Antiangiogenic activity of Tripterygium wilfordii and its terpenoids

Abstract

Ethnopharmacological relevance

Tripterygium wilfordii Hook. f. (Celastraceae) has been traditionally used as folk medicine for centuries in China for the treatment of immune-inflammatory diseases.

Aim of the study

This study aimed to assess the antiangiogenic activities which support the therapeutic use of Tripterygium wilfordii and its terpenoids for angiogenesis disease such as cancer.

Materials and methods

The ethanol extract of Tripterygium wilfordii and subsequent fractions were evaluated on an in vivo antiangiogenic zebrafish embryo model.

Results

Three antiangiogenic terpenoids were isolated by bioassay-guided purification, namely, celastrol (4), cangoronine (5) and triptolide (7). Among them, triptolide manifested the most potent antiangiogenic activity against vessel formation by nearly 50% at 1.2 μM. Semi-quantitative RT-PCR analysis revealed that triptolide dose- and time-dependently reduced the mRNA expression of angiopoietin (angpt)2 and tie2 in zebrafish, indicating the involvement of angpt2/tie2 signaling pathway in the antiangiogenic action of triptolide.

Conclusions

The discovery of an alternative pathway further confirms the value of ethnopharmacological investigations into traditional botanicals for leads for potential drug development.

Introduction

Spouting of new blood vessels from pre-existing ones is called angiogenesis. Excessive angiogenesis is closely related to many human diseases, such as tumor growth and metastasis, retinopathy and inflammation (Folkman, 1995). Since tumor angiogenesis plays an essential role during tumor progression, antiangiogenesis has become a new promising anticancer therapeutic strategy. In 2004, Bevacizumab (Avastin), a humanized antibody against vascular endothelial growth factor (VEGF), was approved by United States Food and Drug Administration as the first angiogenesis-based anticancer agent for clinical use (Gupta and Zhang, 2005).

The success of antiangiogenic therapy for cancer treatment has led to an explosion in the research for antiangiogenic agents. Traditional herbal medicine has long been recognized as a potential source for discovering such agents. Indeed, many herbs and phytochemicals have been shown to have antiangiogenic activities both in vitro and in vivo, e.g. Ulmus davidiana Planchon var. japonica Nakai (Jung et al., 2007), Gastrodia elata Blume (Ahn et al., 2007), Sedum sarmentosum Bunge (Jung et al., 2008), Withania somnifera Dunal (Mathur et al., 2006), and resveratrol, curcumin, genistein, luteolin etc. (Deorukhkar et al., 2007).

Tripterygium wilfordii Hook. f. (Celastraceae), also called Leigongteng (Thunder God's Vine), is a woody wine distributed widely in the Orient (Ma et al., 1999). Its debarked root has long been used in traditional Chinese medicine to treat immune-inflammatory diseases such as rheumatoid arthritis, chronic nephritis, chronic hepatitis and lupus erythematosus (Jia, 1985, Tao et al., 1987). An ethyl acetate (EA) partition from ethanol extract of T. willfordii has been subjected to phase II double blind clinical trial (Mao et al., 1999). Patients with rheumatoid arthritis received with EA partition showed improvement in both clinical manifestations and laboratory findings (Ma et al., 2007). Over 200 chemical constituents are found in this plant (Brinker et al., 2007), which may account for its multiple uses in China in immune-inflammatory diseases, cancer, neurodegenerative diseases, and fertility regulation (Brinker et al., 2007).

Angiogenesis is typically occurred in rheumatoid arthritis, retinopathy and tumor growth, as they are collectively called “angiogenesis diseases” (Folkman, 1995). Thus, drugs used to treat rheumatoid arthritis and other angiogenesis diseases may have the potential to possess antitumor activities through angiogenic inhibition. Although several components of Tripterygium wilfordii were reported to have antiangiogenic activity (Ushiro et al., 1997, Huang et al., 2003, Fan et al., 2006), neither of them were found through bioassay-guided purification, the molecular mechanism remains unclear and the antiangiogenic activity of the extract of Tripterygium wilfordii has not yet been investigated. These prompted us to examine the antiangiogenic activities of Tripterygium wilfordii and further its bioactive components.

Zebrafish angiogenic model is a very promising in vivo model that has been successfully added to the model system for studying antiangiogenic agents (Staton et al., 2004, Taraboletti and Giavazzi, 2004, Norrby, 2006). The reason is a remarkable homology present in the molecular and signaling pathways that drive vessel development between zebrafish and mammals (Fouquet et al., 1997, Liang et al., 1998, Lyons et al., 1998). Hence drugs designed to interact with the functional domains of a zebrafish protein will likely elicit comparable pharmacological effects in human. Consistent with the notion, the effects of several antiagiogenic and proangiogenic agents found in zebrafish were also observed in mammalian systems (Parng et al., 2002, Cross et al., 2003, Seng et al., 2004). The use of zebrafish embryo to study antiangiogenic agents is increasingly reported (Belleri et al., 2005, Isenberg et al., 2007, Wang et al., 2007, Lam et al., 2008). Furthermore, angiogenic zebrafish model is suitable for high-throughput screening for antiangiogenic drugs with lower cost than mammalian models and offers greater relevance to humans as compared with in vitro cell lines and invertebrate models (Stern and Zon, 2003). More recently, a group of researchers has set up automated, quantitative screening system for antiangiogenic compounds using transgenic zebrafish and has successfully discovered three promising antiangiogenic candidates from compound library and proved the activities by mammalian system (Tran et al., 2007). These features and fact make zebrafish embryo an attractive model for identification of antiangiogenic agents.

This study made use of an in vivo model with zebrafish for the detection of antiangiogenesis by Tripterygium wilfordii and its terpenoids. The action mechanism of triptolide was further investigated.