Evaluation of a bimodal, matched pair theranostic agent targeting prostate-specific membrane antigen.

BACKGROUND: Prostate cancer affects 1 in 6 men, and it is the second­leading cause of cancer-related death in American men. Surgery is one of the main treatment modalities for prostate cancer, but it often results in incomplete resection margins or complete resection that leads to nerve damage and undesirable side effects. In the present work, we have developed a new bimodal tracer, NODAGA-sCy7.5 PSMAi (prostate-specific membrane antigen inhibitor), labeled with the true matched theranostic pair 64Cu/67Cu and a near-infrared fluorescent dye. This agent could potentially be used for concomitant PET imaging, optical surgical navigation, and targeted radiopharmaceutical therapy. METHODS: A prostate-specific membrane antigen (PSMA)-targeting urea derivative was conjugated to NODAGA for copper radiolabeling and to the near-infrared fluorophore sulfo-Cy7.5 (sCy7.5). Binding studies were performed in PSMA-positive PC-3 PIP cells, as well as uptake and internalization assays in PC-3 PIP cells and PSMA-negative PC-3 wild type cells. Biodistribution studies of the 64Cu-labeled compound were performed in PC-3 PIP- and PC-3 tumor-bearing mice, and 67Cu biodistributions of the agent were obtained in PC-3 PIP tumor-carrying mice. PET imaging and fluorescence imaging were also performed, using the same molar doses, in the two mouse models. RESULTS: The PSMA conjugate bound with high affinity to PSMA-positive prostate cancer cells, as opposed to cells that were PSMA-negative. Uptake and internalization were rapid and PSMA-mediated in PC-3 PIP cells, while only minimal non-specific uptake was observed in PC-3 cells. Biodistribution studies showed specific uptake in PC-3 PIP tumors, while accumulation in PC-3 tumor-bearing mice was low. Furthermore, tumor uptake of the 67Cu-labeled agent in the PC-3 PIP model was statistically equivalent to that of 64Cu. PET and fluorescence imaging at 0.5 nmol per mouse also demonstrated that PC-3 PIP tumors could be clearly detected, while PC-3 tumors showed no tumor accumulation. CONCLUSIONS: NODAGA-sCy7.5-PSMAi was specific and selective in detecting PSMA-positive, as opposed to PSMA-negative, tumors in mouse models of prostate cancer. This bioconjugate could potentially be used for PET staging with 64Cu, targeted radiopharmaceutical therapy with 67Cu, and/or image-guided surgery with sCy7.5.

Design, characterization and evaluation of a new 99mTc-labeled folate derivative with affinity towards folate receptor.

Folate receptors (FRs) are known to be over-expressed in several human malignancies and therefore serve as an important target for small radiolabeled folate derivatives for non-invasive imaging of tumor, which is an important tool for future treatment recourse. In the present article, we report the synthesis of a new 99mTc-labeled radiotracer for the aforementioned application following the well-established 99mTc-'4+1' chemistry. Formation of the desired [99mTc]Tc-complex with >95% radiochemical purity was confirmed by radio-HPLC and its structure was ascertained by characterizing a natural rhenium analogue of the said complex. Although the ligand exhibited a weaker affinity towards FRs compared to native folic acid (IC50 8.09 µM vs 29.46 nM), the 99mTc-labeled complex was found to bind folate receptor-positive KB cells with high specificity (∼90%). Similar studies in a folate receptor negative cell line viz. A549 further corroborated the receptor-specificity of the synthesized complex. In vivo studies in KB tumor xenograft showed moderate uptake of ∼2.6% upto 3 h post-injection with high specificity (∼80%). The favorable features observed warrant further screening of the current design towards achieving an improved molecular probe for the said application.

Development of technetium-99m labeled ultrafine gold nanobioconjugates for targeted imaging of folate receptor positive cancers.

