Radiosynthesis and Biodistribution of <Superscript>99m</Superscript>Tc-Metronidazole as an <Emphasis Type="Italic">Escherichia coli</Emphasis> Infection Imaging Radiopharmaceutical

Bacterial infection poses life-threatening challenge to humanity and stimulates to the researchers for developing better diagnostic and therapeutic agents complying with existing theranostic techniques. Nuclear medicine technique helps to visualize hard-to-diagnose deep-seated bacterial infections using radionuclide-labeled tracer agents. Metronidazole is an antiprotozoal antibiotic that serves as a preeminent anaerobic chemotherapeutic agent. The aim of this study was to develop technetium-99m-labeled metronidazole radiotracer for the detection of deep-seated bacterial infections. Radiosynthesis of ^99mTc-metronidazole was carried by reacting reduced technetium-99m and metronidazole at neutral pH for 30 min. The stannous chloride dihydrate was used as the reducing agent. At optimum radiolabeling conditions, ~ 94% radiochemical was obtained. Quality control analysis was carried out with a chromatographic paper and instant thin-layer chromatographic analysis. The biodistribution study of radiochemical was performed using Escherichia coli bacterial infection-induced rat model. The scintigraphic study was performed using E. coli bacterial infection-induced rabbit model. The results showed promising accumulation at the site of infection and its rapid clearance from the body. The tracer showed target-to-non-target ratio 5.57 ± 0.04 at 1 h post-injection. The results showed that ^99mTc-MNZ has promising potential to accumulate at E. coli bacterial infection that can be used for E. coli infection imaging.     

Synthesis and biological distribution of 99mTc–norfloxacin complex, a novel agent for detecting sites of infection

The optimization of the radiolabeling yield of ciprofloxacin analogous, norfloxacin, with technetium-99m (^99mTc) was described. Dependence of the labeling yield of ^99mTc–norfloxacin complex on the concentration of norfloxacin, SnCl₂·2H₂O content, pH of the reaction mixture and reaction time was studied. Norfloxacin was labeled with ^99mTc at pH 3 with a labeling yield of 95.4% by using 5 mg norfloxacin, 50 μg SnCl₂·2H₂O and 30 min reaction time. The formed ^99mTc–norfloxacin complex was stable for a time up to 3 h. Biological distribution of ^99mTc–norfloxacin complex was investigated in experimentally induced inflammation rats using Staphylococcus aureus (bacterial infection model) and heat killed Staphylococcus aureus and turpentine oil (sterile inflammation model). In case of bacterial infection, the T/NT value for ^99mTc–norfloxacin complex was found to be 6.9 ± 0.4 which was higher than that of the commercially available ^99mTc–ciprofloxacin under the same experimental condition.     

Radiolabeling of gemifloxacin with technetium-99m and biological evaluation in artificially <Emphasis Type="Italic">Streptococcus pneumoniae</Emphasis> infected rats

In the current investigation complexation of the gemifloxacin (GIN) with technetium-99 m (^99mTc) and its biological evaluation in artificially Streptococcus pneumoniae (S. pneumoniae) infected rats was assessed as potential S. pneumoniae infection radiotracer. Radiochemically the ^99mTc-GIN complex was further analyzed in terms of stability in saline, in vitro stability in serum at 37 °C, in vitro binding with S. pneumoniae and biodistribution in artificially S. pneumoniae (living and heat killed) infected rats. The complex was found 97.25 ± 0.25% radiochemically stable in saline at 30 min after reconstitution. The stability of the ^99mTc-GIN complex was decreased to 90.50 ± 0.20% within 240 min after reconstitution. In serum the ^99mTc-GIN complex showed stable profile with the appearance of 18.85% free tracer within 16 h of incubation. The ^99mTc-GIN complex showed saturated in vitro binding with S. pneumoniae after different intervals. Almost five fold uptake was observed in living S. pneumoniae infected muscle of the rats as compared to the inflamed and normal muscle. No significant difference in the uptake of heat killed S. pneumoniae infected, inflamed and normal muscles of the rats. The high RCP yield in saline, in vitro permanence in serum, in vitro binding with living S. pneumoniae and biodistribution in artificially S. pneumoniae infected rats we recommend the ^99mTc-GIN as potential S. pneumoniae infection radiotracer.     

