72 hours before the study treatment.  
• Cytokine level monitoring and ESR: IL-6, IL-10, CRP, TNF-α, IL-2, IL-8, Fractalkine, 


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IFN-r, MCP-1, IL-5, GM-CSF, and FLT3L. Please refer to 7.2.2.5 for details.  
• Collection of blood samples: please refer to Section 7.2.3 for details.  
• The patient safety should be evaluated according to the NCI-CTCAE4.03.  
If the patient undergoes the screening evaluation within the seven days prior to the study 
treatment and no PD or clinical indication prompts further evaluation, there is no need to 
perform the blood biochemistry, hematology test, thyroid function test, tumor marker 
measurement, lymphocyte subset measurement, cytokine measurement, coagulation profile, 
urine routine test, stool routine test and ECG examination again on the first day of the first 
cycle.  
7.1.4.1.2 Assessment on day eight of each cycle 
• General physical examination: examine all of the systems/organs in the body, 
including body weight, body surface area and vital signs (blood pressure, heart rate, 
respiratory rate and pulse rate).  
• ECOG performance status. 
• Collection of the blood for PBMC expansion: 60-80 ml based on the hematology 
examination results of the patient. If a treatment delay is caused by poor blood cells 
amplification of the patient, the cell therapy can be delayed up to one week. If the delay 
exceeds one week, the Investigator should discuss to decide whether the treatment 
should be continued.  
• Hematology test (including hemoglobin, red blood cell and white blood cell count 
and classification, and platelet count).  
• Blood biochemistry test: liver function test (TBL, ALT, AST, AKP, ALB, A/G and 
LDH), kidney function test (BUN and Cr), and amylase, lipase, blood glucose, blood 
electrolyte (K+, Na+ and Cl-) measurement.  
• Lymphocyte subset distribution: CD3/CD4/CD8/NK. 
• Cytokine level monitoring and erythrocyte sedimentation rate (ESR): IL-6, IL-10, 
CRP, TNF-α, ESR, IL-2 receptor, IL-2, IL-8, Fractalkine, IFN-r, MCP-1, IL-5, GM-
CSF, and FLT3L. Please refer to Section 7.2.1.5 for details.  
• Collection of blood samples: please refer to Section 7.2.3 for details.  
• The tolerability of the prior cycle is evaluated according to the NCI-CTCAE4.03.  
7.1.4.1.3 End of cell therapy 
The visit after the completion of the cell therapy should be conducted within one week 
after the day of the last cell infusion. The time window (± 7 days) is allowed for all of the 
visits and evaluations. AEs and SAEs in all the patients should be followed up within the 28 
days after the last cell infusion, until 24 months after the treatment ends.  


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• General physical examination: examine all of the systems/organs in the body, including 
body weight, body surface area and vital signs (blood pressure, heart rate, respiratory 
raterespiratory rate and pulse rate).  
• ECOG performance status. 
• Hematology test (including hemoglobin, red blood cell, white blood cell count and 
classification, and platelet count).  
• Blood biochemistry: liver function test (TBL, ALT, AST, AKP, ALB, A/G and LDH), 
kidneyfunction test (BUN and Cr), and amylase, lipase, blood glucose, and blood 
electrolyte (K+, Na+ and Cl-) measurements.  
• Thyroid function test: TSH, free T3, and free T4. 
• Tumor markers (CEA, CYFRA21-1 and NSE). 
• Lymphocyte subpopulation: CD3/CD4/CD8/NK. 
• Urine routine test, stool routine test, coagulation profile (aPTT and INR).  
•    Twelve lead ECG. 
• Imaging study: mainly use chest plus abdomen enhanced CT, bone scan (or systemic 
PET-CT) and head contrast-enhanced MRI.  
• Cytokine level monitoring and erythrocyte sedimentation rate (ESR): IL-6, IL-10, CRP, 
TNF-α, ESR, IL-2 receptor, IL-2, IL-8, Fractalkine, IFN-r, MCP-1, IL-5, GM-CSF, and 
FLT3L. Please refer to Section 7.2.1.5 for details.  
• Collection of blood samples: please refer to Section 7.2.3 for details.  
• The tolerability of the prior cycle is evaluated according to the NCI-CTCAE4.03.  
7.1.5   Follow-Up after end of cell therapy 
The patients will go forward into the follow-up phase after cell therapy ends. The tumor 
evaluations of the patients will be followed up according to the scheduled visit in the visit 
form until PD occurs.  The objective response of tumors to the cell therapy is evaluated 
according to RECIST 1.1 and referring to ir RC.  
When the cell therapy is completed, the patient should visit the hospital, undergo safety 
examinations. The safety should be evaluated once a month during the first 12 months, and 
then once every two months during the next 12 months. The time window (± 7 days) is 
allowed for all the visits and evaluations.  
• Complete physical examination: examine all of the systems in the body, including body 
weight, body surface area and vital signs (blood pressure, heart rate, respiratory rate and 
pulse rate). 
• ECOG performance status. 
• Hematology test (including hemoglobin, red blood cell, white blood cell count and 


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Gene Edited T-Cell Therapy for Advanced Lung Cancer. Version 1.9 
 
classification, and platelet count).  
• Blood biochemistry: liver function test (TBL, ALT, AST, AKP, ALB, A/G and LDH), 
kidney function test (BUN and Cr), and amylase, lipase, blood glucose, and blood 
electrolyte (K+, Na+ and Cl-) measurements.  
• Thyroid function test: TSH, free T3, and free T4. 
• Tumor markers (CEA, CYFRA21-1 and NSE). 
• Lymphocyte subpopulation: CD3/CD4/CD8/NK. 
• Urine routine test, stool routine test, coagulation profiles (aPTT and INR).  
•    Twelve lead ECG. 
• Pregnancy test: for all WOCBPs, the negative serum or urine result in pregnancy test 
must be obtained within the 48 hours before the study treatment.  
• Imaging study: mainly chest plus abdomen enhanced CT, bone scan (or systemic PET-
CT) and head contrast-enhanced MRI. Please refer to Section 7.2.2 for details.  
• Cytokine level monitoring and erythrocyte sedimentation rate (ESR): IL-6, IL-10, CRP, 
TNF-α, ESR, IL-2 receptor, IL-2, IL-8, Fractalkine, IFN-r, MCP-1, IL-5, GM-CSF, and 
FLT3L. Please refer to Section 7.2.1.5 for details.  
• Collection of blood samples: please refer to Section 7.2.3 for details.  
• The tolerability of the prior cycle is evaluated according to the NCI-CTCAE4.03.  
7.1.6 End of treatment (EOT) 
When the patient discontinues the study treatment (including study completion and early 
termination), the visit evaluation should be completed within 14 days, and all the evaluations 
listed in the EOT should be performed. The eCRF should be filled in at the closing date and 
reasons for study discontinuation should be recorded. Follow-ups of the AEs and SAEs 
related to treatment should be completed until 24 months after the study ends. For patients 
with early withdrawal, the relevant information should be collected as much as possible. 
• General physical examination: examine all of the systems in the body, including body 
weight, body surface area and vital signs (blood pressure, heart rate, respiratory rate and 
pulse).  
• ECOG performance status. 
• Hematology test (including hemoglobin, red blood cell, white blood cell count, 
differential and platelet count).  
• Blood chemistry test: liver function tests (TBL, ALT, AST, AKP, ALB, A/G and LDH), 
renal function test (BUN and Cr), and amylase, lipase, blood glucose, and blood 
electrolyte (K+, Na+ and Cl-) measurements.  
• Thyroid function test: TSH, free T3 and free T4. 


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• Tumor marker (CEA, CYFRA21-1 and NSE). 
• Lymphocyte subpopulation: CD3/CD4/CD8/NK. 
• Urine routine test, stool routine test, coagulation profiles (aPTT and INR).  
•    Twelve lead ECG. 
• Cytokine level monitoring and erythrocyte sedimentation rate (ESR): IL-6, IL-10, CRP, 
TNF-α, ESR, IL-2 receptor, IL-2, IL-8, Fractalkine, IFN-r, MCP-1, IL-5, GM-CSF, and 
FLT3L. Please refer to Section 7.2.1.5 for details.  
• Collection of blood samples: Please refer to Section 7.2.3 for details.  
• The tolerability of the last cycle is evaluated according to the NCI-CTCAE 4.03.  
• Imaging studies: contrast-enhanced CT imaging of the chest and abdomen, bone 
scanning (or systemic PET-CT) and contrast-enhanced brain MRI. Imaging studies shall 
only be conducted in cases where the discontinuation of the study is due to any reason 
other than PD. If a scanning test is performed within 28 days before the end of the study, 
it is not required to be repeated. Please refer to Section 7.2.2 for details. 
7.1.7 Follow-ups 
7.1.7.1 The safety of follow-up period. 
All patients must undergo safety evaluation on the 30th day (± 7 days) after the EOT visit. 
Once treatment discontinues, all patients should be contacted once to evaluate the safety 
(AEs and/or SAEs), the following anti-tumor treatmentand the survival information. For the 
patients who permanently discontinue the study due to the AEs (including laboratory 
abmormalities), the follow-up must be continued until such events subside orstabiliz (the 
earlier should prevail). If the patients refuse to or fail to return for the safety evaluation visit, 
they should be contacted by telephone to exaluate their physical conditions. The attempts 
made to contact the patient should be recorded in documents (telephone contacts, date of 
registered mail, etc.). AEs and SAEs related to the study treatment should be recorded until 
24 months after the termination of the study or begining new antitumor therapy.  
• General physical examination: examine all of the systems in the body, including body 
weight, body surface area and vital signs (blood pressure, heart rate, respiratory rate and 
pulse rate).  
• ECOG score. 
• Hematology test (including: hemoglobin, red blood cell, white blood cell count and 
differential and blood platelet count). 
• Blood chemistry test: liver function tests (TBL, ALT, AST, AKP, ALB, A/G and LDH), 
renal function test (BUN and Cr), and amylase, lipase, blood glucose, and blood 
electrolyte (K+, Na+ and Cl-) measurements.  


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• Thyroid function test: TSH, free T3, free T4. 
• Tumor marker (CEA, CYFRA21-1 and NSE). 
• Lymphocyte subpopulation: CD3/CD4/CD8/NK. 
• Urine routine test, stool routine test, coagulation profile (aPTT and INR).  
•    Twelve lead ECG. 
• Imaging studies:contrast-enhanced CT imaging of the chest and abdomen, bone 
scanning (or systemic PET-CT) and enhanced brain MRI. Please refer to Section 7.2.2 
for details. 
• Cytokine level monitoring and erythrocyte sedimentation rate (ESR): IL-6, IL-10, CRP, 
TNF-α, ESR, IL-2 receptor, IL-2, IL-8, Fractalkine, IFN-r, MCP-1, IL-5, GM-CSF, and 
FLT3L. Please refer to Section 7.2.1.5 for details.  
• Blood sample collection: please refer to Section 7.2.3 for more details. 
• The tolerability of the prior cycle is evaluated according to the NCI-CTCAE4.03.  
7.1.7.2 PFS follow-up period 
If a patient discontinues the treatment for any reason other than PD, a tumor evaluation 
will be continued asdescribed in Section 7.2.2, until PD, death, or initiation of a new anti-
cancer treatment. For each patient that discontinues the treatment, the investigator or his 
representative will continue to collect the information about the initiation of other anti-cancer 
treatment until the cut-off date of the final data analysis. The patients that lost to be followed 
up should be recorded in the eCRF as censored and, if patients are lost in the follow-up 
period, the investigator should record the attempts to make contact with the patients in the 
original medical records, such as through telephone, registered mail, and so on.  
7.1.7.3 Survival follow-up period  
If the patient develops objective PD, survival follow-up should be performed once every 
two months. The survival information may be collected through several ways, including 
contacting the patients and/or their family by telephone. Information regarding the anti-
tumor therapy, survival information and other ifformations should be collected.  
7.2 Assessment types 
7.2.1 Safety and tolerability assessments 
The safety assessments include the monitoring and laboratory tests of all AEs and SAEs, 
such as the regular monitoring of hematology and urine routine test, regular assessment of 
vital signs, weight, performance status, physical examination, cardiac evaluation, ECG and 
other evaluations.  
For more information about AE collection and report please refer to Section 8.  
7.2.1.1 Physical examination 


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Physical examination should be performed according to the general medical practice, 
including medical history, inspection, palpation, percussion and auscultation. Physical 
examination will be processed at the screening/baseline visits, on day one and day eight of 
each cycle, and at the end of the treatment.  
If a baseline examination has been performed within the 72 hours prior to the first cell 
therapy, repeated physical examination is not required on the first day of the first cycle, which 
is also applicable for the first day examination of the following cycles. 
If clinically indicated, physical examination can be performed more frequently in the 
discretion of the judgment of the investigator. Any important findings recorded prior to 
signing of the ICF must be recorded on the medical history pages in the eCRF. Important 
new findings that are first observed or exacerbated after the signing of the ICF must be 
recorded on the AEs pages in the eCRF.  
7.2.1.2 Vital signs 
Vital signs include temperature, respiratory rate, blood pressure and heart rate. The blood 
pressure and heart rate should be measured in a sitting position and the patients should have 
rested for at least 10 minutes prior to the measurement. Any abnormality with clinical 
significance will be recorded as an AE in the eCRF. 
7.2.1.3 Height and weight 
Height will only be measured at screening and the patient’s weight will be measured at 
the screening/baseline visit, on the first day of the first cycle and the following cycles.  
If a baseline evaluation has been performed within the 72 hours prior to the first cell 
therapy, there is no need to perform it again on the first day of the first cycle.  
7.2.1.4 Performance status 
The ECOG score will be evaluated during the process of screening before the first cycle 
(before cell therapy), on the first day of each following cycle and at the end of the treatment.  
If a baseline evaluation has been performed within the 72 hours prior to the first cell 
therapy, it is not necessary to repeat the evaluation on the first day of the first cycle.  
7.2.1.5 Laboratory assessments 
The testing will be performed at the laboratory of the study center based on the visits 
listed in Table 7-1. Samples for laboratory tests will be collected and evaluated during the 
scheduled visit period, even if the study treatment has been suspended. Additional or 
repeated tests will be performed for safety and/or laboratory abnormalities based on the 
details. The priniciple investigator will take charge of the final evaluation of the laboratory 
findings, the completion of records and the filling in of the eCRF.  
Abnormal laboratory values or findings can be deemed as AEs only when there are 


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clinical signs or symptoms that are meaningful or require treatment. These signs, symptoms 
or relevant diagnoses should be recorded as an AE in the eCRF. In addition, if an independent 
laboratory abnormality is of clinical significance (e.g. resulting in the discontinuation of the 
treatment, as a part of a SAE, or it is the SAE), it should also be recorded as an AE in the 
eCRF. 
Monitoring of CRS:  
The monitoring of CRS will be processed by collectingapproxmately 3 ml peripheral 
blood on the first day of the first cycle, 24 hours after the completion of the last cell infusion 
(day five) of each cycle, day twelve of each cycle (the 7th day after last infusion of each cycle) 
during the study. If a CRS is suspected, additional sampling will be required based on the 
clinical signs.  
The cytokines that are tested include: IL-6, IL-10, CRP, TNF-α, ESR, IL-8, Fractalkine, 
IFN-r, MCP-1, IL-5, GM-CSF and FLT3L. Plasma samples have been retained, and IL-8, 
IL-2, Fractalkine, IFN-r, MCP-1, IL-5, GM-CSF, FLT3L are all exploratory indexes for 
clinical study will be tested at the end of the study. 
If the investigator suspects that a patient’s clinical signs or symptoms are clinically 
significant, additional or repeated test can be conducted based on the conditions of the 
individual case. The principle investigator is responsible for the evaluation and confirmation 
of the lab test results with a detailed record.  
7.2.1.6Tolerability evaluation 
The tolerability will be evaluated based on the cell therapy dose, the treatment course, 
the incidence rate of AEs that cause disruption of the cell therapy (interruption of the 
treatment or dose reduction) and the early-discontinuation of the treatment.  
7.2.2 Measurement of efficacy 
The imaging evaluation results during the screening period are valid within the 28 day 
prior to the treatment. The RECIST1.1 assessment will be applied during the screening (by 
using chest and abdomen CT or MRI); meanwhile, it is necessary to carry out the bone 
scintigraphy/bone scanning and brain scanning/head and neck CT/MRI as clinically 
indicated. All of the scans should be performed according to the schedule from the 
completion day of the treatment (± 7 days). If the patients have brain metastases, the brain 
imaging must be performed during future tumor evaluations. If the patient is confirmed to 
have bone metastases at the time of the baseline assessment, then bone scintigraphy/bone 
scanning should be repeated at an interval of six months (± 7 days) to monitor the new 
lesions. If the imaging test can be used to evaluate the bone metastasis lesion, there is no 
need to repeat the bone scintigraphy/bone scanning.  


