白介素 1 受体辅助蛋白 (IL1RAP) 在急性髓系白血病 (AML) 原始细胞和白血病干细胞 (LSC) 上高表达，但在正常造血干细胞 (HSC) 上不表达，这为靶向和消除疾病提供了机会，同时保留正常造血功能。在此，我们报道了 BIF002（一种新型抗 IL1RAP/CD3 T 细胞接合剂 (TCE)）在 AML 中的活性。通过光电定位和单细胞测序，从免疫小鼠收集的 CD138 B 细胞中分离出 IL1RAP 抗体。生产并表征了个体小鼠单克隆抗体 (mAb)，我们利用 Fab 臂交换从中生成了 BIF002，这是一种抗人 IL1RAP/CD3 TCE。人 IgG1 Fc 中引入突变以减少 FcγR 结合。使用多种细胞系和患者来源的 AML 样本，在体外和体内表征了 BIF002 的抗白血病活性。发现 IL1RAP 在大多数人类 AML 细胞系和原代母细胞中高度表达，包括来自患有两种疾病的患者的富含 CD34 LSC 的亚群。新的和复发/难治性 (R/R) 白血病，但不适用于正常 HSC。在来自健康供体的 T 细胞与 IL1RAPhigh AML 细胞系和原代母细胞的共培养中，BIF002 在亚纳摩尔浓度下诱导剂量和效应器与靶标 (E:T) 比率依赖性 T 细胞激活和白血病细胞裂解。与对照相比，BIF002 与人类 T 细胞一起静脉注射可导致白血病细胞耗竭，并显着延长 IL1RAPhigh MOLM13 或 AML 患者来源的异种移植物的存活时间，且没有脱靶副作用。值得注意的是，BiF002 有效地重新定向 T 细胞以消除 LSC，证据是，与接受 BIF002 T 细胞治疗的供体的骨髓 (BM) 的二次接受者相比，未达到中位生存期；全部存活 > 200 天，二次接受者没有出现疾病。来自载体的 BM 接受者（中位生存期：26 天；p = 0.0004）或同种型对照抗体 T 细胞处理的供体（26 天；p = 0.0002）。新型抗 IL1RAP/CD3 TCE、BIF002 可根除 LSC，并显着延长 AML 异种移植物的存活时间，代表了一种有前景的新型 AML 治疗方法。© 2024。作者。

The interleukin-1 receptor accessory protein (IL1RAP) is highly expressed on acute myeloid leukemia (AML) bulk blasts and leukemic stem cells (LSCs), but not on normal hematopoietic stem cells (HSCs), providing an opportunity to target and eliminate the disease, while sparing normal hematopoiesis. Herein, we report the activity of BIF002, a novel anti-IL1RAP/CD3 T cell engager (TCE) in AML.Antibodies to IL1RAP were isolated from CD138+ B cells collected from the immunized mice by optoelectric positioning and single cell sequencing. Individual mouse monoclonal antibodies (mAbs) were produced and characterized, from which we generated BIF002, an anti-human IL1RAP/CD3 TCE using Fab arm exchange. Mutations in human IgG1 Fc were introduced to reduce FcγR binding. The antileukemic activity of BIF002 was characterized in vitro and in vivo using multiple cell lines and patient derived AML samples.IL1RAP was found to be highly expressed on most human AML cell lines and primary blasts, including CD34+ LSC-enriched subpopulation from patients with both de novo and relapsed/refractory (R/R) leukemia, but not on normal HSCs. In co-culture of T cells from healthy donors and IL1RAPhigh AML cell lines and primary blasts, BIF002 induced dose- and effector-to-target (E:T) ratio-dependent T cell activation and leukemic cell lysis at subnanomolar concentrations. BIF002 administered intravenously along with human T cells led to depletion of leukemic cells, and significantly prolonged survival of IL1RAPhigh MOLM13 or AML patient-derived xenografts with no off-target side effects, compared to controls. Of note, BiF002 effectively redirects T cells to eliminate LSCs, as evidenced by the absence of disease initiation in secondary recipients of bone marrow (BM) from BIF002+T cells-treated donors (median survival not reached; all survived > 200 days) compared with recipients of BM from vehicle- (median survival: 26 days; p = 0.0004) or isotype control antibody+T cells-treated donors (26 days; p = 0.0002).The novel anti-IL1RAP/CD3 TCE, BIF002, eradicates LSCs and significantly prolongs survival of AML xenografts, representing a promising, novel treatment for AML.© 2024. The Author(s).