99mTc-labeling of a dithiocarbamate-DWAY fragment using [99mTcN]2+ core for the preparation of potential 5HT1A receptor imaging agents

1-(2-methoxy phenyl) piperazine fragment of WAY100635 or its phenolic analogue, derived from DWAY is used to design the desired structure of 5HT_1A receptor imaging agents. In this study a DWAY analogue was labeled with ^99mTc-nitrido ([^99mTcN]²⁺) core via dithiocarbamate. 2-(piperazin-1-yl) phenol dithiocarbamate was synthesized by the reaction of 2-(piperazin-1-yl) phenol with an equivalent amount of carbon disulfide in KOH solution then radiolabeled with ^99mTc-nitrido core. The final complex was characterized by HPLC and its radiochemical purity was more than 90 %. In vitro stability studies have shown the complex was stable at least 4 h after labeling at room temperature. The n-octanol/water partition coefficient experiment demonstrated log p = 1.34 for ^99mTcN–OHPP–DTC. Biodistribution results showed that radio tracer had moderate brain uptake (0.39 ± 0.03 %ID/g at 15 min and 0.29 ± 0.02 %ID/g at 120 min) and good retention, suggesting that this complex may lead to a further development of a radiotracer with specific binding to 5-HT_1A receptor.     

Preparation and biological distribution of 99mTc-cefazolin complex, a novel agent for detecting sites of infection

The optimization of the radiolabeling yield of cefazolin with ^99mTc was described. Dependence of the labeling yield of ^99mTc-cefazolin complex on the amounts of cefazolin and SnCl₂·2H₂O, pH and reaction time was studied. Cefazolin was labeled with ^99mTc with a labeling yield of 89.5 % by using 1 mg cefazolin, 5 μg SnCl₂·2H₂O at pH 4 and 30 min reaction time. The radiochemical purity of ^99mTc-cefazolin was evaluated with ITLC. The formed ^99mTc-cefazolin complex was stable for a time up to 3 h, after that the labeling yield decreased 64.0 % at 8 h. Biological distribution of ^99mTc-cefazolin complex was investigated in experimentally induced inflammation mice, in the left thigh, using Staphylococcus aureus (bacterial infection model) and turpentine oil (sterile inflammation model). Both thighs of the mice were dissected and counted and the ratio of bacterial infected thigh/contralateral thigh was then evaluated. In case of bacterial infection, T/NT for ^99mTc-cefazolin complex was 8.57 ± 0.4 after 0.5 h, which was higher than that of the commercially available ^99mTc-ciprofloxacin under the same experimental conditions. The ability of ^99mTc-cefazolin to differentiate between septic and aseptic inflammation indicates that ^99mTc-cefazolin could undergo further clinical trials to be used for imaging sites of infection.     

Synthesis, radiochemistry and stability of the conjugates of technetium-99m complexes with Substance P

Two types of technetium-99m complexes: (i) with the Hynic ligand linked to Substance P(1–11) and (ii) of the type ‘4 + 1’ consisting of tetradentate tripodal chelator tris(2-mercaptoethyl)-amine and monodentate isocyanide ligand previously coupled with Substance P(1–11), have been prepared on the n.c.a. scale. The obtained conjugates exhibit different lipophilicity and high stability in neutral aqueous solutions, even in the presence of excess concentration of histidine/cysteine competitive standard ligands. The conjugate (^99mTc(NS₃)(CN))₂–SP containing two technetium-99m species in the molecule may be expected to be an extremely good diagnostic radiopharmaceutical.     

Labeling of ursodeoxycholic acid with technetium-99m for hepatobiliary imaging

An adopted method for the preparation of high radiochemical purity ^99mTc-ursodeoxycholic acid (UDCA) was conducted with a high radiochemical yield up to 97.5 %. The reaction proceeds well using 2 mg UDCA, 50 μg tin chloride in solution of pH 8 at room temperature for 30 min. The radiochemical yield was up to 97.5 % as pure as ^99mTc-UDCA. Different chromatographic techniques (paper chromatography and electrophoresis) were used to evaluate the radiochemical yield and purity of the labeled product. Biodistribution studies were carried out in Albino Swiss mice at different time intervals after administration of ^99mTc-UDCA. The uptake of ^99mTc-UDCA in the liver gave the chance to diagnose it. The results indicate that the labeled compound cleared from the systematic circulation within 2 h after administration and majority of organs showed significant decrease in uptake of ^99mTc-UDCA. Finally, the liver uptake was high and the results indicate the possibility of using ^99mTc-UDCA for hepatobiliary imaging.     

