An unbiased screen identified the Hsp70-BAG3 complex as a regulator of myosin binding protein C3

Abstract

Objective: We aim to identify regulators of myosin binding protein C3 (MyBP-C) protein homeostasis. Background: Variants in myosin binding protein C3 (MYBPC3) account for approximately 50% of familial hypertrophic cardiomyopathy (HCM). Most pathogenic variants in MYBPC3 are truncating variants that lead to reduced total levels of MyBP-C protein. Elucidation of the pathways that regulate MyBP-C protein homeostasis could uncover new therapeutic strategies that restore normal protein levels. Method: We developed a high-throughput screen to identify compounds that can increase or decrease steady-state levels of MyBP-C in an induced pluripotent stem cell cardiomyocyte (iPSC-CM) model derived from a patient with HCM. To normalize results, we also monitored effects on myosin heavy chain (MYH) and focused on those molecules that selectively modulated MyBP-C levels. Results: Screening a library of 2,426 known biologically active compounds, we identified compounds which either decreased (241/2426, 9.9%) or increased (29/2426, 1.2%) MyBP-C/MYH levels. After a rigorous validation process, including a counter screen for cellular toxicity, two compounds (JG98 and parthenolide) were confirmed as decreasing MyBP-C levels and no compounds were confirmed to increase MyBP-C levels. For further studies, we focused on JG98, which is an allosteric modulator of heat shock protein 70 (Hsp70), inhibiting its interaction with BAG domain co-chaperones. We found that genetic reduction of BAG3 phenocopies treatment with JG98 by reducing MyBP-C protein levels. Conclusion: An unbiased compound screen identified the Hsp70-BAG3 complex as a regulator of MyBP-C stability. Thus, approaches that stimulate function of this complex may be beneficial in the treatment of HCM.

Going Further With Half as Much

“…This would potentially accelerate the deterioration of function and progression of disease. As suggested by Thompson et al 1 , approaches that increased BAG3 protein levels or increased its activity could be therapeutically beneficial in these patients. Supporting this hypothesis, we previously found that adeno-associated virus–mediated overexpression of BAG3 was able to rescue sarcomere function in a mouse heart failure model.…”

“…This is the approach that Thompson et al 1 have taken in this issue of JACC: Basic to Translational Science . The goal of this study was to identify a small molecule that could enhance the level of cMyBPC protein in a model of haploinsufficiency.…”

“…Despite progress in recent years, many steps in these processes remain black boxes, making informed drug development impossible at the moment. Studies such as Thompson et al’s 1 are critical as they reveal some of these mechanistic aspects that bring us closer to such goals, but more work is necessary.…”

Personalized allele-specific CRISPR-Cas9 strategies for myofibrillar myopathy 6

“…Since the disease is dominant and severe, most MFM6 cases result either from de novo mutation or from inheritance of the mutation from a parent who is a genetic mosaic for the mutation [4,[6][7][8]. Although certain small molecule drugs showed improvements in pre-clinical models [9][10][11][12], there are no effective treatments for MFM6.…”

“…Since the disease is dominant and severe, most MFM6 cases result either from de novo mutation or from inheritance of the mutation from a parent who is a genetic mosaic for the mutation [4, 6-8]. Although certain small molecule drugs showed improvements in pre-clinical models [9-12], there are no effective treatments for MFM6. Therefore, we reasoned that advancements in genome engineering may provide alternative routes for effective treatment strategies for MFM6, given its well-known genetic underpinning.…”