Abstract 5436: Targeting oncoproteins via disruption of proteostasis: Identification of oncoprotein destabilizing agents using luciferase tagged oncoproteins

Abstract

Abstract Since many oncoproteins are considered undruggable targets, we have developed and validated a new screening strategy to identify small molecules that inhibit oncoprotein activity by causing their degradation. Recently, the mechanism of action for the class of drugs known as immunomodulators (IMiDs; e.g. thalidomide, lenalidomide and pomalidomide) has uncovered a novel oncogene targeting strategy. IMiDs commandeer an E3 ligase complex that contains the E3 recognition subunit cereblon and divert its activity toward two transcription factors IKZF1 and IKZF3, which play critical roles in hematopoietic cells. This mechanism explains the efficacy of IMiDs in multiple myeloma and may translate into a general approach to target traditionally undruggable oncoproteins. We generated cell lines stably expressing IKZF1as a firefly luciferase fusion protein with coexpression of Renilla luciferase. A pilot HTS campaign was run to uncover novel drugs that can mimic the effects of IMiDs. A cell line expressing the MYC-firefly fusion protein was used for counter-screening. We used pomalidomide as our positive control and it reduced firefly luciferase signal ≈75% with no change in the renilla luciferase signals. This control resulted in a Z prime value typically >0.5. The pilot screen of 3,850 compounds resulted in 39 “hits” that reduced firefly luciferase >35%. More importantly, both thalidomide and lenalidomide were found to be active in the screen. Confirmation testing revealed that 75% of “hits” repeated, but also had activity in the MYC-firefly cell line. Actives were tested in western blots for their ability to decrease fusion protein abundance. Many active compounds did not reduce protein levels in western blots and represent nonspecific hits (e.g. luciferase inhibitor). In conclusion, we present a screening paradigm with the potential to uncover novel proteostasis disruptors by expressing luciferase tagged oncoproteins. Citation Format: Richard E. Middleton, Greg Olsen, Russell McSweeney, Gang Lu, Wenhua Gao, Justin Roberts, Michael R. McKeown, Mark A. Bittinger, Kwok-Kin Wong, James E. Bradner, William G. Kaelin. Targeting oncoproteins via disruption of proteostasis: Identification of oncoprotein destabilizing agents using luciferase tagged oncoproteins. [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 5436. doi:10.1158/1538-7445.AM2015-5436