Synthesis and evaluation of a series of 99mTc-labelled zoledronic acid derivatives as potential bone seeking agents

Developing novel superior bone-seeking radiopharmaceuticals for the detection of malignant bone lesions could further improve the diagnostic value of routine bone scanning. A series of radiolabeled diphosphonates (^99mTc-EIPrDP, ^99mTc-EIBDP and ^99mTc-EIPeDP) have been designed and synthesized successfully in high chemical yields and radiochemical purity. The in vitro and in vivo biological properties were systematically investigated and compared. The biodistribution in mice shows that ^99mTc-EIPrDP has higher bone uptake (13.3 ± 1.23) than those of ^99mTc-EIBDP and ^99mTc-EIPeDP (11.7 ± 0.28 and 8.69 ± 0.04 %ID/g) at 1–2 h post injection. It also has the highest uptake ratio of bone to muscle, spleen and heart, respectively, and faster blood clearance in early times. The present study indicates that ^99mTc-EIPrDP holds great promise as a bone imaging agent.     

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.     

Preparation of 186Re-MIBI complex for myocardial perfusion imaging as potential replacement of analogues 99mTc-MIBI complex

The optimum conditions to label 2-methoxyisobutyl-isonitrile (MIBI) compound with pure ¹⁸⁶Re as a stable contrast agent for myocardial perfusion imaging were investigated. Complexation of MIBI with ¹⁸⁶Re was carried out using anhydrous stannous chloride, gentisic acid and 1 ml of 37 MBq ¹⁸⁶ReO₄ ^? at pH 2 in a boiling water bath for 30 min. The corresponding radiochemical yield was ≈95.5 %. The biodistribution studies in mice indicated that, the complex was cleared from the body by kidneys to urinary bladder and finally into urine. ¹⁸⁶Re-MIBI demonstrated satisfactory heart uptake like to ^99mTc- MIBI (8.94 % dose/organ at 5 min). The obtained data showed that ¹⁸⁶Re-MIBI is a potential replacement of ^99mTc-MIBI for myocardial perfusion imaging.     

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.     

<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.     

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.     

Synthesis of [64Cu]DOTA-ADIBON3-Ala-PEG28-A20FMDV2 via copper-free click chemistry for PET imaging of integrin αvβ6

Catalyst-free click reactions are effective chemical tools for synthesis of radiometal-based radiopharmaceuticals offering advantages towards preparation of non-toxic agents with high specific activity. In the present study the radiotracer [⁶⁴Cu]DOTA-ADIBON₃-Ala-PEG₂₈-A20FMDV2, [⁶⁴Cu]3, was synthesized for positron emission tomography imaging of integrin α_vβ₆ expressing tumors via a strain-promoted click reaction using both a “pre-click” and “post-click” approaches. The radiotracer, prepared in >99 % radiochemical yield, was evaluated in vitro (64.6 ± 2.8 % binding to α_vβ₆-positive cells vs. <5 % to α_vβ₆-negative cells) and in vivo (α_vβ₆-positive tumor uptake: 1.52 ± 0.16 % ID/g, 24 h p.i.). While the high initial renal uptake (76.2 ± 10.7 % ID/g at 1 h p.i.) was comparable to a previously reported radiotracer, [⁶⁴Cu]DOTA-PEG₂₈-A20FMDV2, [⁶⁴Cu]3 showed notably improved renal clearance (11.3 ± 2.5 % ID/g at 24 h p.i.). Thus, the introduction of a chelator-strained alkyne system resulted in improved pharmacokinetics for the present radiotracer, highlighting the attractive prospects of strain-promoted click-based preparations in the construction of radiometalated bioconjugates for targeted molecular imaging and therapy.     

Evaluation of a new radiolabeled bombesin derivative with 99mTc as potential targeted tumor imaging agent

Gastrin-releasing peptide (GRP) receptors are over-expressed in various human tumor including breast and prostate which can be targeted with bombesin for diagnosis and targeted therapy. High abdominal accumulation and the poor in vivo stability of radiolabeled bombesin analogues may represent a limitation for diagnostic imaging and targeted therapy. In this study a new bombesin derivative was labeled with ^99mTc via HYNIC and tricine as a coligand and investigated further. The peptide HYNIC conjugate was synthesized on a solid phase using Fmoc strategy. Labeling with ^99mTc was performed at 100 °C for 10 min and radiochemical analysis involved ITLC and HPLC methods. The stability of radiopeptide was checked in the presence of human serum at 37 °C up to 24 h. Internalization was studied with the human GRP receptor cell line PC-3. The Biodistribution was studied in mice. Labeling yield of >98 % was obtained to correspond a specific activity of ~80.9 GBq/μmol. Radioconjugate internalization into PC-3 cells was high and specific (15.6 ± 1.9 % at 4 h). A high and specific uptake in GRP-receptor-positive organs such as mouse tumor and pancreas (2.11 ± 0.18 and 1.78 ± 0.09 % ID/g after 1 h respectively) was also determined.     