INTRODUCTION: Strategic design and synthesis of nanoparticle based preparations could improve diagnostic screening of several cancer types, thereby facilitating better clinical management of the disease. Towards this, the present work aims to develop and evaluate a radioactive technetium-99m (99mTc) labeled gold nanoparticle (NP) preparation modified with folic acid, so as to diagnose folate receptor positive cancers viz. ovarian, breast, etc. METHODS: 11-Bromoundecanoic acid (UA) was synthetically modified both with folic acid and Hydrazinonicotinic acid (HYNIC) chelate at the carboxylic acid end and subsequently converted to thiol functionality at the bromo terminal to yield folic acid-UA-SH and HYNIC-UA-SH ligands respectively. Gold NPs modified with folic acid and HYNIC chelator were obtained on direct addition of folic acid-UA-SH and HYNIC-UA-SH to chloroauric acid in polysorbate 80 solution under reducing conditions. These NPs were then radiolabeled with 99mTc following HYNIC labeling approach. Both the inactive and 99mTc-labeled gold NPs were then tested for their biological efficacy in folate receptor (FR) positive KB cancer cell lines. Also, biodistribution studies of 99mTc-labeled gold NPs were carried in KB tumor xenografts to ascertain the efficacy towards FR in in vivo system. RESULTS: Polysorbate 80 could stabilize the gold NP preparation with average size <10 nm as determined by TEM. Inhibition of [3H]folic acid with functionalized gold nanoparticle revealed affinity towards FR positive KB cell lines with an IC50 ~ 9 µM. Biodistribution studies of 99mTc-labeled gold NP preparation in SCID mice bearing KB tumor showed an uptake of 1.39 ± 0.18%ID/g in tumor and 5.48 ± 0.72%ID/g in kidneys at 3 h post-injection. In vivo distribution in folic acid pre-treated animals could not establish the specificity towards folate receptors. CONCLUSIONS: Biological evaluation of functionalized gold NP showed affinity towards FR positive cancer cell lines. 99mTc-labeled NP exhibited target uptake in both in vitro and in vivo models, but folic acid inhibition could not establish the target specificity. Nevertheless, in vivo pharmacokinetics envisaged in the present design was achieved using the present gold functionalized NP preparation.

Preparation and evaluation of 99mTc-labeled porphyrin complexes prepared using PNP and HYNIC cores: studying the effects of core selection on pharmacokinetics and tumor uptake in a mouse model.

Porphyrins are tetrapyrrolic macrocyclic ligands known for their affinity towards neoplastic tissues and once radiolabeled with a suitable diagnostic radioisotope could potentially be used for the imaging of tumorous lesions. In the present study, an unsymmetrically substituted porphyrin derivative namely 5-(p-amino-propyloxyphenyl)-10,15,20-tris(carboxymethyleneoxyphenyl)-porphyrin was synthesized and modified further to enable radiolabeling with 99mTc using two different 99mTc-cores viz. 99mTc-HYNIC (hydrazino nicotinic acid) and 99mTc(N)PNP2 (PNP2 = bis-[(2-dimethylphosphino)ethyl]-methoxy-ethylamine) in order to study the effect of employing different 99mTc-cores on tumor affinity and pharmacokinetic behavior of the resultant 99mTc-labeled porphyrin complexes. 99mTc-Porphyrin complexes were characterized by reversed phase HPLC studies and could be prepared with >95% radiochemical purity under optimized radiolabeling conditions. Both 99mTc-complexes were found to be adequately stable in human blood serum till 3 h post-preparation. Bio-distribution studies, carried out in Swiss mice bearing fibrosarcoma tumors, revealed relatively higher tumor uptake for the 99mTc-HYNIC-porphyrin complex (3.95 ± 1.42 and 3.28 ± 0.27% IA per g) compared to that exhibited by the 99mTc(N)PNP-DTC-porphyrin complex (1.52 ± 0.53 and 1.56 ± 0.10% IA per g) at 1.5 and 3 h post-administration, although the former complex exhibited comparatively lower lipophilicity in the octanol-water system. Higher uptake and longer retention in the blood were observed for the 99mTc-HYNIC-porphyrin complex (6.63 ± 0.75 and 4.36 ± 0.25% IA per g) compared to that exhibited by the 99mTc(N)PNP-DTC-porphyrin complex (2.41 ± 0.54 and 2.30 ± 0.16% IA per g) at both 1.5 and 3 h post-administration. However, relatively lower liver uptake was observed for the former complex (19.26 ± 3.48 and 18.45 ± 1.05% IA per g) than that exhibited by the latter one (39.37 ± 3.88 and 34.15 ± 8.25% IA per g) at both 1.5 and 3 h post-administration. This study indicates that the in vivo behavior exhibited by the 99mTc-labeled porphyrins not only depends on their lipophilicity/hydrophilicity but is also governed by the Tc-cores employed for radiolabeling.

