Quinolone antibiotic; DNA gyrase/topoisomerase

The Centers for Disease Control and the World Health Organization have issued a list of priority pathogens for which there are dwindling therapeutic options, including antibiotic-resistant Neisseria gonorrheae, for which novel oral agents are urgently needed. Zoliflodacin, the first in a new class of antibacterial agents called the spiropyrimidinetriones, is being developed for the treatment of gonorrhea. It has a unique mode of inhibition against bacterial type II topoisomerases with binding sites in bacterial gyrase that are distinct from those of the fluoroquinolones. Zoliflodacin is bactericidal, with a low frequency of resistance and potent antibacterial activity against N. gonorrheae, including multi-drug-resistant strains (MICs ranging from ≤0.002 to 0.25 μg/mL). Although being developed for the treatment of gonorrhea, zoliflodacin also has activity against Gram-positive, fastidious Gram-negative, and atypical pathogens. A hollow-fiber infection model using S. aureus showed that that pharmacokinetic/pharmacodynamic index of fAUC/MIC best correlated with efficacy in in vivo neutropenic thigh models in mice. This data and unbound exposure magnitudes derived from the thigh models were subsequently utilized in a surrogate pathogen approach to establish dose ranges for clinical development with N. gonorrheae. In preclinical studies, a wide safety margin supported progression to phase 1 studies in healthy volunteers, which showed linear pharmacokinetics, good oral bioavailability, and no significant safety findings. In a phase 2 study, zoliflodacin was effective in treating gonococcal urogenital and rectal infections. In partnership with the Global Antibiotic Research Development Program (GARDP), zoliflodacin is currently being studied in a global phase 3 clinical trial. Zoliflodacin represents a promising new oral therapy for drug-resistant infections caused by N. gonorrheae[2].

AZD0914 is a new spiropyrimidinetrione bacterial DNA gyrase/topoisomerase inhibitor with potent in vitro antibacterial activity against key Gram-positive (Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus pneumoniae, Streptococcus pyogenes, and Streptococcus agalactiae), fastidious Gram-negative (Haemophilus influenzae and Neisseria gonorrhoeae), atypical (Legionella pneumophila), and anaerobic (Clostridium difficile) bacterial species, including isolates with known resistance to fluoroquinolones. AZD0914 works via inhibition of DNA biosynthesis and accumulation of double-strand cleavages; this mechanism of inhibition differs from those of other marketed antibacterial compounds. AZD0914 stabilizes and arrests the cleaved covalent complex of gyrase with double-strand broken DNA under permissive conditions and thus blocks religation of the double-strand cleaved DNA to form fused circular DNA. Whereas this mechanism is similar to that seen with fluoroquinolones, it is mechanistically distinct. AZD0914 exhibited low frequencies of spontaneous resistance in S. aureus, and if mutants were obtained, the mutations mapped to gyrB. Additionally, no cross-resistance was observed for AZD0914 against recent bacterial clinical isolates demonstrating resistance to fluoroquinolones or other drug classes, including macrolides, β-lactams, glycopeptides, and oxazolidinones. AZD0914 was bactericidal in both minimum bactericidal concentration and in vitro time-kill studies. In in vitro checkerboard/synergy testing with 17 comparator antibacterials, only additivity/indifference was observed. The potent in vitro antibacterial activity (including activity against fluoroquinolone-resistant isolates), low frequency of resistance, lack of cross-resistance, and bactericidal activity of AZD0914 support its continued development[1].

Until recently, the lack of a robust animal model for N. gonorrheae has hampered the ability to understand the PK/pharmacodynamic (PD) and exposure target for this pathogen. Successful colonization is often difficult to achieve and maintain within a therapeutic window due to spontaneous eradication in the host system, which limits the utility of this model in the evaluation of new antibiotics. In vitro dynamic model systems such as the hollow-fiber used to determine the PK/PD driver and evaluate clinic dose regimens have only recently shown successful use with N. gonorrheae. Current therapies and regimens utilized clinically have often relied upon anecdotal evidence provided by clinical use of available agents, demonstrating potent susceptibility to the pathogen. Due to the similarity in the mechanism of action of zoliflodacin to other market topoisomerase inhibitors that have successfully been used to treat gonorrhea, a surrogate pathogen approach with S. aureus was utilized for zoliflodacin. The translation of experimentally derived PK/PD targets was based upon the premise that unbound plasma targets obtained from reference targets in the murine models matched those associated with clinical efficacy. Although the PK/PD driver associated with topoisomerase inhibitors has historically been the area under the concentration curve (AUC)/MIC, this correlation was confirmed with zoliflodacin itself in a series of in vitro hollow-fiber experiments in which unbound drug concentration vs time profiles were simulated and dose fractionated to distinguish among PK/PD drivers of AUC/MIC, Cmax/AUC, and T > MIC. The results of this study utilizing a methicillin-susceptible strain of S. aureus (ARC516) demonstrated AUC/MIC to be the index most closely associated with the activity of zoliflodacin, [R2] = 0.95 (Figure 4).[2]