A perspective on <Superscript>99m</Superscript>Tc and <Superscript>125/131</Superscript>I labeled receptor targeted compounds and their in vitro/in vivo affinities

A novel quinoline derivative, 2,2′-[(5-chloro-8-hydroxyquinoline-7-yl) methylazanediyl] diacetic acid (CHQMADA) was labeled with ^99mTc using SnCl₂·2H₂O as a reducing agent to give a complex with a labeling yield 94 %. Also [^99mTc(H₂O)₃(CO)₃]⁺ was prepared by heating at 100 °C for 30 min using 2 mg CHQMADA at pH 8 to give a labeling yield >99 %. ^99mTc-(CO)₃ CHQMADA and ^99mTc-Sn(II)-CHQMADA showed tissue uptake (target to non target T/NT = 6.80 ± 0.22) and (T/NT = 5.65 ± 0.34) respectively in Escherichia coli induced infection, which is higher than the commercially available ^99mTc-ciprofloxacin (T/NT = 3.80 ± 0.80). In conclusion, both complexes were able to differentiate between septic and aseptic inflammation with superiority of [^99mTc-(CO)₃ CHQMADA].     

Synthesis and biological evaluation of fatty acid derivatives for myocardial imaging containing [<Superscript>99m</Superscript>Tc(CO)<Subscript>3</Subscript>]<Superscript>+</Superscript>

Radiolabeled fatty acids as myocardial metabolic agent are used for detecting ischemic heart disease, however, no ^99mTc-labeled fatty acids have potential use in clinical diagnosis. In this work, five fatty acid analogues labeled with ^99mTc were firstly synthesized and characterized, their biological behaviors were investigated. All Radiotracers had good stability when incubated in rat serum for 3 h at 37 °C. ^99mTc -CpT-12-ODPPA (8b) showed higher initial myocardial uptake (8.17% ID/g at 1 min postinjection) and good heart/blood ratio (2.58 at 30 min postinjection). ^99mTc-11-dpa-OUFA (2b) as positively charged lipophilic compounds had reasonable heart uptake (5.59% ID/g at 1 min postinjection) and good retention (1.89% ID/g at 60 min postinjection). Unfortunately, no great improvement was obtained by the five ^99mTc-labeled fatty acid analogues for the short myocardial retention and poor heart/liver ratios.     

Preliminary studies of 99mTc-PQQ-NMDAR binding and effect of specificity binding by mannitol

Pyrroloquinoline quinone (PQQ) is a powerful neuroprotectant that specifically binds to brain NMDA receptors and inhibits excitotoxicity. Imaging this binding reaction in the brain remains a long sought goal in this field of study, and one of the primary challenges remaining is enabling soluble labeled PQQ to pass the blood–brain barrier (BBB). Previously, our group successfully labeled PQQ with Technetium-99m (^99mTc), a metastable nuclear isomer used in radioactive isotope medical tests. In this work, we determined the specific binding of ^99mTc-PQQ and NMDAR by radioligand receptor assay. Ebselen (EB) and MK-801 both effectively inhibited ^99mTc-PQQ binding. We then investigated methods of opening the BBB using mannitol to enable entry to the brain by ^99mTc-PQQ. Our results showed that 7.5 mL/kg of 20 % mannitol effectively opened the BBB and 20 min was the optimum treatment time. Competition studies showed that mannitol did not affect the specific binding between ^99mTc-PQQ and NMDA receptors. Using this method, the amount of ^99mTc-PQQ uptake and retention was increased most significantly in the hippocampus and cortex, and re-opening the BBB did not affect binding. Together, our results demonstrate that the use of mannitol to open the BBB may contribute significantly to improving image quality by increasing the uptake amount of a water-soluble agent in brain.     