Enhanced skin permeation of Methotrexate from penetration enhancer containing vesicles: In vitro optimization and in vivo evaluation.

Low dose Methotrexate (MTX) therapy is considered a gold standard for Rheumatoid Arthritis (RA). Transdermal drug delivery is hypothesized as an alternative to conventional therapies to alleviate its adverse effects. In our study, MTX was entrapped in deformable liposomes and loaded in a hydroxyethyl cellulose gel. This system was evaluated by the Box Behnken statistical design for optimization. The effect of formulation variables on particle size, entrapment and ex vivo skin permeation was studied. The MTX nanogel was evaluated for its dermal toxicity (acute and repeat dose safety), in vivo biodistribution (using 125I radio-labelled MTX) and therapeutic efficacy (collagen induced arthritis [CIA] model). The optimized formulation demonstrated appreciable nanosize (110 ± 20 nm), drug entrapment (42 ± 1.9%) and high ex vivo transdermal flux (17.37 ± 1.5 µg/cm2/hr). In the dermal toxicity studies, nanogel formulation did not show any signs of irritation or toxicity, whereas in the biodistribution study, the MTX nanogel formulation depicted sustained systemic delivery up to 48 h with low accumulation in its organs of toxicity such as the liver, kidneys and gut. In the CIA model, the MTX nanogel significantly ameliorated hind paw swelling, reduced arthritic score, joint damage (histological, radiological examination) and attenuated the rise in serum cytokines such as TNF-ɑ and IL-6. In conclusion, the optimized MTX nanogel formulation displayed skin biocompatibility, sustained systemic delivery, safety as well as therapeutic efficacy.

Synthesis and evaluation of 99mTc-analogues of [123/131I]mIBG prepared via [99mTc][Tc(CO)3(H2O)3]+ synthon for targeting norepinephrine transporter.

INTRODUCTION: meta-[123/131I]Iodobenzylguanidine (mIBG) is a clinical agent used for imaging neuroendocrine tumors, where uptake in tumor is via active transport mechanism through norepinephrine transporters (NET). Our group in past have evaluated a 99mTc-analogue of the above tracer, based on 99mTc-4 + 1 labeling approach, which exhibited significant affinity for NET but suffered from reduced specific uptake in comparison to reference standard no-carrier-added (n.c.a.) [125I]mIBG. The present work attempts to synthesize two new 99mTc-analogues of the radio-iodinated derivative following [99mTc]Tc(CO)31+ approach with an aim to improve the above specific uptake content. METHODS: Two different precursors, xylylenediamine and 1,3-bis(chloromethyl)benzene, were synthetically modified to yield meta-functionalized benzylguanidine derivatives bearing iminodiacetate (IDA) and aminoethylglycine (AEG) tridentate chelating moieties, respectively. These ligands were labeled with technetium-99m via [99mTc][Tc(CO)3(H2O)3]+ synthon to form desired radioactive complexes 9 and 10. The radiolabeling yields of the complexes obtained were >90% as confirmed by radio-HPLC. The HPLC purified complexes were used for in vitro and in vivo evaluation to understand the true biological efficacy. Structural characterization of the radiolabeled complexes was carried after synthesizing and characterizing their Re-analogues. RESULTS: Cell uptake studies with the radiolabeled complexes in SK-N-SH neuroblastoma cell lines revealed reduced uptake in the cells (<1% of incubated radioactivity/106 cells) in comparison to n.c.a. [125I]mIBG (~12%). However, limited specificity (~60%) was observed for the complexes as ascertained through desmethylimipramine (DMI) inhibition. Biodistribution studies in normal Wistar rats exhibited desired non-target clearance pharmacokinetics for the complexes but in vivo NET efficacy in myocardium for the neutral complex 10 could not be established. CONCLUSIONS: Tridentate [99mTc]Tc(CO)31+ chelation approach severely affects biological behavior of the present small bioactive molecule under study to a significant extent in comparison to monodentate ligation in 99mTc-4 + 1 strategy.

