Noonan Syndrome-Associated Hypertrophic Cardiomyopathy Caused by a Mutation in RIT1 Can Be Partially Rescued by Inhibition of RAS/MAPK Signaling Pathway In Vitro

Objectives: Noonan syndrome (NS) is caused by gain-of-function mutations in various genes encoding components of the RAS/MAPK signaling pathway. Among them, specific mutations of RIT1, such as the base pair substitution c.246T>G resulting in RIT1F82L/+, are strongly associated with hypertrophic cardiomyopathy (HCM). So far, affected individuals can only be treated symptomatically, since the underlying pathophysiology of this HCM is barely understood.

Methods: To elucidate the role of RIT1F82L/+ in myocardial tissue in vitro, cardiomyocytes differentiated from patient-derived induced pluripotent stem cells (iPSCs) were used to generate rod-shaped 3D bioartificial cardiac tissues (BCTs). These were monitored longitudinally for morphological and physiological characterization of the cardiac phenotype and to determine the effect of treatment with the MEK inhibitor trametinib (TB). This evaluation was performed by using a multimodal bioreactor set-up and microscopic assessment. Two individual wild type (WT) iPSC lines were used as controls.

Result: We found that the cross-sectional area was enlarged in RIT1F82L/+ BCTs compared with WT BCTs, by >60% (1.62 vs. 0.97 mm², p < 0.0001), indicating HCM. Moreover, the contractile force and tension of RIT1F82L/+ BCTs were significantly impaired compared with the WT BCTs (2.87 vs. 4.36 mN, p < 0.001 and 1.81 kPa vs. 4.94 kPa, p < 0.0001, respectively). In addition, the characteristics of a single contraction, i.e., time to peak and time to 80% relaxation, were significantly reduced in the RIT1F82L/+ BCTs (0.22 vs. 0.26 milliseconds, p < 0.001 and 0.16 milliseconds vs. 0.24 milliseconds, p < 0.01, respectively). Abnormal contraction characteristics were partially prevented by treatment with TB. Myocardial thickening however was unaffected. The spontaneous contraction frequency of RIT1F82L/+ BCTs was unaltered in comparison to WT BCTs and was not affected by TB. Even after phenotype manifestation, the impairment of the contractile force could have been stopped by TB causing higher contractile tensions as in the untreated BCTs.

Conclusion: In this study, we successfully phenocopied the RIT1F82L/+-associated myocardial phenotype related to NS in vitro including myocardial thickening and impaired contractility. Moreover, our preliminary but promising results show that the phenotype was partially rescued by pharmacological intervention. Thus, this system can now be used for further pharmacological in vitro studies to elucidate the underlying pathomechanism.

Publication History

Article published online:
19 February 2021