Recently, Hebei Provincial Market Supervision Administration organized and formulated the "Technical Specifications for Clinical Operation of Cellular Immunotherapy." This document specifies relevant terms and definitions, basic requirements, risk plans, implementation plan formulation, implementation and operation, treatment of toxic and side effects, patient follow-up, etc. This document is applicable to medical institutions and cell preparation institutions that implement cellular immunotherapy. The specification document will be implemented on February 21, 2021.
Today, the Stem Cell Official Account released the full text of this specification document for reference by counterparts in the cell therapy industry.

Technical specification for clinical operation of cellular immunotherapy

Foreword

This document was drafted in accordance with the rules given in GB/T1.1-2020 "Guidelines for Standardization Work Part 1: Structure and Drafting Rules of Standardization Documents". Please note that some of the contents of this document may involve patents. The issuing agency of this document is not responsible for identifying these patents. This document was proposed by Hebei Provincial Health Commission. This document was drafted by: Nongfuyu Pharmaceutical Hebei Co., Ltd., Hebei Provincial People's Hospital, Hebei Institute of Cell Biotechnology. The main drafters of this document: Cai Jianhui, Li Qingxia, Du Pingping, Zhang Mengya, Deng Xinna, Gao Fei, Zhou Ye, Cai Ziqi. This document is issued for the first time.

Introduction
In recent years, cellular immunotherapy has gradually been applied to the clinic, but the current domestic and provincial clinical operating technical specifications for cellular immunotherapy are still blank. In order to adapt to the development needs of the cellular immunotherapy industry, strengthen the quality control and safety management of the clinical application of cellular immunotherapy, standardize the clinical operation technical process and risk control of cellular immunotherapy, ensure the rights and safety of patients, and promote international and domestic cooperation in the same industry. For communication, it is necessary to formulate technical specifications for the clinical operation of cellular immunotherapy.
 
1 Scope This document specifies relevant terms and definitions, basic requirements, risk plans, formulation, implementation and operation of the implementation plan, treatment of toxic and side effects, patient follow-up, etc. This document is applicable to medical institutions and cell preparation institutions that implement cellular immunotherapy. 2 Normative references There are no normative references in this document. 3 Terms and definitions The following terms and definitions apply to this document. 3.1 Cell preparation institutions cellpreparationinstitution Enterprises, institutions or laboratories with cell preparation professionals, GMP-like cell culture rooms, professional facilities and equipment conditions and qualifications. 3.2 Medical institutions Medicalinstitution A hospital or department that has the personnel, venues, facilities and equipment required for cellular immunotherapy and meets the national qualifications. 3.3 Peripheral blood mononuclear cells peripheralbloodmononuclearcell; PBMC blood has mononuclear cells, mainly including lymphocytes, monocytes, macrophages, dendritic cells, NK cells and a small number of other types of cells. 3.4 Raw material T cell rawmaterialT cell is derived from autologous PBMC, T lymphocytes used to prepare effector cells after separation and/or sorting; or from allogeneic, used for preparation after histocompatibility antigen matching and/or gene editing T lymphocytes of effector cells. 3.5 Effector cells are lymphocytes that are obtained by in vitro processing, preparation, culture and expansion, and capable of non-specific or specific recognition and killing of target cells. 3.6 Chimeric antigen receptor T cells chimericantigenreceptorTcells; CAR-T modified by genetic engineering, which can express the introduced CAR gene containing antigen recognition fragments, T cell receptor activation molecules, costimulatory signals and other signal molecules such as effector T cells. 3.7 T cell receptor chimeric type T cell T cell receptor chimeric type Tcells; TCR-T

