Chemical proteomics reveals HSP70 1A as a target for the anticancer diterpene oridonin in Jurkat cells

J Proteomics. 2013 Apr 26:82:14-26. doi: 10.1016/j.jprot.2013.01.030. Epub 2013 Feb 15.

Abstract

Oridonin, an ent-kaurane diterpene isolated from well known Chinese medicinal plant Isodon rubescens, has been shown to have multiple biological activities. Among them, the anticancer activity has been repeatedly reported by many research groups. The chemopreventive and antitumor effects of oridonin have been related to its ability to interfere with several pathways which are involved in cell proliferation, cell cycle arrest, apoptosis and/or autophagy. Despite the number of studies performed on this diterpene, the molecular mechanism underlying its cellular activity remains to be elucidated. Hence, we tried to mine target protein(s) of oridonin by employing a mass spectrometry-based chemical proteomics approach, providing evidences that oridonin is able to directly bind the multifunctional, stress-inducible heat shock protein 70 1A (HSP70 1A). Oridonin/HSP70 complex formation was confirmed in leukemia-derived Jurkat cells. The characterization of HSP70 inhibition by oridonin was performed using chemical and biological approaches. Moreover, the binding site of oridonin on the chaperone was identified by a mass-based approach combined with Molecular Dynamics simulations.

Biological significance: Although natural products showed high efficiency and several of these agents have now entered in clinical trials, information concerning the mechanisms of action at a molecular level of many of them is very poor or completely missed. Nevertheless, the identification of the molecular target of a drug candidate has several advantages. The most significant is the ability to set up target-based assays and to allow structure-activity relationship studies to guide medicinal chemistry efforts towards lead optimization. The knowledge of drug targets can also facilitate the identification of potential toxicities or side effects, if there is any precedent of toxicities for the identified target. Achieving this in an effective, unbiased and efficient manner subsists as a significant challenge for the new era in drug discovery and optimization. In the present study, we used a chemical proteomic approach aimed to define the possible protein target of the ent-kaurane diterpene oridonin. This natural compound has drawn a rising attention for cancer biologists due to its remarkable anti-tumor activities: accumulating evidence has suggested that oridonin is able to hamper the progression of tumor, mitigate tumor burden and alleviate cancer syndrome, which may improve greatly the survival rates of cancer patients; however molecular mechanisms by which this compound exerts its anti-tumor activities still remained to be discovered. We identified the molecular chaperone HSP70 1A as an oridonin target in Jurkat cells, thus suggesting a mechanism of action for the diterpene consistent with the multiple biological activities described for it. HSP70 inhibition by oridonin might indeed simultaneously result in the impairment of some of client proteins, thus in turn affecting several molecular pathways. Shedding light on the molecular basis of the biological activity of oridonin, our findings may be relevant for possible therapeutic applications of oridonin, such as its use in combination and the design of new therapeutic approaches. In addition, this research demonstrates the effectiveness of chemical proteomic approaches in drug discovery studies and in orphan drug molecular target identification.

MeSH terms

  • Antineoplastic Agents / chemistry
  • Antineoplastic Agents / pharmacology*
  • Diterpenes, Kaurane / chemistry
  • Diterpenes, Kaurane / pharmacology*
  • HSP70 Heat-Shock Proteins / antagonists & inhibitors
  • HSP70 Heat-Shock Proteins / chemistry
  • HSP70 Heat-Shock Proteins / genetics
  • HSP70 Heat-Shock Proteins / metabolism*
  • Humans
  • Jurkat Cells
  • Leukemia / drug therapy*
  • Leukemia / metabolism
  • Molecular Dynamics Simulation
  • Neoplasm Proteins / antagonists & inhibitors
  • Neoplasm Proteins / chemistry
  • Neoplasm Proteins / genetics
  • Neoplasm Proteins / metabolism*
  • Proteomics*

Substances

  • Antineoplastic Agents
  • Diterpenes, Kaurane
  • HSP70 Heat-Shock Proteins
  • Hspa14 protein, human
  • Neoplasm Proteins
  • oridonin