嵌合抗原受体（CAR）-T细胞在急性髓系白血病（AML）的早期临床研究中并未取得重大进展。这种进展不足可能部分归因于AML的免疫抑制微环境，如类单核细胞样髓源性抑制细胞（M-MDSCs）和替代激活巨噬细胞（M2细胞），它们可以抑制CAR-T细胞的抗肿瘤活性。此外，AML细胞通常是异质的，单靶点CAR-T细胞可能无法消除所有AML细胞，导致疾病复发。CD123和NKG2D配体（NKG2DLs）是AML的常用CAR-T治疗靶点，M-MDSCs和M2细胞表达这两种抗原。我们通过各种结构优化筛选开发了双重靶向的CAR-T（123NL CAR-T）细胞，靶向CD123和NKG2DL。我们的研究揭示了123NL CAR-T细胞根除AML细胞并选择性靶向免疫抑制细胞。一种高度紧凑的标记/残杀基因RQR8，可以结合CD34和CD20抗原的靶向表位，也被纳入到CAR结构之前。利妥昔单抗与RQR8的结合导致123NL CAR-T细胞的消除及其细胞毒性的停止。总之，我们成功开发了123NL CAR-T细胞对肿瘤细胞和免疫抑制细胞的双重作用，可以避免靶点逃逸并抵抗免疫抑制微环境的影响。 参考文献：© 2023 The Author(s). Published with license by Taylor & Francis Group, LLC.

Chimeric antigen receptor (CAR)-T cells have not made significant progress in the treatment of acute myeloid leukemia (AML) in earlyclinical studies. This lack of progress could be attributed in part to the immunosuppressive microenvironment of AML, such as monocyte-like myeloid-derived suppressor cells (M-MDSCs) and alternatively activated macrophages (M2 cells), which can inhibit the antitumor activity of CAR-T cells. Furthermore, AML cells are usually heterogeneous, and single-target CAR-T cells may not be able to eliminate all AML cells, leading to disease relapse. CD123 and NKG2D ligands (NKG2DLs) are commonly used targets for CAR-T therapy of AML, and M-MDSCs and M2 cells express both antigens. We developed dual-targeted CAR-T (123NL CAR-T) cells targeting CD123 and NKG2DL by various structural optimization screens. Our study reveals that 123NL CAR-T cells eradicate AML cells and selectively target immunosuppressive cells. A highly compact marker/suicide gene, RQR8, which binds targeting epitopes of CD34 and CD20 antigens, was also incorporated in front of the CAR structure. The binding of Rituximab to RQR8 leads to the elimination of 123NL CAR-T cells and cessation of their cytotoxicity. In conclusion, we successfully developed dual effects of 123NL CAR-T cells against tumor cells and immunosuppressive cells, which can avoid target escape and resist the effects of immunosuppressive microenvironment.© 2023 The Author(s). Published with license by Taylor & Francis Group, LLC.