Evaluation of caspase inhibitor [18F]FET-VAD-FMK as an imaging agent for apoptosis

1348 Objectives Apoptosis is a form of programmed cell death and a common target pathway of anti-cancer drugs. The execution of apoptosis readily involves the induction of a family of cysteine proteases, known as caspases, whose intracellular signalling cascade leads to cell degradation. The ability to monitor apoptosis in vivo would greatly aid treatment stratification in cancer patients. Here we developed a 18F-labelled derivative of VAD-FMK [Val-Ala-DL-Asp(OMe)-fluoromethyl ketone], an irreversible pan-caspase inhibitor, and evaluated this tracer in models of apoptosis. Methods [18F]FET-VAD-FMK was synthesized through a copper mediated 1,3-cycloaddition reaction between [18F]fluoroethyl azide and the terminal alkyne group of the corresponding precursor. Radioligand uptake assays were performed with murine B-cell lymphoma cells, naive and treated with 100μM Cisplatin. Flow cytometry was used to assess the cellular state post-treatment. In vivo tracer characteristics were evaluated in an SD rat model of apoptosis at baseline and 3h post administration of protein synthesis inhibitor cycloheximide (2mg/kg). Results The tracer was obtained in good radiochemical yields (1-3 GBq) and high radiochemical purities (>95%). In vitro cell uptake of [18F]FET-VAD-FMK was low in naive cells and unchanged in cells 2h post-Cisplatin treatment. Tracer uptake was significantly increased by 4-fold at 12h and 8-fold at 24h post treatment with Cisplatin. Tracer uptake positively correlated with increased caspase-activation and loss of plasma membrane integrity at 24h as independently evaluated by flow cytometry. Rodent imaging revealed an increased liver SUV (4.5-fold) of activity following administration of cycloheximide, which was over and above the increase in plasma SUV as indicated by quantitative liver kinetics analysis (Ki at baseline = 0.027; Ki post cycloheximide = 0.05). Conclusions These promising results support further evaluation of [18F]FET-VAD-FMK in vivo.