The bactericidal FabI inhibitor Debio 1453 clears antibiotic-resistant Neisseria gonorrhoeae infection in vivo

Abstract

Gonorrhoea is a prevalent sexually transmitted infection caused by the bacterial pathogen Neisseria gonorrhoeae. N. gonorrhoeae has demonstrated a remarkable capacity to evolve antibiotic resistance, with emerging strains that show resistance to all standard treatment options. The development of new antibiotics for gonorrhoea, especially those with novel targets and no pre-existing resistance, is critical. One such untapped antibacterial target in N. gonorrhoeae is FabI, an enoyl-acyl carrier protein reductase enzyme that is essential for fatty acid biosynthesis in this pathogen. In the current report, structure-based drug design using novel N. gonorrhoeae FabI inhibitor co-crystals guides medicinal chemistry toward increasing potency in the sub-nanomolar range and drives the discovery of Debio 1453. Debio 1453 is optimized for activity against N. gonorrhoeae and is highly active in vitro against diverse N. gonorrhoeae isolates including those resistant to the last remaining treatment options. Additionally, the compound presents a low propensity for selection of mutants with reduced susceptibility. Debio 1453 is efficacious in vivo against N. gonorrhoeae isolates with clinically relevant multi-drug resistance phenotypes in a murine vaginal gonorrhoea infection model underscoring Debio 1453 as a promising candidate for the treatment of gonorrhoea.

Fig. 1. Discovery of Debio 1453.

a Afabicin desphosphono (Debio 1452) with structural characteristics governing FabI inhibition. b Representative key compounds in lead optimisation for Neisseria gonorrhoeae FabI (NgFabI). N. gonorrhoeae minimum inhibitory concentration (MIC)₅₀ values were determined for a screening panel of 14 isolates. IC, inhibitory concentration. Source data are provided as a Source Data file.

Fig. 2. Key structural interactions between Debio 1453 and NgFabI.

a Main interactions of Debio 1453 (bold grey) with Neisseria gonorrhoeae FabI active site (key amino acids in light grey) and NADH co-factor (bold orange) from the co-crystallised ternary structure. H-bonds between Debio 1453 and FabI or NADH are represented by dashed green, while H-bonds between Debio 1453 and NADH via a conserved water network are in solid green. Debio 1453 is also interacting with the enzyme via T-stacking (*) and with the co-factor via Van der Waals hydrophobic interactions (**). b View of the right-hand side methyl group (bold green) of Debio 1453 (bold grey) in the N. gonorrhoeae FabI active site (amino acids and NADH in light grey) with surrounding 3 lipophilic amino acids (bold yellow) from the co-crystallised ternary structure.

Fig. 3. Cumulative minimum inhibitory concentration (MIC) distributions for Debio 1453 and comparator antibiotics for 100 clinical Neisseria gonorrhoeae isolates collected from Sweden in 2023.

Relevant EUCAST resistance breakpoints or epidemiological cutoff (ECOFF) for N. gonorrhoeae: ceftriaxone, >0.125 µg/mL; azithromycin, 1 µg/mL (ECOFF); spectinomycin, >64 µg/mL; tetracycline, >0.5 µg/mL; and ciprofloxacin, >0.06 µg/mL. The dotted line indicates 90% of cumulative isolates (MIC₉₀). Source data are provided as a Source Data file.

Fig. 4. Bactericidal activity of Debio 1453 against Neisseria gonorrhoeae.

Combined in vitro time-kill data for 10 N. gonorrhoeae strains at increasing multiples of Debio 1453 minimum inhibitory concentration (MIC). Data are the mean +/− standard deviation (n = 10). Time-kill curves for each individual strain are presented in Supplementary Fig. 3. Source data are provided as a Source Data file.

Fig. 5. Intracellular killing of Neisseria gonorrhoeae in cultured HeLa229 human cervix carcinoma cells.

a Intracellular killing of N. gonorrhoeae strain ATCC 49226 at increasing multiples of Debio 1453 or azithromycin minimum inhibitory concentration (MIC). b Intracellular killing of N. gonorrhoeae strain WHO X at increasing multiples of Debio 1453 or azithromycin MIC. Data are the mean of biological triplicates +/− standard deviation. Dotted grey lines indicate the limit of detection. CFU colony forming unit, CIP-R ciprofloxacin-resistant, CRO-R ceftriaxone-resistant. Source data are provided as a Source Data file.

Fig. 6. Plasma exposure of Debio 1453 dosed as Debio 1453 P in a Neisseria gonorrhoeae murine vaginal infection model.

Total (black) and free (blue, accounting for mouse plasma protein binding of 93.7%) Debio 1453 concentrations are the mean from three animals per timepoint, with standard deviation. Error bars below zero cannot be displayed due to the logarithmic scale for the x-axis. The lower limit for quantification of total Debio 1453 was 10 ng/mL. The minimum inhibitory concentration (MIC) range for Debio 1453 is highlighted in grey, and the MIC inhibiting 90% of N. gonorrhoeae isolates (MIC₉₀) is represented by the grey dotted line. Pharmacokinetic parameters were determined using Phoenix WinNonlin by non-compartmental analysis. The area under the curve (AUC)last is for total Debio 1453. C_max maximum concentration, t_max time taken to reach C_max, t1/2 half-life. Source data are provided as a Source Data file.

Fig. 7. In vivo efficacy of Debio 1453 for the treatment of Neisseria gonorrhoeae in a murine vaginal infection model.

Bacterial counts (colony forming units, CFU) per millilitre of vaginal lavage fluid for mice infected with N. gonorrhoeae strains a ATCC 700825, b AR Bank0179-15, c AR Bank0181-17, and d WHO X-07, treated with either ceftriaxone (single intraperitoneal dose) or Debio 1453 P (dosed orally twice daily; doses are Debio 1453 equivalent values). Minimum inhibitory concentrations (MICs) were determined using agar dilution and are the mode of multiple measures. Dotted lines indicate the limit of detection, dashed grey lines show mean CFU/mL at the beginning of treatment. ATCC 700825 is resistant to streptomycin, facilitating use in the model, whereas AR Bank0179-15, AR Bank0181-17 and WHO X-07 were engineered for streptomycin resistance from progenitors AR Bank0179, AR Bank0181 and WHO X, respectively. Data for each group is summarised using the mean (black horizontal bars, n = 5 animals per group). Statistically significant differences compared to 0 h were determined using one-way ANOVA with Dunnett’s multiple comparisons test. *P < 0.05, **P < 0.01, ***P < 0.001. AZM-R azithromycin-resistant, CIP-R ciprofloxacin-resistant, CRO-R ceftriaxone-resistant. Source data, including statistical test output, are provided as a Source Data file.

Fig. 8. In vivo efficacy of Debio 1453 in a Staphylococcus aureus neutropenic murine thigh infection model.

The infective strain, S. aureus ATCC 29213, was inoculated 2 h before the start of treatment (0 h). Thighs were harvested at 24 h and 48 h for colony-forming unit (CFU) determinations. Data are summarised by the mean for each group (black horizontal bars, n = 5 animals per group). Debio 1453 was dosed as Debio 1453 P; doses are Debio 1453 equivalents. Debio 1453 P was dosed orally, twice daily. Linezolid was dosed orally three times daily. The dotted grey lines depict the mean CFU/mL at the start of treatment. Statistically significant differences compared to 0 h were determined using one-way ANOVA with Dunnett’s multiple comparisons test. *P < 0.05, **P < 0.01, ***P < 0.001. MIC, minimum inhibitory concentration. Source data, including statistical test output with exact P-values, are provided as a Source Data file.