Clinical Study On Donor-derived Targeting CD19 CAR-T Cells For The Treatment Of Relapsed CD19~+ B-cell Acute Lymphoblastic Leukemia After Hematopoietic Stem Cell Transplantation

Objectives:With the rapid development of biotechnology,biotherapy has made extraordinary achievements in the field of tumor diagnosis and treatment,and has become the main therapy option in cancer treatment.Cancer immunotherapy methods mainly include adoptive cell therapy,immunomodulator,tumor vaccine and immune binding point block therapy,with the certain scope of application.Chimeric antigen receptor-T(CAR-T)therapy has been applied in cancer therapy in recent years,overcoming the disadvantages such as poor specificity of traditional cell therapy.Through gene editing technology,CAR receptors that bind tumor epitopes will be introduced into normal T cells through transfection,etc.,after amplification in vitro,the certain number of CAR-T cells are infused back to identify and eliminate tumor cells through chimeric antigens.Immune cell-based precision targeted immunotherapy has made major breakthroughs.CAR-T-related clinical trials have been carried out on a large scale worldwide.Among them,the second-generation targeted CD19CAR-T cells represented the most widely used technology in hematological malignancies therapy,with the effective rate in treating R/R hematological B cell malignancies could reach60-90%.CAR-T cells are classified into autologous,allogeneic,and stem cell-inducible types according to the source of T cells.Autologous T cells are commonly used at present.For patients who relapse after transplantation,because of the use of strong immunosuppressive agents after transplantation,the activity of patient's own T cells collected for CAR-T cell preparation may be affected,which will reduce the killing leukemia efficacy;at the same time,the process of collecting mononuclear cells is easily to be mixed into leukemia cells,thus affecting the CAR-T preparation efficacy.There are some advantages in using HLA-identical or haplo-identical donor T cells.Donor-derived T cells are a kind of"healthy"T cells,the collection quality is well guaranteed.It is also an important source for the preparation of CAR-T cells.The advantages are as follows:extensive source,contamination of tumor cells can be avoided and strong tumor killing ability.What remains to be observed is its safety,such as graft versus host disease(GVHD)and cytokine releasing syndrome(CRS).Acute lymphoblastic leukemia(ALL)is a common hematologic malignancy,accounting for 15%of all leukemia,30-40%acute leukemia(AL)in adults,and most commonly in children,up to 80%.Complete remission(CR)was 70-90%,and 3-5-year disease free survival(DFS)was 30-60%.Currently,hematopoietic stem cell transplantation(HSCT)is the main approach to cure ALL.However,even if HSCT is carried out,30-60%of patients progress to refractory or relapsed ALL,thus becoming the main cause of death in ALL patients.At present,there is no effective therapy option for refractory/relapsed(R/R)ALL.The current methods include chemotherapy,donor lymphocyte infusion(DLI)and secondary transplantation.However,the effect of DLI on ALL is not significant,and it is likely to induce graft versus host disease(GVHD).The safety of secondary transplantation needs to be seriously considered,and the prognosis of salvage secondary transplantation is relatively poor,with high costs.The 1-year overall survival(OS)rate of DLI after recurrence and secondary transplantation is only 20.74%and 23%,respectively.Therefore,it is urgent to find new effective treatment methods.Therefore,donor-derived CD19 CAR-T cells are expected to become a new treatment option for relapsed ALL.Patients with relapsed acute B lymphoblastic leukemia are treated with donor-derived CD19 CAR-T cells,so as to improve the disease remission rate and long-term survival rate.This study was approved by the Ethics Committee of the Army Medical University,and clinical research numbers approved in Chinese Clinical Trial Register are as follows:Chi CTR-OOC-16008447 and Chi CTR-OIC-17012374.Approval numbers of the Ethics Committee:CHIECRCT-20160022 and XYFY2017-KL033-01.Methods:From July 2015 to March 2019,43 patients from 9 transplant centers in China(including 18 cases from our center)were collected with confirmed B-ALL and underwent hematopoietic stem cell transplantation(HSCT)but relapsed,accepting donor-derived targeted CD19 CAR-T cell therapy.The primary end point of is effectiveness,the secondary end point is the covariates related to safety and effectiveness,monitoring including cytokine releasing syndrome(CRS),immune effector cell-associated neurotoxicity syndrome(ICANS),GVHD,cumulative incidence of relapse(CIR),event-free survival(EFS)and overall survival.The median follow-up time was 18 months(range for 6-47 months).Results:43 patients with relapsed CD19+B-ALL after HSCT were enrolled in this study and treated with donor-derived targeting CD19 CAR-T cells,including 29 males with a median age of 24 years old(4-60 years old).According to different donor types,they were divided into 17 cases receiving human leukocyte antigen(HLA)full-matched donor-derived CAR-T cells and 26 cases receiving HLA half-matched donor-derived CAR-T cells.The recurrent minimal residual disease(MRD)was 0.01-5%in 13 patients,5-50%in 20 patients and>50%in 10 patients.The median time from recurrence to CD19 CAR-T infusion was 42days(35-59 days).The treatment of patients after recurrence included stopping immunosuppression(12 cases),DLI(7 cases),chemotherapy(19 cases)and DLI combined with chemotherapy(5 cases).There were three preinfusion included in immune suppressive regimens:regimen 1:fludarabine,30 mg/m~2/day for 2–4 days and cyclophosphamide,200mg/m~2/day for 2 days;regimen 2:fludarabine,30 mg/m~2/day for 3 days,cyclophosphamide,350 mg/m~2/day for 2 days and cytarabine,and 100 mg/m~2/day for 4 days,and regimen 3:cyclophosphamide,500 mg/m~2/day for 3 days.Overall,34 patients received regimen 1.Regimen 2 was given to 5 younger patients with high tumor burden.Regimen 3 was given to4 older patients with low tumor cells.Expansion and persistence of CAR-T cells were analyzed by flow cytometry or by CAR-T cell DNA copy number.Median dose of infused CAR-T cells was 1.76×10~6/kg.According to the different costimulatory molecules of CAR-T cells,18 cases received CD19-28z CAR-T cells and 25 cases received CD19-BBz CAR-T cells.The median infusion of CAR-T cells was 1.76×10~6/kg.34 patientsachieved complete remission(79.07%).The 1-year EFS and survival rate was 43.33%.Among patients who achieved complete histological remission and did not receive a second transplant,the 1-year CIR was 41.0%,and the 1-year EFS and survival rate was 59.01%.In terms of incidence of adverse reactions,38 patients suffered from CRS(88.37%),of which 7 patients suffered from CRS?grade III;ICANS occurred in 9 patients(20.93%),all of which were?grade II.2patients(4.65%)developed acute GVHD?grade II.2 patients developed severe CAR-T cell-related CRS and multiple organ failure,which eventually resulted in death.Other common adverse effects included fever,agranulocytosis,anemia,thrombocytopenia,neurotoxicity such as headache,photophobia and other symptoms,but no severe brain edema occurs.Other adverse reactions included gastrointestinal symptom(nausea and anorexia),elevated liver enzymes(elevated transaminase and lactate dehydrogenase),electrolyte disturbance,and abnormal coagulation routine(prolonged activated partial prothrombin time(APTT)).CAR-T cell numbers peaked on day 9 after CAR-T cell infusion.Median peak concentration was 4.85×10~5/L.Conclusions:Donor-derived targeting CD19 CAR-T cells is one of the most effective methods to treat relapsed CD19~+B-ALL after HSCT.