Design of a New 99mTc-radiolabeled Cyclo-peptide as Promising Molecular Imaging Agent of CXCR4 Receptor: Molecular Docking, Synthesis, Radiolabeling, and Biological Evaluation.

INTRODUCTION: C-X-C Chemokine receptor type 4 (CXCR4) is often overexpressed or overactivated in different types and stages of cancer disease. Therefore, it is considered a promising target for imaging and early detection of primary tumors and metastasis. In the present research, a new cyclo-peptide radiolabelled with 99mTc, 99mTc-Cyclo [D-Phe-D-Tyr-Lys (HYNIC)- D-Arg-2-Nal-Gly-Lys(iPr)], was designed based on the parental LY251029 peptide, as a potential in vivo imaging agent of CXCR4-expressing tumors. METHODS: The radioligand was successfully prepared using the method of Fmoc solid-phase peptide synthesis and was evaluated in biological assessment. Molecular docking findings revealed high affinity (binding energy of -9.7 kcal/mol) and effective interaction of Cyclo [D-Phe- D-Tyr-Lys (HYNIC)-D-Arg-2-Nal-Gly-Lys(iPr)] in the binding pocket of CXCR4 receptor (PDB code: 3OE0) as well. RESULT: The synthesized peptide and its purity were assessed by both reversed-phase high-performance liquid chromatography (RP-HPLC) and mass spectroscopy. High stability (95%, n = 3) in human serum and favorable affinity (Kd = 28.70 ± 13.56 nM and Bmax = 1.896 ± 0.123 fmol/mg protein) in the B16-F10 cell line resulted. Biodistribution evaluation findings and planar image interpretation of mice both showed high affinity and selectivity of the radiotracer to the CXCR4 receptors. CONCLUSION: Therefore, the findings indicate this designed radioligand could be used as a potential SPECT imaging agent in highly proliferated CXCR4 receptor tumors.

Synthesis, radiolabeling and evaluation of [99mTc][Tc-HYNIC/EDDA]-Met(O) as a early agent for amino acid metabolic imaging in C6 glioblastoma tumor.

Amino acid metabolism is recognized as a target for medical imaging due to its increase in malignant cells. Several radiotracers with primary achievement and possible subsequent chances have been designed and tested to image amino acid metabolism. Here, we report a new amino acid conjugate, with the purpose of extending [99mTc][Tc-HYNIC/EDDA]-Met(O) for single photon emission tomography (SPECT) imaging. The S-oxo-l-methionine (Met(O)) amino acid hydrazinonicotinamide (HYNIC) chelator conjugate (HYNIC-Met(O)) was prepared, using Fmoc solid-phase synthesis, and was radiolabeled with [99mTc]technetium pertechnetate, using tricine and ethylenediamine-N,N-diacetic acid (EDDA) as co-ligands. In vitro cellular uptake profile and saturation binding of radiotracer were determined on C6 glioma cells. Biodistribution and imaging studies were carried out on rat bearing C6 tumor tissue grafts. [99mTc][Tc-HYNIC/EDDA]-Met(O) was prepared in high yield and radiochemical purity (>98 %). The partition coefficient result showed that radioconjugate was very hydrophilic. The radioconjugate indicated both high cell uptake and in vitro internalization. Low nanomolar dissociation constant (66.02 nM) in C6 glioma cells was obtained for it as well. [99mTc][Tc-HYNIC/EDDA]-Met(O) revealed magnificent tumor uptake at early time points, with 1.98 ± 0.33 % injected activity per gram tumor (% IA/g) at 30 min post injection. The tumor uptake continued for 1 and 2 h and was 0.45 ± 0.33 % IA/g at 4 h. The uptake in other organs decreased much more rapidly causing high tumor to normal organ ratios so that the highest ratio of 13.25 of tumor-to-muscle at 60 min after injection was obtained with high contrast in gamma imaging. These results point out a very favorable [99mTc]Tc-labeled amino acid for targeting amino acid metabolism through target system L amino acid transporter (LAT1) in malignant cells especially C6 glioma cells. [99mTc][Tc-HYNIC/EDDA]-Met(O) manifests extremely good distribution, excretion and imaging attributes. So it seems to be an appropriate nominate for clinical imaging.

Inflammation scintigraphy imaging through a novel antimicrobial peptide labeled with technetium-99m in an animal model.

