Abstract 4453: Molecular hallmarks of drug resistance to DNA methylation inhibitors and alternative therapeutic regimen for overcoming resistance

Abstract

Abstract Aberrant DNA methylation perturbing the epigenetic machinery has been continuously underlined as the major hallmark involved in all types of cancer. The archetypal DNA methylation inhibitor, 2’-deoxy-5-azacytidine (5-AZA-CdR) have been approved by FDA as the effective epigenetic drug for the treatment of blood malignancies, over a decade. However, the successful epigenetic therapy remains impeded owing to its chemotherapeutic resistance. To investigate the molecular mechanism of resistance to 5-AZA-CdR, we developed several HCT116 p53 wild-type cell clones, resistant towards 5-AZA-CdR, and principally studied the molecular alterations during the development of resistance, using high throughput RNA sequencing based transcriptomics and mass spectrometry based proteomics. The molecular profiling of HCT116 cells sensitive to 5-AZA-CdR treatment versus 5-AZA-CdR resistant cell clones distinguished the differential expression of various genes and the proteins coded by the genes, compared with untreated HCT116 wild-type cells (ANOVA p<0.05). The major affected cellular pathways were (i) Cell cycle: role of 14-3-3 proteins in cell cycle regulation, G1/S transition and initiation of mitosis (ii) Apoptosis and survival: granzyme A signaling, BAD phosphorylation, p53 dependent apoptosis (iii) Transcription: role of heterochromatin protein I family in transcriptional silencing, role of AP1 in regulation of cellular metabolism (iv) DNA damage: role of SUMO in p53 regulation. Further, we used MTT cytotoxicity assay to determine the cross-resistance or sensitivity of the 5-AZA-CdR resistant cell clones towards the inhibitors of epigenetic “Readers-Writers-Erasers”. The study revealed that 5-AZA-CdR resistant cell clones exhibited cross-resistance towards all the tested epigenetic inhibitors, however, significant sensitivity was exceptionally observed for bromodomain inhibitors. Additionally, the well-known bromodomain inhibitor, (+)-JQ1, showed significant anti-tumor activity sensitizing 5-AZA-CdR resistant HCT116 xenografts, suggesting bromodomain inhibition as the alternative therapeutic regimen for overcoming resistance to DNA methylation inhibitors. Validation of relevant genes and/or proteins as biomarkers for predicting clinical response to 5-AZA-CdR, including mutation analysis and methylome sequencing, and studies validating bromodomains as alternative therapeutic target for re-sensitizing the cancer patients, resistant to DNA methylation inhibitors is currently ongoing. Citation Format: Khushboo Agrawal, Petr Vojta, Dusan Holub, Rastislav Slavkovsky, Ivo Frydrych, Petr Dzubak, Marcela Krecmerova, Marian Hajduch. Molecular hallmarks of drug resistance to DNA methylation inhibitors and alternative therapeutic regimen for overcoming resistance. [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 4453.