Manipulating ß-cell zinc to tatrget regenerative compound activity
Abstract/Contents
- Abstract
- Loss of pancreatic insulin-producing β-cells is characteristic of type 1 and type 2 diabetes mellitus. The recent identification of dual-specificity tyrosine phosphorylation-regulated kinase 1A/B (DYRK1A/B) inhibitors as inducers of β-cell replication raises the possibility of therapeutically regenerating lost β-cell mass. However, before contemplating clinical translation, a strategy for β-cell-selective drug targeting is required to direct the non-selective proliferative drug activity. One potential strategy for β-cell drug targeting is to leverage the uniquely high zinc content of insulin granules to promote preferential drug accumulation and bioactivity. Indeed, conjugating a zinc-chelating moiety (DPA) to the DYRK1A/B inhibitor GNF-4877 (4877-EXT-DPA) improved the β-cell selectivity of drug accumulation and replication induction. Consequently, in Chapter 2, we hypothesized that pharmacologically increasing β-cell zinc content would further enhance the potency and β-cell-selectivity of zinc-binding compounds. Using a chemical screening platform we developed, we identified GR 46611 as a modulator of β-cell zinc content. GR 46611 enhanced β-cell insulin content, diminished unstimulated insulin release, and opposed cAMP signaling. In primary rat and human islet-cell culture, GR 46611 treatment further biased 4877-EXT-DPA-induced (but not GNF-4877) replication towards β-cells. These studies identify a chemical modulator of β-cell zinc content and demonstrate a strategy for enhancing the β-cell selectivity of zinc-chelating DYRK1A/B inhibitors. Chapter 3 discusses pathogenic INS gene mutations that are causative for Mutant INS-gene-induced Diabetes of Youth (MIDY). We characterize a previously undescribed de novo heterozygous INS gene mutation (c.289A> C, p.T97P; INS_Stanford) that presented in an autoantibody negative 5-month-old male infant with severe diabetic ketoacidosis. In silico pathogenicity prediction tools provided contradictory interpretations while our structural modeling indicated a deleterious effect on insulin folding. Transfection of wildtype or INS_Stanford expression constructs demonstrated comparable intracellular proinsulin/insulin levels, but INS_Stanford expression resulted in impaired mature mutant insulin formation and secretion. To assess the effect of INS_Stanford on proinsulin processing and release, we generated reporter constructs where C-peptide was replaced with Gaussia luciferase. Notably, INS_Stanford exhibited approximately 10-fold reduced luciferase secretion, indicating impaired post-translational processing. Notably, proteasome inhibition partially and selectively rescued INS_Stanford-derived luciferase secretion. Additionally, INS_Stanford expression increased intracellular insulin aggregate formation and XBP-1s protein levels, consistent with induction of endoplasmic reticulum stress. We conclude that INS_Stanford is a newly identified pathogenic A-chain variant that is causative for MIDY via disruption of insulin folding and/or proteolytic processing with induction of the endoplasmic reticulum stress response.
Description
| Type of resource | text |
|---|---|
| Form | electronic resource; remote; computer; online resource |
| Extent | 1 online resource. |
| Place | California |
| Place | [Stanford, California] |
| Publisher | [Stanford University] |
| Copyright date | 2021; ©2021 |
| Publication date | 2021; 2021 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Author | Moeller, Hannah Page |
|---|---|
| Degree supervisor | Annes, Justin |
| Thesis advisor | Annes, Justin |
| Thesis advisor | Chen, James Kenneth |
| Thesis advisor | Mochly-Rosen, Daria |
| Degree committee member | Chen, James Kenneth |
| Degree committee member | Mochly-Rosen, Daria |
| Associated with | Stanford University, Department of Chemical and Systems Biology |
Subjects
| Genre | Theses |
|---|---|
| Genre | Text |
Bibliographic information
| Statement of responsibility | Hannah Page Moeller (Fraser). |
|---|---|
| Note | Submitted to the Department of Chemical and Systems Biology. |
| Thesis | Thesis Ph.D. Stanford University 2021. |
| Location | https://purl.stanford.edu/pj726yt2204 |
Access conditions
- Copyright
- © 2021 by Hannah Page Moeller
- License
- This work is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported license (CC BY-NC).
Also listed in
Loading usage metrics...