Dual GLP-1 and GIP receptor agonist Tirzapetide plays an off-target role in the modulation of the β-adrenoceptors and glucose metabolism in hyperglycemic or senescent cardiac cells

BackgroundA dual glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-1 (GLP1) receptor agonist Tirzepatide (TZPD) is a novel cardioprotective agent, particularly in metabolic disturbances-related co-morbidities, however, there is no exact study to emphasize its possible off-target action in cardiac cells. ObjectiveTaking into consideration a relationship between the trafficking of incretin receptors in a manner not anticipated by the standard way of cAMP as a primary actor in TZPD action, together with the role of cAMP depression in cardiac dysfunction, here, we aimed to elucidate a pattern of off-target receptor interactions of TZPD and molecular processes underlying the pleiotropic effects of TZPD through modulation of the {beta}-adrenoceptors ({beta}-ARs) signaling in cardiomyocytes. MethodsTo establish the multifaceted cardioprotective function and underlying mechanisms of TZPD against hyperglycemia (HG)- or senescence (SC)-induced cardiac dysfunction, H9c2 cells were treated with and without TZPD. We also used {beta}3-ARs overexpressed H9c2 cells ({beta}3OE) for comparisons. ResultsThe TZPD intervention ameliorated the HG or SC phenotypes in the cardiac cells via alleviation in protein levels of GLP-1R and GIP-R as well as production of cAMP or cGMP even in the presence of these receptor antagonisms. The TZPD also alleviated the depressed levels of the {beta}1- and {beta}2-ARs with a significant decrease in the activated {beta}3-ARs and PKG being parallel to normalizations in the cAMP and cGMP in the presence of the antagonisms of these receptors. The therapeutic effects of TZPD on the similar parameters of the {beta}3OE group of cells can strongly verify its off-target action among multifaceted effects in either HG or SC cells. In addition, molecular dynamics simulations indicated that TZPD binds with the highest affinity to GLP-1R and {beta}3-ARs rather than GIP-R and then relatively lower but almost similar affinities to {beta}1- and {beta}2-ARs. Furthermore, mechanistically, the cardioprotective effect of TZPD includes significant regulation of the cellular Ca2+, at most, modulating the proteins in {beta}-ARs signaling pathways. Moreover, TZPD could significantly increase not only the depressed protein level but also the translocation of GLUT4 on the sarcolemma, promoting glucose uptake in the HG or SC groups independent of its receptor actions. ConclusionsOur findings indicate that TZPD, with its multifaceted role, has beneficial effects on cardiac cells by positively modulating {beta}-ARs signaling and glucose metabolism rather than on-target receptor action. Furthermore, we demonstrated how TZPD can engage the different targets with distinct signaling motifs at the sarcolemma. Highlights- TZPD has direct cardio-therapeutic effects in cardiac cells under hyperglycemia or senescence, at most, through affecting altered {beta}-ARs signaling in cardiomyocytes with the highest affinity to {beta}3-ARs compared to the others. - The multifaceted roles of TZPD in the HG or SC group of cells include modulation of {beta}- ARs signaling, cellular Ca2+ regulation, and glucose metabolism independent from the insulin signaling pathway. - TZPD could induce translocation of GLUT4 on the membrane and increase its protein level in the HG or SC group of cells independent of its receptor actions. - TZDP could also normalize the depressed level of IRS-1in the HG or SC group of cells. - TZPD activates both GLP-1R and GIP-R in cells, particularly with consideration of the in silico finding on the higher binding affinity of TZPD to GLP-1R rather than GIP-R, it seems an activation of GLP-1R by an agonist stimulates insulin secretion predominantly through the GLP-1R, with an additional contribution of GIP-R activation - Overall, these results demonstrate that the same drug engaging the different targets has distinct signaling motifs at the plasma membrane and provides further information on the role of incretins in cardiac cells under hyperglycemia or senescence.