Myeloid Cell–associated Resistance to PD-1/PD-L1 Blockade in Urothelial Cancer Revealed Through Bulk and Single-cell RNA Sequencing

Abstract

Purpose: To define dominant molecular and cellular features associated with PD-1/PD-L1 blockade resistance in metastatic urothelial cancer. Experimental Design: We pursued an unbiased approach using bulk RNA sequencing data from two clinical trials to discover (IMvigor 210) and validate (CheckMate 275) pretreatment molecular features associated with resistance to PD-1/PD-L1 blockade in metastatic urothelial cancer. We then generated single-cell RNA sequencing (scRNA-seq) data from muscle-invasive bladder cancer specimens to dissect the cellular composition underlying the identified gene signatures. Results: We identified an adaptive immune response gene signature associated with response and a protumorigenic inflammation gene signature associated with resistance to PD-1/PD-L1 blockade. The adaptive immune response:protumorigenic inflammation signature expression ratio, coined the 2IR score, best correlated with clinical outcomes, and was externally validated. Mapping these bulk gene signatures onto scRNA-seq data uncovered their underlying cellular diversity, with prominent expression of the protumorigenic inflammation signature by myeloid phagocytic cells. However, heterogeneity in expression of adaptive immune and protumorigenic inflammation genes was observed among single myeloid phagocytic cells, quantified as the myeloid single cell immune:protumorigenic inflammation ratio (Msc2IR) score. Single myeloid phagocytic cells with low Msc2IR scores demonstrated upregulation of proinflammatory cytokines/chemokines and downregulation of antigen presentation genes, were unrelated to M1 versus M2 polarization, and were enriched in pretreatment blood samples from patients with PD-L1 blockade–resistant metastatic urothelial cancer. Conclusions: The balance of adaptive immunity and protumorigenic inflammation in individual tumor microenvironments is associated with PD-1/PD-L1 resistance in urothelial cancer with the latter linked to a proinflammatory cellular state of myeloid phagocytic cells detectable in tumor and blood. See related commentary by Drake, p. 4139

Tumor-Associated Macrophages in Bladder Cancer: Biological Role, Impact on Therapeutic Response and Perspectives for Immunotherapy

“…Conversely, CD169 + M1-like macrophages in the tumor-draining lymph nodes [48], but not in the tumor [25], are positively correlated with a favorable prognosis in MIBC patients. However, single-cell RNA-seq from immune-infiltrating cells revealed the wide diversity of TAMs in MIBC patients [49]. In this study, six different subsets of monocytes/macrophages were found in MIBC tumor, and their gene signatures did not correlate with the classical M1-like versus M2-like signatures.…”

“…In this study, six different subsets of monocytes/macrophages were found in MIBC tumor, and their gene signatures did not correlate with the classical M1-like versus M2-like signatures. Moreover, the authors also identified another subset of TAM that shares both TAM-like and MDSC-like gene signatures [49], supporting the theory of the MDSC-to-macrophage differentiation. In the future, determining the origin of TAM (from tissue-resident macrophages, monocytes-derived macrophages or MDSC differentiation) could improve knowledge on TAM diversity and complexity but this is still limited by the lack of specific markers for each subtype.…”

“…Zeng et al have analyzed the gene expression in the IMvigor210 cohort (Atezolizumab in BCa patients) and they declared that the M1-like TAM signature was a robust biomarker for predicting the prognosis and response to anti-PD-L1 [92]. Wang and collaborators demonstrated that a pro-tumorigenic inflammation signature was correlated with poor survival in the IMvigor210 and CheckMate275 (Nivolumab in BCa patients) cohorts [49]. They determine a score where TAMs and monocytes were not defined by the classical M1/M2 signature.…”

“…They determine a score where TAMs and monocytes were not defined by the classical M1/M2 signature. With this scoring, they demonstrated that monocytes with a low score were enriched in the peripheral blood of metastatic BCa patients with resistance to anti-PD-L1 [49], suggesting that pro-tumorigenic myeloid cells could be a pertinent biomarker for ICI efficiency.…”

Single-Cell RNA-Seq Analysis Reveals the Acquisition of Cancer Stem Cell Traits and Increase of Cell–Cell Signaling during EMT Progression

“…Solid cancers are rarely composed of similar cells with identical epigenetic traits. Rather, non-genetic determinants diversify cell populations into distinct phenotypes or cell states with various degrees of metastatic potential and resistance to various therapies [ 43 , 44 ]. Specifically, phenotypic heterogeneity along the EMT spectrum, or EM plasticity (EMP), is emerging as a distinctive feature of primary tumors [ 45 ], CTCs [ 46 , 47 ], and in the relation between tumors and CTCs [ 48 ].…”