Soluble glycosaminoglycanases and methods of preparing and using soluble glycosaminoglycanases

• US 20050260186A1
• Filed: 02/23/2005
• Published: 11/24/2005
• Est. Priority Date: 03/05/2003
• Status: Active Grant

• Current Assignee: Halozyme, Inc. (Halozyme Therapeutics, Inc.)
• Original Assignee: Halozyme, Inc. (Halozyme Therapeutics, Inc.)
• Inventors: Frost, Gregory I., Kundu, Anirban, Bookbinder, Louis H., Keller, Gilbert A., Haller, Michael F., Dylan, Tyler M.
• Granted Patent: US 7,871,607 B2
• Time in Patent Office: Days
• US Class Current: 424/94.610
• CPC Class Codes: A61K 2039/505 comprising antibodies
• CPC Class Codes: A61K 2300/00 Mixtures or combinations of active ingredients, wherein at least one active ingredient is fully defined in groups A61K31/00 - A61K41/00
• 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 39/3955 against proteinaceous materials, e.g. enzymes, hormones, lymphokines
• CPC Class Codes: A61K 45/06 Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 27/12 for cataracts
• 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 3/00 Drugs for disorders of the metabolism of the blood or the extracellular fluid A61P7/00
• CPC Class Codes: A61P 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: A61P 31/00 Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
• CPC Class Codes: A61P 31/06 for tuberculosis
• CPC Class Codes: A61P 31/08 for leprosy
• CPC Class Codes: A61P 31/12 Antivirals
• CPC Class Codes: A61P 33/02 Antiprotozoals, e.g. for leishmaniasis, trichomoniasis, toxoplasmosis
• CPC Class Codes: A61P 33/04 Amoebicides
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 7/02 Antithrombotic agents; Anticoagulants; Platelet aggregation inhibitors
• 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: C12N 9/2408 : acting on alpha -1,4-glucosidic bonds
• CPC Class Codes: C12Y 302/01035 : Hyaluronoglucosaminidase (3.2.1.35), i.e. hyaluronidase

Abstract

The invention relates to the discovery of novel soluble neutral active Hyaluronidase Glycoproteins (sHASEGPs), 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 substantially purified glycoprotein, comprising a neutral active soluble hyaluronidase polypeptide and at least one N-linked sugar moiety, wherein the N-linked sugar moiety is covalently attached to an asparagine residue of the polypeptide.

2. The polypeptide of claim 1, wherein the polypeptide is selected from of:

(a) a polypeptide that comprises a sequence of amino acids encoded by the sequence that includes at least about 74% amino acid sequence identity with the sequence of amino acids set forth in SEQ ID NO. 1;

(b) a polypeptide that comprises a sequence of amino acids encoded by the sequence of nucleotides set forth in SEQ ID NO. 6;

(c) a polypeptide that comprises a sequence of amino acids encoded by a sequence of nucleotides that hybridizes along at least 85% of its full-length under conditions of high stringency to the sequence of nucleotides set forth in SEQ ID NO. 6;

or

(d) a polypeptide that is encoded by a sequence of nucleotides that is a splice variant of the sequence of nucleotides that comprises the sequence set forth in SEQ ID NO. 50.

3. The polypeptide of claim 1, wherein said sugar moiety is covalently attached to an asparagine residue selected from amino acids 82, 166, 235, 254, 368, 393 or 490 as set forth in SEQ ID NO. 1.

4. The polypeptide of claim 1, wherein said sugar moiety is covalently linked to said polypeptide through a PNGase sensitive bond.

5. The polypeptide of claim 1, wherein said sugar moiety is of the high mannose type.

6. The polypeptide of claim 1, wherein said sugar moiety is of the complex type.

7. The polypeptide of claim 1, wherein said sugar moiety is a hybrid type.

8. The polypeptide of claim 1, wherein said sugar moiety further comprises at least one terminated with sialic acid.

9. The polypeptide of claim 1, wherein the polypeptide consists essentially of the hyaluronidase domain of human hyaluronidase glycoprotein (sHASEGP) or a catalytically active portion thereof.

10. The polypeptide of claim 1, wherein the hyaluronidase portion of the polypeptide comprises of the hyaluronidase domain of sHASEGP or a catalytically active portion thereof.

11. The polypeptide of claim 1, wherein the polypeptide is modified with a polymer.

12. The polypeptide of claim 11, wherein the polymer is PEG or dextran.

20. The polypeptide of claim 1, comprising greater than about 80% sequence identity with a polypeptide that comprises the sequence of amino acids set forth in SEQ ID NO. 3, wherein the polypeptide is a hyaluronidase.

21. The polypeptide of claim 9, wherein the hyaluronidase domain portion is encoded by a polynucleotide that hybridizes under conditions of high stringency along at least 70% of its full-length to a nucleic acid molecule comprising a sequence of nucleotides set forth in SEQ ID NO. 6 or at least one domain thereof or a catalytically active portion of the domain.

22. The polypeptide of claim 1, wherein the sHASEGP is a human polypeptide.

23. The polypeptide of claim 1, wherein the polypeptide is a soluble polypeptide as set forth in SEQ ID NO. 4.

24. The polypeptide of claim 1, with the proviso that the polypeptide does not comprise the complete sequence set forth in SEQ ID NO. 1 and includes at least amino acids of 1-429 of SEQ ID NO. 4.

25. The polypeptide of claim 1 that is a mutein, wherein up to about 50% of the amino acids are replaced with another amino acid;

and the resulting polypeptide has catalytic activity of at least 10% of the unmutated polypeptide.

26. The polypeptide of claim 25, wherein up to about 10% of the amino acids are replaced with another amino acid.

27. The polypeptide of claim 25, wherein the resulting polypeptide has catalytic activity of at least 50% of the unmutated polypeptide.

28. The polypeptide of claim 25, wherein a free cysteine in the hyaluronidase domain is replaced with another amino acid.

30. The polypeptide of claim 1, wherein a signal sequence capable of directing the polypeptide outside the cell is operably attached to the sHASEGP polypeptide.

38. A nucleic acid molecule, encoding the polypeptide of claim 1.

41. A vector comprising the nucleic acid molecule of claim 38.

42. The vector of claim 41 that is an expression vector.

43. The vector of claim 41 that is a eukaryotic vector.

44. The vector of any of claim 41 that includes a sequence of nucleotides that directs secretion of any polypeptide encoded by a sequence of nucleotides operatively linked thereto.

