BackgroundProstate-specific membrane antigen (PSMA) is frequently overexpressed and upregulated in prostate cancer. To date, various 18F- and 68Ga-labeled urea-based radiotracers for PET imaging of PSMA have been developed and entered clinical trials. Here, we describe an automated synthesis of [18F]DCFPyL via direct radiofluorination and validation in preclinical models of prostate cancer.Methods[18F]DCFPyL was synthesized via direct nucleophilic heteroaromatic substitution reaction in a single reactor TRACERlab FXFN automated synthesis unit. Radiopharmacological evaluation of [18F]DCFPyL involved internalization experiments, dynamic PET imaging in LNCaP (PSMA+) and PC3 (PSMA−) tumor-bearing BALB/c nude mice, biodistribution studies, and metabolic profiling. In addition, reversible two-tissue compartmental model analysis was used to quantify pharmacokinetics of [18F]DCFPyL in LNCaP and PC3 tumor models.ResultsAutomated radiosynthesis afforded radiotracer [18F]DCFPyL in decay-corrected radiochemical yields of 23 ± 5 % (n = 10) within 55 min, including HPLC purification. Dynamic PET analysis revealed rapid and high uptake of radioactivity (SUV5min 0.95) in LNCaP tumors which increased over time (SUV60min 1.1). Radioactivity uptake in LNCaP tumors was blocked in the presence of nonradioactive DCFPyL (SUV60min 0.22). The muscle as reference tissue showed rapid and continuous clearance over time (SUV60min 0.06). Fast blood clearance of radioactivity resulted in tumor-blood ratios of 1.0 after 10 min and 8.3 after 60 min. PC3 tumors also showed continuous clearance of radioactivity over time (SUV60min 0.11). Kinetic analysis of PET data revealed the two-tissue compartmental model as best fit with K1 = 0.12, k2 = 0.18, k3 = 0.08, and k4 = 0.004 min−1, confirming molecular trapping of [18F]DCFPyL in PSMA+ LNCaP cells.Conclusions[18F]DCFPyL can be prepared for clinical applications simply and in good radiochemical yields via a direct radiofluorination synthesis route in a single reactor automated synthesis unit. Radiopharmacological evaluation of [18F]DCFPyL confirmed high PSMA-mediated tumor uptake combined with superior clearance parameters. Compartmental model analysis points to a two-step molecular trapping mechanism based on PSMA binding and subsequent internalization leading to retention of radioactivity in PSMA+ LNCaP tumors.Electronic supplementary materialThe online version of this article (doi:10.1186/s13550-016-0195-6) contains supplementary material, which is available to authorized users.

Antifreeze glycoproteins (AFGPs) are a novel class of biologically significant compounds that possess the ability to inhibit the growth of ice both in vitro and in vivo. Any organic compound that possesses the ability to inhibit the growth of ice has many potential medical, industrial, and commercial applications. In an effort to elucidate the molecular mechanism of action, various spectroscopic and physical techniques have been used to investigate the solution conformations of these glycoproteins. This review examines the characterization of AFGPs and potential biological applications relating to stabilization of lipid membranes and vitrification adjuvants.

Elevated growth in breast cancer (BC) activates hypoxia-inducible factor (HIF1α) and downstream, facilitative glucose transporter 1 (GLUT1), which can be visualized with 2-deoxy-2-[F]fluoro-d-glucose ([F]FDG). GLUT5 (fructose) and GLUT2 (glucose/fructose) might provide alternative targets for BC imaging as to why effects of hypoxia on GLUT1/2/5 levels and function were examined in human BC models. GLUT1/2/5 and HIF1α mRNA was analyzed in BC patient biopsies. In MCF10A, MCF7, and MDA-MB231 cells, [F]FDG, 6-deoxy-6-[F]fluoro-d-fructose (6-[F]FDF) and [F]-fluoroazomycin arabinoside were used in radiotracer experiments, whereas GLUT1/2/5 mRNA was analyzed with real-time PCR and protein levels determined via Western blot/immunohistochemistry. Positron emission tomography imaging was performed in MCF7 and MDA-MB231 tumor-bearing mice. Glucose/fructose/cytochalasin B reduced cellular 6-[F]FDF uptake by 50%, indicating functional involvement of GLUT2. With GLUT5 staining lower than GLUT1, 6-[F]FDF revealed lower uptake than [F]FDG [standardized uptake value (SUV) 0.77 ± 0.06 vs. SUV 1.08 ± 0.07] in MDA-MB231 tumors and was blocked by 20% with cytochalasin B after 10 min. Whereas correspondence between 6-[F]FDF uptake and GLUT5 protein was low, high GLUT2 levels were detected in all cell lines and tumor models. Besides GLUT1, GLUT5 seems to be regulated under hypoxia on the molecular and functional level. Additionally, results strongly support a functional involvement of GLUT2 in fructose metabolism, possibly by compensating for the weaker expression and function of GLUT5 in BC.-Hamann, I., Krys, D., Glubrecht, D., Bouvet, V., Marshall, A., Vos, L., Mackey, J. R., Wuest, M., Wuest, F. Expression and function of hexose transporters GLUT1, GLUT2, and GLUT5 in breast cancer-effects of hypoxia.

The copper-free strain-promoted click chemistry between (18)F-labeled aza-dibenzocyclooctyne [(18)F]FB-DBCO and various azides is described. [(18)F]FB-DBCO was prepared in 85% isolated radiochemical yield (decay-corrected) through acylation of amino aza-dibenzocyclooctyne 1 with N-succinimidyl 4-[(18)F]fluorobenzoate ([(18)F]SFB). [(18)F]FB-DBCO showed promising radiopharmacological profil with fast blood clearance as assessed with dynamic small animal PET studies. Metabolic stability of [(18)F]FB-DBCO was 60% of intact compound after 60 min post injection in normal Balb/C mice and blood clearance half-life was determined to be 53 s based on the time-activity-curve (TAC). Copper-free click chemistry was performed with various azides at low concentrations (1-2 μM) which differed in their structural complexity in different solvents (methanol, water, phosphate buffer and in bovine serum albumin (BSA) solution). Reaction proceeded best in methanol (>95% yield after 15 min at room temperature), whereas reaction in BSA required longer reaction times of 60 min and 40 °C upon completion.

Antifreeze glycoproteins (AFGPs) have many potential applications ranging from the cryopreservation and hypothermic storage of tissues and organs to the preservation of various frozen food products. Since supplying native AFGP for these applications is a labor-intensive and costly process, the rational design and synthesis of functional AFGP analogues is a very attractive alternative. While structure-function studies have implicated specific structural motifs as essential for antifreeze activity in AFGP, the relationship between solution conformation and antifreeze activity is poorly understood. Toward this end, we have analyzed AFGP8 in aqueous solutions using dynamic light scattering (DLS) and circular dichroism (CD). Our results indicate that AFGP8 forms discrete aggregates in solution. These aggregates are predominantly composed of dimers that form at solution concentrations greater than 20 mM. CD spectroscopy indicates that the preferred solution conformation of AFGP8 is consistent with that of random coil. However, significant beta-sheet and alpha-helix character is observed in more concentrated solutions, indicating that these glycopeptides are highly flexible in solution. Aggregation appears to have a minimal effect on the overall solution conformation. Thermal hysteresis (TH) activity of the aggregated solutions is much higher than that of less concentrated solutions that do not form aggregates. While cooperative functioning between lower and higher molecular weight AFGPs has been reported, this is the first instance where cooperative functioning in lower molecular weight AFGPs has been observed.