PURPOSE: Antimicrobial peptides (AMPs), due to their biological properties, have great potential for radiopharmaceutical development. In this research, a new antimicrobial peptide, derived from a 21-residue antimicrobial peptide microcinJ25 (MccJ25), was utilized to diagnose inflamed sites in mice bodies after labeling with [99mTc]Tc. MATERIALS AND METHODS: An antimicrobial peptide derivative was connected with an efficient chelator, hydrazinonicotinamide, and then labeled with [99mTc]Tc. The binding of labeled microcinJ25 conjugate to lymphocyte was investigated in vitro. Turpentine oil-induced inflammation uptake and tissue distribution were assessed through the animal pattern. Detector scanning was done through scintigraphy post injection of [99mTc]Tc-radiopeptide within different time points. RESULTS: High radiochemical purity (>98%) was obtained for [99mTc]Tc-radiopeptide. Lymphocyte binding assessment showed specific cell binding. Binding Inhibition was observed when additional unlabeled conjugate was used. In in vivo biological distribution studies, the uptake for inflamed tissue was 1.52 ± 0.12%ID/g. The inflammation site was visualized by scintigraphy imaging at 1 up to 2 hours. CONCLUSION: Based on our results this new designed [99mTc]Tc-radiopeptide was able to detect inflammation sites early and with high resolution, and could be considered a promising diagnostic candidate in inflammatory diseases.

Evaluation of [99mTc][Tc-HYNIC/EDDA]-Tyr as a target for metabolic tumor imaging in B16F10 melanoma tumor.

Objectives: Clinical interest in metabolic imaging of cancer has been growing in recent years. The increase in protein metabolism of cancer cells is interesting target for metabolic tumor imaging, for which radiolabeled amino acids can be applied. The aim of this study was to evaluate a newly developed radiolabeled amino acid as an imaging protein metabolism in melanoma tumor. Methods: The radiolabeled tyrosine ([99mTc][Tc-HYNIC/EDDA]-Tyr) was prepared and its biological properties was evaluated in B16F10 melanoma tumor. Moreover organs uptake and tumor accumulation were measured in mouse bearing B16F10 melanoma tumor. Results: Radiolabeled tyrosine was attached in B16F10 melanoma cells and showed the cell binding capacity of 13.82±0.73%. In animal study, the accumulation of radiolabeled tyrosine was observed in B16F10 melanoma tumor (2.15±0.09 %ID/g) after 30 min post injection, so that the uptake ratio of tumor to muscle was about 5.11. Through scintigraphy process the melanoma tumor clearly visualized in mice at 30 min post injection. Conclusion: These data suggest that the novel radiotracer ([99mTc][Tc-HYNIC/EDDA]-Tyr) as an protein metabolism imaging agent, is able to transfer into melanoma cells and show great expectation for the clinical application in the imaging of melanoma tumors.

[99m Tc-HYNIC/EDDA]-MccJ25 antimicrobial peptide analog as a potential radiotracer for detection of infection.

Bacterial infections are a serious risk to human health, and therefore techniques for early detection of infectious foci need to be further developed to begin treatment quickly and achieve better results. Antimicrobial peptides labeled with gamma-emission radio nuclides are important diagnostic radiotracers in nuclear medicine. This study was conducted to evaluate the potential of a 99m Tc-labeled MicrocinJ25 (MccJ25) antimicrobial peptide analog for early detection of infection. For this purpose, a HYNIC conjugated cyclic peptide derivative based on the primary structure of MccJ25 peptide was prepared and labeled by 99m Tc with tricine and EDDA as coligands. The [99m Tc-HYNIC/EDDA]-MccJ25 peptide analog showed high radiochemical purity (˃90% (n = 5)) which was stable up to 24 hr after labeling. The radiotracer showed specific uptake to the Escherichia coli (E. coli) bacterial (40.45 ± 5.21%) at 1 hr incubation. High kidneys uptake of radioactivity (4.71 ± 0.84% and 3.76 ± 0.45% ID/g at 1 and 4 hr after injection respectively) demonstrates that most of the whole body clearance was proceeded via the urinary system. Significant radioactivity uptake (1.71 ± 0.34%ID/g) was observed in thigh muscle of mouse with E. coli induced infection at 1 hr after injection. In the blocking test, due to the significant decrease of radioactivity uptake in the infection site (0.62 ± 0.21%ID/g after 1 hr), the specificity of infection uptake was reviled. Despite the high activity of the bladder due to urinary excretion, the infected area was somewhat visible. Hence, the results indicate the potential of this new radiotracer to be used as a diagnostic agent in E. coli infections.

