CD38 标记肝癌中耗尽的 CD8 组织驻留记忆 T 细胞。

INTRODUCTION: Despite recent advances in immunotherapy for hepatocellular carcinoma (HCC), the overall modest response rate underscores the need for a better understanding of the tumor microenvironment (TME) of HCC. We have previously shown that CD38 is widely expressed on tumor-infiltrating leukocytes (TILs), predominantly on CD3 T cells and monocytes. However, its specific role in the HCC TME remains unclear. METHODS: In this current study, we used cytometry time-of-flight (CyTOF), bulk RNA sequencing on sorted T cells, and single-cell RNA (scRNA) sequencing to interrogate expression of CD38 and its correlation with T cell exhaustion in HCC samples. We also employed multiplex immunohistochemistry (mIHC) for validating our findings. RESULTS: From CyTOF analysis, we compared the immune composition of CD38-expressing leukocytes in TILs, non-tumor tissue-infiltrating leukocytes (NIL), and peripheral blood mononuclear cells (PBMC). We identified CD8 T cells as the dominant CD38-expressing TILs and found that CD38 expression was significantly higher in CD8 T in TILs than in NILs. Furthermore, through transcriptomic analysis on sorted CD8 T from HCC tumors, we observed a higher expression of CD38 along with T cell exhaustion genes, including PDCD1 and CTLA4, compared to the circulating memory CD8 T cells from PBMC. This was validated by scRNA sequencing that revealed co-expression of CD38 with PDCD1, CTLA4, and ITGAE (CD103) in T cells from HCC tumors. The protein co-expression of CD38 and PD-1 on CD8 T cells was further demonstrated by mIHC on HCC FFPE tissues, marking CD38 as a T cell co-exhaustion marker in HCC. Lastly, the higher proportions of CD38PD-1 CD8 T cells and CD38PD-1 T were significantly associated with the higher histopathological grades of HCC, indicating its role in the aggressiveness of the disease. CONCLUSION: Taken together, the concurrent expression of CD38 with exhaustion markers on CD8 T underpins its role as a key marker of T cell exhaustion and a potential therapeutic target for restoring cytotoxic T cell function in HCC.

简介：尽管肝细胞癌（HCC）的免疫疗法最近取得了进展，但总体上反应率较低，这突显了人们需要更好地了解 HCC 的肿瘤微环境（TME）。我们之前已经表明，CD38 在肿瘤浸润性白细胞（TIL）上广泛表达，主要在 CD3 T 细胞和单核细胞上。然而，其在 HCC TME 中的具体作用尚不清楚。

方法：在本研究中，我们使用流式细胞术时间飞行（CyTOF）、分选 T 细胞的批量 RNA 测序和单细胞 RNA（scRNA）测序来研究 CD38 的表达及其与 HCC 样本中 T 细胞耗竭的相关性。我们还采用多重免疫组化（mIHC）验证了我们的发现。

结果：从 CyTOF 分析中，我们比较了 TILs、非肿瘤组织浸润性白细胞（NIL）和外周血单核细胞（PBMC）中表达 CD38 的白细胞的免疫组成。我们发现 CD8 T 细胞是 CD38 表达的主要 TILs，并且在 TILs 中 CD8 T 细胞中的 CD38 表达明显高于 NILs。此外，通过对 HCC 肿瘤中分选的 CD8 T 细胞进行转录组分析，我们观察到与循环记忆 CD8 T 细胞相比，CD38 的表达以及 T 细胞耗竭基因（包括 PDCD1 和 CTLA4）的表达在 HCC 肿瘤中的表达更高。这通过 scRNA 测序得到了验证，该测序揭示了 HCC 肿瘤 T 细胞中 CD38 与 PDCD1、CTLA4 和 ITGAE（CD103）的共表达。在 HCC FFPE 组织上的 mIHC 进一步证明了 CD8 T 细胞上 CD38 和 PD-1 的蛋白共表达，将 CD38 标记为 HCC 中的 T 细胞共耗竭标志物。最后，CD38PD-1 CD8 T 细胞和 CD38PD-1 T 的较高比例与 HCC 的较高组织病理学分级显著相关，表明其在疾病侵袭性中的作用。

结论：综上所述，CD8 T 细胞上 CD38 与耗竭标志物的同时表达支持其作为 T 细胞耗竭的关键标志物的作用，并为恢复 HCC 中细胞毒性 T 细胞功能提供了潜在的治疗靶点。