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All of the follow-up imaging scans will be processed based on the schedule from the first 
day of the first treatment cycle. The scan after the first treatment must be processed at weeks 
8 (+1) and 12 (±1) ... After that, the tumor evaluation will be processed once every eight 
weeks (+/-7 days) until PD. Other corresponding examinations will be performed as 
clinically indicated. All of these evaluations must be completed before the next treatment. If 
the imaging study has been performed within the four weeks before the treatment completion, 
there is no need for the re-examination.  
If the treatment for the patients is early discontinued for reasons other than PD, the tumor 
evaluation for those patients should be continued once every two months (according to the 
RECIST1.1 criteria) until the patients' death or the first presence of PD. Final tumor 
evaluation should be conducted between the fourth and sixth weeks after the first PD. All 
known and suspected lesions should be monitored through imaging and evaluated according 
to the RECIST1.1 criterion.  
IrRC will allow re-evaluaton 4 weeks after the pgression as immnotherapy may cause 
short-time tumor flare (pseudoprogression) if general condition is stable. 
7.2.3 Biomarkers 
Table 7-2-3:  Collection of Blood for biomarker tests. 
Sample 
collected 
Time points for collection 
Blood 
sample 
volume  
ctDNA (see  
7.2.3.1 
for 
details)  
baseline, 28th day of second cycle, week 8, week 12, and 
then every eight weeks.  
 
10 ml  
Immune 
repertoire 
(see 
7.2.3.2 
for 
details)  
Pre-A group: baseline, day six of first cycle, first day of 
second cycle. First and sixth days in each cycle if the 
patients continue with the cell infusion, or week 8 and 12 
if the patients discontinue the cell infusion. 
Group A, B and C: baseline, day six of first cycle, first and 
sixth days of second cycle. First and sixth days in each 
cycle if the patients continue with the cell infusion, or 
week 8 and 12 if the patients discontinue the cell infusion. 
5 ml or 10 
ml*  
T 
cell 
in 
vivotracking 
First and 12thday of each treatment cycle  
After treatment: the 28th day of the last treatment cycle, 
three months, six months, and one year after the last cell 
10 ml  


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Gene Edited T-Cell Therapy for Advanced Lung Cancer. Version 1.9 
 
infusion. 
Flow cytometry 
of regulatory T 
cells 
(Treg), 
memory T cells, 
activated T cells 
and PD-1  
Before pre-treatment with cyclophosphamide (blank 
baseline), after pre-treatment with cyclophosphamide 
(medication baseline), 24h after the third cell transfusion 
of each cycle, the end of each cycle (day 28), week eight, 
three months, and six months.  
5 ml  
 
*Notes: A total of 15 ml peripheral blood is collected at the same time point for ctDNA and immune 
repertoire samples; 10 ml peripheral blood is collected at the time point for immune repertoire samples 
only. 
7.2.3.1 ctDNA 
Circulating tumor DNA (ctDNA) refers tothefree partially degradatedendogenous DNA 
in the circulating blood, which is mainly produced by tumor cell necrosis, apoptosis, and 
even the secretion of tumor cells [37].  With regards to the monitoring of anti-cancer treatment, 
the ctDNA level in plasma can reflect the blood tumor mutation burden (bTMB). The ctDNA 
level is highly correlated to the imaging tumor burden, and can be even more sensitive than 
imaging [38]. Tumors have temporal and spatial heterogeneity. Based on the mutation 
spectrum of ctDNA from the peripheral blood, the pedigree of the tumor cell clones can be 
analyzed [39], and the specific responsivity of the tumor clone to the treatment in this study 
can be further explored. 
Ten microliters of peripheral blood will be collected each time at baseline, 28th day of 
second cycle, week 8, week 12, and then every eight weeks. 
7.2.3.2 Immune repertoire 
The diversity of T-lymphocyte and B-lymphocyte receptor repertoires is derived from 
the recombination of receptor genes. The recombination generates a huge diversity of 
receptors, of which, CDR3 has the highest diversity that best represents the specificity of 
receptor repertoires. The diversity of receptor repertoires also demonstrates a potential 
predictive role in the evaluation of the efficacy of immune checkpoint inhibitors. The 
exploration of the dynamic changes in T cell receptors (TCR) during and after the treatment 
and the exploration of the specific T cell clones that are possibly related to efficacy using 
this method have been proposed.  
The peripheral blood will be collected at a volume of 5 ml per time at the baseline of 
overall treatment, 24 hours after the last cell infusion of each treatment cycle. In addition, 5 
ml of peripheral blood will be collected from the patients in each group to analyze the 


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Gene Edited T-Cell Therapy for Advanced Lung Cancer. Version 1.9 
 
diversity of T cell receptors at each of the following time points: Pre-A group: baseline, day 
six of first cycle, first day of second cycle. First and sixth days in each cycle if the patients 
continue with the cell infusion, or week 8 and 12 if the patients discontinue the cell 
infusion.Group A, B and C: baseline, day six of first cycle, first and sixth days of second 
cycle. First and sixth days in each cycle if the patients continue with the cell infusion, or 
week 8 and 12 if the patients discontinue the cell infusion. 
7.2.3.3 Screening of the tumor biomarkers (PD-L1) 
It is required to evaluate the PD-L1 expression of the tumor. Although PD-L1 expression 
is not the best index to predict the efficacy of the PD-1/PD-L1 inhibitors, current data [40,41] 
still reveals that positive PD-L1 expression is related to the efficacy. Additionally, the 
NSCLC patients withpositive PD-L1 have a higher probability to recieve benefits from these 
inhibitors. Furthermore, among the current exploratory prediction indexes, PD-L1 could be 
tested simply and quickly using IHC. Therefore, this study will take PD-L1 positivity as one 
of the inclusion criteria to potentially enhance the therapeutic benefit to patients. PD-L1 
positivity will be identified based on the types of tumor samples in the pathology laboratory 
of the study center. PD-L1 status detection was used by fresh tissue or paraffin-embedded 
tissue. 
7.2.3.4 PDX model 
Patient-derived xenografts (PDX) have been well established by inoculating the tumor 
cells that are derived from the patients to the mice with severe combined immunodeficiency, 
which can highly specifically simulate the in vivo condition of the patients [42, 43]. In addition, 
it has already been reported that the PDX model can be established by utilizing biopsies [44].  
In this study, it has been proposed that fresh tumor specimens from the patients can be 
used to establish the PDX model, test the feasibility ofautologous infusion of the gene-edited 
cells and calculate whether the animal model can predict the clinical efficacy and allow 
analysis of the possible mechanisms. It is proposed that tumor samples from the patients are 
obtained by core needle biopsy before infusing the T cells, and inoculated into NOD SCID 
gamma (NSG) mice in situ, followed by the injection of 107 gene-edited T cells into the mice 
via the caudal vein to observe the effect on the mice. The peripheral blood, bone marrow and 
a tumor specimen will be collected from the mice to analyze the dynamic distribution and 
functional alteration of the cells.  
7.2.3.5 In vivo T cell tracking 
The edited T cells that will be infused in this study is not stem cells that can proliferate 
infinitely. The infused T cells will gradually be dismissed in patients over time. The dynamics 
of edited T cells in vivo, although not clear, may be associated wtih the efficacy and toxicity 


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Gene Edited T-Cell Therapy for Advanced Lung Cancer. Version 1.9 
 
of the treatment. The dynamics of this process is also critical for the determination of the 
optimal treatment period and time of cell infusion. In this program, digital PCR (ddPCR) and 
deep sequencing will be used to dynamically monitor the changes in the cell numbers in the 
peripheral blood of the patients over time, at the intended time points.  
• Baseline and day 12 of each treatment cycle. 
• The 28st day of last treatment cycle, three months, six months, and one year after the 
last cell infusion.  
• Blood volume: 10 ml each time. 
7.2.3.6 Flow cytometery for regulatory T (Treg) cells, memory T cells, activated T cells 
and PD-1.  
Tumor-associated immuno-suppressive cells include Treg cells, myeloid derived 
suppressor cells, tumor-associated macrophages and immature dendritic cells. Treg isthe 
major immuno-suppressive cell type, which can negatively regulate other immune cells 
directly or indirectly, and can inhibit the anti-tumor immune response. Cyclophosphamide is 
a drug that is most commonly used in pre-treatment during tumor immunotherapy. 
Cyclophosphamide can reduce the number of Treg cells, inhibit the function of Treg, and 
enhance anti-tumor immunity function. Therefore, the number of Treg cells inthe peripheral 
blood will be dynamically monitored in this study.  
T memory cells and activated T cells have anti-tumor immunity functions. T-memory 
cells are generated from the proliferation and differentiation of T cells following induction 
by a specific antigen.  T memory cells maintain a long-term immune response activity to 
specific antigens.  Activated T cells are derived from the activation of different T lymphocyte 
subset by antigens or cytokines. Activated T cells play a role in immune regulation. PD-1 is 
an important immuno-suppressive molecule. Overexpression of PD-1 on CD8+T cells 
indicates the effect of tumor immune suppression. In this study, the changes in PD-1 
expression in T memory cells and activated T cells will be dynamically monitored after 
infusion by taking samples of the peripheral blood to evaluate the efficay of the cell therapy. 
The expresssion of PD-1 on pre-edited and post-edited T cells will also be detected. 
Blood collection time points: before pre-treatment with cyclophosphamide (blank 
baseline), after pre-treatment with cyclophosphamide (medication baseline), 24 h after the 
third cell infusion of each cycle, the end of each cycle (day 28), Week eight, three months, 
and six months after the final treatment. Blood collection volume: 5 ml each time.   
7.2.4 Other descriptions 
If the patient’s consent has been obtained, the remaining samples can be used for further 
analysis. The samples can be kept up to five years. These samples can be used to explore the 


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Gene Edited T-Cell Therapy for Advanced Lung Cancer. Version 1.9 
 
efficacy and/or safety (such as off-target analysis) of edited cells. At any phase of this study, 
patients can discontinue the study voluntarily; investigator can determine if patient continue 
or exit the study based on the judgment of the potential benefits to patients.  
Patients will discontinue the study in the following cases: 
• Patient withdraws the ICF  
• Death  
• AEs  
• Laboratory abnormalities or other abnormal findings 
• Misconduct in the study protocol  
• Lost in follow-up  
• PD  
• New anti-cancer treatment  
• Noncompliance of the patients  
For those patients who do not agree to receive further treatment and evaluation, it is 
necessary to clarify whether they are willing to accept the survival follow-up (which can be 
made by telephone). If patients withdraw the ICFs and no longer participate in any study or 
follow-up, including the survival follow-up study, it shall be recorded in detail in the source 
documents and filed in the eCRF.  
7.2.5 Withdrawal of ICF 
Patients will be entitled to exit the study at any period of the study, which neither causes 
the discrimination of patients in following treatment, nor impacts on their decision to 
participate in other studies. 
For patients who withdraw the ICFs, the reasons and whether there is any AE must be 
recorded clearly, and investigators will follow-up regarding the AE after the study.  
7.2.6 Discontinuation of the study 
If significant findings indicate enormous risks for patients, the investigators can 
terminate the study. No matter why the study is terminated, all available data from the 
subjects that terminate the follow-up shall be recorded clearly.  
8. Adverse events 
An adverse event (AE) refers to any new adverse medical event or any worsening disease 
after the patient signs the ICF. An AE may not have a causative relation to the treatment or 
drugs, thus, an AE includes, but is not limited to: the abnormal laboratory test; a symptom 
of clinical significance; a change in an examination result. AEs can be spontenously reported 
by the patients, recorded during open communication with the patients or discovered during 
the examination or evaluation of the patients.  


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       For the existing cases before the ICF is signed, the relevant case history shall be recorded. 
For AEs, the investigators shall trace the details of the AEs to confirm whether the AE is 
related to the study treatment, assess the result of AE and evaluate whether it is a SAE. 
      AEs should be evaluated in accordance with the CTCAE version 4.03. If there is no 
grading for certain AEs in the CTCAE, the AEs shall be graded into grades 1-4, which 
corresponds to mild, moderate, severe and life-threatening.  
An AE should be monitored from the time when the patient signs the prior consent and 
throughout the overall treatment period and safety follow-up period. The safety follow-up 
period is defined as the period from the time when the patients discontinue the cell therapy 
until death. Any SAE identified in the follow-up period should also be reported according to 
the regulation.  
It should be judged by the investigators whether there is a causative relationship between 
any AE and the study treatment. The relationship between an AE and the study treatment 
could be: definitely related, probably related, probably unrelated or definitely unrelated. The 
incidence rate of the AEs that are definitely related and probably related to the study 
treatment will be listed.  
Investigator should follow upthe patients that exit the treatment due to AEs, to follow 
up and record the outcomes of all AEs until such events are fully relieved or subside.  
Investigator must judge whether the AE is related to the study treatment, the measures taken 
during the study treatment (no action, discontinuation of the study treatment, temporary 
interruption for medicine, permanent discontinuation of the study treatment, unknown, etc.), 
and the application of concomitant treatment (with/without concomitant treatment), based 
on the supporting evidence. Investigators should also evaluate the outcomes of the AE 
(disappear/not disappear, recovered, in recovery, whether there are sequelae, death, unknown, 
etc.), and determine whether it is an SAE. 
Abnormal values of laboratory tests: 
The laboratory abnormalities with clinical significance (inducing clinical signs or 
symptoms, requiring concomitant treatment or adjustment of the trial, treatment) should be 
recorded as AEs, with an emphasis on the diagnosis results instead of the signs and symptoms 
(for example, anemia other than low hemoglobin). The laboratory abnormal values that 
recording as AEs should be followed up until they return to normal or can be reasonably 
explained. If an abnormal laboratory value or test result shows the same signs/symptoms as 
a reported AE, it will not be recorded as a different AE. 
Investigator should follow up the patients that exit the study due to AEs, to follow up 
and record the outcomes of any AEs until such events are recovery or stable. Investigator 


