Status: approved and marketed in U.S. and EU

Organizations involved:

Halozyme Therapeutics, Inc. – Manuf.; R&D; Tech

Baxter Healthcare Corp. – USA mark.; Europe mark.; Parent

Baxter Pharm. Solutions LLC – Manuf. other

MidAtlantic Diagnostics, Inc. – World mark.

MediCult A/S – World mark;

Hoffmann-La Roche Ltd. – R&D; World mark.

Avid Bioservices, Inc. – Manuf.

Cook Pharmica LLC – Manuf.

Peregrine Pharmaceuticals Inc. – Parent

University of Connecticut – R&D; Tech.

ProMetic Life Sciences Inc. – Tech.

Sidney Kimmel Cancer Center – Former

ProMetic Life Sciences Inc. – Manuf. other

Cross ref.: See the other hyaluronidase entries, e.g., Wydase (#616) and Amphadase (#617), in the Enzymes section for background information and about currently marketed bovine and ovine hyaluronidase products.

Description: Hylenex (formerly Enhanze SC) is an injectable aqueous formulation and Cumulase is an ex vivo reagent/medical device, both containing soluble recombinant fragment (amino acids 36-482) of PH-20 human hyaluronidase (rHuPH20) glycoprotein expressed by transformed Chilnese hamster ovary (CHO) cells. The glycoprotein is composed of 447 amino acids, with an approximate molecular weight of 61 kDa.

Hylenex is packaged in 2 mL vials each containing 150 USP units of hyaluronidase per mL (300 USP units total), along with 8.5 mg sodium chloride, 1.8 mg sodium phosphate dibasic dihydrate, 4.2 mg sodium hydroxide, 1.0 mg Albumin (human), 1.0 mg edetate disodium dihydrate (EDTA), and 0.4 mg of calcium chloride dihydrate.

Currently-marketed products are Cumulase and SynVitro Cumulase for use in in vitro fertilization (IVF; recently approved); and Hylenex, rHuPH20 used as a spreading agent, e.g., injected intravitreally into the vitreous humor (fluid of the eye) along with a local anesthetic to facilitate spreading of the local anesthetic. Cumulase is an ex vivo formulation to replace the bovine hyaluronidase, Amphadase (which replaced Wydase), currently used for the preparation of oocytes (eggs) prior to in vitro fertilization (IVF) by intracytoplasmic sperm injection (ICSI). Other indications: are as a spreading agent for contrast agents, and for subcutaneous fluid replacement (hypodermoclysis; intravenous fluids extravasation). These indications: for rHuPH20 are essentially the same as for current natural hyaluronidases.

Recombinant human hyaluronidase has more than 100 times higher purity than the active ingredient in Wydase, the original bovine testes-derived hyaluronidase, recently replaced by generically equivalent (comparably purity) Amphadase. See the entries for Wydase (#616) and Amphadase (#617).

SynVitro Cumulase is rHuPH20 (Cumulase) for in vitro fertilization use prepared in proprietary SynVitro media from MediCult.

Halozyme is developing a new, improved manufacturing process for rHuPH20. This is "optimized for productivity and product safety. Once commercialized, it is expected to result in greater than 90% reduction in cost of goods relative to the product produced by the first generation process." This may or likely involves a new CHO cell line, vector constructs, including promoter and, possibly, chaperone, sequences, and downstream purification improvements. As discussed below, rHuPH20 manufactured by the new process has shown no signs of problems in non-humna primate studies.

Nomenclature: Hyaluronidase, rDNA [BIO]; Hylenex [TR]; Enhanze SC [TR former]; Cumulase [TR]; SynVitro Cumulase [TR]; hyaluronidase human injection [FDA]; hyaluronidase (human recombinant) [USAN]; 36-482-hyaluronoglucosaminidase PH20 (human) [CAS]; hyaluronidase 1 (human sperm surface protein PH-20)-(1-447)peptide [CAS]; 57971-58-7 [CAS RN]; rHuPH20 [SY]; hyaluronidase PH-20, recombinant human [SY]; PH-20 hyaluronidase [SY]; E.C. 3.1.25 [EC]; NDC 60977-319-03 [NDC]

Biological.: PH20 hyaluronidase is naturally found in human sperm. Much like use of rHuPH20 (Cumulase) for in vitro fertilization procedures, PH20 breaks down hyaluronic acid in the outer cell membrane of oocytes (eggs), faciliating entry of sperm into the the oocyte.

Cumulase is an ex vivo (used outside of the body) formulation of rHuPH20 to replace the bovine/ovine enzyme currently used for the preparation of oocytes (eggs) prior to in vitro fertilization (IVF) during the process of intracytoplasmic sperm injection (ICSI), in which the enzyme is an essential component. The enzyme strips away the hyaluronic acid that surrounds the oocyte, allowing the clinician to then perform the ICSI procedure, injecting the sperm into the oocyte.

