肿瘤细胞治疗研究
英文原题：Overcoming target epitope masking resistance that can occur on low-antigen-expresser AML blasts after IL-1RAP chimeric antigen receptor T cell therapy using the inducible caspase 9 suicide gene safety switch.
尽管嵌合抗原受体（CAR）T细胞免疫疗法取得了不可否认且明确的成功，但从针对B细胞恶性肿瘤（无论是难治/复发性急性淋巴细胞白血病（ALL）还是淋巴瘤）中CD19抗原的首批临床试验监测中获得的知识，有助于识别出约30-50%的B-ALL中的肿瘤细胞逃逸。
尽管嵌合抗原受体（CAR）T细胞免疫疗法取得了毋庸置疑的成功，但对首批靶向B细胞恶性肿瘤CD19抗原的临床试验（对象包括难治性/复发性急性淋巴细胞白血病（ALL）或淋巴瘤）的监测结果表明，约30%–50%的B-ALL患者会发生肿瘤细胞逃逸。耐药原因包括表面抗原表达丢失（rCD19−），或CAR T细胞过早消失或失活（rCD19+）。近期一例临床病例报告显示，白血病细胞采集样本中的白血病细胞发生意外病毒转导，使CAR在B-ALL细胞表面遮蔽了抗原，导致rCD19−复发。本研究旨在急性髓系白血病（AML）背景下重现这一表位遮蔽耐药模型，采用靶向白血病干细胞所表达白细胞介素-1受体辅助蛋白（IL-1RAP）的免疫治疗CAR T细胞模型。由于AML原代母细胞的IL-1RAP表达水平不一，我们根据抗原位点密度筛选不同AML肿瘤细胞系，并用携带第三代IL-1RAP CAR、iCASP9自杀基因及截短CD19表面基因的慢病毒载体进行转导建模。结果显示，AML原代母细胞易于转导（74.55±21.29%，n=4）；针对IL-1RAP的CAR T细胞毒性与表位遮蔽程度呈反比，而表位遮蔽程度又与IL-1RAP阳性细胞系表面表达的抗原位点数量相关。值得注意的是，体外用Rimiducid处理IL-1RAP+/CAR+白血病细胞系24小时可清除超过85%的细胞。我们证实，IL-1RAP阳性白血病细胞表达IL-1RAP CAR会降低靶抗原的膜表面可及性，从而造成耐药；较高的表位密度则可维持其对CAR T细胞的敏感性。此外，iCASP9/Rimiducid自杀系统安全开关使这一免疫治疗方法有望安全用于未来的I期临床试验。
Although chimeric antigen receptor CAR) T cell immunotherapies are an undeniable and unequivocal success, knowledge obtained from the monitoring of the first clinical trials targeting the CD19 antigen in B malignancies, either refractory/relapsed acute lymphoid leukemia (ALL) or lymphomas, contributed to the identification of tumor cell escape in about 30-50% of B-ALL. Resistance occurred due to loss of surface expression of the antigen (rCD19-) or to the early disappearance or inactivation of CAR T cells (rCD19+). In a recently reported clinical case, rCD19- relapse resulted from masking of the antigen by the CAR at the surface of B-ALL leukemia cells following the unexpected viral transduction of a leukemic cell present in the cytapheresis sample. The objective of this work was to reproduce this epitope-masking resistance model, in the context of acute myeloid leukemia (AML), based on our immunotherapeutic CAR T cell model targeting the accessory protein of the interleukin-1 receptor (IL-1RAP) expressed by leukemic stem cells. As AML primary blasts express different levels of IL-1RAP, we modeled transduction of different AML tumor cell lines screened for density of antigenic sites with our lentiviral vectors carrying a third-generation IL-1RAP CAR, an iCASP9 suicide gene, and a truncated CD19 surface gene. We demonstrated that primary AML blasts can be easily transduced (74.55 21.29%, n = 4) and that CAR T cytotoxicity to IL-1RAP is inversely correlated with epitope masking in relation to the number of antigenic sites expressed on the surface of IL-1RAP+ lines. Importantly, we showed that, in vitro, a 24-h exposure of IL-1RAP+/CAR+ leukemia lines to Rimiducid eliminated >85% of the cells. We confirmed that the expression of IL-1RAP CAR by an IL-1RAP+ leukemic cell, by decreasing the membrane availability of the targeted antigen, can induce resistance while a high epitope density maintains sensitivity to CAR T cells. Moreover, the presence of the iCASP9/Rimiducid suicide system safety switch makes this immunotherapy approach safe for application in a future phase 1 clinical trial.
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