Multiplexed single-cell pathology reveals the association of CD8 T-cell heterogeneity with prognostic outcomes in renal cell carcinoma
Multiplexed single-cell pathology reveals the association of CD8 T-cell heterogeneity with prognostic outcomes in renal cell carcinoma
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DOI：
10.1007/s00262-021-03006-2
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发表时间：
2021-07-14
影响因子：
5.8
通讯作者：
Oya, Mototsugu
中科院分区：
文献类型：
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作者：
Murakami, Tetsushi;Tanaka, Nobuyuki;Oya, Mototsugu
尽管肾细胞癌(RCC)对免疫治疗高度敏感，但RCC已被认为是一种罕见的疾病，CD8(+)T细胞渗入肿瘤床与预后不良有关。为了深入了解肾细胞癌免疫生物学的内部格局，我们进行了多重七色免疫组织化学(CD8、CD39、PD-1、Foxp3、PD-L1和泛细胞角蛋白AE1/AE3与DAPI)，揭示了单个细胞的自动计数和单个细胞到细胞的距离的计算。总共包括186名受试者，其中CD39被用作区分肿瘤特异性(CD39(+))和旁观者(CD39(-))T细胞的标志。我们的透明细胞肾细胞癌队列也显示，如果肿瘤显示CD8(+)T细胞浸润增加，则预后不良。肿瘤内CD8(+)CD39(+)T细胞及其在肿瘤中心区域耗尽的CD8(+)CD39(+)PD-1(+)T细胞使患者能够根据恶性程度进行分组。对抗血管生成治疗后标本的分析显示，增殖的Treg分数Foxp3(+)PD-1(+)细胞显著增加，提示使用抗PD-1抗体后疾病过度进展的潜在机制。我们的逐细胞研究平台提供了肿瘤的空间信息，其中旁观者CD8(+)CD39(-)T细胞在侵袭性边缘区域占主导地位。我们发现CD8(+)CD39(+)PD-1(+)T细胞和Foxp3(+)PD-1(+)Treg细胞之间存在潜在的相互作用，这是由于细胞与细胞之间的接近，形成了一个在PD-1阻断下更专门用于免疫抑制的空间生态位。向免疫抑制环境的范式转变在转移性病变中更为明显；相反，Foxp3(+)和Foxp3(+)PD-1(+)Treg细胞的渗透更为明显。通过这项多元化的单细胞病理学技术，我们进一步揭示了肾细胞癌的免疫生物学地位。
Despite the high sensitivity of renal cell carcinoma (RCC) to immunotherapy, RCC has been recognized as an unusual disease in which CD8(+) T-cell infiltration into the tumor beds is related to a poor prognosis. To approach the inner landscape of immunobiology of RCC, we performed multiplexed seven-color immunohistochemistry (CD8, CD39, PD-1, Foxp3, PD-L1, and pan-cytokeratin AE1/AE3 with DAPI), which revealed the automated single-cell counts and calculations of individual cell-to-cell distances. In total, 186 subjects were included, in which CD39 was used as a marker for distinguishing tumor-specific (CD39(+)) and bystander (CD39(-)) T-cells. Our clear cell RCC cohort also revealed a poor prognosis if the tumor showed increased CD8(+) T-cell infiltration. Intratumoral CD8(+)CD39(+) T-cells as well as their exhausted CD8(+)CD39(+)PD-1(+) T-cells in the central tumor areas enabled the subgrouping of patients according to malignancy. Analysis using specimens post-antiangiogenic treatment revealed a dramatic increase in proliferative Treg fraction Foxp3(+)PD-1(+) cells, suggesting a potential mechanism of hyperprogressive disease after uses of anti-PD-1 antibody. Our cell-by-cell study platform provided spatial information on tumors, where bystander CD8(+)CD39(-) T-cells were dominant in the invasive margin areas. We uncovered a potential interaction between CD8(+)CD39(+)PD-1(+) T-cells and Foxp3(+)PD-1(+) Treg cells due to cell-to-cell proximity, forming a spatial niche more specialized in immunosuppression under PD-1 blockade. A paradigm shift to the immunosuppressive environment was more obvious in metastatic lesions; rather the infiltration of Foxp3(+) and Foxp3(+)PD-1(+) Treg cells was more pronounced. With this multiplexed single-cell pathology technique, we revealed further insight into the immunobiological standing of RCC.