SERCA2a-activating PLB mutant improves calcium handling in dilated cardiomyopathy model of hiPSC-CMs

There is increasing momentum toward the development of gene therapy for heart failure, cardiomyopathy, and chronic cardiac diseases attributed to impaired calcium (Ca) transport and reduced contractility. Here, we established a novel FRET assay using fluorescently-tagged SERCA2a (the cardiac Ca pump) and PLB (its endogenous regulator) that directly tests the capacity of loss-of-function PLB mutants (PLB M ) to compete with wild-type PLB (PLB WT ) to relieve SERCA2a inhibition. The combination of a mutation conferring loss-of-inhibitory-function (L31A-PLB M ) and a mutation associated with increased binding affinity (I40A-PLB M ) resulted in a PLB M (L31A/I40A) capable of displacing PLB WT . Overexpression of L31A/I40A-PLB M increased Ca-ATPase activity in PLBWT-expressing HEK293-6E cells. In human induced pluripotent stem cell derived cardiomyocytes (hiPSC-CMs) from healthy individuals, recombinant adeno-associated virus 2 (rAAV2) driven expression of L31A/I40A-PLB M significantly enhanced Ca-release amplitude and removal. Furthermore, L31A/I40A-PLB M expression in a hiPSC-CM dilated cardiomyopathy fully rescued irregular Ca transients and decreased Ca transport. Taken together, these results suggest that gene transfer of PLB M is a potential therapy for cardiac diseases associated with impaired Ca transport and muscle contractility.