Emprego do peptídeo 99mTc-HYNIC-ßAla-Bombesina(7-14) na identificação de tumor de próstata da linhagem LNCaP em modelo experimental

Abstract

Prostate cancer is one of the most prevalent tumors in men, showing high mortality rates. Current diagnostic methods are not able to identify early prostate carcinoma, often resulting in a late diagnosis with established metastasis. Thus, there is by the scientific community an incessant search for diagnostic methods for early assessment of prostate cancer, facilitating the treatment and increasing the chances of cure. In this context, nuclear medicine provides a diagnostic method which can detect tumors at an early stage, because it is based on biochemical and physiological changes of the tissue, such as overexpression of gastrin releasing peptide receptors (GRPr`s) by prostate cancer cells. Bombesin, a tetradecapeptide isolated from the frog Bombina bombina, has a high affinity for the GRPr`s, since it is analogous to gastrin releasing peptide. Therefore, this study aims to prepare the complex 99mTc-HYNIC-Ala-Bombesin(7-14) and use it for the identification of prostate tumor, LNCaP line, in an experimental model. For in vitro assays, aliquots of 0.026 MBq of the radiopeptide were incubated with 2x106 LNCaP cells in a water bath at 37°C, for 1 and 4 hours, with and without prior addition of cold peptide (n=3). Prostate tumors were induced into the upper right flank of male BALB/c nude mice by subcutaneous injection of 5x106 LNCaP cells resuspended in 150 µL of Matrigel:RPMI-1640 medium (1:1). Biodistribution profile (n=5) and scintigraphic images (n=3) were obtained at 1 and 4 hours after intravenous injection of 7.4 MBq of 99mTc-HYNIC-Ala-Bombesin(7-14). To assess this, healthy male BALB/c mice and tumor-bearing male BALB/c nude mice with 15, 20 and 25 days of tumor development were used. In vitro study results showed that the fraction of the radiopeptide which bound to LNCaP cells was 2.08±0.30% (1 hour) and 2.44±0.18% (4 hours). From the percentage which was bound, the internalized fractions were 25.64±3.14% (1 hour) and 25.27±2.05% (4 hours). On the other hand, prior incubation of LNCaP cells with cold peptide reduced binding percentage in 32.21% (1 hour) and 43.85% (4 hours). Internalization percentage was reduced in 57.88% (1 hour) and 53.70% (4 hours). Biodistribution profile of 99mTc-HYNIC-Ala-Bombesin(7-14) in healthy and tumor-bearing mice showed rapid blood clearance, with high uptake by the kidneys. Percentage of injected dose (%ID) of the radiotracer captured by the tumor focus was significantly higher after 25 days of tumor development, approximately 1% (1 hour) and 0.1% (4 hours). Scintigraphic images have confirmed the radiopharmaceutical biodistribution profile, showing intense uptake of 99mTc-HYNIC-Ala-Bombesin(7-14) by kidneys and bladder, in all investigated groups. In contrast, the tumor focus was best viewed in tumor-bearing mice with 25 days of tumor development, with tumor size around 7.6 mm. In vitro assay results showed that the radiopeptide binds specifically to GRPr`s present in LNCaP cells surface. Biodistribution and scintigraphic images data suggest that 99mTc-HYNIC-Ala-Bombesin(7-14) is able to identify prostate tumor, LNCaP line, by scintigraphic images in experimental model.