See the other hyaluronidase entries, e.g., Wydase (#616) and Amphadase (#617), in the Enzymes section for further background information.

rHuPH20 forms the basis for Halozyme’s platform Enhanze Technology, a proprietary drug enhancement system. When co-formulated and injected subcutaneously with other drugs, rHuPH20 may facilitate drug penetration and dispersion by temporarily opening flow channels under the skin. Molecules as large as 200 nanometers may pass freely through the perforated extracellular matrix, which recovers its normal density within 24 hours, without permanently altering the structure of the skin.

Halozyme is also developing rHuPH20 (Chemophase) as a biologic for adjuvant use with chemotherapy for treatment of solid tumors (using the enzyme to open up channels for spread of chemotherapy drugs into solid tumors).

In the May 2005 issue of Drug Delivery Technology, preclinical animal studies with Enhanze SC were reported showing that rHuPH20 was effective for breaking down the hyaluronic acid of interstitial matrix, allowing subcutaneous, intradermal, and intramuscular administration of biopharmaceuticals, including monoclonal antibodies, and other therapeutics not previously delivered by these routes. Use of rHuPH20 permitted administration of up to five times more volume than normally feasible for subcutaneously injected drugs, e.g., 5-10 mL vs. 1-2 mL, respectively. This may permit conversion of intravenously-administered pharmaceuticals to more desirable local routes, and may even permit patients to self-administer select chronic injectable medicines, reducing the need for hospital visits.

In Sept. 2005, Halozyme reported positive results with rHuPH20 and rHuPH20 conjugated with polyethylene glycol (PEG) in a rodent model of severe ischemic stroke. rHuPH20 alone increased survival at 28 days compared with control (p=0.049, 69% vs. 48%, respectively). With pegylated rHuPH20 , survival increased to 78% (p=0.006).

In April 2006, Halozyme reported that combining intravesicle instillation of mitomycin with Chemophase significantly suppressed tumor growth vs. placebo (p<0.01) in mouse models of bladder cancer, while mitomycin alone did not have a significant response vs. placebo. Combined use was more effective when initial tumor volumes at first treatment were >100 mm3. The treatment was well tolerated with no related toxicity.

In March 2008, Halozyme reported new safety and pharmacokinetic data from three studies in monkeys of a second generation manufacturing process for rHuPH20. The objectives were to investigate the intravenous (IV) and subcutaneous (SC) safety assessment of rHuPH20 administered at significantly higher doses to non-human primates than previously examined. The use of rHuPH20 at higher concentrations in formulations may potentially facilitate the conversion from IV to SC administration of therapeutic proteins by allowing increased injection volumes and local dispersion and absorption of SC co-injected therapeutic proteins into the systemic circulation. In a pharmacokinetic (PK) dose-range finding study, doses up to 3,600,000 U/kg (30 mg/kg) were well-tolerated following either IV or SC dosing. Daily administration of rHuPH20 for one week was well tolerated via either IV or SC delivery at a daily bolus dose of 600,000 U/kg (5 mg/kg). No test article-related changes were observed in any toxicology parameters and no systemic enzyme accumulation was observed. A 3-month interim analysis from an ongoing 39-week chronic toxicity study with weekly dosing up to 240,000 U/kg revealed no changes in standard toxicity parameters or in male fertility assessment..

Companies.: Halozyme Therapeutics, Inc. developed and manufactures rHuPH20, Cumulase and Enhanze SC/Hylenex. With its founding, Halozyme exclusively licensed recombinant PH-20 hyaluronidases from the University of Connecticut. Halozyme was founded in Feb. 1998, and spun out of Sidney Kimmel Cancer Center in 2000. Halozyme completed its initial public offering in March 2004.

MediCult A/S markets Cumulase worldwide as a1X ready-to-use formulation, and MidAtlantic Diagnostics, Inc. markets it worldwide as a 10X concentrate.

In Aug. 2004, Baxter Healthcare Corp. licensed exclusive U.S. marketing and distribution rights for rHuPH20 for ENHANZE SC (spreading indications:, not in combination with other agents) in the U.S. (upon FDA approval). Baxter and Halozyme would be equal partners in commercialization of products. Baxter also received right of first refusal for additional developmental products and territories. Baxter also provides fill and finish (packaging and labeling) services. In Aug. 2005, Baxter exercized its option to market, sell, and distribute Hylenex in the European Union.

In Feb. 2005, Halozyme signed an agreement with Avid Bioservices, Inc., a subsidiary of Peregrine Pharmaceuticals Inc., for contract cGMP manufacture of rHuPH20. In Dec. 2006, this agreement was extended to commercial supplies of rHuPH20. Avid/ Peregrine could manufacture up to 20 runs/year of rHuPH20 over the next five years.

