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Albany 2019: 20th Conversation - Abstracts

category image Albany 2019
Conversation 20
June 11-15 2019
Adenine Press (2019)

Structural studies on reactivation of tumor-related p53 mutants by methylene quinuclidinone (MQ), the active drug spontaneously formed from APR-246

The tumor suppressor p53 is also known as the “guardian of the genome” due to its ability to act as a transcription factor that regulates the expression of a range of target genes in response to genotoxic stress, leading to DNA repair, cell cycle arrest or apoptosis, all essential to prevent cancer (Vogelstein et al., 2000). The function of p53 can be compromised by mutations that lead to p53 inactivation and thus to cancer development. The majority of these mutations are located at the DNA binding core domain of p53 (p53DBD). Among them, six mutational “hotspots” at residues: R175, G245, R248, R249, R273 and R282 (Fig. 1), were shown to occur at high frequency in human cancer (Olivier et al., 2002). The corresponding amino-acid replacements (shown in red) can either affect p53 binding to DNA and referred as DNA-contact mutants: R273H/C, R248Q/W; or cause conformational changes in p53DBD and referred as structural mutants: R175H, G245S, R249S and R282W. Several crystal structures of the p53DBD of such mutants have been reported (Eldar et al., 2013; Suad et al., 2009).

PRIMA-1 and APR-246/PRIMA-1MET are small molecules that are converted into the biologically active compound, methylene quinuclidinone (MQ) which reactivates mutant p53 by binding covalently to cysteines in the p53DBD (Lambert et al., 2009). APR-246 is presently undergoing clinical trials (https://www.clinicaltrials.gov). In our study, we investigate the reactivation mechanisms of mutant p53 by MQ from a series of high-resolution crystal structures of wild-type p53 and several hotspot mutants bound to MQ in the absence and/or presence of DNA. Our data show that MQ can bind to several cysteine residues located at the surface of the core domain at positions: 124, 182, 229, 275, and 277. A detailed comparison between the structures of specific p53 mutants before and after binding to MQ, reveals the role played by MQ in stabilizing p53 and its interaction with DNA, thus providing a structural framework for the design of new molecules for specific targeting of cysteines in p53 mutants.

References

    Eldar, A., Rozenberg, H., Diskin-Posner, Y., Rohs, R., and Shakked, Z. (2013). Structural studies of p53 inactivation by DNA-contact mutations and its rescue by suppressor mutations via alternative protein-DNA interactions. Nucleic Acids Res 41, 8748-8759.

    Kitayner, M., Rozenberg, H., Kessler, N., Rabinovich, D., Shaulov, L., Haran, T.E., and Shakked, Z. (2006). Structural basis of DNA recognition by p53 tetramers. Mol Cell 22, 741-753.

    Lambert, J.M., Gorzov, P., Veprintsev, D.B., Soderqvist, M., Segerback, D., Bergman, J., Fersht, A.R., Hainaut, P., Wiman, K.G., and Bykov, V.J. (2009). PRIMA-1 reactivates mutant p53 by covalent binding to the core domain. Cancer cell 15, 376-388.

    Olivier, M., Eeles, R., Hollstein, M., Khan, M.A., Harris, C.C., and Hainaut, P. (2002). The IARC TP53 database: new online mutation analysis and recommendations to users. Hum Mutat 19, 607-614.

    Suad, O., Rozenberg, H., Brosh, R., Diskin-Posner, Y., Kessler, N., Shimon, L.J., Frolow, F., Liran, A., Rotter, V., and Shakked, Z. (2009). Structural basis of restoring sequence-specific DNA binding and transactivation to mutant p53 by suppressor mutations. J Mol Biol 385, 249-265.

    Vogelstein, B., Lane, D., and Levine, A.J. (2000). Surfing the p53 network. Nature 408, 307-310.
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Figure 1. Six hotspot mutation sites in p53DBD bound to DNA. p53 binds to its DNA target as a tetramer (dimer of dimers). Here is a view of a p53DBD dimer (grey) bound to DNA (blue), showing the six hotspot sites (red). The view is along the DNA helix. The figure is based on coordinates from PDB code 2ac0 (Kitayner et al., 2006).

Oksana Degtjarik 1
Dmitrij Golovenko 1
Yael Diskin-Posner 2
Lars Abrahmsén 3
Haim Rozenberg1
Zippora Shakked1

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Oksana is originally from Belarous, and is currently a post-doctoral fellow with Prof. Zippi Shakked at Weizmann Institute, and will provide a short oral from the platform.

1 Department of Structural Biology
Weizmann Institute of Science
76100 Rehovot, Israel

2 Department of Chemical Research Support
Weizmann Institute of Science
76100 Rehovot, Israel;

3 Aprea Therapeutics AB
17165 Solna, Sweden

Ph: (972)-8-934-3625
Email: oksana.degtjarik@weizmann.ac.il