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With the PK/PD driver established for zoliflodacin against S. aureus in the hollow-fiber model, assessment of the in vivo efficacy of zoliflodacin was established using a murine thigh infection model in both immunocompetent and immunosuppressed mice using a methicillin-susceptible strain of S. aureus (ARC516). Zoliflodacin was highly efficacious in a dose-dependent fashion for both mouse models. Stasis was achieved at an AUC of 20 μg·h/mL. Accounting for the protein binding of zoliflodacin in the mouse (81% bound), a maximum 3 log10 decrease in the CFU/g bacterial burden corresponded to an unbound AUC of 23 μg·h/mL for neutropenic animals. S. aureus ARC516 used for this study had a zoliflodacin MIC of 0.0625 μg/mL with an fAUC/MIC of 368 for >3 log10 kill. Additional S. aureus strains were studied in vivo in the neutropenic thigh infection model including MRSA strains ATCC 33591, USA100 NRS382, and USA300 NR538 in order to establish mean unbound exposure requirements associated with stasis and 1 log10 kill. From the combined in vivo data set, the fAUC/MIC values associated with stasis and 1 log10 kill were 66 and 139, respectively. These values were ultimately used as PK/PD-derived exposure targets for clinical dose justification. The PK/PD index of fAUC/MIC generated in the hollow fiber with zoliflodacin against S. aureus and unbound exposure magnitudes derived from in vivo neutropenic thigh models conducted in mice were subsequently utilized in the surrogate pathogen approach to help establish dose ranges for clinical development with N. gonorrheae. To support the approach, fAUC/MIC magnitudes associated with clinical doses to treat N. gonorrheae and skin infections caused by S. aureus were obtained for reference compounds ofloxacin and ciprofloxacin as shown in Table 6. [2]

[1]. In vitro antibacterial activity of AZD0914, a new spiropyrimidinetrione DNA gyrase/topoisomerase inhibitor with potent activity against Gram-positive, fastidious Gram-Negative, and atypical bacteria. Antimicrob Agents Chemother. 2015 Jan;59(1):467-74.

[2]. Zoliflodacin: An Oral Spiropyrimidinetrione Antibiotic for the Treatment of Neisseria gonorrheae, Including Multi-Drug-Resistant Isolates. ACS Infect. Dis. 2020, 6, 6, 1332–1345

Zoliflodacin has been used in trials studying the basic science and treatment of Gonorrhoea.
In response to the unmet medical need of multi-drug-resistant N. gonorrheae, zoliflodacin (formerly AZD0914, ETX0914), representing a new class of antibacterial agents called the spiropyrimidinetriones, was developed for the treatment of uncomplicated gonorrhea. Zoliflodacin is an orally bioavailable antibacterial agent that is bactericidal via a novel mechanism of action against bacterial type II topoisomerases. This review presents the preclinical and early clinical data that has been generated for this new oral antibiotic.[2]

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Zoliflodacin, a novel spiropyrimidinetrione antibiotic, is being developed by Entasis Therapeutics for the treatment of uncomplicated gonorrhea and represents the first drug in a novel class of topoisomerase inhibitors. Because there is no cross-resistance with fluoroquinolones due to its unique mechanism of action, zoliflodacin could be effective in treating infections caused by fluoroquinolone-resistant strains of N. gonorrheae. This was supported by the safety and efficacy demonstrated during a phase 2 study. Entasis has partnered with the Global Antibiotic Research Development Program (GARDP) to continue the development of zoliflodacin, and the initiation of a global phase 3 study was recently announced (https://gardp.org/news-resources/gardp-and-entasis-therapeutics-initiate-global-phase-3-trial-of-zoliflodacin-a-first-in-class-oral-antibiotic-for-the-treatment-of-gonorrhoea/). This is an open-label trial comparing a single 3 g oral dose of zoliflodacin to a combination of ceftriaxone and azithromycin (2:1 randomization). Similar to the phase 2 study, the phase 3 trial has urogenital infections caused by N. gonorrheae as the criterion for enrollment. However, the presence of extraurogenital infections will also be studied as a secondary end point in the hope that this data will also inform us as to the utility of zoliflodacin in these infections. The clinical study is expected to include approximately 1000 adults with urogenital gonorrhea from sites in the United States, The Netherlands, Thailand, and South Africa (ClinicalTrials.gov identifier NCT03959527), and results are expected in 2021. As we move into the 2020s, a positive outcome of the trial would indicate that zoliflodacin could be used as an oral monotherapy for the treatment of gonorrhea.[2]

C22H22N5O7F

487.438

487.15

C, 54.21; H, 4.55; F, 3.90; N, 14.37; O, 22.98

1620458-09-4

76685216

White to off-white solid powder

1.6±0.1 g/cm3

1.685

1.77

O=C(NC(C12[C@]([C@H](C)O[C@H](C)C3)([H])N3C4=C(C=C5C(ON=C5N6C(OC[C@@H]6C)=O)=C4F)C2)=O)NC1=O

ZSWMIFNWDQEXDT-ZESJGQACSA-N

InChI=1S/C22H22FN5O7/c1-8-7-33-21(32)28(8)17-12-4-11-5-22(18(29)24-20(31)25-19(22)30)16-10(3)34-9(2)6-27(16)14(11)13(23)15(12)35-26-17/h4,8-10,16H,5-7H2,1-3H3,(H2,24,25,29,30,31)/t8-,9+,10-,16+/m0/s1

(4'R,6'S,7'S)-17'-fluoro-4',6'-dimethyl-13'-[(4S)-4-methyl-2-oxo-1,3-oxazolidin-3-yl]spiro[1,3-diazinane-5,8'-5,15-dioxa-2,14-diazatetracyclo[8.7.0.02,7.012,16]heptadeca-1(17),10,12(16),13-tetraene]-2,4,6-trione

Zoliflodacin; ETX0914; AZD-0914; ETX-0914; AZD0914; ETX 0914; AZD 0914; AZD0914; AZD-0914; Zoliflodacin [USAN]; FWL2263R77;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO: ~140 mg/mL (~287.2 mM)

Solubility in Formulation 1: ≥ 2.08 mg/mL (4.27 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.08 mg/mL (4.27 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.08 mg/mL (4.27 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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 (Please use freshly prepared in vivo formulations for optimal results.)