(#21) Establishment of pro-islet amyloid polypeptide as a biomarker in T1D

PRESENTED BY: Yi-Chun Chen

Authors
First NameLast NameAffiliation/Institution
Yi-ChunChen1Department of Surgery, University of British Columbia and BC Children’s Hospital Research Institute
CateSpeakeCenter for Interventional Immunology, Benaroya Research Institute at Virginia Mason
PaolaApaolaza GallegosInstitute of Diabetes Research, Helmholtz Zentrum Muenchen, German Research Center for Environmental Health
TeresaRodriguez-CalvoInstitute of Diabetes Research, Helmholtz Zentrum Muenchen, German Research Center for Environmental Health
PreethiKrishnanIndiana University School of Medicine, Department of Pediatrics, Pediatric Endocrinology and Diabetology, Wells Center for Pediatric Research
FarooqSyedIndiana University School of Medicine, Department of Pediatrics, Pediatric Endocrinology and Diabetology, Wells Center for Pediatric Research
EmilySimsIndiana University School of Medicine, Department of Pediatrics, Pediatric Endocrinology and Diabetology, Wells Center for Pediatric Research
CarmellaEvans-MolinaIndiana University School of Medicine, Department of Pediatrics, Pediatric Endocrinology and Diabetology, Wells Center for Pediatric Research
C. BruceVerchereDepartments of Surgery and Pathology & Laboratory Medicine, University of British Columbia, Centre for Molecular Medicine and Therapeutics, BC Children’s Hospital Research Institute
 

Purpose

Synthesis and secretion of beta cell peptide hormones closely reflect beta cell health, and failure in hormone biosynthesis and processing may result in disproportional secretion of prohormones in diabetes. Indeed, the plasma ratio of proinsulin-to-C-peptide is elevated in type 1 diabetes (T1D) and pre-T1D subjects. Parallel to insulin, islet amyloid polypeptide (IAPP) is first synthesized in beta cells as a 67-amino acid precursor molecule, proIAPP1-67. Through sequential endopeptidase cleavage and amidation, proIAPP is processed into an intermediate form (proIAPP1-48), and eventually becomes mature IAPP (IAPP1-37). Utilizing a sensitive ELISA developed in-house, we previously showed that the ratio of proIAPP1-48 to total IAPP is elevated in plasma from T1D subjects and islet transplant recipients. We aimed to confirm this finding in an additional T1D cohort, to validate the performance of our proIAPP1-48 and IAPP1-37 ELISAs through the JDRF Core for Assay Validation (CAV), and to analyze the relationship between proinsulin and proIAPP levels in T1D plasma and islets from T1D organ donors.
 

Methods

To evaluate the reproducibility of our in-house developed ELISAs, we measured proIAPP1-48 and IAPP1-37 levels in blinded plasma samples from T1D subjects from JDRF CAV, and calculated the percent coefficient of variation (%CV) of triplicate aliquots. To establish the proIAPP1-48-to-total IAPP ratio as a biomarker in T1D, we measured proIAPP1-48 and IAPP1-37 levels in plasma from the Indiana University cohort of new-onset T1D and age-matched subjects, and performed regression analysis on the proinsulin:C-peptide and proIAPP1-48:IAPP ratios. We also examined the expression patterns of proIAPP and proinsulin by performing immunostaining in pancreas sections from T1D and BMI- and age- matched donors.
 

Summary of Results

Both proIAPP1-48 and IAPP1-37 ELISAs displayed good reproducibility (%CV = 16.28 and 12.61, respectively) in intra-assay tests, whereas the proIAPP1-48:IAPP ratio displayed excellent reproducibility (%CV = 3.73). The proIAPP1-48:IAPP ratio was significantly elevated in plasma from new-onset T1D subjects (0.37±0.02 vs. 0.22±0.02; p < 0.0001) in the Indiana University cohort, and ratios of proIAPP1-48:IAPP and proinsulin:C-peptide showed a significant linear correlation (r = 0.445; p < 0.05). Immunostaining of nPOD tissue indicated that proinsulin and proIAPP are co-expressed in beta cells of pancreatic sections from T1D donors.
 

Conclusions

We validated our ELISA for proIAPP1-48 and IAPP1-37 in human plasma and confirmed the proIAPP1-48:IAPP ratio as a biomarker of beta cell dysfunction in T1D. The co-localization of proIAPP and proinsulin in T1D beta cells, and correlation of proinsulin:C-peptide and proIAPP1-48:IAPP ratios in T1D plasma, suggest that the same residual beta cells in T1D likely secrete both proinsulin and proIAPP and are responsible for persistent prohormone secretion in T1D.