Identification of molecules which control in-vitro progenitor to endocrine cell differentiation of hESCs-derived pancreatic cells
Type 1 diabetes is a chronic disease, caused by the loss of insulin-producing β-cells which reside in the pancreatic islets of Langerhans. In-vitro differentiation of human embryonic stem cells into β-cells could serve as a transplantation source, but the high cost and low yield of current protocols make this promising therapy unaffordable for most patients. Although endocrine cells rarely divide, in-vitro differentiation includes an intermediate progenitor cell-state which, if maintained, could serve to expand the differentiating-cells’ number, and reduce costs.
To identify factors which control the progenitor-endocrine balance, we performed a high throughput screen using RNA-sequencing Of Selected Amplicons (ROSA-HTS) which allows targeted sequencing of hundreds of genes from thousands of samples. Of 1280 molecules, we performed full RNA-sequencing on 89 “hits”, and discovered novel effector molecules that shift progenitor-to-endocrine balance. Among these, we discovered acetylcholine-receptor effectors (Nicotinic agonist and Muscarinic antagonist) which induced progenitor gene-expression, whereas PI3K, AKT or mTOR inhibition induced an early endocrine state. In addition, we discovered novel molecular pathways which were not directly targeted by the screen, but whose mediated regulation uncovers a potential influence on pancreatic differentiation. For instance, Interferon-I inhibition seems to induce endocrine state, whereas Interleukins 2 and 18 activation seems to induce progenitor state.
Achieving the optimal combination of molecules which effect the progenitor-to endocrine balance is expected to have a synergic effect, and thus increase yield of existing protocols for β-cell differentiation.