 Modified by genetic engineering, it can express the introduced TCR gene that can recognize the specific MHC antigen complex, and the MHC-limited effector T cell. 3.8 Tumor infiltrating T lymphocytes tumorinfiltrating lymphocytes; specific tumor infiltrating T lymphocytes isolated and amplified from TIL tumor tissue. 3.9 cytotoxic T lymphocytes cytotoxic Tlymphocytes; tumor-specific cytotoxic T lymphocytes activated by mature DC cells loaded with tumor antigens by CTL. 3.10 Therapeutic Dendritic Cell Vaccine (Therapeutic DC Vaccine) Therapeutic DCvaccine ImmatureDCcells are mature DC cells (MatureDCcells) prepared by culturing and expanding and loading tumor antigens. 3.11 Natural killer cells natural killer cells; NKNK cells are non-limiting and non-specific MHC natural killer cells that are cultured and differentiated with the participation of cytokines such as IL-2, IL-12, IFN-α and LR. 3.12 Dendriticcell-cytokine-induced killercells: DC-CIKCD3 monoclonal antibody and a variety of cytokine-induced non-specific killer cells co-cultured with DC cells to obtain a group of heterogeneous cells. Note: For non-specific cellular immunotherapy. However, if co-cultured with antigen-loaded DC cells, the DC-CIK cells obtained have partial specific killing characteristics. 3.13 Transduction The process of introducing foreign genetic material (DNA or RNA) into cells for stable expression by means of viral or non-viral vectors. 3.14 Gene editing gene editing uses genetic engineering technology to knock out specific genomes of cell chromosomes, or to perform random or site-specific gene modification into target genomes. 3.15 Final cell product refers to the final cell culture or harvest made from raw cells through the whole process of in vitro preparation, culture and expansion. 3.16 Label A mark, comment or bar code pasted or attached to an object to distinguish different objects. 3.17 GMP-like cell culture room GMP-likecell laboratory has a cell culture room with more than 10,000-level and localized 100-level specialized cell culture facilities and equipment. 3.18KPS score Kamofskyscore refers to the patient's physical function status score. Note: Generally, a score of 80 or more is considered to be non-dependent, 50 to 70 is considered to be semi-dependent, and a score of less than 50 is considered to be dependent. 3.19 ECOG score Zubrod-ECOG-WHOscore

 Refers to the patient's physical status and treatment tolerance score. Note: Generally, 0~3 points can tolerate treatment, but 4 points or more cannot tolerate treatment. 3.20 Lymphocyte deletion chemotherapy lymphocytedeletionchemotherapy refers to the use of specific chemotherapeutic drugs to perform chemotherapy for patients with non-myeloid lymphocyte deletion. 3.21 Preconditioning refers to the preconditioning of patients with specific drugs for the purpose of enhancing the therapeutic effect or preventing toxic and side effects. 4 Basic requirements
4.1 Personnel requirements
 
4.1.1 The medical institution (hereinafter referred to as the institution) should be equipped with a team of medical staff suitable for its scale. The physician team should include at least three physicians, and at least one physician has senior professional title qualifications (associate senior and above); the nursing team should include at least four nurses, and at least one nurse has intermediate qualifications or above.
4.1.2 All members of the medical care team should have received professional training in cellular immunotherapy, and at least one or more physicians or nurses should have a professional training certificate in cellular immunotherapy.
4.1.3 The institution should have a team of experts for emergency treatment of cellular immunotherapy. The team members should at least include experts in cardiovascular medicine, respiratory medicine, hematology, and ICU and have expert qualifications (associate senior and above), and have multidisciplinary emergency response. Early warning mechanism for handling.
4.2 Institutional qualifications
 
4.2.1 Institutions that implement cellular immunotherapy should have Class III A or equivalent hospital qualifications.
4.2.2 Institutions should have the professional conditions for multidisciplinary emergency treatment required for cellular immunotherapy.
4.2.3 The institution should have an ethics committee, and each cellular immunotherapy technology or product must be ethically identified before it can be implemented in the institution.
4.3 Site, facilities, equipment
 
4.3.1 The institution shall have the premises and facilities that meet the requirements of cellular immunotherapy, including wards and facilities, medical units and facilities, nursing units and facilities, etc.
4.3.2 The organization should have the equipment conditions for emergency treatment, including oxygen inhalation equipment, sputum suction equipment, ECG monitoring equipment, auxiliary breathing equipment, etc.
4.3.3 The organization should have the necessary venues, facilities and equipment for the rescue of critical illnesses.
 5 Risk plan
5.1 The institution shall formulate corresponding risk plans for the implementation of each cellular immunotherapy technology or product.
5.2 The risk plan shall formulate corresponding treatment measures on the basis of comprehensive consideration of the various risks that may occur during the entire implementation process of the implemented technology or product, so as to ensure the safety of patients during the entire implementation process.
 6 Formulation of the implementation plan
6.1 The institution shall formulate a corresponding implementation plan for each cellular immunotherapy technology or product to be implemented to ensure the safety and effectiveness of each treatment.

