Abstract P3145: Enhancing Cardiac Cav1.2 Ion Channel Function By Targeted Deubiquitination

Ventricular excitation-contraction coupling relies on precise spatial arrangement of voltage-gated calcium channels CaV1.2 within T-tubular dyadic junctions proximal to ryanodine receptors (RyR2) that gate Ca2+ release from the sarcoplasmic reticulum. Impaired CaV1.2 trafficking is linked to devastating cardiovascular diseases such as arrythmias and cardiac hypertrophy. Ubiquitination is an important regulatory mechanism that controls ion channel surface trafficking and turnover. Emerging evidence hint at a prominent role for CaV1.2 ubiquitination to underlie heart failure disease progression, that suppresses CaV1.2 currents (ICa,L), resulting in harmful compensatory mechanisms. Targeted protein stabilization (TPS) is a burgeoning therapeutic approach for undruggable targets. We hypothesized that harnessing deubiquitinases using TPS can counteract pathological ubiquitination of CaV1.2, overcoming the need for hormonal remodeling and lead to novel treatment. To this end, we sought to develop engineered deubiquitinases (enDUBs) that can increase functional expression of CaV1.2 channels in the heart. The catalytic domain of ubiquitin specific protease 21 (USP21) fused to nb.F3 (F3-USP21) robustly increased CaV1.2 currents reconstituted in HEK293 cells as well as endogenous ICa,L in mammalian ventricular cardiomyocytes. This increase was within the physiological range of the impact of β-adrenergic stimulation of CaV1.2 and did not alter ventricular action potential duration. The mechanism of nbF3-USP21-induced increase in ICa,L is augmented CaV1.2 surface density. Reassuringly, this approach did not adversely interfere with the correct placement of CaV1.2 channels at dyadic junctions as cardiomyocytes expressing nb.F3-USP21 displayed normal colocalization of CaV1.2 with RyR2 at T-tubules. In conclusion, our study provides evidence that targeting deubiquitinases using TPS could be a viable therapeutic approach for rescuing CaV1.2 functional expression and these findings may pave the way for future development of TPS-based therapies for cardiovascular diseases such as arrhythmias and cardiac hypertrophy.