Targeting conditioned media dependencies and FLT-3 in chronic lymphocytic leukemia.

The importance of the stromal microenvironment in chronic lymphocytic leukemia (CLL) pathogenesis and drug resistance is well established. Despite recent advances in CLL therapy, identifying novel ways to disrupt interactions between CLL and its microenvironment may identify new combination partners for the drugs currently in use. To understand the role of microenvironmental factors on primary CLL cells, we took advantage of an observation that conditioned media (CM) collected from stroma was protective of CLL cells from spontaneous cell death ex vivo. The cytokine in the CM-dependent cells that most supports CLL survival in short term ex-vivo culture was CCL2. Pre-treatment of CLL cells with anti-CCL2 antibody enhanced venetoclax-mediated killing. Surprisingly, we found a group of CLL samples (9 of 23 cases) that are less likely to undergo cell death in the absence of CM support. Functional studies revealed that CM-independent (CMI) CLL cells are less sensitive to apoptosis than conventional stroma-dependent CLL. Additionally, a majority of the CMI CLL samples (80%) harbored unmutated IGHV. Bulk-RNA Seq analysis revealed upregulation of the focal adhesion and Ras signaling pathways in this group, along with expression of FLT3 and CD135 expression. Treatment with FLT3 inhibitors caused a significant reduction in cell viability among CMI samples. In summary, we were able to discriminate and target two biologically distinct subgroups of CLL based on CM dependence with distinct microenvironmental vulnerabilities.