47. An isolated cell, comprising the vector of claim 41.

48. The cell of claim 47 that is a prokaryotic cell.

49. The cell of claim 47 that is a eukaryotic cell.

50. The cell of claim 47 selected from a bacterial cell, a yeast cell, a plant cell, an insect cell or an animal cell.

51. The cell of claim 47 that is a mammalian cell.

52. An antibody that binds to the sHASEGP of claim 1, wherein said antibody is free of reactivity to bovine, ovine or bacterial hyaluronidase.

53. A transgenic non-human animal, wherein an endogenous gene that encodes a polypeptide of claim 1 has been disrupted by homologous recombination or insertional mutagenesis of the animal or an ancestor thereof.

55. A method for producing a soluble polypeptide that contains sHASEGP, comprising:

introducing a nucleic acid that encodes the polypeptide of claim 1 operably linked to a suitable promoter into a cell capable of incorporating N-linked sugar moieties into sHASEGP;

culturing the cell under conditions whereby the encoded polypeptide is expressed by the cell; and

recovering the expressed polypeptide.

57. The method of claim 55, wherein the eukaryotic cell is selected from a mammalian cell, an insect cell, a yeast cell or a plant cell.

58. The method of claim 57, wherein the cell is a CHO cell.

59. A method for producing a soluble polypeptide that contains sHASEGP, comprising:

culturing the cell of claim 47 under conditions whereby the encoded polypeptide is expressed by the cell; and

recovering the expressed polypeptide.

60. A method of ex vivo gene therapy, comprising:

introducing, in vitro, a nucleic acid encoding the polypeptide of claim 1 and operably linked to a suitable promoter into a cell capable of incorporating N-linked sugar moieties into sHASEGP;

thereby generating a genetically modified cell containing the nucleic acid; and

administering the cell comprising the nucleic acid to the subject, whereby the nucleic acid encoding the sHASEGP is transferred to the subject.

61. The method of claim 60, wherein the cell is autologous to the subject.

62. The method of claim 60, wherein the cell is haplotype matched to the subject.

63. A method of preparing a mammalian oocyte for in vitro fertilization said method comprising:

contacting an oocyte with a sHASEGP of claim 1 in amount effective for removing the cumulus matrix.

64. The method of claim 62, wherein the cumulus is effectively removed without manipulation of the oocyte with a narrow bore pipette.

65. A method for generating a sHASEGP polypeptide comprising:

contacting the polypeptide of claim 1 with glycosyltransferase enzymes capable of introducing said N-linked sugar moieties into polypeptide, thereby generating sHASEGP.

66. The method of claim 65 wherein the glycosyltransferase enzymes are derived from canine microsomal membranes.

68. A composition, comprising the polypeptide of claim 1 and a suitable pharmaceutical carrier.

69. A method for treating a subject having an excess of sHASEGP substrate, comprising:

administering an amount of sHASEGP polypeptide of claim 1 sufficient to remove said sHASEGP substrate.

70. The method of claim 69, wherein said excess substrate is produced from a scar tissue.

71. The method of claim 70, wherein said scar tissue is a glial scar resulting from spinal cord injury.

72. The method of claim 70, wherein said scar tissue is a result of surgery.

73. The method of claim 70, wherein said scar tissue is a keloid scar.

74. The method of claim 69, wherein said substrate is associated with a herniated disk.

80. A method of treating a cardiovascular disorder comprising:

administration of a sHASEGP polypeptide of claim 1 to a subject in an amount sufficient to remove excess glycosaminoglycans.

81. The method of claim 80, wherein said cardiovascular disorder is selected from a myocardial infarct, congestive heart failure or arteriosclerosis.

82. A method of treating a tumor comprising:

administration of a sHASEGP polypeptide of claim 1 to a subject in an amount sufficient to remove excess glycosaminoglycans.

83. A method of delivering a molecule to a tissue containing excess amounts of glycosaminoglycan, comprising:

administering a sHASEGP polypeptide of claim 1 to a tissue in an amount sufficient to degrade glycosaminoglycans sufficiently to open channels less than about 500 nm in diameter; and

administering a molecule to the tissue comprising the degraded glycosaminoglycans.

84. The method of claim 83, wherein the polypeptide is modified with a polymer.

85. The method of claim 84, wherein the polymer is PEG or dextran.

86. The method of claim 83, wherein the sHASEGP polypeptide is administered prior to, simultaneously with or following administration of the molecule.

87. The method of claim 83, wherein the sHASEGP is administered at a site different from the site of administration of the molecule.

88. The method of claim 83, wherein the sHASEGP is administered at a site the same as the site of administration of the molecule.

89. A pharmaceutical composition, comprising, substantially purified soluble hyaluronidase glycoprotein (sHASEGP) polypeptide of claim 1 and a pharmaceutical carrier.

90. A pharmaceutical composition, comprising the substantially purified sHASEGP polypeptide of claim 1.

91. The pharmaceutical composition of claim 89, further comprising a pharmaceutically active or pharmacologic agent.

92. The pharmaceutical composition of claim 91, wherein the pharmaceutically active or pharmacologic agent is selected from the group of agents consisting of 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 antiarthritics agent, an anti-fungal agent, an antihypertensive agent, an antipyretic agent, an anti-parasitic agent, an antihistamine agent, an alpha-adrenargic agonist agent, an alpha blocker agent, an anesthetic agent, a bronchial dilator agent, a biocide agent, a bactericide agent, a bacteriostatic 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 psychic energizer agent, a sedative agent, a sleep inducer, a sympathomimetic agent, a tranquilizer agent, a urinary agent, a vaginal agent, a viricide agent, a vitamin agent, a non-steroidal anti-inflammatory agent, an angiotensin converting enzyme inhibitor agent, a polypeptide, a protein, a nucleic acid, a drug, or an organic molecule.

93. The pharmaceutical composition of claim 92, wherein the chemotherapeutic agent is a toxin or a tumor necrosis factor.

94. The pharmaceutical composition of claim 92, wherein the anesthetic agent is lignocaine or bupivicaine.

95. The pharmaceutical composition of claim 92, further comprising a hormonal agent.

96. The pharmaceutical composition of claim 95, wherein the hormonal agent is epinephrine.

97. The pharmaceutical composition of claim 89, wherein the pharmaceutical carrier comprises a stabilizing solution.

98. The pharmaceutical composition of claim 97, wherein the stabilizing solution comprises, NaCl and a metal selected from CaCl₂ and MgCl₂.

99. The pharmaceutical composition of claim 97, wherein the stabilizing solution further comprises ethylenediamine tetra-acetic acid (EDTA).

