Functional reconstitution of adrenocortical transdifferentiation from human pluripotent stem cells

Proper prenatal adrenal cortex development is essential to prevent life-threatening adrenal insufficiency. This process relies on self-renewing progenitor cells within a capsule (Cap)-derived niche of the definitive zone (DZ) and trans-differentiation into cortisol-producing transitional and androgen-producing fetal zones. However, species-specific limitations in rodent models hinder mechanistic insights. To address this, we developed a human iPSC-derived organoid system that mimics prenatal adrenal development. Our advanced platform efficiently generates DZ-like cells (DZLCs) and Cap-like cells (CapLCs), revealing that DZLCs arise from adrenal primordium-like cells (APLCs) and are sustained via CapLC-derived RSPO3/WNT signaling. These DZLCs trans-differentiate into transitional and fetal zone-like cells (FZLCs) through ACTH and RSPO3 signaling. Notably, NR0B1 loss impairs DZLC specification while driving direct APLC-to-FZLC conversion, recapitulating NR0B1 mutation-associated X-linked adrenal hypoplasia congenita. Notably, DZLCs encapsulated with CapLCs form ACTH-responsive, cortisol-producing cortical tissue in vivo, demonstrating therapeutic potential. These organoid models offer a powerful platform for adrenal research and lay the groundwork for cell-based therapies for adrenal insufficiency.