Abstract 7332: Investigating genes and pathways determining sensitivity or resistance to anti-L1cam ADCs in lymphomas

Abstract Background: L1 cell adhesion molecule (L1CAM) is a glycoprotein initially characterized for its role in neuronal migration, cell adhesion, axonal growth, and synaptogenesis. In recent years, it has been identified in various solid tumors, in which it promotes invasion, metastasis, and resistance to therapies. We previously demonstrated the expression of L1CAM in a subset of lymphomas, mainly mantle cell lymphoma (MCL) and cutaneous T-cell lymphomas, determining in vitro and in vivo sensitivity to anti-L1CAM antibody-based therapies (Spriano et al., AACR-NCI-EORTC 2023). This project aimed to define genes and pathways contributing to the anti-lymphoma activity of L1CAM targeting antibody-drug conjugates (ADCs) in L1CAM-positive lymphomas. We used the Z138 MCL cell line as a model system exposed to two ADCs, sharing the same antibody backbone but with different payloads, the DNA targeting SG3199 (ADC6) and the tubulin targeting MMAE (ADC4), respectively. Methods: We generated stable Cas9 or dCas9-VP64 expressing Z138 cells, then infected them with the Brunello (LOF) or the Calabrese (GOF) library. After the appropriate selection, cells were split to obtain the baseline representation of sgRNA distribution (T0) and then left untreated or treated with either of the two anti-L1CAM ADCs (exp arms) for two weeks. MAGeCK-MLE was used to model sgRNA read counts and assess the sgRNA β values and statistical enrichment (FDR<0.1). Combination treatments were performed, exposing cells to increasing concentrations of both drugs for three days, followed by an MTT assay. Results: L1CAM expression was strongly associated with sensitivity to ADC4 but not to ADC6. As already reported and demonstrated also by us, the SG3199 payload is highly potent, rendering its activity less dependent on L1CAM levels. Among the genes sustaining resistance to ADC4, we identified FBXW7 and SOX11 (known L1CAM regulators), kinesins involved in microtubule assembly, MDR1, BCL2, CASP9, and ARID1A; TP53, BAX, CD20, CD19, RC3H1, and FCGR1B were identified as genes providing sensitivity to ADC4. Among the genes promoting resistance to ADC6, we identified RAD51B, ATR, BCL2, CHD4, and genes involved in the mitochondrial respiratory chain. Among the genes related to ADC6 sensitivity, we identified CDKN2B, RIPK1, MAPK9, RC3H1, NFKB1A, and RIF1. Finally, due to the observed involvement of ATR and BCL2 in the resistance to ADC6, we tested the addition of an ATR inhibitor (elimusertib) or a BCL2 inhibitor (venetoclax) to ADC6 in the Z138, achieving a strong synergism and additivity/synergism, respectively. Conclusion: LOF and GOF CRISPR-Cas9 screenings identified genes related to resistance or sensitivity to anti-L1CAM ADCs that could be potentially used as therapeutic targets or biomarkers. Citation Format: Filippo Spriano, Giulio Sartori, Chiara Tarantelli, Luciano Cascione, Sara Napoli, Flavio Mehli, Gunther Spohn, Francesco Bertoni. Investigating genes and pathways determining sensitivity or resistance to anti-L1cam ADCs in lymphomas [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 7332.