In Dec. 2006, Halozyme entered into an agreement with Hoffmann-La Roche to apply Enhanze/rHuPH20 to unspecified Roche biopharmaceuticals to improve their subcutaneous delivery. Roche paid Halozyme $20 million as an initial upfront payment for use of rHuPH20 with three pre-defined Roche biologic targets. Over the next ten years, Roche will have the option to exclusively develop and commercialize rHuPH20 with an additional ten targets. Roche may pay Halozyme further milestones which could potentially reach a value of up to $111 million as well as royalties on potential product sales for the first three targets. For each of the additional ten targets, Roche may pay Halozyme further upfront and milestone payments of up to $47 million per target. The Roche Venture Fund made an $11 million equity investment, representing ~5% of Halozyme’s outstanding common stock. Roche also obtained access to Halozyme’s expertise in developing and applying rHuPH20 to Roche targets. Roche will have a worldwide, exclusive license to develop and commercialize product combinations of rHuPH20 and Roche target compounds resulting from the collaboration.

In Feb. 2007, Baxter expanded its licensing agreement with Halozyme to include the use of Hylenex with unspecified recombinant and small molecule drug therapeutics. Baxter paid an initial upfront payment of $10 million, made a $20 million equity investment, and will make milestone payments. Halozyme will receive royalties on Hylenex as a standalone product and on any kits and co-formulations of Hylenex with Baxter’s or other small molecule drugs. The agreement does not include combinations of HYLENEX recombinant with cytostatic and cytotoxic chemotherapeutic agents, the rights to which have been retained by Halozyme. In addition, Baxter also assumed all development, manufacturing, clinical, regulatory, sales and marketing costs. An expanded launch of Hylenex was planned for later in 2007.

In Nov. 2009, Halozyme entered into a long-term agreement with ProMetic Life Sciences Inc. for supply of the proprietary synthetic ligand affinity adsorbent manufactured by ProMetic's UK subsidiary, ProMetic BioSciences Ltd., used for affinity purification of rHuPH20. The agreement is for an initial five-year term with the opportunity for future extension. ProMetic's synthetic ligand adsorbent was used by Halozyme throughout the development of rHuPH20 and for manufacture of this FDA-approved product.

In April 2010, Halozyme concluded two commercial supply agreements with its existing manufacturing partners. Halozyme amended its existing commercial supply agreement with Avid Bioservices, Inc., entered into a new commercial supply agreement with Avid and entered into a new commercial supply agreement with Cook Pharmica LLC. These three agreements werre expected to cover the commercial production of r(rHuPH20) to be used in the manufacture of both current products as well as certain partnered product candidates, such as subcutaneous Herceptin and GAMMAGARD, with rHuPH20. Subcutaneous Herceptin and GAMMAGARD with rHuPH20 were both then in Phase III trials.

Halozyme appears to have been rather quickly and inexpensively developed. Halozyme reports spending just $4.6 million for R&D from inception in 1998 to mid-2004, with ~90% of this for Cumulase and Enhanze SC. At this rate, Enhanze may have set a record for speed and low expenses for development of a new biopharmaceutical. As noted by the company chairman, “Filing a first NDA is a major milestone for any company, but filing one merely 21 months after vector construction, as Halozyme has achieved, is quite rare.”

Manufacture: As described in U.S. 5,721,348, which may only partially reflect large-scale manufacture, a full-length clone for PH-20 is subcloned into an E. coli expression vector, pMAL-c. PH-20 is expressed as a fusion protein, the N-terminal fusion partner being the maltose binding (MBP) protein of E. coli. The PH-20-MPB fusion protein is found in the cytoplasm and does not form disulfides. Using pMAL-c carrying strains of E. coli, expression of PH-20 is high (the fusion protein is the major band in an E. coli extract on a Coomassie blue-stained SDS-PAGE gel). To purify the human PH-20 fusion protein, the MBP-PH-20 fusion protein was bound to an amylose resin (to which MBP binds) and eluted with maltose.

As part of its manufacturing process, Hylenex is purified using proprietary synthetic-ligand affinity chromatography matrix manufactured by ProMetic BioSciences Ltd. based on the company’s Mimetic Ligand technology.

FDA class: Medical device for Cumulase, 510(k) filing; Biologic for Enhanze SC, BLA filing

Approvals: Date = 20050419; Cumulase 510(k) approval (equivalent to one or more currently-approved medical devices, e.g., Amphadase and Wydase); Indication = for the treatment of oocytes to facilitate certain in vitro fertilization (IVF) procedures

Date = 200501205; Hylenex original NDA approved under 505(b)2 generic drug regulations; Indications: = see below

Date = 20050400; 510(k) approval for SynVitro Cumulase; Indication = for the treatment of oocytes to facilitate certain IVF procedures

Indications: [full text of the Hylenex "INDICATIONS AND USAGE” section of product insert/labeling]:

Hylenex recombinant is indicated as as adjuvant to increase the absorption and dispersion of other injected drugs; for hypodermoclysis; and as an adjunct in subcutaneous urography for improving resorption of radiopaque agents.

