Soluble hyaluronidase glycoprotein (sHASEGP), process for preparing the same, uses and pharmaceutical compositions comprising thereof

• US 8,202,517 B2
• Filed: 02/20/2009
• Issued: 06/19/2012
• Est. Priority Date: 03/05/2003
• Status: Active Grant

• Current Assignee: Halozyme, Inc. (Halozyme Therapeutics, Inc.)
• Original Assignee: Halozyme, Inc. (Halozyme Therapeutics, Inc.)
• Inventors: Bookbinder, Louis H., Kundu, Anirban, Frost, Gregory I.
• Primary Examiner(s): Kosson, Rosanne
• Application Number:
  US12/378,969
• Publication Number:
  US 20090181013A1
• Time in Patent Office: 1,215 Days
• Field of Search: 424/94.62, 435/201, 435/69.1, 530/350
• US Class Current: 424/94.62
• CPC Class Codes: A61K 2039/505 comprising antibodies
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: A61K 38/47 acting on glycosyl compounds (3.2), e.g. cellulases, lactases
• CPC Class Codes: A61K 39/395 Antibodies agglutinins A61K38/36 ; as drug carriers A61K47/50; Immunoglobulins; Immune serum, e.g. antilymphocytic serum
• CPC Class Codes: A61K 9/0048 Eye, e.g. artificial tears
• CPC Class Codes: A61P 17/00 Drugs for dermatological disorders
• CPC Class Codes: A61P 17/02 for treating wounds, ulcers, burns, scars, keloids, or the like
• CPC Class Codes: A61P 19/00 Drugs for skeletal disorders
• CPC Class Codes: A61P 19/02 for joint disorders, e.g. arthritis, arthrosis
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 25/16 Anti-Parkinson drugs
• CPC Class Codes: A61P 25/24 Antidepressants
• CPC Class Codes: A61P 27/02 Ophthalmic agents
• CPC Class Codes: A61P 29/00 Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• CPC Class Codes: A61P 31/00 Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
• CPC Class Codes: A61P 31/04 Antibacterial agents
• CPC Class Codes: A61P 31/10 Antimycotics
• CPC Class Codes: A61P 31/12 Antivirals
• CPC Class Codes: A61P 33/00 Antiparasitic agents
• CPC Class Codes: A61P 33/04 Amoebicides
• CPC Class Codes: A61P 33/06 : Antimalarials
• CPC Class Codes: A61P 35/00 : Antineoplastic agents
• CPC Class Codes: A61P 43/00 : Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 7/04 : Antihaemorrhagics; Procoagulants; Haemostatic agents; Antifibrinolytic agents
• CPC Class Codes: A61P 7/10 : Antioedematous agents; Diuretics
• CPC Class Codes: A61P 9/00 : Drugs for disorders of the cardiovascular system
• CPC Class Codes: A61P 9/04 : Inotropic agents, i.e. stimulants of cardiac contraction; Drugs for heart failure
• CPC Class Codes: A61P 9/10 : for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
• CPC Class Codes: A61P 9/12 : Antihypertensives
• CPC Class Codes: C12N 9/2474 : Hyaluronoglucosaminidase (3.2.1.35), i.e. hyaluronidase
• CPC Class Codes: C12Y 302/01035 : Hyaluronoglucosaminidase (3.2.1.35), i.e. hyaluronidase
• CPC Class Codes: Y02A 50/30 : Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Abstract

The invention relates to the discovery of novel soluble neutral active Hyaluronidase Glycoproteins (sHASEGP'"'"'s), methods of manufacture, and their use to facilitate administration of other molecules or to alleviate glycosaminoglycan associated pathologies. Minimally active polypeptide domains of the soluble, neutral active sHASEGP domains are described that include asparagine-linked sugar moieties required for a functional neutral active hyaluronidase domain. Included are modified amino-terminal leader peptides that enhance secretion of sHASEGP. The invention further comprises sialated and pegylated forms of a recombinant sHASEGP to enhance stability and serum pharmacokinetics over naturally occurring slaughterhouse enzymes. Further described are suitable formulations of a substantially purified recombinant sHASEGP glycoprotein derived from a eukaryotic cell that generate the proper glycosylation required for its optimal activity.

Claims

1. A method for delivering a therapeutically effective drug or pharmaceutical agent to a subject, comprising:

a) administering a hyaluronidase glycoprotein to a tissue in the subject in an amount sufficient to degrade glycosaminoglycans, wherein;

the hyaluronidase glycoprotein consists of the sequence of amino acids set forth as amino acids 36-477, 36-478, 36-479, 36-480, 36-481, 36-482, or 36-483 of SEQ ID NO;

1, or contains amino acid substitutions in the sequence of amino acids set forth as amino acids 36-477, 36-478, 36-479, 36-480, 36-481, 36-482 or 36-483 of SEQ ID NO;

1, whereby the amino-acid substituted hyaluronidase glycoprotein consists of a sequence of amino acids that has at least 95% amino acid sequence identity with the sequence of amino acids set forth as amino acids 36-477, 36-478, 36-479, 36-480, 36-481, 36-482 or 36-483 of SEQ ID NO;

1;

the glycoprotein is neutral active; and

the glycoprotein contains at least one sugar moiety that is covalently attached to an asparagine (N) residue of the polypeptide; and

b) administering a therapeutically effective amount of the drug or pharmaceutical agent to the subject.

