Figure 2. Schematic of PEGylated insulin formulations.

To assess the effect of host–guest binding affinity on the pharmacokinetics of dynamically PEGylated insulin, two non-covalently PEGylated insulin systems were tested. (A) The N-terminal B1 phenylalanine of insulin can bind to cucurbit[7]uril (CB[7]) with a K_eq~10⁶ M⁻¹. A PEG_20k-CB[7] can thus be used to non-covalently PEGylate the phenylalanine on insulin. (B) To test host–guest motifs with higher binding affinity, a covalent insulin-CB[7] conjugate was combined with a guest-linked PEG_20k. To depict the conjugation between CB[7] and insulin, the chemical structures of CB[7], bicyclo[6.1.0]nonyne (BCN), and the PEG linker used are shown (left). PEG_20k-xylylenediamine (K_eq~10⁹ M⁻¹) or PEG_20k-O-adamantane (K_eq~10¹⁰ M⁻¹) were selected as guests. Bond lifetime (τ), is inversely related to the dissociation rate k_off and was calculated based on the following relationships and assumptions: K_eq = k_on/k_off; k_on~10⁷; τ = 1/k_off. See Table S1 for a complete list of values.