[unofficial indications:, for Cumulase]

for the preparation of oocytes prior to in vitro fertilization (IVF) during the process of intracytoplasmic sperm injection (ICSI)

Status: A 510(k) application for Cumulase to facilitate certain in vitro fertilization (IVF) procedures was filed on Sept. 14, 2004. It was approved on April 19, 2005. U.S. launch was on June 20, 2005.

Cumulase received CE Mark (medical device) approval in the European Union on Dec. 28, 2004 for the treatment of oocytes to facilitate certain in vitro fertilization (IVF) procedures. Cumulase was launched in European Union countries in early 2005.

A New Drug Application (NDA) seeking approval under 505(b)(2) generic drug regulations for Enhanze SC/Hylenex was filed on March 23, 2005. Hylenex received priority review (PDUFA/target day Sept. 23, 2005), and was designated a new chemical entity (NCE), indicating that any marketing exclusivity granted to other hyaluronidases (Amphadase and Vitrase) do not apply to Hylenex, and that approval of Hylenex does not include designation of therapeutic equivalence to any other approved hyaluronidase product. Thus, unlike most generic drugs, which receive approvals including recognition of therapeutic substitutability with one or more comparable drugs, with pharmacists allowed to substitute generics and often little or no marketing needed, Baxter will have to market Hylenex to physicians inorder to gain market share.

NDA Approval of Hylenex as a New Chemical Entity (NCE) under 505(b)2 was granted on Dec. 5, 2005 (approval time = ~8.5 months). However, as a NCE, Hylenex was not rated as therapeutically equivalent to other hyaluronidase products (which are all animal-derived). NCE status gives Hylenex five years of market exclusivity in the U.S., i.e., FDA will not approval an equivalent product for five years. However, this means that unlike most other generic drug approvals, Hylenex is not considered equivalent and substitutable for other hyaluronidase products in the filling of prescriptions. Thus, it will have to be marketed as a branded rather than generic drug. However, this NCE-based exclusivity has little meaning in actually, since the currently-approved animal-derived hyaluronidase products also were approved as NCEs and similarly received so-called five-year exclusivity, any other new animal-derived products will also likely receive NCE status, and any new recombinant products during this period will be effectively blocked by Halozyme’s patents. Hylenex was launched in the U.S. on June 27, 2006.

In many respects, the FDA approval of Hylenex under 505(b)(2) generic drug regulations was similar to biosimilar, biocomparable, follow-on protein (FOP), etc. approvals, with approvals based on abbreviated trials showing comparability and bioequivalence to a comparator/reference product, rather than full Phase III safety and efficacy trials. To many, FDA’s appproval of Hylenex based on a much abbreviated appliction shows that the agency effectively does not need new regulations to approve what are essentially biogeneric products based on abbreviated applications. For example, the Hylenex NDA was primarily supported by a safety study in 100 patients and preclinical data, with FDA requiring no efficacy or Phase III studies. See the Trials section for further discussio of clinical trials. FDA’s approval of Hylenex strengthened the arguments of biogeneric advocates that FDA has both legal authority and precedents for approving recombinant therapeutic proteins based on far less data than are required for conventional NDAs or BLAs. However, because Hylenex was given NCE status, its approval gave no indication of how FDA will consider biogenerics to be equivalent to innovator/reference products (allowing substitution when filling prescriptions), as is the case with most generic drugs, including others approved under 505(b)(2). FDA’s recent 505(b)(2) biogeneric-like approvals, including other hyaluronidase products and recombinant calcitonin and glucagon, which all received NCE status, suggest FDA will continue to consider such products to be NCEs (at least, until it implements new biogeneric/follow-on protein regulations).

rHuPH20 has characteristics that many opponents of development of formal biogeneric regulations consider to be substantial hurdles or even barring formal biogeneric-type approvals, including it being a large complex molecule (about double the size of erythropoietin/EPO), and the molecule is glycosylated or having variable carbohydrate polymer chains attached at often varying amino acids of the glycoproteins sequence. In fact, Baxter, which now markets Hylenex, had submitted a citizen’s petition to FDA in 2000 after Wyeth halted manufacture of Wydase (see related entry) supporting the view of innovator/established companies regarding biogenerics. Baxter specifically argued that approval of follow-on products to Wydase must be based on full safety and efficacy trials. FDA responded in May 2004 that a 505(b)(2) application “may rely for approval on investigations that were not conducted by or for the 505(b)(2) applicant and to which the applicant has not obtained a right of reference.” Although the animal-derived hyaluronidase products have been and continue to not be well characterized, a U.S. Pharmacopeia assay “is a valid surrogate for enzymatic activity and thus may be used to establish the effectiveness of new hyaluronidase products.”