Preparation,molecular modeling and biodistribution of 99mTc-phytochlorin complex

Phytochlorin [21H, 23H-Porphine-7-propanoicacid, 3-carboxy-5-(carboxymethyl)13-ethenyl-18-ethyl-7,8-dihydro-2,8,12,17-tetramethyl-,(7S,8S)] was labeled with ^99mTc and the factors affecting the labeling yield of ^99mTc-phytochlorin complex were studied in details. At pH 10, ^99mTc-phytochlorin complex was obtained with a high radiochemical yield of 98.4 ± 0.6 % by adding ^99mTc to 100 mg phytochlorin in the presence of 75 μg SnCl₂·2H₂O after 30 min reaction time. The molecular modeling study showed that the structure of ^99mTc-phytochlorin complex presents nearly linear HO–Tc–OH unit with an angle of 179.27° and a coplanar Tc(N1N2N3N4) unit. Biodistribution of ^99mTc-phytochlorin complex in tumor bearing mice showed high T/NT ratio (T/NT = 3.65 at 90 min post injection). This preclinical study showed that ^99mTc-phytochlorin complex is a potential selective radiotracer for solid tumor imaging and afford it as a new radiopharmaceutical suitable to proceed through the clinical trials for tumor imaging.     

Practicality of Tetragonal Nano-Zirconia as a Prospective Sorbent in the Preparation of 99Mo/99mTc Generator for Biomedical Applications

The feasibility of using tetragonal nano-zirconia (t-ZrO₂) as an effective sorbent for developing a ⁹⁹Mo/^99mTc chromatographic generator was demonstrated. The structural characteristics of the sorbent matrix were investigated by different analytical techniques such as XRD, BET surface area analysis, FT-IR, TEM etc. The material synthesized was nanocrystalline, in tetragonal phase with an average particle size of ~7 nm and a large surface area of 340 m² g^?1. The equilibrium sorption capacity of t-ZrO₂ is >250 mg Mo g^?1. The present study indicates that ⁹⁹Mo is both strongly and selectively retained by t-ZrO₂ at acidic pH and ^99mTc could be readily eluted from it, using 0.9% NaCl solution. A 9.25 GBq (250 mCi) t-ZrO₂ based chromatographic ⁹⁹Mo/^99mTc generator was developed and its performance was repeatedly evaluated for 10 days. ^99mTc could be eluted with >85% yield having acceptable radionuclidic, radiochemical and chemical purity for clinical applications. The compatibility of the product in the preparation of ^99mTc labeled formulations such as ^99mTc-EC and ^99mTc-DMSA was evaluated and found to be satisfactory.     

Formulation and preclinical evaluation of 99mTc–gemcitabine as a novel radiopharmaceutical for solid tumor imaging

The aim of this study is the formulation of a new radiopharmaceutical for imaging solid tumor bearing. Gemcitabine is a nucleoside analogue used as chemotherapeutic agent. Gemcitabine was formulated and radiolabeled with one of the most important diagnostic radioactive isotopes (technetium-99m) to be investigated in solid tumor imaging. The labeling parameters such as gemcitabine amount, stannous chloride amount, pH of the reaction mixture, and reaction time were optimized. ^99mTc–gemcitabine was prepared at pH 9 with a maximum labeling yield of 96 ± 0.3 % without any notable decomposition at room temperature over a period of 8 h. The preclinical evaluation and biodistribution in solid tumor bearing mice showed that ^99mTc–gemcitabine had solid tumor selectivity, preclinical high biological accumulation in tumor cells and high retention. Tumor/normal muscle (T/NT) ratios increased with time showing high T/NT ratio (T/NT = 4.9 ± 0.27 at 120 min post injection) and high Tumor/Blood ratio (3.4 ± 0.06), suggesting ^99mTc–gemcitabine as a novel solid tumor imaging agent.     