100. The pharmaceutical composition of claim 97, wherein the stabilizing solution further comprises a carrier selected from the group comprising albumin, detergent or a surfactant.

101. The pharmaceutical composition of claim 97, wherein the stabilizing solution comprises phenol red, human serum albumin, HEPES, NaCl and a metal selected from CaCl₂ and MgCl₂.

102. The pharmaceutical composition of claim 101, wherein the concentration of the glycoprotein is about 80 Units/ml.

103. The pharmaceutical composition of claim 89, further comprising a surfactant.

104. The pharmaceutical composition of claim 103, wherein the surfactant is selected from sorbitan monolaurate;

sorbitan monooleate;

sorbitan palmitate;

sorbitan monostearate;

sorbitan sesquitolate;

sorbitan trioleate;

polyoxyethylene oleic acid ester derivatives;

polyoxyethylene lauryl amine derivatives;

polyoxyethylene stearyl amine derivatives;

polyoxyethylene oleyl amine derivatives;

polyoxyethylene castor oil derivatives;

polyoxyethylene hydrogenated castor oil derivatives;

polyoxyethylene bis phenol ether derivatives;

polyoxyethylene glycols;

sorbitan fatty acid ester derivatives;

polyoxyethylene sorbitan fatty acid ester derivatives;

polyoxyethylene-polyoxypropylene derivatives;

polyethylene glycol hexadecyl ether (Brij™

     56);

polyethylene glycol octadecyl ether (Brij™

72);

polyoxyethylene 10 oleyl ether (Brij™

     97);

t-Octylphenoxypolyethoxyethanol (TRITON™

X

     100);

polyethylene glycol sorbitan monolaurate (TWEEN

     20);

polyoxyethylenesorbitan monopalmitate (TWEEN

     40);

polyethylene glycol sorbitan monostearate (TWEEN™

     60);

polyoxyethylenesorbitan Tristearate (TWEEN™

     65);

polyethylene glycol sorbitan monooleate (TWEEN

     80);

polyoxyethylenesorbitan Trioleate (TWEEN

     85);

tris(hydroxymethyl) aminomethane lauryl sulfate (TRIZMA™

dodecyl sulfate);

block copolymer of polyethylene and polypropylene glycol (Pluronic™

F68);

or albumin.

105. The pharmaceutical composition of claim 89, wherein the concentration of the glycoprotein is between about 1 Unit/mg and 5000 Units/mg.

106. The pharmaceutical composition of claim 103, wherein the concentration of the glycoprotein is between about 1 Unit/mg and 300 Units/mg.

107. The pharmaceutical composition of claim 103, wherein the concentration of the glycoprotein is between about 1 Unit/mg and 150 Units/mg.

108. The pharmaceutical composition of claim 89, further comprising a metal ion selected from calcium and magnesium.

109. The pharmaceutical composition of claim 89 that is formulated as a timed release formulation or that is released upon contact with the site of application or targeted tissue.

110. The pharmaceutical composition of claim 89 that is formulated for topical, local, enteric, parenteral, intracystal, intracutaneous, intravitreal, subcutaneous, intramuscular, or intravenous administration.

111. The pharmaceutical composition of claim 89 that is formulated as spray, foam or aerosol.

112. The pharmaceutical composition of claim 89 that is formulated as a tablet or capsule.

113. The pharmaceutical composition of claim 108, further comprising an enteric coating.

114. The pharmaceutical composition of claim 108, wherein the coating is selected from cellulose acetate phthalate, polyethylene glycol, polyoxyethylene sorbitan, castor oil, ethyl cellulose pseudolatex, phenyl salicylate, n-butyl stearate, stearic acid and carnauba wax.

115. The pharmaceutical composition of claim 81 that is a lyophilized powder.

116. The pharmaceutical composition of claim 110, wherein the lyophilized powder is produced by a process, comprising:

(a) transferring the pharmaceutical composition into a buffer solution;

(b) sterile-filtering the resulting solution; and

(c) lyophilizing the filtered solution under standard conditions to produce a sterile powder.

117. The pharmaceutical composition of claim 116, wherein the buffer solution is selected from HEPES, Phosphate, Citrate and Bicarbonate.

118. The pharmaceutical composition of claim 117, wherein the buffer solution further comprises a sugar or carbohydrate.

119. The pharmaceutical composition of claim 118, wherein the sugar or carbohydrate is dextrose or lactose.

120. The pharmaceutical composition of claim 119, wherein the lactose is present at a concentration of between about 1 mg/ml to about 15 mg/ml.

121. The pharmaceutical composition of claim 116, wherein the buffer further comprises a surfactant.

122. The pharmaceutical composition of claim 116, wherein the surfactant is selected from sorbitan monolaurate;

sorbitan monooleate;

sorbitan palmitate;

sorbitan monostearate;

sorbitan sesquitolate;

sorbitan trioleate;

polyoxyethylene oleic acid ester derivatives;

polyoxyethylene lauryl amine derivatives;

polyoxyethylene stearyl amine derivatives;

polyoxyethylene oleyl amine derivatives;

polyoxyethylene castor oil derivatives;

polyoxyethylene hydrogenated castor oil derivatives;

polyoxyethylene bis phenol ether derivatives;

polyoxyethylene glycols;

sorbitan fatty acid ester derivatives;

polyoxyethylene sorbitan fatty acid ester derivatives;

polyoxyethylene-polyoxypropylene derivatives;

polyethylene glycol hexadecyl ether (Brij™

     56);

polyethylene glycol octadecyl ether (Brij™

72);

polyoxyethylene 10 oleyl ether (Brij™

     97);

t-Octylphenoxypolyethoxyethanol (TRITON™

X

     100);

polyethylene glycol sorbitan monolaurate (TWEEN™

     20);

polyoxyethylenesorbitan monopalmitate (TWEEN™

     40);

polyethylene glycol sorbitan monostearate (TWEEN™

     60);

polyoxyethylenesorbitan Tristearate (TWEEN™

     65);

polyethylene glycol sorbitan monooleate (TWEEN™

     80);

polyoxyethylenesorbitan Trioleate (TWEEN™

     85);

tris(hydroxymethyl) aminomethane lauryl sulfate (TRIZMA™

dodecyl sulfate);

block copolymer of polyethylene and polypropylene glycol (Pluronic™

F68);

or albumin.

123. A combination comprising,

a sterile vial and the pharmaceutical composition of claim 89 wherein the composition is contained in the vial.

124. The combination of claim 123, wherein the sterile vial contains an amount of the pharmaceutical composition that is for single dose administration.

125. A combination comprising,

a sterile vial containing the pharmaceutical composition of claim 110.