In May 2006, MediCult A/S received 510(k) approval from (FDA) for SynVitro Cumulase for the treatment of oocytes to facilitate certain in vitro fertilization (IVF) procedures. The media-formulated SynVitro Cumulase product contains the same active recombinant enzyme used in Cumulase(R)-10X. SynVitro Cumulase is a formulation of rHuPH20 prepared in MediCult’s proprietary SynVitro media, and is ready to use in ex vivo IVF procedures.

On April 27, 2007, Baxter formally launched Hylenex in the U.S. for the ophthalmology market for use in enhancing the absorption and dispersion of other injected drugs.

In Oct. 2009, Baxter launched Hylenex in the U.S. for use in pediatric rehydration.

In May/June 2010, Baxter International Inc reported "manufacturing failures" in making Hylenex. On May 17, 2010, Halozyme announced the voluntary withdrawal of affected lots of 150U Hylenex from distribution after Halozyme and Baxter confirmed the presence of glasss particles in a limited number of vials. The company estimates that there were approximately 3,500 vials in the marketplace.

In Aug. 2010, Halozyme announced the completion of its root cause investigation regarding HYLENEX manufacturing and identified a corrective action plan and regulatory strategy to reintroduce Hylenex to the market. Baxter's senior management reiterated its commitment to the successful commercialization of Hylenex. As a result of this progress, Halozyme has lifted the notice of breach (or contracty) that was delivered to Baxter on May 16, 2010.

In Aug. 2012, FDA delayed (refused) approval of HyQ, Halozyme's hyaluronidase in combination with immunoglobulin from Baxter for treatment for primary immunodeficiency. FDA requested additional preclinical data, including more animal studies regarding non-neutralizing antibodies, reproduction, fertility, and fetal development. This was expected to significantly delay approval. Also, questions were raised regardng patients developing antibodies to rHuPH20.

Tech. transfer: U.S. 5,721,348, “DNA encoding PH-20 proteins,” DNA encoding PH-20 proteins, Feb. 24, 1998, assigned to the University of Connecticut, has been exclusively licensed by Halozyme Therapeutics. This patent concerns PH-20 DNA isolated from mammalian (including human) sperm, cloned DNA, and recombinant constructs for expression of PH-20 hyaluronidase enzymes. This patent is primarily directed towards use of PH-20 as an immunogen for contreptive (sperm-attacking) vaccines. Mammalian PH-20 protein and the discovery that the DNA encoding PH-20 in one mammalian species is cross-reactive (i.e., hybridizable) with genomic DNA from all other mammals tested.

Halozyme has reported that it may become involved in interference proceedings in the U.S. to determine the priority of its licensed patent(s). U.S. 6,123,938, “Human urinary hyaluronidase,” assigned to the University of California, expiring Feb 24, 2015, concerns purification and cloning of the major hyaluronidase activity of human plasma, termed HYAL1 (also known as hpHAse), with 40% homology to PH-20. Incyte Pharmaceuticals, Inc. (Palo Alto, CA) has received patents covering recombinant hyaluronidases with homology to PH-20 including 6,057,110; 5,958,750; and 5,854,046; each entitled, “Human hyaluronidase.”

In Aug. 2010, Halozyme received U.S 7,767,429, "Soluble hyaluronidase glycoprotein (sHASEGP), process for preparing the same, uses and pharmaceutical compositions comprising thereof," including claims for its recombinant purified hyaluronidase glycoprotein (rHuPH20). In the U.S., the issued patent will expire in March 5, 2024, but the term may be extended until Sept. 23, 2027 if the addition of patent term adjustment time of 1,297 days is approved by the PTO. Patents with similar claims had been granted in India, Japan, Eurasia, Hong Kong, Mexico, New Zealand, Singapore and South Africa.

Halozyme has received EP 1603541 B1, SOLUBLE HYALURONIDASE GLYCOPROTEIN (sHASEGP), PROCESS FOR PREPARING THE SAME, USES AND PHARMACEUTICAL COMPOSITIONS COMPRISING THEREOF, expiring in 2024, including Halozyme noting this.

In Sept. 2007, Baxter entered into an agreement with Halozyme potentially worth $47 million to use Enhanze technology to help develop a subcutaneous formulation of immune globulin (Gammagard Liquid 10%).

Trials: In June 2005, results were reported from an IVF post-marketing study involving 26 patients and 402 oocytes. The rate of fertilization following intracytoplasmic sperm injection (ICSI) in eggs treated with Cumulase was 81.4% (162 fertilized eggs out of 199) vs. 71.9% (146 fertilized eggs out of 203) in eggs treated with the traditional bovine-derived extract, a 13% increase in fertilization rate, and above the average ICSI fertility rates in U.S. fertility clinics, currently 70-75%.