Preparation of radioiodinated ritodrine as a potential agent for lung imaging

Ritodrine (a beta-2 adrenergic receptor agonist) was successfully labeled with ¹²⁵I via direct electrophilic substitution reaction at ambient temperature. ¹²⁵I-ritodrine was obtained with a maximum labeling yield of 97 ± 0.163 % and in vitro stability up to 24 h. Biodistribution studies showed that maximum in vivo uptake of ¹²⁵I-ritodrine in lungs was 20.4 ± 0.22 % injected activity/g tissue at 1 h post-injection, whereas the clearance from mice appeared to proceed mainly via the renal pathway. ¹²⁵I-ritodrine is not a blood product and so it is more safe than the currently available ^99mTc-MAA, and its lung uptake is higher than that of the recently discovered ^99mTc(CO)₅I and ^99mTc-DHPM. As a conclusion, radioiodinated ritodrine could be used as a novel radiopharmaceutical for lung perfusion scan safer than the currently available ^99mTc-MAA and more potential than the recently discovered ^99mTc(CO)₅I and ^99mTc-DHPM.     

Radiochemical determination of technetium-99

The radiochemistry of technetium-99 is reviewed and the different measurement techniques are compared. Experimental results on sorption of technetium on two different types of ion exchange resins using^99mTc and^95mTc as chemical yield tracers are presented. Spectra calibrations of liquid scintillation counter using^95mTc as chemical yield tracer of⁹⁹Tc are discussed.     

<Superscript>99m</Superscript>Tc-roxifiban: a potential molecular imaging agent for the detection and localization of acute venous thrombosis

^99mTc-roxifiban was obtained in a high radiochemical yield (98.4%) by complexing ~750 MBq ^99mTc with 2.5 mg roxifiban in the presence of 150 µg SnCl₂·2H₂O. Factors affecting the labelling yield were investigated and optimized. The complex was lipophilic and stable in saline and serum for more than 8 h. The complex structure prediction and molecular docking to its target activated GPIIb/IIIa receptor were performed. The tracer in vitro binding to activated platelets was high (27–32%). In vivo evaluation was performed through clearance, biodistribution and imaging studies in rats. All results supported the usefulness of the tracer as thrombus imaging agent.     

In vitro evaluation of apoptosis detection by 99mTc-tetrofosmin in MCF-7 breast cancer cell line

Radiolabeled molecules have an important role to evaluate tumor characteristics such as aggressiveness, and to identify the effectiveness of cancer treatments such as chemotherapy and radiotherapy. Various radionuclide (¹⁸F, ^99mTc, ¹²⁴I) labeled molecules can be used apoptosis detection by estimating decrescendos cell viability after therapy. ^99mTc-tetrofosmin which is used as a myocardial perfusion imaging agent in routine and at the same time is known to accumulate in various tumors including breast tumor. The aim of this study was to assess the utility of ^99mTc-tetrofosmin for monitoring the early response of MCF-7 breast cancer to chemotherapy. To evaluate the role of ^99mTc-tetrofosmin in vitro chemotherapy, the uptake ratio was determined using MCF-7 breast cancer line after the cells had been treated with cisplatin. When we examined the apoptotic ratios which induced with different dose of cisplatin in MCF-7 breast cancer cells by using Annexin V and TUNEL methods, it was observed that the rate of apoptosis increased with soaring dose. The uptake rates of ^99mTc-tetrofosmin in MCF-7 cell line in the chemotherapeutic groups were lower than it is in the control group (p < 0.01). The negative correlation between uptake ratios and apoptotic rates shows that ^99mTc-tetrofosmin may be used a radiopharmaceutical for evaluating chemotherapy response. ^99mTc-tetrofosmin might be probably useful as an imaging agent for estimation of early chemotherapy response in breast cancer.     

Derivatization of dihydrotetrabenazine for technetium-99m labelling towards a radiotracer targeting vesicular monoamine transporter 2