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Gene Edited T-Cell Therapy for Advanced Lung Cancer. Version 1.9 
 
must judge whether the AE is related to the study treatment, the measures taken during the 
study treatment (no action, discontinuation of the study treatment, temporary interruption for 
medicine, permanent discontinuation of the study treatment, unknown, etc.), and the 
application of concomitant treatment (with/without concomitant treatment), based on the 
supporting evidence. Investigators should also evaluate the outcomes of the AE 
(disappear/not disappear, recovered, in recovery, whether there are sequelae, death, unknown, 
etc.), and determine whether it is an SAE. 
8.1 Serious adverse events 
8.1.1 Definition of serious adverse events 
A severe adverse event (SAE) refers to any adverse medical events as follows:  
• Cause death;  
• Threaten life (risk of sudden death);  
• Hospitalization or prolongation of current hospital stay;  
• Cause a permanent or significant disability/incapacity (heavily damage the ability to 
perform daily activities);  
• Cause a congenital abnormality/birth defect.  
Hospitalization for the following reasons should not be reported as a SAE:  
a)  Hospitalization due to signs and symptoms of PD;  
b)  Regular treatment or monitoring for study indication, without exacerbation;  
c)  Social reasons and temporary hospitalization, without exacerbation;  
d)  Elective or advance treatment to pre-existing disease irrelevant to the indication in 
this study, without exacerbation after the ICF is signed.  
If the patients receive treatment in outpatient or emergency departments without 
hospitalization, and the time involved does not meet the above definition of SAE, the 
involved events shall not be regarded as AEs.  
8.1.2 Report 
An SAE should be reported in the patients after signing the ICFs (on molecular 
screening). For those patients who have signed the main ICFs, no matter whether the 
screening fails or not, the reporting of an SAE will start from the time when the main ICF 
has been signed.  
To ensure the safety, any SAE that occurs from the time when the ICF is signed to the 
time 28 days after the termination the treatment should be reported, no matter whether it is 
suspected to be related to the study treatment or not.  
When an SAE that occurs after the EOT is suspected to have a causative relation with 


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Gene Edited T-Cell Therapy for Advanced Lung Cancer. Version 1.9 
 
the study treatment, information about the initiation, closure, complication and progression 
of the SAE must be reported to the relevant departments within 24h after the information 
was received. During different periods, an SAE should be reported as a new event if the SAE 
is completely unrelated to the previously reported SAE. All SAE information should be 
collected and recorded on the SAE report form; as a complete clinical report, all applicable 
sections on this form must be filled in.  
In the follow-up section, the following items shall be described: if the event has subsided 
or continues; if any treatment has been given and how the treatment is applied; if the patient 
is continuing the study or has dropped off.  
Any SAE shall be reported to the ethics committee and the relevant management 
department within 24 hours after it is identified (Appendix 2). 
8.2 Pregnancy 
In order to ensure the safety of patients, the pregnancy events during the study should be 
reported within 24h once recognized. The pregnancy should be followed up and the final 
outcomes should be recorded, including spontaneous termination, a detailed description 
during childbirth, birth defects, congenital abnormalities, and complications for the mother 
and/or the newborn.  
The enrolled patients should take a strict and effective contraception measure. If the 
patients are pregnant during the study period, the cell therapy should be terminated 
immediately.  
The female patients of bearing-age are required to use effective birth control during the 
cell therapy and for 18 months after stoping the cell therapy.  
Efficient birth control includes:  
•    Female sterilization 
•    Male sterilization (at least six months before screening)  
•    The combination of any two of the following methods (a+b, a+c, or b+c):  
a. Oral, injectable or implantable hormone, or other hormone with equivalent effect 
(failure rate < 1%), for contraception, for example, using hormone pessulum or transdermal 
hormone for contraception  
b. Intrauterine devices (IUD) or intrauterine systems (IUS)  
c. 
Barrier 
methods 
of 
contraception: 
condom 
or 
spermicidal 
foam/glue/film/cream/vaginal pessary (diaphragm or cervical/vaginal fornix cap) 
9. Data management and analysis 
9.1 Data confidentiality 


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Gene Edited T-Cell Therapy for Advanced Lung Cancer. Version 1.9 
 
      Information about study subjects will be kept confidential and managed under the 
applicable laws and regulations.  
9.2 Site monitoring 
During the study, the monitor will visit the site regularly to check the completeness of 
patient records, the accuracy of entries on the CRFs, the adherence to the protocol to Good 
Clinical Practice, the progress of enrollment, and to ensure that study product is being stored, 
dispensed, and accounted for according to specifications. Key study personnel must be 
available to assist the field monitor during these visits.  
The investigators must maintain source documents for each patient in the study, 
consisting of case and visit notes (hospital or clinic medical records) containing demographic 
and medical information, laboratory data, electrocardiograms, and the results of any other 
tests or assessments. All information recorded on CRFs must be traceable to source 
documents in the patient's file. The investigator must also keep the original signed informed 
consent form (a signed copy is given to the patient).  
9.3 Data collection 
For study using Electronic Data Capture (EDC), the designated investigator staff will 
enter the data required by the protocol into the Electronic Case Report Forms (eCRF).  
The principal investigator is responsible for assuring that the data entered into eCRF is 
complete, accurate, and that entry and updates are performed in a timely manner. 
9.4 Statistical methods and data analysis 
Analysis and reporting of study data will be based on the data from all patients. The 
following rules will be followed for reporting results unless stated otherwise:  
For the categorical data, the frequency and percentage will be presented. For continuous 
data, mean, standard deviation, median, minimum, and maximum will be provided.  
The treatment groups are specified based on the dose levels of the cell therapy. Each 
treatment group includes all of the patients who are assigned to receive the dose at the 
specific level. All summaries, tables, figures and analyses are displayed by treatment group.  
Screening failure patients are referred to as those who signed the ICFs but did not meet 
the eligibility criteria. For these patients, the eCRF data collected according to Section 7.2 
will not be included in the analysis, but will be reported as separate listings in the CSR.  
9.4.1 Analysis data sets 
Full analysis set:  
The full analysis set (FAS) includes all of the patients who received at least one infuson 
of the cell therapy. The FAS will be the default analysis set unless specified otherwise.  


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Safety set:  
The safety set includes all of the patients who received at least one infusion of the cell 
therapy and had safety evaluation. Patients will be analyzed according to the study treatment 
they actually received. The safety set will be used for safety summary of the study.   
Pharmacokinetic analysis set:  
The pharmacokinetic Analysis Set (PAS) consists of all of the patients in the FAS who 
have at least one blood sample that can provide evaluable pharmacokinetic data. The PAS is 
used for all pharmacokinetic analyses.  
9.4.2 Patient demographics / other baseline characteristics 
Demographic and other baseline data (including disease characteristics) will be listed and 
summarized descriptively by treatment group using FAS.  
9.4.3 Treatments (study treatment, concomitant therapies, compliance) 
Study treatment  
The actual dose and the relative dose (the ratio of actual received dose to planned dose) 
will be listed and summarized with descriptive statistics by treatment group. The reasons for 
dose delay, reductions, interruption and discontinuation will be listed and summarized.  
Concomitant therapies 
Concomitant medications and significant non-drug therapies prior to and after the the 
treatment will be listed by patient and summarized by ATC (anatomical therapeutic chemical 
classification system) term and treatment group. 
Compliance  
Compliance to the protocol will be assessed by the number and proportion of patients 
with protocol deviations. Protocol deviations will be identified prior to database lock and 
will be listed and summarized by treatment group/disease group. Compliance to the study 
drug will be assessed by the number of dose delay, reductions and dose interruptions.  
9.4.4 Primary objective 
To evaluate the safety and tolerability after autologous transfusion of CRISPR-Cas9 
knockout PD-1 T cells.  
9.4.4.1 Variable 
Safety analysis varibles include the incidence of dose limiting toxicities (DLTs) in the 
first cycle of treatment, incidence rate and severity of AEs and SAEs including changes in 
laboratory parameters, vital signs, and ECG results.  
9.4.4.2 Safety analysis 
The primary analysis will include the summary of observed DLTs by treatment group, 
and the summary of the frequency, severity of AEs, changes in laboratory findings, physical 


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Gene Edited T-Cell Therapy for Advanced Lung Cancer. Version 1.9 
 
examinations, vital signs, ECGs. The safety set will be used for all safety evaluation. 
Adverse events will be coded using the Medical Dictionary for Regulatory Activities 
(MedDRA) terminology. Severity of AE will be coded based on the National Cancer 
Institute’s Common Terminology Criteria for Adverse Events (CTCAE 4.03). All AEs 
recorded during the study will be listed. The incidence of treatment-related AEs (definitely 
or probably related to the treatment) will be summarized by system organ class (SOC) and 
preferred term (PT), severity (based on CTCAE grades), relation to study treatment. 
The number and percentage of patients with DLTs will be tabulated by SOC and PT 
severity. All observed DLTs will be listed. 
Deaths reportable as SAEs and non-fatal serious adverse events will be listed by patient 
and tabulated by type of adverse event and treatment group.  
All laboratory values will be graded by using CTCAE 4.03. For laboratory tests covered 
by CTCAE, a Grade 0 will be assigned for all non- missing values not graded as 1 or higher. 
Grade 5 will not be used. For laboratory tests where grades are not defined by CTCAE, 
results will be graded by the low/normal/high classifications based on laboratory normal 
ranges. 
If the lower limits of normal ranges used in CTCAE definitions are missing, then they 
have to be replaced by a clinical meaningful limit. 
The following summaries will be generated separately for hematology, biochemistry and 
urinary laboratory tests: 
• frequency table for newly occurring on-treatment grades 3 or 4; 
• shift tables using CTCAE grades to compare baseline to the worst on-treatment 
value; for laboratory tests where CTCAE grades are not defined, shift tables using 
the low/normal/high/(low and high) classifications to compare baseline to the worst 
on- treatment value;  
• Listing of all laboratory data with values flagged to show the corresponding 
CTCAE grades and the classifications relative to the laboratory normal ranges.  
The number and percentage of patients with ECG and vital signs abnormalities will be 
provided. All ECG and vital signs of patients with the abnormalities will also be listed. 
9.4.4.3 Tolerability 
Tolerability of the study treatment will be evaluated by dose interruptions, reductions 
and dose intensity.  
9.4.5 Secondary objective analysis 
9.4.5.1 Efficacy analysis 
Efficacy analysis will use FAS.  


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The best objective response (BOR), objective response rate (ORR), 8-week disease 
control rate (DCR), time to response (TTR), duration of response (DOR), progression free 
survival (PFS) and overall survival (OS) will be analyzed toassess the efficacy of the study 
treatment. The tumor response will be evaluated by investigators based on the RECIST 1.1 
and immune-related response criteria (irRC) (refer to Section 12.3).  
The individual lesion and overall tumor response will be listed by patient and assessment 
date. BOR, TTR, DOR, PFS and OS will be listed by patients. 
The BOR will be summarized as appropriate, and the estimates rate of each type of 
response will be provided along with 95% exact confidence interval (CI) by treatment group. 
The best change in tumor size from baseline will be graphically presented.  
9.4.5.2 In vivo activity status and duration of PD-1 edited T cells 
The number of PD-1 edited T cells in blood will be listed and summarized by treatment 
group and time point. Descriptive statistics will include the arithmetic mean and geometric 
mean, median, standard deviation, arithmetic coefficient of variation (CV), geometric 
coefficient of variation, minimum and maximum. In geometric mean calculations, 
concentration 0 is not included. Patient level and averaged concentration of PD-1 edited T 
cell measures will be displayed using longitudinal plots. Graphical presentation of arithmetic 
mean (±SD) concentrations at each scheduled time point will be provided by treatment group. 
9.4.5.3 Immunological factors and biomarkers 
Baseline level of PD-L1 will be listed, Soluble immune and inflammatory cytokines (e.g. 
IL-10, interferon gamma, IL-6, IL-6 receptor, CRP, and ferritin) will be listed by patient and 
may be summarized by treatment group and time point. 
9.4.6 Exploratory analysis 
The frequency of TCR, TCR diversity and bTMB among patients may be summarized 
and displayed in graphics by treatment group at each time point before and during the 
treatment. TCR diversity and bTMB were compared between patients with and without 
disease control benefit using the exact Wilcoxon test. Statistical significance was set at p ≤ 
0.05 (2-sided). 
The relationship between the efficacy and safety variables and changes in the level of 
other molecules may be explored through plots. If any potential relationship is explicated in 
the graphics, Further analysis with statistical modeling may be performed. Additional 
exploratorty analysis may be performed as appropriate. 
9.4.7 Interim analysis 
      No formal interim analysis is planned for the study. During the dose-escalation, decisions 
will be made based on the collected data following the study procedure. For each dose cohort, 


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when all patients complete the DLT observation, the dose level for the next cohort will be 
determined based on the observed data.  
9.4.8 Sample size calculation 
In this dose-escalation study, each dose cohort will enroll three to six patients. Each dose 
level is allowed to be tested in more than one cohort of patients sequentially. About 20 
patients are planned to enroll for this study.  
10 Ethical considerations 
10.1 Regulatory and ethical compliance 
The design, implementation and reporting of this clinical study will follow the ICH 
tripartite coordination guiding principles of good clinical practice, applicable to local laws 
and regulations and ethical principles derived from the Declaration of Helsinki.  
10.2 Responsibilities of the investigators and IRB 
Before the study initiation, the Declaration of Helsinki (2000 edition) must be followed 
and the relevant Chinese medical study norms and regulations must be implemented. The 
clinical study will be conducted only after the ethics committee of the study unit has 
approved the study protocol and the proposed ICF. Before each patient is chosen for the study, 
the investigator will be responsible for introducing the objective, procedures and possible 
risks completely and comprehensively to the patient or his representive in writing. Patients 
should be aware that they have the right to exit the study at any time. Each patient should be 
given a written patient ICF prior to screening and enrollment. It is the investigator's 
responsibility to obtain the ICF before the patient is screened and enrolled, and the ICF 
should be retained for future reference as a clinical study document. Before the study 
initiation, the investigator needs to sign on the signature page of the study protocol to confirm: 
He/she agrees to carry out the study in accordance with all of the guidelines and procedures 
in these documents and the study protocol, and permits the examination of related data and 
records by CRA, ethics and the competent authority as needed.  
10.3 Informed consent procedures 
Only eligible patients who have provided a written, ethically approved ICF can be 
enrolled in the study.  
The ICF must be collected prior to the execution of any study procedure (i.e., all of the 
procedures described in the study protocol), especially the molecular screening. The process 
of collecting the ICF should be recorded in the patient's original medical record. The date of 
ICF should be recorded in the patient's eCRF.  
The investigator will prepare ICF that is appropriate for the study and complies with the 


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Gene Edited T-Cell Therapy for Advanced Lung Cancer. Version 1.9 
 
ICH GCP guidelines and regulatory requirements.  
Women of childbearing age should be informed that there is an unknown risk to the fetus 
if pregnancy occurs during the study, and the patient can only be enrolled in the study after 
she agrees to abide by the contraceptive requirements. The patient will not be enrolled in the 
study if she is unable to comply with the contraceptive requirements reliably.  
10.4 Discontinuation of the study 
     The sponsor reserves the right to suspend the study under the conditions specified in the 
clinical study protocol.  
10.5 Publication of study protocol and results 
The sponsor ensures that the key design content of the study protocol will be published 
in a publicly accessible database (e.g. clinicaltrials.gov.). In addition, the study results will 
be submitted for publication and/or published in a publicly accessible clinical trial database 
upon the completion of the study and the conclusion of the study report.  
10.6 Study documentation, record keeping and retention of documents 
The study center will retain the appropriate medical records and study records of the 
study. The study center will permit the competent authority to conduct reviewing, auditing 
and assessing of study safety and progress for quality assurance, and to review the clinical 
record (copy the clinical record as applicable to legal regulations).  
The original data are the information necessary for clinical studies to rebuild and 
evaluate the study, including original record of clinical findings, observation and other 
activities. Examples of these original documents and data records include, but are not limited 
to: hospital records, clinical and medical records, laboratory records, memos, patient diaries 
or assessment lists, drug relapse records, automated instrumental recordings, copies or 
transcripts with accuracy and completion validated, microfilms, negative films, microfilm 
strip and magnetic media records, X-ray recordings and patient documentation, and records 
kept by pharmacies, laboratories, and medical technological department participating in the 
study.  
Data must be collected by the researchers that are authorized by the investigator. The 
primary data collection tool for this study is the experiment case report form (eCRF). The 
investigator should ensure the accuracy, completeness, clarity and timeliness of the data in 
the eCRF and all other required reports. The data in the eCRF that is derived from the original 
document should be consistent with the original document and the inconsistent data should 
be interpreted. All data required in the eCRF must be recorded and all missing data must be 
interpreted. All changes or corrections to the eCRF must be confirmed by signing the date 


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and providing initials, and the initial records must be reflected in the original medical records. 
An audit trail record should be maintained in the system for the eCRF. The investigator 
should retain records of changes and corrections in the eCRF.  
The investigator/institution should maintain the study documentation in accordance with 
the requirements of the essential documents of the clinical study regulations and applicable 
legal regulations and/or guidelines. Investigators/institutions should take measures to 
prevent accidental damage or premature destruction of the documents.  
10.7 Confidentiality of study documents and patient records 
The investigator must ensure that the patients are anonymous and must not identify the 
patients by name in any of the documents submitted to other partners. The strict 
confidentiality of signing the ICFs and patient selection must be maintained so as to identify 
patients in the studycenter.  
10.8 Compensation and insurance 
10.8.1 Payment for transportation expenses, etc 
The costs involved in the observations and inspections during the therapy will be borne 
by the investigator. The study institution should pay the patient for transportation and other 
expenses after approval from the EC.  
10.8.2 Indemnity and insurance 
If the patient has suffered health damage due to AEs as a result of participating in the 
study, the investigator will provide compensation for the damage unless the health damage 
is caused by the patient’s himself. The investigator will take the necessary measures (such 
as insurance coverage) to undertake the relevant responsibilities. 
11. Protocol compliance 
The investigators declare that they will do their duty to avoid protocol deviations. The 
protocol should be amended if the investigator believes that deviating from the study 
protocol will improve the performance of the study. The protocol amendment can be 
implemented only after approving by EC. All major protocol deviations should be recorded 
and reported in the CSR.  
11.1 Study protocol amendment 
Any changes or additions to the study protocol should be documented in writing in the 
study protocol amendments, and must be approved by health authorities (if required) and the 
EC. Only amendments that are necessary for patient safety can be implemented prior to EC 
approval. Although approval is required for study protocol amendment, the investigator can 
take any required action immediately for the safety of the patients that are enrolled in the 


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study, even if the measure deviates from the study protocol. In this case, the investigator 
should inform the study center EC in time.  


