Abstract 3509: Novel dual inhibitors of LSD1-HDAC for treatment of cancer
Authors: Sivanandhan Dhanalakshmi , Rajagopalan Sridharan , Nair Sreekala , Dewang Purushottam , Kumar Durga Prasanna , Mulakala Chandrika , Mahadevan Lavanya , Skariah Neema , Kavuru Venkata Giri , Kuntrapaku Damodara , Sajja Suchitra , Zainuddin Mohammad , V Krishnakumar , Singh Ritu , V Swarnakumari , Gosu Ramachandraiah , Aiyar Jayashree , Iyer Pravin , Rajagopal Sriram 
Published: August 2015

Introduction: LSD1 is a flavin adenine dinucleotide (FAD)-dependent amine oxidase that removes methyl groups from mono- or dimethylated histone H3 lysine 4. LSD1 has been reported to be overexpressed in many malignant tumors, including breast, colon, prostate, lung, gastric cancers and others. Studies have shown that LSD1 may contribute to acute myelogenous leukaemia pathogenesis and patients that are non responders to ATRA have responded to LSD1-ATRA combination treatment in clinic. Recent studies have shown that there is cross-talk between two components of CoREST complex, LSD1 and HDAC which provides an advantage to cancer cell proliferation and survival by regulating key signalling pathways. Accordingly, combined inhibition of LSD1 and HDAC has been shown to be more efficacious in inhibiting the growth of glioblastoma, AML and breast cancer. Therefore, dual inhibitors targeting both LSD1 and HDAC could be useful in treating several cancers effectively without enhancing systemic toxicity mediated by administration of multiple drugs. To test the hypothesis, we have developed a set of molecules that either have LSD1 activity alone or dual activity on LSD1 and HDAC.

Methods: Computational chemistry approaches were used to design LSD1 specific and LSD1-HDAC dual inhibitors. To assess in vitro LSD1 potency, TR-FRET assay was used. For assessing in vitro HDAC activity fluorescence based pan-HDAC activity assay was performed. Western blotting was used to assess biomarkers of LSD1 and HDAC inhibition. Alamar blue cytotoxicity assay was used to assess cell proliferation.

Results: In vitro potency of these compounds on LSD1 ranged between,0.007 to 0.2 μM, which was comparable to literature molecules. Pan-HDAC potency of these molecules in the in vitro assay was comparable to the approved drug, Vorinostat. H3K4 di- and mono-methyl markers and H3K9 acetylation levels respectively, were used as target engagement biomarkers for LSD1and HDAC inhibition. Treatment of leukaemia cells with LSD1or LSD1-HDAC inhibitors resulted in a dose-dependent increase in the levels of H3K4 di-and mono methylation. In addition, compounds with LSD1-HDAC dual activity showed a clear increase in H3K9 acetylation levels as well. In the cell proliferation assay, compounds with LSD1 activity alone did not show any effect up to 72 h of treatment. However, compounds with LSD1-HDAC dual activity showed a strong effect in inhibiting cell proliferation in multiple cell lines, with IC50s ranging from 0.02 to 2 μM, with leukaemia cell lines being generally more sensitive than solid tumors.

Conclusion: The data obtained demonstrate that it is feasible to design dual LSD1-HDAC inhibitors that retain individual activity and potently inhibit cell proliferation, and such inhibitors could serve as powerful therapeutic agents for cancer. Further understanding of mechanisms to identify the right tool compound is in progress.

Citation Format: Dhanalakshmi Sivanandhan, Sridharan Rajagopalan, Sreekala Nair, Purushottam Dewang, Durga Prasanna Kumar, Chandrika Mulakala, Lavanya Mahadevan, Neema Skariah, Venkata Giri Kavuru, Damodara Kuntrapaku, Suchitra Sajja, Mohammad Zainuddin, Krishnakumar V, Ritu Singh, Swarnakumari V, Ramachandraiah Gosu, Jayashree Aiyar, Pravin Iyer, Sriram Rajagopal. Novel dual inhibitors of LSD1-HDAC for treatment of cancer. [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 3509. doi:10.1158/1538-7445.AM2015-3509