Nitroimidazole Derivatives Research Articles

Methoxy poly (ethylene glycol)-block-poly (glutamic acid)-graft-6-(2-nitroimidazole) hexyl amine nanoparticles for potential hypoxia-responsive delivery of doxorubicin

Tumor microenvironment-responsive nano drug delivery vehicles are gaining mounting attention in the field of biomedical sciences. The hypoxic response of the tumorous cells due to very low partial pressure of oxygen (some time less than 2.5 mm of Hg) in the tumor tissues makes hypoxia-responsive drug delivery system as the more appealing in cancer chemotherapy. Based on these considerations, we synthesized hypoxia-responsive polymeric materials methoxy poly (ethylene glycol)-block-poly (glutamic acid)-graft-6-(2-nitroimidazole) hexyl amine (mPEG-b-PLG-g-NID) by conjugation of the hydrophobic nitro imidazole derivative (NID)[6-(2-nitroimidazole) hexyl amine] with the pendant carboxylic group of poly (ethylene glycol)-block-poly (L-glutamic acid)(mPEG-b-PLG). The structure and degree of substitution were confirmed by proton NMR, FTIR, and UV–Vis spectroscopy. The degree of substitution was found to enhance with the increase in NID to polymer ratio. The hypoxia response of the material was evaluated by UV–Vis spectroscopy and zeta potential measurements. Doxorubicin was hydrophobically encapsulated in the micellar core of the hypoxia-responsive nanoparticles. The drug-loaded micelles showed faster release in hypoxic condition as compared to normoxic conditions. Moreover, the developed polymeric system was found non-toxic to MCF-7 cell line, thus suggesting its biocompatibility and suitability as drug delivery device.

Nitroimidazole-Containing H2dedpa and H2CHXdedpa Derivatives as Potential PET Imaging Agents of Hypoxia with (68)Ga.

(68)Ga is an attractive radiometal for use in positron emission tomography (PET) imaging. The success of (68)Ga-based agents is dependent on a chelator that exhibits rapid radiometal incorporation, and strong kinetic inertness to prevent transchelation of (68)Ga in vivo. The linear chelating agents H2dedpa (1,2-[[6-carboxypyridin-2-yl]methylamino]ethane) and H2CHXdedpa (CHX = cyclohexyl/cyclohexane) (N4O2) have recently been developed that bind Ga(3+) quickly and under mild conditions, ideal properties to be incorporated into a (68)Ga PET imaging agent. Herein, nitroimidazole (NI) derivatives of H2dedpa and H2CHXdedpa to investigate specific targeting of hypoxic tumor cells are investigated, given that NI can be reduced and retained exclusively in hypoxic cells. Nine N,N'-bis-alkylated derivatives of H2dedpa and H2CHXdedpa have been synthesized; they have been screened for their ability to bind gallium, and cyclic voltammetry of nonradioactive complexes was performed to probe the redox cycling mechanism of NI. The compounds were radiolabeled with (67)Ga and (68)Ga and show promising radiolabeling efficiencies (>99%) when labeled at 10(-5) M for 10 min at room temperature. Moreover, stability studies (via apo-transferrin challenge, 37 °C) show that the (67)Ga complexes exhibit exceptional stability (86-99% intact) after 2 h. In vitro uptake studies under hypoxic (0.5% O2) and normoxic (21% O2) conditions in three cancerous cell lines [HT-29 (colon), LCC6(HER-2) (breast), and CHO (Chinese hamster ovarian)] were performed. Of the four H2dedpa or H2CHXdedpa NI derivatives tested, all showed preferential uptake in hypoxic cells compared to normoxic cells with hypoxic/normoxic ratios as high as 7.9 ± 2.7 after 120 min. The results suggest that these novel bis-alkylated NI-containing H2dedpa and H2CHXdedpa ligands would be ideal candidates for further testing in vivo for PET imaging of hypoxia with (68)Ga.

The biological effect of the nitroimidazole derivative of a polypyridyl ruthenium complex on cancer and endothelial cells.

