Trichosanthin - The Ancient Remedy Reveals Additional Modern Uses

An Effective Abortion Drug

For centuries tien hua fen, the tuber of a humble climbing plant Trichosanthes kirilowii which grows wild in eastern China, has been used by the Chinese to induce abortion. According to the Compendium of Materia Medica compiled by Li Shi Zhen in the Ming dynasty, tien hua fen can reset menstruation and expel retained placentas. Chinese scientists conducting chemical and pharmacological analyses on the plant in the 1970s identified a protein to be responsible for inducing the abortion, and named it trichosanthin.


The climbing plant
Trichosanthes Kirilowii

In the late 70s, researchers from CUHK began to conduct research on trichosanthin and discovered that it can suppress the growth of certain tumour cells.

As a medicinal protein, trichosanthin is not without its shortcomings. Its curative effects are short-lived as it remains in the body only for a limited period of time. Besides, it defeats its own purpose by inducing the production of antibodies. It can also cause skin allergies, fever, muscle pains, and even the sudden lowering of blood pressure.

Research on Trichosanthin

Dr. P.C. Shaw of the Department of Biochemistry has since 1987 been working on the protein engineering of trichosanthin so as to improve its pharmacological performance and elucidate the role of constituent amino acids. His work is supported by a direct grant and three earmarked grants from the Research Grants Council (see table below). A related plant protein, a-momorcharin, with functions similar to trichosanthin, is the research subject of another team from the same department, headed by Prof. W.K.K. Ho.

Research projects on the functions and mechanisms of trichosanthin
YearProject titleAmount received
Investigators
1987Molecular Cloning of alpha-momorcharin HK$50,000Dr. P.C. Shaw,
and Prof. W.K.K. Ho
1989Protein Engineering of Trichosanthin and alpha-momorcharin: Molecular Cloning and ExpressionHK$610,000Dr. P.C. Shaw,
Prof. W.K.K. Ho,
and Dr. H.W. Yeung
1991Protein Engineering of Trichosanthin and alpha-momorcharin: Modification of Antigenic Determinants and
Structure-function Studies		HK$2,290,000Dr. P.C. Shaw,
Prof. W.K.K. Ho,
Dr. Y.S. Moon,
Dr. H.W. Yeung,
Prof. T.B. Ng,
and Dr. H.W. Leung*
1995Trichosanthin Derivatives with Reduced
Immunogenicity and Nephrotoxicity and Long Plasma Half-life 		HK$581,000Dr. P.C. Shaw,
and Prof. M.S.C. Tam
* Baptist University staff


Trichosanthin Production in the Laboratory -- An Important Breakthrough

Trichosanthin for research purposes used to be extracted from the tuber of Trichosanthes kirilowii. As the Hong Kong climate is not conducive to the growth of this plant, researchers often had to purchase it from mainland China. This technical problem is now solved by isolating the relevant DNA sequence from the tuber of the plant and inserting it into the bacteria Escherichia coli to enable the rapid production of the protein. The trichosanthin so derived has the same biological functions as that extracted from the plant. This successful attempt at 'synthesizing' trichosanthin made it possible for the protein to be produced in large quantities in laboratories in a purified form.

An Interdisciplinary Effort to Strengthen Curative Properties and Minimize Side Effects

In 1989 Dr. H.W. Yeung of the Biochemistry Department and his collaborators in the US found that trichosanthin can inhibit the reproduction of the AIDS virus in infected cells without affecting normal cells. Clinical trials also showed that the symptoms of AIDS in certain patients were alleviated by the protein. This discovery aroused great international interest and provided a fresh impetus for research on trichosanthin at The Chinese University.

As protein is made up of amino acids, an understanding of the latter's functions is crucial to the understanding of the protein's mechanisms. In order to learn the roles of different amino acids in the biochemical activities of trichosanthin and to identify those that trigger off antibody production, the researchers acquired a silicon graphic workstation with high processing power for manipulating large proteins in real time. Dr. Y.S. Moon of the CUHK Department of Computer Science and Engineering and Dr. H.W. Leung from Baptist University helped set up the workstation and input information.

