Abstract 5141: Macrophage transcriptomic signature validation in scRNA seq and overall survival differences in urothelial carcinoma
Authors: Bukavina Laura , Bell Spencer , Geynisman Daniel , Isali Ilaha , Ranti Daniel , Sfakianos John , Valentine Henkel , Calaway Adam , Kutikov Alexander , Correa Andres , Uzzo Robert , Ponsky Lee , Abbosh Philip 
Published: April 2023

Introduction: While the prognostic value of immune system biomarkers has been well explored, role of cancer associated macrophages in bladder cancer(BC) is unclear due to lack of consistent results, coupled with tissue nonspecific transcriptomic signature.

Methodology: Data acquisition from 3,936 patients of The Cancer Genome Atlas (TCGA) in addition to Gene Expression Omninus (GEO) data sets GSE13507, GSE16945, GSE48277, GSE32894, GSE149582, as well as European Genotype Phenotype EGAS000001004507 was obtained. Tumor associated macrophages M1 and M2 were defined utilizing 188 and 159 gene expression profiles via xCell computational algorithm. To obtain bladder tissue residence macrophage signatures, scRNA seq was performed on tissue collected from surgical resection of 5 HG bladder cancer patients and 2 healthy controls. CellRanger software package with default parameters was utilized giving a total of 8,068 cells. FindAllMarkers was utilized for integration of DEG genes (cancer vs control) among the cluster and identification of BC macrophage specific markers. Endpoint of OS were measured in months from the time of cystectomy to follow up.

Results: Surprisingly, presence of M1 infiltration was found to be associated with improved OS in two out of eight cohorts only (GSE32894, GSE70691, p&lt;0.001), while the remainder detected no significant difference. Similarly, no association with OS was detected among all 8 cohorts with M2 infiltration. To improve our understanding of tissue specific markers of macrophage population in bladder, we then analyzed macrophage clusters detected within scRNAseq of BC patients compared to healthy. Gene ontology enrichment analysis of functions within tumor specific macrophages demonstrated an exaggerated expression of mTORC1 signaling, PI3K/AKT/mTOR signaling, TGF beta and EGF receptor pathways compared to non-cancer controls. Reanalysis of 22 tissue specific markers for M1/M2 infiltration showed no difference in any of the 8 cohorts with M1 high vs low infiltration. Similarly, addition of M2 infiltration as predictive marker yielded no further associations with OS in all but one cohort (GSE32894, p=0.00041), where high M2 tumor presence was associated with improved OS.

Conclusion: Our study represents the largest TAMs evaluation of BC across 8 cohorts with additional scRNA seq exploration of tissue specific signatures, demonstrating no association with OS in bladder cancer.

Citation Format: Laura Bukavina, Spencer Bell, Daniel Geynisman, Ilaha Isali, Daniel Ranti, John Sfakianos, Henkel Valentine, Adam Calaway, Alexander Kutikov, Andres Correa, Robert Uzzo, Lee Ponsky, Philip Abbosh. Macrophage transcriptomic signature validation in scRNA seq and overall survival differences in urothelial carcinoma. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5141.