Preparation and assessment of a new radiotracer technetium-99m-6-hydrazinonicotinic acid-tyrosine as a targeting agent in tumor detecting through single photon emission tomography.

The goal of investigation was to bring up an impressive way to synthesize technetium-99 m-6-hydrazinonicotinic acid-tyrosine (99mTc-HYNIC-Tyr), a newfound radiotracer, and to assess the capacity of being as a tumor scintigraphy agent. The conjugate was prepared by solid phase method using tritely chloride resin. The precursor HYNIC-Tyr was labeled with 99mTc which was accomplished at 100 °C through the coligands tricine and EDDA. Furthermore, the serum albumin binding, cellular attachment, organs uptake and tumor accumulation were measured. C6 glioma cells were used for cellular and tumor uptake studies. 99mTc-HYNIC-Tyr was prepared with labeling yield of >99% (n = 3). Radiotracer showed stability in serum proteins in incubates temperature. Specific cellular attachment of radiotracer was noticeable in C6 glioma cells with dissociation constant in nano molar range (21.03 ± 1.54 nM). The amount of uptake in C6 rat glioma xenograft was 2.61 ± 0.12 percent of injection dose per gram after 30 min. In whole-body scintigraphy, C6 glioma tumor was easy to be traced and interpreted at 1 h after administration of radiotracer. Our data suggest that 99mTc-HYNIC-Tyr, a new radiotracer based on amino acid, efficiently differentiates the tumor cells and goes into them. Our results indicate that this radiotracer has excellent capacity to detect tumor cells in rat and to be included as a radiopharmaceutical for detecting cancer tumors.

Design of peptide-based inhibitor agent against amyloid-ß aggregation: Molecular docking, synthesis and in vitro evaluation.

Formation of the amyloid beta (Aß) peptide aggregations represents an indispensable role in appearing and progression of Alzheimer disease. ß-sheet breaker peptides can be designed and modified with different amino acids in order to improve biological properties and binding affinity to the amyloid beta peptide. In the present study, three peptide sequences were designed based on the hopeful results of LIAIMA peptide and molecular docking studies were carried out onto the monomer and fibril structure of amyloid beta peptide using AutoDock Vina software. According to the obtained interactions and binding energy from docking, the best-designed peptide (d-GABA-FPLIAIMA) was chosen and synthesized in great yield (%96) via the Fmoc solid-phase peptide synthesis. The synthesis and purity of the resulting peptide were estimated and evaluated by Mass spectroscopy and Reversed-phase high-performance liquid chromatography (RP-HPLC) methods, respectively. Stability studies in plasma and Thioflavin T (ThT) assay were performed in order to measure the binding affinity and in vitro aggregation inhibition of Aß peptide. The d-GABA-FPLIAIMA peptide showed good binding energy and affinity to Aß fibrils, high stability (more than 90%) in human serum, and a reduction of 20% in inhibition of the Aß aggregation growth. Finally, the favorable characteristics of our newly designed peptide make it a promising candidate ß-sheet breaker agent for further in vivo studies.

Preclinical study of a new 177Lu-labeled somatostatin receptor antagonist in HT-29 human colorectal cancer cells.

