Abstract
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Objectives: Resection of lymph node metastases using image-guided surgery may have high clinical impact on the outcome of oligometastatic prostate cancer patients. Therefore, dual-labeled PSMA-inhibitors based on PSMA-11 have been developed. This combines preoperative staging by means of PET/CT, followed by fluorescence-guided surgery further improving the accuracy of detecting PSMA-positive tumor lesions by merging the strengths of both techniques.
Methods: The optical-dye conjugates PSMA-HBED-CC-FITC, -AlexaFluor488, -STARRED and -IRDye800CW were synthesized by conjugating the dyes to PSMA-11. Their PSMA-binding properties were analyzed in a competitive cell binding assay followed by internalization experiments in human PSMA expressing LNCaP cells. To further investigate internalization and intracellular distribution characteristics fluorescence was detected by microscopy. To determine specific tumor uptake and pharmacokinetic properties of the conjugates in vivo studies were performed in LNCaP tumor-bearing mice (BALB/c nu/nu) and healthy pigs.
Results: Comparative cell binding experiments revealed a high binding affinity to PSMA expressing cells for all conjugates. The radiolabeled fluorescent-dye conjugates showed PSMA-specific cell uptake and effective internalization into LNCaP cells, which has also been visualized by microscopy. First in vivo results indicated slightly varying pharmacokinetic properties depending on the fluorescent dye. The FITC- and AlexaFluor488-conjugates revealed a higher tumor uptake [10.86 ± 0.94 % ID/g (68Ga-PSMA-HBED-CC-FITC), 9.10 ± 6.70 % ID/g (68Ga-PSMA-HBED-CC-AlexaFluor488)] compared to 68Ga-PSMA-11 (4.83 ± 1.34 % ID/g), while a similar uptake was detected for the 68Ga-STARRED- (5.60 ± 2.30 %ID/g) and 68Ga-IRDye800CW-conjugates (3.80 ± 1.00 % ID/g). A specific fluorescent signal in PSMA-expressing tissue could be detected in vivo for the clinical relevant candidate PSMA-HBED-CC-IRDye800CW.
Conclusion: Conjugation of a fluorescent dye to the well-established imaging agent PSMA-11 showed rather minor dye-dependent impact on cell binding properties, tumor uptake and the pharmacokinetic characteristics. As demonstrated in these preclinical results and in vivo studies, dual-labeled PSMA-inhibitors also show a PSMA-specific fluorescence signal in a surgical setting. This emphasizes the potential of dual-labeled PSMA-inhibitors to provide sensitive pre-, intra- and post-therapeutic detection of cancer lesions, thereby improving the treatment of prostate cancer. Research Support: We thank PD Dr. med. Arianeb Mehrabi (University Hospital Heidelberg) and his team for kindly supporting the animal studies.