The ruthenium polypyridyl complexes [Ru(dip)2(bpy/bpy-2-nitroIm)](2+) (dip = 4,7-diphenyl-1,10-phenanthroline, bpy = 2,2'-bipyridine, bpy-2-nitroIm = 4-[3-(2-nitro-1H-imidazol-1-yl)propyl]) were found to be ca. ten times more cytotoxic against breast cancer (4T1) and human lung adenocarcinoma epithelial cells (A549) than a well-known anticancer drug, cisplatin. Even though the Ru complexes were quite cytotoxic towards FVB mouse lung microvascular endothelial cells (MLuMEC FVB) their efflux from these non transformed cells was much more efficient than from cancer ones. Both Ru complexes accumulated in cells. The cellular uptake of both Ru complexes occurs through passive diffusion while the nitroimidazole derivative is also endocytosed. They arrest cell growth in the S-phase and induce apoptosis. Such cell response can result from activation of oxidative stress by Ru complexes. The modulation of the mRNA expression profile for genes which might be involved in metastasis and angiogenesis processes by Ru complexes was analyzed for both cancer (4T1) and endothelial (MLuMEC FVB) cells. Ru complexes appeared to have a distinct impact on cell adhesion and migration as well as they affect endothelial cell vasculature. They are not only cytotoxic but are also potentially invasive and anti-metastatic agents. This work illustrates the putative future development of polypyridyl ruthenium.

Multifocal fixed drug eruption due to ornidazole

Ornidazole, a synthetic nitroimidazole derivative, is widely used in clinical practice. Ornidazol has low rate of side effects, but sometimes may cause significant allergic reactions such as fixed drug eruption (FDE). We describe a case of multifocal FDE caused by ornidazole.

Evaluation of saliva levels and therapeutic effects of morinidazole, a new 5-nitroimidazole derivative, in patients with periodontal diseases

The aim of this study was to evaluate and compare the saliva and gingival crevice fluid concentrations, as well as therapeutic effects of morinidazole (MNZ) and ornidazole (ONZ) in patients with periodontitis after intravenous (i.v.) infusion administration. 142 outpatient subjects (80 women and 62 men) were enrolled in the phase II clinical trial, and the patients were randomly assigned to receive intravenous infusions of either Ornidazole or Morinidazole, twice daily for 5 days (500 mg each time, infusion period was 50 min). A saliva was collected prior to and again after finishing the first i.v. dose at intervals of 15, 30, 45, 60, 90 min on day 1. Bacteriological studies were undertaken to determine the effectiveness of the MNZ or ONZ. Reverse phase high performance liquid chromatography (RP-HPLC) method with UV detection at 318 nm was carried out, using a mixture of KH2PO4, acetonitrile and methanol (55:15:30, v/v/v) as a mobile phase, with a flow rate of 1.0 ml/min. There was no significant difference between the MNZ and ONZ groups. The cure rate of morinidazole was found to be slightly higher than that of ONZ, just like the result of concentration in saliva, but no significant differences between the two agents were apparent. During our trial, we found that the clinical resistance of anaerobes to morinidazole was proven to be extremely rare. Key words: Periodontitis, morinidazole, ornidazole, tinidazole, saliva.

A chiral high-performance liquid chromatography–tandem mass spectrometry method for the stereospecific determination of morinidazole in human plasma

Morinidazole is a novel 5-nitroimidazole derivative used for the treatment of amoebiasis, trichomoniasis, and anaerobic bacterial infections. Morinidazole possesses a chiral carbon and is clinically administered as a racemate. In the present study, an enantioselective and sensitive liquid chromatography-tandem mass spectrometry method of determining morinidazole enantiomers in human plasma was developed and validated to characterize the stereoselective pharmacokinetics. Plasma samples were processed by liquid-liquid extraction using tert-butyl methyl ether. Chiral separation was optimized within 8.5min on a cellulose column using an isocratic mobile phase of methanol/water (80:20, v/v). Detection was using mass spectrometry in multiple reaction monitoring mode, using the transitions of m/z 271→144 for morinidazole enantiomers, and m/z 275→148 for d4-morinidazole (internal standard). The calibration curves were linear over 5.00-6000ng/mL for each enantiomer. The lower limit of quantification for each enantiomer was established at 5.00ng/mL. Intra- and inter-day precisions were less than 6.4% for each enantiomer in terms of relative standard deviation, and accuracies were between -2.5% and 6.4% in terms of relative error for each enantiomer. No chiral inversion was observed during sample storage, preparation procedure and analysis. Major glucuronide and sulfate conjugates were not observed to interfere with the determination of morinidazole enantiomers. The method was applied to study the stereoselective pharmacokinetics of morinidazole in humans. Moderate stereoselectivity was observed in healthy subjects and patients with severe renal impairment.

