The Lys-Specific Molecular Tweezer, CLR01, Modulates Aggregation of the Mutant p53 DNA Binding Domain and Inhibits Its Toxicity

Abstract

The tumor suppressor p53 plays a unique role as a central hub of numerous cell proliferation and apoptotic pathways and its malfunction due to mutations is a major cause of various malignancies. Therefore, it serves as an attractive target for developing novel anti-cancer therapeutics. Due to its intrinsically unstable DNA-binding domain, p53 unfolds rapidly at physiological temperature. Certain mutants shift the equilibrium towards the unfolded state and yield high-molecular-weight, non-functional, and cytotoxic β-sheet-rich aggregates that share tinctorial and conformational similarities with amyloid deposits found in various protein-misfolding diseases. Here, we examined the effect of a novel protein-assembly modulator, the lysine (Lys)-specific molecular tweezer, CLR01, on different aggregation stages of misfolded mutant p53 in vitro and on the cytotoxicity of the resulting p53 aggregates in cell culture. We found that CLR01 induced rapid formation of β-sheet-rich, intermediate-size p53 aggregates, yet inhibited further p53 aggregation and reduced cytotoxicity of the resulting aggregates. Our data suggest that aggregation modulators, such as CLR01, could prevent formation of toxic p53 aggregates.

Aggregation and Prion-Like Properties of Misfolded Tumor Suppressors: Is Cancer a Prion Disease?

“…Whereas CLR01 induced rapid formation of p53 aggregates of intermediate sizes, it inhibited additional p53 aggregation and reduced the cytotoxicity of the amyloid aggregates (Herzog et al 2015). To some extent, this behavior is similar to that found for the PrP protein, in which some compounds, such as polyanions, stimulate or inhibit aggregation depending on the condition (Gomes et al 2008;Silva et al 2008;Vieira et al 2014).…”

“…In a recent study, a protein assembly modulator (CLR01) showed an intriguing effect on p53 DNA-binding domain aggregation (Herzog et al 2015). Whereas CLR01 induced rapid formation of p53 aggregates of intermediate sizes, it inhibited additional p53 aggregation and reduced the cytotoxicity of the amyloid aggregates (Herzog et al 2015).…”

Molecular tweezers for lysine and arginine – powerful inhibitors of pathologic protein aggregation

“…In differentiated PC-12 cells, no toxicity was observed up to 200 μM and ~15% reduced viability (MTT assay) occurred at 400 μM. 36 No reduction in viability (XTT assay) was found up to 100 μM in human H1299 non-small cell lung carcinoma cells 82 and up to 500 μM in the HIV reporter cell line TZM-bl. 40 …”

Structural, morphological, and functional diversity of amyloid oligomers

“…Partially disordered Ab oligomers formed in the presence of polyphenols were non-toxic [31,136,137], whereas more structured Ab oligomers were toxic. Lysine-binding small molecules ('molecular tweezers') were shown to promote formation of non-toxic oligomers from a variety of amyloidogenic proteins and peptides [138][139][140][141][142] by modulating the oligomer structure. In case of Ab, they have been shown to compact the structures of dimers and tetramers and prevented formation of more toxic higher-order oligomers [143].…”