126. The combination of claim 123, wherein the sterile vial further contains an amount of sterile water for injection;

and the final concentration of the glycoprotein is between about 1 and 5000 Units/ml.

127. A kit comprising the combination of claim 123 and at least one of:

(a) a packaging material;

or

(b) instructions for using the kit for use of the pharmaceutical composition.

128. The kit of claim 127, wherein the packaging material includes ice, dry ice, styrofoam, foam, plastic, cellophane, shrink wrap, bubble wrap, paper, cardboard, starch peanuts, twist ties, metal clips, metal cans, drierite, glass and rubber.

129. A combination comprising,

a sterile syringe containing the pharmaceutical composition of claim 89.

130. The combination of claim 129, wherein the sterile syringe has a volume of between about 5 to 50 μ

l.

131. The combination of claim 129, wherein the final concentration of the glycoprotein is between about 1 and about 5000 Units/ml.

132. A combination comprising, a sterile syringe containing the pharmaceutical composition of claim 110.

133. The combination of claim 132, wherein the final concentration of the glycoprotein is between about 1 and about 5000 Units/ml.

134. The combination of claim 132, wherein the pharmaceutical composition is dissolved in about 5111 to about 50 μ

l of sterile water.

135. The combination of claim 132, further comprising a second syringe containing a pharmaceutically effective agent.

136. The combination of claim 132, wherein the pharmaceutically effective or pharmacologic agent is selected from 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 antiarthritics agent, an anti-fungal agent, an antihypertensive agent, an antipyretic agent, an anti-parasitic agent, an antihistamine agent, an alpha-adrenargic agonist agent, an alpha blocker agent, an anesthetic agent, a bronchial dilator agent, a biocide agent, a bactericide agent, a bacteriostatic 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 ophthalnic agent, a parasympathomimetic agent, a psychic energizer agent, a sedative agent, a sleep inducer, a sympathomimetic agent, a tranquilizer agent, a urinary agent, a vaginal agent, a viricide agent, a vitamin agent, a non-steroidal anti-inflammatory agent, or an angiotensin converting enzyme inhibitor agent, a polypeptide, a protein, a nucleic acid, a drug, or an organic molecule.

137. The combination of claim 135, wherein the second syringe comprises a viscoelastic.

138. A kit comprising the combination of claim 132 and one or more of the following:

(a) packaging material; and

(b) instructions for using the kit for the use of the pharmaceutical composition.

139. The kit of claim 138, wherein the packaging material includes ice, dry ice, styrofoam, foam, plastic, cellophane, shrink wrap, bubble wrap, paper, cardboard, starch peanuts, twist ties, metal clips, metal cans, drierite, glass and rubber.

144. The glycoprotein of claim 1 wherein said N-linked glycan demonstrates a degree of polymerization of about 15.6.

145. The glycoprotein of claim 1 wherein said N-linked glycan demonstrates a degree of polymerization of about 13.7.

146. The glycoprotein of claim 1 wherein said N-linked glycan demonstrates a degree of polymerization of about 11.6.

147. The glycoprotein of claim 1 wherein said N-linked glycan demonstrates a degree of polymerization of about 10.0.

148. The glycoprotein of claim 1 wherein said N-linked glycan demonstrates a degree of polymerization of about 8.37.

149. The glycoprotein of claim 1 wherein said N-linked glycan demonstrates a degree of polymerization of about 7.32.

150. The glycoprotein of claim 1 wherein said N-linked glycan demonstrates a degree of polymerization of about 6.1.

151. The glycoprotein of claim 1 wherein said N-linked glycan demonstrates a degree of polymerization of about 5.6.

152. The glycoprotein of claim 1 wherein said N-linked glycan demonstrates a degree of polymerization of about 3.8.

153. The glycoprotein of claim 1 wherein said N-linked glycan demonstrates a degree of polymerization of about 3.2.

156. A method of inducing liquefaction of vitreous humor to treat a disorder of a mammalian eye comprising contacting the vitreous humor with an amount of a protein of claim 1 effective to liquefy said vitreous humor whereby the disorder is treated.

157. The polypeptide of claim 156, wherein the polypeptide is modified with a polymer.

158. The polypeptide of claim 157, wherein the polymer is PEG or dextran.

13. A polypeptide, consisting essentially of a hyaluronidase domain or a catalytically active portion thereof of a human hyaluronidase protein.

14. The polypeptide of claim 13, wherein the hyaluronidase is super-sialated.

15. The polypeptide of claim 13, wherein the hyaluronidase domain comprises the sequence of amino acids set forth as amino acids 1-450 of SEQ ID NO. 3.

16. The polypeptide of claim 13, wherein the hyaluronidase domain comprises the sequence of amino acids set forth as amino acids 1-438 of SEQ ID NO. 3.

17. The polypeptide of claim 13, wherein the hyaluronidase domain comprises the sequence of amino acids set forth as amino acids 1-459 of SEQ ID NO. 3.

18. The polypeptide of claim 13, wherein the polypeptide is modified with a polymer.

19. The polypeptide of claim 18, wherein the polymer is PEG or dextran.

29. An isolated substantially pure polypeptide that consists essentially of the hyaluronidase domain of sHASEGP.

31. An isolated soluble human hyaluronidase glycoprotein (sHASEGP), wherein the potency of the sHASEGP is greater than 40,000 USP Units/mg protein.

32. The sHASEGP of claim 31, wherein the hyaluronidase is sialated.

33. The sHASEGP of claim 31, wherein the potency is greater than about 45,000, 55,000, 60,000, 80,000, 90,000, 95,000 or 100,000 Units/mg.

34. The sHASEGP of claim 31, wherein the potency is about 60,000 to 80,000 Units/mg.

35. The sHASEGP of claim 31, wherein the polypeptide is modified with a polymer.

36. The sHASEGP of claim 35, wherein the polymer is PEG or dextran.

37. A composition consisting essentially of a glycoprotein of claim 31.

45. The vector of claim 33 that is a Pichia vector or an E. coli vector.

46. The vector of claim 33, wherein the vector is a viral vector.

39. A nucleic acid molecule, comprising a sequence selected from:

(a) a sequence as set forth in SEQ ID NO. 6;

(b) a sequence that hybridizes under high stringency to at least about 70% of the full-length to the sequence of nucleotides set forth in SEQ ID NO. 6;

(c) a sequence of nucleotides that encodes the polypeptide set forth in SEQ ID No. 1, 3, 4, 5 and 50;

(d) a sequence that is a splice variant of (a), (b) or (c);

(e) a sequence that encodes the hyaluronidase domain or a catalytically active portion thereof that includes a sequence of nucleotides having at least about 60% sequence identity with the sequence set forth in SEQ ID No. 6 or 50; and

(f) a sequence comprising degenerate codons of (a), (b), (c), (d) or (e).

