Proceedings book download - 5Z.com

30 min at pH 7. Radiochemical yields (RCYs), as determined by TLC and HPLCchromatography, were around 90%.The second method is a one-step procedure: Na 99m TcO 4 (50.0 MBq-3 GBq) was addedto a vial containing SDH, SnCl 2 , ethanol, NAP-NS1/2 and PNPn. RCYs determined byTLC and HPLC, after 60 min at 80 °C, were around 90%.The [ 99m Tc(NNAP-NS1/2)(PNPn)] + complexes isolated by HPLC were concentratedon a Sep Pack C18 column rinsed with H 2 O and eluted using a mixture of EtOH/phosphatebuffer 0.2 M, pH 7.4, 80/20. The second fraction containing all the activity was utilized forin vitro and in vivo studies. After purification, radiochemical purities (RCPs) evaluated byTLC and HPLC were > 90% [4].The in vitro stability of the complexes was evaluated by transchelation experiments inpresence of cysteine and glutathione solutions, at 37°C for 24 hours. At fixed times aliquotsof the reaction mixture were withdrawn and the RCPs analysed by TLC and HPLC. Wealso assessed the in vitro stability monitoring the RCPs at different time points in presenceof human serum (HS), rat serum (RS) and in rat liver and kidney homogenates (RLH,RKH). Complexes evidenced a good stability in challenge experiments and in human andrat sera while a partial degradation was observed in homogenates. Biodistribution studieswere performed in healthy female S.D. rats with [ 99m Tc(N)(NAP-NS1/2)(PNPn)] + toinvestigate their organ uptake and excretion pathways. Complexes present an extremelyfast elimination from the blood and from significant organs; the renal excretion wasextremely rapid and the activity was manly eliminated through the urinary tract.NAP-NS1/2 were also tested for their affinity toward MC1R overexpressed on B16murine melanoma cells. Cells were incubated in Avidin-ITC and the fluorescence intensitymeasured at increasing peptide concentration. Jurkat Human lymphoma cells were used asa reference. Peptides evidenced a selective binding to B16 melanoma cells (Figure 2).160014001200B16 cellsJurkat cellsMFI value100080060040020000 10 20 30 40ug peptideFig. 2. Binding of NAP-NS1 toward B16 cells.Future studies will be focused on the evaluation in vitro and in vivo of the 99m Tc-complexesbinding affinities in B16 melanoma cell line.AcknowledgmentsSupported by the Italian MIUR grant PRIN 2008 (2008F5A3AF_002).References1. Bolzati, C., et al. Current Medicinal Chemistry 17, 2656-2683 (2010).2. Cheng, Z., et al. Bioconjugate Chem. 18, 765-772 (2007).3. Grieco, P., et al. J. Peptide Res. 57, 250-256 (2001).4. Agostini, S., et al. J. Pept. Sci. 13, 211-219 (2007).519