Development and Validation of a Discriminating In Vitro Dissolution Method for Oral Formulations Containing Satranidazole

The development of a meaningful dissolution procedure for drug products with limited water solubility has been a challenge to the pharmaceutical industry. Satranidazole (BCS Class II drug) is a new nitroimidazole derivative with potent antiamoebic action. There is no official dissolution medium available in the literature. In the present study, parameters such as saturation solubility in different pH medium, dissolution behavior of formulations, influence of sink conditions, stability, and discriminatory effect of dissolution testing were studied for the selection of a proper dissolution medium. Results of solubility data revealed that solubility of Satranidazole decreases with an increase in pH. Satranidazole showed better sink condition in 0.1 N HCl as compared to other media. The drug and marketed formulations were stable in the dissolution media used. An agitation speed of 75 rpm showed a more discriminating drug release profile than 50 rpm. Using optimized dissolution parameters (paddle at 75 rpm, 900 mL 0.1 N HCl) greater than 80% of the label amount is released over 60 minutes. UV-spectroscopic method used was validated for the specificity, linearity, precision, robustness, and solution stability. The method was successfully applied to granular formulations and also to marketed tablets containing 300 mg Satranidazole.

Synthesis and preliminary biological evaluation of 99mTc(CO)3-labeled pegylated 2-nitroimidazoles

This work reports the synthesis, radiolabeling and preliminary biodistribution results in tumor-bearing mice of Tc-99m(CO)(3)-labeled pegylated (PEG) 2-nitroimidazoles for tumor hypoxia imaging. The novel 2-nitroimidazole derivatives were successfully synthesized by conjugation of tridendate chelators to 2-nitroimidazole via PEG(3) linker. Radiolabeling was performed in high yield with [Tc-99m(CO)(3)](+) core to get cationic [Tc-99m(CO)(3)(BPA-PEG(3)-NIM)](+), neutral [Tc-99m(CO)(3)(AOPA-PEG(3)-NIM)] and anionic [Tc-99m(CO)(3)(IDA-PEG(3)-NIM)](-) respectively, all of which were hydrophilic and stable at room temperature. Biodistribution studies in tumor-bearing mice showed that Tc-99m(CO)(3)-labeled pegylated 2-nitroimidazoles accumulated in the tumor with low uptake. Tc-99m-chelate and charge had significant impact on partition coefficient, radiotracer tumor uptake and pharmacokinetic properties. The results indicate the need for synthetic modification of the parent 2-nitroimidazole derivatives and the Tc-99m-chelate with a view to improve the tumor targeting efficacy and in vivo kinetic profiles.

Design and synthesis of 2-styryl of 5-Nitroimidazole derivatives and antimicrobial activities as FabH inhibitors

A series of 2-Styryl-5-Nitroimidazole derivatives (25–48) have been synthesized and their biological activities were also evaluated against two Gram-negative bacterial strains: Escherichia coli and Pseudomonas aeruginosa and two Gram-positive bacterial strains: Bacillus subtilis and Bacillus thuringiensis as potential FabH inhibitors. All the compounds were structurally determined by 1H NMR, MS, and elemental analysis. E. coli β-ketoacyl-acyl carrier protein synthase III inhibitory assay and docking simulation indicated that compound 33 with IC50 of 9.0–36.4 μg/mL and compound 47 with IC50 of 6.3–34.3 μg/mL against bacterial strains were most potent inhibitors of E. coli FabH. And more, compounds 33 and 47 which possessed a broad-spectrum of antibacterial activities didn't exhibit any toxicity towards macrophage.

Development and Evaluation of Taste Masked Granular Formulation of Satranidazole by Melt Granulation Technique.

Drugs from nitroimidazole category are generally bitter in taste. Oral formulation with bitter taste is not palatable. Geriatrics and pediatrics patients usually suffer from swallowing difficulties. Many other patients in some disease conditions avoid swallowing tablets. Satranidazole is a new nitro-imidazole derivative with bitter taste and is available in market as film coated tablet. The purpose of this research was to mask the bitter taste of Satranidazole by coating complexation with low melting point wax and Eudragit EPO. Different types of wax (glyceryl monostearate, stearic acid and cetyl alcohol) were tried for taste masking. The drug to stearic acid ratio 1 : 2 was found to be optimum on the basis of taste evaluation and in vitro release. The formulated granules were found to possess good flow property. FTIR studies confirmed that there was no interaction between drug and excipients. Scanning Electron Microscopy of drug and the optimized batch of granules was performed. The in vitro release of drug from granules was compared with marketed tablet formulation. The taste masked granules of optimized batch showed 87.65% release of drug in 1 hr which is comparable to that of marketed tablet formulation.

