Soluble glycosaminoglycanases and methods of preparing and using soluble glycosaminogly ycanases

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.

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.  
     
     
         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.  
     
     
         20 . (canceled)  
     
     
         21 . (canceled)  
     
     
         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 . (canceled)  
     
     
         25 . (canceled)  
     
     
         26 . (canceled)  
     
     
         27 . (canceled)  
     
     
         28 . (canceled)  
     
     
         29 . (canceled)  
     
     
         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.  
     
     
         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 . (canceled)  
     
     
         34 . (canceled)  
     
     
         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 . (canceled)  
     
     
         38 . A nucleic acid molecule, encoding the polypeptide of  claim 1 .  
     
     
         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.  
     
     
         41 . A vector comprising the nucleic acid molecule of  claim 38 .  
     
     
         42 . (canceled)  
     
     
         43 . (canceled)  
     
     
         44 . (canceled)  
     
     
         45 . (canceled)  
     
     
         46 . (canceled)  
     
     
         47 . An isolated cell, comprising the vector of  claim 41 .  
     
     
         48 . (canceled)  
     
     
         49 . (canceled)  
     
     
         50 . (canceled)  
     
     
         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.  
     
     
         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.    
     
     
         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.    
     
     
         56 . (canceled)  
     
     
         57 . (canceled)  
     
     
         58 . (canceled)  
     
     
         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 . (canceled)  
     
     
         62 . (canceled)  
     
     
         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 . (canceled)  
     
     
         67 . A composition, comprising a substantially purified sHASEGP glycoprotein polypeptide and a suitable pharmaceutical carrier.  
     
     
         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 . (canceled)  
     
     
         73 . The method of  claim 70 , wherein said scar tissue is a keloid scar.  
     
     
         74 . (canceled)  
     
     
         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.  
     
     
         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 . (canceled)  
     
     
         88 . (canceled)  
     
     
         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 a gent, 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 . (canceled)  
     
     
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         119 . (canceled)  
     
     
         120 . (canceled)  
     
     
         121 . (canceled)  
     
     
         122 . (canceled)  
     
     
         123 . A combination comprising, 
 a sterile vial and the pharmaceutical composition of  claim 89  wherein the composition is contained in the vial.    
     
     
         124 . (canceled)  
     
     
         125 . (canceled)  
     
     
         126 . (canceled)  
     
     
         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 . (canceled)  
     
     
         129 . A combination comprising, 
 a sterile syringe containing the pharmaceutical composition of  claim 89 .    
     
     
         130 . (canceled)  
     
     
         131 . (canceled)  
     
     
         132 . A combination comprising, 
 a sterile syringe containing the pharmaceutical composition of  claim 110 .    
     
     
         133 . (canceled)  
     
     
         134 . (canceled)  
     
     
         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 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, 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 . (canceled)  
     
     
         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.    
     
     
         144 . (canceled)  
     
     
         145 . (canceled)  
     
     
         146 . (canceled)  
     
     
         147 . (canceled)  
     
     
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         153 . (canceled)  
     
     
         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.  
     
     
         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 . (canceled)  
     
     
         158 . (canceled)  
     
     
         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 . (canceled)  
     
     
         161 . (canceled)  
     
     
         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 . (canceled)  
     
     
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         203 . (canceled)  
     
     
         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™), Fosphenytoins (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™), Inmunoglobulins (including mixtures of immunoglobulins such as GAMMAGARD™ and the like, as well as any of a variety of purified 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 . (canceled)  
     
     
         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 . (canceled)  
     
     
         215 . (canceled)  
     
     
         216 . (canceled)  
     
     
         217 . (canceled)  
     
     
         218 . (canceled)  
     
     
         219 . (canceled)  
     
     
         220 . (canceled)  
     
     
         221 . (canceled)  
     
     
         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.  
     