Use of ESI-MS for semi-quantitative estimation of inactive precursor in no-carrier-added 131I- meta-Iodobenzylguanidine radiopharmaceutical preparation.

No-carrier-added (nca)-131I-meta-Iodobenzylguadine (mIBG) is a clinical agent used for the therapy of Neuroendocrine tumors. It is prepared by reaction of radioiodine with precursors that are chemically different from mIBG. The precursor in few cases is structurally similar and may co-elute along during purification step. Presence of these precursors in final radiolabeled formulation may affect the clinical behaviour of the radiopharmaceutical. The present paper describes the use of Electron-spray ionization-Mass Spectrometry (ESI-MS) where up to nano-molar concentrations of these precursors could be estimated with high precision in the final radiolabeled formulation.

Synthesis and biodistribution studies of 99m Tc labeled fatty acid derivatives prepared via "Click approach" for potential use in cardiac imaging.

123 I-Iodophenylpentadecanoic acid (IPPA) is a metabolic agent used in nuclear medicine for diagnosis of myocardial defects. Efforts are underway worldwide to develop a 99m Tc substitute of the above radiopharmaceutical for the aforementioned application. Herein, we report synthesis and biodistribution studies of 99m Tc labeled fatty acids (8, 11, and 15 carbons) obtained via "click chemistry" for its potential use in myocardial imaging. ω-Bromo fatty acids (8C/11C/15C) were synthetically modified at bromo terminal to introduce a heterocyclic triazole with glycine sidearm in a five step procedure. Modified fatty acids were subsequently radiolabeled with preformed [99m Tc(CO)3 ]+ synthon to yield the desired fatty acid complexes which were evaluated in Swiss mice. All the radiolabeled complexes were obtained with radiochemical purities >80%, as characterized by HPLC. Biodistribution studies of all three complexes in Swiss mice showed myocardial uptake of ~6-9% ID/g at 2 minutes post-injection, close to* I-IPPA (~9% ID/g). Complexes exhibited significant retention in the myocardium up to 30 minutes (~1% ID/g) but were lower to the standard agent (~7% ID/g). Similar uptake of activity in myocardium for the newly synthesized complexes in comparison to 125 I-IPPA along with favorable in vivo pharmacokinetics merits potential for the present "click" design of complexes for myocardial imaging.

Synthesis and evaluation of 68Ga labeled palmitic acid for cardiac metabolic imaging.

This work evaluates the potential of a 68Ga labeled long chain 16C fatty acid for cardiac metabolic imaging. For radiolabeling with 68Ga, hexadecanedioic acid was coupled with the chelator p-NH2-Bn-NOTA. Under the optimized conditions, NOTA-hexadecanoic acid could be radiolabeled with 68Ga in ≥95% yields. In biodistribution studies carried out in Swiss mice, 68Ga-NOTA-hexadecanoic acid showed low myocardial uptake at 2 min p.i. (3.7 ±â€¯1.3%ID/g). While 68Ga-NOTA-hexadecanoic acid cleared rapidly from non-target organs such as blood, lungs, intestine and kidney, wash out from liver was slow. Radio-HPLC analyses of myocardial extracts of rats injected with 68Ga-NOTA-hexadecanoic acid confirmed its metabolic transformation in the myocardium.

Bulk Scale Formulation of Therapeutic Doses of Clinical Grade Ready-to-Use 177Lu-DOTA-TATE: The Intricate Radiochemistry Aspects.

