Abstract C048: Chemoproteomic-enabled fragment-based drug discovery target identification in small cell lung cancer

Abstract Small cell lung cancer (SCLC) has a 5-year survival rate of 7%. Most patients are diagnosed with metastatic or extensive-stage disease and present a good response to treatment, but relapse within one year. The lack of therapeutic options reflects the scarcity of known targets and specific drugs. Our study uses high-throughput fragment-based ligand discovery to unveil new targets and provide the basis for drug development. 4 SCLC and 2 non-small cell lung cancer (NSCLC) cell lines were employed in a cell viability screen with two small-molecule libraries, comprising 640 and 15,000 compounds. 83 compounds reduced viability in one (<20%) or 2 SCLC cell lines (30% both) without affecting NSCLC cell lines (>70%) and were filtered by scaffold clustering and similarity. Selected hits were validated through viability assays in 11 SCLC and 2 NSCLC cell lines. 5 probes displayed consistent selectivity and potency towards SCLC and were employed in chemoproteomics. For this, specific competitors (CPT) and the control probe (CP) were synthesized and used in the affinity enrichment assays. SCLC cell lines, H1048 and H82, were treated with CP, Probe or Probe+CPT, followed by UV crosslink, click chemistry, pulldown and analysis of tryptic peptides through liquid chromatography-tandem mass spectrometry. Our stringent comparative and statistical analysis elicited 60 high-confidence targets. CRISPR modulation (activation and inhibition) screenings were carried out to determine the effect of those targets on cell viability. H1048 and H82 cells were transduced with lentiviral particles for stable expression of dCas9-A (transcriptional activator; CRISPRa) and dCas9-I (transcriptional repressor; CRISPRi). Although none of the CRISPRa targets displayed effect on cell viability, targeting NAMPT through CRISPRi leads to a significant decrease in cell viability. NAMPT essentiality in SCLC cells was confirmed through RNAi and CRISPRi complementary assays. Pulldown, followed by Western Blot assays, confirmed the interaction between NAMPT and probe 30 (P30), which was competed for pretreatment with NAMPT inhibitors (NAMPTi), FK866, and GNE617. These results suggest that P30 binds to the NAMPT catalytic site, which is supported by docking studies. Despite the SCLC dependency on NAMPT, demonstrated by IC50 of 1-3nM for NAMPTi, media supplemented with niacin lead to cell survival upon 100nM of NAMPTi treatment, bypassing NAMPT essentiality. Surprisingly, niacin supplementation did not affect the sensitivity of SCLC to P30 to the same extent, suggesting P30 may act by inhibiting an additional target. Further validation of the NAMPT/P30 interaction is ongoing. Together, our findings highlight NAMPT as an essential target in SCLC and identify P30 as a novel NAMPT inhibitor that may exert its effects through additional mechanisms. These results provide a foundation for future target validation and medicinal chemistry efforts to expand therapeutic options in SCLC. Citation Format: Vanessa Camara Fernandes, Micheal Lamptey, Fumi Kinose, Mina Iskandar, Yi Liao, Nada Elkholy, Simon Bayle, Derek R Duckett, Eric B Haura, Andrii Monastyrskyi, Uwe Rix. Chemoproteomic-enabled fragment-based drug discovery target identification in small cell lung cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2025 Oct 22-26; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2025;24(10 Suppl):Abstract nr C048.