In Vitro Activities of Arylomycin Natural-Product Antibiotics against Staphylococcus epidermidis and Other Coagulase-Negative Staphylococci

The coagulase-negative staphylococci (CoNS) are a heterogeneous group of at least 15 different species of Gram-positive bacteria that have emerged in recent decades as important nosocomial pathogens (10, 31). A particularly problematic species is Staphylococcus epidermidis, which is responsible for a growing number of infections among hospital patients with compromised immune systems and is especially notorious for forming biofilms that adhere to surgical equipment and other hospital surfaces and indwelling devices (3, 18). Methicillin was traditionally the first-line antibiotic against CoNS, but its widespread use has resulted in resistance in 50% to 80% of CoNS infections and 75 to 90% of nosocomial S. epidermidis infections (18). As a result, vancomycin is now the first line agent for treating CoNS infections; however, isolates with reduced susceptibility to vancomycin have also been observed (11, 26), and the emergence of enterococci harboring mobile elements that confer vancomycin resistance has raised concerns that resistance might be transferred to S. epidermidis and/or other CoNS (17, 28). These concerns continue to motivate the search for new antibiotics that are active against CoNS, especially S. epidermidis. The arylomycins (Fig. ​(Fig.1)1) are a novel class of natural-product antibiotics that act by inhibiting bacterial type I signal peptidase (SPase) (19, 25). SPase is a Ser-Lys dyad protease that removes N-terminal signal sequences from preproteins following their translocation across the cytoplasmic membrane (5, 20). SPase is an attractive target for antibiotic therapy because it is conserved, essential, and located in the relatively accessible outer leaflet of the cytoplasmic membrane. Furthermore, because bacterial SPase acts via a catalytic mechanism that is distinct from that of its eukaryotic homologues, the arylomycins are unlikely to exhibit mechanistic toxicity in humans (5, 20). FIG. 1. Structures of arylomycin A2 (R = iso-C12) and arylomycin C16 (R = iso-C16) [iso-C12 = CH2(CH2)7CH(CH3)2 and iso-C16 = CH2(CH2)11CH(CH3)2]. Despite the apparent accessibility, essentiality, and conservation of SPase, initial reports suggested that the arylomycins were active against only a few Gram-positive bacteria, including Streptococcus pneumoniae, Rhodococcus opacus, and Brevibacillus brevis (15, 25), and not against other important Gram-positive pathogens or against any Gram-negative bacteria. However, after reporting the first synthesis of an arylomycin, arylomycin A2, as well as the synthetic derivative arylomycin C16 (Fig. ​(Fig.1),1), we found that each potently inhibits the growth of S. epidermidis (24) and that S. epidermidis evolves resistance to the arylomycins by mutating residue 29 of one of its two SPases, SpsIB, from Ser (Ser29) to Pro (Pro29) (29). Moreover, a Pro residue is naturally present at the analogous position in the homologous SPases of the pathogens Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa, and we showed that it imparts resistance by reducing the affinity with which the arylomycins bind. Furthermore, we found that a remarkably diverse range of both Gram-positive and Gram-negative bacteria whose SPases lack a Pro at the analogous position are susceptible to the arylomycins, including Staphylococcus epidermidis, Streptococcus pyogenes, Helicobacter pylori, Chlamydia trachomatis, and some strains of Francisella tularensis (29). In total, the results suggest that the identified SPase polymorphism is a major contributor to naturally occurring arylomycin resistance. However, we also showed that Yersinia pestis and some strains of S. aureus are susceptible to the arylomycins despite the presence of an analogous Pro, while others, such as many of the Lactobacillales, Clostridia, and Bacteriodetes, are resistant despite its absence, implying that in some cases, susceptibility must depend on additional factors, such as variable levels of toxicity associated with the inhibition of protein secretion. The potent activity of the arylomycins against a strain of S. epidermidis (RP62A) suggests that they might be useful in the treatment of this and perhaps other CoNS. Here, to examine the spectrum of activity of the arylomycins against clinical isolates of S. epidermidis and other CoNS, we report the activity of arylomycin C16 against two panels of isolates from hospitals in geographically diverse locations and compare the activity to that of vancomycin. The results reveal that the arylomycins have potent antibacterial activity against a range of important CoNS species whose SpsIB orthologs lack the previously identified resistance-conferring Pro, while less activity is observed against species where Pro is present. While we generally observed similar susceptibilities for different isolates within a species, significant differences were observed in several cases, with one atypical instance of susceptibility resulting from the presence of a Ser in place of the resistance-conferring Pro. Significant differences in susceptibility between isolates of the same species are usually observed with clinically deployed antibiotics where selection for resistance has occurred during therapy (2, 9, 14, 21), and therefore, these results may be relevant to understanding the natural evolution of arylomycin resistance in nature.