Hematopoietic Tet2 inactivation enhances the response to checkpoint blockade immunotherapy

Somatic mutations inactivating TET2 are among the most common drivers of clonal hematopoiesis (CH). While TET2 inactivation is associated with monocyte-derived inflammation and improved chimeric antigen-receptor-T cell function, its impact on immunotherapy response is unknown. In murine models, hematopoietic Tet2 mutation enhanced the immune checkpoint blockade (ICB) response via the combined presence of phagocytes, CD4 and CD8 T cells. Mechanistically, in Tet2-mutant tumor-infiltrating leukocytes (TILs), ICB preferentially restricted cell states linked to tumor progression while inducing anti-tumor states. Tet2-mutant monocytes activated costimulatory programs, while Tet2-mutant T cells showed enhanced T cell memory signatures, alongside decreased exhaustion and regulatory phenotypes. This murine data was clinically relevant, since tumors from colorectal cancer and melanoma patients with TET2 driver mutation-CH (TET2-CH) showed enhanced immune infiltration, inflammation, and T cell activation. In melanoma patients treated with ICB, TET2-CH was associated with 6-fold greater odds of clinical benefit. Collectively, our data establishes that hematopoietic Tet2 inactivation primes leukocytes for anti-tumor states associated with immunotherapy response and provides a potential biomarker for personalized therapy. 9 weeks after rescue with Tet2wt or Tet2het bone marrow cells, C57Bl6/J mice were subcutaneously implanted with 250,000 MC-38 cells in the right flank. 17 days post-tumor implantation Tet2wt or Tet2het rescued mice were matched by tumor-size and assigned to receive 300 μg of anti-PD-1 blocking antibody (5 mice per genotype) or rat IgG2a isotype control (3 mice per genotype) every 2 days for 3 doses. Mice were euthanized 48 hours after the final dose and live, CD45+ tumor-infiltrating leukocytes were isolated using Fluorescence-Activated Cell Sorting. Single cells were processed using Chromium 10X 5’ Single Cell and Chromium Single Cell Mouse TCR Amplification kits.

功能失活型TET2体细胞突变是克隆性造血（clonal hematopoiesis, CH）最常见的驱动因素之一。尽管TET2功能失活与单核细胞介导的炎症反应及嵌合抗原受体T（chimeric antigen-receptor-T, CAR-T）细胞功能改善相关，但其对免疫治疗应答的影响仍未明确。在小鼠模型中，造血系统Tet2突变通过协同招募吞噬细胞、CD4及CD8 T细胞，增强了免疫检查点阻断（immune checkpoint blockade, ICB）的应答效果。从机制层面来看，在携带Tet2突变的肿瘤浸润白细胞（tumor-infiltrating leukocytes, TILs）中，ICB可优先抑制与肿瘤进展相关的细胞状态，同时诱导抗肿瘤细胞状态。Tet2突变的单核细胞可激活共刺激通路，而Tet2突变的T细胞则表现出增强的T细胞记忆特征，同时耗竭及调节性表型有所降低。该小鼠研究数据具有临床相关性：携带TET2驱动突变型克隆性造血（TET2-CH）的结直肠癌及黑色素瘤患者的肿瘤组织中，免疫浸润、炎症反应及T细胞活化水平均有所升高。在接受ICB治疗的黑色素瘤患者中，TET2-CH与临床获益几率升高6倍显著相关。综上，本研究数据证实，造血系统Tet2功能失活可将白细胞预编程为与免疫治疗应答相关的抗肿瘤状态，并为个性化治疗提供了潜在的生物标志物。在通过Tet2野生型（Tet2wt）或Tet2杂合型（Tet2het）骨髓细胞进行造血重建后的第9周，将25万个MC-38细胞皮下接种于C57Bl6/J小鼠右侧背部皮下。肿瘤接种后第17天，按肿瘤体积对Tet2wt及Tet2het重建小鼠进行匹配分组，分别给予300μg抗PD-1阻断抗体（每种基因型5只小鼠）或大鼠IgG2a同型对照（每种基因型3只小鼠），每2天给药1次，共给药3次。末次给药48小时后处死小鼠，通过荧光激活细胞分选（Fluorescence-Activated Cell Sorting, FACS）分离存活的CD45+肿瘤浸润白细胞。采用Chromium 10X 5’单细胞及Chromium单细胞小鼠TCR扩增试剂盒对单细胞进行测序前处理。