Detection of tumor cell apoptosis by PET: Comparison of [¹⁸F]C-SNAT, [^99mTc]HYNIC-Annexin V and [¹⁸F]ML-10

Objectives It has been shown recently that [¹⁸F]C-SNAT can be used to detect early response to therapy in tumors by PET via a mechanism of caspase 3-triggered cyclization and nanoaggregation. Here we compared cell culture uptake of [¹⁸F]C-SNAT to clinically validated radiotracers, [^99mTc]HYNIC-Annexin V and [¹⁸F]ML-10, for the detection of drug-induced apoptosis.

Methods Murine lymphoma EL-4 cells were treated with 15 µM etoposide to induce apoptosis. 18h post either drug or vehicle treatment, temporal [¹⁸F]C-SNAT, [^99mTc]HYNIC-Annexin V and [¹⁸F]ML-10 cell retention were assessed in vitro. In addition, radiotracer uptake was measured in cell mixtures containing 0%, 25%, 50%, 75% and 100% etoposide-treated cells 1h after radiotracer incubation. Cell-associated radioactivity was compared to levels of cell death as determined by flow cytometric analysis of the same cell mixtures (n=3 independent experiments).

Results Etoposide treatment led to a 24-fold increase in [^99mTc]HYNIC-Annexin V cell-associated radioactivity, from 7±2 %ID/mg to 173±15 %ID/mg in vehicle versus etoposide-treated cells (P<0.001). For [¹⁸F]C-SNAT, cell uptake increased 8-fold with treatment (P<0.001). Cell-associated radioactivity was 2 orders of magnitude lower than [^99mTc]HYNIC-Annexin V (P<0.001), at 0.25±0.10 %ID/mg and 1.96±0.15 %ID/mg for vehicle and etoposide-treated cells, respectively. No significant change in [¹⁸F]ML-10 retention was measured in drug-treated cells (0.10±0.04 %ID/mg and 0.12±0.05 %ID/mg vehicle and etoposide-treated cells respectively). With cell mixtures, both [^99mTc]HYNIC-Annexin V and [¹⁸F]C-SNAT correlated with % apoptotic cells (R²=0.989 and 0.986, respectively), whereas there was no correlation between [¹⁸F]ML-10 uptake and % apoptosis (R²=0.029).

Conclusions In this systematic comparison study, [¹⁸F]C-SNAT and [^99mTc]HYNIC-Annexin V correlated with apoptosis, whereas no correlation was found for [¹⁸F]ML-10. In vivo evaluation of the three radiotracers in EL-4 xenografts ± therapy is currently in progress.