Three dimensional structure of trichosanthin

Assistance was also obtained from Prof. Y.C. Dong of the Institute of Biophysics of the Academia Sinica in Beijing to elucidate the three-dimensional structures of trichosanthin and alpha-momorcharin. Prof. Dong spent six months at the University, during which he used the workstation to draw three-dimensional models of the interactions between the two proteins and their targets. The models help identify the amino acids that are crucial to the functions of trichosanthin as well as those that cause side effects. Researchers were able to change certain DNA sequences of trichosanthin and reinsert them into Escherichia coli to obtain modified versions of the protein. Prof. Y.B. Ke and Mr. S.K. Ma, both experts in trichosanthin from the Shanghai Institute of Cell Biology of the Academia Sinica, helped the researchers develop a new assay system for studying the effects of the modified trichosanthin. The effects of the modified proteins on various biological systems were studied by Prof. T.B. Ng (Department of Biochemistry) and Prof. W.Y. Chan (Department of Anatomy) at CUHK. Some of the modified proteins were given to Prof. Dong for three-dimensional analysis in Beijing.

The joint efforts of these scientists resulted in publications in prestigious journals, such as Science in China and Journal of European Biochemistry.

Dr. Shaw then worked with Dr. C.C. Wong and Prof. M.S.C. Tam of the Department of Physiology on a project to improve trichosanthin as a medicinal protein by lengthening its time inside the human body and reducing its allergic effects. The amino acids responsible for side effects were first identified and then modified in the laboratory. Some of these 'second generation' proteins exhibit a significantly reduced immunological effect, and different trichosanthin derivatives can be produced by manipulating the 'culprit' amino acids in different ways. It is hoped that an ideal drug can eventually be developed through systematic manipulations.

In the course of the research, Dr. Shaw also discovered that trichosanthin can inhibit the propagation of certain plant viruses. Working with Prof. Y.S. Wong of the CUHK Department of Biology, Dr. R.N.S. Wong of the Department of Biology at Baptist University, and Prof. B. Wang of the Institute of Genetics of the Academia Sinica in Beijing, Dr. Shaw and Dr. Yeung found that when inserted with a trichosanthin gene, tabacco can produce offsprings that demonstrate strengthened antiviral properties. Subsequently, under the sponsorship of the Beijing - Hong Kong Academic Exchange Centre, Ms X.B. Li, a senior engineer of the Institute of Genetics of the Academia Sinica, visited the University to exchange experience with Dr. Shaw. After her return to Beijing, and with assistance from Dr. Shaw, Ms Li and her colleague Mr. R.Z. Fu obtained a grant from the National Natural Science Funds to conduct research on the introduction and expression of the trichosanthin gene in potatoes and on the toxicity of trichosanthin in transgenic potato plants.

An Overview of the Significance of Trichosanthin Research

Research on trichosanthin has not only increased the understanding of the functions and mechanisms of this protein but also illustrated the importance of multidisciplinary collaboration. The research has grown in scope and depth through the contributions of various departments from local universities as well as from the Academia Sinica of mainland China. The experience has also enhanced scientific and technological exchange between Hong Kong and China.

Dr. P.C. Shaw graduated with first class honours from The Chinese University's Department of Biology in 1981. After receiving his M.Phil. in 1983, he pursued his doctoral studies at the Imperial College of Science and Technology, University of London, on a Croucher Foundation scholarship. Dr. Shaw was a postdoctoral fellow at the Imperial College before joining his alma mater as a lecturer in biochemistry in 1987. He was promoted to senior lecturer rank in 1995. Dr. Shaw has broad interest in scientific research and its application. Another research of his on the screening and cloning of restriction enzymes was reported in the SpringSummer issue of the Chinese University Bulletin in 1995.