Synthesis of a Fluorescently Labeled ⁶⁸Ga-DOTA-TOC Analog for Somatostatin Receptor Targeting

Abstract

Fluorescently labeled imaging agents can identify surgical margins in real-time to help achieve complete resections and minimize the likelihood of local recurrence. However, photon attenuation limits fluorescence-based imaging to superficial lesions or lesions that are a few millimeters beneath the tissue surface. Contrast agents that are dual-labeled with a radionuclide and fluorescent dye can overcome this limitation and combine quantitative, whole-body nuclear imaging with intraoperative fluorescence imaging. Using a multimodality chelation (MMC) scaffold, IRDye 800CW was conjugated to the clinically used somatostatin analog, Ga-DOTA-TOC, to produce the dual-labeled analog,Ga-MMC(IRDye 800CW)-TOC, with high yield and specific activity. pharmacological assays demonstrated retention of receptor-targeting properties for the dual-labeled compound with robust internalization that was somatostatin receptor (SSTR) 2-mediated. Biodistribution studies in mice identified the kidneys as the primary excretion route forGa-MMC(IRDye 800CW)-TOC, along with clearance via the reticuloendothelial system. Higher uptake was observed in most tissues compared to Ga-DOTA-TOC but decreased as a function of time. The combination of excellent specificity for SSTR2-expressing cells and suitable biodistribution indicate potential application ofGa-MMC(IRDye 800CW)-TOC for intraoperative detection of SSTR2-expressing tumors.

Specific Targeting of Somatostatin Receptor Subtype-2 for Fluorescence-Guided Surgery

“…Radiolabeling was performed using a cation exchange cartridge as previously described (12). Following Sep-Pak Light C18 (Waters) purification, the product was diluted with PBS and analyzed by radio-HPLC.…”

“…Radioactive uptake studies were performed as previously described (12). Briefly, HCT116-SSTR2, NCI-H69, and non-SSTR2-expressing HCT116-WT cells (200,000 cells/well) were incubated with a 10 nmol/L solution of 68 Ga-MMC(IR800)-TOC, 68 Ga-DOTA-TOC, or a mixture of radiotracer with a 100-fold excess of octreotide at 37 C for 1 hour.…”

“…Fluorescence imaging of Ga-MMC(IR800)-TOC cellular uptake was performed as previously described (12). Briefly, HCT116-SSTR2 and HCT116-WT cells were plated (50,000 cells/well), allowed to attach for 48 hours, and incubated with 5 mmol/L Ga-MMC(IR800)-TOC for 1 hour at 37 C. NCI-H69 cells were identically processed with the exception of being in suspension.…”

“…Moreover, radiolabeled somatostatin analogs are currently used preoperatively for surgical planning and postoperatively for surveillance. In order to extend the utility of PET into the operating room, our group developed the first bioactive fluorescent analog of a clinical radiotracer using 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid-Tyr 3 -octreotide ( 68 Ga-DOTA-TOC) as a model (12). We introduced a synthesis scheme that enabled conversion of 68 Ga-DOTA-TOC into a dual-labeled counterpart, 68 Ga-MMC(IR800)-TOC, where MMC represents a multimodality chelator that replaces the standard chelating agent, DOTA (Fig.…”

New Developments in Dual-Labeled Molecular Imaging Agents

“…3B) that could be blocked in the presence of unlabeled octreotide, indicating receptor-mediated binding. The authors then synthesized a second-generation agent using an MMC analog with a modified pendant arm that contained a carboxyl group for stable chelation of 68 Ga (22). In vitro studies confirmed retention of SSTR2binding properties as shown by the ability of the agent to maximally inhibit cyclic adenosine monophosphate formation (EC 50 , 0.066 6 0.012 nM) and stimulate receptor internalization (EC 50 , 48.7 6 9.9 nM), as well as receptor-mediated uptake that was demonstrated by confocal microscopy and quantitative uptake assays.…”