RNA Polymerase Inhibitors with Activity against Rifampin-Resistant Mutants of Staphylococcus aureus

Abstract

A collection of rifampin-resistant mutants of Staphylococcus aureus with characterized RNA polymerase ␤-subunit (rpoB) gene mutations was cross-screened against a number of other RNA polymerase inhibitors to correlate susceptibility with specific rpoB genotypes. The rpoB mutants were cross-resistant to streptolydigin and sorangicin A. In contrast, thiolutin, holomycin, corallopyronin A, and ripostatin A retained activity against the rpoB mutants. The second group of inhibitors may be of interest as drug development candidates.Bacterial DNA-dependent RNA polymerase is an attractive drug target because RNA chain elongation is essential for bacterial growth (6, 16). Among those antibiotics discovered in the last 50 years, there are several known, or suspected, inhibitors of bacterial DNA-dependent RNA polymerase that have not been developed and could be candidates for new drugs. These agents include thiolutin (18), holomycin (B. Oliva, A. O'Neill, J. M. Wilson, P. J. O'Hanlon and I. Chopra, submitted for publication), streptolydigin (6, 16), the ripostatins, corallopyronins and sorangicins (10-12, 23) (Fig. 1). However, bacterial resistance has already developed to the rifamycins, the only class of RNA polymerase inhibitor that is in use clinically (21). Therefore, before considering whether other RNA polymerase inhibitors might be developed, it is important to establish whether resistance to rifamycins, such as rifampin, also confers cross-resistance to the other agents. Some attempts to address this issue have been made (9,12,13,15,24). However, the data are incomplete and the genetic basis of resistance to rifamycins in those strains used for cross-screening has rarely been determined. Furthermore, some data are contradictory; e.g., cross-resistance between rifampin and streptolydigin has been observed by some authors (13) but not by others (9,15).To assist the evaluation of these older agents we crossscreened them against a collection of rifampin-resistant mutants of Staphylococcus aureus, generated in an isogenic background, with defined RNA polymerase ␤-subunit (rpoB) gene mutations. These S. aureus strains, which provide a model for rpoB mutations occurring in naturally occurring isolates of staphylococci and other organisms (1,7,8,15,22,28,29), have allowed us to correlate susceptibility with specific rpoB genotypes.The antibiotics used here were either purchased from Sigma (rifampin and streptolydigin) or were gifts from H. Reichenbach, Gesellschaft für Biotechnologische Forschung, Braunschweig, Germany (corallopyronin A, ripostatin A, and sorangicin A); P. O'Hanlon, SmithKline Beecham Pharmaceuticals, Harlow, United Kingdom (holomycin and thiolutin); and Pharmacia & Upjohn (rifabutin). Spontaneous rifampin-resistant mutants of S. aureus 8325-4 (20) were isolated by plating approximately 10 8 CFU onto Iso-Sensitest agar (Oxoid, Basingstoke, United Kingdom) containing 0.032 g of rifampin/ml (four times the MIC). A number of rifampin-resistant mutants were picked at random, and their MICs of rifampin were d...

Myxobacterial secondary metabolites: bioactivities and modes-of-action

“…This requirement is met by myxopyronin, corallopyronin and ripostatin, but resistance to the rifamycins confers cross-resistance to sorangicin. 182 The structural basis for the lack of cross-resistance to the three metabolites was recently revealed by an elegant study carried out by Arnold, Ebright and colleagues, 183 in which they elucidated in detail the binding interaction between myxopyronin and bacterial RNAP. The overall shape of both bacterial and eukaryotic RNAPs is reminiscent of a crab claw, wherein the two pincers define the active site cleft (Fig.…”

Structural Perspective on Mutations Affecting the Function of Multisubunit RNA Polymerases

“…It has also been observed that some rifampin-resistant mutations can confer sorangicin resistance, suggesting largely overlapping binding sites for these two drugs (31,127,145). However, the structures of these drugs are different and observation that some rifampin-resistant mutants are sorangicin sensitive, while most sorangicin-resistant mutations confer rifampin resistance, suggests that subtle differences exist with regard to their modes of action (145).…”

“…8B). More specifically, they are found in the protrusion, fork and external 1 and 2 domains of ␤ (14,31,81,82,97,108,120,127,129,130,156,160,174,178,186,201,202,207). Thus, these replacements most probably affect the conformation of the binding pocket and lower its affinity for rifampin.…”

Novel Hybrid-Type Antimicrobial Agents Targeting the Switch Region of Bacterial RNA Polymerase

“…Holomycin (4) also possessed antibacterial properties and provided moderate but broad antibiotic spectrum against 12 In addition, 4 showed the inhibition of RNA synthesis, 11 and the calculate log P (ClogP) did not exceed 2.0.…”

“…11,12 In addition, 4 showed the inhibition of RNA synthesis, 11 and the calculate log P (ClogP) did not exceed 2.0. 13 These properties fascinated us to hybridize myxopyronins with the holothin molecule to expect the efficacy beyond our anticipations.…”