Abstract 2866: NAPRTsilencing in rhabdomyosarcoma confers therapeutic vulnerabilities to NAD+depletion
Authors: Zhao Sophia J. , Noronha Katelyn J. , Bhardwaj Prateek , Lucas Karlie N. , Sundaram Ranjini K. , Heer Collin D. , Morotti Raffaella , Spurrier Josh , Raponi Mitch , Bindra Ranjit S. , Vasquez Juan C. 
Published: March 2024

Abstract
Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma among children, and new treatments are needed to improve survival. Cells primarily synthesize nicotinamide adenine dinucleotide (NAD) through the Preiss-Handler and Salvage pathways driven by nicotinic acid phosphoribosyltransferase (NAPRT) and nicotinamide phosphoribosyltransferase (NAMPT), respectively. NAMPT inhibitors (NAMPTi) have been tested in clinical trials, but their efficacy has been limited by (a) lack of appropriate biomarkers for patient selection, and (b) significant dose-limiting toxicities (DLTs), including myelosuppression and retinal toxicity, although the latter has been described only in preclinical studies. Previous work from our group indicates that loss of NAPRT expression by tumor-specific promoter CpG island methylation may provide a novel biomarker for sensitivity to NAMPTi in a variety of tumors. Here, we sought to determine if RMS harbors NAPRT silencing that confers a synthetic lethal interaction with NAMPTi. We probed methylation data within the Cancer Dependency Map (DepMap) and found a high frequency of NAPRT gene promoter hypermethylation in RMS models. Predicted NAPRT silencing was confirmed at the protein level by western blot in a panel of RMS cell lines. In vitro growth delay assays demonstrated that NAPRT-silenced cells are exquisitely sensitive to NAMPTi even in the presence of nicotinic acid (NA) supplementation, while NAPRT-expressing cells were completely rescued. This sensitivity to NAMPTi was further mirrored by changes in NAD levels. In vivo, NAMPTi induced significant tumor regression in a NAPRT-silenced RMS xenograft model. To further establish the translational relevance, NAPRT protein expression in a TMA of 51 human RMS tumors was assessed via IHC staining using a newly validated NAPRT monoclonal antibody (4A5D7). Based on a semi-quantitative scoring system accounting for the percentage of positive tumor cells and staining intensity, 28% of RMS samples demonstrated loss of NAPRT protein expression. Overall, these data suggest that NAPRT loss may serve as a therapeutic target in RMS by inducing a synthetic lethal interaction with NAD+ depleting agents. Future studies in patient-derived xenograft (PDX) models will determine the optimal cut-off of NAPRT expression that portends NAMPTi sensitivity, paving the way for the development of biomarker-driven clinical trials in RMS. Ongoing studies will also evaluate the efficacy of combining NAMPTi with existing cytotoxic and DNA damaging chemotherapy in these difficult-to-treat tumors.
Citation Format: Sophia J. Zhao, Katelyn J. Noronha, Prateek Bhardwaj, Karlie N. Lucas, Ranjini K. Sundaram, Collin D. Heer, Raffaella Morotti, Josh Spurrier, Mitch Raponi, Ranjit S. Bindra, Juan C. Vasquez. NAPRTsilencing in rhabdomyosarcoma confers therapeutic vulnerabilities to NAD+depletion [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2866.