p53Lab, Agency for Science, Technology and Research, Singapore, Singapore.
PLoS One. 2013 Apr 30;8(4):e62564. doi: 10.1371/journal.pone.0062564. Print 2013.
HDM2 binds to the p53 tumour suppressor and targets it for proteosomal degradation. Presently in clinical trials, the small molecule Nutlin-3A competitively binds to HDM2 and abrogates its repressive function. Using a novel in vitro selection methodology, we simulated the emergence of resistance by evolving HDM2 mutants capable of binding p53 in the presence of Nutlin concentrations that inhibit the wild-type HDM2-p53 interaction. The in vitro phenotypes were recapitulated in ex vivo assays measuring both p53 transactivation function and the direct p53-HDM2 interaction in the presence of Nutlin. Mutations conferring drug resistance were not confined to the N-terminal p53/Nutlin-binding domain, and were additionally seen in the acidic, zinc finger and RING domains. Mechanistic insights gleaned from this broad spectrum of mutations will aid in future drug design and further our understanding of the complex p53-HDM2 interaction.
HDM2 与抑癌蛋白 p53 结合，并将其靶向到蛋白酶体降解。目前正在进行临床试验的小分子 Nutlin-3A 与 HDM2 竞争性结合，从而阻断其抑制功能。我们使用一种新的体外选择方法，通过进化能够在抑制野生型 HDM2-p53 相互作用的 Nutlin 浓度下结合 p53 的 HDM2 突变体，模拟耐药性的出现。在存在 Nutlin 的情况下测量 p53 反式激活功能和直接 p53-HDM2 相互作用的体外表型在 ex vivo 测定中得到了重现。赋予耐药性的突变不仅局限于 p53/Nutlin 结合域的 N 端，而且还存在于酸性、锌指和 RING 结构域中。从这种广谱突变中获得的机制见解将有助于未来的药物设计，并进一步加深我们对复杂的 p53-HDM2 相互作用的理解。
