 Chemical Structure.png)
Avibactam (NXL104; NXL-104; Avycaz) is a novel, covalent but reversible inhibitor of β-lactamase belonging to the the non-β-lactam antibiotic, inhibiting β-lactamase TEM-1 and CTX-M-15 with IC50s of 8 nM and 5 nM, respectively. It is a component of the approved drug combination (avibactam + ceftazidime; trade name Avycaz), which was approved by the FDA on February 25, 2015, for treating complicated urinary tract and complicated intra-abdominal Infections caused by antibiotic resistant-pathogens, including those caused by multi-drug resistant gram-negative bacterial pathogens.
Physicochemical Properties
| Molecular Formula | C7H11N3O6S |
| Molecular Weight | 265.24374 |
| Exact Mass | 265.036 |
| Elemental Analysis | C, 31.70; H, 4.18; N, 15.84; O, 36.19; S, 12.09 |
| CAS # | 1192500-31-4 |
| Related CAS # | Avibactam sodium;1192491-61-4;Avibactam sodium hydrate;2938989-90-1;Avibactam sodium dihydrate |
| PubChem CID | 9835049 |
| Appearance | White to off-white solid powder |
| Density | 1.9±0.1 g/cm3 |
| Index of Refraction | 1.679 |
| LogP | -3.2 |
| Hydrogen Bond Donor Count | 2 |
| Hydrogen Bond Acceptor Count | 6 |
| Rotatable Bond Count | 3 |
| Heavy Atom Count | 17 |
| Complexity | 457 |
| Defined Atom Stereocenter Count | 2 |
| SMILES | O=S(ON1[C@]2([H])CC[C@@H](C(N)=O)[N@@](C2)C1=O)(O)=O |
| InChi Key | NDCUAPJVLWFHHB-UHNVWZDZSA-N |
| InChi Code | InChI=1S/C7H11N3O6S/c8-6(11)5-2-1-4-3-9(5)7(12)10(4)16-17(13,14)15/h4-5H,1-3H2,(H2,8,11)(H,13,14,15)/t4-,5+/m1/s1 |
| Chemical Name | (2S,5R)-2-carbamoyl-7-oxo-1,6-diazabicyclo[3.2.1]octan-6-yl hydrogen sulfate |
| Synonyms | Avibactam Free Acid; NXL-104; NXL104; Avibactam; 1192500-31-4; Avibactam free acid; AVE-1330A free acid; Avibactam (free acid); Nxl-104 free acid; [(2S,5R)-2-carbamoyl-7-oxo-1,6-diazabicyclo[3.2.1]octan-6-yl] hydrogen sulfate; NXL 104 |
| HS Tariff Code | 2934.99.9001 |
| Storage |
Powder-20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
| Shipping Condition | Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs) |
Biological Activity
| Targets | CTX-M-15(IC50=5 nM);TEM-1(IC50=8 nM ) |
| ln Vitro |
Monetary antibacterial activity is low for avibactam, which inhibits class A and C β-lactamases but not metallotypes or Acinetobacter OXA carbapenemases[2]. With MIC50 and MIC90 for both 8 mg/L, ceftazidime (HY-B0593)-avibactam (0-256 mg/L) inhibits the growth of 16 blaKPC-2 positive and 1 blaOXA-232 positive Klebsiella pneumonia[4]. |
| ln Vivo | Ceftazidime-Avibactam (0.375 mg/g; s.c.; every 8 hours for 10 days) significantly affects the bacteria and has been shown to have some therapeutic efficacy in an infected mouse model with K. pneumoniae strain Y8[3]. Avibactam (64 mg/kg; s.c.; once) infected neutropenic mice with lung infection exhibits a mean estimated half-life in plasma in the terminal phase of 0.24 h[3]. |
| Enzyme Assay | In a 200 μL reaction volume, 1 μM TEM-1 is incubated with and without 5 μM Avibactam for 5 min at 37°C and subjected to two ultrafiltration cartridge (UFC) steps to remove excess inhibitor (Ultrafree-0.5 with Biomax membrane, 5-kDa cutoff). Centrifugation at 10,600× g for 8 min is performed at 4°C. After each ultrafiltration step, 20 μL retentate are diluted with 180 μL assay buffer to restore the original enzyme concentration. After two UFC treatments, the amount of free Avibactam is quantified by liquid chromotography/MS/MS and found to be<5% of the original concentration. Loss of protein during UFC is assessed by measuring TEM-1 activity (on 4,000-fold dilution) in the acyl-enzyme sample compare with non-UFC-treated enzyme, and loss is found to be <5%[1]. |
| Cell Assay |
Cells (~109 cfu) from overnight broth culture are spread on Mueller-Hinton agar supplemented with either (i) Ceftaroline plus Avibactam (1 or 4 mg/L) at 1-16× the MICs or (ii) Ceftaroline at 1 or 4 mg/L plus Avibactam at 1-8× the concentration needed to reduce the Ceftaroline MIC to 1 or 4 mg/L. Colonies are counted after overnight incubation and representatives are retained[2].