Synthesis and evaluation of a phenylbenzothiazole-based 99mTc(CO)3-radiotracer for possible application in imaging of β-amyloid plaques in Alzheimer’s disease

The 2-phenyl benzothiazole pharmacophore is known to have high affinity for amyloid beta (Aβ) and is therefore derivatized, to [N-(4′-benzothiazol-pyridin-2-yl-methyl-amino)-acetic acid (BTPAA)] for radiolabeling with [^99mTc(CO)₃(H₂O)₃]⁺ precursor. The radiotracer, ^99mTc(CO)₃–BTPAA is evaluated in vitro and in vivo to determine its binding with the Aβ and ability to cross the blood brain barrier. The radiotracer prepared in >95 % radiochemical yield, showed ~25 % inhibition in presence of thioflavin-T, indicating its specificity towards aggregated Aβ protein. The radiotracer also showed brain uptake of 0.25 ± 0.04 % injected dose/g at 2 min post injection, indicating its ability to cross the blood brain barrier.     

Synthesis of techentium-99m labeled clinafloxacin (99mTc�CCNN) complex and biological evaluation as a potential Staphylococcus aureus infection imaging agent

In the present study synthesis of the ^99mTc?CCNN complex and its efficacy as a prospective Staphylococcus aureus (S. aureus) infection imaging agent was assessed. The ^99mTc?CCNN complex was characterized in terms of stability in saline, serum, in vitro binding with S. aureus and in vivo percent absorption in male Wister rats (MWR) infected with live and heat killed S. aureus. Radiochemically the ^99mTc?CCNN complex showed stable behavior in saline and serum at different intervals. At 30 min after reconstitution the complex showed maximum radiochemical purity (RCP) yield of 97.55 ± 0.22%. The RCP yield decreased to 90.50 ± 0.18% within 240 min. In serum, 18.15% unwanted side product was appeared within 16 h of the incubation. In vitro saturated binding with S. aureus was observed at different intervals with a 62.00% maximum at 90 min. Normal percent in vivo uptake was observed in MWR artificially infected with live S. aureus with a five times higher in the infected muscle as compared to the inflamed and normal muscles. No difference in the percent uptake of the complex in MWR infected with heat killed S. aureus in the infected, inflamed and normal muscles were observed. Based on the promising in vitro and in vivo radiochemical and biological characteristics, we recommend the ^99mTc?CCNN complex for in vivo localization of the S. aureus infectious foci.     

In vivo biodistribution of pancreatic-derived factor using 18F-labeled PANDER PET imaging

The purpose of this study was to investigate in vivo biodistribution and potential target tissues of pancreatic-derived factor (PANDER, FAM3B) using ¹⁸F-labeled PANDER positron emission tomography (PET) imaging. ¹⁸F-Labeled PANDER ([¹⁸F]FB-PANDER) was prepared by reaction of PANDER and N-succinimidyl-4-[¹⁸F]fluorobenzoate ([¹⁸F]SFB). The uncorrected radiochemical yield of [¹⁸F]FB-PANDER was 15.2 ± 3.4 % (n = 4) based on [¹⁸F]SFB within the total synthesis time of 30 min. In vivo biodistribution of [¹⁸F]FB-PANDER in nomal mice and PET imaging demonstrated high uptake of the radiotracer in urinary bladder, kidneys and gall bladder, and fast clearance from kidneys and gall bladder. Also, moderate uptake in blood, liver, pancreas, small intestine and bone, low uptake in brain and muscle, and almost no uptake in S180 fibrosarcoma tissue were observed. The results indicated that the major excretion route of PANDER was through renal-urinary bladder and biliary system, and no obvious binding targets of PANDER in the main organs and S180 fibrosarcoma tissue were found.     

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.     

