Preparation and evaluation of chlorin e6-PSMA inhibitor constructs as multi-purpose and isostructural molecular imaging and therapy agents for prostate cancer

Objectives To develop and evaluate a PSMA targeted chlorin e6 (Ce6) derivative as an isostructural molecular imaging probe and targeted photosensitizer for photodynamic therapy (PDT).

Methods Ce6 was linked to a potent Lys-Urea-Glu based inhibitor of prostate specific membrane antigen (PSMA), which is a transmembrane protein overexpressed in malignant prostate cancer. The binding affinity of the conjugate with PSMA was determined through a LNCaP cell competition binding assay. PSMA-targeted Ce6 was subsequently incubated with both LNCaP cells (PSMA-positive) and PC3 cells (PSMA-negative), and flow cytometry and fluorescence microscopy were used to observe the differential fluorescence.

Results The IC₅₀ for the lead construct was determined to be 74 nM with respect to the potent PSMA inhibitor ¹²⁵I-TAAG-PSMA. Both fluorescence imaging and flow cytometry showed greater uptake of the PSMA-targeted Ce6 in LNCaP cells as compared to PC3 cells, with flow cytometry indicating a seven-fold greater fluorescence signal, in reference to non-targeted Ce6. As expected, Ce6 alone which is likely taken up into cells via LDL-specific endocytosis, had a fluorescence signal much greater than the PSMA-targeted Ce6 in LNCaP cells. The evidence collected indicated that the uptake of the conjugate is largely governed by the targeting vector.

Conclusions The work reported here demonstrates that Ce6 can be effectively linked to targeting vectors including inhibitors of PSMA. Studies in PSMA positive LNCaP cells showed a greater uptake of the agent compared to the negative controls. However, significantly higher fluorescence was observed for non-targeted Ce6. In addition to the potential utility for prostate cancer imaging and therapy, the reported constructs represent a versatile synthon that can be used to prepare isostructural molecular imaging probes and the associated PDT agents.

Research Support Canadian Institutes of Health Research, Canadian Cancer Society, Ontario Institute for Cancer Research, Natural Sciences and Engineering Research Council of Canada