 
6.2 The formulation of the implementation plan should fully take into account the characteristics of the technology or product implemented, and comply with relevant regulations and guidelines such as the "Administrative Measures for Clinical Research and Transformation Application of Somatic Cell Therapy", "Technical Guidelines for Research and Evaluation of Cell Therapy Products".
 7 Implementation and operation
7.1 Patient evaluation
 
7.1.1 The evaluation of patients with autologous cellular immunotherapy includes evaluation before peripheral blood PBMC collection and evaluation before reinfusion therapy:
a) The content of patient assessment before PBMC collection includes but is not limited to:
1) No active infectious diseases;
2) Cardiopulmonary function, liver function, and kidney function are basically normal;
3) KPS score>50 or ECOG score below 3;
4) White blood cell (WBC) count>4.0×109/L, lymphocyte count>2.0×109/L, hemoglobin (HGB)>90g/L, platelet (PLT) count>100×109/L; bilirubin ≤1.5 Times the upper limit of normal value, alanine aminotransferase (ALT) ≤ 2 times the upper limit of normal value, and aspartate aminotransferase (AST) ≤ 2 times the upper limit of normal value;
5) Infectious disease test: hepatitis B surface antigen, hepatitis B E antigen, antibody, hepatitis B core antibody, hepatitis C (HCV) antibody, AIDS (HIV) antibody, syphilis antibody are all negative. Those who are positive for any of them need to be specially marked and notify the cell preparation agency to prepare cells in an independent preparation space to prevent cross-contamination.
b) The content of patient evaluation before reinfusion treatment includes but not limited to:
1) No infectious disease;
2) Cardiopulmonary function, liver function, and kidney function are basically normal;
3) KPS score>50 or ECOG score below 3 points.
7.1.2 The evaluation of allogeneic cell immunotherapy includes evaluation of T cell donors and evaluation of patients before reinfusion:
a) The evaluation of raw material T cell donors should strictly follow the "Technical Guidelines for Cell Therapy Product Research and Evaluation" and "CAR-T Cell Therapy Product Quality Control Testing Research and Non-clinical Research Considerations" and other relevant regulations;
b) The content of patient assessment before reinfusion treatment is the same as above.
 
7.2 Collection and transportation of peripheral blood PBMC
 
7.2.1 The blood collection department approved by the health administrative department should use an automatic blood component separator to set up a mononuclear cell separation program to collect peripheral PBMC-enriched blood; or use qualified medical equipment to collect patient peripheral blood through a vein.
7.2.2 The collected peripheral blood PBMC should be treated with anticoagulation.
7.2.3 The transfer of peripheral blood PBMC should be delivered to a cell preparation facility equipped with a GMP-like cell culture room within 3 hours at 4°C.
 
7.3 Preparation and transportation of effector cell preparations
 
7.3.1 The preparation of effector cell preparations should be completed in a cell preparation facility equipped with a GMP-like cell culture room.
7.3.2 The preparation process of different effector cell preparations must strictly follow the "Technical Guidelines for the Research and Evaluation of Cell Therapy Products (Trial)", "Immune Cell Preparation Quality Management Standards", and "CAR-T Cell Preparation Quality Management Standards" (Draft for Solicitation of Comments)" and other norms or guidelines are implemented.
7.3.3 Each batch of effector cell preparations shall undergo strict period inspection and release inspection for quality control. The quality control indicators shall include but not limited to: cell number, cell viability, cell phenotype, functional molecule expression, cytokine secretion Function, in vitro killing function, pH value, osmotic pressure, endotoxin, rapid sterility test, bacterial and fungal culture test, mycoplasma culture test, harmful residue test, and each batch of cell preparation samples should be frozen for traceability.
7.3.4 The final product of effector cell preparations can adopt two methods: fresh cell preparations or frozen cell preparations.
7.3.5 The transport of effector cell preparations shall adopt the following methods:
a) Fresh effector cell preparations should be transported to the ward at 4°C. The transfer to reinfusion treatment generally does not exceed 12 hours;
b) The effector cell preparations cryopreserved in liquid nitrogen should be transported in a gas-phase liquid nitrogen tank to avoid the risk of liquid nitrogen overflow or contamination of the frozen storage bag rupture.
 