OBJECTIVES: Somatostatin receptor-positive neuroendocrine tumors have been targeted using various peptide analogs radiolabeled with therapeutic radionuclides for years. The better biomedical properties of radioantagonists as higher tumor uptake make these radioligands more attractive than agonists for somatostatin receptor-targeted radionuclide therapy. In this study, we tried to evaluate the efficiency of Luthetium-177 (177Lu) radiolabeled DOTA-Peptide 2 (177Lu-DOTA-Peptide 2) as a new radioantagonist in HT-29 human colorectal cancer in vitro and in vivo. METHODS: DOTA conjugated antagonistic peptide with the sequence of p-Cl-Phe-Cyclo(D-Cys-L-BzThi-D-Aph-Lys-Thr-Cys)-D-Tyr-NH2 (DOTA-Peptide 2) was labeled with 177Lu. In vitro assays (saturation binding assay and internalization test) and animal biodistribution were performed in human colon adenocarcinoma cells (HT-29) and HT-29 tumor-bearing nude mice. RESULTS: 177Lu-DOTA-Peptide 2 showed high stability in acetate buffer and human plasma (>97%). Antagonistic property of 177Lu-DOTA-Peptide 2 was confirmed by low internalization in HT-29 cells (<5%). The desired dissociation constant (Kd =11.14 nM) and effective tumor uptake (10.89 percentage of injected dose per gram of tumor) showed high binding affinity of 177Lu-DOTA-Peptide 2 to somatostatin receptors. CONCLUSION: 177Lu-DOTA-Peptide 2 demonstrated selective and high binding affinity to somatostatin receptors overexpressed on the surface of HT-29 cancer cells, which could make this radiopeptide suitable for somatostatin receptor-targeted radionuclide therapy.

Evaluation of a New 99mTc-labeled GnRH Analogue as a Possible Imaging Agent for Prostate Cancer Detection.

INTRODUCTION: Prostate cancer is a serious threat to men's health so it is necessary to develop technics for early detection of this malignancy. The purpose of this research was the evaluation of a new99mTc-labeled GnRH analogue as an imaging probe for tumor targeting of prostate cancer. METHODS: 99mTc-labeled-DLys6-GnRH analogue was prepared based on HYNIC as a chelating agent and tricine/ EDDA as coligands for labeling with 99mTc. HYNIC was coupled to epsilon amino group of DLys6 through aminobutyric acid (GABA) as a linker. Radiochemical purity and stability in normal saline and serum, were determined by TLC and HPLC methods. Furthermore, calculation of protein-binding and partition coefficient constant were carried out for 99mTc labeled peptide. The cellular experiments including receptor binding specificity and affinity were studied using three prostate cancer cell lines LN-CaP, DU-145 and PC-3. Finally, the animal assessment and SPECT imaging of radiolabeled GnRH analogue were evaluated on normal mice and nude mice bearing LN-CaP tumor. RESULTS: The GnRH conjugate was labeled with high radiochemical purity (~97%). The radiolabeled peptide showed efficient stability in the presence of normal saline and human serum. The in vitro cellular assays on three prostate cancer cell lines indicated that the radiotracer was bound to LN-CaP cells with higher affinity compared to DU-145 and PC-3 cells. The Kd values of 99mTc- HYNIC (tricine/ EDDA)-Gaba-D-Lys6GnRH were 89.39±26.71, 93.57±30.49 and107.3±18.82 in LN-CaP, PC-3 and DU-145 cells respectively. The biodistribution studies in normal mice and LN-CaP tumor-bearing nude mice showed similar results including rapid blood clearance and low radioactivity accumulation in non-target organs. High kidney uptake proved that the main excretion route of radiopeptide was through the urinary system. The tumor uptake was 1.72±0.45 %ID/g at 1h p.i. decreasing to 0.70±0.06%ID/g at 4h p.i. for 99mTc-HYNIC-Gaba-D-Lys6GnRH. The maximum tumor/ muscle ratio was 2.30 at 1h p.i. Pre-saturation of receptor using an excess of unlabeled peptide revealed that the tumor uptake was receptor mediated. The results of the SPECT image of LN-CaP tumor were in agreement with the biodistribution data. CONCLUSION: Based on this study, we suggest LN-CaP as a favorable cell line for in vivo studies on GnRH analogues. Moreover, this report shows that 99mTc-HYNIC (tricine/EDDA)-Gaba-D-Lys6GnRH may be a suitable candidate for further evaluation of prostate cancer.

Synthesis, biological evaluation and preclinical study of a novel 99mTc-peptide: A targeting probe of amyloid-ß plaques as a possible diagnostic agent for Alzheimer's disease.

