Interaction between CLL and its microenvironment is a major factor in drug resistance. Bone marrow derived stroma cells provide a protective and supportive niche to allow CLL cells to survive drug-induced apoptosis. Therefore, a current challenge is to develop new strategies to target the microenvironment. To understand the role of tumor microenvironment in CLL survival, we studied conditioned media (CM) derived from NKTert cells. Human NKTert stroma cells provide a survival advantage to primary CLL cells by protecting them from spontaneous apoptosis in ex-vivo culture. We identified three cytokines (IL6, IL8 and CCL2) from CM as candidates for mediating survival. We exposed CLL cells to neutralizing antibodies against each cytokine and found significant decrease in CLL viability in presence of anti-CCL2 suggesting its role in CLL survival. BH3 profiling revealed an increase in apoptotic priming with anti-CCL2 treatment as well. Using dynamic BH3 profiling (DBP), we found that anti-CCL2 increased dependence on BCL-2 as well as BCL-xL in primary CLL cells suggesting a potential role of CCL2 on CLL survival. To our surprise, we also identified a CLL group (CM independent, CMI) that were resilient to cell death in the absence of stroma support in ex-vivo culture. In addition to this, anti-CCL2 antibody showed no effect on their survival. Next, we exposed CMI CLL cells to venetoclax and found that these CLL cells were less sensitive to apoptosis compared to the CLL samples that are stroma dependent (CM dependent, CMD). BH3 profiling showed that CMI CLL cells are less primed for apoptosis. Consistent with our findings, DBP with anti-CCL2, venetoclax or the combination didn’t increase any drug induced apoptotic priming in CMI CLL samples. In our study, 80% of CMI harbor unmutated IgHV. To provide additional insight to the molecular mechanism, we performed western blot analyses for BCL-2 family proteins and found decreased BAX ad BCL-2 expression in CMI samples. This result suggests altered BCL-2 family proteins expression might be a mechanism for differential priming. Bulk RNA seq analysis revealed upregulation of focal adhesion genes and Ras signaling pathway in CMI samples. In summary, our study suggest that in the CMD subset of CLL, anti-CCL2 therapy enhances apoptotic priming and venetoclax efficacy. Furthermore, we have also functionally defined two biologically distinct CLL subgroups based on stroma dependence.

Citation Format: Salma Parvin, Aditi Aryal, Shanye Yin, Matthew S. Davids, Geoffrey G. Fell, Catherine J. Wu, Anthony Letai. Targeting tumor-microenvironmental dependencies as a therapeutic approach in Chronic Lymphocytic Leukemia (CLL) [abstract]. In: Proceedings of the Third AACR International Meeting: Advances in Malignant Lymphoma: Maximizing the Basic-Translational Interface for Clinical Application; 2022 Jun 23-26; Boston, MA. Philadelphia (PA): AACR; Blood Cancer Discov 2022;3(5_Suppl):Abstract nr A39.