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12. Appendix 
12.1 ECOG performance status 
0 
Fully active, able to carry on all pre-disease performance without restriction. 
1 
Restricted in physically strenuous activity but ambulatory and able tocarry out 
light work or sedentary activity, eg, light house work, office work. 
2 
Ambulatory and capable of all self-care but unable to carry out any workactivities. 
Up and about more than 50% of waking hours. 
3 
Only limited self-care ability, confined to bed or chair more than 
50% of waking hours. 
4 
Completely disabled. No self-care ability. Totally confined to 
bed or chair. 
5 
Dead. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 


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12.2  
RECIST 1.1 Guideline45 
Glossary  
CR  
Complete remission  
CSR  
Clinical study report  
CT  
Computed tomography  
DFS  
Disease free survival  
eCRF  
Electronic case report form  
FPFV  
The first visit of the first patient  
LPLV  
The last visit of the last patient  
MRI  
Magnetic resonance imaging  
OS  
Overall survival  
PD  
Progressive Disease  
PFS  
Progression free survival  
PR  
Partial remission  
RAP  
Report and analysis plan  
RECIST  
Remission evaluation criteria for solid tumor
SD  
Stability of disease  
SOD  
Sum of diameters  
TTF  
Time to treatment failure  
TTP  
Time to progress  
UNK  
Unknown  


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1 Background  
Omitted 
2 Objective  
Omitted  
3   Tumor measurability at baseline level 
3.1. Definitions 
At baseline, tumour lesions/lymph nodes will be categorized measurable or non-
measurable as follows: 
3.1.1. Measurable 
Tumour lesions: Must be accurately measured in at least one dimension (longest 
diameter in the plane of measurement is to be recorded) with a minimum size of: 
• 10 mm by CT scan (CT scan slice thickness no greater than 5 mm). 
• 10 mm caliper measurement by clinical exam (lesions which cannot be accurately 
measured with calipers should be recorded as non-measurable). 
• 20 mm by chest X-ray. 
• Malignant lymph nodes: To be considered pathologically enlarged and measurable, a 
lymph node must be P15 mm in short axis when assessed by CT scan (CT scan slice 
thickness recommended to be no greater than 5 mm). At baseline and in follow-up, only the 
short axis will be measured and followed. 
3.1.2. Non-measurable 
Other lesions include small lesions (longest diameter <10 mm or pathological lymph node 
short diameters ≥ 10 mm to <15 mm) and immeasurable lesions. Unmeasured lesions include: 
meningeal disease, ascites, pleural or pericardial effusion, inflammatory breast cancer, 
cancerous lymphangitis of the skin/lungs, abdominal mass that cannot be diagnosed and 
followed-up by imaging study, and cystic lesions.  
3.1.3. Special considerations regarding lesion measurability 
Bone lesions, cystic lesions, and lesions previously treated with local therapy require 
particular comment:  
Bone lesions: 
• Bone scan, PET scan or plain films are not considered adequate imaging techniques to 
measure bone lesions. However, these techniques can be used to confirm the presence or 
disappearance of bone lesions. 
• Lytic bone lesions or mixed lytic-blastic lesions, with identifiable soft tissue components, 
that can be evaluated by cross. sectional imaging techniques such as CT or MRI can be 
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• Blastic bone lesions are non-measurable. 
Cystic lesions: 
• Lesions that meet the criteria for radiographically defined simple cysts should not be 
considered as malignant lesions (neither measurable nor non-measurable) since they are, by 
definition, simple cysts. 
• ‘Cystic lesions’ thought to represent cystic metastases can be considered as measurable 
lesions, if they meet the definition of measurability described above. However, if noncystic 
lesions are present in the same patient, these are preferred for selection as target lesions. 
Lesions with prior local treatment: 
• Tumour lesions situated in a previously irradiated area, or in an area subjected to other 
loco-regional therapy, are usually not considered measurable unless there has been 
demonstrated progression in the lesion. Study protocols should detail the conditions under 
which such lesions would be considered measurable. 
3.2. Specifications by methods of measurements 
3.2.1. Measurement of lesions 
All measurements should be recorded in metric notation, using calipers if clinically 
assessed. All baseline evaluations should be performed as close as possible to the treatment 
start and never more than 4 weeks before the beginning of the treatment. 
3.2.2. Method of assessment 
The same method of assessment and the same technique should be used to characterize 
each identified and reported lesion at baseline and during follow-up. Imaging based 
evaluation should always be done rather than clinical examination unless the lesion(s) being 
followed cannot be imaged but are assessable by clinical exam. 
Clinical lesions: Clinical lesions will only be considered measurable when they are 
superficial and P10 mm diameter as assessed using calipers (e.g. skin nodules). For the case 
of skin lesions, documentation by color photography including a ruler to estimate the size of 
the lesion is suggested. As noted above, when lesions can be evaluated by both clinical exam 
and imaging, imaging evaluation should be undertaken since it is more objective and may 
also be reviewed at the end of the study. 
Chest X-ray: Chest CT is preferred over chest X-ray, particularly when progression is an 
important endpoint, since CT is more sensitive than X-ray, particularly in identifying new 
lesions. However, lesions on chest X-ray may be considered measurable if they are clearly 
defined and surrounded by aerated lung. 
CT, MRI: CT is the best currently available and reproducible method to measure lesions 
selected for response assessment. This guideline has defined measurability of lesions on CT 


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scan based on the assumption that CT slice thickness is 5 mm or less. As is described in 
Appendix II, when CT scans have slice thickness greater than 5 mm, the minimum size for 
a measurable lesion should be twice the slice thickness. MRI is also acceptable in certain 
situations (e.g. for bodyscans).  
Ultrasound: Ultrasound is not useful in assessment of lesion size and should not be used 
as a method of measurement. Ultrasound examinations cannot be reproduced in their entirety 
for independent review at a later date and, because they are operator dependent, it cannot be 
guaranteed that the same technique and measurements will be taken from one assessment to 
the next (described in greater detail in Appendix II). If new lesions are identified by 
ultrasound in the course of the study, confirmation by CT or MRI is advised. If there is 
concern about radiation exposure at CT, MRI may be used instead of CT in selected instances. 
Endoscopy, laparoscopy: The utilization of these techniques for objective tumour 
evaluation is not advised. However, they can be useful to confirm complete pathological 
response when biopsies are obtained or to determine relapse in studys where recurrence 
following complete response or surgical resection is an endpoint. 
Tumour markers: Tumour markers alone cannot be used to assess objective tumour 
response. If markers are initially above should be performed as close as possible to the 
treatment start and never more than 4 weeks before the beginning of the treatment. 
4. Tumour response evaluation 
4.1. Assessment of overall tumour burden and measurable disease 
To assess objective response or future progression, it is necessary to estimate the overall 
tumour burden at baseline and use this as a comparator for subsequent measurements. Only 
patients with measurable disease at baseline should be included in protocols where objective 
tumour response is the primary endpoint. Measurable disease is defined by the presence of 
at least one measurable lesion. In studies where the primary end point is tumour progression 
(either time to progression or proportion with progression at a fixed date), the protocol must 
specify if entry is restricted to those with measurable disease or whether patients having non-
measurable disease only are also eligible. 
4.2. Baseline documentation of ‘target’ and ‘non-target’ lesions 
When more than one measurable lesion is present at baseline all lesions up to a maximum 
of five lesions total (and a maximum of two lesions per organ) representative of all involved 
organs should be identified as target lesions and will be recorded and measured at baseline 
(this means in instances where patients have only one or two organ sites involved a 
maximum of two and four lesions respectively will be recorded).  
Lymph nodes merit special mention since they are normal anatomical structures which 


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may be visible by imaging even if not involved by tumour. As noted in Section 3, 
pathological nodes which are defined as measurable and may be identified as target lesions 
must meet the criterion of a short axis of P15 mm by CT scan. Only the short axis of these 
nodes will contribute to the baseline sum. The short axis of the node is the diameter normally 
used by radiologists to judge if a node is involved by solid tumour. Nodal size is normally 
reported as two dimensions in the plane in which the image is obtained (for CT scan this is 
almost always the axial plane; for MRI the plane of acquisition may be axial, sagittal or 
coronal). The smaller of these measures is the short axis. For example, an abdominal node 
which is reported as being 20 mm · 30 mm has a short axis of 20 mm and qualifies as a 
malignant, measurable node. In this example, 20 mm should be recorded as the node 
measurement All other pathological nodes (those with short axis P10 mm but <15 mm) 
should be considered non-target lesions. Nodes that have a short axis <10 mm are considered 
non-pathological and should not be recorded or followed. 
A sum of the diameters (longest for non-nodal lesions, short axis for nodal lesions) for 
all target lesions will be calculated and reported as the baseline sum diameters. If lymph 
nodes are to be included in the sum, then as noted above, only the short axis is added into 
the sum. The baseline sum diameters will be used as reference to further characterize any 
objective tumour regression in the measurable dimension of the disease. 
All other lesions (or sites of disease) including pathological lymph nodes should be 
identified as non-target lesions and should also be recorded at baseline. Measurements are 
not required and these lesions should be followed as ‘present’, ‘absent’, or in rare cases 
‘unequivocal progression’ (more details to follow). In addition, it is possible to record 
multiple nontarget lesions involving the same organ as a single item on the case record form 
(e.g. ‘multiple enlarged pelvic lymph nodes’ or ‘multiple liver metastases’). 
4.3. Response criteria 
This section provides the definitions of the criteria used to determine objective tumour 
response for target lesions. 
4.3.1. Evaluation of target lesions 
Complete Response (CR): Disappearance of all target lesions. Any pathological lymph 
nodes (whether target or non-target) must have reduction in short axis to <10 mm. 
Partial Response (PR): At least a 30% decrease in the sum of diameters of target lesions, 
taking as reference the baseline sum diameters. 
Progressive Disease (PD): At least a 20% increase in the sum of diameters of target 
lesions, taking as reference the smallest sum on study (this includes the baseline sum if that 
is the smallest on study). In addition to the relative increase of 20%, the sum must also 


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demonstrate an absolute increase of at least 5 mm. (Note: the appearance of one or more new 
lesions is also considered progression). 
Stable Disease (SD): Neither sufficient shrinkage to qualify for PR nor sufficient 
increase to qualify for PD, taking as reference the smallest sum diameters while on study. 
4.3.2. Special notes on the assessment of target lesions 
Lymph nodes. Lymph nodes identified as target lesions should always shave the actual 
short axis measurement recorded (measured in the same anatomical plane as the baseline 
examination), even if the nodes regress to below 10 mm on study. This means that when 
lymph nodes are included as target lesions, the ‘sum’ of lesions may not be zero even if 
complete response criteria are met, since abnormal lymphnodeis defined as having a short 
axis of <10 mm. Case report forms or other data collection methods may therefore be 
designed to have target nodal lesions recorded in a separate section where, in order to qualify 
for CR, each node must achieve a short axis <10 mm. For PR, SD and PD, the actual short 
axis measurement of the nodes is to be included in the sum of target lesions. 
Target lesions that become ‘too small to measure’. While on study, all lesions (nodal and 
non-nodal) recorded at baseline should have their actual measurements recorded at each 
subsequent evaluation, even when very small (e.g. 2 mm). However, sometimes lesions or 
lymph nodes which are recorded as target lesions at baseline become so faint on CT scan 
that the radiologist may not feel comfortable assigning an exact measure and may report 
them as being ‘too small to measure’. When this occurs, it is important that a value be 
recorded on the case report form. If it is the opinion of the radiologist that the lesion has 
likely disappeared, the measurement should be recorded as 0 mm. If the lesion is believed 
to be present and is faintly seen but too small to measure, a default value of 5 mm should be 
assigned (Note: It is less likely that this rule will be used for lymph nodes since they usually 
have a definable size when normal and are frequently surrounded by fat such as in the 
retroperitoneum; however, if a lymph node is believed to be present and is faintly seen but 
too small to measure, a default value of 5 mm should be assigned in this circumstance as 
well). This default value is derived from the 5 mm CT slice thickness (but should not be 
changed with varying CT slice thickness). The measurement of these lesions is potentially 
non-reproducible, therefore providing this default value will prevent false responses or 
progressions based upon measurement error. To reiterate, however, if the radiologist is able 
to provide an actual measure, that should be recorded, even if it is below 5 mm. 
Lesions that split or coalesce on treatment. when non-nodal lesions ‘fragment’, the 
longest diameters of the fragmented portions should be added together to calculate the target 
lesion sum. Similarly, as lesions coalesce, a plane between them may be maintained that 


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would aid in obtaining maximal diameter measurements of each individual lesion. If the 
lesions have truly coalesced such that they are no longer separable, the vector of the longest 
diameter in this instance should be the maximal longest diameter for the ‘coalesced lesion’. 
4.3.3. Evaluation of non-target lesions 
This section provides the definition soften criteria used to determine the tumour response 
for the group of non-target lesions. While some non-target lesions may actually be 
measurable, they need not be measured and instead should be assessed only qualitatively at 
the time points specified in the protocol. 
Complete Response (CR): Disappearance of all non-target lesions and normalization of 
tumour marker level. All lymph nodes must be non-pathological in size (<10 mm short axis). 
Non-CR/Non-PD: Persistence of one or more non-target lesion(s) and/or maintenance of 
tumour marker level above the normal limits. 
Progressive Disease (PD): Unequivocal progression (see comments below) of existing 
non-target lesions. (Note: the appearance of one or more new lesions is also considered 
progression). 
4.3.4. Special notes on assessment of progression of non-target disease 
The concept of progression of non-target disease requires additional explanation as 
follows: When the patient also has measurable disease. In this setting, to achieve 
‘unequivocal progression’ on the basis of the non-target disease, there must be an overall 
level of substantial worsening in non-target disease such that, even in presence of SD or PR 
in target disease, the overall tumour burden has increased sufficiently to merit 
discontinuation of therapy. A modest ‘increase’ in the size of one or more non-target lesions 
is usually not sufficient to quality for unequivocal progression status. The designation of 
overall progression solely on the basis of change in non-target disease in the face of SD or 
PR of target disease will therefore be extremely rare. 
When the patient has only non-measurable disease. This circus stance arises in some 
phase III studys when it is not acroterion study entry to have measurable disease. The same 
general concepts apply here as noted above, however, in this instance there is no measurable 
disease assessment to factor into the inter predation of an increase in non-measurable disease 
burden. Because worsening in non-target disease cannot be easily quantified (by definition: 
if all lesions are truly non-measurable) a useful test that can be applied when assessing 
patients for unequivocal progression is to consider if the increase in overall disease burden 
based on the change in non-measurable disease is comparable in magnitude to the increase 
that would be required to declare PD for measurable disease: i.e. an increase in tumour 
burden representing an additional 73% increase in ’volume’ (which is equivalent to a 20% 


