Computer-aided drug design to generate a unique antibiotic family

Abstract

The World Health Organization has identified antibiotic resistance as one of the three greatest threats to human health. The need for antibiotics is a pressing matter that requires immediate attention. Here, computer-aided drug design is used to develop a structurally unique antibiotic family targeting holo-acyl carrier protein synthase (AcpS). AcpS is a highly conserved enzyme essential for bacterial survival that catalyzes the first step in lipid synthesis. To the best of our knowledge, there are no current antibiotics targeting AcpS making this drug development program of high interest. We synthesize a library of > 700 novel compounds targeting AcpS, from which 33 inhibit bacterial growth in vitro at ≤ 2 μg/mL. We demonstrate that compounds from this class have stand-alone activity against a broad spectrum of Gram-positive organisms and synergize with colistin to enable coverage of Gram-negative species. We demonstrate efficacy against clinically relevant multi-drug resistant strains in vitro and in animal models of infection in vivo including a difficult-to-treat ischemic infection exemplified by diabetic foot ulcer infections in humans. This antibiotic family could form the basis for several multi-drug-resistant antimicrobial programs.

Fig. 1. Rationale design of AcpS active site inhibitors.

a AcpS catalyzes the transfer of a 4’-phosphopantetheine moiety from coenzyme A onto a conserved Ser residue of apo-ACP producing 3’5’-adenosine bisphosphate and holo-ACP. b Exemplar compounds from the three generations of thienyltetrazole-focused library. Compounds in Generation 1 (Gen 1) were optimized to find a plateau of AcpS potency and were physiochemically optimized through Generation 2 (Gen 2) for bacterial cell permeability (via MIC) as denoted by the double circle, to arrive at Generation 3 (Gen 3) compounds that can both inhibit AcpS and permeate bacterial membranes. Average values of each property are shown left to right across the library optimization. Several compounds, as the focused library evolved, rose to the potency of low μM inhibitors of AcpS IC₅₀ and low μg/ml MIC against the test organism, MRSA. c Predicted top pose (after minimization) of DNM0547 in the AcpS active site. Proximal cationic and lipophilic residues are prominently labeled: Lys63 A chain; Phe41, Arg46, Phe50, Arg54, Lys87 B chain.

Fig. 2. In vivo efficacy of DNM0547.

a DNM0547 activity against MRSA (ATCC 33591) in a rabbit ischemic ear model of infection. Control contained excipient cream only while DNM0547 was 2% (w/w) in excipient; control (n = 6) and DNM0547 (n = 10). Data are represented as mean ± SEM. Significance was determined using a two-way ANOVA with Geisser-Greenhouse correction with Šidák’s multiple comparisons test, P 0.05. b CFUs per gram of tissue were determined at the end of a rabbit ischemic ear model of infection; control (n = 4) and 2% DNM0547 (n = 7). Data are represented as individual data points with the line representing the mean and error as SEM. Significance was determine using a one-sided unpaired student’s t-test with Welch’s correction, ***P 0.008. c Mupirocin activity against MRSA (ATCC 33591) in a rabbit ischemic ear model of infection; control (n = 6) wounds and 2% mupirocin treated (n = 5). Data are represented as mean ± SEM. Significance was determined using a two-way ANOVA with Geisser-Greenhouse correction with Šidák’s multiple comparisons test. Differences between control and mupirocin treated animals were not statistically significant. d DNM0547 activity against MRSA (ATCC 33591) in a mouse cellulitis model of infection; control (n = 5) and 2% DNM0547 treated (n = 5). Data are represented as mean ± SEM. Significance was determined using a two-way ANOVA with Geisser-Greenhouse correction with Šidák’s multiple comparisons test, *P 0.05. e CFUs per gram of tissue were determined at the end of the mouse cellulitis model of infection; control (n = 5) and 2% DNM0547 treated (n = 5). Data are represented as individual data points with the line representing the mean. Significance was determine using a one-sided unpaired student’s t-test, ***P 0.0001.