Fast Fluorine-18 labeling and preclinical evaluation of novel Mucin1 and its Folate hybrid peptide conjugate for targeting breast carcinoma

Abstract

Background There is a need to develop new and more potent radiofluorinated peptide and their hybrid conjugates for multiple-receptors targeting properties that overexpress on many cancers. Methods We have synthesized MUC1-[18F] SFB and MUC1-FA-[18F] SFB hybrid conjugates using a convenient and one-step nucleophilic displacement reaction. In vitro cell binding and in vivo evaluation in animals were performed to determine the potential of these radiolabeled compounds. Results Radiochemical yields for MUC1-[18F] SFB and MUC1-FA-[18F] SFB conjugates were greater than 70% in less than 30 min synthesis time. Radiochemical purities were greater than 97% without HPLC purification, which makes these approaches amenable to automation. In vitro studies on MCF7 breast cancer cells showed that the significant amounts of the radiofluorinated conjugates were associated with cell fractions and held good affinity and specificity for MCF7 cells. In vivo characterization in Balb/c mice revealed rapid blood clearance with excretion predominantly by urinary as well as hepatobiliary systems for MUC1-[18F] SFB and MUC1-FA-[18F] SFB, respectively. Biodistribution in SCID mice bearing MCF7 xenografts, demonstrated excellent tumor uptake (12% ID/g) and favorable kinetics for MUC1-FA-[18F] SFB over MUC1-[18F]SFB. The tumor uptake was blocked by the excess co-injection of cold peptides suggesting the receptor-mediated process. Conclusion Initial PET/CT imaging of SCID mice with MCF7 xenografts, confirmed these observations. These results demonstrate that MUC1-FA-[18F] SFB may be a useful PET imaging probe for breast cancer detection and monitoring tumor response to the treatment.

Synthesis, Radiolabeling, and Biological Evaluation of 68Ga-Mucin1 and Its Folate Hybrid Peptide Conjugates for the Diagnosis of Ovarian Cancer

“…e MUC1 peptide analog was prepared using the method reported previously [28,31], briefly by solid-phase peptide synthesis (on a CS Bio peptide synthesizer, CA, USA) following standard Fmoc (9-fluorenylmethoxycarbonyl) chemistry, using Rink amide methylbenzhydrylamine (MBHA) resin on a 0.2 mmol scale. After incorporating all the desired amino acids, the N-terminal Fmoc-protecting group was removed and the peptide was cleaved from the resin followed by the removal of the other side-chain protecting groups using a mixture of TFA/H 2 O/dithiothreitol (DTT) 95 : 2.5 : 2.5 for 2 h at room temperature.…”

“…After partial separation of the phases by gravity, 0.7 mL of each phase was transferred to separate tubes and centrifuged at 5000 rpm for 5 min. Duplicate 0.2 mL aliquots of each phase were taken for radioactivity measurement and the partition coefficient was determined by the function: partition coefficient � Log 10 (counts in n-octanol layer/ counts in the aqueous layer) [28,29].…”

“…For stability in plasma, the purified 68 Ga-NODAGA-MUC1 and 68 Ga-NODAGA-MUC1-FA hybrid peptide conjugates (100 µL, 20 µCi each) were incubated with human plasma (500 µL) in duplicate at 37°C for 2 h. is was followed by precipitation using a mixture of ACN/EtOH (400 µL, 1 : 1 v/v) and centrifugation at 5000 rpm for 5 min [28,29]. e supernatant layer was then analyzed by HPLC under the conditions described above.…”

“…24 h before conducting the cell-binding assay, media was replaced with RPMI-1640 without further addition of FBS. Confluent cultures were harvested by trypsinization, and 6 × 10 6 cells were suspended in 1.8 mL of sterile saline for binding assay [28,29]. Approximately 300,000 cells (in 0.3 mL of sterile saline) were incubated with various amounts of the purified 68 Ga-NODAGA-MUC1 and 68 Ga-NODAGA-MUC1-FA hybrid peptide conjugates ranging from 0.3 to 18 nM in duplicate for 60 min at room temperature.…”

“…In the previous studies, we have described the preclinical evaluation of 18 F-labeled Mucin1 as well as its folate conjugate for targeting breast cancer [28,29]. Based on the encouraging results of these studies, it seems interesting to further explore the potential of this MUC1 and its folate conjugate beyond breast cancer targeting.…”