With respect to the main role of amyloid-ß (Aß) plaques as one of the pathological hallmarks in the brain of Alzheimer's patients, the development of new imaging probes for targeted detection of Aß plaques has attracted considerable interests. In this study, a novel cyclopentadienyl tricarbonyl Technetium-99 m (99mTc) agent with peptide scaffold, 99mTc-Cp-GABA-D-(FPLIAIMA)-NH2, for binding to the Aß plaques was designed and successfully synthesized using the Fmoc solid-phase peptide synthesis method. This radiopeptide revealed a good affinity for Aß42 aggregations (Kd = 20 µM) in binding affinity study and this result was confirmed by binding to Aß plaques in brain sections of human Alzheimer's disease (AD) and rat models using in vitro autoradiography, fluorescent staining, and planar scintigraphy. Biodistribution studies of radiopeptide in AD and normal rats demonstrated a moderate initial brain uptake about 0.38 and 0.35% (ID/g) 2 min post-injection, respectively. Whereas, AD rats showed a notable retention time in the brain (0.23% ID/g at 30 min) in comparison with fast clearance in normal rat brains. Normal rats following treatment with cyclosporine A as a p-glycoprotein inhibitor showed a significant increase in the radiopeptide brain accumulation compared to non-treated ones. There was a good correlation between data gathered from single-photon emission computed tomography/computed tomography (SPECT/CT) imaging and biodistribution studies. Therefore, these findings showed that this novel radiopeptide could be a potential SPECT imaging agent for early detection of Aß plaques in the brain of patients with AD.

99m Tc-(EDDA/tricine)-HYNIC-GnRH analogue as a potential imaging probe for diagnosis of prostate cancer.

Prostate cancer is a serious threat to men's health, so it is necessary to develop the techniques for early detection of this malignancy. Radiolabeled peptides are the useful tools for diagnosis of prostate cancer. In this research, we designed a new HYNIC-conjugated GnRH analogue and labeled it by 99m Tc with tricine/EDDA as coligands. We used aminohexanoic acid (Ahx) as a hydrocarbon linker to generate 99m Tc-(tricine/EDDA)-HYNIC-Ahx-[DLys6 ]GnRH. The radiopeptide exhibited high radiochemical purity and stability in solution and serum. Two human prostate cancer cell lines LN-CaP and DU-145 were used for cellular experiments. The binding specificity and affinity of radiopeptide for LN-CaP were superior to DU-145 cells. The Kd values for LN-CaP and DU-145 cells were 41.91 ± 7.03 nM and 55.96 ± 10.56 nM, respectively. High kidney uptake proved that the main excretion route of radiopeptide was through the urinary system. The tumor/muscle ratio of 99m Tc-HYNIC-Ahx-[DLys6 ]GnRH was 4.14 at 1 hr p.i. that decreased to 2.41 at 4 hr p.i. in LN-CaP tumor-xenografted nude mice. The blocking experiment revealed that the tumor uptake was receptor-mediated. The lesion was visualized clearly using 99m Tc-[DLys6 ]GnRH at 1 hr p.i. Accordingly, this research highlights the capability of 99m Tc-(tricine/EDDA)-HYNIC-Ahx-[DLys6 ]GnRH peptide as a promising agent for GnRHR-expressing tumor imaging.

Design, preparation and biological evaluation of a 177Lu-labeled somatostatin receptor antagonist for targeted therapy of neuroendocrine tumors.

Somatostatin receptor-targeted radionuclide therapy has become an effective treatment in patients with neuroendocrine tumors. Recently, investigations on the development of antagonistic peptides are increasing with possible superior biological properties as opposed to the agonists. Herein, we have reported the development of a new somatostatin receptor peptide ligand labeled with 177Lu to achieve a therapeutic ligand for tumor treatment. The interactions of selected and drown ligands using Avogadro software were docked on somatostatin receptor by Dink algorithm. The best docked peptide-chelator conjugate (DOTA-p-Cl-Phe-Cyclo(d-Cys-l-BzThi-d-Aph-Lys-Thr-Cys)-d-Tyr-NH2) (DOTA-Peptide 2) was synthesized using the Fmoc solid-phase method. DOTA-Peptide 2 was radiolabeled with the 177Lu Trichloride (177LuCl3) solution at 95 °C for 30 min and radiochemical purity (RCP) of 177Lu-DOTA-Peptide 2 solution was monitored by radio-HPLC and radio-TLC procedures. The new radiolabeled peptide was evaluated for stability, receptor binding, internalization, biodistribution and single-photon emission computed tomography (SPECT) imaging using C6 glioma cells and C6 tumor-bearing rats. DOTA-Peptide 2 was obtained with 98% purity and efficiently labeled with 177Lu (RCP > 99%). 177Lu-DOTA-Peptide 2 showed a high value of stability in acetate buffer (91.4% at 312 h) and human plasma (>97% at 24 h). Radioconjugate exhibited low internalization (<5%) and high affinity for somatostatin receptors (Kd = 12.06 nM, Bmax = 0.20 pmol/106 cells) using saturation binding assay. Effective tumor uptake of 7.3% ID/g (percentage of injected dose per gram of tumor) at 4 h post-injection and fast clearance of radiopeptide from blood and other organs led to a high tumor-to-normal organ ratios. SPECT/CT imaging clearly showed the activity localization in tumor. The favorable antagonistic properties of 177Lu-DOTA-Peptide 2 on the somatostatin receptors can make it a suitable candidate for peptide receptor radionuclide therapy (PRRT). In the future study, the therapeutic application of this radiopeptide will be evaluated.