In Oct. 2004, investigators from Stanford University reported results from a study comparing Cumulase and current animal-derived hyaluronidase extracts in certain IVF procedures. After extended exposure to Cumulase, murine oocytes (eggs) had fertilization rates similar to natural IVF, but in the same time period, oocytes exposed to bovine hyaluronidase experienced ~50% decrease in viability relative to natural IVF or Cumulase-treated oocytes. The purity of Cumulase was more than 100-fold higher than the animal preparations.

In Oct. 2005, Halozyme initiated its first Phase I trial, an open-label study, in about 5-10 patients with Chemophase to enhance the delivery of cancer chemotherapy. This evaluated a single intravesical (into the bladder) administration of Chemophase along with the widely used anticancer drug mitomycin in patients with superficial bladder cancer. Co-delivery of Chemophase is expected to increase the penetration of mitomycin throughout tumors, allowing the drug to better reach residual tumor cells that otherwise might develop into recurrent tumors.

The NDA under 505(b)(2) was approved based on a single double-blinded trial in 100 healthy human volunteers comparing Hylenex to a saline control that showed the absence of allergic reactions and significantly reduced injection site discomfort compared to the saline control. Thus, FDA’s approval of Hylenex is similar to recent approvals of animal-derived hyaluronidase products, with each approval as an NCE based on preclinical data and a single Phase I trial designed to rule out an incidence of hypersensitivity reactions of greater than 10%. Subjects in the trial were injected intradermally with Hylenex in one forearm and saline control in the other forearm, and evaluated for allergic responses and injection site side effects. None of the 100 subjects had evidence of allergenicity. With regard to side effects, 28 subjects experienced discomfort (e.g., stinging, burning, other discomfort) at the site of saline injection, while significantly fewer (only 3) experienced such side effects at the site of injection. The lack of allergic response was important, since the most frequently reported adverse experiences with currently available, animal-derived hyaluronidase products are local injection site reactions. The significant reduction in injection site discomfort is consistent with the rapid-onset mechanism of action of rHuPH2.

In Feb. 2006, Halozyme and Baxter reported results from the pivotal Phase IIIb INFUSE-LR (INcreased Flow Utilizing Subcutaneously-Enabled Lactated Ringer’s) trial of subcutaneous (sub-Q) infusion for subcutaneous fluid replacement (hypodermoclysis) with Hylenex. This involves the introduction of fluids under the skin to replace inadequate intake or excessive loss of water and electrolytes during illness or operation, and pertains to Hylenex use for hydration. The trial studied Hylenex with subcutaneous infusions of Lactated Ringer’s solution using gravity flow without an infusion pump. INFUSE-LR was designed to determine the subcutaneous infusion flow rate of Lactated Ringer’s (LR) solution with and without Hylenex, the sub-Q infusion flow rate dose response to Hylenex over one order of magnitude of dose, and assess safety and tolerability. This prospective, double-blind, randomized, placebo-controlled, within-subject, dose-comparison study enrolled 54 volunteer subjects who received Sub-Q infusions simultaneously in both upper arms through 24 gauge catheters. The use of Hylenex preceding sub-Q infusion accelerated the flow rate by approximately four-fold versus the Sub-Q infusion with placebo, while causing less edema and was preferred by both investigator (for 92% of subjects) and study subjects (92%). Hylenex accelerated flow versus placebo in every subject studied, and by an overall mean ratio of approximately four-fold. The overall mean flow rate for sub-Q infusion with Hylenex was 464 mL/hr versus 118 mL/hr with placebo (p < 0.0001) The faster flow rates did not result in an increase in edema. The sub-Q infusion rate, when preceded by Hylenex, was closer in flow to the IV infusion (ratio of 2.6) than to the Sub-Q infusion rate with placebo (ratio of 2.9). There were no serious or severe adverse events (AE), and Hylenex was well tolerated. Doses of 750 and 1,500 unit doses produced faster flow rates than a 150 unit dose, but the study was not statistically powered to detect a dose response. This study demonstrated that Hylenex with sub-Q infusions of Lactated Ringer’s solution using gravity flow without an infusion pump achieves substantial flow rates with acceptable tolerability. In Dec. 2005, Halozyme initiated a Phase IV (post-approval) INFUSE-LR trial.

Results from the INFUSE-LR trial were published in in the Dec. 2008 issue of the Journal of Palliative Medicine (19(6): 1312-1320).