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increase diameter in a measurable lesion). Examples include an increase in a pleural effusion 
from ‘trace’ to ‘large’, an increase in lymphangitic disease from localized to widespread, or 
may be described in protocols as ‘sufficient to require a change in therapy’. If ‘unequivocal 
progression’ is seen, the patient should be considered to have had overall PD at that point. 
While it would be ideal to have objective criteria to apply to non-measurable disease, the 
very nature of that disease makes it impossible to do so, therefore the increase must be 
substantial. 
4.3.5. New lesions 
The appearance of new malignant lesions denotes disease progression; therefore, some 
comments on detection of new lesions are important. There are no specific criteria for the 
identification of new radiographic lesions; however, the finding of a new lesion should be 
unequivocal: i.e. not attributable to differences in scanning technique, change in imaging 
modality or findings thought to represent something other than tumour (for example, some 
‘new’ bone lesions may be simply healing or flare of pre-existing lesions). This is 
particularly important when the patient’s baseline lesions show partial or complete response. 
For example, necrosis of a liver lesion may be reported on a CT scan report as a ‘new’ cystic 
lesion, which it is not. 
A lesion identified on a follow-up study in an anatomical location that was not scanned 
at baseline is considered a new lesion and will indicate disease progression. An example of 
this is the patient who has visceral disease at baseline and while on study has a CT or MRI 
brain ordered which reveals metastases. The patient’s brain metastases are considered to be 
evidence of PD even if he/she did not have brain imaging at baseline. 
If a new lesion is equivocal, for example because of its small size, continued therapy and 
follow-up evaluation will clarify if it represents truly new disease. If repeat scans confirm 
there is definitely a new lesion, then progression should be declared using the date of the 
initial scan. 
While FDG-PET response assessments need additional study, it is sometimes reasonable 
to incorporate the use of FDG-PET scanning to complement CT scanning in assessment of 
progression (particularly possible ‘new’ disease). New lesions on the basis of FDG-PET 
imaging can be identified according to the following algorithm: 
a. Negative FDG-PET at baseline, with a positive l FDG-PET at follow-up is a sign of PD 
based on a new lesion. 
B. No FDG-PET at baseline and a positive FDG-PET at follow-up: 
If the positive FDG-PET at follow-up corresponds to a new site of disease confirmed by 
CT, this is PD. 


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If the positive FDG-PET at follow-up is not confirmed as a new site of disease on CT, 
additional follow-up CT scans are needed to determine if there is truly progression occurring 
at that site (if so, the date of PD will be the date of the initial abnormal FDG-PET scan). 
If the positive FDG-PET at follow-up corresponds to a pre-existing site of disease on CT 
that is not progressing on the basis of the anatomic images, this is not PD. 
4.4. Evaluation of best overall response 
The best overall response is the best response recorded from the start of the study 
treatment until the end of treatment taking into account any requirement for confirmation. 
On occasion a response may not be documented until after the end of therapy so protocols 
should be clear if post-treatment assessments are to be considered in determination of best 
overall response. Protocols must specify how any new therapy introduced before progression 
will affect best response designation. The patient’s best overall response assignment will 
depend on the findings of both target and non-target disease and will also take into 
consideration the appearance of newlesions. Furthermore, depending on the nature of the 
study and the protocol requirements, it may also require confirmatory measurement (see 
Section 4.6). Specifically, in non-randomized trials where response is the primary endpoint, 
confirmation of PR or CR is needed to deem either one the ’best overall response’. This is 
described further below. 
4.4.1. Time point response 
It is assumed that at each protocol specified time point, a response assessment occurs. 
Table 1 on the next page provides a summary of the overall response status calculation at 
each time point for patients who have measurable disease at baseline.  
Table 1 Response of time point: Patients with Target Lesions (with/without non-target 
lesions)  


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When patients have non-measurable (therefore non-target) disease only, Table 2 is to 
be used. 
Table 2 Response of Time point-the Patients with only Non-target Lesions  
 
4.4.2. Missing assessments and unevaluable designation 
When no imaging/measurement is done at all at a particular time point, the patient is not 
evaluable (NE) at that time point. If only a subset of lesion measurements are made at an 
assessment, usually the case is also considered NE at that time point, unless a convincing 
argument can be made that the contribution of the individual missing lesion(s) would not 
change the assigned time point response. This would be most likely to happen in the case of 
PD. For example, if a patient had a baseline sum of 50 mm with three measured lesions and 


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at follow-up only two lesions were assessed, but those gave a sum of 80 mm, the patient will 
have achieved PD status, regardless of the contribution of the missing lesion. 
4.4.3. Best overall response: all time points 
The best overall response is determined once all the data for the patient is known. 
Best response determination in trials where confirmation of complete or partial response 
IS NOT required: Best response in these trials is defined as the best response across all time 
points (for example, a patient who has SD at first assessment, PR at second assessment, and 
PD on last assessment has a best overall response of PR). When SD is believed to be best 
response, it must also meet the protocol specified minimum time from baseline. If the 
minimum time is not met when SD is otherwise the best time point response, the patient’s 
best response depends on the subsequent assessments. For example, a patient who has SD at 
first assessment, PD at second and does not meet minimum duration for SD, will have a best 
response of PD. The same patient lost to follow-up after the first SD assessment would be 
considered unevaluable. 
Best response determination in trials where confirmation of complete or partial response 
IS required: Complete or partial responses may be claimed only if the criteria for each are 
met at a subsequent time point as specified in the protocol (generally 4 weeks later). In this 
circumstance, the best overall response can be interpreted as in Table 3. 
 
Table 3. Optimal Total Remission with CR and PR Effects to Be Confirmed  
 
4.4.4. Special notes on response assessment 
When nodal disease is included in the sum of target lesions and the nodes decrease to 
‘normal’ size (<10 mm), they may still have a measurement reported on scans. This 
measurement should be recorded even though the nodes are normal in order not to overstate 
progression should it be based on increase in size of the nodes. As noted earlier, this means 
that patients with CR may not have a total sum of ‘zero’ on the case report form (CRF). 
In trials where confirmation of response is required, repeated ‘NE’ time point assessments 


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may complicate best response determination. The analysis plan for the trials must address 
how missing data/assessments will be addressed in determination of response and 
progression. For example, in most trials it is reasonable to consider a patient with time point 
responses of PR-NE-PR as a confirmed response. 
Patients with a global deterioration of health status requiring discontinuation of treatment 
without objective evidence of disease progression at that time should be reported as 
‘symptomatic deterioration’. Every effort should be made to document objective progression 
even after discontinuation of treatment. Symptomatic deterioration is not a descriptor of an 
objective response: it is a reason for stopping study therapy. The objective response status 
of such patients is to be determined by evaluation of target and non-target disease as shown 
in Tables 1–3. 
Conditions that define ‘early progression, early death and in evaluability’ are study 
specific and should be clearly described in each protocol (depending on treatment duration, 
treatment periodicity). 
In some circumstances it may be difficult to distinguish residual disease from normal 
tissue. When the evaluation of complete response depends upon this determination, it is 
recommended that the residual lesion be investigated (fine needle aspirate/biopsy) before 
assigning a status of complete response. FDG-PET may be used to upgrade a response to a 
CR in a manner similar to a biopsy in cases where a residual radiographic abnormality is 
thought to represent fibrosis or scarring. The use of FDG-PET in this circumstance should 
be prospectively described in the protocol and supported by disease specific medical 
literature for the indication. However, it must be acknowledged that both approaches may 
lead to false positive CR due to limitations of FDG-PET and biopsy resolution/sensitivity. 
For equivocal findings of progression (e.g. very small and uncertain new lesions; cystic 
changes or necrosis in existing lesions), treatment may continue until the next scheduled 
assessment. If at the next scheduled assessment, progression is confirmed, the date of 
progression should be the earlier date when progression was suspected. 
4.5. Frequency of tumour re-evaluation 
Frequency of tumour re-evaluation while on treatment should be protocol specific and 
adapted to the type and schedule of treatment. However, in the context of phase II studies 
where the beneficial effect of therapy is not known, follow-up every 6–8 weeks (timed to 
coincide with the end of a cycle) is reasonable. Smaller or greater time intervals than these 
could be justified in specific regimens or circumstances. The protocol should specify which 
organ sites are to be evaluated at baseline (usually those most likely to be involved with 
metastatic disease for the tumour type under study) and how often evaluations are repeated. 


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Normally, all target and non-target sites are evaluated at each assessment. In selected 
circumstances certain non-target organs may be evaluated less frequently. For example, bone 
scans may need to be repeated only when complete response is identified in target disease 
or when progression in bone is suspected. 
After the end of the treatment, the need for repetitive tumour evaluations depends on 
whether the trial has as a goal the response rate or the time to an event (progression/death). 
If ‘time to an event’ (e.g. time to progression, disease-free survival, progression-free survival) 
is the main endpoint of the study, then routine scheduled re-evaluation of protocol specified 
sites of disease is warranted. In randomized comparative trials in particular, the scheduled 
assessments should be performed as identified on a calendar schedule (for example: every 
6–8 weeks on treatment or every 3–4 months after treatment) and should not be affected by 
delays in therapy, drug holidays or any other events that might lead to imbalance in a 
treatment arm in the timing of disease assessment. 
4.6. Confirmatory measurement/duration of response 
4.6.1. Confirmation 
In non-randomized trials where response is the primary endpoint, confirmation of PR 
and CR is required to ensure responses identified are not the result of measurement error. 
This will also permit appropriate interpretation of results in the context of historical data 
where response has traditionally required confirmation in such trials (see the paper by 
Bogaerts et al. in this Special Issue 10 ). However, in all other circumstances, i.e. in 
randomized trials (phase II or III) or studies where stable disease or progression are the 
primary end points, confirmation of response is not required since it will not add value to 
the interpretation of trial results. However, elimination of the requirement for response 
confirmation may increase the importance of central review to protect against bias, in 
particular in studies which are not blinded. 
In the case of SD, measurements must have met the SD criteria at least once after study 
entry at a minimum interval (in general not less than 6–8 weeks) that is defined in the study 
protocol. 
4.6.2. Duration of overall response 
The duration of overall response is measured from the time measurement criteria are first 
met for CR/PR (whichever is first recorded) until the first date that recurrent or progressive 
disease is objectively documented (taking as reference for progressive disease the smallest 
measurements recorded on study). The duration of overall complete response is measured 
from the time measurement criteria are first met for CR until the first date that recurrent 
disease is objectively documented. 


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4.6.3. Duration of stable disease 
Stable disease is measured from the start of the treatment (in randomized trials, from 
date of randomization) until the criteria for progression are met, taking as reference the 
smallest sum on study (if the baseline sum is the smallest, this is the reference for calculation 
of PD). 
The clinical relevance of the duration of stable disease varies in different studies and 
diseases. If the proportion of patients achieving stable disease for a minimum period of time 
is an endpoint of importance in a particular trial, the protocol should specify the minimal 
time interval required between two measurements for determination of stable disease. Note: 
The duration of response and stable disease as well as the progression-free survival are 
influenced by the frequency of follow-up after baseline evaluation. It is not in the scope of 
this guideline to define a standard follow-up frequency. The frequency should take into 
account many parameters including disease types and stages, treatment periodicity and 
standard practice. However, these limitations of the precision of the measured endpoint 
should be taken into account if comparisons between studys are to be made. 
4.7. Progression-free survival/proportion progression-free 
4.7.1. Phase II trials 
This guideline is focused primarily on the use of objective response endpoints for phase 
II trials. In some circumstances, ‘response rate’ may not be the optimal method to assess the 
potential anticancer activity of new agents/regimens. In such cases ‘progression-free 
survival’ (PFS) or the ‘proportion progression-free’ at landmark time points, might be 
considered appropriate alternatives to provide an initial signal of biologic effect of new 
agents. It is clear, however, that in an uncontrolled trial, these measures are subject to 
criticism since an apparently promising observation may be related to biological factors such 
as patient selection and not the impact of the intervention. Thus, phase II screening trials 
utilizing these endpoints are best designed with a randomized control. Exceptions may exist 
where the behavior patterns of certain cancers are so consistent (and usually consistently 
poor), that a non-randomized trial is justifiable. However, in these cases it will be essential 
to document with care the basis for estimating the expected PFS or proportion progression-
free in the absence of a treatment effect. 
 
 
12.3 irRC evaluation criteria 
Experts from the International Association of Immunotherapy have formally put forward 
the new immune-related response criteria in Clinical Cancer Research in 2009, which is 


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known as irRC.  
SPD (the product of two maximum vertical diameters of the measurable lesion)is 
adopted in irRC to estimate the size of tumor. Different from the respective WHO assessment 
standard, which assess every lesion separately; irRC uses the total tumor burdens of all 
measurable lesions. Tumor burden is defined as the sum of SPD of all lesions, including 
SPD of new lesions.  
Measurable lesions of irRC are defined as lesions greater than 5mm*5mm on CT scan, 
with maxium 5 lesions occuring on each visceral organ, no more than 10 lesions on all 
visceral organs, and no more than 5 lesions presenting on body surface.  
Effect assessment of irRC: The curative effect evaluation in irRC is based on the increase 
or decrease in the overall tumor burden at the observation time as compared with the time 
of baseline assessment, confirmed by repeating the comparison between two consecutive 
operation time points intervened by at least 4 weeks, and classified.  
Specifically, it is divided into four categories as follows:  
irCR - all the lesions disappear completely;  
irPR - in the continuous detections, tumor burden decreases by more than or equal to 50% 
compared with the baseline tumor burden;  
irSD - inconformity to irCR and irPR standard, and no appearance of irPD;  
irPD –tumor burden increases by more than or equal to 25% compared with the baseline 
tumor burden.  
Comparison between irRC and Traditional WHO Standard  
Item  
WHO  
irRC  
Newly 
found 
measurable lesion 
(for 
example, 
≥5mm*5mm)  
Always indicates rogression disease 
(PD)  
The overall tumor burden 
should be included to assess 
whether it is PD  
Newly 
found 
ummeasurable 
lesion 
(for 
example, 
≤5mm*5mm)  
Always indicates rogression disease 
(PD)   
Not defined as PD  
CR 
Disappearance of all the lesions is 
confirmed 
at 
two 
consecutive 
observation points with an interval 
of at least 4 weeks. 
 


