Quinolone
Enoxacin targets bacterial DNA gyrase and topoisomerase IV, enzymes essential for bacterial DNA replication. By inhibiting these enzymes, it interferes with DNA replication and exerts its antibacterial activity. It also targets TAR RNA-binding protein 2 (TRBP)-mediated microRNA processing, acting as a cancer-specific growth inhibitor.

Enoxacin hydrate (also known as Enoxacin sesquihydrate) improves in a dose-dependent manner, with a median effective concentration (EC50) of approximately 30 µM, siGFP-mediated gene knockdown mediated by siRNA against EGFP in HEK293 cells-based reporter system, while having no effect on the cells expressing GFP alone. In HEK293 cells, enoxacin (50 µM) facilitates siRNA duplex loading onto RISCs and miRNA processing[3].
Enoxacin has no impact on the way that Dicer alone processes pre-let-7 or pre-miR-30a. On the other hand, the combination of Enoxacin and TRBP can improve let-7 or pre-miR-30a processing[3].
Enoxacin inhibits 90% of the following microorganisms: Escherichia coli, Aeromonas sp., Enterobacter spp., Serratia spp., Proteus mirabilis, and Morganella morganii at less than or equal to 0.8 micrograms/ml[5].
In vitro, Enoxacin hydrate is a potent inhibitor of bacterial DNA gyrase (IC50=126 µg/mL) and topoisomerase IV (IC50=26.5 µg/mL). It has potent activities against gram-positive and -negative bacteria. It is a miRNA processing activator that enhances siRNA-mediated mRNA degradation and promotes endogenous miRNA biogenesis. It is a cancer-specific growth inhibitor that acts by enhancing TRBP-mediated microRNA processing.

Enoxacin hydrate (Enoxacin sesquihydrate; 100 µM; 2 µl; injected into ear once a day for 3 consecutive days (days 12, 13 and 14)) improves the efficiency of Lv-siGFP-induced GFP mRNA knockdown (from 80% to 60%; 40% GFP mRNA level remained), while alone has no effect on GFP expression in the GFP transgenic line C57BL/6-Tg(ACTB-EGFP)1Osb/J (10 d old) using lentivirus expressing shGFP (Lv-siGFP; injected into ear for 10 days)[3].
In vivo, Enoxacin has been used clinically for the treatment of urinary tract infections, respiratory tract infections, and sexually transmitted diseases. It is a broad-spectrum 6-fluoronaphthyridinone antibacterial agent. Its antibacterial efficacy in vivo is well-documented. Detailed dose-response and pharmacokinetic data are available in the clinical literature.

For cell-free assays, the inhibitory activity of Enoxacin against DNA gyrase and topoisomerase IV can be measured using standard enzyme activity assays. DNA gyrase activity can be assessed by measuring the relaxation of supercoiled DNA. Topoisomerase IV activity can be assessed by measuring DNA decatenation. The compound is incubated with the purified enzyme and DNA substrate, and the inhibition of enzyme activity is measured. IC50 values are calculated from dose-response curves.

For in vitro cellular assays, the antibacterial activity of Enoxacin is assessed in bacterial cultures using broth microdilution methods to determine MIC values. Its cancer-specific growth inhibitory activity can be assessed in cancer cell lines by measuring cell viability using MTT or SRB assays. Its effect on microRNA processing can be assessed by measuring miRNA levels by qPCR.

In vivo, Enoxacin is administered orally for the treatment of bacterial infections. Its efficacy is evaluated in animal models of infection or in clinical trials. In animal models, endpoints include reduction in bacterial load and survival.

Absorption, Distribution and Excretion
Enoxacin is rapidly absorbed after oral administration, with an absolute oral bioavailability of approximately 90%. Metabolism/Metabolites Metabolized in the liver. Enoxacin inhibits certain isoenzymes of the hepatic microsomal cytochrome P-450 enzyme system. Following a single dose, over 40% of the drug is excreted unchanged in the urine within 48 hours. Biological Half-Life The plasma half-life is 3 to 6 hours.
Enoxacin hydrate (CAS 84294-96-2) has a molecular formula of C15H17FN4O3·1.5H2O and a molecular weight of 347.34 g/mol. It is also known as Enoxacin sesquihydrate. Appearance: typically a powder. Solubility: DMSO and other solvents. Storage: typical for fluoroquinolones (desiccated, protected from light, room temperature). Purity: typically >98% for research use.

Medication Use During Pregnancy and Lactation ◉ Overview of Medication Use During Lactation
Traditionally, fluoroquinolones are not recommended for use in infants due to concerns about adverse effects on the developing joints of infants. However, recent studies suggest the risk is minimal. Calcium in breast milk may prevent the absorption of small amounts of fluoroquinolones in breast milk, but there is currently insufficient data to confirm or refute this claim. Enoxacin may be acceptable for breastfeeding women, but close monitoring of the infant's gut microbiota is necessary to prevent adverse reactions such as diarrhea or candidiasis (thrush, diaper rash). However, it is best to use alternative medications with available safety information.
◉ Effects on Breastfed Infants
No published information found as of the revision date.
◉ Effects on Lactation and Breast Milk
No published information found as of the revision date.

