Abstract B21: Novel pateamine A analogs to target pro-survival proteins in chronic lymphocytic leukemia

Abstract

Abstract Introduction: Dysregulation of protein translation is a common feature of cancer. Because the great majority of oncoproteins turn-over rapidly, thus have an acute need for mRNA translation compared to housekeeping proteins, protein synthesis is a promising avenue to explore for targeted cancer therapy. The marine natural product pateamine A (PatA) inhibits cap-dependent translation initiation by binding to the eukaryotic initiation factor 4A (eIF4A). We have previously synthesized a family of PatA-based small molecules. The lead compound, des-methyl, des-amino pateamine A (DMDAPatA), is more stable and easier to synthesize than the parent natural product, and demonstrated anti-cancer activities in both cell lines and animal models. In this study, we extended the investigations of the activities of DMDAPatA in B cell malignancies, focusing on the primary chronic lymphocytic leukemia (CLL) cells. We propose to target the biological rewiring of the CLL cells, namely the overexpression of short-lived pro-survival proteins that keep these cells from undergoing apoptosis. We hypothesize that a transient inhibition of translation will cause a lethal decrease in those labile pro-survival proteins in CLL cells that will initiate apoptosis. Experimental Procedures: Primary CLL cells were used to screen the PatA analogs for their anti-leukemia activity. Cell viability was analyzed by flow cytometry. Protein translation was measured by tritium labeled leucine incorporation. Protein expression was determined by immunoblotting. Results: Our data showed that DMDAPatA blocked protein synthesis in the CLL cells. This action reduced the short-lived anti-apoptotic proteins Mcl-1 and XIAP, and initiated apoptosis in as short as 4 hr. The levels of the more stable anti-apoptotic proteins Bcl-2 and Bcl-XL were not affected. This is consistent with the selective activity of translation inhibitors toward proteins with rapid turn-over rates. Since DMDAPatA depleted Mcl-1 without affecting Bcl-2, and the Bcl-2 antagonist ABT-199 inhibits Bcl-2 activity but spares Mcl-1, the combination of DMDAPatA and ABT-199 targeted both arms of apoptosis control and killed CLL cells synergistically. Dose reduction index analysis demonstrated mutual potentiation of ABT-199 and DMDAPatA. However, in the presence of human plasma proteins, DMDAPatA lost its potency substantially. A plasma protein binding analysis showed over 99% binding in human plasma, which may hamper its development as a therapeutic agent. To address this issue, we synthesized a new series of PatA analogues with the goal of improving the physical properties and CLL potency. We found that any alteration on the binding domain of PatA, either on eastern half of the macrolide ring or the side chain, resulted in dramatic loss of activity toward the CLL cells. The scaffolding domain on the western side of the ring was more tolerable to modifications which made it feasible to improvement. We proposed to introduce an amino group at either the C2 (alpha-amino variant) or C3 position (beta-amino variant) of the macrolide ring to lower lipophilicity and reduce binding to plasma proteins. This strategy led to the identification of three new leads with potent inhibition of protein synthesis. They exhibited greater cytotoxic potency toward CLL cells than DMDAPatA and lower human plasma protein binding. They also showed better selectivity towards CLL cells over normal lymphocytes compared to the parental compound PatA, demonstrating improved therapeutic index. Conclusions: We have successfully synthesized three new PatA analogs with potent anti-leukemia activities. These compounds hold promise for application to cancers with the right biological context, such as CLL, in which the leukemia cells are addicted to the sustained expression of short-lived oncoproteins for survival. Citation Format: Rong Chen, Mingzhao Zhu, Yuling Chen, Wesley Skillern, Qun Qin, William G. Wierda, Kenneth G. Hull, Daniel Romo, William Plunkett. Novel pateamine A analogs to target pro-survival proteins in chronic lymphocytic leukemia. [abstract]. In: Proceedings of the AACR Special Conference on Translational Control of Cancer: A New Frontier in Cancer Biology and Therapy; 2016 Oct 27-30; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2017;77(6 Suppl):Abstract nr B21.