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Gene Edited T-Cell Therapy for Advanced Lung Cancer. Version 1.9 
 
PR 
The diameters of all the measurable 
lesions decreasing more than 50% 
compared to the baseline, which are 
confirmed 
at 
two 
continuous 
observation points with an interval 
of at least 4 weeks. No new lesion or 
other 
disease 
development 
is 
observed.  
The overall tumor burden 
decreasing more than 50% 
(irPR), compared to the 
baseline, is confirmed at 
two continuous observation 
points with an interval of at 
least 4 weeks.  
SD 
The lesion diameter increasing 
detected 
at 
two 
continuous 
observation points is less than 25%. 
No new lesion or other disease 
development is observed.  
As 
confirmed 
at 
two 
continuous 
observation 
points, the overall tumor 
burden decreasing is less 
than 50% or increasing is 
less 
than 
25% 
(irSD), 
compared to the baseline.  
PD 
The lesion diameter increasing ≥ 
25% compared to baseline is 
detected at any observation point. 
And (or) new lesion and (or) other 
disease development are observed. 
The overall tumor burden 
increasing ≥ 25% (irPD), 
compared to the baseline, is 
confirmed at any time 
between the two continuous 
observation points with an 
interval of at least 4 weeks.  
In accordance to irRECIST, if a primary PD is identified by tumor imaging the tumor 
should be revaluated after at least four weeks to confirm the PD. During the period from 
identification to confirmation of PD, if the patient agrees to continue the cell infusion therapy, 
the patient may continue to receive the cell infusion treatment. If the next imaging 
examination is scheduled within 4 weeks, it is not necessary for the patients examined by 
confirmation tumor imaging to take the next appointed tumor imaging test.  If the clinical 
conditions of the patient are stable, the tumor imaging examination can be re-initiated at an 
appointed time point and continued until PD is verified by imaging assessment.  Given that 
tumors may burst within the first several months after the initial cell infusion therapy and 
then relieve in some patients, patients are allowed to continue the cell infusion therapy after 
the first presence of PD until the next PD presents. For the patients with unstable clinical 
conditions, it is not necessary to repeat the imaging examination to confirm PD. Tumor 
bursting includes: 


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Gene Edited T-Cell Therapy for Advanced Lung Cancer. Version 1.9 
 
• The original target lesion worsens 
• The original non-target lesion worsens 
• The origin of new lesions 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 


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Gene Edited T-Cell Therapy for Advanced Lung Cancer. Version 1.9 
 
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Design of the ARCTIC Study. Clin Lung Cancer 2016; 17:232-+. 
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Life Sci 2016; 59:468-79. 
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in B cell acute lymphoblastic leukemia. Science translational medicine 2014; 6:224ra25. 
36. Weber JS, Yang JC, Atkins MB, Disis ML. Toxicities of Immunotherapy for the Practitioner. Journal 
of clinical oncology: official journal of the American Society of Clinical Oncology 2015; 33:2092-9. 
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the blood. Nature reviews Clinical oncology 2013; 10:472-84. 
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DNA with broad patient coverage. Nature medicine 2014; 20:548-54. 
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England journal of medicine 2016; 374:1864-73. 
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Cancer Ther 2015; 14:847-56. 
41. Deng L, Gyorffy B, Na F, et al. Association of PDCD1 and CTLA-4 Gene Expression with 
Clinicopathological Factors and Survival in Non-Small-Cell Lung Cancer: Results from a Large and 
Pooled Microarray Database. J Thorac Oncol 2015; 10:1020-6. 
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Nat Rev Cancer 2015; 15:311-6. 
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Patient-Derived Xenografts from Ultrasound-Guided Transbronchial Needle Aspirates. PloS one 2015;10. 
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89 
 
Amendments 
Table of Changes (Version 1.5, Oct 10, 2016) 
Section 
No. 
Section 
Title 
Description of Change (s) 
Rationale 
7.1.2; 
7.1.4.2 
Screening; 
Day 8 of 
each cycle 
The following sentences were added: 
Collection of blood for PBMC expansion: 60-80 ml. The actual blood volume is confirmed according to the 
routine blood test result of the patient at that time. 
According to protocol design 
for details. 
7.2.3.5  
 
In vivo T 
cell tracking 
The following sentences were added: 
• Baseline and Day 12 of each treatment cycle  
•  After end of treatment: the 28st day of last treatment cycle, three months, six months, and one year after 
the last cell infusion.  
• 
 Blood sampling volume: 10 ml each time.  
Notes: the treatment of different groups may vary from two to four cycles; therefore, the number of blood 
sampling collection points are: eight (two cycles), 10 (three cycles), or 12 (four cycles). 
In order to track the edited T 
cells in vivo. 
13.5.17 
Off-target 
effect 
Off-target will be assessed for edited cells before the infusion of patient. 
Updated to the following sentence: 
Blood sample with 10 ml was collected for assessment of off-targeton baseline, end of study and unscheduled 
timepoint based on investigator ajudgement. 
A consideration of safety. 
 
 
 
 


 
 
 
90 
 
Table of Changes (Version 1.6, Jan 24, 2017) 
 
Section 
NO. (s) 
Section Title 
(s) 
Description of Change (s) 
Rationale 
All 
sections 
General 
Replaced “21 Days/Cycle” with “28 Days/Cycle” 
Updated protocol about the start 
of the time of T cell culture. 
Protocol 
summary, 
3.1; 
5.1.2.1 
Trial and 
design 
 
The underlined words were added:  
Pre-A: 2x107 PD-1 edited T cells per kilogram of body weight, after the first treatment cycle, observe 28 Days 
and then evaluate the patient; if the patient does not have DLTs, one more treatment cycle will be allowed. 
Group A: 1x107 PD-1 edited T cells per kilogram of body weight  
2 treatment cycles; 
Group B: 2x107 PD-1 edited T cells per kilogram of body weight 
2 treatment cycles; 
Group C: 4x107 PD-1 edited T cells per kilogram of body weight  
2 treatment cycles; 
According to the national 
regulation of “Guidelines for 
Research and Assessment of 
Cell Product” published in Jan 
2017. 
2.3 
Primary 
endpoint 
The underlined wordswere added:  
The rate of dose-limiting toxicity (DLT) of the cell therapy during the period of the first treatment cycle. 
To clarify the observed period 
of DLT. 
3.1.1.1 
Molecular 
pre-
screening  
PD-L1 detection will be further confirmed with the positive PD-L1 staining by Roche Ventana PD-L1 (SP142, 
SP263), which is performed in the pathology department of the trial center with Leica bondmax automatic 
immunohistochemistry machine. 
The following text was added: 
After the tumor being confirmed to be PD-L1 positive, the patient can sign the ICF, and the screening/baseline 
visit can be initiated. 
Remarks: If the patients have been confirmed to have PD-L1 positive tumors through staining with Roche 
Ventana PD-L1 test (SP142, SP263) and Dako PD-L1 test (Dako28-8, Dako22c3) prior to their participation in 
the screening for this clinic trial, they candirectly sign the ICF for the trial, and the screening/baseline visit can be 
To clarify that the enrolled 
patient was permitted to confirm 
by any PD-L1 test mentioned in 
protocol, because there are four 
standarded PD-L1 detest 
technology according to the 
literature. 


 
 
 
91 
 
initiated.  
Protocol 
summary; 
4.1 
Inclusion 
criteria 
 
4. NSCLC, stage IIIB/IV confirmed by histology or cytology; All patients must be screened for EGFR gene 
mutation and ALK gene fusion; For the patients with a positive EGFR mutation or ALK gene fusion, treatment 
failure of EGFR-TKI or ALK-TKI must have been experienced.   
8.  Life expectancy ≥ 3 months. 
Updated the protocol 
Protocol 
summary; 
4.2 
Exclusion 
criteria 
 
12. Patients receivedprior therapy with an anti-PD-1, anti-PD-L1, anti-PD-L2, anti-CD137 or anti-CTLA-4 
antibodies. 
Available to screen patients 
5.1.2.1 
Cell 
infusion 
procedure  
Cell infusion procedure #6 was revised: 
6. Cell infusion duration: At the first time of cells infusion to the patient, the drops at the beginning should be 
adjusted to about 15-20 drops/min for 10-15min to evaluate safety. If adverse events are not observed, the 
infusion speed will be adjusted to around 60 drops/min. For the elderly patients and the patients with cardio-
pulmonary function insufficiency, the infusion speed should be slow down with close monitoring to the 
patient. The speed of infusion can be adjusted accordingly.  
For more standardization. 
5.3.2 
Tentative 
dose  
The initial dose of treatment group A: 1x107 PD-1 edited T cells per kilogram of body weightin total.  
A dose of PD-1 edited T cells splited into three doses of 20%, 30%, and 50% will be conducted on the 1st, 3rd, 
and 5th day of each cycle, 28 days/cycle. Patients will be treated for a total of two cycles. Furthermore, the cell 
dose will gradually increase to 2.0x107 and 4.0x107 PD-1 edited T cells per kilogram of body weight per cycle in 
the next two treatment groups.  
For details. 
5.3.7 
Definition 
of DLT 
Unless otherwise specified, NCI CTCAE4 .03 will be applied to assess the toxicity. DLT is defined as the AE or a 
laboratory abnormal value that is probably attributable to the T cell infusion. An abnormal laboratory value can 
be attributed to the T cell infusion if it happens within one cycle after the first cell infusion (≤ 28 days) or occurs 
in thefirst cycle but is later confirmed in the second cycle, and meets any criterion of the following conditions 
For details. 
6.4.3.3 
Disposal 
The following text was added: 
For details. 


 
 
 
92 
 
and 
destruction 
 
If the cells are not completely infused for some special reason, the residual cells should be sealed, packed and 
frozen/refrigerated for storage strictly according to the operation standards, and further disposal conditions 
should be documented clearly.   
7 
Visit 
schedule 
and 
assessments  
 
Visit schedule and assessments #1, #2, #4 and #5 were revised: 
1.Deleted “a. Questionnaires for the patient - EORTC QLQ-C30 and EORTC QLQ-LC13” of the section 
screening; 
2. The term of lymphocyte subpopulation was changed from “CD56” to “NK”. 
4. The total volume of the blood collection for PBMC expansion was revised:  
Group A: 40ml; Group B/C: 60-80ml. 
based on the hematology examination results of the patient at the time. 
5. The cytokines that are tested were added as follow: IL-6, IL-10, CRP, TNF-α, ESR, IL-8, IL-2, Fractalkine, 
IFN-r, MCP-1, IL-5, GM-CSF and FLT3L. 
This is phase I study with small 
size of patients, questionnaires 
had not meaning. The text was 
updated to further clarify and to 
add more cytokines as 
exploratoryindexes. 
7.2.2.5 
Laboratory 
assessments 
 
The following text was added: 
Cytokine level monitoring: IL-6、IL-10、CRP、TNF-α、ESR、IL-8、Fractalkine、 IFN-r、MCP-1、IL-5、 
GM-CSF、FLT3L (plasma samples have been collected, and IL-8, IL-2, Fractalkine, IFN-r, MCP-1, IL-5, GM-
CSF, FLT3L are all exploratory indexes which will be tested at the end of trial.) 
According to protocol design for 
details. 
7.2.3 
Biomarker 
 
The following text was added: 
*Notes: A total of 15 ml peripheral blood sample is collected at the same time point for ctDNA and immune 
repertoire; 10 ml peripheral blood sample is collected at the time point for immune repertoire only. 
10ml blood could meet the 
detect of ctDNA and immune 
repertoire, when the detect of 
ctDNA and immune repertoire 
were at the same time point. 
7.2.3 
Biomarker 
 
The following text was added: 
Flow cytometery analysis of regulatory T (Treg) cells, memory T cells, activated T cells and PD1 expression. 
In order to explore the dynamic 
change of regulatory T (Treg) 


 
 
 
93 
 
Blood collection time points: before pretreatment with cyclophosphamide (blank baseline), after pretreatment 
with cyclophosphamide (medication baseline), 24 h after the third cell infusion of each cycle, the end of each 
cycle (day 21), week eight, three months, and six months after the last infusion. Blood sampleing collection 
volume: 5 ml each time.  
cells, memory T cells, activated 
T cells and PD1 expression after 
T cell infusion. 
 


 
 
 
94 
 
Table of Changes (Version 1.7, June 05, 2017) 
 
Section 
Number 
(s) 
Section Title (s) 
Description of Change (s) 
Rationale 
7; 7.1.2 
Visit schedule 
and assessments: 
Table 7-1 
Flowchart of 
Trial Visits; 
Screening 
The blood collection for PBMC pre culture 
According to protocol design. 


 
 
 
95 
 
Table of Changes (Version 1.8, Jan 30, 2018) 
 
Section 
Number 
(s) 
Section Title (s) 
Description of Change (s) 
Rationale 
Home 
page 
Protocol Title 
A single-arm, open, prospectivephase I clinical trial of PD-1 knockout engineered T cells treating patients 
with advanced non-small cell lung cancer 
 
Protocol 
summary; 
2.2 
Secondary 
objective 
2) The potential immunological indexes and biomarkers. 
Update to: 
2) To investigate the changes of the potential immunological indexes and biomarkers. 
3) To track the PD-1 KO T cells in patients and investigate the correlation of the clinical response. 
Update to the protocol 
Protocol 
summary; 
2.4 
Secondary 
endpoint 
 4) To evaluate the duration of the first confirmed stable disease (SD) (the time from the first confirmed SD to 
the first PD or death caused by any factor). 
The PFS is secondary endpoint. 
Protocol 
summary; 
4.2 
Exclusion 
criteria 
13. The patients who accepted the anti-tumor vaccines or otherimmunostimulatory anti-tumor drugs. 
The primary end point of the 
clinical trial was safety. 
3.1 
Description of 
the study 
design 
 
Pre-A: Patients will receive A dose of 2×107PD-1 edited T cells per kilogram of body weight splited into three 
doses of 20%, 30%, and 50% will be conducted on the 1st, 3rd, and 5th day of each cycle. After finishing the 
first cycle and resting for 28 days, the patients will be allowed to receive another cycle of transfusion. 
Around 20 patients with advanced PD-L1 positive NSCLC that show progression after the multiline 
treatments will be enrolled in this dose escalation trial. Patients will be assigned into three groups, with a 
treatment cycle of 28 days. 
The text was updated for more 
precise. 


 
 
 
96 
 
3.1; 5.0 
Summary 
schema of the 
trial design  
Summary schema of the trial design was updated:
 
 
5.1.1 
Pretreatment 
 
Add the underlined words in text: 
A single dose (20 mg/kg) of cyclophosphamide will be administrated as a pre-treatment intending  intending 
for  lymphocytes depletionon the third day before the cell transfusionof the first treatment cycle.  
The clear definition of 
application of 
cyclophosphamide in study. 
5.1.2.1 
Treatment group 
 
 
1) If the patient decides to discontinue the cell infusion therapy, the patient will be followed upaccording to 
scheduled visits (7 visits and evaluation) until end of the study. Additionally, the patient has the right to 
choose other clinical therapeutic strategies to treat the cancer.  
If the patient agrees to continue the cell infusion therapy, she orhe can receive the cell infusion therapy until 
the end of the study. Based on the irRECIST criteria, if a primary PD is identified through tumor imaging, 
another evaluation should be conducted after at least four weeks to confirm the PD status. During the period 
from identification to confirmation of the PD, if the patient’s general condition does not worsen from the 
baseline (ECOG：0-2 scores), or no intolerable therapeutic side effects/other conditions that hold for further 
cell infusion therapy occur, and the patient agrees to continue the cell infusion therapy, the patient may 
continue to receive the cell infusion treatment. If the next imaging examination is scheduled around four 
The text was updated based on 
irRECIST. 


