Allele-Specific p53 Mutant Reactivation

The Cancer Institute of New Jersey, New Brunswick, NJ 08903, USA.
Cancer cell (Impact Factor: 23.52). 05/2012; 21(5):614-25. DOI: 10.1016/j.ccr.2012.03.042
Source: PubMed


Rescuing the function of mutant p53 protein is an attractive cancer therapeutic strategy. Using the National Cancer Institute's anticancer drug screen data, we identified two compounds from the thiosemicarbazone family that manifest increased growth inhibitory activity in mutant p53 cells, particularly for the p53(R175) mutant. Mechanistic studies reveal that NSC319726 restores WT structure and function to the p53(R175) mutant. This compound kills p53(R172H) knockin mice with extensive apoptosis and inhibits xenograft tumor growth in a 175-allele-specific mutant p53-dependent manner. This activity depends upon the zinc ion chelating properties of the compound as well as redox changes. These data identify NSC319726 as a p53(R175) mutant reactivator and as a lead compound for p53-targeted drug development.

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Available from: Alexei Vazquez, Jul 21, 2014
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    • "Among the various approaches, targeting the p53 for improved and efficient anticancer therapies is the restoration of wild-type p53 function in tumors that have lost p53 tumor suppressor activity [17]. Several small-molecule screening studies have led to the identification of compounds such as the PRIMA-1 [18], MIRA-1 [19], CP-31398 [20], STIMA-1 [21], SCH529074 [22], NSC319726 [23] and others [24] [25] with the ability to reactivate the mutant p53 protein and confer biological functions such as the activation of the target gene expression. A common property of many of these compounds is they possess chemically active, highly electrophilic double bonds that participate in reactions of nucleophilic addition and in a cellular milieu; as such, they are capable of inducing oxidative stress and redox imbalance in human cells (Fig. 1A). "
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    ABSTRACT: Small molecules that can restore biological function to the p53 mutants found in human cancers have been highly sought to increase the anticancer efficacy. In efforts to generate hybrid anticancer drugs that can impact two or more targets simultaneously, we designed and developed piperlongumine (PL) derivatives with an aryl group inserted at the C-7 position. This insertion bestowed a combretastatin A4 (CA4, an established microtubule disruptor) like structure while retaining the piperlongumine configuration. The new compounds exhibited potent antiproliferative activities against eight cancer cell lines, in particular, were more cytotoxic against the SKBR-3 breast cancer cells which harbor a R175H mutation in p53 suppressor. KSS-9, a representative aryl PL chosen for further studies induced abundant ROS generation and protein glutathionylation. KSS-9 strongly disrupted the tubulin polymerization in vitro, destabilized the microtubules in cells and induced a potent G2/M cell cycle block. More interestingly, KSS-9 showed the ability to reactivate the p53 mutation and restore biological activity to the R175H mutant protein present in SKBR3 cells. Several procedures, including immunocytochemistry using conformation-specific antibodies for p53, immunoprecipitation combined with western blotting, electrophoretic shift mobility shift assays showed a reciprocal loss of mutant protein and generation of wild-type like protein. p53 reactivation was accompanied by the induction of the target genes, MDM2, p21cip1 and PUMA. Mechanistically, the redox-perturbation in cancer cells by the hybrid drug appears to underlie the p53 reactivation process. This anticancer drug approach merits further development.
    European Journal of Medicinal Chemistry 11/2015; DOI:10.1016/j.ejmech.2015.10.052 · 3.45 Impact Factor
    • "Recently, two compounds, NSC319726 (Yu et al., 2012) and stictic acid (Wassman et al., 2013), were reported as mutant Figure 6. Binding of CTM to the Hsp40 Protein Is Required for the CTM-Mediated Reactivation of Mutant p53 R175H (A) Hsp40 expression is increased and its binding capacity to mutant p53 is enhanced upon CTM treatment in CAL-33 (R175H) cells. "
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    ABSTRACT: TP53 is the most frequently mutated gene in human cancer, and small-molecule reactivation of mutant p53 function represents an important anticancer strategy. A cell-based, high-throughput small-molecule screen identified chetomin (CTM) as a mutant p53 R175H reactivator. CTM enabled p53 to transactivate target genes, restored MDM2 negative regulation, and selectively inhibited the growth of cancer cells harboring mutant p53 R175H in vitro and in vivo. We found that CTM binds to Hsp40 and increases the binding capacity of Hsp40 to the p53 R175H mutant protein, causing a potential conformational change to a wild-type-like p53. Thus, CTM acts as a specific reactivator of the p53 R175H mutant form through Hsp40. These results provide new insights into the mechanism of reactivation of this specific p53 mutant. Copyright © 2015 Elsevier Ltd. All rights reserved.
    Chemistry & biology 08/2015; 22(9). DOI:10.1016/j.chembiol.2015.07.016 · 6.65 Impact Factor
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    • "If ZMC1 is a Zn 21 ionophore and the source of the Zn 21 it delivers is extracellular, as suggested by our kinetic experiments in Zn 21 -free media, then depleting the extracellular Zn 21 from complete media should inhibit ZMC1's function. To test this prediction, we took advantage of ZMC1's known ability to induce a conformational change in p53- R175H using the conformation specific antibodies PAB240 and PAB1620 in complete media with and without Zn 21 chelators (Fig. 5A) (Yu et al., 2012). Consistent with previous results, ZMC1 treatment shifted the p53-R175H immunophenotype from misfolded (PAB240) to WT-like (PAB1620) in TOV112D cells in untreated media (Yu et al., 2012). "
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    ABSTRACT: p53 is a Zn(2+)-dependent tumor suppressor inactivated in >50% of human cancers. The most common mutation, R175H, inactivates p53 by reducing its affinity for the essential zinc ion, leaving the mutant protein unable to bind the metal in the low [Zn(2+)]free environment of the cell. The exploratory cancer drug ZMC1 was previously demonstrated to reactivate this and other Zn(2+)-binding mutants by binding Zn(2+) and buffering it to a level such that Zn(2+) can repopulate the defective binding site, but how it accomplishes this in the context of living cells and organisms is unclear. Here, we demonstrate that ZMC1 increases intracellular [Zn(2+)]free by functioning as a zinc ionophore. ZMC1 binds Zn(2+) in the extracellular environment, diffuses across the plasma membrane as the neutral complex, and releases zinc into the cell once again as the Zn(2+) ion. It raises intracellular [Zn(2+)]free in cancer (TOV112D) and non-cancer (HEK293) cell lines to 15.8 and 18.1 nM, respectively, with half times of 2-3 min. These [Zn(2+)]free are predicted to result in ~90% saturation of p53-R175H, thus accounting for its observed reactivation. This mechanism is supported by the x-ray crystal structure of the [Zn(ZMC1)2] complex, which demonstrates structural and chemical features consistent with those of known metal ionophores. These findings provide a physical mechanism linking ZMC1's in vitro and in vivo activities, and define the remaining critical parameter necessary for developing synthetic metallochaperones for clinical use. The American Society for Pharmacology and Experimental Therapeutics.
    Molecular pharmacology 02/2015; 87(5). DOI:10.1124/mol.114.097550 · 4.13 Impact Factor
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