Medical Science: Novel Therapeutic Platforms

by Mihai Mihut, MD

"Use of transgenic animals that produce complex human proteins for prevention
or treatment of serious health hazards will save lives and improve quality of life
for millions around the globe."

Use of transgenic animals that produce complex human proteins for prevention or treatment of serious health hazards will save lives and improve quality of life for millions around the globe.

While there is a general understanding of the pathophysiological aspects of diseases, the therapeutic act is often limited by a lack of available treatments or by a prohibitive cost incurred in order to obtain those medicinal substances. The difficulty in obtaining active pharmaceutical ingredients (APIs) resides for example in a complex structure that cannot be synthesized chemically, or cannot be correctly reproduced in early biotechnological systems such as transgenic bacteria.

A representative case is treating patients who suffered exposure to nerve gas agents. These substances (Soman, Sarin, Tabun, VX, etc.) are considered chemical warfare (Sarin was used in the Tokyo subway attack in 1995); relatively small quantities of these volatile gases may have lethal effects on scores of people, in a very short time. While there are some symptomatic and pathogenic treatments available, these can only be administered after the fact, during a very short time window; if the exposure was severe, even with the best treatment the patient will likely be lost. Therefore, the therapeutic arsenal is missing an important component: an etiological treatment. We will discuss in the following chapters the path that was followed that would soon allow a novel medicine (Protexia™) to be made available to those in need.

The human body produces its own medicine

Human butyrylcholinesterase (BChE) is a naturally occurring enzyme existing in blood in very small amounts (2 mcg/mL). BChE is a natural bioscavenger. As such, it exerts both a biochemical and a physicochemical effect, through binding and then degrading the nerve agents while still in the bloodstream. At physiological concentrations it does not offer adequate protection against exposure, but large quantities injected into the body (as proven in test animals) offer protection and reverse the effect of nerve gas agents.  

When the solution looks like a goat

The main problem the scientists faced was the source of API for large-scale manufacturing of a medicine. The existing method filters human blood to extract this scarce enzyme. In order to achieve a therapeutic dose of 500 mg BChE, at least 500 L of blood must be processed. The quantity of blood required to obtain enough medication in case of a terrorist attack over a medium size city, where for example 100,000 people would need treatment, the quantity of blood to be filtered approaches 50,000,000 L. As the production process would take a very long time and the stability of the molecule would not be infinite, it is easy to understand the need for a more robust production process (more reliable source of API).

The specialists looked at alternative methods to manufacture this substance. The one that received special attention was in-vitro production by transgenic bacteria, a method used in producing other human proteins. After several attempts, major impediments blocked any further development into that direction. The amounts secreted by bacteria were insignificant (quantity issue) and the ¡°polishing¡± of this complex enzyme was not adequate (quality issue), due to significant differences in the protein synthesis apparatus between bacterial and human cells.

At that point in time a multi-disciplinary team based in Canada had the idea of producing human recombinant BChE (HuBChE) in a transgenic mammal. This was meant to address both the issues of quality (mammals have very similar protein synthesis pathways) and quantity (mammals lactate, so they could produce the protein in a very simple and natural way, by secreting it into their milk). Because of the shorter reproduction cycle and large volume of milk produced, the goat was selected as the most suitable animal to become a transgenic platform.

A multitude of scientists from different fields contributed to the development of this program. Geneticists, biologists, worked together to isolate the human gene, and transfer it into an oocyte. The gene had to be combined with another DNA fragment that would activate it in the mammary gland, and allow it to be secreted in milk.

Once that was done, the new cell had to be developed into a new goat, and copied in multiple organisms. This required intense and specialized work from embryologists, and novel techniques such as somatic cell nuclear transfer (cloning), in-vitro fertilization, artificial insemination. Veterinarians had to ensure the safety and well being of the goats, while also screening and treating them for any disease that could be transmitted to humans via the source material (milk goat).

Collaboration from medical doctors was required at every step to identify any potential health risks for human, due to unknown characteristics of the novel molecule, e.g. its immunogenicity. Since there was only one other such biological substance produced in transgenic animals at the same time, extensive discussions with regulatory agencies took place, to create the framework that would allow this technology to be applied, and medicines produced through it to be used in humans.

It became very important that the Quality personnel that were overseeing the whole process be well versed in human medicine, animal health, embryology and biochemistry.

Chemists, biologists, microbiologists, MDs and other specialists had to create valid assays to measure all the important characteristics of the protein. The purification process required qualified engineers and research personnel.

After all the efforts, the first results were visible: the milk produced by the transgenic goats contained sizable amounts of HurBChE, in amounts thousands of times higher than the concentration in blood.

The development is not over. It is currently being tested in healthy volunteers, to asses its safety. But one thing is sure, when this new novel product receives final approval: the source will be reliable, reproducible, available on large scale and therefore affordable.

All this required learning, science classes and collaboration between many different fields. In the end, it is the patient who will benefit. And the patient may be any of us, at some point.