Tumor Associated Antigen Specific T Cells Clinical Research For The Treatment Of Acute Myeloid Leukemia

Background Acute myeloid leukemia is a kind of malignant tumor of hematopoietic system,for clinical treatment and its molecular biological characteristics,in addition to-acute promyelocytic leukemia have more in-.depth understanding;able to take target gene differentiation therapy,other myeloid leukemia is still dominated by combined chemotherapy,followed by autologous hematopoietic stem cell transplantation and allogeneic hematopoietic stem cell transplantation.Some patients received complete remission after chemotherapy,but the majority of patients with cancer in 5 years of continuous complete remission(CCR)probability is low,the main cause of treatment failure is residual leukemia relapse.After allogeneic hematopoietic stem cell transplantation tumor recurrence rate is low,but it is only suitable for patients under the age of 50,HLA matched donor,more complications,high cost,can receive allogeneic hematopoietic stem cell transplantation in the treatment of patients is less,and some patients still relapse.Therefore,the need of develop new therapies for the treatment of a variety of hematologic malignancies,including acute leukemia.Objective Tumor associated antigen specific cytotoxic T lymphocytes(TAA-CTL)are activated by tumor antigens and confered specificity to the tumor.A variety of methods can be used in vitro generation of tumor associated antigen specific immune activity of CTL at present,and in clinical trials to demonstrate its preliminary anti-tumor effect.However,because of diversity,different individuals and different tumor antigen presentation of tumor antigen is effective and specific tumor antigen identification and isolation technology and other conditions,the generation ofTAA-CTL still need to be further improved,but also limited its wide application.The purpose of this study was to establish a TAA-CTL therapeutic system,and to explore the safety and efficacy of autologous TAA-CTL therapy targeting a variety of AML associated antigens(NY-ESO-1,MAGEA3,PRAME,Survivin ? WT-1).Methods 9 cases with AML were retrospectively analyzed.100ml of peripheral blood was collected from patients,peripheral blood mononuclear cells(PBMCs)were separated,dendritic cells(DCs)were cultured and loaded with leukemia-related antigen mixed peptides(NY-ESO-1,MAGEA3,PRAME,Survivin and WT-1).DCs were co-cultured with autologous PBMCs to prepare leukemia antigen-specific T cells before re-infusing to these patients three weeks later.After reinfusion,they also received subcutaneous injection of IL-2 10-15×105 IU/day×14 days.All of these nine patients suffered from AML(non-M3).Before TAA-CTL infusion,there were two cases with relapsing,three cases with MRD+(flow cytometry)and four cases with MRD-(flow cytometry);the median patient age was 46 years old(30-68 years old).Detect the TAA-CTL phenotype by flow-cytometry.Collect the eripheral blood from the patients weekly after the infusion for the first month.using the mononuclear cells extracted for ELISpot assay,according to the level of interferon gamma secretion evaluate the TAA-CTL activity.Results Each patient received TAA-CTL cell re-infusion for one to three times,1.04-5.08×107 cells/m2 for each time.None of them had obvious adverse reactions during infusion and after infusion;2 patients had fever and swelling at the injection site after using IL-2,but the symptoms disappeared after stopping it.Among these 9 patients,4 MRD-patients(adjuvant therapy)were still MRD-(3 to 11 months,during follow-up,with no other treatments).For 2 patients who turned from MRD-to MRD+,their MRD was persistently negative(9 months and 11 months,during follow-up,with no other treatments)after one course of TAA-CTL cell treatment.For 1 patient whose MRD was still positive after receiving multiple courses of treatment,MRD turned negative after one course of treatment with decitabine(10mg/m2,d1-5)prior to re-infusing TAA-CTL cells,but the patient had a relapse 5 months later.Now TAA-CTL treatment is undergoing now.Among 2 patients with relapse,1 patient remained stable after one course of TAA-CTL treatment,then TAA-CTL was re-infused againtwo months later,and decitabine combined with CAG chemotherapy scheme was also given until complete remission(CR)was reached;the other patient reached CR after one course of treatment through TAA-CTL and decitabine + CAG chemotherapy scheme.Phenotyping of the ex vivo-expanded T-cell lines showed a mean CD3+content of 93.5%,(range,56.5%-99%)and varying distribution of CD4+(mean37.4%,range3.2%-57.1%,)and CD8+(mean,43.2%;range,5.5%-64%)T cells,few natural killer cells(CD3-CD56+:mean4.9%,range,0.3%-9.8%)and rare residual B cells(CD19+:mean,0.3%,range0-0.6%,).The majority of both CD4+(mean,46.7%;range,5.6%-63.3%)and CD8+(mean,50.2%;range,3.9%-78%)T cells were composed of CD45RO+CD62L-CCR7-T cells in accordance with an effector-memory phenotype.Very few naiveCD45RA+CD62L+CCR7+cells(CD4+:mean,3.9%,range,0-8.9%;CD8+:mean,2.1%,range,0.5%-3.9%)and central-memory CD45RO+CD62L+CCR7+cells(CD4+:mean,5.7%,range,0-12.3%;CD8+:mean,13.5%,range,0.8%-21%)were present after two restimulations.For elispot assay,the patients who receiving once TAA-CTL treatment,the number of SFC(spot forming cells),suggesting that over time,the SFC curve increased gradually then fall,the average peak occurred at 14-21 days after the infusion,for the patients,combined with the clinical manifestations,who receiving twice or thrice treatments,the persistent remission ones have a larger number of up to 253 SFC/2×105 cells,and the patiens who recurrence within months or sustained non-remission with low SFC number,flat curve.Conclusion Autologous TAA-CTL targeting multiple tumor-associated antigens(NY-ESO-1,MAGEA3,PRAME,Survivin and WT-1)has good safety and preliminarily demonstrates its clinical efficacy.Its long-term efficacy and whether its combination with decitabine and other chemotherapies can further enhance the efficacy of autologous TAA-CTL should be further studied.