INTRODUCTION: 177Lu-DOTA-TATE is a clinically useful and promising therapeutic radiopharmaceutical for peptide receptor radionuclide therapy of neuroendocrine tumors (NETs) overexpressing somatostatin receptors. Currently, the radiopharmaceutical is prepared in-house at nuclear medicine centers, thereby restricting its use to limited centers only. In this article, the authors describe systematic studies toward bulk scale formulation of "ready-to-use" 177Lu-DOTA-TATE using medium specific activity 177Lu (740-1110 GBq/mg) at a centralized radiopharmacy facility. METHODS: In an optimized protocol, 177Lu-DOTA-TATE synthesis was carried out by direct heating of 177LuCl3 (Sp. act. 740-1110 GBq/mg) with DOTA-TATE peptide (1.5-3.0 equivalents) in ammonium acetate buffer (0.2 M) containing 2,5-dihydroxy benzoic acid (gentisic acid). Thereafter, the crude labeled product was purified using a Sep-Pak® C18 column and diluted with acetate buffer-gentisic acid (1.5% w/v) solution to final radioactive concentration of 740 MBq/mL. This was further sterilized and dispensed as 7.4 GBq patient dose/vial with 2 days postformulation calibration. RESULTS: A peptide/metal ratio of 1.5-3.0 is essential for complexation wherein radiolabeling yields >90% are obtained minimizing free 177Lu waste. For formulation of 7.4 GBq patient dose (2 days postproduction), even specific activity of about 555 GBq/mg was found to be adequate for the radiometal. The ready-to-use 740 MBq/mL 177Lu-DOTA-TATE formulation with gentisic acid (1.5% w/v) is observed to be safe for human use for more than 1 week (radiochemical purity >98%) from the day of production when stored at -70°C. However, the target specificity may get affected beyond 2 days as the total peptide content for 7.4 GBq dose may exceed the critical peptide limit of 300 µg. Patient treatment carried with several batches of present formulation in diseased NET patients exhibited desired distribution at the tumor and its metastatic site. CONCLUSIONS: A ready-to-use formulation of 177Lu-DOTA-TATE was successfully prepared and optimized for regular bulk scale production and supply to distant nuclear medicine centers.

Radiosynthesis and evaluation of a 99mTc-folic acid radiotracer prepared using [99mTcN(PNP)]2+ metal fragment.

Folate receptors (FR) are over-expressed on a wide variety of tumor cells and are a potential molecular target for radiolabeled folates. In this respect, several SPECT and PET based radiofolates have been evaluated in the past albeit with their high renal uptake posing limitation towards their clinical use. To overcome this, a new 99mTc labeled folic acid was synthesized via the use of [99mTcN(PNP)]2+ metal fragment, where the presence of the latter pharmacophore redirects in vivo clearance via the hepatobiliary pathway. In this respect, folic acid was derivatized at the γ-acid group with a cysteine BFCA (bifunctional chelating agent) and subsequently reacted with the preformed [99mTcN]2+ intermediate in presence of PNP2 (bisphosphine) ligand, to yield the final complex. While preliminary, in vivo distribution of the complex exhibited high association of activity with liver and intestines and provided support to the rationality of the present design as clearance of labeled folic acid could be effected via the hepatic route, the in vitro studies of the folic acid-cysteine conjugate carried out in KB-31 cells, did not show much promise with reduction in receptor affinity in comparison with the native folic acid. The route followed herein to prepare a folic-acid based radiotracer constitutes the first report of radiolabeling folic acid using the [99mTcN(PNP)]2+ as a radiosynthon. Modification in the structure of conjugate by linking the BFCA through a long-chain linker can be envisaged to improve the affinity of [99mTcN(PNP)]-folic acid complex towards FRs.

68Ga labeled fatty acids for cardiac metabolic imaging: Influence of different bifunctional chelators.

