Diabetes mellitus is an escalating global health problem. Both Type 1 and Type 2 diabetes lead to the gradual loss of insulin-producing beta cells. Our long-term goal is to develop beta cell replacement strategies to overcome the insulin deficiency in diabetic patients. To achieve this goal, we have developed novel, safer approaches to convert patient fibroblasts to induced pluripotent stem cells (iPSCs) using chemicals together with genetic factors. These iPSCs can be differentiated into multipotent pancreatic progenitors, the precursors to beta cells, using protocols developed for human embryonic stem cells (hESCs). However, our understanding of the basic biology of pancreatic progenitor cells is still rudimentary, which presents hurdles in the development of effective strategies to expand and further differentiate these progenitor cells for therapy. Evidence emerged from our own studies and from those of other laboratories suggests that pancreatic progenitor cells are heterogeneous, and likely consist of subpopulations with different physiological functions. Yet, molecular and functional distinctions between these subpopulations are poorly understood. It is unclear which subpopulation(s) in the developing pancreas is(are) capable of generating functional beta cells, and to which extent a specific subpopulation is capable of self-renewal. Even less is known about the physiological functions and self-renewal properties of pancreatic progenitor cells differentiated from hESCs.
We aim to address these critical questions in this proposal. We hypothesize that the pancreatic progenitor cell compartment in vivo contains heterogeneous subpopulations: (i) a common multipotent pancreatic progenitor responsible for generating all three pancreatic lineages; and (ii) distinct lineage-specific progenitors responsible for generating one or two specific pancreatic lineages. Based on this hypothesis, the overall objective of this proposal is to define pancreatic progenitor subpopulations in terms of their molecular characteristics and physiological functions, as well as to identify mechanisms for self-renewal. We anticipate that our study will yield the following outcomes.
Aim 1 will reveal the presence of distinct pancreatic progenitor subpopulations both during embryo development and in hESC differentiation culture. This will identify the progenitor subpopulation(s) capable of generating functional cells.
Aim 2 will identify cell surface markers for enrichment of the appropriate progenitor subpopulation(s) for further ? differentiation into ? cells, and set the stage for studies on expansion of progenitor cells.
Aim 3 will elucidate the mechanism of self-renewal of distinct progenitor subpopulations, which are critical for the development of novel, effective strategies to expand pancreatic progenitors for beta cell replacement therapy. Additionally, the ability to generate a large quantity of human pancreatic progenitor cells will provide a new way to study the biology of these cells to complement mouse genetics approaches. Broadly, the proposed research will lead to novel findings to fill in critical gaps in our current knowledge of human pancreatic development.

Public Health Relevance

Generating functional pancreatic beta cells from human pluripotent stem cells offers a promising cure for diabetes, a growing global health problem resulting from the loss of beta cells. However, a major challenge in fulfilling such a promise lies in the rudimentary understanding of the distinct types of pancreatic progenitor cells, which are the precursors that generate mature, insulin-secreting beta cells during embryo development. The goal of this research is to use combined stem cell and developmental biology approaches to understand pancreatic progenitor subtypes in terms of their molecular characteristics, physiological functions, and self-renewal properties, which will form the foundation for developing rational approaches to effective beta cell replacement therapy.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK096239-04
Application #
8853275
Study Section
Cellular Aspects of Diabetes and Obesity Study Section (CADO)
Program Officer
Sato, Sheryl M
Project Start
2012-07-01
Project End
2016-06-30
Budget Start
2015-07-01
Budget End
2016-06-30
Support Year
4
Fiscal Year
2015
Total Cost
Indirect Cost
Name
Sloan-Kettering Institute for Cancer Research
Department
Type
DUNS #
064931884
City
New York
State
NY
Country
United States
Zip Code
10065
Verma, Nipun; Pan, Heng; Doré, Louis C et al. (2018) TET proteins safeguard bivalent promoters from de novo methylation in human embryonic stem cells. Nat Genet 50:83-95
Teijeiro, Virginia; Yang, Dapeng; Majumdar, Sonali et al. (2018) DICER1 Is Essential for Self-Renewal of Human Embryonic Stem Cells. Stem Cell Reports 11:616-625
Shukla, Abhijit; Huangfu, Danwei (2018) Decoding the noncoding genome via large-scale CRISPR screens. Curr Opin Genet Dev 52:70-76
Amin, Sadaf; Cook, Brandoch; Zhou, Ting et al. (2018) Discovery of a drug candidate for GLIS3-associated diabetes. Nat Commun 9:2681
Pulecio, Julian; Verma, Nipun; Mejía-Ramírez, Eva et al. (2017) CRISPR/Cas9-Based Engineering of the Epigenome. Cell Stem Cell 21:431-447
Verma, Nipun; Zhu, Zengrong; Huangfu, Danwei (2017) CRISPR/Cas-Mediated Knockin in Human Pluripotent Stem Cells. Methods Mol Biol 1513:119-140
Soh, Chew-Li; Huangfu, Danwei (2017) CRISPR/Cas9-Mediated Mutagenesis of Human Pluripotent Stem Cells in Defined Xeno-Free E8 Medium. Methods Mol Biol 1498:57-78
Shi, Zhong-Dong; Lee, Kihyun; Yang, Dapeng et al. (2017) Genome Editing in hPSCs Reveals GATA6 Haploinsufficiency and a Genetic Interaction with GATA4 in Human Pancreatic Development. Cell Stem Cell 20:675-688.e6
Wang, Qiong; Zou, Yilong; Nowotschin, Sonja et al. (2017) The p53 Family Coordinates Wnt and Nodal Inputs in Mesendodermal Differentiation of Embryonic Stem Cells. Cell Stem Cell 20:70-86
Shi, Zhong-Dong; Soh, Chew-Li; Zhu, Zengrong et al. (2017) Genome Editing and Directed Differentiation of hPSCs for Interrogating Lineage Determinants in Human Pancreatic Development. J Vis Exp :

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