40. A vector having a sequence as set forth in SEQ ID NO:

51.

54. A method for producing a soluble polypeptide that contains sHASEGP, comprising:

introducing a nucleic acid as set forth in SEQ ID NO. 6 operably linked to a promoter into a cell that incorporates N-linked sugar moieties into sHASEGP;

culturing the cell under conditions whereby the encoded polypeptide is expressed by the cell; and

recovering the expressed polypeptide.

56. The method of claim 54, wherein the cell is a eukaryotic cell.

67. A composition, comprising a substantially purified sHASEGP glycoprotein polypeptide and a suitable pharmaceutical carrier.

75. A method for increasing the diffusion of a therapeutic substance or other molecule or macromolecular complex (each an Agent) smaller than about 0.5 microns in diameter in a subject, comprising:

administering to a subject a sHASEGP polypeptide in an amount sufficient to open or to form channels smaller than about 0.5 microns in diameter and an Agent, whereby the diffusion of the Agent is increased.

76. The method of claim 75, wherein said sHASEGP polypeptide is injected at a dose between 0.1 and 1500 Units.

77. The method of claim 75 wherein said sHASEGP polypeptide is mixed with the therapeutic substance or other Agent prior to injection.

78. A kit, comprising:

(a) a sHASEGP polypeptide at a dose between 0.1 and 1500 Units/ml in an acceptable carrier;

(b) at least one therapeutic substance or other molecule or macromolecular complex (each an Agent) in an acceptable carrier; and

(c) optionally, instructions for delivering therapeutic substances or other Agents.

79. The kit of claim 78, wherein the sHASEGP polypeptide and the therapeutic substance or other Agent are provided as a mixture.

140. A method for the treatment of a pathologic accumulation of glycosaminoglycans, comprising:

administration of a recombinant sHASEGP, wherein said sHASEGP is free of bovine, ovine or bacterial enzymes, in an amount sufficient to ameliorate or reduce the accumulated glycosaminoglycan.

141. The method of claim 140, wherein the accumulation is selected from cardiovascular, cerebral, paraphimosis, myxedema, scleromyxedema and lymphedema.

142. A method of facilitating revascularization of necrotic tissue, comprising administration of a recombinant sHASEGP in an amount sufficient to induce the growth of neovasculature into said necrotic tissue.

143. A method for removal of glycosaminoglycans from the vitreous humor comprising;

administration of a sHASEGP.

154. A process for producing a substantially purified neutral active soluble hyaluronidase polypeptide and at least one N-linked sugar moiety, wherein the N-linked sugar moiety is covalently attached to an asparagine residue of the polypeptide, comprising growing Chinese Hamster Ovary cells in a suitable growth media and in the presence of 0.1-1 mM Sodium Butyrate under conditions suitable for production of the polypeptide.

155. A process for purifying a sHASEGP, said process comprising;

contacting sHASEGP containing media under low ionic strength with an anion exchange resin at neutral pH, and eluting said sHASEGP with about 400 mM of a salt;

contact of said sHASEGP with hydrophobic interaction chromatography resin in the presence of about 0.5M ammonium sulfate;

contact of said sHASEGP in ammonium sulfate with phenyl boronate resin;

eluting said sHASEGP in low salt at neutral pH;

contact said sHASEGP with Hydroxyapatite resin, and elution of said sHASEGP with about 100 mM Na Phosphate, resulting in a sHASEGP wherein said sHASEGP is substantially purified.

159. A method of reducing intraocular pressure in the eye of a subject comprising administering to a subject in need thereof, human sHASEGP characterized as

being substantially free of bovine, ovine or bacterial enzymes;

being greater than about 40,000 USP Units/mg protein; and

thereby reducing intraocular pressure in the subject.

160. The method of claim 159, further being free of IgG.

161. The method of claim 159, wherein the sHASEGP is administered to the anterior chamber of the eye.

162. A method of promoting delivery of a sHASEGP or other glycosaminoglycanase (GAG Enzyme) into a first tissue within a mammal, comprising introducing a volume of liquid comprising the GAG Enzyme into the first tissue and allowing the volume of liquid and catalytic activity of the GAG Enzyme to promote delivery of the GAG Enzyme into the tissue.

163. The method of claim 162, wherein the volume of liquid is sufficient to cause localized distension of the first tissue at or near the site of introduction of the volume of liquid.

164. The method of claim 162, further comprising introducing one or more additional pharmacologic or other agents (A-X) into the first tissue.

165. The method of claim 164, wherein A-X is selected from the group consisting of a detectable molecule or other diagnostic agent, an anesthetic or other tissue-modifying agent, a pharmacologic or pharmaceutically effective agent, and a cosmetic or other esthetic agent.

166. The method of claim 164, wherein A-X is a pharmacologic or pharmaceutically effective agent.

167. The method of claim 164, wherein A-X is introduced into the first tissue by inclusion within the volume of liquid comprising the GAG Enzyme.

168. The method of claim 164, wherein A-X is introduced into the first tissue by co-administration with the volume of liquid comprising the GAG Enzyme.

169. The method of claim 164, wherein A-X is introduced into the first tissue by causing its prior introduction into the bloodstream supplying the first tissue.

170. The method of claim 169, wherein the step of causing prior introduction of A-X into the bloodstream supplying the first tissue is initiated prior to the step of introducing the volume of liquid comprising the GAG Enzyme into the first tissue.

171. The method of claim 169, wherein the step of causing prior introduction of A-X into the bloodstream supplying the first tissue is initiated within about one to three hours following the step of introducing the volume of liquid comprising the GAG Enzyme into the first tissue.

172. The method of claim 169, wherein the step of causing prior introduction of A-X into the bloodstream supplying the first tissue is initiated within about twelve hours following the step of introducing the volume of liquid comprising the GAG Enzyme into the first tissue.

173. The method of claim 169, wherein the step of causing prior introduction of A-X into the bloodstream comprises the step of intravenous injection or infusion of A-X.

174. The method of claim 169, wherein the step of causing introduction into the bloodstream comprises the step of oral or other non-intravenous administration of A-X followed by its absorption into the bloodstream.

175. The method claim 162, wherein the GAG Enzyme permeates the first tissue and is dispersed into one or more distal tissues.

176. The method claim 164, wherein A-X is introduced into the first tissue and is also dispersed into one or more distal tissues.

