High-level semi-synthetic production of the potent antimalarial artemisinin.

In 2010 there were more than 200 million cases of malaria, and at least 655,000 deaths. The World Health Organization has recommended artemisinin-based combination therapies (ACTs) for the treatment of uncomplicated malaria caused by the parasite Plasmodium falciparum. Artemisinin is a sesquiterpene endoperoxide with potent antimalarial properties, produced by the plant Artemisia annua. However, the supply of plant-derived artemisinin is unstable, resulting in shortages and price fluctuations, complicating production planning by ACT manufacturers. A stable source of affordable artemisinin is required. Here we use synthetic biology to develop strains of Saccharomyces cerevisiae (baker's yeast) for high-yielding biological production of artemisinic acid, a precursor of artemisinin. Previous attempts to produce commercially relevant concentrations of artemisinic acid were unsuccessful, allowing production of only 1.6 grams per litre of artemisinic acid. Here we demonstrate the complete biosynthetic pathway, including the discovery of a plant dehydrogenase and a second cytochrome that provide an efficient biosynthetic route to artemisinic acid, with fermentation titres of 25 grams per litre of artemisinic acid. Furthermore, we have developed a practical, efficient and scalable chemical process for the conversion of artemisinic acid to artemisinin using a chemical source of singlet oxygen, thus avoiding the need for specialized photochemical equipment. The strains and processes described here form the basis of a viable industrial process for the production of semi-synthetic artemisinin to stabilize the supply of artemisinin for derivatization into active pharmaceutical ingredients (for example, artesunate) for incorporation into ACTs. Because all intellectual property rights have been provided free of charge, this technology has the potential to increase provision of first-line antimalarial treatments to the developing world at a reduced average annual price.

Piperine inhibits gastric emptying and gastrointestinal transit in rats and mice.

Piperine (1), an alkaloid of black and long peppers, inhibited gastric emptying (GE) of solids/liquids in rats and gastrointestinal transit (GT) in mice in a dose and time dependent manner. Compound 1 significantly inhibited GE of solids and GT at the doses extrapolated from humans (1 mg/kg and 1.3 mg/kg p.o. in rats and mice, respectively). However, at the same dose the effect was insignificant for GE of liquids. One week oral treatment of 1 mg/kg and 1.3 mg/kg in rats and mice, respectively, did not produce a significant change in activity as compared to single dose administration. GE inhibitory activity of 1 is independent of gastric acid and pepsin secretion.

Antidiarrhoeal activity of piperine in mice.

Peppers are common food ingredients used worldwide. They are also added in traditional antidiarrhoeal formulations of different herbs. Piperine (1) is an alkaloidal constituent of black and long peppers recently established as a bioavailability enhancer of drugs and other substances. As a part of efforts to study its effects on the gastrointestinal tract, the experiments were performed to determine the rationale, if any, for its use in traditional antidiarrhoeal formulations. Antidiarrhoeal activity of 1 against castor oil, MgSO4 and arachidonic acid was studied in mice. It significantly inhibited diarrhoea produced by these cathartics at 8 and 32 mg/kg p.o. dose. Inhibition of castor oil induced enteropooling by 1 suggests its inhibitory effect on prostaglandins. The results validate the rationale for its use in traditional antidiarrhoeal formulations.

Development of liposomal amphotericin B formulation.

A considerable effort has been spent in the past three decades to investigate various aspects of liposomes as novel drug delivery systems. In 1990, the first amphotericin B (AmB) liposomal preparation (L-AmB) under the brand name AmBisome was introduced into the market by Vestar. The successful marketing of the product moved liposomes out of the stage of experimental obscurity to the realistic stage of clinical utility. The launch of AmBisome sparked off the introduction of other lipid-based AmB products marketed by Liposome Technology (Amphocil) and The Liposome Co. (Abelcet). The drive behind the development of a modified formulation of AmB was to improve the therapeutic index of this drug with respect to its major drawback associated with both acute and chronic toxic effects. In a 30-year-long experience with AmB, several reports were recorded in the literature of acute adverse effects, such as fever, rigors, vomiting, cardiotoxicity and hypotension occurring during infusion; while long-term therapy was reported to be associated with hypokalemia, renal dysfunction and hematological abnormalities. Another serious problem encountered with the drug had been the poor response obtained in immunocompromised patients like those with AIDS, neutropenia and cancer patients on chemotherapy. The encapsulation of amphotericin B in liposomal vesicles was hence targeted not only to obtain an improvement in the therapeutic index but also to see if it was useful in eradicating deep-seated fungal infections in immunocompromised patients. The liposomal AmB was found to have a better therapeutic index and lower toxicity than the commercial AmB preparations. The LD50 of AmBisome in mouse was 175 mg/kg compared with 3.7 mg/kg for Fungizone, the commercial preparation of AmB. Additionally, L-AmB has prolonged circulation time, and extravasates into the site of infection and delivers the drug directly to the site, with no nephrotoxicity and neurotoxicity as experienced with AmB. This review traces the course of development of L-AmB and discusses the rationale behind the development of its liposomal preparation. The results in in vitro, in vivo and clinical studies, mechanism of action, biodistribution, and formulation considerations of L-AmB are described. The clinical experience with the marketed preparation is reviewed.