HLA-DR expression in melanoma microenvironment

The emergence of immune checkpoint inhibitors has dramatically changed the therapeutic landscape for patients with advanced melanoma. However, relatively low response rates and a high incidence of severe immune-related adverse events have prompted the search for predictive biomarkers. A positive predictive value has been attributed to the aberrant expression of Human Leukocyte Antigen-DR (HLA-DR) by melanoma cells, but it remains unknown why this is the case. In this study, we have examined the microenvironment of HLA-DR positive metastatic melanoma samples using a multi-omics approach. First, using spatial, single-cell mapping by multiplexed immunohistochemistry, we found that the microenvironment of HLA-DR positive melanoma regions was enriched by professional antigen presenting cells, including classical dendritic cells and macrophages, while a more general cytotoxic T cell exhaustion phenotype was present in these regions. In parallel, transcriptomic analysis on micro dissected tissue from HLA-DR positive and HLA-DR negative areas showed increased IFN? signalling, enhanced leukocyte adhesion and mononuclear cell proliferation in HLA-DR positive areas. Finally, multiplexed cytokine profiling identified an increased expression of germinal center cytokines CXCL12, CXCL13 and CCL19 in HLA-DR positive metastatic lesions, which, together with IFN? and IL4 could serve as biomarkers to discriminate tumour samples containing HLA-DR overexpressing tumour cells from HLA-DR negative samples. Overall, this suggests that HLA-DR positive areas in melanoma attract the anti-tumour immune cell infiltration by creating a dystrophic germinal center-like microenvironment where an enhanced antigen presentation leads to an exhausted microenvironment, nevertheless representing a fertile ground for a better efficacy of anti-PD-1 inhibitors due to high levels of PD-1 expression.免疫检查点抑制剂的出现，极大地重塑了晚期黑色素瘤患者的治疗格局。然而，相对较低的临床应答率与高发的严重免疫相关不良反应，推动了预测性生物标志物的筛选与研究。既往研究认为黑色素瘤细胞人类白细胞抗原-DR（Human Leukocyte Antigen-DR, HLA-DR）的异常表达具有阳性预测价值，但该现象的潜在机制仍未阐明。本研究采用多组学方法，对HLA-DR阳性转移性黑色素瘤样本的肿瘤微环境展开分析。首先，借助多重免疫组化技术开展空间单细胞图谱分析，我们发现HLA-DR阳性黑色素瘤区域的微环境富集有专职抗原呈递细胞，包括经典树突状细胞与巨噬细胞；该区域同时呈现更为广泛的细胞毒性T细胞耗竭表型。随后，对显微切割获取的组织样本开展转录组分析，结果显示HLA-DR阳性区域的干扰素γ（IFN-γ）信号通路活性上调、白细胞黏附功能增强，且单核细胞增殖活跃。最后，多重细胞因子谱分析显示，HLA-DR阳性转移性病灶中，生发中心细胞因子CXCL12、CXCL13与CCL19的表达水平显著升高；上述细胞因子与干扰素γ及白细胞介素4（IL4）联合，可作为区分过表达HLA-DR的肿瘤样本与HLA-DR阴性样本的生物标志物。综上，本研究结果表明，黑色素瘤中的HLA-DR阳性区域通过构建退行性生发中心样微环境，招募抗肿瘤免疫细胞浸润：该微环境中抗原呈递功能增强，但最终导致局部免疫耗竭；不过，由于PD-1表达水平较高，该区域反而成为抗PD-1抑制剂发挥更佳临床疗效的理想场景。