Forbes Medi-Tech Granted Patent on Microbial Conversion of Phytosterols to Steroid Drugs
Overcomes difficult solubility problems of older steroid processes.
Forbes Medi-Tech Inc. (FMT; Vancouver, BC, Canada) has been awarded a patent on its process for using microbes to convert plant steroids to industrially important androstenedione (AD) and androstadienedione (ADD). The patent, US 6,071,714 (Process for the microbial conversion of phytosterols to androstenedione and androstadienedione), was issued on June 6 and listed James Kutney and Radka Milanova of FMT as its inventors, as well as three Bulgarian researchers.
AD and ADD are essential intermediates used to make a host of steroid-based pharmaceutical products such as oral contraceptives, anti-inflammatories, synthetic anabolic steroids, and estrogenic hormones. The combined worldwide market for AD and ADD, according to FMT, is approximately US $750 million annually, but that's just for raw materials. Products derived from these parent compounds are valued at nearly $4 billion per year.
Biosynthetic pathway from phytosterols (plant sterols; left panel) to pharmaceutical steroids (far right panel) through intermediates AD and ADD (middle panel). Plant steroids have the same structure as cholesterol, a 27-carbon molecule, except for an alkyl group (R) at C-24, which may be ethyl (sitosterol), ethylidene (fucosterol, isofucosterol), etc. Side chains may also contain a double bond between C-22 and C-23. Mycobacterium species oxidize the sidechain down to a ketone and also oxidize the A-ring to the ene-one (AD) or diene-one (ADD).
"This patent gives Forbes strong protection for its pharmaceutical steroid business, effectively safeguarding the Company's interest as we move to commercial production of AD and ADD," said Jack Miller, senior vice president and chief operating officer of FMT.
AD and ADD are commercially produced by fermentation of plant steroids—phytosterols. Raw materials used by Forbes are essentially a byproduct of the wood pulping process known as "tall oil soap." Other phytosterol sources include common vegetable oils such as soy, rapeseed, corn, cottonseed, sunflower, olive, linseed, and rice bran. Forbes' AD/ADD production process uses a mutant strain of Mycobacterium for fermentation process. Once the fermentation is complete, the AD or ADD material is extracted and purified.
That in itself would not be of tremendous interest. However, the new patent covers a series of highly effective solubilizing agents developed by Forbes that allow the process to work, in very high yield and with reduced fermentation time, on process streams containing high phytosterol concentrations.
"The microbial conversion process patented by Forbes will enable us to economically produce higher yields of superior quality AD and ADD on a commercial scale," said James Kutney, vice president of scientific research and development at Forbes.
The patented process is currently being optimized in pilot (5,000 liter) and commercial scale (50,000 liter) bioreactors for future commercial production of AD and ADD. Forbes believes that this novel technology will provide an attractive alternative to the production process currently in commercial use.
In addition to its steroid intermediate fermentation technology, Forbes has developed laboratory-level technologies for the production of higher value steroid compounds derived from AD and ADD, e.g. progestational, adrenocortical, estrogenic, and contraceptive agents, and is working on processes for additional products.
A Bit of History
The microbial conversion of phytosterols, typically from soybeans, by Mycobacterium species has been used for commercial production of steroids since the mid 1970's (see references). The process, although quite clean, is time-consuming due to steroid precursors' insolubility in water, which is where the transforming bacteria like to live.
Inadequate solubility means the process works from very low concentrations of substrate in the nutrient medium, resulting in poor contact with the micro-organism and generally leading to low yields.
Another problem associated with the known phytosterol bio-conversion process is that the bioconversion end-product consists of both AD and ADD, which are similar in structure and therefore difficult to separate. A further problem is the high cost of the media in which the Mycobacteria are grown. The patent addresses both concerns with one chemical modification.
By adding glycol (i.e. polypropylene glycol) or silicones to the broth, FMT is able to achieve very high concentration of steroid precursors in a clear solution. In the right proportions, FMT's proprietary glycerol or silicone formulation also promote proliferation of Mycobacterium, in this case M. MB 3683.
Thus, FMT achieves substrate concentrations of 30 g/l, vs. just 10 g/l using conventional fermentation media. Glycol solubilization results in preferential conversion of starting materials to AD, whereas silicones promote ADD production. Yields are in the 80% to 90% range.
References
- S. Kraychy, and R. D. Muir, U.S. Pat. No. 3,684,657 (1972). W. J. Marsheck, S. Kraychy and R. D. Muir, Appl. Microbiol., 23, 72 (1972).
- A. H. Conner, M. Nagaoka, J. W. Rowe and D. Perlman, Appl. and Environ. Microbiol., 32, 310 (1976).
- K. Kieslich, J. Basic Microbiol., 25, 461 (1985).
For more information: Martin Livingston, Director of Investor Relations, Forbes Medi-Tech, Suite 200 - 750 West Pender St., Vancouver, BC, Canada V6C 2T8. Tel: 604-681-8976.
By Angelo DePalma