Researchers uncover the structure of anti-TB drug target, and identify a new weapon against the deadly disease

The fight against TB has taken several promising steps forward – scientists from Rutgers University have not only uncovered vital new information about how rifampicin, the frontline anti-TB drug, binds to its target, but have also discovered a completely new class of compounds that specifically kill TB bacteria.

The paper was published in the journal Molecular Cell.

TB is one of the biggest threats to global health. Caused by the bacteria Mycobacterium tuberculosis, it’s estimated to infect approximately 10 million people around the world each year, and kill almost 2 million, according to a 2016 World Health Organisation (WHO) report.

Currently, our main weapon against the disease is the drug rifampicin. This antibiotic works by targeting Mtb RNA Polymerase (Mtb RNAP), an enzyme in the bacteria which is crucial in the process of bacterial protein production. By interfering with how the enzyme works, rifampicin essentially stops the bacteria from making important proteins, leading to cell death.

The problem is, resistance to rifampicin has been on the rise. The WHO have reported about 0.6 million new cases of resistant strains each year. Usually, this happens when a mutation causes a change in structure at the site where rifampicin would bind. With a change in shape, the drug can’t bind anymore and isn’t able to exert its effect.

Now the 3D structure of Mtb RNAP, both alone and bound to rifampicin, has been revealed by the research team from Rutgers University. Data in this area was extremely limited, but by using a process known as X-ray crystallography, they’ve been able to give a much better idea of how rifampicin interacts on a structural level with the binding site on the enzyme.

Using this information, it should theoretically be much easier to develop new drugs, derived from rifampicin, which are able to show an improved ability to inhibit Mtb RNAP.

Mycobacterium tuberculosis, the bacteria which causes TB, has been exhibiting resistance towards the frontline drug rifampicin at an increasing rate, with approximately 0.6 million new cases each year. Image credit: Harvard Medical School

By screening 114,000 synthetic compounds, the team also discovered a brand-new group of inhibitors, unrelated to rifampicin, called Nɑ-aroyl-N-aryl-phenylalaninamides (or, more simply, AAPs). These compounds act on the same enzyme as rifampicin, Mtb RNAP, but crucially they have a completely different binding site.

The reason this is important is that even if a mutation in a strain of M. tuberculosis resulted in rifampicin’s binding site becoming altered, this shouldn’t have an effect on the AAPs binding to their target site.

When the team tested these compounds, they found they exhibited strong, selective action against TB bacteria, and didn’t appear to damage other bacterial or mammalian cells. Given that one of the risks of using antibiotics is the potential to kill our cells and the bacteria forming our microbiota, the fact that AAPs can avoid this ‘friendly fire’ effect is fairly promising.

They also found that AAPs and rifampicin worked extremely well as partners against M. tuberculosis – when they were both applied, their anti-mycobacterial ability saw an increase, and they were able to supress the emergence of resistant strains to below detectable levels.

“AAPs represent an entirely new class of Mtb RNAP inhibitors and are, without question, the most promising Mtb RNAP inhibitors for anti-TB drug development since rifampicin,” said Professor Richard H. Ebright, one of the researchers behind the work. “We are actively pursuing AAPs. We have synthesised and evaluated more than 600 novel AAPs and have identified AAPs with high potencies and favourable intravenous and oral pharmacokinetics.”

Nader Fotouhi, Chief Scientific Officer at the Global Alliance for TB Drug Development praised the work at an earlier press conference. “The discovery of an alternative binding site and the AAPs represents a significant step towards the identification of a novel RNAP inhibitor that would behave like rifampicin but be devoid of any pre-existing resistance.

The co-crystal structure of AAP and Mtb RNAP is a critical stepping stone for the design of next generation RNAP inhibitors.”

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