Bacteria develop resistance against a drug only when they are exposed to it or when the drug is misused. But now, a team of researchers from India has found whether and how drug resistance can develop against a candidate drug called Augmentin even before the drug is approved for treating patients with drug-resistant TB. Augmentin is currently undergoing clinical trials in patients with drug-resistant TB; it is already being used for common bacterial infections.
Besides deciphering the mechanism by which TB bacteria can develop resistance against Augmentin, the researchers have found ways of overcoming this potential resistance mechanism, thereby making Augmentin a potentially powerful drug to treat both multidrug-resistant TB (MDR-TB) and extensively drug-resistant TB (XDR-TB).
The beta-lactam class of antibiotics such as penicillin, ampicillin and amoxicillin is one of the most widely used class of antibacterial drugs. Despite its ability to kill several types of bacteria, the beta-lactam antibiotics have never been used against TB bacteria. This is because TB bacteria are naturally resistant to this class. TB bacteria inherently produce an enzyme called beta-lactamase which breaks down beta-lactam class of antibiotics (through hydrolysis) and makes the drug ineffective against TB disease.
Making of Augmentin
One of the strategies of getting around using the beta-lactam class of antibiotics is developing an inhibitor against beta-lactamase enzyme. Clavulanic acid is one such inhibitor, which blocks the beta-lactamase enzyme. Augmentin, which is a combination of a beta-lactam antibiotic (amoxicillin) and beta-lactamase inhibitor (clavulanic acid), can thus be an effective drug against TB bacteria.
“Till now no one knew the exact mechanism of how the combination of beta-lactam and beta-lactamase inhibitor was killing TB bacteria and how resistance against the combination can emerge in future,” says Dr. Amit Singh from the Centre for Infectious Disease Research at the Indian Institute of Science (IISc), Bengaluru, and the corresponding author of the paper published in the journal eLife. “Our study was able to provide insights into how resistance against Augmentin can emerge.”
The team used integrated experimental technology and computer tools to understand the mechanism by which resistance against Augmentin can set in.
Deciphering the mechanism
The first thing that the researchers asked was how the TB bacterium senses the presence of the drug combination in and around it. “We found the bacterium when exposed to this drug combination changes its metabolism and respiration, which led to the production of sub-lethal amount of reactive oxygen species (ROS). The ROS acts as a danger signal for the bacteria to mount a defence mechanism against Augmentin,” Dr. Singh explains.
The defence mechanism is through a protein called WhiB4, which is normally present in bacteria and is responsible for regulating the production of beta-lactamase enzyme. When the WhiB4 protein senses the ROS signal, it produces large amounts of beta-lactamase enzyme in the TB bacteria. “This could be one method by which the bacteria can become resistant to Augmentin,” Dr. Singh says.
Besides producing beta-lactamase enzyme, the WhiB4 protein also controls the production of an antioxidant molecule called mycothiol. The main role of mycothiol is to reduce the excessive increase in ROS so that ROS level is kept in balance; excessive ROS can kill bacteria by damaging proteins, DNA, and cell wall lipids.
“The WhiB4 protein can detect the ROS signal produced by antibiotics and direct the production of both beta-lactamase and mycothiol, which work together and contribute to bacterium’s ability to resist augmentin,” says Saurabh Mishra from the Centre for Infectious Disease Research at IISc and the first author of the paper.
Making Augmentin powerful
The researchers demonstrated that it is possible to kill MDR-TB and XDR-TB by simply changing the levels of the regulator, WhiB4, and/or increasing the ROS levels inside the bacteria. “When we knocked out mycothiol production, the level of ROS increased inside the bacteria and ultimately resulted in efficient killing of drug-resistant TB bacteria,” he says.
There are certain antibiotics (such as clofazimine) that work by increasing the ROS levels inside bacteria. The researchers are currently testing if using such antibiotics along with Augmentin can efficiently kill drug-resistant TB bacteria. Augmentin and clofazimine antibiotics can together elevate the production of ROS. The excessive ROS inside the bacteria can then kill all forms of drug-resistant TB bacteria.