177. The method of claim 176, wherein the first tissue comprises the skin, the subcutaneous layer or a muscle.

178. The method of claim 176, wherein the one or more distal tissues comprise the bloodstream.

179. The method of claim 176, wherein A-X is an agent that modifies the physiology of one or more cells within the first tissue and/or the one or more distal tissues.

180. The method of claim 176, wherein A-X is an anti-cancer agent or an anti-infective agent

181. The method of claim 176, wherein A-X is a molecule or macromolecular complex of between about 20 nm and 200 nm in diameter.

182. The method of claim 164, wherein the first tissue comprises the dermis and A-X is a vaccine.

183. The method of claim 178, wherein A-X is a blood-modifying pharmacologic agent.

184. The method of claim 178, wherein A-X is a pharmacologic or pharmaceutically effective agent delivered to a tissue within the body which is supplied by the bloodstream.

185. The method claim 162, wherein the GAG Enzyme is introduced into the first tissue by transmucosal, transdermal, intradermal, subcutaneous, intramuscular, periorbital, intraorbital or intrathecal injection.

186. The method of claim 162, wherein the first tissue is selected from the group consisting of skin, a subcutaneous layer, a muscle and the eye.

187. The method of claim 162, wherein introducing comprises intradermal delivery.

188. The method of claim 187, wherein the intradermal delivery comprises introducing a needle into the dermis.

189. The method of claim 162, wherein introducing comprises transdermal delivery.

190. The method of claim 162, wherein introducing comprises subcutaneous delivery.

191. The method of claim 190, wherein the subcutaneous delivery comprises introducing a needle into the subcutaneous layer.

192. The method of claim 162, wherein introducing comprises intramuscular delivery.

193. The method of claim 162, wherein introducing comprises periorbital or intraorbital delivery.

194. The method of claim 193, wherein introducing comprises introduction into the episcleral space.

195. The method of claim 194, wherein A-X is delivered across the sclera to the choroid and/or choroid.

196. The method of claim 193, wherein A-X is an agent that modifies the physiology of one or more cells or tissues within the eye.

197. The method of claim 196, wherein A-X is an anesthetic agent or an anti-inflammatory agent.

198. The method of claim 196, wherein A-X is an anti-infective agent.

199. The method of claim 164, wherein A-X is introduced into the first tissue by transmucosal, transdermal, intradermal, subcutaneous, intramuscular, periorbital, intraorbital or intrathecal injection.

200. The method of claim 164, wherein the GAG Enzyme and A-X are co-formulated in the liquid and introduced together into the first tissue by transmucosal, transdermal, intradermal, subcutaneous, intramuscular, periorbital, intraorbital or intrathecal injection.

201. The method of claim 164, wherein the GAG Enzyme and A-X are co-administered into the first tissue by transmucosal, transdermal, intradermal, subcutaneous, intramuscular, periorbital, intraorbital or intrathecal injection.

202. The method of claim 164, wherein A-X is a molecule selected from the group consisiting of a small molecule, a protein, and a nucleic acid.

203. The method of claim 164, wherein A-X is a macromolecular complex comprising combinations of molecules selected from the group consisting of small molecules, proteins, nucleic acids, lipids and amphiphilic molecules.

204. The method of claim 164, wherein A-X is selected from the group consisting of a chemotherapeutic or anticancer 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 antiarthritics agent, an anti-fungal agent, an antihypertensive agent, an antipyretic agent, an anti-parasitic agent, an antihistamine agent, an alpha-adrenargic agonist agent, an alpha blocker agent, an anesthetic agent, a bronchial dilator agent, a biocide agent, a bactericide agent, a bacteriostatic agent, a beta adrenergic blocker agent, a calcium channel blocker agent, a cardiovascular drug agent, a contraceptive agent, a cosmetic or esthetic 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 psychic energizer agent, a sedative agent, a sleep inducer, a sympathomimetic agent, a tranquilizer agent, a urinary agent, a vaginal agent, a viricide agent, a vitamin agent, a non-steroidal anti-inflammatory agent, or an angiotensin converting enzyme inhibitor agent.

205. The method of claim 164, wherein A-X is selected from the group consisting of Adalimumabs (e.g. HUMIRA™

), Agalsidase Betas (e.g. FABRAZYME™

), Aldesleukins (PROLEUKIN™

IL-2), Alefacepts (e.g. AMEVIVE™

), Ampicillins (e.g. UNASYN™

Injection), Anakinras (e.g. KINERET™

), Antipoliomyelitic Vaccines (e.g. PEDIARIX™

), Anti-Thymocytes (e.g. THYMOGLOBULIN™

), Azithromycins (e.g. ZITHROMAX™

IV), Becaplermins (e.g. REGRANEX™

), Caspofungins (e.g. CANCIDAS™

), Cefazolins (e.g. ANCEF™ and

CEFAZOLIN™

), Cefepimes (e.g. MAXIPIME™

), Cefotetans (e.g. CEFOTAN™

), Ceftazidimes (e.g. FORTAZ™

), Ceftriaxones (e.g. ROCEFIN™

), Cetuximabs (e.g. ERBITUX™

), Cilastatins (e.g. PRIMAXIN™

IV), Clavulanic Acids (e.g. in conjunction with Amoxicillins such as in AUGMENTIN™

), Clindamycins (e.g. CLEOCIN™

), Darbepoetin Alfas (e.g. ARANESP™

), Deaclizumabs (e.g. ZENAPAX™

), Diphtheria Toxoids (typically in combinations e.g. DAPTACEL™

, INFANRIX™ and

PEDIARIX™

), Efalizumabs (e.g. RAPTIVA™

), Epinephrines (e.g. EPIPEN™

), Erythropoietin Alphas (e.g. EPOGEN™

And PROCRIT™

), Etanercepts (e.g. ENBREL™

), Filgrastims (e.g. NEUPOGEN™

), Fluconazoles (e.g. DIFLUCAN™

Injection), Follicle-Stimulating Hormones such as Follitropin Alphas (e.g. GONAL-F™

