Abstract 4232: Small molecule phenotypic targeting of aggressive B-cell lymphomas

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Diffuse large B-cell lymphoma (DLBCL) is a heterogeneous disease. Combination of gene expression and metabolic profiling of DLBCLs uncovered two biologically relevant entities: BCR-DLBCL (∼60% of DLBCL pts) characterized by a glycolytic metabolism and B-cell receptor (BCR) signaling activation; and OXPHOS-DLBCL characterized by enhanced mitochondrial energy transduction and increased glutathione levels. To identify small molecules that can serve as biological probes and potential therapeutics for these subgroups, we conducted a phenotypic screening in BCR-DLBCLs vs. OXPHOS-DLBCLs. We first tested cell growth inhibition capacity of 84,000 drug-like small molecules in one BCR (OCI-Ly7) vs. OXPHOS (Toledo) cell lines for 48 h at 20 µM. We identified 3,684 compounds that inhibited >90% the growth of either or both cell lines. In a secondary screening we selected compounds that showed a dose-response effect lower than 10 µM. We identified 31 BCR-selective, 8 OXPHOS-selective and 5 non-selective compounds. We then determined potency and phenotypic-selectivity of active compounds in an expanded panel of BCR-glycolytic (OCI-Ly1, OCI-Ly3, OCI-Ly7, OCI-Ly10, SU-DHL6) vs. OXPHOS (OCI-Ly4, Karpas422, Toledo, WSU-DLCL2) cell lines. Five compounds were validated as BCR-glycolytic specific and 3 as bi-specific and their GI50 values (50% growth inhibition) were determined. We further characterized the most selective (>5-fold) and potent (low µM) BCR-glycolytic specific small molecules, termed LI-2 and LI-5. These compounds showed GI50 between 500 nM - 3 µM in BCR-glycolytic vs 9-20 µM in OXPHOS DLBCLs. LI-2 and LI-5 selectively induced 60-80% caspase-dependent apoptosis (p≤0.05) after 24 h at 2.5 µM in BCR-glycolytic but not in OXPHOS. To determine phenotypic changes upon treatment with LI-2 and LI-5, we performed RNA-sequencing on BCR-glycolytic DLBCL OCI-Ly7 and SUDHL-6 cells. LI-2 signature included 91 up- and 76 down-regulated transcripts (FC>1.2, FDR adjusted p<0.05), whereas LI-5 comprised 493 up- and 248 down-regulated genes. Computational analysis revealed that both compounds affected survival and cell cycle pathways (p=5.2x10-7) as well as amino acid and glucose metabolism pathways (p=2.4x10-8). LI-2 specifically affected DNA mismatch repair and replication pathways (p=3.9x10-5), and LI-5 specifically affected cytokine and growth factors signaling pathways (p=2.4x10-24). When compared to other known drug effects, LI-2 showed an expression signature similar to valinomycin (an OXPHOS uncoupler) and to pyrvinium (that kills cancer cells upon glucose starvation), suggesting that activity of TCA and glycolytic pathways are both critical for BCR-glycolytic cells survival. LI-5 relates to the protein synthesis inhibitor anisomycin and the cell cycle inhibitor podophyllotoxin. In sum, phenotypic screening provides a basis for discovery of distinct susceptibilities of DLBCL subtypes that could be capitalized towards individualized therapies. Citation Format: Mariano G. Cardenas, Matt R. Teater, Nieves Calvo-Vidal, Shao Ning Yang, J F. Glickman, Ari Melnick, Leandro Cerchietti. Small molecule phenotypic targeting of aggressive B-cell lymphomas. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4232. doi:10.1158/1538-7445.AM2014-4232