Carbon-11 Isotopic Radiolabeling of CP31398 and Development of a Fluorine-18 Derivative to Target Protein p53 with PET Imaging

TL;DR

Researchers developed two PET radiotracers based on CP31398 to target mutant p53 in cancer, but found they bind to an unknown target instead.

Contribution

The study introduces novel carbon-11 and fluorine-18 radiolabeled derivatives of CP31398 for potential p53 imaging.

Findings

Both [11C]CP31398 and [18F]FG-CP31398 were synthesized with acceptable radiochemical yields and molar activities.

Binding experiments showed [18F]FG-CP31398 binds to cells with some specificity but not directly to p53.

Autoradiography revealed unexpected binding to p53-negative cells, suggesting an alternative target.

Abstract

Mutant protein p53,
a central driver of pro-oncological deregulations,
is widely recognized as a biomarker of cancer aggressiveness and therapy
resistance. In a personalized medicine perspective, positron emission
tomography (PET) imaging of mutant p53 would be a powerful tool for
patient stratification and drug development. However, to date, no
PET radiotracers directly targeting mutant p53 have been reported.
Inspired by the CP31398 drug, which stabilizes p53 conformation and
treats tumors expressing mutant p53, we designed two novel PET radiotracers
labeled with either carbon-11 by isotopic labeling or fluorine-18,
namely [11C]CP31398 and [18F]FG-CP31398. The nonradioactive fluorinated analogue was synthesized,
as well as the two radiolabeling precursors. Optimization of the radiomethylation
with carbon-11 of the phenol precursor was achieved, and automated
radiosynthesis of [11C]CP31398 afforded the
ready-to-inject radiotracer with 40 ± 13% radiochemical yield
and 39 ± 12 GBq/μmol (n = 6) molar activity
after quality control. Automated radiofluorination with 18F-fluoride by aliphatic SN2 of a tosylate precursor afforded
the ready-to-inject [18F]FG-CP31398 in 10
± 4% radiochemical yield (RCY) and 80 ± 39 GBq/μmol
(n = 4) molar activity after quality control. Binding
experiments performed with [18F]FG-CP31398 on HEK-293T cells transfected with a green fluorescent protein-p53
wild-type plasmid, with or without presaturation with CP31398, demonstrated that despite the chemical modification performed on
this compound, [18F]FG-CP31398 was still able
to bind specifically to the cell with ca. 20% nonspecific
binding. However, autoradiography experiments performed after the
incubation of either [11C]CP31398 or [18F]FG-CP31398 on H358 (p53 positive) and A549
(p53 negative) tumor slices derived from human lung cancer cells revealed
that both tracers were not able to bind the p53-positive cells. Surprisingly,
a specific fixation demonstrated by presaturation experiments was
observed for both radiotracers on the p53-negative A549 cells. Overall,
our findings indicate that neither CP31398 nor FG-CP31398 binds directly to p53 but instead interacts with
an unidentified target. While unsuitable for p53 imaging, these radiotracers
may serve as valuable tools to unravel the controversial mechanism
of action of CP31398.

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