7.4 Cell handover
 
7.4.1 After the cells are transported to the ward, the full-time medical staff and transport personnel need to check, inspect and hand over the cell preparations together.
7.4.2 Check items include, but are not limited to: the patient's name, gender, hospitalization number, cell batch number, production date, release inspection result, signature of the inspector and other information shown in the cell packaging bag (bottle) mark. The inspection items include but are not limited to: whether the cell packaging bag (bottle) is damaged, leaking, leaking, and whether the content is abnormal.
7.4.3 If any problems are found, immediately contact the cell preparation institution for re-check; if there are any uncertain factors, it should be returned to the cell preparation institution and discarded immediately.
7.4.4 When transferring cell preparations from multiple patients, special care should be taken to avoid confusion.
7.4.5 Fresh cell preparations should be temporarily stored at 4°C during the time period from the completion of handover to reinfusion treatment, and the room temperature should not exceed 30 minutes; cryopreserved cell preparations should be placed as soon as possible during the time period from the completion of handover to reinfusion treatment The liquid nitrogen tank should be frozen and stored at room temperature for no more than 30 minutes after re-thawing.
7.5 Lymphocyte-depleting chemotherapy preconditioning
 Commonly used regimen is cyclophosphamide 300mg/m2～400mg/m2 intravenous infusion once a day, on the 1st to 2nd day; fludarabine 20mg/m2～25mg/m2 intravenous infusion 1 time per day, on the 1st to 4th day day. The purpose is to reduce the level of immunosuppressive cells and factors in the tumor microenvironment through the deletion of non-myeloid lymphocytes to improve clinical efficacy. Cell reinfusion therapy should be implemented within 2 to 7 days after pretreatment.
7.6 Preparation before reinfusion treatment
 
7.6.1 When infection, cardiopulmonary dysfunction, hypotension and other toxic side effects occur after lymphocyte depleting chemotherapy pretreatment, the cell reinfusion treatment should be appropriately delayed, and the reinfusion treatment time should be determined or abandon the treatment as appropriate.

7.6.2 Preparation of first aid equipment and medicines: necessary rescue equipment such as oxygen inhalation, sputum suction, ECG monitoring, ventilator, etc. must be prepared at the bedside, as well as anti-allergic drugs, cortisol drugs (such as methylprednisolone, dexamethasone) and other first aid Drugs; when gene-edited T cell preparations are reinfused, the institution should have IL-6 receptor inhibitors (such as tocilizumab) and TNF-α receptor inhibitors (such as etanercept) as a backup; unless IL-6 receptors are present If the inhibitor is ineffective or life-threatening, cortisol drugs are generally not used.
7.6.3 Diphenhydramine and antipyretic analgesics should be given 30min to 60min before reinfusion treatment to prevent acute allergic reactions.
7.6.4 Preparation of cell preparations: After fresh cell preparations are taken out from 4°C, the room temperature should not exceed 30 minutes; frozen cells should be placed in a 37°C water bath immediately after being taken out of the liquid nitrogen tank. Shake repeatedly and quickly re-thaw. After being completely melted, the room temperature should be kept for no more than 30 minutes. If any abnormalities such as breakage, liquid leakage, air leakage, etc. occur during the re-melting process, discard this batch of cells and notify the preparation institution for medical waste disposal.
7.7 Cell reinfusion therapy
 
7.7.1 The route of reinfusion treatment is generally intravenous or intraperitoneal injection. Intravenous reinfusion therapy should use a blood transfusion set without a filter to prevent the loss of effective cell components and reduce the efficacy.
7.7.2 The procedure for reinfusion treatment is generally: flushing the tube with 100ml of normal saline → reinfusion of cells → flushing the tube with 100ml of normal saline; the speed of the reinfusion of cells is generally controlled at 2ml/min～3ml/min in the first 15 minutes, and it is changed when there is no adverse reaction. 5ml/min～10ml/min.
7.7.3 After the treatment is completed, it is necessary to closely observe for at least 1 to 2 days and actively deal with any toxic and side effects that occur. Some special treatments (such as CAR-T) may have serious side effects (such as CRS, off-target effects, or neurotoxicity) about a week after treatment. Therefore, the patient's hospitalization observation time after treatment is completed depends on different treatment plans.

