Increased Expression of CD36 and CD163 in Clear Cell Renal Cell Carcinoma Suggests an Association Between Lipid Transport and an "M2-like" Macrophage Phenotype

Yosra Bouraoui¹Rahma SAID¹Christina Bruss^2,3Agnieszka Martowicz⁴Kilian Wagner²Florian Weber⁵Simon Engelmann⁶Sebastian Kaelble⁶Marcus Höring⁷Gerhard Liebisch⁷Roman Mayr^6,8Peter J. Siska^9*

• ¹Universite de Jendouba Institut Superieure de Biotechnologie de Beja, Beja, Tunisia
• ²Universitatsklinikum Regensburg Klinik und Poliklinik fur Innere Medizin III, Regensburg, Germany
• ³Institute of Functional Genomics, University of Regensburg, Regensburg, Germany, Regensburg, Germany
• ⁴Department of Internal Medicine V, Haematology & Oncology, Comprehensive Cancer Center Innsbruck (CCCI), Innsbruck, Austria
• ⁵Universitat Regensburg Institut fur Pathologie, Regensburg, Germany
• ⁶Caritas-Krankenhaus St Josef Regensburg, Regensburg, Germany
• ⁷Institute of Clinical Chemistry and Laboratory Medicine, University Hospital Regensburg, Regensburg, Germany, Regensburg, Germany
• ⁸Urology, Faculty of Medicine, University of Augsburg, Augsburg, Germany, Augsburg, Germany
• ⁹University Medical Center Regensburg, Regensburg, Germany

Background. Clear cell renal cell carcinoma (ccRCC) is marked by intracellular lipid accumulation and dysregulated lipid metabolism, features that contribute to tumor progression and possibly immune suppression. Tumor-associated macrophages (TAMs) in ccRCC are abundant and phenotypically diverse, with CD68 marking general macrophage presence and CD163 indicating alternatively activated, immunosuppressive (M2-like) subsets. The fatty acid transporter CD36 and the metabolic regulator CD147 can be found in tumors and may influence macrophage polarization, but their associations with TAM phenotypes and tumor lipid content in ccRCC remain unclear. Methods. Samples from 23 ccRCC patients were analyzed. Oil Red O staining quantified lipid accumulation. Immunohistochemistry for CD36, CD147, CD68, CD163, and CD3 was evaluated using both automated area-based quantification and semi-quantitative observer scoring. Flow cytometry of freshly resected tumors was used to assess the expression of CD36, CD147 and the infiltration with macrophages and CD8 T cells. Lipidomic analyses were performed from ccRCC tissues (n=5). RNA expression data from TCGA (n=533) were used for validation. Single-cell RNA sequencing data from two published datasets were analyzed to characterize cell-type-specific expression of lipid metabolism and macrophage markers. Results. ccRCC tumors showed an increased lipid accumulation and the expression of CD36 was detected on both CD45-negative cells and macrophages. Macrophages expressed CD163, suggesting a M2-like phenotype and CD163 expression was higher in larger ccRCC tumors. CD163 on macrophages positively correlated with CD36 on CD45-negative cells from ccRCC tumors, while CD8 T cells showed a trend to lower numbers in tumors with high CD36 expression on CD45-negative cells. CD147 was broadly expressed on CD45-negative and CD45-positive cells and positively correlated with CD36 on CD45-negative cells as well as with CD163 on macrophages. Tumors accumulated triacylglycerols, which correlated with CD163. Single-cell RNA-seq revealed CD163 expression across all TAM subsets and a compartmentalized distribution of lipid metabolism genes. Conclusions. Lipid metabolic markers CD36 and CD147 are expressed in ccRCC tumors and correlations with immune cell subsets suggest their role in suppressing anti-tumor immunity. These findings suggest the existence of a metabolic-immune axis in ccRCC and provide a rationale for targeting TAM metabolism to enhance immunotherapeutic efficacy.