In May 2006, Halozyme published a clinical study in Fertility & Sterility showing increased fertilization rates in oocytes (eggs) that were treated with Cumulase(R) prior to IVF using the ICSI procedure. In this prospective, randomized, blinded, sibling oocyte trial, the per-patient rate of fertilization following ICSI in mature eggs treated with Cumulase was 85.3% versus 70.0% for eggs treated with a traditional bovine-derived extract enzyme, a 22% increase in fertilization rate (p<0.05). A trend was observed toward higher quality embryos with Cumulase (43% of excellent quality) compared to the bovine-derived extract product (25%). A subsequent, randomized, blinded, parallel group, multicenter trial is currently underway to further characterize the benefits of Cumulase, including an evaluation of longer-term clinical outcomes.

In Jan. 2007, Halozyme reported improved absorption and bioavailability of an unspecified commercially-available recombinant protein therapeutic using Enhanze/rHuPH20. The open-label, dose escalation, within-patient controlled study used escalating dose cohorts of rHuPH20 (1,600-12,800 U) subcutaneous (SC) injection of the therapeutic with one SC injection of the therapeutic agent combined with Enhanze. The study compared the bioavailability and other PK parameters, along with safety and tolerability, of the two SC injections, one with and one without Enhanze. For the primary endpoint of area under the curve (AUC) for plasma concentration of the therapeutic, the AUC over the 14 days following injection was higher with rHuPH20 compared to without rHuPH20 for 100% (15/15) of the patients. Addition of rHuPH20 increased the AUC above baseline (average trough level) of the therapeutic for all cohorts combined by an average of 58%. The company reported, “The findings from this first clinical trial of Enhanze Technology provide proof of concept that rHuPH20 may also have clinical application for large molecule therapeutics.”

In Feb. 2007, Halozyme and Baxter reported positive results from the INFUSE-Morphine (INcreased Flow Utilizing Subcutaneously-Enabled Morphine) trial of Hylenex with morphine. This Phase IIIB trial showed that subcutaneous administration of morphine with Hylenex accelerated the time to maximal blood levels of morphine by 33% versus morphine with placebo, and appeared safe and well-tolerated. Morphine is a widely used drug for pain management and is currently approved for both intravenous and subcutaneous administration. The study indicated that clinical effects, such as analgesia, could be achieved more rapidly by subcutaneous injection, without the need for intravenous infusion. This double-blind, randomized, crossover, placebo-controlled trial determined the time to maximal blood levels of morphine after subcutaneous administration with and without Hylenex compared to determine the time to maximal blood levels after intravenous administration of morphine, and to assess safety and tolerability. Average time to maximal plasma concentration (Tmax) of morphine was reduced from 13.8 minutes when injected subcutaneously with the saline placebo to 9.2 minutes when injected with Hylenex, a 33% reduction in the time to maximal plasma concentration (p<0.05). The most common adverse events were mild injection site redness, rash, swelling, and itching. No Hylenex toxicity was apparent. These results suggest that SC morphine plus Hylenex provides pharmacokinetic characteristics that are superior to SC morphine alone and closer to intravenous morphine.

In April 2007, following the successful completion of an initial U.S. Phase I trial of single-administration of Chemophase and mitomycin, a Phase I/IIa dose-ranging trial began with Chemophase to enhance the delivery of chemotherapy drugs. In the Phase I trial, pharmacokinetic data were obtained from the plasma of five bladder cancer patients dosed intravesically (instillation into the bladder) with 20,000 units of Chemophase along with 40 mg of mitomycin. The observed plasma levels of mitomycin were less than 1/40th of the levels reported to be predictive of suppression of white blood cell production. The new trial involves multiple intravesical administrations of Chemophase along with mitomycin in patients with superficial bladder cancer. This Phase I/IIa study is enrolling up to 36 evaluable patients with superficial bladder cancer to determine the maximum tolerated dose and dose-limiting toxicities, if any, of escalating doses of Chemophase in combination with mitomycin administered as weekly intravesical instillations for five weeks according to the usual standard of care; will establish the optimal dose of Chemophase with mitomycin for future studies; and will observe patients for any preliminary evidence of anti-tumor activity.

In Jan. 2009, Baxter International and Halozyme started a Phase III clinical trial of Baxter's Gammagard Liquid [Immune Globulin Intravenous] 10% (IGIV; see related entry) with rHuPH20 for the treatment of primary immunodeficiency (PID). The purpose of this trial is to evaluate the safety and efficacy and gain regulatory approval for the treatment of PID using IVIG and rHuPH20 via subcutaneous injection at a single site. Gammagard Liquid is currently administered intravenously (IV). Subcutaneous (SC) administration could allow patients to receive a full monthly dose of IVIG in a single injection site in their home setting.

Medical: For increased absorption and dispersion of injected therapeutics, 50-300 U, most typically 150 U hyaluronidase, is added to the injection solution.

For hypodermoclysis, the needle for fluid injection is inserted ith tip lying free and movable between skin and muscle, clysis (fluid administration) is started, and then Hylenex is injected into rubber tubing close to needle. An alternate method is to inject Hylenex under skin prior to clysis. Typically, 150 U will facilitate absorption of 1,000 mL or more of solution. Hylenex may also be added to small volumes of solution (up to 200 mL), such as small clysis for infants or solutions of drugs for subcutaneous injection. For infants and children less than 3 years old, the volume of a single clysis.

