Abstract LB-C19: The investigation of structural determinants that mediate selective inhibition of UBA5, the UFM1 activating enzyme

Abstract

Abstract Protein conjugation with the UFM1 small protein modifier promotes cellular survival in times of ER stress, which is a common occurrence within diseased cells. We have developed the first selective inhibitor of UBA5, the E1 activating enzyme responsible for initiating UFM1 conjugation. Based on the crystal structure of human UBA5, we identified acidic residues in close proximity to the ATP pocket that we chose to target using a zinc-coordinated polyazamacrocycle. This coordination complex was appended to adenosine through alkyl linkers of varying length in order to probe for optimal inhibition of UBA5 activity. Through this small structure-activity relationship (SAR), we identified our lead inhibitor 5C-Z that disrupts UBA5-mediated UFM1 conjugation in vitro through a non-competitive mode of action (IC50 = 4.1 μM, 95% Confidence Interval (C.I.) = 2.1-7.7 μM). Our compound demonstrates selectivity for UBA5 over other E1 enzymes in vitro, as well as over nearly 100 human kinases as evaluated using a kinome screen, despite containing adenosine within its structure. Currently, we are investigating the structural determinants that are indispensible for inhibitory activity. We are making iterative changes by varying each component of 5C-Z including the metal centre, the polyazamacrocycle, the linker, the ribose as well as the nitrogenous base. To date, we have shown that other transition metals are not as active as the zinc (II) centre, increasing the size of the macrocycle abolishes activity and reducing the linkage from the polyazamacrocycle to the linker also significantly abrogates activity. Investigating the role of each functional group on 5C-Z will help delineate the mechanism by which it selectively inhibits UBA5 activity and may shed light in progressing towards a more drug-like small molecule inhibitor. Citation Format: Stacey-Lynn Paiva, Sara da Silva, Mathew Bancerz, Mulu Geletu, Andrew M. Lewis, Jijun Chen, Yafei Cai, Julie L. Lukkarila, Honglin Li, Patrick T. Gunning. The investigation of structural determinants that mediate selective inhibition of UBA5, the UFM1 activating enzyme. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr LB-C19.