Preparation, Characterization, and Radiolabeling of Anti-HER2 scFv With Technetium Tricarbonyl and Stability Studies.

Breast cancer is the most common diagnosed cancer, and the second cause of cancer death among women, worldwide. HER2 overexpression occurred in approximately 15% to 20% of breast cancers. Invasive biopsy method has been used for detection of HER2 overexpression. HER2-targeted imaging via an appropriate radionuclide is a promising method for sensitive and accurate identification of HER2+ primary and metastatic lesions. 99mTc-anti-HER2 scFv can specifically target malignancies and be used for diagnosis of the cancer type and metastasis as well as treatment of breast cancer. We radiolabeled anti-HER2 scFv that was expressed in Escherichia coli and purified through Ni-NTA resin under native condition with 99mTc-tricarbonyl formed from boranocarbonate. HER2-based ELISA, BCA, TLC, and HPLC were used in this study. In the current study, anti-HER2 scFv was lyophilized before radiolabeling. It was found that freeze-drying did not change the binding activity of anti-HER2 scFv to HER2. Results demonstrated direct anti-HER2 scFv radiolabeling by 99mTc-tricarbonyl to hexahistidine sequence (His-tag) without any changes in biological activity and radiochemical purity of around 98%. Stability analysis revealed that 99mTc-anti-HER2 scFv is stable for at least 24 h in PBS buffer, normal saline, human plasma proteins, and histidine solution.

Synthesis and Radiolabeling of Glu-Urea-Lys with 99mTc-Tricarbonyl-Imidazole-Bathophenanthroline Disulfonate Chelation System and Biological Evaluation as Prostate-Specific Membrane Antigen Inhibitor.

Background: The Glu-Urea-Lys (EUK) pharmacophore as prostate-specific membrane antigen (PSMA)-targeted ligand was synthesized, radiolabeled with 99mTc-tricarbonyl-imidazole-BPS chelation system, and biological activities were evaluated. The strategy [2 + 1] ligand is applied for tricarbonyl labeling. (5-imidazole-1-yl)pentanoic acid as a monodentate ligand and bathophenanthroline disulfonate (BPS) as a bidentate ligand formed a chelate system with 99mTc-tricarbonyl. EUK-pentanoic acid-imidazole and EUK were evaluated for PSMA active site using AutoDock 4 software. Materials and Methods: EUK-pentanoic acid-imidazole was synthesized in two steps. BPS was radiolabeled with 99mTc-tricarbonyl at 100°C for 30 min. The purified 99mTc(CO)3(H2O)BPS was used to radiolabel EUK-pentanoic acid-imidazole at 100°C, 30 min. Radiochemical purity, Log P, and stability studies were carried out within 24 h. Affinity of 99mTc(CO)3BPS-imidazole-EUK was performed in the saturation binding studies using LNCaP cells at 37°C for 1 h with a range of 0.001-1000 nM radiolabeled compound range. Internalization studies were performed in LNCaP cells with 1000 nM radiolabeled compound incubated for (0-2) h at 37°C. Biodistribution was studied in normal male Balb/c mice. The artificial intelligence predicts the uptake of radiolabeled compound in tumor. Results: The structures of synthesized compounds were confirmed by mass spectroscopy. Radiochemical purity, Log P, and protein binding were ≥95%, -0.2%, and 23%, respectively. The radiolabeled compound was stable in saline and human plasma within 24 h with radiochemical purity ≥90%. There was no release of 99mTc within 4 h in competition with histidine. The affinity was 82 ± 26.38 nM, and the activity increased inside the cells over time. Biodistribution studies showed radioactivity accumulation in kidneys less than 99mTc-HYNIC-PSMA. There was a moderate accumulation of radioactivity in the liver and intestine. Conclusion: Based on the results, 99mTc(CO)3BPS-imidazole-EUK can potentially be used as an imaging agent for studies at prostate bed and distal areas. The chelate system can be potentially labeled with rhenium for imaging studies (fluorescent or scintigraphy) and therapy.