For subcutaneous urography urographic contrast media is injected by the subcutaneous route when intravenous administration cannot be successfully accomplished, particularly in infants and small children. With the patient prone, 75 U of HYLENEX recombinant is injected subcutaneously over each scapula, followed by injection of the contrast medium at the same sites.

Disease: There were an estimated early 760,000 total IVF cycles worldwide in 2004. The potential market for Cumulase consists of an estimated 500,000 or more intracytoplasmic sperm injection (ICSI) procedures worldwide in 2004. About 90,000 ICSI procedures are performed annually in the U.S., with this growing at ~11% annually. About 400,000 ICSI procedures were completed in 2000 outside the U.S., also growing at 11% annually. Halozyme has reported, “The total Cumulase market consists of an estimated 500,000 ICSI cycles worldwide in 2005, with nearly 90,000 of those performed in the U.S.”

According to data from the American Cancer Society, National Cancer Institute, American Urological Association, and Southwest Oncology Group Study, more than 100,000 patients have new or recurrent superficial bladder cancer in the U.S. every year. All of these patients may be potential candidates for Chemophase in the event it is approved as first-line treatment with mitomycin.

Market: The 2007 Average Wholesale Price for Hylenex is $273/60/vial (Red Book,.2007).

Halozyme sales for the year ending Dec. 31, 2005, with products still being launched, were negligible, with company total 2005 sales revenue of $127,209.

Analysts with Brean Murray & Co. have projected the market for Hylenex (for its currently approved indications:) to be $10-20 million/year     

Halozyme has estimated the worldwide market for hyaluronidase in IVF procedured to be about $10 million. There are about 400 in vitro fertilization clinics in the U.S., and about 1,000 in the European Union. In May 2006, Halozyme reported, “The total Cumulase market consists of an estimated 500,000 ICSI cycles worldwide in 2005, with nearly 90,000 of those performed in the U.S.” In Oct. 2006, Halozyme reported, “Fifty-seven percent of IVF procedures performed in the U.S. are ICSI or include ICSI.”

The potential market for Hylenex includes the ~6.4 million local eye anesthesia procedures (or 45% of the 14.2 million total estimated cataract surgery procedures) worldwide in 2004. Hylenex will replace natural hyaluronidase products for many of these procedures.

Halozyme has estimated the market for Hylenex use in subcutaneous infusions is $200 million to $300 million.

As part of marketing Cumulase, Halozyme plans to “raise public awareness of the current risk of using animal-derived products in IVF applications among industry professionals and the general public through direct contact with target audience.”

rHuPH20 could attain blockbuster (sales >$1 billion/year), if it is adopted for other uses, e.g., for adjuvant treatment of solid tumors and broad drug delivery applications.

Competition: Recombinant hyaluronidase competes with ovine and bovine hyaluronidase products. See related entries in the Enzymes section.

Ongoing: In May 2005, preclinical safety and pharmacology animal studies with rHuPH20 showed that when hyaluronic acid products (see #925 and related product entries) commonly used in cataract surgery were left in the eyes of animals, injection of rHuPH20 enzyme into the front of the eye significantly reduced the incidence and severity of intraocular pressure (IOP) rises. Such pressure “spikes” occur when viscoelastic materials used in the surgical procedure clog up the eye’s drainage canals and prevent fluid from properly draining out, a process that can potentially result in permanently dilated pupils, persistent glare, pain, and discomfort, and retinal and optic nerve vascular occlusion. Injection of rHuPH20 into the front of the eye caused no toxicity to the corneal endothelial cells in the eye. About 13 million cataract procedures are performed worldwide every year. Presumably, Halozyme/Baxter will conduct clinical trials for this indication, and/or ophthamologists may use rHuPH20 off-label for this indication (particularly, once Enhanze SC is approved for ophthalmic use).

R&D: In Sept. 2005, Halozyme reported positive results from a study with pegylated rHuPH20 (i.e., polyethylene glycol polymer strand(s) attached to the molecule) in animal models of ischemic stroke. As with other pegylated proteins, rHuPH20-PEG had a dramatically longer serum half-life compared with the unmodified form, and both rHuPH20 and rHuPH20-PEG showed increased survival in stroke models. Survival at 28 days postinfarction in the same rat MCAO model improved to 69% with rHuPH20 and to 78% with rHuPH20-PEG. The enzymes appeared to prevent death in treated animals by reducing edema (or swelling) within the brain. Brain edema can occur after severe strokes when large regions of the brain are deprived of blood flow. This results in increased edema, similar to that may result from cardiovascular disease, tumors and organ transplantation.