Development and Validation of Stability Indicating RP-HPLC Method for Estimation of Satranidazole from Its Formulation

Satranidazole is a new nitroimidazole derivative with potent antiamoebic action and is available in market in the form of tablet and dry syrup either alone or in combination with Ofloxacin. The present study involves the development of simple, accurate, precise, and reproducible reversed phase high performance liquid chromatography (RP-HPLC) method for determination of Satranidazole from its granular dosage form. Isocratic elution at a flow rate of 1.0 mL/min was employed on BDS Hypersil C18 (250 mm × 4.6 mm, 5 μm) column at 25°C temperature. The mobile phase consists of 0.16% v/v orthophosphoric acid solution, pH 3: acetonitrile in the ratio of 60 : 40 v/v. The UV detection wavelength was 320 nm, and 20 μL sample was injected. The retention time for Satranidazole was about 4.3 minutes. The method was validated for various parameters such as system suitability, precision, recovery, robustness, and ruggedness as per ICH guidelines. The validated RP-HPLC method was found to be specific, linear, precise, and accurate and can be successfully employed for the assay of Satranidazole taste masked granules coated with Eudragit E100 and marketed tablets.

Influence of the Electron Transfer Rate on Electrochemically Controlled Hydrogen Bonding

Hydrogen bonding is an essential interaction in natural and artificial systems. Its strength can be modulated by employing process known as Electrochemically Controlled Hydrogen bonding (ECHB). Although these processes are assumed to operate under thermodynamic control no experimental evidence for kinetic control exists. In this work, ECHB processes where studied using electrogenerated radical anions from 5-nitroimidazole derivatives as receptor molecules and 1,3-diethylurea as hydrogen bond donor species. Results revealed that kinetic control occurs due to an increase in the internal reorganization energy of the receptor molecule, which cause a decrease in electron transfer rate. Electronic structure calculations and experimental Kb values suggested that kinetic limitations were the product of a competition between intra and intermolecular hydrogen bonding formation during the global process.

Hypoxia-responsive polymeric nanoparticles for tumor-targeted drug delivery

Hypoxia is a condition found in various intractable diseases. Here, we report self-assembled nanoparticles which can selectively release the hydrophobic agents under hypoxic conditions. For the preparation of hypoxia-responsive nanoparticles (HR-NPs), a hydrophobically modified 2-nitroimidazole derivative was conjugated to the backbone of the carboxymethyl dextran (CM-Dex). Doxorubicin (DOX), a model drug, was effectively encapsulated into the HR-NPs. The HR-NPs released DOX in a sustained manner under the normoxic condition (physiological condition), whereas the drug release rate remarkably increased under the hypoxic condition. From in vitro cytotoxicity tests, it was found the DOX-loaded HR-NPs showed higher toxicity to hypoxic cells than to normoxic cells. Microscopic observation showed that the HR-NPs could effectively deliver DOX into SCC7 cells under hypoxic conditions. In vivo biodistribution study demonstrated that HR-NPs were selectively accumulated at the hypoxic tumor tissues. As consequence, drug-loaded HR-NPs exhibited high anti-tumor activity in vivo. Overall, the HR-NPs might have a potential as nanocarriers for drug delivery to treat hypoxia-associated diseases.

