Abstract 4729: Class-selective histone deacetylase inhibitors differentially promote translation-dependent HER2 transcript decay in HER2-positive breast cancer cells
Authors: Scott Gary K. , Frazier Katya , Malik Sadaf , Alejo Mariah , Schilling Birgit , Benz Christopher 
Published: July 2016

Abstract
Dysregulation of translational control is emerging as an essential aspect of tumorigenesis ripe for targeted therapeutic intervention. We have recently shown that polysome assembly can be differentially targeted by class IIB/HDAC6 selective histone deacetylase (HDAC) inhibitors (Wilson-Edell et al., Oncotarget, 2014), while translation-dependent and 3’UTR-targeted decay of oncogenic transcripts like HER2 is induced by pan-HDAC inhibition (Scott et al., Mol Cancer Res, 2008). To learn more about the HDAC dependency of oncogenic transcript stability, HER2-positive SKBr3 breast cancer cells were treated (2-24 h) with a pan-inhibitor of both class I and II HDACs (trichostatin-A/TSA), a class IIB &gt; class I selective inhibitor (ACY-1215/Ricolinostat), class I/HDAC1-3 selective inhibitors (Entinostat, ACY-1035), and class IIB/HDAC6 selective inhibitors (ACY-775, Tubacin) for their respective abilities to induce HER2 mRNA decay (Northern blotting), reduce HER2 protein levels, induce acetylation of histone 2B and/or tubulin (Western blotting), and to reduce cell culture growth (MTT cell viability). In parallel studies, protein candidates potentially mediating HER2 transcript decay were identified by mass spectrometry of complexes specifically binding to the 3’ UTR sense strand of HER2 mRNA. Polysome profiling demonstrated that pan-HDAC inhibition promoted protein candidates hnRNP-A2/B1 and the DEAD Box Helicase 5 (DDX5) to co-migrate with the 40S ribosome, and candidate hnRNP-K to co-migrate with higher (&gt;60S) polysome fractions. Additionally, immunoprecipitates of these candidates were found to be acetylated following pan-HDAC inhibitor treatment. With regard to reducing cell viability (at 72 h) and HER2 protein levels (at 24 h), TSA and ACY-1215 proved to be the most potent inhibitors, causing &gt;50% reductions at &gt;0.5 uM doses, while equimolar doses of the HDAC6-selective inhibitors showed minimal effects and the class I inhibitors (Entinostat, ACY-1035) showed only intermediate effects on both cell viability and HER2 protein reduction. Interestingly, only the pan-HDAC inhibitor TSA and the class I/IIB selective inhibitor ACY-1215 appeared capable of activating HER2 mRNA decay within 6 h of treatment; and both of these HDAC inhibitors produced similar enhancement of hnRNP-A2/B1 and DDX5 co-association with the 40S ribosome fraction. In summary, while the translational machinery including ribosome (40S, 60S) and polysome protein complexes can be rapidly dysregulated by either class IIB/HDAC6 selective or pan-HDAC inhibitors, it appears that the rapid induction of HER2 mRNA decay requires inhibition of both class I and class IIB/HDAC6 HDACs. These findings suggest that the investigational HDAC inhibitor ACY-1215/Ricolinostat possesses optimal class-selective properties for use as an adjunct to treat HER2-positive breast cancers.
Citation Format: Gary K. Scott, Katya Frazier, Sadaf Malik, Mariah Alejo, Birgit Schilling, Christopher Benz. Class-selective histone deacetylase inhibitors differentially promote translation-dependent HER2 transcript decay in HER2-positive breast cancer cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4729.