) and Follitropin Betas (e.g. FOLLISTIM™

), Fosphenyloins (e.g. CEREBYX™

), Fluconazoles (e.g. DIFLUCAN™

), Gadodiamides (e.g. OMNISCAN™

), Gadopentetates (e.g. MAGNEVIST™

), Gatifloxacins (e.g. TEQUIN™

), Glatiramers (e.g. COPAXONE™

), GM-CSF'"'"'s (e.g. LEUKINE™

), Goserelins (e.g. ZOLADEX™

), Granisetrons (e.g. KYTRIL™

), Haemophilus Influenza B'"'"'s (e.g. COMVAX™ and

HibTITER™

), Haloperidols (e.g. HALDOL™

), Hepatitis A Vaccines (e.g. HAVRIX™

, TWINRIX™ and

VAQTA™

), Hepatitis B Vaccines (e.g. recombinants COMVAX™

, ENGERIX™

-B, RECOMBIVAX™

-HB, TWINRIX™

, and non-recombinants BAYHEP™

-B and NABI™

-HB), Ibritumomab Tiuxetans (e.g. ZEVALIN™

), Immunoglobulins, Influenza Virus Vaccines (e.g. FLUMIST™

), Infliximabs (e.g. REMICADE™

), Insulins (e.g. HUMALOG™

, HUMALOG™

MIX 75/25™

, HUMULIN™

products (incl. 50/50, 70/30, Regular, NPH, Ultra and Ultralente), and NOVOLIN™

), Insulin Glargines (e.g. LANTUS™

), Interferon Alfa-2a'"'"'s (ROFERAN™

-A), Interferon Alfa-2b'"'"'s (e.g. INTRON-A™

), Interferon Alfacon-1'"'"'s (e.g. INFERGEN™

), Interferon Alfa-n3s (e.g. ALFERON N™

), Interferon Betas (e.g. BETASERON™

And BETAFERON™

), Interferon Beta-1a'"'"'s (e.g. AVONEX™

And REBIF™

), Interferon Gammas (e.g. ACTIMMUNE™

), Iodixanols (e.g. VISIPAQUE™

), Iohexols (OMNIPAQUE™

), Iopamidols, Ioversols (e.g. OPTIRAY™

), Ketorolacs (e.g. TORADOL™

), Laronidases (e.g. ALDURAZYME™

), Levofloxacins (e.g. LEVAQUIN™

), Lidocaines, Linezolids (e.g. ZYVOX™

), Lorazepams (e.g. ATIVAN™

), Measles Vaccines (e.g. ATTENUVAX™

), Measles-Mumps-Rubella Virus Vaccines (e.g. M-M-R™

II), Medroxyprogesterones (e.g. DEPO-PROVERA™

), Meropenems (e.g. MERREM™

IV), Methylprednisolones (e.g. SOLU-MEDROL™

), Midazolams (e.g. VERSED™

), Morphines (e.g. ASTRAMORPH/PF™

), Octreotides (e.g. SANDOSTATIN™

), Omalizumabs (e.g. XOLAIR™

), Ondansetrons (e.g. ZOFRAN™

), Palivizumabs (e.g. SYNAGIS™

), Pantoprazoles (e.g. PROTONIX™

), Pegaspargases (e.g. ONCASPAR™

), Pegfilgrastims (e.g. NEULASTA™

), Peg-Interferon Alfa-2a'"'"'s (e.g. PEGASYS™

), Peg-Interferon Alfa-2b'"'"'s (e.g. PEG-INTRON™

), Pegvisomants (e.g. SOMAVERT™

), Pertussis vaccines, Piperacillins (e.g. ZOSYN™

), Pneumococcal Vaccines (e.g. PNEUMOVAX™

     23) and Pneumococcal Conjugate Vaccines (e.g. PREVNAR™

), Promethazines (e.g. PHENERGAN™

), Reteplases (e.g. RETAVASE™

), Somatropins (e.g. GENOTROPIN™

, HUMATROPE™

, NORDITROPIN™

, NUTROPIN™

, SAIZEN™

, SEROSTIM™

, ZORBTIVE™

), Sulbactams, Sumatriptans (e.g. IMITREX™

), Tazobactams, Tenecteplases (e.g. TNKASE™

), Tetanus Purified Toxoids, Ticarcillins (e.g. TIMENTIN™

), Tositumomabs (e.g. BEXXAR™

), Triamcinolone Acetonides, Vancomycins (e.g. VANCOCIN™

), Varicella vaccines (e.g. VARIVAX™

), and other vaccines.

206. The method of claim 164, wherein A-X is an antibiotic agent or combination of agents selected from the group consisting of Aminoglycosides;

Amphenicols;

Ansamycins;

Carbacephems;

Carbapenems;

Cephalosporins or Cephems;

Cephamycins;

Clavams;

Cyclic lipopeptides;

Diaminopyrimidines;

Ketolides;

Lincosamides;

Macrolides;

Monobactams;

Nitrofurans;

Oxacephems;

Oxazolidinones;

Penems, thienamycins and miscellaneous beta-lactams;

Penicillins;

Polypeptides antibiotics;

Quinolones;

Sulfonamides;

Sulfones;

Tetracyclines; and

other antibiotics (such as Clofoctols, Fusidic acids, Hexedines, Methenamines, Nitrofurantoins Nitroxolines, Ritipenems, Taurolidines, Xibomols).

207. The method of claim 200, wherein the volume of liquid further comprises an additional pharmacologic agent.

208. The method of claim 162, wherein the GAG Enzyme is a sHASEGP.

209. The method of claim 208, wherein the sHASEGP is a human sHASEGP.

210. The method of claim 209, wherein the human sHASEGP is a PH20 sHASEGP.

211. The method of claim 210, wherein the sHASEGP is a soluble polypeptide as set forth in SEQ ID NO. 4.

212. The method of claim 208, wherein the sHASEGP is a pegylated or super-sialated sHASEGP.

213. The method of claim 211, wherein the sHASEGP is a pegylated human PH20 sHASEGP.

214. The method of claim 162, wherein the liquid is a pharmaceutical excipient

215. The method of claim 162, wherein the volume is between about 1 to 10 ml.

216. The method of claim 162, wherein the first tissue is itself or is proximal to a tissue comprising an excess of a glycosaminoglycan.

217. The method of claim 162, wherein the first tissue is itself or is proximal to a tissue exhibiting an elevated interstitial or oncotic pressure.

218. The method of claim 162, wherein the first tissue is itself or is proximal to a tumor.

219. The method of claim 162, wherein the first tissue is itself or is proximal to a tissue which is edemic.

220. The method of claim 162, wherein introducing the GAG Enzyme into the first tissue increases interstitial flow within the tissue.

221. The method of claim 162, wherein introducing the GAG Enzyme into the first tissue increases permeability within the tissue.

222. A method of increasing the permeability of a target tissue of a mammal, comprising introducing a sHASEGP into the tissue.

223. The method of claim 222, wherein the sHASEGP is introduced into the tissue by injection of the sHASEGP into the tissue or into a nearby tissue.

