Rifamycin: Definition, Mechanism and Uses

What are rifamycin antibiotics?

Rifamycin is a complex isolated from the metabolites of Mediterranean streptomycin by Sensi et al. in 1959, and subsequently isolated A, B, C, D, E, etc. In 1962, rifamycin B was chemically converted to rifamycin SV and was first used clinically. Rifamycin is a class of broad-spectrum antibiotics mainly used to treat infections caused by Gram-positive bacteria, tuberculosis bacilli and certain drug-resistant Staphylococcus aureus. It is widely used in the treatment of mycobacterium infection such as tuberculosis and leprosy. The parent nucleus of rifamycin class is macrocyclic lactam, which contains a naphthalene nucleus, and the naphthalene ring is connected with two non-adjacent carbon atoms of an aromatic group by a fat chain, forming an ansa bridge structure, so it belongs to ansamycin class antibiotics.

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Rifamycin mechanism of action

Rifamycin achieves bactericidal effect mainly by inhibiting ribonucleic acid polymerase activity in bacteria. This drug has strong antibacterial activity against gram-positive bacteria and tuberculosis bacteria, etc. Specifically, rifamycin's mechanisms of action include:

Inhibition of RNA polymerase: Rifamycin prevents the formation of phosphodiester bonds by specifically binding to the beta subunit of bacterial DNA-dependent RNA polymerase (RNAP), thereby inhibiting the elongation of RNA chains, resulting in the blockage of bacterial protein synthesis, and thus affecting bacterial growth and reproduction. These antibiotics consist of a naphthalene core, which is brided by a polyketone ansa. This structure presents a unique three-dimensional structure, which binds to the RNA polymerase through a series of hydrogen bonds attached to the C21 and C23 of the naphthalene nuclei and ansa bridges. This binding occurs not at the enzymatically active site of template-directed RNA synthesis, but at the RNA exit channel, thereby blocking the production of full-length RNA.

Inhibition of DNA cyclotase: Rifamycin can interfere with the process of bacterial DNA replication, so that the bacterial DNA chain can not be connected properly, thus affecting the growth and reproduction of bacteria.

Rifamycin class and uses

The most well-known rifamycin compounds include rifampicin, rifapentine, rifabutin and rifaximin.

Rifampicin (rifampin) is a semi-synthetic broad-spectrum rifamycin antibacterial agent, which has obvious bactericidal effect on mycobacterium tuberculosis and some non-tuberculous mycobacteria (including mycobacterium leprae) both in and out of host cells. Rifampicin is bactericidal at low concentrations and bactericidal at high concentrations. The principle of action is that rifampin binds firmly to the beta subunit of DNA-dependent RNA polymerase, which inhibits the synthesis of bacterial RNA, but has no effect on mammalian enzymes. Rifuren infiltrates body tissues and body fluids, can enter the cerebrospinal fluid, mainly used for the treatment of tuberculosis and extrapulmonary tuberculosis.

The rifamycin family of antibiotics. (Surette, M. D., 2021)

Rifapentine is a cycloamyl-substituted rifamycin, a broad-spectrum full-effect bactericide with an action strength of 2-10 times that of rifampicin. It is mainly used for the treatment of primary and secondary tuberculosis.

Rifabutin is a semi-synthetic derivative of rifamycin developed and marketed in Italy in 1993. It was originally approved for use in AIDS patients with Mycobacterium avium intracellularis infection. Compared with rifampicin, rifabutin has stronger antibacterial activity, longer duration of action, and lower toxicity. Studies have shown that the antibacterial effect of rifabutin on Mycobacterium tuberculosis is about 4 times that of rifampicin, and about 30% of rifampicin-resistant strains are still sensitive to rifabutin. Therefore, rifampicin-resistant strains should no longer be selected for rifapentine, but rifabutin can be used as appropriate.

