Development and Evaluation of 99mTc Tricarbonyl Complexes Derived from Flutamide with Affinity for Androgen Receptor.

With the objective to develop a potential 99mTc radiopharmaceutical for imaging the androgen receptor (AR) in prostate cancer, four ligands bearing the same pharmacophore derived from the AR antagonist flutamide were prepared, labeled with 99mTc, and their structures corroborated via comparison with the corresponding stable rhenium analogs. All complexes were obtained with high radiochemical purity. Three of the complexes were highly stable, and, due to their favorable physicochemical properties, were further evaluated using AR-positive and AR-negative cells in culture. All complexes exhibited considerable uptake in AR-positive cells, which could be blocked by an excess of flutamide. The efflux from the cells was moderate. They also showed significantly lower uptakes in AR-negative cells, indicating interactions with the AR receptor. However, the binding affinities were considerably reduced by the coordination to 99mTc, and the complex that exhibited the best biological behavior did not show sufficient specificity towards AR-positive cells.

[99m Tc]Tc-HYNIC-EcgDf21: A defensin short analogue with potential application in infection foci imaging.

Opportunistic infections are a problem of great relevance in public health and the precise detection and localization of infection in the early stages of the disease is of great importance for patient management as well as cost containment. Our proposal seeks to contribute to developing a new agent that meets the needs of diagnosis and follow-up of fungal and bacterial infections, focused on the design of a radiotracer with the potential for recognition of hidden infection foci. Defensins are plant antimicrobial peptides that not only show activity against plant pathogens but also against human ones. A short analogue of EcgDf1 defensin, EcgDf21d (NH2 -ERFTGGHCRGFRRRCFCTKHC-COOH), was labelled through the formation of a 99m Tc-HYNIC complex which was assessed for physicochemical and biological behaviour both in vitro and in vivo. The [99m Tc]Tc-HYNIC-EcgDf21 labelling procedure rendered a single product with remarkably high RCP and stability in the labelling milieu. The Log p value indicated that [99m Tc]Tc-HYNIC-EcgDf21 has a hydrophilic behaviour, confirmed by the biodistribution profiles. The optimal uptake value was obtained for Candida albicans infection model reaching a lesion/muscle ratio of 3, this correlates with in vitro binding studies, and the lesion can be definitely observed in the scintigraphic images.

Development and characterization of two novel 68 Ga-labelled neuropeptide Y short analogues with potential application in breast cancer imaging.

In vivo receptor targeting with radiolabelled peptide-based probes is an attractive approach for the development of novel radiotracers for molecular imaging. This work presents the development and characterization of two novel neuropeptide Y analogues labelled with a positron emitter 68 Ga, for potential use in breast cancer imaging. Both analogues share the same amino acid sequence and were derivatized with NOTA through either a lysine linker (L1) or an acetylated lysine (L2). In both cases, a single product with radiochemical purity higher than 95% was obtained. The two complexes were hydrophilic, showed remarkable in vitro stability, good cellular uptake, binding affinity in the nanomolar range and high cellular internalization rate. Biodistribution studies revealed low blood uptake and elimination through the urinary tract. The addition of an acetyl group in the spacer increased the lipophilicity of C2 and modified the reactivity of the ε-amino group of the lysine which resulted in lower protein binding and lower percentage of injected dose in bladder and urine. The tumour versus muscle ratio was (3.8 ± 0.4) for 68 Ga-L1 and (4.7 ± 0.4) for 68 Ga-L2. These results encourage performing further studies in order to complete the evaluation of both tracers as potential radiopharmaceutical for breast cancer imaging.

Development and characterization of a 99m Tc-labeled Neuropeptide Y short analog with potential application in breast cancer imaging.

The aim of this work was to develop and evaluate a 99m Tc-labeled neuropeptide Y derivative with affinity toward Y1-receptor. The selected amino acid sequence included nine amino acids derived from the C-terminal portion of the NPY complemented with the addition of one cysteine-mercaptoacetic acid moiety to bind the radiometal. Labeling was achieved through the preparation of a 3 + 1 nitrido complex. Physicochemical evaluation, cell uptake, internalization and externalization studies, and competitive assays were performed. Biodistribution experiments were carried out in normal and tumor-bearing mice. A single product with radiochemical purity >90% and high stability was obtained. In vitro analysis showed specific cellular uptake, IC50 of 73.2 nM, and a high internalization rate (80%). Biodistribution studies showed low blood and renal uptake and combined hepatobiliary and urinary elimination. Preliminary studies in mice bearing induced breast tumors rendered promising uptake values.

