Spatial analysis of the tumor immune microenvironment in papillary renal cell carcinoma.

Abstract

471 Background: Spatial analysis of the tumor immune microenvironment (TIME) has yet to be explored in papillary renal cell carcinoma (pRCC). We utilized multiplex immunofluorescence (mIF) and spatial transcriptomics using spatial molecular imaging (SMI) to evaluate TIME properties in pRCC and contrasted these results with clear cell RCC (ccRCC). Methods: Tumor specimens were obtained from localized RCC tumors. mIF was performed on regions of interest (ROIs) selected from matched compartments from tumor, stroma, and tumor/stromal subsets of the interface. Two antibody panels were used for markers against T cells and B cells/macrophages. Marker abundance and clustering differences between pRCC and ccRCC were evaluated across ROIs. Select markers were also explored across pathologic tumor staging in pRCC. The SMI platform used for validation utilized probes against 959 transcripts. Cells were phenotyped using InSituType using the Kidney Cell Atlas as a reference. Cell clustering was quantified by univariate and bivariate Ripley’s K using the spatialTIME package in R. Results: mIF was performed on 1178 ROIs from 16 pRCC tumors and 70 ccRCC tumors. Compared to ccRCC, pRCC immune cell abundance was statistically lower amongst many T cell types and M2-like macrophages (Figure). M1-like macrophages were the only cell line seen at higher levels in interface compartments only. Increased macrophage clustering was observed in pRCC, including doubly positive M2-like macrophages in interface compartments (p=0.001 and 0.007). Higher abundance of CD8+ and FOXP3+ T cells in pRCC was associated with worse clinical stage, but no trend was seen with marker clustering. Four ROIs from 2 pRCC patients underwent SMI validation. On SMI of the tumor compartment, T cells were clustered with other T cells, B cells, and M1 macrophages. Conclusions: Compared to ccRCC, pRCC has fewer T cells and macrophages but more macrophage clustering. Using spatial transcriptomics, we found significant clustering between T cells, macrophages, and B cells in pRCC.