Rifaximin, a semi-synthetic derivative of rifamycin SV, was marketed in Italy in 1987 as an anti-infectious diarrhea drug. It was later approved by the FDA for the treatment of diarrhea form of irritable bowel syndrome (IBS-D) in adults. It works by targeting bacteria in the gut that can cause IBS symptoms such as abdominal pain, bloating and diarrhoea. Rifaximin is modified at the C3C4 site of rifamycin's naphthalene nucleus to exhibit zwitterionic properties at physiological pH. Pyridine nitrogen is positively charged, imidazole nitrogen is negatively charged, and the opposite charged nitrogen, together with the presence of phenolic hydroxyl groups, results in molecular ionization at all pH values along the digestive tract, thus not being absorbed throughout the digestive tract. Rifaximin has high antibacterial activity against a variety of gram-positive, gram-negative aerobic and anaerobic bacteria. Only 1% of the oral dose is absorbed through the gastrointestinal tract, which only plays a role in the intestine and has almost no systemic effect, so the adverse reactions are light and safe. Unlike other intestinal antibiotics, rifaximin has little killing effect on normal intestinal flora, and can inhibit bacterial virulence (inhibit bacterial adhesion, internalization, and translocation), down-regulate inflammatory response, and positively regulate intestinal microbiota. Rifaximin can increase the abundance of beneficial bacteria in the gut while keeping the overall composition of the gut microbiome stable.

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Mechanism of rifamycin resistance

Rifamycin (Rif) is an important antibiotic used to treat tuberculosis. However, bacterial resistance to rifamycin has become a serious problem. Resistance to rifamycin mainly stems from point mutations in RNA polymerase, which reduce the affinity of antibiotics for the RNA exit channel binding site. Among these microorganisms, rifamycin resistance involves many different enzyme mechanisms that directly modify and alter antibiotics to inactivate them, including ADP ribosyltransferases, glycosyltransferases, phosphotransferases, and monooxygenases.

Rifampin ADP ribosyltransferase (ARR): This enzyme neutralizes antibiotic activity by ribosylation at a specific hydroxyl group on the ansa bridge of rifamycin. This process has been found in mycobacteria.

Rifampin phosphotransferase (RPH): This enzyme changes its ability to bind to RNA polymerase by adding a phosphate group to the ansa chain of rifamycin.

Rifampin glycosyltransferase (RGT): This enzyme adds sugar groups at specific locations on rifamycin, thereby changing its structure and function, making bacteria resistant to drugs.

Rifampin monooxygenase (ROX): This enzyme breaks down rifamycin and reduces its concentration, allowing bacteria to grow at otherwise lethal concentrations.

To avoid antibiotic resistance, researchers needed a drug similar to Rif that could bind to RNAP even in the presence of mutations. Typically many scientists choose to synthesize such molecules in the laboratory, but Sean F. Professor Brady turned his eyes to nature. He said: "Rifamycin is naturally produced by a bacterium. So I wanted to find out if nature had also made an analogue of Rif, a molecule that looked like rifamycin but was slightly different." To identify the substance, Brady's laboratory genetically sequenced microorganisms in soil samples collected across the United States. They hope to find antibiotics related to the Rif gene, but with small mutations that allow them to bind to mutated RNAP. Researchers found a group of natural antibiotics in soil samples called kanglemycin (kang), which shares most of the genes with rifamycin. In addition, analysis by postdoctoral researcher James Peek showed that these antibiotics were effective against bacteria that did not respond to Rif. By analyzing their structure, the research team found that extra sugar could stop kang in RNAP 'pockets' that are not available to other drugs. The discovery of this parking location provides researchers with a new strategy for developing more powerful antibiotics.

References

1. Surette, M. D., et al. The enzymes of the rifamycin antibiotic resistome. Accounts of Chemical Research. 2021, 54(9): 2065-2075.
2. Peek, J., et al. Rifamycin congeners kanglemycins are active against rifampicin-resistant bacteria via a distinct mechanism. Nature Communications. 2018, 9(1): 4147.