Abstract 5436: MED8-orchestrated C24 sphingolipid metabolism promotes tumor progression through enhanced tumor cell membrane fluidity and mitochondrial oxidation

Abstract Mediator complex is a large protein complex containing up to 30 proteins that coactivates the RNA polymerase II (Pol II) enzyme to transcribe all protein-coding and most non-coding RNA genes. Recently, it has been reported that various subunits of the complex may possess distinct functions in cancer progression. After comparing the expression of all the subunits between normal and tumor tissue and analyzing their prognostic potential, we found that mediator complex 8 (MED8) was the most oncogenic gene that consistently predicted poor survival in liver hepatocellular carcinoma (LIHC), kidney renal clear cell carcinoma (KIRC), sarcoma (SARC), and uterine corpus endometrial carcinoma (UCEC) according to TCGA database. Next, we aimed to explore the common functions and mechanisms of MED8 involved in the development of the aforementioned tumor types. Pathway analysis revealed a strong association between MED8 and sphingolipid metabolism. To further study the role of MED8 in LIHC, KIRC, SARC, and UCEC, we established shMED8 stable clones using cell lines of each tumor type followed by metabolomics profiling. Notably, a significant correlation was observed between MED8 levels and a cluster of C24 lipids. ELOVL1, which specifically produced C24 acyl-coAs and linked to C24 sphingolipid synthesis, was confirmed to be positively regulated by MED8. Also, as one of the main groups of C24 sphingolipid, C24 ceramide was found to be reduced by the knockdown of MED8 indicated by the mitigatory phosphorylation of LYN, further underscoring the regulatory role of MED8 in sphingolipid metabolism. Although ceramide is known to initiate apoptosis and suppress tumor development, the metabolism of ceramide into other forms of sphingolipid including sphingomyelin (SM), sphingosine (Sph), and sphingosine-1-phosphate (S1P) has been reported to promote tumor progression. Specifically, C24-SM has been found to increase cell membrane fluidity, contributing to the distant metastasis of tumor cells. Furthermore, S1P has been reported to promote cytochrome c oxidase (COX IV) and mitochondrial respiration. In our studies, MED8 was validated to enhance tumor cell membrane fluidity, COX IV expression, and migratory ability, likely mediated by altered C24-SM and C24-S1P metabolism. Besides being converted into other metabolites, ceramide is involved in small extracellular vesicles (sEV) biogenesis via the complex TSG101-CERT (ceramide transport). Decreased formation of such complex and sEV secretion were observed in shMED8 tumor cells. In addition, MED8-enriched sEV endowed recipient cells with enhanced invasive ability by increasing their membrane fluidity and respiration. Citation Format: Zhixian Chen, Xiaoxin Zhang, Judy Wai Ping Yam. MED8-orchestrated C24 sphingolipid metabolism promotes tumor progression through enhanced tumor cell membrane fluidity and mitochondrial oxidation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5436.