Faiella Laura, Piaz Fabrizio Dal, Bisio Angela, Tosco Alessandra, De Tommasi Nunziatina
Dipartimento di Scienze Farmaceutiche, Università di Pisa, via Bonanno 33, 56126 Pisa, Italy.
Mol Biosyst. 2012 Oct;8(10):2637-44. doi: 10.1039/c2mb25171j.
Clerodane diterpenoids are a class of naturally occurring molecules widely distributed in the Lamiaceae family. Neo-clerodane diterpenoids from Salvia ssp were recently described as compounds inhibiting the proliferation of human cancer cell lines. To gain new insights into molecular mechanism(s) underlying the antitumor potential of this class of compounds, we used a chemical proteomics approach to analyse the cellular interactome of hardwickiic acid (HAA) selected as a representative molecule. HAA was linked to an opportune 1,1'-carbonyldiimidazole modified by 1,12-dodecanediamine and then immobilized on a matrix support. The modified beads were then used as bait for fishing the potential partners of HAA in a U937 cell lysate. We identified heat shock protein 27 (Hsp27), an ATP-independent antiapoptotic chaperone characterized for its tumorigenic and metastatic properties and now referenced as a major therapeutic target in many types of cancer, as a major HAA partner. Here, we also report the study of HAA-Hsp27 interaction by means of a panel of chemical and biological approaches, including surface plasmon resonance measurements limited proteolysis, and biochemical assays. Our data suggest that HAA could provide a potential tool to develop strategies for the discovery of Hsp27 chemical inhibitors.
克罗烷二萜类化合物是一类广泛分布于唇形科的天然存在的分子。最近，来自鼠尾草属植物的新克罗烷二萜类化合物被描述为抑制人类癌细胞系增殖的化合物。为了深入了解这类化合物抗肿瘤潜力的分子机制，我们采用化学蛋白质组学方法分析了被选为代表性分子的硬叶酸（HAA）的细胞相互作用组。HAA与经1,12 - 十二烷二胺修饰的合适的1,1'-羰基二咪唑连接，然后固定在基质支持物上。然后将修饰后的珠子用作诱饵，在U937细胞裂解物中钓取HAA的潜在相互作用分子。我们鉴定出热休克蛋白27（Hsp27），一种不依赖ATP的抗凋亡伴侣蛋白，以其致瘤和转移特性而著称，目前被视为多种癌症的主要治疗靶点，是HAA的主要相互作用分子。在此，我们还通过一系列化学和生物学方法，包括表面等离子体共振测量、有限蛋白酶解和生化分析，报告了对HAA - Hsp27相互作用的研究。我们的数据表明，HAA可能为开发发现Hsp27化学抑制剂的策略提供一个潜在工具。
