Discovery of retinoic acid receptor agonists as proliferators of cardiac progenitor cells through a phenotypic screening approach

Abstract

Identification of small molecules with the potential to selectively proliferate cardiac progenitor cells (CPCs) will aid our understanding of the signaling pathways and mechanisms involved and could ultimately provide tools for regenerative therapies for the treatment of post-MI cardiac dysfunction. We have used an in vitro human induced pluripotent stem cellderived CPC model to screen a 10,000-compound library containing molecules representing different target classes and compounds reported to modulate the phenotype of stem or primary cells. The primary readout of this phenotypic screen was proliferation as measured by nuclear count. We identified retinoic acid receptor (RAR) agonists as potent proliferators of CPCs. The CPCs retained their progenitor phenotype following proliferation and the identified RAR agonists did not proliferate human cardiac fibroblasts, the major cell type in the heart. In addition, the RAR agonists were able to proliferate an independent source of CPCs, HuES6. The RAR agonists had a time-of-differentiation-dependent effect on the HuES6-derived CPCs. At 4 days of differentiation, treatment with retinoic acid induced differentiation of the CPCs to atrial cells. However, after 5 days of differentiation treatment with RAR agonists led to an inhibition of terminal differentiation to cardiomyocytes and enhanced the proliferation of the cells. RAR agonists, at least transiently, enhance the proliferation of human CPCs, at the expense of terminal cardiac differentiation. How this mechanism translates in vivo to activate endogenous CPCs and whether enhancing proliferation of these rare progenitor cells is sufficient to enhance cardiac repair remains to be investigated.

Phenotypic Screen with the Human Secretome Identifies FGF16 as Inducing Proliferation of iPSC-Derived Cardiac Progenitor Cells

“…In contrast, FGF9 had a more general proliferative effect on cardiac progenitor cells, cardiac fibroblasts, and kidney fibroblasts (Figure 2). We previously used human iPS-derived CPCs in phenotypic high-content screening assays in our team to identify small-molecule compounds that induce proliferation [19,23]. In the presented work, we focused on screening a human secretome library in a proliferation assay with hCPCs.…”

Cardiac Stem Cell Therapy, Resident Progenitor Cells and the role of Cellular Signalling; a Review

“…All-Trans Retinoic Acid (ATRA) and AM580 were 2 RAR agonists identified and shown to increase proliferation of induced pluripotent stem cell derived -CPCs. Additionally, the ATRA compounds did not increase proliferation of terminal cardiac cell types [19]. Whether ATRA and AM580 hold therapeutic value in future treatments remains to be investigated.…”

“…A recent study suggests that retinoic acid (RA), as well as being crucial for guiding correct development of the embryo via intercellular signalling, may also play a role in cardiac repair. Retinoic acid receptor (RAR) agonists were shown to be potent proliferators of CPCs [19]. Importantly, new CPCs formed maintained their progenitor phenotype.…”

Bioactive Lipid Signaling in Cardiovascular Disease, Development, and Regeneration

“…More recent reports have expanded upon these findings. Drowley et al [ 297 ] utilized a hiPSC-derived cardiac progenitor cell (CPC) assay, described previously [ 298 ], to screen 10,000 different compounds for their proliferative effects. They found that two agonists of the retinoic acid receptor (RAR)—ATRA (all-trans retinoic acid) and AM580—increased the number of CPC nuclei without altering their inherent progenitor cell identity, an effect that was attenuated by RAR antagonists.…”

“…They found that two agonists of the retinoic acid receptor (RAR)—ATRA (all-trans retinoic acid) and AM580—increased the number of CPC nuclei without altering their inherent progenitor cell identity, an effect that was attenuated by RAR antagonists. When applied to CPCs on day 5 of the cardiac differentiation protocol, RA inhibited terminal cardiomyocyte differentiation and activated human CPC proliferation instead, demonstrating the stage-specificity of this effect [ 297 ]. Another recent study utilized an inhibitor of retinoic acid synthesis in developing mouse embryos to show that a deficiency in RA signaling delayed the migration of embryonic epicardial cells into the myocardium, with the opposite effect being seen under conditions of excess RA production.…”