The microdilution broth method was performed to analyze the minimal inhibitory concentration (MIC). The time-kill curve assay of ceftazidime-avibactam at various concentrations was conducted in 16 strains of KPC-2 and 1 strain of OXA-232 carbapenemase-producing Klebsiella pneumoniae. The in vitro synergistic bactericidal effect of ceftazidime-avibactam combined with aztreonam was determined by checkerboard assay on 28 strains of NDM and 2 strains of NDM coupled with KPC carbapenemase-producing Klebsiella pneumoniae. According to calculating grade, the drugs with synergistic bactericidal effect were selected as an inhibitory concentration index. The in vitro bactericidal tests of ceftazidime-avibactam combined with aztreonam were implemented on 12 strains among them.[3] Objectives: Ceftaroline + avibactam (NXL104) is a novel inhibitor combination active against Enterobacteriaceae with class A and C β-lactamases. We investigated its risk of mutational resistance. Methods: Single- and multi-step mutants were sought and characterized from Enterobacteriaceae with extended-spectrum β-lactamases (ESBLs), AmpC β-lactamases and KPC β-lactamases. Results: Overgrowth occurred on agar with low MIC multiples of ceftaroline + avibactam, but frequencies for single-step mutants were <10(-9). Most mutants were unstable, with only three remaining resistant on subculture. For one, from an CTX-M-15-positive Escherichia coli, the ceftaroline + avibactam MIC was raised, but the organism had reduced resistance to ceftaroline and lost resistance to other oxyimino-cephalosporins, with this profile retained when the mutant bla(CTX-M-15) was cloned into E. coli DH5α. Sequencing identified a Lys237Gln substitution in the CTX-M-15 variant. The other two stable single-step mutants were from an AmpC-derepressed Enterobacter cloacae strain; these had unaltered or slightly reduced resistance to other β-lactams. Both had amino acids 213-226 deleted from the Ω loop of AmpC. Further stable mutants were obtained from AmpC-inducible and -derepressed E. cloacae in multi-step selection, and these variously had reduced expression of OmpC and OmpF, and/or Asn366His/Ile substitutions in AmpC. Conclusions: Stable resistant mutants were difficult to select. Those from AmpC-derepressed E. cloacae had porin loss or AmpC changes, including Ω loop deletions. A Lys237Gln substitution in CTX-M-15 conferred resistance, but largely abolished ESBL activity.[2] |
| Animal Protocol |
Animal Model: Six-week-old BALB/c mice (female), K. pneumoniae strain Y8 infection model[4] Dosage: 0.375 mg/g in combination with Ceftazidime Administration: Subcutaneous injection given every 8 hours for 10 days starting 4 hours after infection Result: Within 4 days, 70% of the mice in the infection group perished, and in 13 days, every mouse in the PBS group perished. When the antibiotic was given every eight hours for ten days after infection, all of the mice in the treatment group survived; however, when the antibiotic treatment was stopped, all of the mice in the control group perished in four days. When compared to the infected group, the treatment group mice's liver and spleen had reduced CFU counts. |
| ADME/Pharmacokinetics |
Absorption, Distribution and Excretion Avibactam and ceftazidime are excreted mainly by the kidneys. The steady state volumes of distribution of avibactam and ceftazidime is 22.2L and 17L respectively. Avibactam and ceftazidime has a clearance of ~12L/h and ~7L/h respectively. Metabolism / Metabolites No metabolism of avibactam was observed in human liver preparations. Unchanged avibactam is the major drug-related component in human plasma and urine. 80-90% of ceftazidime is eliminated as unchanged . Biological Half-Life Ceftazidime-avibactam has a half life of ~2.7-3.0 hours. |
| Toxicity/Toxicokinetics |
Protein Binding 5.7%-8.2% of avibactam is bound to plasma protein, and less than 10% of ceftazidime is protein bound. |
| References |