Recent advances in the design and applications of amyloid-ß peptide aggregation inhibitors for Alzheimer's disease therapy.

Alzheimer's disease (AD) is an irreversible neurological disorder that progresses gradually and can cause severe cognitive and behavioral impairments. This disease is currently considered a social and economic incurable issue due to its complicated and multifactorial characteristics. Despite decades of extensive research, we still lack definitive AD diagnostic and effective therapeutic tools. Consequently, one of the most challenging subjects in modern medicine is the need for the development of new strategies for the treatment of AD. A large body of evidence indicates that amyloid-ß (Aß) peptide fibrillation plays a key role in the onset and progression of AD. Recent studies have reported that amyloid hypothesis-based treatments can be developed as a new approach to overcome the limitations and challenges associated with conventional AD therapeutics. In this review, we will provide a comprehensive view of the challenges in AD therapy and pathophysiology. We also discuss currently known compounds that can inhibit amyloid-ß (Aß) aggregation and their potential role in advancing current AD treatments. We have specifically focused on Aß aggregation inhibitors including metal chelators, nanostructures, organic molecules, peptides (or peptide mimics), and antibodies. To date, these molecules have been the subject of numerous in vitro and in vivo assays as well as molecular dynamics simulations to explore their mechanism of action and the fundamental structural groups involved in Aß aggregation. Ultimately, the aim of these studies (and current review) is to achieve a rational design for effective therapeutic agents for AD treatment and diagnostics.

Evaluation of 99m Tc-MccJ25 peptide analog in mice bearing B16F10 melanoma tumor as a diagnostic radiotracer.

OBJECTIVES: Despite recent advances in treatment modalities, cancer remains a major source of morbidity and mortality throughout the world. Currently, the development of sensitive and specific molecular imaging probes for early diagnosis of cancer is still a problematic challenge. Previous studies have been shown that some of the antimicrobial peptides (AMPs) exhibit a broad spectrum of cytotoxic activity against cancerous cells in addition to their antimicrobial activities. MicrocinJ25 (MccJ25) is an antimicrobial peptide that is produced by Escherichia coli (E. coli) strain. The aim of this study was to investigate the potential of a new peptide radiopharmaceutical derived from MccJ25 for diagnosis of melanoma tumor bearing C57BL/6 mice. METHODS: A 14 amino acid analog of MccJ25 was labeled with technetium-99m (99mTc) through hydrazinonicotinamide (HYNIC) chelator and tricine as coligand. In vivo tumor uptake and tissue distribution were evaluated. The in vivo biodistribution studies were determined in C57BL/6 mice bearing B16F10 tumor. RESULTS: The amount of non-peptide related 99mTc-impurities that measured by thin layer chromatography (TLC) did not exceed 5% of the total radioactivity. The in vitro binding to B16F10 cells was 30.73 ± 0.9% after 1 h incubation at 37°C, and saturation binding experiments showed good affinity for radio-complex (Kd=47.98±6.25 nM). The melanoma tumor was clearly visible up 1 h post-injection by gamma camera imaging. CONCLUSION: The results showed that 99mTc-labeld peptide could be a promising candidate as a targeting radiopharmaceutical for melanoma tumor imaging in mice.

New 99m Tc(CO)3 -radiolabeled arylpiperazine pharmacophore as potent 5HT1A serotonin receptor radiotracer: Docking studies, chemical synthesis, radiolabeling, and biological evaluation.