New BATO complexes for brain and myocardial imaging

Two new BATO complexes^99mTcC1 (dmg)₃BC₆H₄CH₃ (Cholor[bis {2,3-butanedionedioxime(1-)-O} {2,3-butanedionedioximato (2-)-N,N',N",N'",N",N"'} (m-tolueneborato) technetium]) and^99m TcCl (4-MCDO)₃MeB, ([bis{4-methyl-1,2-cyclohexanedioximato (1-)-O} {4-methyl-cyclohexane-1, 2-dione-dioximato (2-)-O} {methyl-borato (2-)-N,N', N", N'", N", N"'} chlorotechnetium]), generally called BATO (Boronic Acid Adducts of Technetium Dioximes, had been synthesized and evaluated for potential use in brain and myocardial perfusion imaging. Their labeling conditions were also investigated. In their biodistribution studyies they showed higher radiochemical stability and rapid brain uptake and myocardial uptake in mice. After i.v administration, the first complex had 0.87%ID in the brain and 1.02% ID in the heart at 2 min and it had a longer retention in brain (0.62%ID was maintained at 15 min postinection) but rapidly cleared from heart (0.33%ID postinjection). For the second complex, it showed very rapid blood clearance. The uptake of heart, lung and blood in mice at 2 min respectively were: 1.32%ID, 2.48%ID and 6.66%ID. These two complexes formation were 2rapid, simple and of high yield(91%). The processes were easy to kit formulation.     

99mTc-labeled bombesin analog for breast cancer identification

Bombesin is a tetradecapeptide that binds specifically to gastrin releasing peptide receptors in humans. Several forms of cancer, including lung, prostate, breast, and colon express receptors for bombesin-like peptides. Radiolabeled bombesin analogs with a high affinity for these receptors might therefore be used for scintigraphic imaging of these tumor types. A truncated bombesin derivative (HYNIC-βAla-Bombesin_(7–14)) was radiolabeled with technetium-99m using EDDA and tricine as coligands. In vitro stability was evaluated in presence of plasma and excess of cysteine. The receptor-binding affinity assays was evaluated in MDA-MB-231 cancer cell line. In addition, in vivo biodistribution was performed in nude mice bearing breast tumor. In vitro assay showed a good affinity for the MDA-MB-231 cell line, showing 20.0 % of internalization at 4 h post-administration. ^99mTc-HYNIC-βAla-Bombesin_(7–14) biodistribution revealed a rapid clearance and a significant renal excretion. In addition, tumor uptake was higher than non-excretory organs, such as the spleen, the liver, and muscles. Tumor-to-muscle and tumor-to-blood ratios for ^99mTc-HYNIC-βAla-Bombesin_(7–14) showed high values at 4 h post-injection (5.34 and 4.55, respectively). Furthermore, blocked studies using cold bombesin peptide were performed, which demonstrated an important decrease in tumor uptake, indicating a tumor specificity for ^99mTc-HYNIC-βAla-Bombesin_(7–14). The ^99mTc-HYNIC-βAla-Bombesin_(7–14) displayed suitable radiochemical characteristics, and adequate affinity to breast tumor cells (MDA-MB-231). Therefore, this analog can be considered as a candidate for the identification of bombesin-positive tumors.     

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,radiolabelling and biodistribution of HYNIC-Tyr<Superscript>3</Superscript> octreotide: a somatostatin receptor positive tumour imaging agent

In vivo imaging of tumours using radiolabelled somatostatin (SST) analogues has become an accepted clinical tool in oncology. HYNIC-Tyr³ octreotide and Tyr³ octreotide were synthesized by FMOC solid-phase peptide synthesis using a semi-automated synthesizer. These were analyzed and purified by RP-HPLC, mass spectroscopy, IR spectroscopy, ¹H NMR and ¹³C NMR. The prochelator 6-BOC-HYNIC was also synthesised and characterised indigenously. HYNIC-Tyr³ octreotide was labelled with ^99mTc using Tricine and EDDA as coligand by SnCl₂ method. Labelling with ^99mTc was performed at 100 °C for 15 min and radiochemical analysis by ITLC and HPLC methods. The radiochemical purity of the complex was over 98% and log p value was found to be −1.27 ± 0.12. The stability of radiolabelled peptide complex was checked at 37 °C up to 24 h. Blood clearance and protein-binding study was also performed. In vivo biodistribution studies in rat showed uptake of ^99mTc-HYNIC-TOC in kidney than any other organs. The blood clearance was faster with rapid excretion through kidneys and relatively low uptake in liver.     