224. The method of claim 222, wherein the sHASEGP is introduced into the tissue by prior introduction of the sHASEGP into the bloodstream supplying the tissue.

225. The method of claim 224, wherein the prior introduction into the bloodstream is mediated by intravenous injection or infusion of the sHASEGP.

226. The method of claim 224, wherein the prior introduction into the bloodstream is mediated by non-intravenous parenteral administration of the sHASEGP.

227. The method of claim 222, further comprising introducing A-X into the tissue.

228. The method of claim 227, wherein A-X is introduced into the tissue by injection into the tissue or into a nearby tissue.

229. The method of claim 227, wherein A-X is introduced into the tissue by prior introduction of the PH20 into the bloodstream supplying the tissue.

230. The method of claim 229, wherein the prior introduction into the bloodstream is mediated by intravenous injection or infusion of the molecule or macromolecular complex.

231. The method of claim 229, wherein the prior introduction into the bloodstream is mediated by non-intravenous parenteral administration of the molecule or macromolecular complex.

232. The method of claim 227, wherein the step of introducing A-X into the tissue is initiated within three hours following the step of introducing the sHASEGP into the tissue.

233. The method of claim 227, wherein the step of introducing A-X into the tissue is initiated within twelve hours following the step of introducing the sHASEGP into the first tissue.

234. The method of claim 227, wherein A-X is a molecule selected from the group consisiting of a small molecule, a protein, and a nucleic acid.

235. The method of claim 227, wherein A-X is a macromolecular complex comprising combinations of molecules selected from the group consisting of small molecules, proteins, nucleic acids, lipids and amphiphilic molecules.

236. The method of claim 227, wherein A-X is selected from the group consisting of a detectable molecule or other diagnostic agent, an anesthetic or other tissue-modifying agent, a pharmacologic or pharmaceutically effective agent, and a cosmetic or other esthetic agent.

237. The method of claim 227, wherein A-X is a pharmacologic or pharmaceutically effective agent.

238. The method of claim 227, wherein A-X is selected from the group consisting of a chemotherapeutic or anticancer 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 antiarthritics agent, an anti-fungal agent, an antihypertensive agent, an antipyretic agent, an anti-parasitic agent, an antihistamine agent, an alpha-adrenargic agonist agent, an alpha blocker agent, an anesthetic agent, a bronchial dilator agent, a biocide agent, a bactericide agent, a bacteriostatic agent, a beta adrenergic blocker agent, a calcium channel blocker agent, a cardiovascular drug agent, a contraceptive agent, a cosmetic or esthetic 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 psychic energizer agent, a sedative agent, a sleep inducer, a sympathomimetic agent, a tranquilizer agent, a urinary agent, a vaginal agent, a viricide agent, a vitamin agent, a non-steroidal anti-inflammatory agent, or an angiotensin converting enzyme inhibitor agent.

239. The method of claim 227, wherein A-X is an anticancer agent or combination of agents selected from the group consisting of Aclacinomycins, Actinomycins, Adriamycins, Ancitabines, Anthramycins, Azacitidines, Azaserines, 6-Azauridines, Bisantrenes, Bleomycins, Cactinomycins, Carmofurs, Carmustines, Carubicins, Carzinophilins, Chromomycins, Cisplatins, Cladribines, Cytarabines, Dactinomycins, Daunorubicins, Denopterins, 6-Diazo-5-Oxo-L-Norleucines, Doxifluridines, Doxorubicins, Edatrexates, Emitefurs, Enocitabines, Fepirubicins, Fludarabines, Fluorouracils, Gemcitabines, Idarubicins, Loxuridines, Menogarils, 6-Mercaptopurines, Methotrexates, Mithramycins, Mitomycins, Mycophenolic Acids, Nogalamycins, Olivomycines, Peplomycins, Pirarubicins, Piritrexims, Plicamycins, Porfiromycins, Pteropterins, Puromycins, Retinoic Acids, Streptonigrins, Streptozocins, Tagafurs, Tamoxifens, Thiamiprines, Thioguanines, Triamcinolones, Trimetrexates, Tubercidins, Vinblastines, Vincristines, Zinostatins, And Zorubicins.

240. The method of claim 227, wherein A-X is an antibody or other protein having anti-cancer or chemotherapeutic activity.

241. The method of claim 227, wherein A-X is an antibiotic agent or combination of agents selected from the group consisting of Aminoglycosides;

Amphenicols;

Ansamycins;

Carbacephems;

Carbapenems;

Cephalosporins or Cephems;

Cephamycins;

Clavams;

Cyclic lipopeptides;

Diaminopyrimidines;

Ketolides;

Lincosamides;

Macrolides;

Monobactams;

Nitrofurans;

Oxacephems;

Oxazolidinones;

Penems, thienamycins and miscellaneous beta-lactams;

Penicillins;

Polypeptides antibiotics;

Quinolones;

Sulfonamides;

Sulfones;

Tetracyclines; and

other antibiotics (such as Clofoctols, Fusidic acids, Hexedines, Methenamines, Nitrofurantoins Nitroxolines, Ritipenems, Taurolidines, Xibomols).

242. The method of claim 222, wherein the sHASEGP is a human sHASEGP.

243. The method of claim 242, wherein the human sHASEGP is a PH20 sHASEGP.

244. The method of claim 243, wherein the sHASEGP is a soluble polypeptide as set forth in SEQ ID NO. 4.

245. The method of claim 222, wherein the sHASEGP is a pegylated or super-sialated sHASEGP.

246. The method of claim 244, wherein the sHASEGP is a pegylated human PH20 sHASEGP.

253. The method of claim 246, wherein the sHASEGP is a pegylated or super-sialated sHASEGP.

255. The method of claim 246, wherein the mammal is a human and the sHASEGP is a pegylated sHASEGP.

247. A method of ameliorating the consequences of stroke or other CNS injury in a mammal, comprising introducing a sHASEGP into a mammal following said stroke or CNS injury.

248. The method of claim 247, wherein said step of introducing a sHASEGP comprises intravenous administration of a sHASEGP.

249. The method of claim 247, wherein said step of introducing a sHASEGP comprises intrathecal administration of a sHASEGP.

250. The method of claim 247, wherein the sHASEGP is a human sHASEGP.

251. The method of claim 250, wherein the human sHASEGP is a PH20 sHASEGP.

252. The method of claim 251, wherein the sHASEGP is a soluble polypeptide as set forth in SEQ ID NO. 4.

254. The method of claim 252, wherein the sHASEGP is a pegylated human PH20 sHASEGP.