SBIR-STTR Award

Therapeutic peptides are used to treat human diseases ranging from HIV to diabetes and have some of the best features of small molecule and recombinant protein drugs. Therapeutic peptides account for $13 billion of annual pharmaceutical sales and are part of a growing sector of the biopharmaceutical market. Unfortunately, therapeutic peptides typically suffer from poor stability and short half-lives in the human body, which limits their value. The requirement for high dosing and frequent injections follows, which can be inconvenient, expensive, and dangerous for patients. While there have been methods developed to address these issues, they either: (i) hinge on in vitro or recombinant attachment of a large polymer chain, which dramatically impacts peptide activity or (ii) require in vitro processing steps which increase manufacturing costs and complicate purification. It is now well-established that the stability and half-life of peptide drugs can be greatly improved by conjugation to oligosaccharides that are nonimmunogenic in the human body. Several therapeutic peptides (e.g., Exenatide, Glucagon-like peptide 1) have benefitted significantly from glycosylation with small, human-like glycans by increasing protease resistance, prolonging activity, and improving biodistribution. However, this requires multiple complicated in vitro reactions and purifications which have kept this promising concept from reaching the industrial scale. Glycobia has developed a transformative solution to this growing, unsolved problem by engineering bacteria as a platform for the biosynthesis of therapeutic glycopeptides. These novel strains of Escherichia coli are useful for the expression of recombinant peptides conjugated to nonimmunogenic, human-like oligosaccharides. The hypothesis to be tested here is that non-pathogenic, glycoengineered strains of E. coli can produce affordable recombinant peptide drugs with improved stability, biocompatibility, and prolonged half-life in serum. The main objective of this Phase I project is to identify and characterize peptide drug candidates for animal and preclinical studies in Phase II of this project. This will be accomplished through the following specific aims: (1) express a panel of therapeutic peptide glycoconjugates and screen for expression and glycosylation efficiency; and (2) screen glycoconjugate drug candidates for biophysical properties and in vitro activity. The panel of peptides to be studied here includes seven FDA- approved therapeutic peptides that will be evaluated for expression in the glycoengineered E. coli system. Of these seven peptides, five are currently produced using recombinant expression systems and account for over $2.6 billion in annual sales. Each of the targets will be assayed for solubility, stability, and in vitro activity. This information will be considered along with the commercial potential of each peptide, leading to the identification of candidate peptides for Phase II of this project.

Public Health Relevance Statement:


Public Health Relevance:
Therapeutic peptides account for $13 billion of pharmaceutical sales annually and are used to treat human diseases ranging from HIV to diabetes. Many approved and emerging peptide drugs suffer from poor stability and have short half-lives in the human body, which can be problematic and dangerous for patients. The goal of this work is to produce stable, soluble therapeutic peptides in glycoengineered Escherichia coli.

NIH Spending Category:
Biotechnology

Project Terms:
ABO blood group system; Accounting; Address; Anabolism; Animal Model; Animals; Bacteria; bacterial H antigen; base; Biodistribution; Biological Assay; Biological Availability; Biological Products; biomaterial compatibility; bivalirudin; Blood groups; Blood Transfusion; Blood typing procedure; candidate identification; Cells; cost; cost effective; Diabetes Mellitus; Dose; drug candidate; Engineering; Escherichia coli; Evolution; exenatide; FDA approved; glucagon-like peptide; Glycoconjugates; Glycopeptides; Glycoproteins; glycosylation; Goals; Half-Life; HIV; Hormones; Human; Human body; human disease; Immune; improved; In Vitro; in vitro activity; inhibitor/antagonist; Injection of therapeutic agent; Insulin; Laboratories; Lead; Light; Link; Manufacturer Name; Marketing; Methods; Modification; novel; Oligosaccharides; Patients; Peptide Hydrolases; Peptides; Pharmaceutical Preparations; Pharmacologic Substance; Phase; phase 2 study; Polymers; Polysaccharides; Positioning Attribute; pre-clinical; preclinical study; Preclinical Testing; Process; Production; Property; Proteins; public health relevance; Reaction; recombinant peptide; Recombinant Proteins; Recombinants; Resistance; Sales; Serum; small molecule; Solubility; Solutions; sugar; System; Technology; Teriparatide; Testing; Therapeutic; Thrombin; Viral Tumor Antigens; Work