Development of 68Ga labeled fatty acids is of immense interest due to the availability of 68Ga through a generator and its superiority over SPECT based tracers in carrying out dynamic imaging on a PET scanner. Our present work explores the influence of different chelators on the cardiac uptake and pharmacokinetics of the 68Ga-labeled fatty acids. Two new 68Ga labeled fatty acids were synthesized by conjugation of 11-aminoundecanoic acid with the bifunctional chelators (BFCs) viz. p-SCN-Bn-DTPA (S-2-(4-isothiocyanatobenzyl)-diethylenetriaminepentaacetic acid) and p-SCN-Bn-NODAGA (S-2-(4-isothiocyanatobenzyl)-1,4,7-triazacyclononane-1-glutaric acid-4,7-acetic acid) and their comparison was carried out with the previously reported 68Ga-NOTA-undecanoic acid. Both the conjugates were radiolabeled with 68Ga in high yields and purities (>95%). Their formation was established by preparation and characterization of their inactive analogs with natGa at macroscopic levels. Biodistribution studies of the complexes in Swiss mice showed lower initial myocardial uptake for 68Ga-NODAGA-undecanoic acid (3.8±0.6%ID/g) and 68Ga-DTPA-undecanoic acid (1.3±0.5%ID/g) complexes in comparison to previously reported 68Ga-NOTA-undecanoic acid complex (7.4±2.8%ID/g) at 2min p.i. However, significant retention of the tracer in the myocardium was observed in the case of 68Ga-NODAGA-undecanoic complex, which led to improved heart/non-target ratios of the complex over time in comparison to the other 68Ga complexes. Similarly, the DTPA complex exhibited increased washout from the liver in comparison to other 68Ga derivatives. The ß oxidation mechanism in myocytes was investigated by isolating the myocardial extract post intravenous injection of the respective 68Ga complexes and analyzing them by radio-HPLC, which showed metabolic transformation of the parent fatty acid complex peak in all the three complexes. This study has provided an insight into the design characteristics of 68Ga labeled fatty acids to achieve the desired myocardial imaging characteristics.

Synthesis and evaluation of a (68)Ga labeled folic acid derivative for targeting folate receptors.

Present work evaluates the potential of a newly synthesized (68)Ga-NOTA-folic acid conjugate for PET imaging of tumors over-expressing folate receptors (FRs). NOTA-folic acid conjugate was synthesized and characterized. It was radiolabeled with (68)Ga in ≥ 95% radiolabeling yields. In vitro cell binding studies showed a maximum cell uptake of 1.7±0.4% per million KB cells which was completely blocked on addition of cold folic acid showing specificity towards the FRs. However, further studies in tumor xenografts are warranted in order to assess the potential of (68)Ga-folic acid complex for imaging tumors over-expressing FRs.

A (99m)Tc-labeled misonidazole analogue: step toward a (99m)Tc-alternative to [18F]fluromisonidazole for detecting tumor hypoxia.

The PET radiopharmaceutical [(18)F]Fluromisonidazole ([(18)F]FMISO) is presently the agent of choice for the clinical imaging of tumor hypoxia. Considering the logistic advantages of (99m)Tc and wider availability of SPECT machines, a (99m)Tc-radiopharmaceutical for this purpose constitutes an attractive choice. In the work presented here, a misonidazole analogue was radiolabeled with (99m)Tc(CO)3 core and the complex was evaluated in Swiss mice bearing fibrosarcoma tumor. The results obtained are compared with the biodistribution of [(18)F]FMISO carried out in the same tumor-bearing animal model. Misonidazole-(99m)Tc(CO)3 complex showed significant uptake and retention in tumor. Notably, the rate of clearance of misonidazole complex from the tumor was slower than that of [(18)F]FMISO. The maximum tumor/muscle ratio obtained with misonidazole-(99m)Tc(CO)3 complex was significantly higher than that of [(18)F]FMISO. The study constitutes a positive step toward the development of a (99m)Tc-analogue of [(18)F]FMISO.

Development of 68Ga-labeled fatty acids for their potential use in cardiac metabolic imaging.