[1]. Avibactam is a covalent, reversible, non-β-lactam β-lactamase inhibitor. Proc Natl Acad Sci U S A. 2012 Jul 17;109(29):11663-8. [2]. Characterization of β-lactamase and porin mutants of Enterobacteriaceae selected with ceftaroline + avibactam (NXL104). J Antimicrob Chemother. 2012 Jun;67(6):1354-8. [3]. WO 2017104691 A1. [4]. In vitro and in vivo bactericidal activity of ceftazidime-avibactam against Carbapenemase-producing Klebsiella pneumoniae. Antimicrob Resist Infect Control. 2018 Nov 21;7:142. |
| Additional Infomation |
Avibactam is a member of the class of azabicycloalkanes that is (2S,5R)-7-oxo-1,6-diazabicyclo[3.2.1]octane-2-carboxamide in which the amino hydrogen at position 6 is replaced by a sulfooxy group. Used (in the form of its sodium salt) in combination with ceftazidime pentahydrate for the treatment of complicated urinary tract infections including pyelonephritis. It has a role as an antibacterial drug, an antimicrobial agent and an EC 3.5.2.6 (beta-lactamase) inhibitor. It is a monocarboxylic acid amide, a member of ureas, an azabicycloalkane and a hydroxylamine O-sulfonic acid. It is a conjugate acid of an avibactam(1-). Avibactam is a non-β-lactam β-lactamase inhibitor that is available in combination with ceftazidime (Avycaz). This combination was approved by the FDA on February 25, 2015 for the treatment of complicated intra-abdominal infections in combination with metronidazole, and the treatment of complicated urinary tract infections, including pyelonephritis caused by antibiotic resistant-pathogens, including those caused by multi-drug resistant gram-negative bacterial pathogens. As there is limited clinical safety and efficacy data, Avycaz should be reserved for patients over 18 years old who have limited or not alternative treatment options. Avibactam is a beta Lactamase Inhibitor. The mechanism of action of avibactam is as a beta Lactamase Inhibitor. Drug Indication AVYCAZ (ceftazidime-avibactam), in combination with metronidazole, is indicated for the treatment of complicated intra-abdominal infections caused by the following susceptible microorganisms: Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis, Providencia stuartii, Enterobacter cloacae, Klebsiella oxytoca, and Pseudomonas aeruginosa in patients 18 years or older. AVYCAZ is also indicated for the treatment of complicated urinary tract infections including pyelonephritis caused by the following susceptible microorganisms: Escherichia coli, Klebsiella pneumoniae, Citrobacter koseri, Enterobacter aerogenes, Enterobacter cloacae, Citrobacter freundii, Proteus spp., and Pseudomonas aeruginosa in patients 18 years or older. FDA Label Mechanism of Action Avibactam is a non-β lactam β-lactamase inhibitor that inactivates some β-lactamases (Ambler class A β-lactamases, including Klebsiella pneumoniae carbapenemases, Ambler class C and some Ambler class D β-lactamases) by a unique covalent and reversible mechanism, and protects ceftazidime from degradation by certain β-lactamases. Avibactam rapidly reaches the periplasm of bacteria at high enough concentrations to restore activity of ceftazidime against ceftazidime-resistant, β-lactamase-producing strains. Avibactam does not decrease the activity of ceftazidime against ceftazidime susceptible organisms. Avibactam is a β-lactamase inhibitor that is in clinical development, combined with β-lactam partners, for the treatment of bacterial infections comprising gram-negative organisms. Avibactam is a structural class of inhibitor that does not contain a β-lactam core but maintains the capacity to covalently acylate its β-lactamase targets. Using the TEM-1 enzyme, we characterized avibactam inhibition by measuring the on-rate for acylation and the off-rate for deacylation. The deacylation off-rate was 0.045 min(-1), which allowed investigation of the deacylation route from TEM-1. Using NMR and MS, we showed that deacylation proceeds through regeneration of intact avibactam and not hydrolysis. Other than TEM-1, four additional clinically relevant β-lactamases were shown to release intact avibactam after being acylated. We showed that avibactam is a covalent, slowly reversible inhibitor, which is a unique mechanism of inhibition among β-lactamase inhibitors.