Hybrid Tracers Based on Cyanine Backbones Targeting Prostate-Specific Membrane Antigen: Tuning Pharmacokinetic Properties and Exploring Dye–Protein Interaction

Abstract

Prostate cancer surgery is currently being revolutionized by the use of prostate-specific membrane antigen (PSMA)-targeted radiotracers, for example, 99m Tc-labeled PSMA tracer analogs for radioguided surgery. The purpose of this study was to develop a second-generation 99m Tc-labeled PSMA-targeted tracer incorporating a fluorescent dye. Methods: Several PSMA-targeted hybrid tracers were synthesized: glutamic acid-urea-lysine (EuK)-Cy5-mas 3 , EuK-(SO 3)Cy5-mas 3 , EuK-Cy5(SO 3)-mas 3 , EuK-(Ar)Cy5-mas 3 , and EuK-Cy5(Ar)-mas 3 ; the Cy5 dye acts as a functional backbone between the EuK targeting vector and the 2-mercaptoacetyl-seryl-seryl-seryl (mas 3) chelate to study the dye's interaction with PSMA's amphipathic entrance funnel. The compounds were evaluated for their photophysical and chemical properties and PSMA affinity. After radiolabeling with 99m Tc, we performed in vivo SPECT imaging, biodistribution, and fluorescence imaging on BALB/c nude mice with orthotopically transplanted PC346C tumors. Results: The dye composition influenced the photophysical properties (brightness range 0.3-1.5 • 10 4 M −1 • cm −1), plasma protein interactions (range 85.0% ± 2.3%-90.7% ± 1.3% bound to serum, range 76% ± 0%-89% ± 6% stability in serum), PSMA affinity (half-maximal inhibitory concentration [IC 50 ] range 19.2 ± 5.8-175.3 ± 61.1 nM) and in vivo characteristics (tumorto-prostate and tumor-to-muscle ratios range 0.02 ± 0.00-154.73 ± 28.48 and 0.46 ± 0.28-5,157.50 ± 949.17, respectively; renal, splenic, and salivary retention). Even though all tracer analogs allowed tumor identification with SPECT and fluorescence imaging, 99m Tc-EuK-(SO 3)Cy5-mas 3 had the most promising properties (e.g., half-maximal inhibitory concentration, 19.2 ± 5.8, tumor-to-muscle ratio, 5,157.50 ± 949.17). Conclusion: Our findings demonstrate the intrinsic integration of a fluorophore in the pharmacophore in PSMA-targeted smallmolecule tracers. In this design, having 1 sulfonate on the indole moiety adjacent to EuK (99m Tc-EuK-(SO 3)Cy5-mas 3) yielded the most promising tracer candidate for imaging of PSMA.

A DROP-IN beta probe for robot-assisted 68Ga-PSMA radioguided surgery: first ex vivo technology evaluation using prostate cancer specimens

“…thus only a small urine layer must be considered (24,31)). Nonetheless, acknowledgement of this effect by radiochemists (32,33) and the reduced renal clearance of for example [ 18 F]F-PSMA tracers (34,35) may in the future help to overcome these issues. In addition, the in uence of renal clearance might also be overcome by using b-emitting isotopes that have a longer half-life, allowing the tumor resection to take place after all renal clearance of non-bound tracer is realized, e.g.…”

Robotic surgery using a DROP-IN beta probe – feasibility study with 68Ga-PSMA in prostate cancer specimens

“…In fact, when detecting beta radiation, only particles originating from few millimeters around the detector can give a signal, and thus only a very small urine layer must be considered (28)(29). Nonetheless, acknowledgement of this effect by radiochemists (30)(31) and the reduced renal clearance of for example 18 F-PSMA tracers (32-34) may in the future help to further minimize this potential limitation. In addition, the influence of renal clearance might also be overcome by using β-emitting isotopes that have a longer half-life, allowing the tumor resection to take place after all renal clearance of non-bound tracer is realized, e.g.…”

Cyanine Conjugate‐Based Biomedical Imaging Probes

“…Cyanine with a single reactive group can be directly coupled to a chelator for radiolabeling, as is the case for 111 In-DTPA-HQ4 (76); [113] while two reactive groups allow the introduction of an extra targeting warhead, e.g., 99m Tc-KuE-(SO 3 )Cy5mas 3 (77). [114] Besides, a subset of SPECT/NIRF dual-modal probes for targeted imaging were assembled based on a lysine scaffold: -NH 2 for cyanine fluorophores, -NH 2 for 111 In-DTPA and -COOH for various targeting peptides including cRGD (78), [115] antimicrobial peptide UBI 29−41 (79), [116] and CXCR4 targeting peptide Ac-TZ14011 (80). [117] Unlike most short-lived PET radionuclides, 111 In is suitable for the development of antibody-guided probes owing to its better compatibility with mAbs pharmacokinetics.…”