While [(11)C]palmitate continues to be a promising tracer for cardiovascular Positron Emission Tomography (PET) imaging, unfavourable logistics due to the short half-life of (11)C (20 min) and cumbersome labeling methodologies are the major impediments that limit its widespread use. In order to circumvent such limitations, an attempt has been made to explore the potential of (68)Ga-labeled fatty acid analogs for metabolic imaging owing to the availability of (68)Ga through a (68)Ge/(68)Ga generator on an on-demand basis. In this study, two fatty acid conjugates were synthesized by conjugation of p-SCN-benzyl NOTA with the ω-amino group of 11-amino undecanoic acid and 12-amino dodecanoic acid, respectively, under alkaline conditions. Both derivatives were radiolabeled in high yields with (68)Ga obtained from an in-house (68)Ge/(68)Ga generator. Biodistribution studies in Swiss mice showed reasonable myocardial uptake at 2 min for both derivatives (7.4 ± 2.8% ID/g for 11-carbon fatty acid-NOTA conjugate and 6.4 ± 2.1% ID/g for 12-carbon fatty acid-NOTA conjugate), which cleared rapidly over 30 min. However, significant activity was found in blood for both tracers, with heart/blood ratios observed to be below 0.5 at all time points, diminishing the potential of the synthesized complexes for cardiac imaging.

A study on nitroimidazole-99mTc(CO)3 complexes as hypoxia marker: some observations towards possible improvement in in vivo efficacy.

INTRODUCTION: Hypoxia plays a negative role in the clinical management of cancer. Detection of hypoxic status of a cancer is important for selecting patients for hypoxia directed therapy. Though [(18)F]fluoromisonidazole ([(18)F]FMISO), a PET radiopharmaceutical, is presently being used in the clinic for the detection of hypoxia, considering the logistical advantages of (99m)Tc and wider availability of SPECT scanners, a radiopharmaceutical based on this isotope may find wider applicability. METHODS: Nine nitroimidazole (2-, 4- and 5-nitroimidazole) ligands were synthesized and radiolabeled using [(99m)Tc(CO)3(H2O)3](+) precursor to obtain a group of complexes possessing different single electron reduction potential (SERP), overall charge and lipophilicity, the three attributes which decide the efficacy of the complex to detect hypoxic cells in vivo. The nitroimidazole-(99m)Tc(CO)3 complexes as well as [(18)F]FMISO were evaluated in fibrosarcoma tumor bearing mice. RESULTS: The (99m)Tc(CO)3 complexes of nitroimidazole iminodiacetic acid (IDA) showed better tumor uptake and retention than nitroimidazole diethylenetriamine (DETA) and nitroimidazole aminoethylglycine (AEG) complexes. Tumor uptake observed with [(18)F]FMISO was higher than any of the nitroimidazole-IDA- (99m)Tc(CO)3 complexes. However, [(18)F]FMISO clearance from tumor was found to be faster compared to 2-nitroimidazole-IDA-(99m)Tc(CO)3 complex. Observed tumor uptake and retention of the radiotracers evaluated could be correlated to its blood clearance pattern and SERP. CONCLUSIONS: Results of the present study indicated that uptake of the radiotracer in tumor is closely associated with its rate of clearance from blood. The study also indicated that along with SERP, clearance of radiotracer from blood (net effect of charge and lipophilicity) is a critical factor which decides the in vivo efficacy of the hypoxia detecting radiopharmaceutical.

Preparation and evaluation of a 99mTcN-PNP complex of sanazole analogue for detecting tumor hypoxia.

A sanazole derivative, having a favorable single electron reduction potential (SERP) value compared to that of misonidazole, was synthesized and radiolabeled with [(99m)TcN(PNP)] precursor to evaluate its potential as a hypoxia imaging agent. The complex, which was lipophilic, could be prepared in good yields and challenging studies with cysteine showed stability of the complex against trans-chelation. However, despite being lipophilic as well as possessing favorable SERP value, biodistribution studies of this complex in fibrosarcoma tumor bearing Swiss mice showed low uptake in tumor. This observation is possibly attributed to fast clearance of the complex from blood, whereby the complex spends insufficient time in tumor to get reduced and trapped. Though uptake in tumor was low, slow clearance of activity from tumor suggests reduction and trapping of the complex in hypoxic cells. The present (99m)Tc-complex demonstrated acceptable values of tumor to blood (TBR) and tumor to muscle (TMR) ratios. However, low uptake in tumor which may not be indicative of the actual hypoxic status of the tumor, limit the utility of the complex to detect tumor hypoxia.

