Supplementary Methods from The Pan-RAF–MEK Nondegrading Molecular Glue NST-628 Is a Potent and Brain-Penetrant Inhibitor of the RAS–MAPK Pathway with Activity across Diverse RAS- and RAF-Driven Cancers

Ryan, Meagan B.; Quade, Bradley; Schenk, Natasha; Fang, Zhong; Zingg, Marshall; Cohen, Steven E.; Swalm, Brooke M.; Li, Chun; Özen, Ayşegül; Ye, Chaoyang; Ritorto, Maria Stella; Huang, Xin; Dar, Arvin C.; Han, Yongxin; Hoeflich, Klaus P.; Hale, Michael; Hagel, Margit

doi:10.1158/2159-8290.26417057.v1

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Supplementary Methods from The Pan-RAF–MEK Nondegrading Molecular Glue NST-628 Is a Potent and Brain-Penetrant Inhibitor of the RAS–MAPK Pathway with Activity across Diverse RAS- and RAF-Driven Cancers

journal contribution

posted on 2024-07-31, 18:00 authored by Meagan B. Ryan, Bradley Quade, Natasha Schenk, Zhong Fang, Marshall Zingg, Steven E. Cohen, Brooke M. Swalm, Chun Li, Ayşegül Özen, Chaoyang Ye, Maria Stella Ritorto, Xin Huang, Arvin C. Dar, Yongxin Han, Klaus P. Hoeflich, Michael Hale, Margit Hagel

Supplementary methods with detailed protein production, mass spectrometry, in vivo, and in vitro methodology

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ARTICLE ABSTRACT

Alterations in the RAS–MAPK signaling cascade are common across multiple solid tumor types and are a driver for many cancers. NST-628 is a potent pan-RAF–MEK molecular glue that prevents the phosphorylation and activation of MEK by RAF, overcoming the limitations of traditional RAS–MAPK inhibitors and leading to deep durable inhibition of the pathway. Cellular, biochemical, and structural analyses of RAF–MEK complexes show that NST-628 engages all isoforms of RAF and prevents the formation of BRAF–CRAF heterodimers, a differentiated mechanism from all current RAF inhibitors. With a potent and durable inhibition of the RAF–MEK signaling complex as well as high intrinsic permeability into the brain, NST-628 demonstrates broad efficacy in cellular and patient-derived tumor models harboring diverse MAPK pathway alterations, including orthotopic intracranial models. Given its functional and pharmacokinetic mechanisms that are differentiated from previous therapies, NST-628 is positioned to make an impact clinically in areas of unmet patient need.Significance: This study introduces NST-628, a molecular glue having differentiated mechanism and drug-like properties. NST-628 treatment leads to broad efficacy with high tolerability and central nervous system activity across multiple RAS- and RAF-driven tumor models. NST-628 has the potential to provide transformative clinical benefits as both monotherapy and vertical combination anchor.See first author Meagan B. Ryan discuss this research article, published simultaneously at the AACR Annual Meeting 2024: https://vimeo.com/932606757/4d0bd5aa98