The development of technetium-99m-labelled dihydrotetrabenazine (DTBZ) derivative for vesicular monoamine transporter 2 (VMAT2) tracing could be a benefit for single photon emission computed tomography (SPECT) imaging due to easy labelling chemistry and great availability through nuclide generator system. Here, we successfully prepared a technetium-99m-labelled DTBZ derivative and subsequently evaluated its biological activity targeting VMAT2. A novel combination of the bisaminoethanethiol (BAT) chelator scaffold with the biologically active DTBZ vector was performed to synthesize the labelling precursor BAT-P-DTBZ, and it was accomplished in six steps. The technetium-99m labelling was carried out in the radiochemical study of BAT-P-DTBZ conjugate, and the radiolabelling conditions were investigated and optimized. Under the optimized labelling condition, ^99mTc-BAT-P-DTBZ was acquired with a good radiochemical purity of above 95 %. The quality control test showed that ^99mTc-BAT-P-DTBZ is stable over 6 h and it has a suitable lipophilicity, suggesting successful appositeness for the needs of routine biological evaluation experiments. The in vitro biological evaluation revealed that ^99mTc-BAT-P-DTBZ could bind to VMAT2 sites. The in vivo biodistribution study clearly indicated that the pancreas (VMAT2-enriched region) displays relatively high uptake of ^99mTc-BAT-P-DTBZ among all organs in mice. The specific VMAT2 binding signal of ^99mTc-BAT-P-DTBZ was separately detected in the in vitro and in vivo biological evaluation. Therefore, ^99mTc-BAT-P-DTBZ might be a potential imaging agent for monitoring VMAT2 binding sites in the pancreas.     

Synthesis and preliminary evaluation of 99mTc-spermine as a tumor imaging agent

Polyamines are essential for the growth and survival of all cells with biosynthesis and transportation of polyamines being very active in tumors. With the aim of developing a new tumor imaging agent, the endogenous polyamine, spermine was labeled with ^99mTc, and its characters were also evaluated via in vitro and in vivo studies. ^99mTc-labeled spermine probe (^99mTc-spermine) was synthesized by the direct pretinning procedure and the labeling procedure was optimized with regard to the pH, reaction time, amounts of spermine and SnCl₂. The stability of the ^99mTc-spermine and its capacity to accumulate into 4T1 tumor cells were also evaluated. Biodistribution of ^99mTc-spermine was studied in 4T1 tumor-bearing mice. In the optimal conditions, the whole radiosynthesis was accomplished within 10 min with a decay-corrected yield of 96.5 ± 1.3 % and radiochemical purity of >95 %.^99mTc-spermine was stable at both 37 and 4 °C for at least 6 h. In vitro tests revealed that the ability of ^99mTc-spermine to penetrate in 4T1 tumour cells and an excess of spermine blocked the accumulation of the compound in the models. Biodistribution studies showed a high tumor uptake peaked at 30 min post-injection with 1.82 ± 0.19 % ID%/g. The tumor to muscle uptake ratios of the probe were 3.60 ± 0.51, 4.48 ± 0.29, 4.82 ± 0.18, 5.64 ± 0.10, respectively at 30 min, 1, 2 and 4 h postinjection. Block studies indicated that ^99mTc-spermine had specific binding of tumor via polyamine transport systems. ^99mTc-spermine is a promising radiopharmaceutical in tumor imaging. Further studies are required to determine the usability of ^99mTc–spermine for diagnosis purposes.     

Chemical and biological properties of verapamil labeled with technetium-99m: A potential myocardial imaging agents

On the base of property to enter into myocardial cells as a calcium channel blocker, verapamil was labeled with technetium-99m in order to investigate the possibility to obtain new radiopharmaceutical for myocardial imaging. The conditions of labeling verapamil with technetium-99m for different ammounts of stannous(II) ion, mannitol, cystein and pH range 2.5–3.5 were examined. Investigation of radiochemical purity (>95%) and biodistribution of ^99mTc-verapamil in rats showed that it was stable during 2 hours after labeling. Accumulation of ^99mTc-verapamil in heart was 1.2% and in liver 9.4%, 5 minutes after injection. Biodistribution of ^99mTc-verapamil in rats in conditions of stress, pharmacologically caused by dipiridamol, showed that the elimination of ^99mTc-verapamil from the heart was slower related to the control group. In the group of rats previously treated with isoproterenol uptake of ^99mTc-verapamil in heart was lower related to the control group (0.7% versus 1.0%) 5 minutes after injection. Lipophilicity of ^99mTc-verapamil was examined by determination of partition coefficient (log P = 0.62) and protein binding (79%). Imaging studies on dogs provided relatively good myocardial images with partially overlap of activity in the lung and liver.