Cancer-Stem-Cell Phenotype-Guided Discovery of a Microbiota-Inspired Synthetic Compound Targeting NPM1 for Leukemia

The human microbiota plays an important role in human health and disease, through the secretion of metabolites that regulate key biological functions. We propose that microbiota metabolites represent an unexplored chemical space of small drug-like molecules in the search of new hits for drug discovery. Here, we describe the generation of a set of complex chemotypes inspired on selected microbiota metabolites, which have been synthesized using asymmetric organocatalytic reactions. Following a primary screening in CSC models, we identified the novel compound UCM-13369 (4b) whose cytotoxicity was mediated by NPM1. This protein is one of the most frequent mutations of AML, and NPM1-mutated AML is recognized by the WHO as a distinct hematopoietic malignancy. UCM-13369 inhibits NPM1 expression, downregulates the pathway associated with mutant NPM1 C+, and specifically recognizes the C-end DNA-binding domain of NPM1 C+, avoiding the nucleus-cytoplasm translocation involved in the AML tumorological process. The new NPM1 inhibitor triggers apoptosis in AML cell lines and primary cells from AML patients and reduces tumor infiltration in a mouse model of AML with NPM1 C+ mutation. The disclosed phenotype-guided discovery of UCM-13369, a novel small molecule inspired on microbiota metabolites, confirms that CSC death induced by NPM1 inhibition represents a promising therapeutic opportunity for NPM1-mutated AML, a high-mortality disease.This work was supported by grants PID2022-138797OB-I00, PGC2018-096049-B-I00 and PID2021-126663NB-I00 funded by MCIN/AEI/10.13039/501100011033 and by “ERDF A way of making Europe”; grant PID2019-106279RB-I00 funded by MCIN/AEI/10.13039/501100011033; grant PDC2022-133488-I00 funded by MCIN/AEI/10.13039/501100011033 and by the “European Union NextGenerationEU/PRTR”; grants PI21/00191 and CP19/00140 funded by Instituto de Salud Carlos III; CNIO agreements 2017-2020, 2020-2023 funded by Foundation CRIS contra el Cancer; grants BIO-198 and P18-FR-3487 funded by Junta de Andalucía; VI PPIT program funded by Universidad de Sevilla; and by Ramón Areces Foundation. The authors acknowledge technological support from NMR, mass spectrometry and elemental analysis CAIs (Complutense University of Madrid), Biointeractomicts Platform (cicCartuja, Seville), and the Services at CITIUS (University of Seville). S.A., A.S.-M., I.A.-A. and R.L.G.-A. are grateful to Ministerio de Ciencia e Innovación and Complutense University of Madrid for predoctoral fellowships; M.V.-E. to European Union’s Horizon 2020 for Marie Sklodowska-Curie grant; and P.A.-G. to Fundación Española de Hematología y Hemoterapia for grant. The authors thank Dr. Adrián Velázquez-Campoy at the University of Saragossa for helping in fitting ITC analysis and Prof. Miguel A. De la Rosa at the University of Seville for critical reading of the manuscript.Peer reviewe