In spite of previous efforts, there is lack of a radiotracer for imaging the 5HT1A receptor density in human brain, which is involved in several neurological brain disorders. The aim of this study was to prepare a new derivative of 1-(2-methoxyphenyl)piperazine (MPP) as a main chemical structure of 5HT1A receptor antagonist with 3-carbon linker and radiolabeled by [99m Tc][Tc(CO)3 (H2 O)3 ]+ precursor. Docking studies before chemical synthesis showed similar fashion of interaction for both WAY100635 (potent 5HT1A receptor antagonist) and new designed ligand, despite of addition of 99m Tc(CO)3 group in the structure of new ligand. MPP-(CH2 )3 -N3 was synthesized via three efficient and reliable chemical synthesis steps (more than 80% yield) then radiolabeled by addition of 2-ethynylpyridine and [99m Tc][Tc(CO)3 (H2 O)3 ]+ precursor in one pot procedure (more than 95% radiochemical efficiency) through click chemistry method. After incubation, radiotracer was found stable in vitro up to 2 hours. Binding assays showed about 33% specific binding of radiotracer to the 5HT1A receptors. Brain biodistribution studies indicated (0.26 ± 0.05)% ID/g hippocampus uptake at 30 minutes post injection, which its specificity was verified through blocking studies. These results suggested that new designed radioligand might serve as a potent SPECT imaging agent to estimate status of 5HT1A receptors.

Synthesis of a Peptide Derivative of MicrocinJ25 and Evaluation of Antibacterial and Biological Activities.

MicrocinJ25 (MccJ25) is a small ribosomally synthesized antimicrobial peptide that is produced by Enterobacteriacea family especially E. coli. The present study focuses on preparation and evaluation of in-vitro antimicrobial and biological properties of a new peptide derived from MccJ25. We prepared a MccJ25-derived peptide containing 14 amino acids and a single intra-molecular disulfide bond according to solid phase synthesis strategy. The purified peptide was characterized by Liquid chromatography-mass spectrometry (LC-MS) and Fourier Transform Infrared (FTIR) spectroscopy. 96-well microdilution plate assay was exerted for determination of minimum inhibitory concentration (MIC) of peptide against different bacterial strains. Cytotoxicity of the peptide derivative on HT-29 cell line assayed using MTT test. The final peptide successfully was prepared with purity more than 99.8% as determined by analytical HPLC. The evaluation of antibacterial activity of the peptide against Gram-positive and Gram- negative bacteria revealed that the peptide was very effective against E. coli 35218 with minimum inhibitory concentration (MIC) at dose 3.9 µM. The hemolytic activity toward human erythrocytes was very minimal below 0.3%. The cell viability percentage of HT-29 cell line after 24 h of contact with the peptide was more than 83%. The high sensitivity of E. coli strain to this new peptide derived from MccJ25 and through minimal toxicity to cancerous cell, suggesting that above synthesized peptide could be considered as a bioactive compound for further investigations.

The Value of Technetium-99m Labeled Alpha-Melanocyte-Stimulating Hormone (99mTc-α-MSH) in Diagnosis of Primary and Metastatic Lesions of Malignant Melanoma.

OBJECTIVES: Malignant melanoma is the most lethal type of skin cancers with unfavorable prognosis. Alpha-MSH peptide analogues have a high affinity for melanocortine-1 (MC1) receptors on melanocytes over expressing in malignant melanoma cells. Pre-clinical studies have shown promising results for radiolabeled MSH imaging in this malignancy. The purpose of this study is to assess the diagnostic value of 99mTc-α-MSH imaging in malignant melanoma. METHODS: Twenty-one patients (13 men) with pathologically confirmed malignant melanoma with or without metastatic distribution were included in this study. 740-1110 MBq 99mTc-α-MSH was injected and whole body scans were performed 20, 120 and 240 minutes post injection and were assessed both qualitatively and semi-quantitatively using target (T) to background (BG) ratio. RESULTS: The T/BG ratio for the primary tumor bed was 2.51±2.26, 2.56±2.48 and 1.92±1.79 minutes in the whole body scans 20, 120 and 240 minutes post injection, respectively. The sensitivity, specificity, negative and positive predictive values were 75%, 80%, 50% and 92% for primary lesion and 25%, 100%, 68% and 100% for distant metastasis, respectively. CONCLUSION: 99mTc-α-MSH is a newly introduced agent for diagnosis of tumoral lesions in malignant melanoma. Our study showed a high sensitivity with this modality in primary lesions as well as lymph node involvements. However the detection rate was not high in distant metastasis. The preliminary results are promising especially as a new complementary imaging method in management of malignant melanoma.