8 Treatment of side effects
8.1 Off-target effect
 Specific cellular immunotherapy mostly has clear targets, but in addition to tumor cells expressing these targets, normal tissues may also express them, so toxic and side effects caused by off-target effects may occur. Such side effects are more common in lung function damage, and patients may have clinical manifestations such as dyspnea and decreased blood oxygen. Treatment is based on symptomatic treatment measures such as oxygen inhalation, sputum suction, ventilator support, and anti-infection.
8.2 Cytokine release syndrome (cytokinerelease syndrome, CRS)
 The occurrence of CRS is due to the large number of proliferation and targeted killing of effector cells after returning to the body to release a large number of cytokines, forming a cytokine "storm" and causing multiple organ damage and failure. The main clinical manifestations are high fever and chills, decreased cardiopulmonary function, hypotension, hypoxia and pulmonary edema, acute kidney injury and other symptoms of multiple organ dysfunction. Because the clinical symptoms are more dangerous, treatment needs to be timely and active. Generally, reasonable use of cytokine antagonists such as tocilizumab and other drugs can be relieved on the basis of active symptomatic treatment. Cortisol hormone therapy can also be considered when cytokine antagonists are ineffective. .
8.3 Neurotoxicity
 Severe neurotoxicity may occur after CAR-T or TCR-T cell immunotherapy. The mechanism may be the neurological damage caused by the large amount of cytokines entering the blood-brain barrier. The clinical manifestations are headache, mental changes, disturbance of consciousness, delirium, Hallucinations, etc. In severe cases, epilepsy may occur. Because tocilizumab has a large molecular weight and cannot easily pass through the blood-brain barrier, severe neurotoxicity needs to be treated with hormone drugs such as dexamethasone.
8.4 Allergic reactions
 The reinfusion treatment of various cell preparations may cause fever, rash and other symptoms similar to allergic reactions. Generally, the incidence of high fever (>38.5℃) is 30%-40%, and the incidence of low fever (<38.5℃) is 40%-50%. %, the incidence of skin rash (5%-10%). Clinical treatment is mainly based on symptomatic treatment, and treatment measures such as anti-allergic drugs, antipyretic drugs, and physical cooling can be used. Generally, it can return to normal within a few hours or 1 to 2 days.
8.5 Tumor lysis syndrome
 Because a large number of tumor cells are killed by effector cells within a short period of time, a large amount of cytokines and inflammatory factors are released, which damages the organ function. It is more common in hematological tumors and less common in solid tumors. Clinical treatment can generally adopt symptomatic treatment measures such as allopurinol, alkalization of urine, and diuresis, and pay attention to actively correcting hyperphosphate, hyperkalemia, and hypocalcemia to maintain renal function.
8.6 Cross-reactive antigen toxicity
 The effector cells represented by CAR-T or TCR-T may recognize the cross antigens expressed by organ tissues while recognizing and killing target cells, thus causing damage to organ tissues other than tumors. As some of the newly discovered target antigen-related cross-reactive antigens are still unclear, it is difficult to predict the toxicity of cross-reactive antigens. It is necessary to closely observe and detect changes in the function of various organs during treatment and give corresponding symptomatic treatment in time to ensure cellular immunity The safety of treatment.
8.7 Graft versus host disease (graft-versus-hostdisease, GVHD)
 Acute GVHD mostly occurs within 3 months after treatment, mainly manifested as skin, gastrointestinal mucosa and liver function damage and infection; chronic GVHD can occur 3 to 6 months after treatment, mainly manifested as systemic organ damage Such as xeroderma, scleroderma, chronic liver disease and infections. The clinical need to attach great importance to the treatment of acute GVHD. At this time, the patient should be treated with the maintenance of cell therapy as a supplement. Sequential treatment of hormones and immunosuppressants and anti-infective treatment can be considered.

 9 Patient follow-up The content and time limit of follow-up and follow-up are also different due to different technologies and products used in cellular immunotherapy. In the later follow-up, attention should be paid to follow-up of virus replication, bacterial replication, and genetic variant diseases or secondary tumors that may be caused by random gene insertion. Follow-up is also required to observe chronic GVHD and delayed allergic reactions. Therefore, patients receiving treatment should complete a one-year follow-up and record as much as possible, and if any related damage is found, necessary treatment measures should be taken in time and reported to the ethics committee for the record.

 References [1] "Immune Cell Preparation Quality Management Self-Regulations" issued by China Medical Biotechnology Association in October 2016 [2] "Technical Guidelines for Research and Evaluation of Cell Therapy Products" State Food and Drug Administration (2017 No. 216) No.) In December 2017 [3] "Chimeric Antigen Receptor Modified T Cell (CAR-T Cell) Preparation Quality Management Regulations" promulgated by China Medical Biotechnology Association in May 2018 [4] "CAR-T Cell Therapy" Product quality control testing research and non-clinical research considerations" China Food and Drug Control Research Institute promulgated in June 2018[5] "Somatic Cell Therapy Clinical Research and Transformation Application Management Measures (Trial)" promulgated by the National Health Commission in March 2019