Central to Neugenesis' work with Neurospora crassa has been the development of a versatile, cost effective fungal platform for protein expression. This proprietary platform has been chosen by multiple clients as the vehicle for the production of monoclonal antibodies, enzymes, and virus like particles.
Neugenesis has developed a solid technical and intellectual property position around Neurospora crassa for use as a host for production of biopharmaceuticals. This host is a very safe organism (no toxic secondary metabolites) and has long been eaten as a food source in some tropical countries. It has been used academically as a model system for over seven decades and a plethora of genetic, biochemical and molecular biological tools are available for its use including a fully sequenced and annotated genome, and a functional genomics project with individual strains available carrying knock-outs of virtually all of the genes of the fungus.
Production of biopharmaceuticals using this system has a very low cost. Neurospora, in addition to being highly productive, grows very rapidly on simple, defined media derived from inexpensive, non-animal sources. As a secretory system, capital requirements for purification, often the largest contributor to COGS, is significantly reduced.
The NeuBIOS™ platform is safer than bacterial production systems, since there is no endogenous source of endotoxin. Neurospora does not use viruses, and produces no toxins or adventitious agents eliminating the need for inactivation steps in the production process, thus reducing the costs of manufacture and increasing the potency of the final product. Related filamentous fungi are used to produce an enormous number of products for direct human use, e.g. virtually all the citric acid used for food and soft-drink manufacture worldwide, as well as many other protein products used in food and wine manufacture. In addition, a wide variety of proteins like lipases, cellulases and proteases are used in washing powders and clothing manufacture as well as in cosmetics for direct human use. Most importantly, fungal production (via fermentation) is responsible for approximately half of all pharmaceutical and biotech products on the market. Because of the ubiquitous nature of these products, much work is, and has been put into identifying potential toxins, allergens and immunogens for removal by genetic or process development means. We currently are collaborating with Dr. Jay Slater, of CBER at the US FDA, for examination of Neurospora production broths to identify allergens.
Synergistic Technologies for Product Development
The company has developed additional technology that adds value to the production platform, including:
- Unique high-throughput screening methodology.
- Novel approaches to gene diversification for product improvement.
- Combinatorial biology technology that allows simplified production of multimeric protein complexes (such as monoclonal antibodies) as well as multivalent vaccine antigens and virus-like particle (VLP) preparations.
Recombinant Vaccine Products
The human vaccine market outlook has improved significantly over the past decade, with the rejuvenation of the industry by new Federal liability exemptions, recommendations for expanded recipient populations, and a new wave of approved recombinant vaccines. Recombinant vaccine antigens provide significant improvement in terms of product characterization and safety compared to traditional attenuated or inactivated vaccine approaches. Newer production approaches also provide significantly reduced cost of goods compared to traditional expression systems. Fungal production systems such as yeast provide advantages similar to bacterial production systems, but in addition allow larger proteins to be produced, including native, correctly folded eukaryotic protein antigens important for the efficacy of many vaccines. Recombinant yeast-produced vaccine products now on the market include Engerix®-B (GSK), Recombivax HB® and Gardasil™ (Merck). A logical next step is to use the NeuBIOS™ platform for these biopharmaceuticals to exploit the dramatically increased productivity of filamentous fungi in comparison to the single-celled yeast.
Soluble subunit antigens can be purified by standard chromatography techniques, dependent upon the biochemical characteristics of the antigen. For most human biologics, a histidine tag is not recommended for regulatory reasons. For veterinary applications, however, a tagged approach may be appropriate. In general, the complexity of the clarified broth feedstock is significantly lower in secretory systems such as NeuBIOS™ vs. platforms that require cell lysis, like most bacterial or plant systems. This simplifies the purification and reduces overall costs. Products are also more soluble and refolding is rarely necessary, improving yields.
Neugenesis is currently contracted by PATH, a non-profit dedicated to healthcare in the developing world, to create VLP-based influenza vaccine candidates for in vivo testing. After this first-year proof of concept project, a second stage contract in 2010 will provide support to Neugenesis for the preclinical development and early-stage manufacturing to take these candidate vaccines to Phase I clinical trials in 2011. Additional opportunities to develop other vaccines with PATH are possible when the influenza program demonstrates proof of concept in Q2 2010.
The NeuBIOS™ expression system's advantages over traditional vaccine production methods fall into three categories: yield, development time, and cost per dose. Of the three, increased yield is the most significant, as can be seen below.
| Numbers based on production of monovalent bulk and 15 ug HA dose.
*MMC information from the Novartis website. **NeuBIOS™ calculations use a conservative yield estimate for the system | |||
| Production Method | HA mg/L Final Yield |
Vaccine Doses/L |
L to generate 10 MM doses |
|---|---|---|---|
| Eggs | 7 | 470 | 21,277 |
| Mammalian Cell Culture * | 2.7 | 180 | 55,000 |
| NeuBIOS™ ** | 100 | 6,667 | 1,500 |
The veterinary vaccine market is even more cost sensitive than the market for human influenza vaccines. The potential market, however, is both large and varied with many unmet needs. Neugenesis is exploring veterinary opportunities, building off our human recombinant vaccine development program.
Recombinant Monoclonal Antibody Products
The Neugenesis approach to production of monoclonal antibodies offers some unique advantages. Our platform offers the ability to fold complex mammalian proteins correctly, to glycosylate proteins, and to secrete them into inexpensive media. The system does not express toxins and has no known viruses. Our proprietary intellectual property (IP) position is free from dominating or competing IP. The system is readily scalable and requires significantly less infrastructure than traditional methods.
Neugenesis is also under contract with DARPA to develop low cost, rapid response manufacture of biotherapeutic antibodies for the Department of Defense. The company was awarded a multi-million dollar Stage I contract by the Defense Advanced Research Projects Agency (DARPA) in July of 2007 to demonstrate the platform's capabilities in biodefense and the management of pandemic outbreaks. Successful completion of this first phase has led to the current Phase II contract to quickly produce monoclonal antibodies at commercial scale.
Under this program, the company is focused on the development of a unique, broad spectrum anti-viral monoclonal antibody with a potentially large market. This product is of particular value in that it is very poorly expressed in standard mammalian and insect cell culture systems.