[1] Background: In recent years, the incidence of carbapenem-resistant Enterobacteriaceae (CRE) infections has increased rapidly. Since the CRE strain is usually resistant to most of antimicrobial agents, patients with this infection are often accompanied by a high mortality. Therefore, it instigates a severe challenge the clinical management of infection. In this study, we study the in vitro and in vivo bactericidal activity of ceftazidime-avibactam administrated either alone or in combination with aztreonam against KPC or NDM carbapenemase-producing Klebsiella pneumoniae, and explore a new clinical therapeutic regimen for infections induced by their resistant strains. [3] Methods: The microdilution broth method was performed to analyze the minimal inhibitory concentration (MIC). The time-kill curve assay of ceftazidime-avibactam at various concentrations was conducted in 16 strains of KPC-2 and 1 strain of OXA-232 carbapenemase-producing Klebsiella pneumoniae. The in vitro synergistic bactericidal effect of ceftazidime-avibactam combined with aztreonam was determined by checkerboard assay on 28 strains of NDM and 2 strains of NDM coupled with KPC carbapenemase-producing Klebsiella pneumoniae. According to calculating grade, the drugs with synergistic bactericidal effect were selected as an inhibitory concentration index. The in vitro bactericidal tests of ceftazidime-avibactam combined with aztreonam were implemented on 12 strains among them. Effect of ceftazidime-avibactam antibiotic against KPC carbapenemase-producing K. pneumoniae strain Y8 Infection was performed in the mouse model. [3] Results: The time-kill assays revealed that ceftazidime-avibactam at various concentrations of 2MIC, 4MIC and 8MIC showed significant bactericidal efficiency to the resistant bacteria strains. However, in 28 strains of NDM and 2 strains of NDM coupled with KPC carbapenemase- producing Klebsiella pneumoniae, only 7 strains appeared the susceptibility to ceftazidime-avibactam treatment, MIC50 and MIC90 were 64 mg/L and 256 mg/L, respectively. Antimicrobial susceptibility testing of ceftazidime-avibactam combined with aztreonam disclosed the synergism of two drugs in 90% (27/30) strains, an additive efficiency in 3.3% (1/30) strains, and irrelevant effects in 6.6% (2/30) strains. No antagonism was found. The subsequent bactericidal tests also confirmed the results mentioned above. Therapeutic efficacy of Ceftazidime-Avibactam against K. pneumoniae strain Y8 infection in mouse indicated 70% of infection group mice died within 4 days, and all mice in this group died within 13 days. Bacterial load testing results showed that there was no significant difference in the amount of bacteria in the blood between the infected group and the treatment group. However, the spleen and liver of treatment group mice showed lower CFU counts, as compare with infected group, indicating that ceftazidime-avibactam has a significant effect on the bacteria and led to a certain therapeutic efficacy. [3] Conclusion: This study indicated ceftazidime-avibactam therapy occupied significant bactericidal effects against KPC-2 and OXA-232 carbapenemase-producing Klebsiella pneumoniae. While combined with aztreonam, the stronger synergistic bactericidal effects against NDM carbapenemase-producing Klebsiella pneumoniae were achieved.[3] |
Solubility Data
| Solubility (In Vitro) | DMSO : ~125 mg/mL (~471.27 mM) |
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.08 mg/mL (7.84 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. Solubility in Formulation 2: ≥ 2.08 mg/mL (7.84 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. Solubility in Formulation 3: ≥ 2.08 mg/mL (7.84 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. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 3.7702 mL | 18.8509 mL | 37.7017 mL | |
| 5 mM | 0.7540 mL | 3.7702 mL | 7.5403 mL | |
| 10 mM | 0.3770 mL | 1.8851 mL | 3.7702 mL |