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Protein Binding
In healthy subjects, enoxacin binds to plasma proteins at a rate of approximately 40%; in patients with impaired renal function, the binding rate is approximately 14%.
Enoxacin has a well-established clinical safety profile. Common side effects include gastrointestinal disturbances, headache, and dizziness. Less common but serious side effects include tendon rupture and peripheral neuropathy. It is contraindicated in children and pregnant women due to effects on cartilage development.

[1]. Small molecule enoxacin is a cancer-specific growth inhibitor that acts by enhancing TAR RNA-binding protein 2-mediated microRNA processing. Proc Natl Acad Sci U S A. 2011 Mar 15;108(11):4394-9.

[2]. Target preference of 15 quinolones against Staphylococcus aureus, based on antibacterial activities and target inhibition. Antimicrob Agents Chemother. 2001 Dec;45(12):3544-7.

[3]. A small molecule enhances RNA interference and promotes microRNA processing. Nat Biotechnol. 2008 Aug;26(8):933-40.

[4]. Small molecules with big roles in microRNA chemical biology and microRNA-targeted therapeutics. RNA Biol. 2019 Jun;16(6):707-718.

[5]. Chin, N.-X. and H.C. Neu, In vitro activity of enoxacin, a quinolone carboxylic acid, compared with those of norfloxacin, new beta-lactams, aminoglycosides, and trimethoprim. Antimicrobial agents and chemotherapy, 1983. 24(5): p. 754-763.

Enoxacin is a 1,8-naphthidine derivative with the structure 1,4-dihydro-1,8-naphthidine, with an ethyl group at position 1, a carboxyl group at position 3, an oxygen substituent at position 4, a fluorine substituent at position 5, and a piperazine-1-yl group at position 7. It is an antibacterial drug used to treat urinary tract infections and gonorrhea. Enoxacin has antibacterial and DNA synthesis-inhibiting effects. It is a monocarboxylic acid, amino acid, 1,8-naphthidine derivative, N-arylpiperazine, quinolone antibiotic, and fluoroquinolone antibiotic.
A broad-spectrum 6-fluoronaphthidine ketone antibacterial agent (fluoroquinolone), its structure is related to nalidixic acid.
A broad-spectrum 6-fluoronaphthidine ketone antibacterial agent, its structure is related to nalidixic acid.
See also: Enoxacin sesquihydrate (its active moiety).

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Indications
For the treatment of the following infections caused by susceptible strains of specified microorganisms in adults (≥18 years): (1) uncomplicated urethral or cervical gonorrhea caused by Neisseria gonorrhoeae; (2) uncomplicated urinary tract infections (cystitis) caused by Escherichia coli; Staphylococcus epidermidis or Staphylococcus saprophyticus; and (3) complicated urinary tract infections caused by Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis, Pseudomonas aeruginosa, Staphylococcus epidermidis, or Enterobacter cloacae.

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Mechanism of Action
Enoxacin exerts its bactericidal effect by inhibiting the bacterial essential enzyme DNA gyrase (DNA topoisomerase II).

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Pharmacodynamics
Enoxacin is a quinolone/fluoroquinolone antibiotic. Enoxacin has a bactericidal effect by binding to an enzyme called DNA gyrase to block bacterial DNA replication, thereby preventing the DNA double helix from unwinding and thus preventing DNA replication into two double helices. Enoxacin is a broad-spectrum antibiotic effective against both Gram-positive and Gram-negative bacteria. However, enoxacin may induce resistance to drugs with different mechanisms of action.
Enoxacin is an approved antibiotic in many countries for the treatment of urinary tract infections, respiratory tract infections, and sexually transmitted diseases. Its mechanism of action involves inhibition of bacterial DNA gyrase and topoisomerase IV. It is also being studied for its cancer-specific growth inhibitory activity through enhancement of microRNA processing.

C₁₅H₁₇FN₄O₃.₃/₂H₂O

347.34

320.128

84294-96-2

84294-96-2 (Enoxacin hydrate;Enoxacin sesquihydrate; AT-2266 hydrate; CI-919 hydrate); 74011-58-8 (free)

3229

White to yellow solid powder

226 °C

0.992

2

8

3

23

521

0

IDYZIJYBMGIQMJ-UHFFFAOYSA-N

InChI=1S/C15H17FN4O3/c1-2-19-8-10(15(22)23)12(21)9-7-11(16)14(18-13(9)19)20-5-3-17-4-6-20/h7-8,17H,2-6H2,1H3,(H,22,23)

1-ethyl-6-fluoro-4-oxo-7-piperazin-1-yl-1,8-naphthyridine-3-carboxylic acid

Enoxacin sesquihydrate; AT-2266 hydrate; CI-919 hydrate; Enoxacin hydrate

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture and light.

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

1M NaOH: ~100 mg/mL (~287.9 mM)
DMSO: ~2.78 mg/mL (~8.0 mM)
H2O: ~1 mg/mL (~2.9 mM)

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*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.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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