O5 HTD4010: analogous peptide of INGAP-PP with greater stimulatory power on function and insular mass ß

Abstract

Introduction: INGAP-PP, a peptide derived from insular neogenesis-associated protein (INGAP), positively modulates β-cell mass and function in normal and diabetic rats. Its use in a clinical study increased the release of C-peptide in people with DM1 and improved glycemic control in people with DM2, but the study had to be interrupted due to the appearance of skin lesions at the injection site. Searching for more powerful analogues, capable of being used in lower quantities and injection frequency, HTD4010 emerged, a peptide with greater stability in plasma and potential favorable impact at the island level.

Objectives: Determine the effect of HTD4010 on insulin secretion and gene expression of factors modulating ß mass and function in rat islets.

Materials and methods: isolated islets (digestion with collagenase) from normal male Wistar rats were cultured for 4 days in RPMI, forming 5 groups: Control (C); with the addition of INGAP-PP 1µg/ml (I1) to the medium; addition of HTD4010 0.01µg/ml (H0.01); 0.1µg/ml (H0.1) or 1µg/ml (H1). After culture, the islets were incubated for one hour with 3.3 or 16.6 mM glucose to measure insulin secretion by RIA. In the islets of groups C, I1 and H0.01 we isolated total RNA to determine gene expression levels (qRT-PCR) related to insular function, insulin signaling, neogenesis and apoptosis.

Results: we did not find differences in the insulin released against 3.3 mM glucose, but against 16.6 mM glucose, the islets of groups I1, H0.01, H0.1 and H1 secreted significantly more insulin than those of C (p<0.05); Furthermore, HTD4010 produced the same effect as INGAP-PP with a dose 100 times lower (H0.01 vs. I1). In H0.01 islets the gene expression of insulin and PDX1 (islet function) increased (p<0.05),
Ngn-3 (neogenesis), from RI, IRS1 and 2, PI3K and Akt (insulin signaling), from Bcl2 (anti-apoptotic) and Bad (pro-apoptotic). In parallel, caspases 8, 9 and 3 (apoptosis) decreased (p<0.05) at H0.01. The Bcl2/Bad ratio showed a 30% greater antiapoptotic effect in H0.01 compared to I1 islets.

Conclusions: HTD4010 is 100 times more potent than INGAP-PP in its potentiating effect on insulin secretion, an effect that is associated with increased expression of genes involved in the function and modulation of ß cell mass. Consequently, HTD4010 could be a good candidate to be used in new clinical trials in people with DM2.