January 11, 2007
Engineered biosynthesis – a path to new and improved antibiotics

It’s an old adage that you sometimes have to take the good with the bad. This is regrettably the case with therapies for many diseases. In reality, many treatments that help a patient can also do some measure of harm. With antibiotics, side-effects can range from manageable, like nausea, fatigue and dizziness, to severe such as irreversible organ damage.

The antifungal antibiotic amphotericin B is one of the most notoriously toxic drugs used in clinical medicine. Its many side effects include damage to kidneys, heart and brain. On the other hand, amphotericin B is powerfully active against most fungal pathogens and resistance has been slow to emerge over the past fifty years. For these reasons, amphotericin B remains the drug of choice for the treatment of life-threatening systemic fungal infections.

Amphotericin B is a complex natural product obtained from a bacterium called Streptomyces nodosus. The active compound is extracted from fermentation cultures on an industrial scale. The toxicity of this material can be reduced by chemical modification or by encapsulation of the drug into liposomes, but neither of these solutions is economically favourable.

At the UCD School of Biomolecular and Biomedical Science, Dr Patrick Caffrey, a Centre for Synthesis and Chemical Biology (CSCB) and UCD Conway Investigator, leads a team of microbiologists working on engineering biosynthesis of less toxic amphotericin derivatives. Initial work involved cloning and sequencing of amphotericin biosynthetic genes and developing methods for manipulating these genes within Streptomyces nodosus.

"We believed that this would allow for the production of analogues with reduced toxicity,” explains Dr Caffrey. He continues “About 150 kb of chromosomal DNA is dedicated to amphotericin production. Sequence analysis revealed that over ninety enzymes are involved in the biosynthetic pathway. Genes for key enzymes were eventually knocked out to generate mutant strains that produce improved versions of amphotericin B. These compounds cannot be easily made by chemical synthesis.”

The new amphotericin derivatives are produced in moderate to good yields. Chemical analysis was carried out with the assistance of Dr Dilip Rai of the CSCB mass spectrometry facility at UCD. Detailed structural analysis was carried out in collaboration with Dr Bernard Rawlings of the Department of Chemistry at the University of Leicester.

The most promising new analogues show antifungal activity comparable to amphotericin B and a dramatic reduction in haemolytic activity, a measure of toxicity. Work is in progress to scale up fermentation and extraction of these compounds.

In addition to their value as antifungal agents, Dr Caffrey is optimistic that non-toxic amphotericin analogues and formulations may also be useful in other therapeutic areas. “Amphotericin B is effective against HIV, Leishmania parasites and prion diseases but toxicity has prevented exploitation of these additional activities. The availability of less toxic analogues may allow for the full potential of amphotericin-based drugs to be realised.”

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