Extramedullary disease (EMD) is a high-risk feature of multiple myeloma (MM) and remains a poor prognostic factor, even in the era of novel immunotherapies. Here, we applied spatial transcriptomics (RNA tomography for spatially resolved transcriptomics [tomo-seq] [n = 2] and 10x Visium [n = 12]) and single-cell RNA sequencing (n = 3) to a set of 14 EMD biopsies to dissect the 3-dimensional architecture of tumor cells and their microenvironment. Overall, infiltrating immune and stromal cells showed both intrapatient and interpatient variations, with no uniform distribution over the lesion. We observed substantial heterogeneity at the copy number level within plasma cells, including the emergence of new subclones in circumscribed areas of the tumor, which is consistent with genomic instability. We further identified the spatial expression differences between GPRC5D and TNFRSF17, 2 important antigens for bispecific antibody therapy. EMD masses were infiltrated by various immune cells, including T cells. Notably, exhausted TIM3+/PD-1+ T cells diffusely colocalized with MM cells, whereas functional and activated CD8+ T cells showed a focal infiltration pattern along with M1 macrophages in tumor-free regions. This segregation of fit and exhausted T cells was resolved in the case of response to T-cell–engaging bispecific antibodies. MM and microenvironment cells were embedded in a complex network that influenced immune activation and angiogenesis, and oxidative phosphorylation represented the major metabolic program within EMD lesions. In summary, spatial transcriptomics has revealed a multicellular ecosystem in EMD with checkpoint inhibition and dual targeting as potential new therapeutic avenues.

中文翻译：

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空间转录组学揭示了髓外骨髓瘤的严重亚克隆异质性和 T 细胞功能障碍


髓外疾病 （EMD） 是多发性骨髓瘤 （MM） 的高危特征，即使在新型免疫疗法时代，它仍然是一个不良的预后因素。在这里，我们将空间转录组学（用于空间分辨转录组学的 RNA 断层扫描 [tomo-seq] [n = 2] 和 10x Visium [n = 12]）和单细胞 RNA 测序 （n = 3） 应用于一组 14 个 EMD 活检，以剖析肿瘤细胞的 3 维结构及其微环境。总体而言，浸润免疫细胞和基质细胞表现出患者内和患者间的变化，在病灶上没有均匀的分布。我们在浆细胞内拷贝数水平上观察到很大的异质性，包括在肿瘤的限定区域出现新的亚克隆，这与基因组不稳定性一致。我们进一步确定了 GPRC5D 和 TNFRSF17 之间的空间表达差异，这是双特异性抗体治疗的 2 种重要抗原。EMD 肿块被各种免疫细胞浸润，包括 T 细胞。值得注意的是，耗竭的 TIM3 + / PD-1 + T 细胞与 MM 细胞弥漫共定位，而功能性和活化的 CD8 + T 细胞在无肿瘤区域与 M1 巨噬细胞一起表现出局灶性浸润模式。在对 T 细胞结合双特异性抗体有反应的情况下，拟合 T 细胞和耗竭 T 细胞的这种分离得到了解决。MM 和微环境细胞嵌入到影响免疫激活和血管生成的复杂网络中，氧化磷酸化代表了 EMD 病变内的主要代谢程序。综上所述，空间转录组学揭示了 EMD 中的多细胞生态系统，其中检查点抑制和双重靶向是潜在的新治疗途径。