The sensitivity and specificity of 99mTc-IgG radiotracer to differentiate infection lesions induced by Staphylococcus aureus and sterile inflammation lesions induced by carrageenan assay in rat's foot.

BACKGROUND: Detection and identification of infection from sterile inflammation foci has a crucial role in diagnosis and therapy of patients in clinical practice. OBJECTIVE: To assess the efficiency of labeled human polyclonal immunoglobulin with technetium 99m in order to detect septic or aseptic lesions which were induced in a rat model. METHODS: The freeze-dried IgG kits have been reconstituted by 99mTc. The radio conjugate yield, radiochemical impurities and stability radio complex were performed by ITLC (Instant Thin Layer Chromatography) and Gel filtration assays. Twenty adult, male NMRI (Naval Medical Research Institute) rats were randomly divided into two groups equally. Infection was induced by Staphylococcus aureus and sterile inflammation created by Carrageenan test. All lesions were created in the rat's foot. Then radioisotope investigations were undertaken. RESULTS: Labeling yield was approximately 98%. The radio complex showed good stability in normal saline. All affected feet could be easily visualized by imaging in qualitative study. The value of target to non-target ratio at the infection (n=10) and sterile inflammation (n=10) were 2.81±0.16 and 1.54±0.15 with p<0.007. Therefore, the radiotracer uptake at the septic lesions was significantly higher than the aseptic lesions. CONCLUSION: Imaging with 99mTc-IgG is highly sensitive to localized infection or inflammation foci. The increased accumulation of radiotracer at the infection versus inflammation foci may be helpful to interpret the image.

Preparation and evaluation of 188 Re sulfide colloidal nanoparticles loaded biodegradable poly (L-lactic acid) microspheres for radioembolization therapy.

Radioembolization with radioactive microspheres has been an effective method for the treatment of liver lesions. The aim of this study was to prepare carrier-free 188 Re loaded poly (L-lactic acid) (PLLA) microspheres through 188 Re sulfide colloidal nanoparticles (188 Re-SC nanoparticles). The formation of 188 Re-SC nanoparticles was confirmed by ultraviolet-visible spectrophotometry. The labeling yield of 188 Re-SC nanoparticles was verified using the RTLC method. Effects of synthesis parameters on morphology and size of prepared 188 Re-sulfide colloidal-PLLA microspheres (188 Re-SC-PLLA microspheres) were studied by scanning electron microscopy. In vitro stability of 188 Re-SC-PLLA microspheres was investigated in normal saline at room temperature and in human serum at 37°C. In vivo distribution studies and gamma camera imaging were performed in healthy BALB/c mice. The microspheres could be prepared with sizes between 13 and 48 µm (modal value 29 µm) and radiolabeling efficiency >99%. After incubation, the microspheres were found stable in vitro up to 72 hours. The biodistribution after intravenous injection in healthy BALB/c mice showed high accumulation in lung as a first capture pathway organ for microsphere followed by great retention over 48 hours for these microspheres. These data show that 188 Re-SC-PLLA microspheres are suitable candidate for clinical studies.

Development of a 99mTc-Labeled CXCR4 Antagonist Derivative as a New Tumor Radiotracer.

Due to high expression of CXCR4 (CXC chemokine receptor type 4) receptors in many tumors and metastasis, synthesis and labeling of CXCR4 receptor-targeted analogs as tumor imaging agents have been encouraged. Herein, CXCR4 receptor-targeted peptide antagonist was prepared and thereafter its labeling with 99mTc by a bifunctional chelating agent and tricine coligand was developed. Radiotracer purity, stability, and tumor cell binding were assessed. Bioevaluation of radiotracer was performed in mice bearing xenograft tumor. More than 95% labeling yield and stability up to 24 hours were observed. Radiotracer-related tumor accumulation was 3.61 ± 0.15% ID/g at 1 hour postinjection. High stability and specific tumor uptake are important characteristics of the radiotracer that could nominate this as a targeted imaging agent in the future.