2. The method of claim 1, wherein the drug or pharmaceutical agent is selected from among a small molecule and a protein.

3. The method of claim 1, wherein the drug or pharmaceutical agent is selected from among a chemotherapeutic agent, an analgesic agent, an anti-inflammatory agent, an antimicrobial agent, an amoebicidal agent, a trichomonocidal agent, an anti-Parkinson agent, an anti-malarial agent, an anticonvulsant agent, an anti-depressant agent, an anti-arthritic agent, an anti-fungal agent, an antihypertensive agent, an antipyretic agent, an anti-parasite agent, an antihistamine agent, an alpha-adrenergic agonist agent, an alpha blocker agent, an anesthetic agent, a bronchial dilator agent, a biocide agent, a bactericide agent, a bacteriostat agent, a beta adrenergic blocker agent, a calcium channel blocker agent, a cardiovascular drug agent, a contraceptive agent, a decongestant agent, a diuretic agent, a depressant agent, a diagnostic agent, an electrolyte agent, a hypnotic agent, a hormone agent, a hyperglycemic agent, a muscle relaxant agent, a muscle contractant agent, an ophthalmic agent, a parasympathomimetic agent, a sedative agent, a sympathomimetic agent, a tranquilizer agent, a viricide agent, a vitamin agent, a non-steroidal anti-inflammatory agent, an angiotensin converting enzyme inhibitor agent, a cytokine, an antibody, antibody fragment, an insulin and a sleep inducer.

4. The method of claim 3, wherein the chemotherapeutic agent is a toxin or a tumor necrosis factor.

5. The method of claim 3, wherein the anesthetic agent is lidocaine or bupivacaine.

6. The method of claim 1, wherein the drug or pharmaceutical agent is an insulin.

7. The method of claim 1, wherein the drug or pharmaceutical agent is a cytokine.

8. The method of claim 1, wherein the drug or pharmaceutical agent is an antibody or antibody fragment.

9. The method of claim 8, wherein the antibody or antibody fragment is a monoclonal antibody.

10. The method of claim 1, further comprising administering a hormonal agent.

11. The method of claim 10, wherein the hormonal agent is epinephrine.

12. The method of claim 10, wherein the hormonal agent is administered simultaneously or sequentially with the hyaluronidase glycoprotein.

13. The method of claim 1, wherein the hyaluronidase glycoprotein consists of a sequence of amino acids set forth as amino acids 36-477, 36-478, 36-479, 36-480, 36-481, 36-482, or 36-483 of SEQ ID NO:

1.

14. The method of claim 1, wherein a composition comprising the hyaluronidase glycoprotein that consists of the sequence of amino acids set forth as amino acids 36-482 of SEQ ID NO:

1 is administered.

15. The method of claim 1, wherein the hyaluronidase glycoprotein consists of the sequence of amino acids that contains amino acid substitutions in the sequence of amino acids set forth as amino acids 36-477, 36-478, 36-479, 36-480, 36-481, 36-482, or 36-483 of SEQ ID NO:

1, whereby the amino-acid substituted hyaluronidase glycoprotein consists of a sequence of amino acids that has at least 95% sequence identity with the sequence of amino acids set forth as amino acids36-477, 36-478, 36-479, 36-480, 36-481, 36-482, or 36-483 of SEQ ID NO;

1.

16. The method of claim 1, wherein the hyaluronidase glycoprotein is produced in and secreted from a mammalian cell by a method comprising:

introducing an expression vector comprising a polynucleotide that consists of a sequence of nucleotides set forth as nucleotides 106-1446 of SEQ ID NO;

6 or degenerates thereof or the sequence of nucleotides set forth in SEQ ID NO;

48 or degenerates thereof inserted between flanking sequences in the vector into a mammalian cell;

culturing the mammalian cell under conditions whereby the glycoprotein is expressed and secreted by the cell; and

recovering a composition comprising the expressed and secreted hyaluronidase glycoprotein.

17. The method of claim 16, wherein the polynucleotide has the sequence of nucleotides set forth as SEQ ID NO:

48.

18. The method of claim 16, wherein the mammalian cell is a CHO cell.

19. The method of claim 1, wherein the hyaluronidase glycoprotein is modified with a polymer.

20. The method of claim 19, wherein the polymer is a polyethylene glycol (PEG) or a dextran.

21. The method of claim 1, wherein the hyaluronidase glycoprotein is administered in an amount that is or is between 1 and 10,000 Units/ml.

22. The method of claim 1, wherein the hyaluronidase glycoprotein and molecule are provided in the same composition or as separate compositions.

23. The method of claim 1, wherein the hyaluronidase glycoprotein is administered prior to, simultaneously with or following administration of the molecule.

24. The method of claim 1, wherein the hyaluronidase glycoprotein is administered at a site that is different from the site of administration of the molecule.

25. The method of claim 1, wherein the hyaluronidase glycoprotein is administered at a site that is the same as the site of administration of the molecule.

26. The method of claim 1, wherein the hyaluronidase glycoprotein and/or drug or pharmaceutical agent is administered by parenteral administration.

27. The method of claim 26, wherein parenteral administration is selected from among subcutaneous, intravenous and intramuscular administration.

28. The method of claim 1, wherein the drug or pharmaceutical agent is a therapeutic agent.