 
 
 
97 
 
weeks, it is not necessary for the patient to take an additional imaging test.  If the clinical conditions of the 
patient are stable, the tumor imaging examination can be re-initiated at an appointed time point and continued 
until PD is verified through imaging assessment. Given that tumors may flare within the first several months 
after the initial cell infusion therapy and then shrink in some patients, patients are allowed to continue the cell 
infusion therapy after the first presence of PD until the confirmation of next PD. For the patients with unstable 
clinical conditions, it is not necessary to repeat the imaging examination to confirm PD. Tumor flares 
includes: 
• 
Worsening of the original target lesion 
• 
Worsening of the original non-target lesion  
• 
Identification of new lesions 
5.3.5 
Dose escalation 
of the patients  
The escalation dose is from 1x107 to 2x107 and 4x107 PD-1 edited T cells per kilogram of body weight in the 
three treatment groups A, B, and C. Engineered T-cells splitting into three applications (20%, 30% and 50%) 
will be conducted on the 1st, 3rd, and 5th day of each cycle. The patient must be able to tolerate the lowest 
dose, and the effect of the prior dose has been assessed, the patient can receive the sequential treatments.  
To appropriately reflect the 
protocol. 
5.5 
Supportive 
therapy 
The cell therapy treated patients may as recommended by the investigators, be treated with some appropriate 
supportive therapies, based on the guidelines for immune-related advert events (irAE). 
To appropriately reflect the 
protocol. 
6.4.1 
Package and 
label of the trial 
cells 
Package and label of the trial cells was updated as follow: 
 


 
 
 
98 
 
 
8. 
AEs 
 
Abnormal values of laboratory tests: 
The laboratory abnormalities with significance (clinically significant, inducing clinical signs or symptoms, 
requiring concomitant treatment or adjustment of the trail, treatment) should be recorded as AEs, with an 
emphasis on the diagnosis results instead of the signs and symptoms (for example, anemia other than low 
hemoglobin). The laboratory abnormal values that conform the AE should be followed up until those AEs 
recove stable. If an abnormal laboratory value or test result shows the same signs/symptoms as a reported AE, 
it should not be recorded as aseperate AE. 
Investigator should follow-up the patients that exit the study due to AEs, to observe and record the outcomes 
of any AEs until such events are completely relieved or stable. Investigatormust judge whether the AE is 
related to the treatment, the measures taken after AE is observed (no action, discontinuation of the trial 
treatment, temporary interruption for medicine, permanent termination of the trial treatment, unknown, etc.), 
and the application of concomitant treatment (with/without concomitant treatment), based on the supporting 
evidence. Investigator should also evaluate the outcomes of the AE (disappear/not disappear, recovered, in 
recovery, whether there are sequelae, death, unknown, etc.), and determine whether it is an SAE. 
Add the precisedescription 
about AEs. 


 
 
 
99 
 
9. 
Data 
monitoring 
committee 
The independent data monitoring committee will be set up, consisting of 3 independent medical consultants 
who are responsible for overseeing the safety of the trial. According to the analysis of available clinical and 
laboratory data, the committee may suggest the trial to be continued, revised, and/or suspended, based on the 
data. In the event of some new safety issues, especially a serious incident (such as death) during the trial or a 
safety notification regarding the trial treatment, a meeting shall be convened upon the request of the sponsors. 
The independent data 
monitoring committee in our 
hospital did not set up at that 
time. This phase I study has 
been supervised by Clinical 
Research Administrative Board, 
West China Hospital. 
9.4.4.1 
Variable  
Safety analysis varibles include the incidence of dose limiting toxicities (DLTs) in 3 week and during the 
treatment  
Safety analysis varibles include the incidence of dose limiting toxicities (DLTs) in the first cycle of treatment, 
incidence rate and severity of AEs and SAEs including changes in laboratory parameters, vital signs, and 
ECG results.  
Definition of DLT was changed 
in protocol. 
9.4.4.2 
Safety analysis 
The total observation period can be divided into three phases:  
1. Pre-treatment period: From the day that the patient signs the ICF to the day before the first trial treatment is 
given  
2. Treatment period period: From the day that first trial treatment is given to the day that 150 days after the 
last dose is given  
3. Post-treatment period: From the day that 151 days after the last dose is given. 
The description of content was 
repeated. 
10.4.6.2 
Relationship 
between the 
clinical factors 
and therapeutic 
effect  
Clinical factors include the pathogenesis, pre-therapeutic regimens, and patient performance status. Conduct a 
tentative trial by graphics on the relationship between efficacy indexes and clinical factors. Depending on 
specific circumstances, conduct further trial using statistical modeling if any potential relationship is 
identified. Specific analysis will be recorded in a separate report. Describe any additional data analysis 
depending on specific circumstances in a separate analysis-planning document. 
This is an evaluation of safty 
other than efficacy 
9.4.8 
Sample size 
In the dose escalation trial, one dose group will enroll three to six evaluable patients receiving the 
The text was updated in case of 


 
 
 
100 
 
 
calculation  
 
recommended dose 
DLT. 
13.4 
Safe treatment 
process on tumor 
immunology 
drugs 
13.4Safe treatment process on tumor immunology drugs 
We figure out a brochure 
regarding AE treatment to 
replace the description. 


Table of Changes (Version 1.9, Dec 04, 2018) 
Section 
Number 
(s) 
Section Title 
(s) 
Description of Change (s) 
Rationale 
3.1 
Description of 
the study 
design  
The following text was added: 
This clinical study was a first-in-human trial using CRISPR technology knockouting PD-1 gene in T 
cell to treat lung cancer patients. Although there were no exact dosage/regimen and safety data, 
starting dose was selected referring to cell therapy or CART treatment for considering benefit-risk 
of cancer patients (see section 5.3.1 Starting dose rationale). Pre-A: 2x107 PD-1 edited T cells per 
kilogram of body weight (percentage of PD-1 knockout T cell may vary depending on technology 
efficiency). The study defines the Pre-A group as a preliminary exploratory dose group. The Pre-A 
group has a 28 days-observation after the first treatment cycle. If no DLT was observed in Pre-A 
group during the defined observation period, the study will continue other treat groups according to 
provisional dosage; If DLT or other unexpected toxicity occurs in the Pre-A group, the investigator 
will adjust the cell dose and interval of sequential cell therapy based on data from Pre-A group. 
More precise 
description about the 
dose of Pre-A group 
and give a clear 
interpret the choosing 
dose of 2x107 PD-1 
edited T cells per 
kilogram of body 
weight. 
5.5.1  
Cytokine 
release 
syndrome 
(CRS) 
Cytokine release storm (CRS) 
Corrected: Cytokine release 
syndrome. 
7.1.1; 
7.2.3.3 
Molecular 
screening; 
Screening of 
the tumor 
biomarkers 
The underlined is added: 
PD-L1 status detection was used by fresh tissue or paraffin-embedded tissue. Before the molecular 
screening, the candidate patients are asked to sign the ICF for molecular screening. 
 


 
 
 
102 
 
8.1.2 
Report 
An SAE should be reported in the patients with an unknown condition of PD-L1 after the ICF (on 
molecular screening) is signed. Based on the evaluation by investigator, the event can be reported as 
an SAE only if it is suspected to have a causative relationship to the trial process (such as traumatic 
operation or biopsy). An SAE should be followed up until it subsides, or a clinically significant 
improvement or stabilization is observed.  
For those patients with a known PD-L1 condition who have signed the main ICF, no matter whether 
the screening fails or not, the reporting of an SAE will start from the time when the main ICF has 
been signed. 
The deletion was replaced by underline content for concise: 
An SAE should be reported in the patients after the ICF (on molecular screening) is signed. For 
those patients who have signed the main ICF, no matter whether the screening fails or not, the 
reporting of an SAE will start from the time when the main ICF has been signed.  
To ensure the safety of patients, any SAE that occurs during the time when the ICF is signed to the 
28th day after the patients terminate the treatment should be reported, no matter whether it is 
suspected to be related to the study treatment or not. 
To accurately report the 
initiation and termination of 
SAE. 
12.2 
RECIST 1.1
 
 
Guideline 48  
 
The following text was added: 
Eisenhauer EA. Et al. New response evaluation criteria in solid tumours: revised RECIST guideline 
(version 1.1). Eur J Cancer. 2009 Jan;45(2):228-47 
 


 
Gene Edited T-Cell Therapy for Advanced Lung Cancer.  
 
Final statistical analysis plan  
(Version 2, May 04, 2018) 
 


 
 
 
104 
 
Table of contents 
 
1 
Introduction ................................................................................................................ 107 
2 
Study objectives and study design ............................................................................. 107 
2.1 Primary objective ............................................................................................................. 107 
2.2 Secondary objectives ....................................................................................................... 107 
2.3 Study design .................................................................................................................... 107 
3 
Subjects and treatments .............................................................................................. 108 
3.1 Analysis sets .................................................................................................................... 108 
3.2 Treatment groups ............................................................................................................. 109 
4 
Statistical analysis ...................................................................................................... 109 
4.1 Patient disposition ........................................................................................................... 109 
4.2 Demographic and baseline characteristics ....................................................................... 110 
4.3 Medical History ............................................................................................................... 110 
4.4 Antineoplastic therapy ..................................................................................................... 110 
4.5 Concomitant therapy........................................................................................................ 111 
4.6 Study medication ............................................................................................................. 111 
4.7 Safety evaluation ............................................................................................................. 111 
4.7.1 Adverse events............................................................................................ 112 
4.7.2 Laboratory test abnormalities ..................................................................... 114 
4.7.3 Vital signs and weight ................................................................................ 114 
4.7.4 Electrocardiogram (ECG) ........................................................................... 114 
4.7.5 Other safety data ......................................................................................... 115 
4.8 Efficacy evaluation .......................................................................................................... 115 
4.8.1 Best overall response (BOR) ...................................................................... 115 
4.8.2 Overall response rate (ORR) and disease control rate (DCR) .................... 115 
4.8.3 Time to response (TTR) and duration of response (DOR) ......................... 115 
4.8.4 Progression free survival (PFS) and overall survival (OS) ........................ 115 
4.9 Pharmacokinetic (PK) evaluations .................................................................................. 115 
4.10 Immunological factors and biomarkers ........................................................................... 116 
4.11 Interim analyses ............................................................................................................... 116 
4.12 Determination of sample size .......................................................................................... 117 
5 
General definitions ...................................................................................................... 117 
5.1 Assessment windows, baseline and post baseline definitions, missing data handling .... 117 
5.1.1 Date of first administration of study drug/treatment .................................. 117 
5.1.2 Date of last administration of study drug/treatment ................................... 117 
5.1.3 Study day, baseline, study visit and window .............................................. 117 
5.2 Details of demographic and baseline characteristics analyses......................................... 118 


 
 
 
105 
 
5.3 Details of efficacy analyses ............................................................................................. 118 
5.3.1 Overall response rate (ORR) ...................................................................... 118 
5.3.2 Disease control rate (DCR) ........................................................................ 118 
5.3.3 Time to response (TTR) ............................................................................. 118 
5.3.4 Duration of response (DOR) ...................................................................... 119 
5.3.5 Progression free survival (PFS) .................................................................. 119 
5.3.6 Overall survival (OS) ................................................................................. 119 
5.3.7 Date of last adequate tumor assessment ..................................................... 119 
5.3.8 Construction of waterfall plot ..................................................................... 119 
5.3.9 Clinical progression .................................................................................... 120 
5.4 Imputation rule of partial or missing dates ...................................................................... 120 
5.5 General presentation of descriptive summaries ............................................................... 120 


 
 
 
106 
 
List of abbreviations 
Abbreviation 
Detail 
AE 
Adverse Event 
ATC 
Anatomic Therapeutic Chemical (Classification) 
BMI 
Body Mass Index 
CI 
Confidence Interval 
CR 
Complete Response 
CTC 
Common Terminology Criteria 
CTCAE 
Common Terminology Criteria for Adverse Events 
DCR 
Disease Control Rate 
DLT 
Dose Limiting Toxicity 
DOR 
Duration of Response 
ECOG 
Eastern Cooperative Oncology Group 
eCRF 
Electronic Case Report/Record Form 
FAS 
Full Analysis Set 
LLOQ 
Lower Limit of Quantification 
MedDRA 
Medical Dictionary for Regulatory Activities 
NSCLC 
Non-Small Cell Lung Cancer 
ORR 
Overall Response Rate 
OS 
Overall Survival 
PAS 
Pharmacokinetic Analysis Set 
PD 
Progressive Disease 
PFS 
Progression Free Survival 
PR 
Partial Response 
PT 
Preferred Term 
RAP 
Report Analysis Plan 
RECIST 
Response Evaluation Criteria in Solid Tumors 
SAE 
Serious Adverse Event 
SOC 
System Organ Class 
SS 
Safety Set 
TTR 
Time to response 
 
 
 
 
 
 
 
 
 
 


 
 
 
107 
 
1 Introduction 
This document provides detailed statistical methodology for the analysis of data from study 
MHC-001. 
Data will be analyzed according to the data analysis section 9 of the study protocol version 
1.8.  Important information is given in the following sections. 
2 Study objectives and study design 
2.1 Primary objective 
The primary objective is to investigate the safety and tolerability of autologous T cells with 
PD-1 knocked out by CRISPR-CAS9-OPT (PD-1 KO T cell) in patients with PD-L1 positive 
advanced non-small cell lung cancer (NSCLC) who have progressed after multiple-line 
standard treatment. 
2.2 Secondary objectives 
• To preliminarily investigate the anti-tumor activity of PD-1 KO T cell infusion therapy 
• To investigate the changes in the potential immunological factors and biomarkers 
related to PD-1 KO T cell infusion therapy 
• In vivo tracking of PD-1 KO T cells in patients and investigate the mechanism of the 
clinical response 
2.3 Study design 
This is an investigator-initiated Phase I open-label dose escalation study. Patients were those 
having PD-L1 positive NSCLC who have progressed after multiple-line standard treatment.  
Patients in each dose cohort would receive two cycles of treatment (28-day cycle) as planned, 
and could continue treatment in agreement with the investigator and patient if the treatment 
was deemed beneficial and tolerable, as long as there is no evidence of disease progression, 
intolerable toxicity, withdrawal of consent, or discontinuation for any other reason.  Patients 
will be continually reassessed for evidence of toxicity.  


 
 
 
108 
 
Each dose cohort was planned to enroll and treat 3 patients. A clinical synthesis of the 
available toxicity information, including dose limiting toxicity (DLTs) and adverse events 
(AEs) other than DLTs in and post Cycle 1, pharmacokinetic (PK) data, efficacy information 
and other relevant data, would be reviewed by the investigator to determine whether to 
escalate, stay or deescalate for the next cohort.   
3 Subjects and treatments 
This study enrolled patients with histologically/cytologically confirmed advanced NSCLC, 
who were refractory to currently available therapies or had tumors for which no effective 
treatment was available, and had PD-L1 positive in tumor molecular test. 
The study included the following planned dose cohorts: 
• Pre-A testing cohort: 2x107/kg (one cycle of interruption between the first treatment 
cycle and the second treatment cycle) 
• Cohort A: 1x107/kg 
• Cohort B: 2x107/kg 
• Cohort C: 4x107/kg 
Each dose was split into 3 infusions: 20%, 30% and 50% of the total dose on D1, D3 and D5 
of each cycle, respectively.  
Statistical analyses results will be presented by treatment groups specified according to dose 
level unless otherwise specified. 
3.1 Analysis sets 
The analysis sets to be used are defined as below.  
Screening Set: The Screened Set consists of all patients who have signed informed 
consent/assent and screened in the study.  
Full Analysis Set(FAS): The FAS consists of all patients to whom study treatment has been 
assigned, and have received at least one infusion of PD-1 KO T cells.  


