A tri-modal molecular imaging agent for sentinel lymph node mapping

Abstract

IntroductionWe report an “instant kit” method to radiolabel fluorescent-tilmanocept with 68Ga and 99mTc for tri-modal molecular imaging of sentinel lymph nodes (SLNs). MethodsSolutions of sodium acetate, 68GaCl3 and Na99mTcO4 were added successively to a “kit vial” containing lyophilized 800CW-tilmanocept, SnCl2, trehalose and ascorbic acid. After a 30-min incubation, the pH was neutralized with PBS. No purification was required. Radiochemical and fluorescence purity was measured by HPLC and ITLC techniques. In vitro stability was measured by standing gel chromatography (SGC) and ITLC by a 100-fold dilution 0.25h after radiolabeling. In vivo stability was measured by SGC and ITLC after an 11h incubation in human plasma. A dose (0.1nmol, ~1MBq 68Ga, ~25MBq 99mTc) was injected to the footpad of 4 mice. Popliteal SLNs were imaged by PET and fluorescence imaging systems at 0.5, 24, 48, 72h, then excised and assayed for 99mTc. ResultsRadiochemical and fluorescent purity exceeded 98%. The in vitro stability assay demonstrated high irreversibility of both radiolabels and the fluorescent label, and in vivo stability assay demonstrated high stability of the technetium and fluorescent labels to plasma metabolism. Popliteal SLNs were identified by PET and fluorescence imaging within 0.5h of injection. SLN fluorescence intensity remained constant for 72h, when ~1% of the injected dose resided in the SLN. ConclusionsFluorescent-labeled tilmanocept can be radiolabeled with 68Ga and 99mTc by the sequential addition of each generator eluate to a lyophilized kit. The resulting tri-modal agent provides: PET images for pre-operative SLN mapping, fluorescence imaging up to 72hours after injection, and quantitative radiometric measurement of SLN accumulation after excision.