On the structural modification of 2-nitroimidazole-(99m)Tc(CO)(3) complex, a hypoxia marker, for improving in vivo pharmacokinetics.

INTRODUCTION: A 2-nitroimidazole-(99m)Tc(CO)(3) complex reported earlier showed promise with respect to its uptake and retention in hypoxic tumor. However, significant uptake and slow clearance from liver imposed severe limitations towards advocating its possible practical utility. In an attempt to improving its liver clearance, an ether linkage, which is known to help in liver clearance, was introduced in the molecule. METHODS: The 2-nitroimidazole iminodiacetic acid (IDA) derivative containing an ether linkage was synthesized in a five step procedure from 2-nitroimidazole. This derivative was radiolabeled using [(99m)Tc(CO)(3)(H(2)O)(3)](+) precursor complex. The corresponding Re(CO)(3) analogue was also synthesized in the macroscopic level for structural characterization. The (99m)Tc(CO)(3) complex was evaluated in an animal model bearing fibrosarcoma tumor. RESULTS: The in vivo evaluation of the complex indicated that, as envisaged, introduction of the ether linkage has improved clearance from the liver. The complex also showed higher retention in tumor compared to the 2-nitroimidazole-IDA-(99m)Tc(CO)(3) complex reported earlier. Though the tumor to muscle ratio improved with time, the tumor to blood ratio did not show any significant improvement. Despite improved liver clearance, there was significant liver activity present even at 3h p.i. attributable to gradual accumulation of activity cleared from muscle and blood. CONCLUSIONS: Though the introduction of ether linkage improved liver clearance of the modified 2-nitroimidazole complex, it was found that a single ether linkage was not sufficient to achieve the desirable level of clearance. Probably, a linker with multiple ether groups, such as a di- or tri-ethylene glycol spacer, may be a possible solution to this issue.

Efficient production of therapeutic doses of [131I]-metaiodobenzylguanidine for clinical use.

[(131)I]-metaiodobenzylguanidine (mIBG) is a known radiopharmaceutical used for the treatment of neuroendocrine tumors. The development of therapeutic [(131)I]-mIBG doses at production level is highly challenging due to rapid product degradation and high radiation exposures to the production plant personnel. In the present work, a working module for the production of 10 doses (100 mCi each) in a single operation was developed following copper (I) assisted isotope exchange. The labeled product complies with the pharmaceutical specifications suitable for in-vivo patient use.

On the isolation and evaluation of a novel unsubstituted 5-nitroimidazole derivative as an agent to target tumor hypoxia.

The presence and extent of hypoxic regions in cancerous tissue bears a negative influence on the effectiveness of radiation therapy and chemotherapy of the cancer hence estimation of hypoxia is an important problem. Several (99m)Tc-labeled nitroimidazole-based non-invasive agents have been tried for this purpose but none had optimal characteristics and the search continues. Herein we report, for the first time to the best of our knowledge, the isolation, (99m)Tc(CO)(3) labeling and evaluation of an unsubstituted 5-nitroimidazole derivative obtained as a side product during the synthesis of 4-nitroimidazole derivative. The (99m)Tc(CO)(3)-labeled complex of 5-nitroimidazole derivative could be prepared in excellent yield under mild conditions. Its evaluation in fibrosarcoma tumor bearing Swiss mice showed uptake and slow clearance of injected activity from tumor. The tumor-to-muscle ratio was found to be very high but tumor-to-blood ratio greater than 1 could not be obtained throughout the limited time point study. The study revealed that complex under investigation has features similar to other 2-nitroimidazole complexes so far as the retention of injected activity in tumor is concerned.