Therapeutic peptides are used to treat human diseases ranging from HIV to diabetes and have some of the best features of small molecule and recombinant protein drugs. Therapeutic peptides account for $13 billion of annual pharmaceutical sales and are part of a growing sector of the biopharmaceutical market. Unfortunately, therapeutic peptides suffer from poor stability and short half-lives in the human body, which limits their value. The requirement for high dosing and frequent injections can be inconvenient, expensive, and dangerous for patients. While there have been methods developed to address these issues, they either: (i) hinge on the in vitro or recombinant attachment of a large polymer chain, which dramatically impacts peptide activity or (ii) require in vitro processing steps which increase manufacturing costs and complicate purification. It is now well- established that the stability and half-life of peptide drugs can be greatly improved by conjugation to humanlike oligosaccharides. Several therapeutic peptides (e.g., Exenatide, Glucagon-like peptide 1) have benefitted significantly from glycosylation with small human-like glycans by increasing protease resistance, prolonging activity, and improving biodistribution. However, this requires multiple complicated in vitro reactions and purifications which have kept this promising concept from reaching the industrial scale. At Glycobia we have developed novel strains of Escherichia coli for the expression of recombinant peptides conjugated to humanlike oligosaccharides. In Phase I of this project, we applied our glycoengineered bacteria as a platform for the biosynthesis of therapeutic glycopeptides by: (1) expressing a panel of therapeutic peptide glycoconjugates and screening for glycosylation efficiency and (2) screening glycoconjugate drug candidates for physical properties and in vitro activity. We show proof-of-concept of several recombinant peptides with improved stability and/or activity when modified with glycosylation. Now having identified lead candidates for Phase II of this project, the objective of this proposal is to synthesize and advance our first drug targets from glycoengineered E. coli into preclinical testing by: (1) expressing, scaling up, purifying, and characterizing a glycosylated human peptide drug from in E. coli and (2) testing pharmacology, calcemic response and pharmacokinetics of a glycosylated human peptide drug in animal models. We will attach two different humanlike glycans to the drug and compare performance to an aglycosylated version of the drug. The benchmark of success for this project is the generation of positive preclinical validation data to further advance commercialization of this glycoengineering technology. Our bacterial expression platform represents a transformative solution to the unanswered biomedical challenge of producing improved therapeutic peptides for patients.

Public Health Relevance Statement:


Public Health Relevance:
Therapeutic peptides account for over $13 billion of pharmaceutical sales and are part of a rapidly growing market with at least 600 peptide drug candidates in clinical testing or development. Many approved and emerging peptide drugs suffer from poor stability and have short half-lives in the human body, which can be problematic and dangerous for patients. The proposed studies focus on - for the first time - producing stable, longer-lasting, glycosylated therapeutic peptides in the simple bacterium Escherichia coli and advancing them into preclinical testing.

Project Terms:
ABO blood group system; Accounting; Address; Anabolic Agents; Anabolism; analog; Animal Model; Animals; Bacteria; bacterial H antigen; Benchmarking; Biochemical Reaction; Biodistribution; Biological; Biological Products; Bioreactors; Blood groups; Blood Transfusion; Blood typing procedure; Chemicals; Clinical; Collaborations; commercialization; cost; cost effective; Data; Development; Diabetes Mellitus; Dose; drug candidate; Drug Kinetics; Drug Targeting; Engineering; Escherichia coli; exenatide; FDA approved; Frequencies (time pattern); Generations; Glucagon; glucagon-like peptide; Glycoconjugates; Glycopeptides; glycosylation; Goals; Half-Life; HIV; Hormones; Human; Human body; human disease; human PTH protein; Hypercalcemia; Hypoparathyroidism; Immune; improved; In Vitro; in vitro activity; Industry; Injection of therapeutic agent; Lead; Marketing; Methods; microorganism; mouse model; novel; Oligosaccharides; Osteoporosis; Parathyroid gland; pathogenic Escherichia coli; Patients; Peptide Hydrolases; Peptides; Performance; Pharmaceutical Preparations; Pharmacologic Substance; Pharmacology; Phase; physical property; Polymers; Polysaccharides; Positioning Attribute; pre-clinical; Preclinical Testing; Process; Production; Property; Proteins; public health relevance; Reaction; recombinant peptide; Recombinant Proteins; Recombinants; Relative (related person); research clinical testing; Resistance; response; Sales; Sampling; scale up; screening; Serum; small molecule; Solubility; Solutions; Subcutaneous Injections; success; sugar; Synthesis Chemistry; Technology; Teriparatide; Testing; Therapeutic; Time; Treatment Efficacy; Validation