Site-specific insulin-polymer conjugates
AssigneeAntriaBio, Inc.
InventorsKenneth Hinds, Danny Lewis, Paul Schmidt, Kathleen M. Campbell
Life of this patent
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Abstract
The invention is directed to a single-step method for rapidly and efficiently preparing protein-polymer conjugates, including an insulin-polymer conjugate. According to the method of the present invention, a protein and hydrophilic polymer are contacted in the presence of at least one organic solvent and at least one metal chelator, under conditions that promote the formation of a conjugate of the protein and polymer. Thus, the invention is directed to the site-specific modification of selected proteins, such as insulin, with poly(ethylene glycol) at residue PheB1. The invention also provides a pharmaceutical formulation for encapsulating the conjugate in a biodegradable polymer.
Claims18
An insulin-PEG conjugate comprising PEG linked to insulin at the PheB1 amino terminus via a secondary amine, wherein the conjugate comprises an unreacted GlyA1 amino terminus and an unreacted LysB29 amino terminus, wherein the insulin comprises human insulin having the amino acid sequence of SEQ ID NO:1, and wherein the insulin retains insulin activity.
A poly(lactide-co-glycolide) (PLGA) microsphere encapsulating an insulin-PEG conjugate, wherein the insulin-PEG conjugate comprises PEG linked to insulin at the PheBl amino terminus via a secondary amine, wherein the insulin comprises human insulin having the amino acid sequence of SEQ ID NO:1, and wherein the insulin retains insulin activity.
An insulin-PEG conjugate produced according to a method comprising:
- contacting insulin with PEG in the presence of at least one organic solvent and at least one metal chelator, under conditions that promote the formation of a conjugate of the insulin and PEG, wherein the formation of the insulin-PEG conjugate is by a predominant reaction of insulin at residue PheBl without protecting insulin at GlyAl and LysB29 residues;
- and
- isolating the conjugate, wherein the insulin comprises human insulin having the amino acid sequence of SEQ ID NO:1, and wherein the insulin retains insulin activity.
A F5000 insulin-PEG conjugate comprising PEG linked to insulin at the PheBl amino terminus via a secondary amine, wherein the insulin comprises human insulin having the amino acid sequence of SEQ ID NO:
- 1, and wherein the insulin retains insulin activity.
A composition for preparing a pharmaceutical formulation, the composition comprising:
- an organic solvent;
- a metal chelator;
- a first mono-PEGylated insulin conjugate comprising PEG linked to insulin at the PheBl amino terminus via a secondary amine;
- a second mono-PEGylated insulin conjugate comprising PEG linked to insulin at either the GlyAl amino terminus or the LysB29 amino terminus;
- an unreacted insulin protein;
- and
- a di-PEGylated insulin conjugate, wherein:
- the first mono-PEGylated insulin conjugate comprises 95% or greater of the total of the first mono-PEGylated insulin conjugate and the second mono-PEGylated insulin conjugate, wherein the insulin comprises human insulin having the amino acid sequence of SEQ ID NO:
- 1, and wherein the insulin retains insulin activity.
Dependencies are inferred from the claim text.