Abstract 111: Adaptation to histone deacetylase inhibitors induces switching of histone H3 lysine 27 modifications, reprograming of cancer cell gene expression, and attenuation of the malignant phenotype
Authors: Lin Hsin-Ching , Vengsarkar Diana , Markert Elke , Rabson Arnold B. 
Published: August 2015

Background: Vorinostat a histone deacetylase inhibitor (HDACi) approved for treatment of cutaneous T cell lymphomas, inhibits the growth of cancer cell lines and human tumor xenografts. HDACi's increase acetylation of many proteins including histones, and lead to changes in cellular gene expression, however their mechanisms and optimal therapeutic uses are not known. HDACi's have multiple effects on cancer cell lines, ranging from induction of differentiation to apoptosis, senescence, and autophagy. We hypothesized that chronic treatment of malignant cells with HDAC inhibitors would attenuate the malignant phenotype by inducing reprogramming of cancer cell gene expression.

Methods: We derived a subclone of SW480 colon cancer cells (S3 cells, with K-Ras, p53 and APC mutations), which were treated with step-wise increases of vorinostat (to 3μM) resulting in vorinostat-adapted cells (SH80). SH80 cells were studied for growth rate, colony formation, and tumorigenicity in nude mice. Gene expression arrays (Affymetrix U133A2.0) were performed and gene expression changes validated by qPCR. Histone modifications were analyzed by Western blot and chromatin immunoprecipitation assays.

Results: SH80 vorinostat-adapted cells exhibited a marked decrease in cell growth, colony formation and tumorigenicity in nude mice, as well as features of differentiation including gland formation, increased intestinal alkaline phosphatase production, and increased cell size. Decreased growth rate and colony formation were seen even following drug withdrawal (for &gt;12weeks), suggesting that drug-adaptation resulted in a relatively stable phenotype. Gene expression microarrays demonstrated &gt;2 fold changes in expression of more than 10% of transcripts. Increased expression of colonocyte differentiation markers, putative tumor antigens and tumor suppressor genes was observed, as was decreased expression of several oncogenes and colonocyte-associated transcription factors. SH80 cells exhibited activation of polycomb repressive complex 2 (PRC2) target genes, associated with reduction of histone H3 lysine 27 trimethylation and increased H3K27 acetylation at the enhancer/promoter regions of these genes. These changes were distinct from those observed in SAHA-resistant cells obtained as survivors of acute drug treatment, which maintained oncogenic characteristics despite exhibiting resistance to vorinostat. Similarly, long term low dose (1uM) vorinostat treatment of HCT116 colon cancer cells also resulted in decreased growth rate and reduced colony formation.

Conclusions: Chronic HDAC inhibitor treatment induces a reprogramming of cancer cell epigenetic modifications and gene expression, even in the presence of multiple oncogenic mutations, leading to attenuation of the malignant phenotype.

Citation Format: Hsin-Ching Lin, Diana Vengsarkar, Elke Markert, Arnold B. Rabson. Adaptation to histone deacetylase inhibitors induces switching of histone H3 lysine 27 modifications, reprograming of cancer cell gene expression, and attenuation of the malignant phenotype. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 111. doi:10.1158/1538-7445.AM2015-111