Fluorescent Image–Guided Surgery with an Anti-Prostate Stem Cell Antigen (PSCA) Diabody Enables Targeted Resection of Mouse Prostate Cancer Xenografts in Real Time

Abstract

Purpose The inability to visualize cancer during prostatectomy contributes to positive margins, cancer recurrence, and surgical side effects. A molecularly targeted fluorescent probe offers the potential for real-time intra-operative imaging. The goal of this study was to develop a probe for image-guided prostate cancer surgery. Experimental Design An antibody fragment (cys-diabody, cDb) against prostate stem cell antigen (PSCA) was conjugated to a far-red fluorophore, Cy5. The integrity and binding of the probe to PSCA was confirmed by gel electrophoresis, size exclusion and flow cytometry, respectively. Subcutaneous models of PSCA-expressing xenografts were used to assess the biodistribution and in vivo kinetics, while an invasive intramuscular model was utilized to explore the performance of Cy5-cDb-mediated fluorescence guidance in representative surgical scenarios. Finally, a prospective, randomized study comparing surgical resection with and without fluorescent guidance was performed to determine if this probe could reduce the incidence of positive margins. Results Cy5-cDb demonstrated excellent purity, stability and specific binding to PSCA. In vivo imaging showed maximal signal-to-background ratios at 6 hours. In mice carrying PSCA+ and − dual xenografts, the mean fluorescence ratio of PSCA+/− tumors was 4.4:1. In surgical resection experiments, residual tumors <1mm that were missed on white light surgery were identified and resected using fluorescence guidance, which reduced the incidence of positive surgical margins (0/8) compared to white light surgery alone (7/7). Conclusions Fluorescently labeled cDb enables real-time in vivo imaging of prostate cancer xenografts in mice, and facilitates more complete tumor removal than conventional white light surgery alone.

Antibody Fragment and Affibody ImmunoPET Imaging Agents: Radiolabelling Strategies and Applications

“…Such dual‐modality optical‐PET imaging is advantageous over single modality imaging, as the much higher spatial resolution of optical imaging methods relative to PET can aid in more accurately localising the probe. Dual optical‐PET and optical fragment‐based probes may further help in the diagnosis and staging of cancer, as well as in providing greater delineation of healthy and cancerous tissue during surgery …”

“…Typically,t hey range in size from 7 to 100 kDa, depending on the specific type of fragment ( Figure 1). Antibody fragments have much shorter circulation times (hours rather than weeks), deeper penetration into tissue, are therefore better matched with the more clinically applieds horter-lived PET radionuclides (e.g., 18 F, 68 Ga) and can enable same-dayi maging. [9] This should, in theory,l ead to improvedi mages as blood clearancew ill be faster giving ab etter signal-to-noise ratio for the specifically bound radiolabelled fragment.…”

18F-Positron Emitting/Trimethine Cyanine-Fluorescent Contrast for Image-Guided Prostate Cancer Management

“…PSMA-targeted fluorophores have been proposed to improve intraoperative decision-making by using intraoperative fluorescence (FL) guidance and tumor margin delineation at high resolution. 13–15 …”

Surgical Guidance in Prostate Cancer: “From Molecule to Man” Translations

“…See related article by Sonn et al, p. 1403 In this issue of Clinical Cancer Research, Sonn and colleagues report on a prostate stem cell antigen (PSCA)-specific imaging agent for fluorescence-guided surgery (1). Herein, the authors rightfully reason that surgical guidance toward the location of prostate cancer may help to improve surgical outcome and to reduce procedureassociated side effects (2).…”

“…While shifting to a near-infrared dye may improve the latter, the Cy5-dye used by Sonn et al has a superior signal intensity. However, by having two dyes on a single diabody (1), this signal could be quenched to a certain extent. Of course, optimization of the optical properties should be supplemented by detailed evaluation in vitro (receptor specificity) and in vivo (pharmacokinetics).…”