PET-MR bei Patienten mit Glioblastoma multiforme

Glioblastoma multiforme (GBM) is the most common and most aggressive primary tumor of the brain. In recent years newer therapeutic approaches have been developed. To allow for an optimized treatment planning it is important to precisely delineate necrotic tissue, edema and vital tumor tissue and to identify the most aggressive parts of the GBM. The magnetic resonance (MR) portion of an MR-positron emission tomography (PET) examination in patients with GBM should consist of both structural and functional sequences including diffusion-weighted and perfusion sequences. The use of (18)F-fluorodeoxyglucose ((18)F-FDG) is limited in patients with gliomas as glucose metabolism is already physiologically high in parts of the brain but (18)F-FDG is nevertheless a commonly used radiopharmaceutical for neuro-oncological questions. (18)F-fluorothymidine reflects the cellular activity of thymidine kinase 1 and correlates with the expression of KI-67 as an index of mitotic activity. The nitroimidazole derivatives (18)F-fluoromisonidazole and (18)F-fluoroazomycin arabinoside ((18)F-FAZA) allow the detection of hypoxic areas within the tumor. In recent years amino acid tracers, such as (18)F-fluoroethyltyrosine are increasingly being used in the diagnosis of gliomas. The simultaneous PET-MR image acquisition allows new approaches, e.g. motion correction by the simultaneous acquisition of MR data with a high temporal resolution and an improved quantification of the PET signal by integrating the results of functional MR sequences. Moreover, the simultaneous acquisition of these two time-consuming methods leads to reduced imaging times for this, often severely ill patient group.

Synthesis and biological evaluation of a novel 99mTc labeled 2-nitroimidazole derivative as a potential agent for imaging tumor hypoxia

Tumor hypoxia plays a major role in reducing the efficacy of therapeutic modalities like chemotherapy and radiation therapy in combating cancer. In order to target hypoxic tissues, a tripeptide ligand having a 2-nitroimidazole moiety, as a bioreductive species, was synthesized. The latter was radiolabeled with 99mTc for imaging hypoxic regions of tumors and was characterized by means of its rhenium analogue. The biodistribution and scintigraphic image of the corresponding 99mTc-complex showed accumulation in tumor and these results suggest that it could be a marker for imaging tumor hypoxia.

Synthesis, in vitro and in vivo characterization of two novel 68Ga-labelled 5-nitroimidazole derivatives as potential agents for imaging hypoxia

Hypoxia imaging is an important field in radiopharmaceutical research since hypoxic cells are very resistant to radiation treatment and diffusional limitations restrict the efficacy of chemotherapy. Gallium-68 is a widely used radionuclide for positron emission tomography (PET) due to the availability of the (68)Ge/(68)Ga-generator. With the aim to develop new potential [(68)Ga]-radiopharmaceuticals for imaging hypoxia, we have synthesized and evaluated two novel (68)Ga-labelled 5-nitroimidazole derivatives. Two 5-nitroimidazole derivatives, and were synthesized. Preparation of [(68)Ga]complexes [(68)Ga]-Nit1 and [(68)Ga]-Nit2 was performed at pH 4.5 and 95 °C during 15 minutes and radiochemical purity (RP) was evaluated by reverse phase HPLC. Stability, lipophilicity and plasma protein binding were studied. Biological behaviour in HCT-15 cells both in normoxia and hypoxia has been assessed. Biodistribution in animals bearing induced 3LL Lewis murine lung carcinoma was studied. Comparison with [(18)F]FMISO is also presented. Nit1 and Nit2 have been successfully synthesized. Labelling in high radiochemical purity was achieved for both ligands. Complexes are stable in labelling milieu for at least four hours and in human plasma or in the presence of an excess of DTPA for at least two hours. Both compounds showed high uptake in hypoxic cells in vitro and a very favourable biodistribution profile in mice bearing induced tumours. Results are comparable to those obtained for [(18)F]FMISO. Selective uptake and retention in tumour together with favourable tumour/muscle ratio make these compounds promising candidates for further evaluation as potential hypoxia imaging agents.

Synthesis and antiplasmodial and antimycobacterial evaluation of new nitroimidazole and nitroimidazooxazine derivatives.

The synthesis and antiplasmodial and antimycobacterial evaluation of two new series of nitroimidazole and nitroimidazooxazine derivatives is described. The majority of these compounds, especially hybrids 9d, 9f, and 14b, exhibited potent activity against the chloroquine-resistant K1 strain of Plasmodium falciparum. Furthermore, a notable number from the tetrazole series were significantly more active against M. tuberculosis than kanamycin, a standard TB drug.

