br Materials and methods br Results br

Materials and methods
Results
Discussion Employment of GIPR signaling, specifically the direction of this manipulation, as a strategy to improve body weight remains a contentious subject of intense scientific investigation [22], [32], [33]. Despite the physiological benefits of GIP on Levodopa control, the initial direction in drug design was directed towards attenuating GIP action with peptide antagonists for the treatment of patients with T2D and obesity [34], and that assumption currently continues in scientific research [35]. However, it has yet to be shown that an elevated level of circulating GIP is a pathophysiological driver of T2D. Furthermore, inhibiting an endogenous insulinotropic response seems counterintuitive for pharmacotherapies aimed to improve glucose control. Despite these incongruences, it is important to note that the studies comprising this report were not designed to address the question of which direction to manipulate GIPR signaling for glycemic benefit in diabetic rodent models, but rather to specifically address the question of which direction to manipulate GIPR signaling for body weight benefit in non-diabetic, genetically wild-type obese mice. The proposition for GIPR antagonists as opposed to GIPR agonists for the treatment of obesity largely originates from the report that congenital global Gipr−/− mice are protected from diet-induced and genetic-induced obesity [10]. Importantly and seemingly forgotten is the comparable phenotype displayed by congenital Glp1r−/− mice [36]. Furthermore, the [Pro3] GIP analog was shown to decrease body weight in obese mice [37], and at the time of its original report, the body weight lowering effect was believed to be the product of GIPR antagonism. However, this peptide has since been thoroughly tested and is now re-characterized as a weak partial agonist at mGIPR [31]. Thus, those historical results on body weight should be interpreted with caution but support mGIPR agonism in lowering body weight. Therefore, to directly address this dichotomy as to which direction to manipulate the GIP system for therapeutic benefits in obesity, a series of precise chemical reagents were developed and studied for their pharmacological effects in obese rodents. There are appreciable opportunities to improve the pharmacological profile of GIP by chemical optimization. As such, the GIP analogs reported herein were specifically advanced to address the question of how GIPR agonism affects body weight in DIO mice with chronic dosing. Through a series of diagnostic studies measuring the acute effects on basal glucose levels in wild-type, Gipr−/−, and Glp1r−/− mice, we demonstrate that these GIP analogs are selective for mouse GIPR and suitable for studies to specifically address the effects of GIPR mono-agonism on body weight in obese mice. Chronic administration to DIO mice caused meaningful body weight loss in numerous DIO mouse studies using several structurally different analogs, but relatively high doses were required. This latter notion is important as chronic, twice daily administration of D-Ala2 GIP analogs to DIO mice did not result in significant body weight change [26], likely due to using subthreshold doses, which notably were lower than doses used here with the Aib2 GIP analogs. Our results also indicate that the human and mouse sequences demonstrate differential potency at mGIPR, but not hGIPR. This largely recapitulates previous results [31], and the pivotal amino acid change influencing potency among the three differences between human and mouse GIP is His18Arg. Protection against DDP-IV cleavage by incorporation of Aib2 also increased in vitro and in vivo potency. Noteworthy is our discovery that Aib2 enhances in vitro potency at mGIPR when incorporated into the human GIP sequence. One potential explanation for the lack of body weight lowering effects of other reported GIPR agonists [21], [25] could be due to this species divergence and subsequent reduced potency at mGIPR as that pharmacological aspect was not previously studied in those reports.