Variability in the effect of subcutaneously administered insulin represents a major challenge in insulin therapy where precise dosing is required in order to achieve targeted glucose levels. Since this variability is largely influenced by the absorption of insulin, a deeper understanding of the factors affecting the absorption of insulin from the subcutaneous tissue is necessary in order to improve glycaemic control and the long-term prognosis in people with diabetes. These factors can be related to either the insulin preparation, the injection site/patient, or the injection technique. This review highlights the factors affecting insulin absorption with special attention on the physiological factors at the injection site. In addition, it also provides a detailed description of the insulin absorption process and the various modifications to this process that have been utilized by the different insulin preparations available.

Here, we describe the molecular engineering of insulin icodec to achieve a plasma half-life of 196 h in humans, suitable for once-weekly subcutaneously administration. Insulin icodec is based on re-engineering of the ultra-long oral basal insulin OI338 with a plasma half-life of 70 h in humans. This systematic re-engineering was accomplished by (1) further increasing the albumin binding by changing the fatty diacid from a 1,18-octadecanedioic acid (C18) to a 1,20-icosanedioic acid (C20) and (2) further reducing the insulin receptor affinity by the B16Tyr → His substitution. Insulin icodec was selected by screening for long intravenous plasma half-life in dogs while ensuring glucose-lowering potency following subcutaneous administration in rats. The ensuing structure–activity relationship resulted in insulin icodec. In phase-2 clinical trial, once-weekly insulin icodec provided safe and efficacious glycemic control comparable to once-daily insulin glargine in type 2 diabetes patients. The structure–activity relationship study leading to insulin icodec is presented here.

Recently, the clinical proof of concept for the first ultra-long oral insulin was reported, showing efficacy and safety similar to subcutaneously administered insulin glargine. Here, we report the molecular engineering as well as biological and pharmacological properties of these insulin analogues. Molecules were designed to have ultra-long pharmacokinetic profile to minimize variability in plasma exposure. Elimination plasma half-life of ~20 h in dogs and ~70 h in man is achieved by a strong albumin binding, and by lowering the insulin receptor affinity 500-fold to slow down receptor mediated clearance. These insulin analogues still stimulate efficient glucose disposal in rats, pigs and dogs during constant intravenous infusion and euglycemic clamp conditions. The albumin binding facilitates initial high plasma exposure with a concomitant delay in distribution to peripheral tissues. This slow appearance in the periphery mediates an early transient hepato-centric insulin action and blunts hypoglycaemia in dogs in response to overdosing.

FSH is a key component in assisted reproductive technologies. Because of rapid clearance of the hormone, patients have to be treated with daily injections. To address this problem, a long-acting FSH mutein was created by introduction of additional N-linked glycosylation into the molecule. New glycosylation sites were introduced by two different approaches: structure-aided, site-directed introduction of sites within the FSH molecule and addition of N-terminal extensions. A mutein with the extension sequence ANITVNITV at the N terminus of the alpha-chain (FSH1208) was efficiently glycosylated at both new sites. This resulted in a molecule with increased size and charge, factors known to reduce renal clearance of proteins. FSH1208 was found to have a 3- to 4-fold increased serum half-life, compared with wild-type recombinant FSH. Furthermore, in spite of a lower in vitro activity, FSH1208 had a markedly increased in vivo potency, as shown by increased ability to augment the ovarian weight and stimulate the serum estradiol levels in rats. These characteristics make FSH1208 a possible candidate for improved infertility treatment.

Recently, the first basal oral insulin (OI338) was shown to provide similar treatment outcomes to insulin glargine in a phase 2a clinical trial. Here, we report the engineering of a novel class of basal oral insulin analogues of which OI338, 10, in this publication, was successfully tested in the phase 2a clinical trial. We found that the introduction of two insulin substitutions, A14E and B25H, was needed to provide increased stability toward proteolysis. Ultralong pharmacokinetic profiles were obtained by attaching an albumin-binding side chain derived from octadecanedioic (C18) or icosanedioic acid (C20) to the lysine in position B29. Crucial for obtaining the ultralong PK profile was also a significant reduction of insulin receptor affinity. Oral bioavailability in dogs indicated that C18-based analogues were superior to C20-based analogues. These studies led to the identification of the two clinical candidates OI338 and OI320 (10 and 24, respectively).