Kit-like 18F radiolabeling of caspase activatable molecular probe for in situ noninvasive imaging of drug-induced apoptosis

Objectives: Positron emission tomography (PET) imaging for apoptosis is very important for the early evaluation of the therapeutic efficacy of tumors. A stimuli-response apoptosis-imaging probe based on the peptide sequence of Asp-Glu-Val-Asp (DEVD), ¹⁸F-AmBF₃-CBT-Cys(StBu)-DEVD ([¹⁸F]1), was designed and synthesized. This study aims to establish a rapid radiosynthesis method for the probe and fully examine its early response to tumor therapy.

Methods: The labeling precursor of AmBF₃-CBT-Cys(StBu)-DEVD (1) was obtained by multi-step reactions. After radiolabelled in the buffer of pH = 2.5, the radiolabelling yield, radiochemical purity and stability of the probe [¹⁸F]1 in PBS and serum were investigated by HPLC. The octanol/water partition coefficient (log P) of [¹⁸F]1 was determined at pH = 7.0. The detection of drug-induced cell death with [¹⁸F]1 in HeLa cells and tumor were fully evaluated, respectively. The level of activated caspase-3 in treated cancer cells was histologically analyzed.

Results: The radiosynthesis and purification process for the ¹⁸F-fluorination of 1 is around 25 min with the radiochemical yield of 50%, the specific radioactivity of 1.45 ± 0.4 Ci/µmol and the radiochemical purity over 99%. It is very stable in both PBS and mouse serum within 4 h. The retained radioactivity [¹⁸F] in doxorubicin (DOX)-treated HeLa cells was 2.2 times of that in untreated cells. The uptake ratio of tumor to muscle (T/M) increased from 1.74 to 2.18 in the untreated tumor with the injection of [¹⁸F]1, from 1.88 to 10.52 in treated tumor with the injection of only [¹⁸F]1, and from 3.08 to 14.81 in treated tumor with the co-injection of [¹⁸F]1 and 1.

Conclusions: An activatable PET tracer was designed and radiolabeled in a kit-like manner with a simple, rapid, safe and economic methodology. It showed significantly increased uptake in tumor as well as remarkably enhanced tumor-to-muscle uptake ratio in the mouse model of tumor chemotherapy, which correlated well with the level of drug-induced apoptosis. This early readout of treatment response ensured that it is a promising PET imaging agent for in situ noninvasive imaging of apoptosis. Financial supports: This work was financially supported by National Natural Science Foundation of China (21371082) and Natural Science Foundation of Jiangsu Province (BK20151118).