Development and evaluation of a 99mTc(V)-nitrido complex derived from estradiol for breast cancer imaging.

Estrogen receptors are overexpressed in 70% of breast cancer and identification of their presence is important to select the appropriate treatment. This work proposes the preparation and evaluation of an estradiol derived as potential ER imaging agent. Ethinylestradiol was derivatized to introduce a dithiocarbamate function for Tc coordination. Labeling was achieved through the formation of a symmetric Tc(V)-nitrido complex with a radiochemical purity (RCP) > 95%. Physicochemical evaluation, cell uptake, biodistribution in normal animals and in nude mice bearing induced ER + breast tumors showed promising results.

Evaluation of a Labelled Bacteriophage with 99mTc as a Potential Agent for Infection Diagnosis.

BACKGROUND AND OBJECTIVE: The design of target-specific molecular imaging probes to determine infection sites are mainly based on the biochemistry of the inflammatory response that may lead to an ideal agent for infection imaging. Infectious diseases timely and specifically diagnosed can be clinically challenging but essential for the patient's recovery. Laboratory tests can detect the responsible microorganism but cannot discriminate between sterile inflammatory disease and truly infectious disease. On the other hand, scintigraphic images, can pinpoint the infection in the body. METHODS: Bacteriophages (phages) are viruses that infect specific bacterial strains. Given the composition of the protein capsid, they could be used as radiopharmaceuticals to diagnose bacterial infection. In this case, PP7 phage was labelled and evaluated as a specific tracer for Pseudomonas aeruginosa infections. 99mTc-Phage synthesis used HYNIC as a bifunctional agent. Physicochemical evaluation included studies such as stability in time, ligand exchange, lipophilicity and bacterial binding assay. Three groups of animals namely; healthy, infected with Pseudomonas aeruginosa and induced sterile inflammation were used to conduct biological evaluation Results: The radiolabelling process required size exclusion purification of the 99mTc-Phage, which was obtained with a radiochemical purity higher than 90%, during 18 hours post labelling. The collective accumulation in the stomach, small intestine and large intestine and thyroid of 99mTc-Phage was negligible, indicating no in vivo reoxidation. The complex presented urinary elimination. Target/ non-target ratio (T/NT) was determined both for sterile inflammation and for infection. Values were 2.5 ± 0.4 and 4.2 ± 0.3 respectively. These values indicate significant differences between sterile inflammation and infection by Pseudomonas aeruginosa (p<0.05 unpaired two sided t-test). CONCLUSION: Targeted biodistribution profile and good T/NT ratios, indicate that this complex presents enough specificity to discriminate between infection caused by Pseudomonas aeruginosa and sterile inflammation.

Preparation and biological evaluation of (99m)Tc-labelled phenazine dioxides as potential tracers for hypoxia imaging.

The aim of this study was to investigate the capability of phenazine dioxides, recognized bioreductive antitumour agents, as carriers for (99m)Tc in order to generate potential theranostic radiopharmaceuticals towards hypoxic solid tumours. Two different phenazine dioxides were used as ligands for the (99m)Tc-tricarbonyl core in order to prepare the potential radiopharmaceuticals. The main physicochemical and biological properties were evaluated. Biodistribution of the two radiotracers was studied at different time points after intravenous injection in tumour bearing animals. Both compounds were obtained in high yield and radiochemichal purity. They were stable in labelling milieu, in human plasma and in the presence of histidine. Biodistribution studies in mice were characterized by slow blood clearance and persistent liver uptake, results that correlate with the values of lipophilicities and protein binding. Both the complexes showed good tumour uptake, which remained constant during the studied period. Tumour/muscle ratios proved very favourable, comparable to those of FMISO in the same animal model. On the other hand, tumour/blood ratios were low due to high blood uptake. The use of phenazine dioxides as ligands for the preparation of potential (99m)Tc-radiopharmaceuticals towards solid tumours is possible since tumour uptake and retention are promising although high blood and liver uptake are drawbacks worth consideration.