     
         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 . (canceled)  
     
     
         249 . (canceled)  
     
     
         250 . (canceled)  
     
     
         251 . (canceled)  
     
     
         252 . (canceled)  
     
     
         253 . (canceled)  
     
     
         254 . (canceled)  
     
     
         255 . (canceled)  
     
     
         256 . The polypeptide of any one of claims  12 ,  19  or  158 , wherein the polymer is PEG.  
     
     
         257 . The polypeptide of  claim 256 , wherein the polypeptide comprises at least about three PEG moieties per polypeptide.  
     
     
         258 . The polypeptide of  claim 256 , wherein said PEG moieties are branched.  
     
     
         259 . The polypeptide of  claim 256 , wherein said PEG moieties are selected from the group consisting of mPEG-SBA (5 kDa), mPEG-SBA (20 kDa), mPEG-SBA 30 kDa), mPEG-SMB (20 kDa) mPEG-SMB (30 kDa), mPEG-butyrldehyde (30 kDa), mPEG-SPA (20 kDa), mPEG-SPA (30 kDa), mPEG2-NHS (10 kDa branched), mPEG2-NHS (20 kDa branched), mPEG-NHS (40 kDa branched), mPEG2-NHS (60 kDa branched), PEG-NHS-biotin (5 kDa biotinylated), PEG-p-nitrophenyl-carbonate (30 kDa) and PEG-propionaldehyde (30 kDa).  
     
     
         260 . A method of preparing a PEGylated polypeptide, comprising reacting a polypeptide of any one of claims  12 ,  19  or  158  with a molar excess of a PEG reagent selected from the group consisting of a PEG-succinimidyl butanoate, a PEG-succinimidyl methylbutanoate, a PEG-succinimidyl proprionate, a PEG-N-hydroxysuccinimide, a PEG-aldehyde, a PEG-carbonate, a PEG-propionaldehyde and a PEG-maleimide.  
     
     
         261 . The method of  claim 260  wherein the PEG reagent is selected from the group consisting of mPEG-SBA (5 kDa), mPEG-SBA (20 kDa), mPEG-SBA 30 kDa), mPEG-SMB (20 kDa) mPEG-SMB (30 kDa), mPEG-butyrldehyde (30 kDa), mPEG-SPA (20 kDa), mPEG-SPA (30 kDa), mPEG2-NHS (10 kDa branched), mPEG2-NHS (20 kDa branched), mPEG-NHS (40 kDa branched), mPEG2-NHS (60 kDa branched), PEG-NHS-biotin (5 kDa biotinylated), PEG-p-nitrophenyl-carbonate (30 kDa) and PEG-propionaldehyde (30 kDa).  
     
     
         262 . A PEGylated polypeptide produced according to the method of  claim 260 .  
     
     
         263 . A PEGylated polypeptide produced according to the method of  claim 261 .  
     
     
         264 . A PEGylated hyaluronidase, wherein the hyaluronidase comprises at least about three PEG moieties per polypeptide.  
     
     
         265 . The hyaluronidase of  claim 264 , wherein said PEG moieties are branched.  
     
     
         266 . The hyaluronidase of  claim 264 , wherein said PEG moieties are selected from the group consisting of mPEG-SBA (5 kDa), mPEG-SBA (20 kDa), mPEG-SBA 30 kDa), mPEG-SMB (20 kDa) mPEG-SMB (30 kDa), mPEG-butyrldehyde (30 kDa), mPEG-SPA (20 kDa), mPEG-SPA (30 kDa), mPEG2-NHS (10 kDa branched), mPEG2-NHS (20 kDa branched), mPEG-NHS (40 kDa branched), mPEG2-NHS (60 kDa branched), PEG-NHS-biotin (5 kDa biotinylated), PEG-p-nitrophenyl-carbonate (30 kDa) and PEG-propionaldehyde (30 kDa).  
     
     
         267 . The hyaluronidase of  claim 264 , wherein the hyaluronidase is a PEGylated sHASEGP.  
     