 
 
 
109 
 
Safety Set (SS): The Safety Set consists of all patients who received at least one infusion of 
PD-1 KO T cells and had at least one valid post-baseline safety assessment. The statement 
that a patient has no adverse events (on the Adverse Events eCRF) constitutes a safety 
assessment.  
Patients will be classified according to the study treatment they actually received, where 
treatment received is defined as (i) the treatment assigned (dose-level and schedule planned) 
if it was received at least once, or (ii) the total dose received when starting therapy if assigned 
dose level is never received completely. Each patient will be classified into and analyzed 
consistently within one (and only one) treatment group. 
Pharmacokinetic Analysis Set (PAS): The PAS consists of all patients in FAS who have 
at least one sample providing evaluable PK data.  
3.2 Treatment groups 
Dose levels investigated during the study will be considered to identify treatment groups. 
For example, the following may be considered: 1x107/kg, 2x107/kg, and 4x107/kg.  
4 Statistical analysis 
Data will be summarized using descriptive statistics (continuous data) and/or contingency 
tables (categorical data) for demographic and baseline characteristics, efficacy 
measurements, safety measurements, and all relevant pharmacokinetic measurements. 
Unless otherwise described, all data will be listed appropriately by using FAS. 
4.1 Patient disposition 
The FAS will be used for the following summaries. 
The number and percentage of patients who were still on-treatment at the date of data cut-
off and who discontinued the study will be summarized by treatment group. Among patients 
who discontinued the treatment, primary reason for the discontinuation will also be 
summarized. 


 
 
 
110 
 
Number and percentage of patients in each analysis set will be summarized by treatment 
group.  
4.2 Demographic and baseline characteristics 
The FAS will be used for all baseline and demographic summaries demographic 
characteristics include age and sex. Baseline characteristics include weight, height, body 
mass index (BMI) and ECOG performance score at baseline. Disease history and 
characteristics include primary site of cancer, time from most recent relapse/progression to 
first PD-1 KO T-cells administration in month, histological grade, predominant 
histology/cytology, stage at initial diagnosis, types of lesions at baseline (target only/non-
target only/both target and non-target/unknown).   
In addition, the following category for age will be used to calculate the number and 
percentage of patients included in each category: 
• 
Age: < 65 and ≥ 65 years. 
4.3 Medical History 
The FAS will be used for the following summaries and listings. 
Relevant medical histories and current medical condition will be listed and summarized by 
primary system organ class, preferred term and treatment group. Medical history/current 
medical conditions will be coded using Medical Dictionary for Regulatory Activities 
(MedDRA) terminology. 
4.4 Antineoplastic therapy 
The FAS will be used for the following summaries and listings. 
Prior antineoplastic therapy will be listed in three separate listings: medications, 
radiotherapy, and surgery. 


 
 
 
111 
 
The summary of prior antineoplastic medication will include the total number of regimens 
used, and setting, best response, and reason for discontinuation at last medication. All prior 
antineoplastic medications will be summarized by ATC class, preferred term and treatment 
group. 
The summary of prior antineoplastic radiotherapies will include locations and setting at last 
radiotherapy. 
The summary of prior antineoplastic surgeries will include the procedure and residual 
disease at last surgery. 
Antineoplastic therapies since discontinuation of study treatment will be tabulated by 
treatment group, and will be listed by patient. 
4.5 Concomitant therapy 
The safety set will be used for the followingsummaries and listings. 
Concomitant medicationswill be listed and summarized by ATC class and preferred term. 
These summaries will include medications starting on or after the start of study treatment or 
medications starting prior to the start of study treatment and continuing after the start of 
study treatment. Any prior medications starting and ending prior to the start of study 
treatment will be listedand summarized by ATC class and preferred term.  
4.6 Study medication 
The study drug dosing information will be listed by treatment group for the safety set. 
4.7 Safety evaluation 
The primary objective is to characterize the safety and tolerability of PD-1 KO T cell 
infusion in patients with PD-L1 positive NSCLC. Therefore, the primary analysis will 
include the summary of observed DLTs by treatment group, and the summary of the 
frequency, severity of AEs, changes in laboratory findings, physical examinations, vital 
signs, ECGs. The safety set will be used for all safety evaluation. 


 
 
 
112 
 
The number and percentage of patients with DLTs will be tabulated by system organ class 
(SOC), preferred term (PT) and treatment group. All observed DLTs will be listed. 
The assessment of safety is based mainly on the type and frequency of adverse events (AEs) 
and on the number of laboratory values that fall outside of pre-determined ranges (Common 
Toxicity Criteria [CTC] grading limits or normal ranges as appropriate). Other safety data 
(e.g., electrocardiogram, vital signs, and special tests) will be considered as appropriate. 
The safety summary tables will include assessments collected during on-treatment period 
starting from the date of the first study treatment administration until 28 days after the end 
of the last treatment cycle. Those collected during the pre-treatment and post-treatment 
periods are to be flagged. 
4.7.1 Adverse events 
Definition of adverse event and reporting 
An AE is defined as the appearance of (or worsening of any pre-existing) undesirable sign(s), 
symptom(s), or medical condition(s) that occur after patient’s signed informed consent for 
the main study (or for molecular screening for the document unavailable patient in the 
expansion part) has been obtained. Abnormal laboratory values or test results occurring after 
informed consent constitute AEs only if they induce clinical signs or symptoms, are 
considered clinically significant, require therapy (e.g., hematologic abnormality that 
required transfusion or hematological stem cell support), or require changes in study 
medication(s). 


 
 
 
113 
 
Analyses of AEs 
All AEs recorded during the study will be listed. Treatment-related AEs will be summarized. 
Adverse events will be coded using the Medical Dictionary for Regulatory Activities 
(MedDRA) terminology. Severity of AE will be coded based on the National Cancer 
Institute’s Common Terminology Criteria for Adverse Events (CTCAE 4.03). If CTCAE 
grading does not exist for an adverse event, the severity of mild, moderate, severe, and life-
threatening, corresponding to Grades 1 to 4, will be used. CTCAE Grade 5 (death) will not 
be used in this study; rather, information about deaths will be collected in the Death eCRF 
page.  
The following AEs will be summarized: 
• DLTs by SOC, PT and treatment group 
• AEs regardless of study drug relationship by SOC, PT and treatment group 
• AEs with suspected relationship to study drug by SOC, PT and treatment group 
• SAEs regardless of study drug relationship by SOC, PT and treatment group 
• SAEs with suspected relationship to study drug by SOC, PT and treatment group 
• Grade 3 or 4 AEs regardless of study drug relationship by SOC, PT and treatment 
group 
• Grade 3 or 4 AEs with suspected relationship to study drug by SOC, PT and treatment 
group 
• AEs leading to study drug discontinuation regardless of study drug relationship by 
SOC, PT and treatment group 
• AEs regardless of study drug relationship by PT and treatment group 
• Deaths by SOC, PT and treatment group 
All AEs, SAEs, AEs leading to study drug discontinuation, and deaths will be listed. 


 
 
 
114 
 
4.7.2 Laboratory test abnormalities 
All laboratory values will be graded by using CTCAE 4.03. Parameters for which a 
grading does not exist will be classified into low/normal/high group by means of 
laboratory normal ranges. 
Analyses of laboratory values 
The following summarieswill be produced for laboratory parameters in hematology, 
biochemistry, and urinalysis: 
• Shift table using CTCAE grade to compare baseline to the worst post-baseline value 
• For laboratory tests where CTCAE grades are not defined, shift table using the 
low/normal/high/(lowand high) classification to compare baseline to the worst post-
baseline value 
The following listings will be produced for all laboratory parameters: 
• Listing of all laboratory data with values flagged to show the corresponding CTCAE 
grades and the classifications relative to the laboratory normal ranges 
4.7.3 Vital signs and weight 
All vital signs including systolic blood pressure, diastolic blood pressure and pulse and 
weight will be listed.  
The number and percentage of patients with abnormal vital signs and weight change will be 
provided. Patients with abnormal vital signs will also be listed. 
4.7.4 Electrocardiogram (ECG) 
ECG data will be listed. Patients with abnormal ECG values will be listed.  


 
 
 
115 
 
4.7.5 Other safety data 
Data from other tests will be listed, notable values will be flagged, and any other 
information collected will be listed as appropriate. 
4.8 Efficacy evaluation 
Tumor response will be defined as per RECIST v1.1, and refer to the irRC. 
The FAS will be used for the following analyses. 
4.8.1 Best overall response (BOR) 
BOR will be summarized by treatment group. The waterfall plot will be used to depict anti-
tumor activity. Details of waterfall plot will describe in section 5.3.8.  
4.8.2 Overall response rate (ORR) and disease control rate (DCR) 
Definitions of ORR and DCR are described in sections 5.3.1 and 5.3.2. 
ORR and DCR will be presented as point estimate (at each treatment group) and 
corresponding 95% exact confidence interval according to Clopper-Pearson method. 
4.8.3 Time to response (TTR) and duration of response (DOR) 
Definitions of TTR and DCR are described in sections 5.3.3 and 5.3.4. Due to the small 
sample size, for patients who achieved best overall response of CR or PR, the TTR and DOR 
will be listed. 
4.8.4 Progression free survival (PFS) and overall survival (OS) 
Definitions of PFS and OS are described in sections 5.3.5 and 5.3.6. Due to the small sample 
size. The Kaplan-Meier method will be used to estimate overall median PFS and median OS. 
PFS and OS will be presented by patient in listings.  
4.9 Pharmacokinetic (PK) evaluations 
Patients in the PAS with at least one evaluable measurement of PD-1KO T cell concentration 
will be included in the PK analysis.  


 
 
 
116 
 
The concentration of PD-1KO T cells in blood will be listed and summarized by treatment 
group and time point. Descriptive statistics include arithmetic mean and geometric mean, 
median, standard deviation, arithmetic coefficient of variation (CV), geometric coefficient 
of variation, minimum and maximum. In geometric mean calculation, concentration 0 is not 
included.  
Patient level and averaged concentration of PD-1KO T cell measures will be displayed using 
longitudinal plots. Graphical presentation of arithmetic mean (±SD) concentrations at each 
scheduled time point will be provided by treatment group. 
4.10 Immunological factors and biomarkers 
Baseline level of PD-L1 will be listed, and may also graphed against clinical response status 
using strip plots.  
Soluble immune and inflammatory cytokines (e.g. IL-10, interferon gamma, IL-6, IL-6 
receptor, CRP, and ferritin) will be listed by patient and summarized by treatment group and 
time point. Baseline and absolute and relative change (percent and or fold change) from 
baseline will be calculated for each treatment group and time point and summarized using 
sample size, mean, standard deviation, median, minimum and maximum. If both the baseline 
and post baseline values are below LLOQ, absolute, percent and fold change from baseline 
will not be imputed and reported as missing. The maximum change from baseline measure 
for each cytokine may also graphed against clinical response status and AEs of special 
interest using strip plots. Patient level and averaged cytokine measures and percent change 
from baseline may be displayed using longitudinal plots.  
TCR diversity and bTMB were compared between patients with and without disease control 
benefit using the exact Wilcoxon test. Statistical significance was set at p ≤ 0.05 (2-sided). 
4.11 Interim analyses 
Not applicable. 


 
 
 
117 
 
4.12 Determination of sample size 
No formal statistical power calculations to determine sample size were performed for this 
study. Cohorts of at least 3 evaluable patients per dose level will be enrolled.  
5 
General definitions 
5.1 Assessment windows, baseline and post baseline definitions, missing 
data handling 
5.1.1 Date of first administration of study drug/treatment 
The date of first administration of study drug/treatment will be derived as the first date when 
a non-zero dose of study drug is administered and recorded on the dose administration record 
eCRF. 
The date of first administration of study treatment is also referred to as start of study 
treatment. 
5.1.2 Date of last administration of study drug/treatment 
The date of last administration of study drug/treatment is defined as the last date and time 
when a non-zero dose of study drug is administered and recorded on the dose administration 
record eCRF. 
5.1.3 Study day, baseline, study visit and window 
Study day 
The study day for all assessments (both efficacy and safety) is calculated as the difference 
between the date of the event (visit date, onset date of an event, assessment date, etc.) and 
the start of study treatment. The first day of study treatment is study day 1.Note that for this 
study there is no study day 0, so the day immediately prior to study day 1 is study day -1. 
For any events on or after the first dose of study drug, study day is calculated as: event date 
– date of first administration of study treatment + 1. As such, the first dose date was study 
day 1.  


 
 
 
118 
 
For any events before the first dose date, study day is calculated as: event date – date of first 
administration of study treatment. As such, one day before first dose date was study day -1.  
Baseline 
For comparison against baseline (e.g. for laboratory parameters), baseline is considered to 
be the last available assessment or value before or at the date of first study drug 
administration. Allowance for the baseline visit is within 28 days prior to first dosing.  
Patients with no data on a particular parameter before first treatment will be deemed to have 
a missing baseline for that parameter. 
5.2 Details of demographic and baseline characteristics analyses 
BMI will be calculated as follows: 
• BMI (kg/m2) = Weight (kg) / [Height (m)]2. 
Time from [Date A] to [Date B] in month will be calculated as follows: 
• ([Date B] – [Date A] + 1) / 30.4375. 
Therefore, times such as time since initial diagnosis of primary cancer to first study drug 
administration in month will be calculated based on the method above. Demographic 
characteristics will be summarized by dose level.  
5.3 Details of efficacy analyses 
5.3.1 Overall response rate (ORR) 
ORR is defined as the proportion of subjects with a best overall response of CR or PR.  
5.3.2 Disease control rate (DCR) 
DCR is defined as the proportion of subjects with a best overall response of CR, PR or SD. 
5.3.3 Time to response (TTR) 
TTR is defined as the duration from the date of first PD-1 KO T cell infusion to the date 
when the response criteria of CR or PR is first met. 


 
 
 
119 
 
5.3.4 Duration of response (DOR) 
DOR is defined as the duration from the date when the response criteria of CR or PR is first 
met to the date of relapse or death due to underlying cancer. 
5.3.5 Progression free survival (PFS) 
PFS is the duration from date of first PD-1 KO T cell infusion to the date of relapse or death 
due to any reason.  
5.3.6 Overall survival (OS) 
OS is the duration from date of first PD-1 KO T cell infusion to the date of death due to any 
reason.  
5.3.7 Date of last adequate tumor assessment 
The last adequate tumor assessment is the last assessment with a result other than unknown. 
The last assessment is the assessment made before an event or a censoring occurred.   
Note that tumor assessments made after commencement of a new antineoplastic therapy will 
not be used in the assessment of time related efficacy endpoints. 
5.3.8 Construction of waterfall plot 
The waterfall plot will be used to depict anti-tumor activity. It displays the best percentage 
change from baseline in the sum of the longest diameter of all target lesions for each patient. 
To distinguish treatment group, different bar pattern will be used by treatment group.It will 
be based on the investigator reported overall lesion/radiological response. 
Patient with missing/unknown best percentage change from baseline will be represented by 
a special symbol (e.g.,$) in the waterfall plot. Patient with unknown/missing best percentage 
change from baseline and with unknown overall response will be excluded. 
The following display will be used (from left to right): 
1. Bars under the horizontal axis representing tumor shrinkage 
2. Bars above the horizontal axis representing tumor growth 


 
 
 
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3. “Zero” bars with stars ($) symbol for patient with missing best percentage change 
5.3.9 Clinical progression 
Standard RECIST CRF pages do not allow for the capture of clinical progression. Clinical 
progression can only be reported in the treatment discontinuation form by using the “Disease 
progression” reason for drug discontinuation. 
By default, the clinical progression will not be considered in the efficacy analyses based on 
RECIST. 
5.4 Imputation rule of partial or missing dates 
As a general rule, when a date will be recorded as a partial date, the missing day will be 
imputed to the 15th of the month (e.g., DEC2007 imputed to 15DEC2007), and if the day 
and month will be both missing then to July 1st of that year (e.g., 2007 imputed to 
01JUL2007).  
5.5 General presentation of descriptive summaries 
Categorical data will be summarized by frequency count and percentages. Percentages will 
be calculated using the number of subjects in the relevant population as the denominator. 
Continuous data will be summarized by appropriate descriptive statistics (i.e. mean, standard 
deviation, median, minimum and maximum). 
All data will be listed appropriately. 
The imputed dates are used only to allow the analysis of partial data, and are not displayed 
in data listings.