对免疫疗法的异质耐药性仍然是癌症治疗的主要挑战，常常导致疾病进展和死亡。使用 CITE-seq 和匹配的 40 重 PhenoCycler 组织成像，我们对具有先天性耐药、获得性耐药或对免疫治疗有反应的转移性黑色素瘤患者的肿瘤进行纵向多模式单细胞分析。我们建立了多模式整合工具包来对齐转录组特征、细胞表位和空间信息，以提供对肿瘤更深入的了解。通过纵向分析，我们确定了一种“免疫奋斗”的肿瘤微环境，其特征是肿瘤周围淋巴聚集、肿瘤中 T 细胞的低浸润以及治疗后 MITF SPARCL1 和 CENPF 黑色素瘤亚克隆的出现。淋巴聚集体分子组成中 B 细胞相关特征的富集与更好的生存相关。这些发现为建立微环境细胞相互作用和空间结构的分子组成提供了进一步的见解，可以为治疗干预提供信息。版权所有 © 2024 作者。由爱思唯尔公司出版。保留所有权利。

Heterogeneous resistance to immunotherapy remains a major challenge in cancer treatment, often leading to disease progression and death. Using CITE-seq and matched 40-plex PhenoCycler tissue imaging, we performed longitudinal multimodal single-cell analysis of tumors from metastatic melanoma patients with innate resistance, acquired resistance, or response to immunotherapy. We established the multimodal integration toolkit to align transcriptomic features, cellular epitopes, and spatial information to provide deeper insights into the tumors. With longitudinal analysis, we identified an "immune-striving" tumor microenvironment marked by peri-tumor lymphoid aggregates and low infiltration of T cells in the tumor and the emergence of MITF+SPARCL1+ and CENPF+ melanoma subclones after therapy. The enrichment of B cell-associated signatures in the molecular composition of lymphoid aggregates was associated with better survival. These findings provide further insights into the establishment of microenvironmental cell interactions and molecular composition of spatial structures that could inform therapeutic intervention.Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.