Synthesis and biological evaluation of <Superscript>99m</Superscript>Tc labeled aryl piperazine derivatives as cerebral radiotracers

The aim of this study is to develop cerebral radiotracers for central nervous system receptors imaging. The synthesis, characterization and biological evaluation of two aryl piperazine ^99mTc-radiocomplexes based on the piano stool motif [CpM(CO₃)] (Cp = cyclopentadienyl, M = ^99mTc/Re) is reported. The ^99mTc-radiocomplexes were obtained quickly (time < 5 min) with high radiochemical yields. The ^99mTc-radiocomplexes characterized by high performance liquid chromatography comparison with the rhenium surrogates have both a suitable lipophilicity and are able to cross the blood brain barrier with 0.43 ± 0.05 and 1.96 ± 0.06% ID/g of brain uptake, at 10 min post injection.     

Synthesis of 99mTc-oxybutynin for M3-receptor-mediated imaging of urinary bladder

Radiolabeling of oxybutynin, a muscarinic acetylcholine (mACh) receptor antagonist agent with ^99mTc is of considerable interest for imaging of urinary bladder. This study is aimed to optimize radiolabeling yield of oxybutynin with ^99mTc using SnCl₂·2H₂O as a reducing agent with respect to factors that affect the reaction conditions such as oxybutynin amount, stannous chloride amount, reaction time and pH of the reaction mixture. In vitro stability of the radiolabeled complex was checked and it was found to be stable for up to 8 h. ^99mTc-oxybutynin was injected via subcutaneous and intravenous administration routes into normal Sprague?CDawley rats. Biodistribution studies have revealed that ^99mTc-oxybutynin exhibits high affinity and specificity for the muscarinic M₃ subtype located on the smooth muscle of urinary bladder relative to the M₁ and M₂ subtypes of the G protein coupled receptor (GPCR) superfamily. In vivo uptake of subcutaneous ^99mTc-oxybutynin in urinary bladder was 19.6 ± 0.42% ID at 0.5 h, whereas in intravenous administration route the accumulation in the urinary bladder was found to be 9.4 ± 0.31% ID at 0.5 h post injection. Administration of cold oxybutynin effectively blocked urinary bladder uptake and further confirms the high specificity of this complex for the M₃ receptor.     

Synthesis and biodistribution of <Superscript>99m</Superscript>TcN-isopentyl xanthate as a potential myocardial perfusion imaging agent

A novel ^99mTc nitrido xanthate complex ^99mTcN(IPEXT)₂ (IPEXT: isopentyl xanthate) has been synthesized by the reduction of ^99mTcO₄ ⁻ into [^99mTcN]²⁺ with stannous chloride in the presence of succinic dihydrazide and propylenediamine tetraacetic acid, followed by the addition of the corresponding xanthate ligand. The radiochemical purity of the complex was over 90% as measured by thin layer chromatography (TLC). No decomposition of the complex at room temperature was observed over a period of 6 hours. Its partition coefficient indicated that it was a lipophilic complex. The electrophoresis results showed the complex was neutral. Biodistribution in mice showed that the ^99mTcN(IPEXT)₂ complex accumulated in the heart with high uptake. The heart uptake (%IDg) was 8.00% at 5-minute post-injection, but the heart/lung, heart/liver and heart/blood ratios were not high, thereby, restricting the use of the complex as a good myocardial imaging agent.     

Synthesis,radiolabeling and biological distribution of a new dioxime derivative as a potential tumor imaging agent

A novel dioxime derivative (2E,2′E,3E,3′E)-3,3′-(pyrimidine-4,5-diylbis(azanylylidene))bis(butan-2-one)dioxime was synthesized with a yield of 65%. IR, elemental analysis, mass spectroscopy and ¹H-NMR were used to characterize the structure of the synthesized compound. ^99mTc-dioxime was radio-synthesized with a high radiochemical yield of 97.8 ± 0.5% and in vitro stability of 6 h under the optimum conditions. The preclinical evaluation of ^99mTc-dioxime in solid tumor-bearing mice showed high accumulation in solid tumor cells with a high Target/Non-Target ratio of 5.14 at 30 min post-injection. This study suggests that ^99mTc-dioxime derivative is a promising candidate as a new ^99mTc-based tumor-imaging agent after further preclinical studies.