Program: Oral and Poster Abstracts
Session: 602. Disordered Gene Expression in Hematologic Malignancy, including Disordered Epigenetic Regulation: Poster I
In this study we identified a key set of microRNA (miR147b, miR-210, miR425, miR1253, miR4267 and miR4667-3p) that targeted BTK. Ectopic expression of each individual miRNA reduced BTK expression by 30-50% whereas a pool of these miRNA reduced BTK by 80%. When the expression of the BTK targeting microRNAs was compared in the CLL cells from 80 patients with a pool of healthy B-cells (5 donors) we found that CLL cells expressed lower levels of the BTK targeting microRNA compared to healthy B cells. Chromatin immunoprecipitation followed by deep sequencing as well as real time-PCR experiments identified that a gene repressive complex, consisting of HDAC1, HDAC2 and KDM1, became recruited to the promoters of these microRNA to silence their expression. Consequently, targeting the HDACs with the novel 2nd generation HDAC inhibitor Abexinostat induced the expression of BTK targeting miRNAs by 5-200 fold with reciprocal decreases in the levels of BTK protein. Similarly RNAi against HDAC1 induced the BTK targeting miRNA as well as decreased BTK in primary CLL cells as well as in Mec2, a CLL cell line. We then demonstrated that HDAC inhibition with Abexinostat synergized with ibrutinib to dually target BTK (where abexinostat decreased total BTK protein and ibrutinib inhibits kinase activity of remaining BTK to achieve a more complete inhibition of BTK signaling and synergistic cytotoxicity. Importantly, we also demonstrated that HDAC inhibitors retain the ability to induce the BTK targeting microRNA and reduce BTK protein in ibrutinib-resistant CLL cells that developed the mutant p.C481S BTK.
Together, our results show that the over-expression of HDACs in CLL causes epigenetic silencing of BTK targeting miRNAs leading to a higher expression of BTK in these patients. HDAC inhibition restores the expression of these miRNAs which in turn target BTK effecting a quantitative reduction. Since Abex is well toleratd in phase 1 clinical trials for heme malignancies, it makes an attractive therapeutic strategy to target BTK as part of a combination therapy as well as in Ibrutinib resistant patients.
Disclosures: Byrd: Acerta Pharma BV: Research Funding .
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