Rigosertib reverses hypertrophic cardiomyopathy in RAF1-associated Noonan syndrome

Abstract

RASopathies are rare genetic disorders caused by mutations in genes within the canonical Ras/MAPK signaling pathway, affecting cell growth and differentiation. These syndromes, which include conditions like Noonan syndrome (NS), are characterized by developmental delays, distinctive facial dysmorphia, and a variety of cardiac defects, notably hypertrophic cardiomyopathy (HCM). Despite their prevalence and impact, therapeutic options for RASopathies remain limited. Rigosertib, a novel dual Ras/MAPK and PI3K/AKT pathway inhibitor, is currently in clinical trials for treatment of melanoma and recessive dystrophic epidermolysis bullosa. Here, we identify rigosertib as a candidate therapy for RAF1-associated HCM. Our Drosophila screen of clinically relevant drugs and compounds identified rigosertib as broadly effective across a panel of transgenic RASopathy fly transgenic models, indicating that rigosertib may be effective against multiple disease isoforms. Analysis of a Drosophila model targeting a RAF1L613V transgene to the heart found that rigosertib reduced aspects of cardiac hypertrophy. Rigosertib treatment prevented or regressed cellular hypertrophy in human induced pluripotent stem cell- (iPSC-) derived cardiomyocytes homozygous for the NS-associated RAF1S257L allele. We extended these findings to a mammalian model, using Raf1L613V/+ KI mice to explore the therapeutic implications of rigosertib on RAF1-driven HCM. Longitudinal six-week treatment with rigosertib in these mice resulted in significant improvement in left ventricular chamber dimension and posterior wall thickness, total heart mass, size of individual cardiomyocytes (CMs), as well as reversal of cardiac hypertrophy. Rigosertib treatment also led to normalized fetal gene expression and inhibition of ERK and AKT pathway activities in primary CMs isolated from Raf1L613V/+ mice. Cardiac function, as assessed by echocardiography, showed significant improvement in ejection fraction and fractional shortening, with molecular studies confirming downregulation of hypertrophic markers and signaling pathways. Together with the Drosophila data, these mammalian results support the potential and use for rigosertib to reverse pathological hypertrophy in NS through targeted pathway inhibition in patients. Moreover, in addition to its effects in the heart, rigosertib treatment in mice also significantly improved other NS-associated syndromic features, including increasing bone growth and correcting craniofacial abnormalities. Our findings suggest rigosertib effectively normalizes and reverses RASopathy-associated HCM as well as other NS-associated syndromic features, supporting its potential for development as a promising treatment for RAF1-associated HCM and, potentially, other RASopathies-dependent pathologies. This study not only highlights the therapeutic potential of rigosertib but also demonstrates the utility of an integrated approach using Drosophila, iPSC and mammalian models to elucidate drug effects across complex biological systems.

Competing Interest Statement

R.L.C., B.D.G., and M.I.K. received grant funding support for this project from Onconova Therapeutics (now Traws Pharma), the developer of rigosertib; Onconova, however, was not involved in the study design, execution or interpretation. M.I.K. is also a consultant for BioMarin Pharmaceutical Inc., but this work is independent of the project conducted herein. B.D.G. is a named inventor on issued patents related to PTPN11, SHOC2, RAF1, and SOS1 NS mutations. Mount Sinai has licensed the patent to several diagnostics companies and has received royalty payments, some of which are distributed to B.D.G. Previously, B.D.G. received financial compensation as a consultant for Day One Therapeutics and BioMarin, companies focused on developing a MEK inhibitor as potential therapy for NS and other RASopathies. B.D.G. also previously received grant funding from Day One Therapeutics for a study of MEK inhibition as a treatment for the RASopathies.