Abstract 2375: TFAM drives metabolic plasticity and cisplatin resistance in bladder cancer
Authors: Iqbal Maheen , Xu Tong , Lu Yi-Tsung , Goldkorn Amir 
Published: June 2022

Abstract
Background: First-line therapy for locally advanced and metastatic bladder cancer (BC) consists of cisplatin-based chemotherapy, but its efficacy is limited by the emergence of resistance. We previously showed that resistance can emerge rapidly, in a mutation-independent manner, through epigenetic and transcriptional reprogramming associated with a metabolic shift towards oxidative phosphorylation (OxPhos). Based on these findings, we hypothesized that genes associated with mitochondrial activation may drive the transition to chemoresistance.
Methods: Using our previously published (PMID: 21518726, 23784558, 32017074) model of bladder cancer plasticity, we FACS-sorted BC cell lines into highly aggressive, stem-like, drug-resistant side population (SP) and non-stem-like drug-sensitive non-side population (NSP). SP and NSP cells were analyzed by RNA-seq, and differentially expressed genes were entered into the Gene Transcription Regulation Database (GTRD; http://gtrd.biouml.org/) to identify candidate upstream regulators associated with OxPhos. For functional analysis, T24 and J82 BC cells were treated with cisplatin, with or without siRNA knockdown of candidate driver genes, followed by qRT-PCR, Western blot, FACS, and cell counts.
Results: Differentially expressed genes between SP and NSP were analyzed by GTRD, which predicted Transcription Factor A, Mitochondrial (TFAM) as a key upstream driver of the phenotypic transition between the two cell states. Overexpression of TFAM in SP cells was confirmed by qRT-PCR (1.8-fold, p value &lt;0.001). Cisplatin treatment (10μM) induced higher expression of TFAM mRNA and protein (1.6-fold, p value &lt;0.001 and 1.7-fold, p value &lt;0.001, respectively, at 72hr). Conversely, siRNA-mediated TFAM knock-down at 48hr reduced the number of SP cells (0.5-fold, p value 0.03), depleted mitochondrial DNA (0.7-fold, p value &lt;0.001), and synergized with cisplatin to kill BC cells (coefficient of drug interaction: 0.57, p value &lt;0.001). TCGA analysis revealed significantly higher TFAM expression in primary BC tumors vs. normal (p value &lt;0.001) and in higher histological grade (p value 0.006).
Conclusion: We demonstrate that drug-resistant BC cells overexpress TFAM, a known mitochondrial activator, and that more aggressive BC in TCGA is also marked by TFAM elevation. Importantly, cisplatin exposure induces TFAM expression which in turn drives a rapid shift towards a drug-resistant phenotype. Hence, TFAM may play a key role in non-mutation-mediated metabolic plasticity, permitting a subset of BC cells to persist in the face of cisplatin exposure. Further analysis of TFAM and its partners may elucidate new therapeutic targets to delay or avert drug resistance in BC.
Citation Format: Maheen Iqbal, Tong Xu, Yi-Tsung Lu, Amir Goldkorn. TFAM drives metabolic plasticity and cisplatin resistance in bladder cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2375.