Abstract 4885: Diverse microenvironmental agonists induce de novo resistance to Bcl-2 and BTK targeted combination therapy and cancer cell proliferation in B cell malignancies
Authors: Jayappa Kallesh D. , Portell Craig A. , Gordon Vicki L. , Bender Timothy P. , Williams Michael E. , Weber Michael J. 
Published: August 2018

Abstract
The Bruton's Tyrosine Kinase (BTK) inhibitor ibrutinib (IBR) is FDA approved for Chronic Lymphocytic Leukemia (CLL) and Mantle Cell Lymphoma (MCL), but de novo resistance and rapid recurrence are common. We showed that venetoclax (VEN), an inhibitor of Bcl-2, induced synergistic cytotoxicity with IBR in cell lines and patient samples ex vivo (Axelrod et al. Leukemia, 2014; Jayappa et al. Blood Adv., 2017) and have initiated a clinical trial to test this combination. However, we noted variable sensitivity to IBR+VEN among CLL/MCL patient samples ex vivo, indicating a high frequency of de novo resistance. Resistance was evident in CLL cells showing an “activation” phenotype (CD5+/CD19+/CD69+) that occurs from interactions with the microenvironment in vivo, suggesting these interactions could induce resistance. This hypothesis was supported when we found that the combination of CpG-ODN, soluble CD40L (sCD40L), and IL10 (“agonist mix”) generated resistance to IBR+VEN in most CLL/MCL samples.
In this study, we explored the role of innate immune receptors and other microenvironmental agonists in drug resistance and cancer cell proliferation, singly and in combination. We analyzed induction of apoptosis by IBR+VEN in CLL/MCL patient PBMCs cultured with agonists of various TLRs, NOD1/2, CD40, and IL10R by FACS. Significant resistance to apoptosis by IBR+VEN was noted in most patient samples treated with TLR9 agonist CpG-ODN (10/12) or sCD40L (9/12). IL10 induced modest resistance in a few samples (4/12). TLR1/2, TLR7, and NOD1/2 agonists generated moderate resistance only in MCL samples (N=4). Our preliminary FACS data suggest that CLL cells expressing ZAP-70, a marker of poor disease prognosis, responded with enhanced proliferation to CpG-ODN, sCD40L, and agonist mix as compared to ZAP-70- cells (3/3). In addition, prior exposure to CpG-ODN enhanced the proliferation and drug resistance response to sCD40L and vice versa (4/4), predicting mutually reinforcing microenvironmental interactions in vivo. CpG-ODN or agonist mix induced NF-kB dependent over expression of Mcl-1 and Bcl-xL in CLL/MCL cells, and increased ratio of these proteins to the cognate pro-apoptotic proteins (Puma, BIM, and BAD), indicating enhanced dependence of leukemic B cells for Mcl-1 and Bcl-xL. This presumably caused resistance by lowering cancer cell dependence for VEN target Bcl-2 in presence of agonists.
In conclusion, several microenvironmental agonists, particularly the agonist of TLR9 and CD40, induce resistance to IBR+VEN and cell proliferation in CLL/MCL. Response to these agonists is enhanced in ZAP-70+ cells and when agonist exposure occurs combinatorially. Drug resistance by these diverse agonists was generated by a convergent mechanism of NF-kB dependent over-expression of Mcl-1 and Bcl-xL, which provide additional targets to overcome drug resistance.
Citation Format: Kallesh D. Jayappa, Craig A. Portell, Vicki L. Gordon, Timothy P. Bender, Michael E. Williams, Michael J. Weber. Diverse microenvironmental agonists induce de novo resistance to Bcl-2 and BTK targeted combination therapy and cancer cell proliferation in B cell malignancies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4885.