Table 2.
Summary of reports modeling monogenic mutations associated with permanent neonatal diabetes mellitus (PNDM) or pancreatic agenesis.
| Gene studied | Pancreatic defects reported in humans | Effects recapitulated in mice | Genome editing approach | Differentiation protocol | Type of human pluripotent stem cell | In vitro phenotypes | Ref. |
|---|---|---|---|---|---|---|---|
| GATA6 | GATA6 heterozygous inactivating mutations result in pancreatic agenesis. | Gata6 heterozygous mice are fertile and phenotypically normal. Gata6 null mice are embryonic lethal. Biallelic loss of Gata6 and its paralog Gata4 result in a phenotype similar to human PNDM GATA6-mutated patients. | CRISPR/Cas9-mediated genome editing. | Adapted from (52). | Patient-derived hiPSCs and genome-edited hESCs. Isogenic, mutation-corrected, hiPSCs were used as controls. | GATA6 homozygous mutations lead to impaired DE differentiation. Rescue of DE defects in these cells by re-expression of other GATA family members allows β-like cell production with a lower efficiency. hPSCs with GATA6 heterozygous mutations show defects in DE differentiation. β-like cells produced in both cases are defective in the GSIS and in insulin processing. | (114) |
| CRISPR/Cas9-mediated genome editing. | (61, 62, 111), with some modifications. | Genome-edited hESCs (H1 and HUES8). | Differentiation of GATA6 -/- hPSCs revealed impaired DE commitment and pancreatic endocrine differentiation. No defects in DE differentiation from GATA6 +/- hPSCs, but a lower number of PDX1+ NKX6.1+ pancreatic progenitors and β-like cells was produced. | (115) | |||
| TALENs | (55), adapted from (52). | hiPSCs derived from pancreatic agenesis patients with GATA6 heterozygous mutations. Genome-edited hESCs (H9) and hiPSCs. Non-mutated hESCs and hiPSCs were used as isogenic controls. | GATA6 heterozygous hPSCs present a modest decrease in the generation of DE, which differentiate less efficiently into MPCs and EPs. GATA6-null hPSCs fail to enter the DE lineage. | (116) | |||
| CRISPR-CAS9-mediated genome editing. | Adapted from (61, 62, 54). | hiPSCs derived from a patient with pancreatic agenesis. Isogenic, mutation-corrected hiPSCs were used as control. | hiPSCs with GATA6 heterozygous mutations present reduced efficiency for generation of pancreatic progenitor cells in vitro. Correction of these mutations allowed identifying a non-coding SNP that additionally contributes to the phenotype observed. | (117) | |||
| PDX1 | Homozygous mutations in PDX1 result in pancreatic agenesis. PDX1 heterozygous patients exhibit diabetes caused by defects in β cell function and/or the maintenance of β cell mass in adults. | Homozygous mutations in Pdx1cause pancreatic agenesis, while heterozygous animals develop a pancreas but become diabetic in adulthood due to β cell apoptosis. | TALEN ad CRISPR/Cas9. | Adapted from (52, 54). | Genome-edited hESCs (HUES8). | Differentiation of PDX1 +/- mutant hESCs present a 65% reduction of INS+ cells at the β-like cell stage, which are mainly polyhormonal cells using the protocol described in this study. | (111) |
| RFX6 | Patients carrying biallelic RFX6 inactivating mutations present a reduction in the pancreas size and obstruction of the small intestine. These patients present defects in the formation of pancreatic progenitors and their further differentiation into functional endocrine cells. | Similar to humans, Rfx6-null mice show variable degrees of pancreatic hypoplasia and premature death. | TALEN ad CRISPR/Cas9. | Adapted from (52, 54). | Genome-edited hESCs (HUES8). | Differentiation of RFX6 -/- mutant hESCs show a reduction in the number of PDX1+ pancreatic progenitor cells. Severe reduction in β-like cells and complete absence of α cells. | (111) |
| CRISPR/Cas9-mediated genome editing. | Adapted from (62) | hiPSCs were derived from patients with MRS and from their healthy, heterozygous father. hESCs (H9) was used as control. | hiPSCs with RFX6 homozygous mutations show normal DE and PFG differentiation, but fail to robustly activate PDX1. MPCs and endocrine-competent progenitors differentiate less efficiently from these cells. | (118) | |||
| PTF1A | Homozygous inactivating mutations in PTF1A cause pancreatic and cerebellar agenesis. | Ptf1a-null mice present a complete absence of exocrine pancreatic tissue, but all islet endocrine cell types are present until the late stages of embryogenesis. | TALEN ad CRISPR/Cas9. | Adapted from (52, 54). | Genome-edited hESCs (HUES8). | Differentiation of PTF1A -/- mutant hESCs do not present defects in pancreatic endocrine differentiation using the protocol described. | (111) |
| GLIS3 | Biallelic mutations of GLIS3 underlie a rare clinical syndrome, characterized by neonatal diabetes and congenital hypothyroidism. | Global Glis3 -/- mice die of severe neonatal diabetes shortly after birth. Minor differences in gene dosage of Glis3 produce substantive changes in the expression levels of Ngn3 and Ins1, leading to a variable phenotype among the multiple Glis3-KO mouse lines. | TALEN ad CRISPR/Cas9. | Adapted from (52, 54). | Genome-edited hESCs (HUES8). | Differentiation of GLIS3 -/- mutant hESCs do not present defects in pancreatic endocrine differentiation using the protocol described. | (111) |
| CRISPR/Cas9-mediated genome editing. | (64) | Genome-edited hESCs. | Differentiation of GLIS3 -/- mutant hESCs show impaired expression of pancreatic endocrine-associated genes, including PDX1, NEUROD1, NKX6.1, and MAFA, and present increased β-like cell death. A chemical screen identified a drug candidate that rescues mutant GLIS3-associated β-cell death both in vitro and in vivo. | (64) | |||
| MNX1 | Homozygous mutations in MNX1 are associated with the occurrence of diabetes in infancy without evidence of exocrine pancreatic dysfunction. Reduced number of pancreatic endocrine cells, including β cells. | Mnx1-deficient mice show pancreatic dorsal-lobe agenesis and smaller pancreatic islets, while Mnx1 gain-of-function in the pancreas leads to aberrant pancreatic development. | TALEN ad CRISPR/Cas9. | Adapted from (52, 54). | Genome-edited hESCs (HUES8). | Differentiation of MNX1 −/− mutant hESCs do not present defects in pancreatic endocrine differentiation using the protocol described. | (111) |
DE, definitive endoderm; MPC, multipotent pancreatic progenitor cells; PFG, posterior foregut; GSIS, glucose-stimulated insulin secretion; MRS, Mitchell-Riley syndrome; KO, knock out; ER, endoplasmic reticulum.