We routinely treat the cells with Nutlin-3 which prevents the negative regulator MDM2 to do its job, stabilizing/activating TP53.

Article Small-Molecule MDM2 Antagonists as a New Therapy Concept for...

Hi,

There is plenty of literature describing p53 stabilization in response to many different types of stress. The most "popular" techniques include (assuming that you are working with wt p53 cell line):

- treatment with doxorubicin (0.75 uM working concentration). p53 is significantly stabilized in 3h. Maximum of stabilization partially depends on the cell line used but I would say 8-10 h post Dox addition may be the right time.

- Cisplatin (10-20 ug/ml working concentration). Overall stabilization is less pronounced when compared to Dox, but still clearly there 10-12h post Cis addition.

- Nutlin-3a (10 uM working concentration). Works very fast, p53 is significantly stabilized in less than 3h in many cell lines. Overall levels of stabilized p53 (maximum) are very high. Keep in mind that Nutlin-3a is not a genotoxic drug and its effect on p53 stabilization is based on direct disrupting p53-Mdm2 circuit. Therefore, if you are looking at p53 specific covalent modifications, it may be tricky.

-NCS (neocarcinostatin, 300 ng/ml working concentration). This drug is kind of unique and not used very often but treatment with NCS results in rapid, though relatively minor (as compared to Dox or Nutlin) p53 stabilization: in 2-3 hours you can see p53 stabilized and after 5-6 h p53 levels are back to normal.

You can also use IR, H2O2 treatment and many other drugs/stresses as well.

In any case, I would recommend to do a time course for each drug at fixed concentration to find about the p53 stabilization curve.

Good luck

Thanks for your valuable suggestions .........surely it will help me thanks Jo and Oleg.

R.i.t.a.