SP6616 as a new Kv2.1 channel inhibitor efficiently promotes β-cell survival involving both PKC/Erk1/2 and CaM/PI3K/Akt signaling pathways

Abstract

Kv2.1 as a voltage-gated potassium (Kv) channel subunit has a pivotal role in the regulation of glucose-stimulated insulin secretion (GSIS) and pancreatic β-cell apoptosis, and is believed to be a promising target for anti-diabetic drug discovery, although the mechanism underlying the Kv2.1-mediated β-cell apoptosis is obscure. Here, the small molecular compound, ethyl 5-(3-ethoxy-4-methoxyphenyl)-2-(4-hydroxy-3-methoxybenzylidene)-7-methyl-3-oxo-2,3-dihydro-5H-[1,3]thiazolo[3,2–a]pyrimidine-6-carboxylate (SP6616) was discovered to be a new Kv2.1 inhibitor. It was effective in both promoting GSIS and protecting β cells from apoptosis. Evaluation of SP6616 on either high-fat diet combined with streptozocin-induced type 2 diabetic mice or db/db mice further verified its efficacy in the amelioration of β-cell dysfunction and glucose homeostasis. SP6616 treatment efficiently increased serum insulin level, restored β-cell mass, decreased fasting blood glucose and glycated hemoglobin levels, and improved oral glucose tolerance. Mechanism study indicated that the promotion of SP6616 on β-cell survival was tightly linked to its regulation against both protein kinases C (PKC)/extracellular-regulated protein kinases 1/2 (Erk1/2) and calmodulin(CaM)/phosphatidylinositol 3-kinase(PI3K)/serine/threonine-specific protein kinase (Akt) signaling pathways. To our knowledge, this may be the first report on the underlying pathway responsible for the Kv2.1-mediated β-cell protection. In addition, our study has also highlighted the potential of SP6616 in the treatment of type 2 diabetes.

Calmodulin and PI3K Signaling in KRAS Cancers

“…Via CaM, increase of intracellular free Ca 2+ can activate PI3K/Akt, localizing Akt in the plasma membrane [34–36]. CaM also modulates epidermal growth factor receptor (EGFR)’s tyrosine kinase activity [37] which activates Ras and PI3Kα and has essential roles in programmed cell death and autophagy [28].…”

ETA as a novel Kv2.1 inhibitor ameliorates β‐cell dysfunction and hyperglycaemia

“…According to a previous report, the platform based on membrane potential (FLIPR Membrane Potential Assay Kit) by FlexStation II 384 was applied to screen Kv2.1 inhibitor candidates against the lab compound library. As shown in Figure B, Stromatoxin‐1 (ScTx‐1,100 nmol/L), a known Kv2.1 inhibitor, obviously inhibited the membrane potential in overexpressed Kv2.1 cells (CHO‐Kv2.1), suggesting the credibility for the screening of Kv2.1 inhibitor candidates.…”

“…Thus, the effects of ETA on GSIS and β‐cell survival were investigated in INS‐832/13 cells. In the GSIS assay, 16.8 mmol/L of glucose was used to stimulate insulin secretion, and ScTx‐1 (100 nmol/L) was used as a positive control . As shown in Figure A, ETA (10, 20 and 40 μmol/L) activated insulin secretion in response to glucose stimulation.…”

“…Next, we investigated the effect of ETA against β‐cell viability by 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide (MTT) assay. In the assay, STZ (0.4 mmol/L) was used to stimulate cell apoptosis, and SP6616 (10 μmol/L) was used as a positive control . As shown in Figure B, ETA (10, 20 and 40 μmol/L) had no effect on β‐cell viability in INS‐832/13 cells.…”

“…Kv2.1 overexpression potentiates apoptosis in INS‐1 β‐cells, and Kv2.1 inhibition or knockdown abrogates the pro‐apoptotic effects in neuron cells . In addition, Kv2.1 inhibitors including vindoline and SP6616 are beneficial to the promotion of GSIS and β‐cell survival …”