     
         268 . The hyaluronidase of  claim 264 , wherein the hyaluronidase is a PEGylated rHuPH20.  
     
     
         269 . The hyaluronidase of  claim 264 , wherein the hyaluronidase is a PEGylated animal-derived or bacterial-derived hyaluronidase.  
     
     
         270 . A method of preparing a-PEGylated hyaluronidase, comprising reacting a hyaluronidase with a molar excess of a PEG reagent selected from the group consisting of a PEG-succinimidyl butanoate, a PEG-succinimidyl methylbutanoate, a PEG-succinimidyl proprionate, a PEG-N-hydroxysuccinimide, a PEG-aldehyde, a PEG-carbonate, a PEG-propionaldehyde and a PEG-maleimide.  
     
     
         271 . The method of  claim 270  wherein the PEG reagent is selected from the group consisting of mPEG-SBA (5 kDa), mPEG-SBA (20 kDa), mPEG-SBA 30 kDa), mPEG-SMB (20 kDa) mPEG-SMB (30 kDa), mPEG-butyrldehyde (30 kDa), mPEG-SPA (20 kDa), mPEG-SPA (30 kDa), mPEG2-NHS (10 kDa branched), mPEG2-NHS (20 kDa branched), mPEG-NHS (40 kDa branched), mPEG2-NHS (60 kDa branched), PEG-NHS-biotin (5 kDa biotinylated), PEG-p-nitrophenyl-carbonate (30 kDa) and PEG-propionaldehyde (30 kDa).  
     
     
         272 . A PEGylated hyaluronidase produced according to the method of  claim 270 .  
     
     
         273 . A PEGylated hyaluronidase produced according to the method of  claim 271 .  
     
     
         274 . A method of delivering a molecule to a tissue containing excess amounts of glycosaminoglycan, comprising: administering a PEGylated hyaluronidase of  claim 264  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.  
     
     
         275 . A pharmaceutical composition, comprising a PEGylated hyaluronidase of  claim 264  and a pharmaceutical carrier.  
     
     
         276 . A method for increasing the diff-usion 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 PEGylated hyaluronidase of  claim 264  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.    
     
     
         277 . The method of  claim 276 , wherein the 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.  
     
     
         278 . A kit, comprising: 
 (a) a PEGylated hyaluronidase of  claim 264  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.    
     
     
         279 . A method for removal of glycosaminoglycans from the vitreous humor comprising; 
 administration of a PEGylated hyaluronidase of  claim 264 .    
     
     
         280 . 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 PEGylated hyaluronidase of  claim 264  effective to liquefy said vitreous humor whereby the disorder is treated.  
     
     
         281 . A method of ameliorating the consequences of stroke or other CNS injury in a mammal, comprising introducing a PEGylated hyaluronidase of  claim 264  into a mammal following said stroke or CNS injury.  
     
     
         282 . A method of introducing one or more pharmacologic or other agents (A-X) into a tissue of the body comprising administering a PEGylated hyaluronidase of  claim 264  to said tissue prior to or coincident with administering said agent.  
     
     
         283 . A method of  claim 282 , 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.  
     
     
         284 . The method of  claim 282 , wherein A-X is an anti-cancer agent or an anti-infective agent.  
     
     
         285 . The method of  claim 282 , wherein A-X is a blood-modifying pharmacologic agent  
     
     
         286 . The method of  claim 282 , wherein A-X is a molecule or macromolecular complex of between about 20 nm and 200 nm in diameter.  
     
     
         287 . The method of  claim 282 , 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™), Fosphenytoins (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 (including mixtures of immunoglobulins such as GAMMAGARD™ and the like, as well as any of a variety of purified 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-la's (e.g. AVONEX™ And REBIF™), Interferon Gamrnas (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. PHTNERGAN™), 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.  
     
     
         288 . The method of  claim 282 , 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).  
     
     
         289 . The method of  claim 282 , 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.