Validation of a modified fluorimetric assay for the screening of trichomonacidal drugs

A fluorimetric microassay that uses a redox dye to determine the viability of the flagellate Trichomonas vaginalis has been optimised to provide a more sensitive method to evaluate potential trichomonacidal compounds. Resazurin has been used in recent years to test drugs against different parasites, including trichomonadid protozoa; however, the reproducibility of these resazurin-based methods in our laboratory has been limited because the flagellate culture medium spontaneously reduces the resazurin. The objective of this work was to refine the fluorimetric microassay method previously developed by other research groups to reduce the fluorescence background generated by the media and increase the sensitivity of the screening assay. The experimental conditions, time of incubation, resazurin concentration and media used in the microtitre plates were adjusted. Different drug sensitivity studies against T. vaginalis were developed using the 5-nitroimidazole reference drugs, new 5-nitroindazolinones and 5-nitroindazole synthetic derivatives. Haemocytometer count results were compared with the resazurin assay using a 10% solution of 3 mM resazurin dissolved in phosphate buffered saline with glucose (1 mg/mL). The fluorimetric assay and the haemocytometer counts resulted in similar percentages of trichomonacidal activity in all the experiments, demonstrating that the fluorimetric microtitre assay has the necessary accuracy for high-throughput screening of new drugs against T. vaginalis.

Dose–response relationships of FMISO between trace dose and various macro-doses in rat by ultra-performance liquid chromatography with mass spectrometry and radioactivity analysis

Screening the pharmacokinetics of candidates using liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) may be efficacious and safe for the research and development of new PET imaging agents. However, the PET imaging agent is administered as trace dose and the sensitivity of LC-MS/MS is often insufficient. If the dose was increased to be quantifiable, it should be necessary to prove whether the pharmacokinetics between trace and macro-doses is consistent or not. In this paper, fluoromisonidazole (FMISO), a tumor PET imaging agent, was chosen to evaluate the dose-response pharmacokinetics by administering various single intravenous doses (0.1, 0.4, 1.6 and 6.4 mg/kg) in male Sprague-Dawley rats. The plasma concentration of FMISO was determined by an ultra-performance liquid chromatography-tandem mass spectrometric (UPLC-MS/MS) method, and the blood radioactivity of [(18)F]FMISO was detected by a gamma counter. By calculating and comparing the pharmacokinetic parameters, the total area under the plasma concentration-time curve from time zero to infinity (AUC(0-∞)) and peak plasma concentration (C(max)) values increased with the selected FMISO doses, and showing linear dose-dependent. On the other hand, some parameters related to time, such as the elimination half-lives (t(1/2)) and elimination rate constant (K(e)) were dose-independent, and there is no significant deference between trace dose and various macro-doses. The data should be useful to evaluate the novel 2-nitroimidazole derivatives as potential PET tumor imaging agents.

Synthesis, in vitro and in vivo characterization of novel 99mTc-‘4+1’-labeled 5-nitroimidazole derivatives as potential agents for imaging hypoxia

The evaluation of oxygenation status of solid tumors is an important field of radiopharmaceutical research. With the aim to develop new potential 99mTc-radiopharmaceuticals for imaging hypoxia, we have synthesized two novel isocyanide derivatives of metronidazole, which has demonstrated high affinity for hypoxic tumors in vitro and in vivo. Metronidazole derivatives 4-isocyano-N-[2-(2-methyl-5-nitro-1H-imidazol-1-yl)ethyl]butanamide (M1) and 1-(4-isocyanobutanoyl)-4-[2-(2-methyl-5-nitro-1H-imidazol-1-yl)ethyl]piperazine (M2) were synthesized, and labeling was performed through preparation of their corresponding 99mTc-(4+1) complexes, 99mTc-NS3M1 and 99mTc-NS3M2. The structure of the technetium complexes was corroborated by preparation and characterization of the corresponding rhenium complexes. We have studied the main physicochemical properties (stability, lipophilicity and plasma protein binding). Biological behavior in HCT-15 cells both in oxia and in hypoxia was assessed. Biodistribution in normal mice and in animals bearing induced 3LL Lewis murine lung carcinoma was also studied. Metronidazole derivatives were successfully synthesized. Labeling with high radiochemical purity was achieved for both ligands. 99mTc complexes were stable in labeling milieu and human plasma. However, presence of the piperazine linker in M2 resulted in higher lipophilicity and protein binding. Although cell uptake in hypoxic conditions was observed for both radiotracers, 99mTc-NS3M2 biodistribution was considered unsuitable for a potential radiopharmaceutical due to high liver uptake and poor blood clearance. However, 99mTc-NS3M1 demonstrated a very favorable in vivo profile both in normal mice and in mice bearing induced tumors. Selective uptake and retention in tumor together with favorable tumor/muscle ratio make 99mTc-NS3M1 a promising candidate for further evaluation as potential hypoxia imaging agent in tumors.