Dual targeting of oncogenic microtubules and mitochondria in PDAC

ABSTRACT

Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest cancers. Microtubule inhibition is a promising therapeutic target as microtubule dynamics play a critical role in growth of metastases due to over expression of several β-tubulin subtypes compared to normal cells. Previous studies have shown that decreased expression of βIII- and βIVb-tubulin is associated with decreased PDAC cell growth. Bromodomain and Extra-Terminal domain (BET) proteins are transcription factors that regulate mitochondria proteins. In this study, we hypothesize that SB-216 and Veru-111 (related novel compounds) inhibit cell growth via suppression of oncogenic βIII- and βIVb-tubulin subtypes and mitochondria function via suppression of BRD4, the most active BET protein. PDAC cell growth was analyzed with the IncuCyte Live-Cell Analysis system. mRNA expression of βIII- and βIVb-tubulin was evaluated with quantitative real time PCR. Western blot analysis was performed for βIII, βIVb-tubulin, and BRD4 protein expression and expression of autophagy and mitophagy markers LC3B and p62/SQSTM1. Mitochondrial function/respiration was measured using a Seahorse XF-24 Flux Analyzer. Cell growth was greatly inhibited across all doses in multiple PDAC cell lines (p < .0001). mRNA expression of TUBB3 (βIII subtype) and TUBB4 (βIVb subtype) was significantly decreased (p < .05). BRD4 protein expression was reduced in with compensatory increase in mRNA expression. Treated PDCL had reduced mitochondrial respiration. Autophagy markers were increased in treated PDAC cells. Our data demonstrates that SB-216 effectively inhibits PDAC cell growth through inhibiting oncogenic microtubules and mitochondrial function. This novel approach simultaneously targets two hallmarks of cancer and patient demise.