A novel insulin analog with unique properties: [Lys.sup.B3], [Glu.sup.B29] insulin induces prominent activation of insulin receptor substrate 2, but marginal phosphorylation of insulin receptor substrate 1

The potentially enhanced mitogenic activity of insulin analogs represents a safety risk that requires detailed analysis of new analogs considered for therapeutic applications. We assessed the signaling properties and mitogenic potency of two novel rapid-acting insulin analogs, [Lys.sup.B3],[Glu.sup.B29] insulin (HMR 1964) and [Lys.sup.B3],[Ile.sup.B28] insulin (HMR 1153) using myoblasts and cardiomyocytes. In myoblasts, both binding and internalization were two- to threefold higher for [Asp.sup.B10] insulin and HMR 1153 when compared with HMR 1964 and regular insulin. This finding correlated with a prominent Shc/ IGF-I receptor interaction, tyrosine phosphorylation of She, activation of extracellular signal-regulated protein kinase (ERK)-I and -2, and stimulation of DNA synthesis by HMR 1153 and [Asp.sup.B10] insulin. In contrast, HMR 1964 produced a marginal activation of the Shc/ERK kinase cascade and was equipotent to insulin in stimulating DNA synthesis in myoblasts. Further, the in vivo growth-promoting activity of this analog was round to be identical to that of regular human insulin. In myoblasts, HMR 1964 produced a miner activation of insulin receptor substrate (IRS)-I tyrosine phosphorylation, but a prominent activation of IRS-2, with a significantly stronger effect than insulin in human myoblasts. Predominant activation of IRS-2 was also observed in adult cardiomyocytes where HMR 1964 increased 3-O-methyl-glucose transport and the activation of Akt and glycogen synthase kinase-3 to the same extent as human insulin. We concluded that 1) the mitogenic properties of insulin analogs may result from a series of initial receptor interactions, including internalization and phosphorylation; 2) the mitogenic and metabolic potential of HMR 1964 is identical to that of insulin; and 3) predominant activation of IRS-2 may open new avenues for optimized insulin therapies.