## 4 Anti-Cancer Activities of Tanshinones
### Inhibition of Angiogenesis  
Angiogenesis is a process of new vessel formation from existing ones and plays a crucial role in tumor growth and metastasis. Endothelial cells can migrate toward the angiogenesis stimuli released from tumor cells and proliferate and differentiate to form a new vessel [212]. Neo-angiogenesis not only supplies the necessary nutrients and oxygen [213], but also allows tumor cells to escape into the circulation and metastasize to distant sites [214]. Recently studies suggest that tanshinones can suppress angiogenesis through inhibiting endothelial proliferation and angiogenic differentiation, in association with modulation of angiogenic regulators including but not limited to VEGF, HIF-1\(\upalpha\), c-Myc, matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs). In 2005, tanshinones were reported to inhibit the proliferation of fetal bovine serum (FBS)-induced proliferation of cultured rat vascular smooth muscle cells ( VSMCs) in a concentration-dependent manner, in association with inactivation of ERK1/2, increased P21 and down-regulated cyclin D\({}_{1}\) abundance [215]. Later, [PERSON] _et al._[216] used human umbilical vein endothelial cells (HUVECs) as a model and found that TIIA at 1, 5, 10 and 20 \(\upmu\)M concentration-dependently inhibited FBS-induced migration, TNF-\(\upalpha\)-induced invasion, and extracellular matrix (ECM)-induced tube formation. They also showed TIIA suppressed phorbol-12-myristate-13-acetate (PMA)-induced angiogenesis in chicken embryo chorioallantoic membrane (CAM) assay. The anti-angiogenesis activity of TIIA was associated with regulating MMP and TIMP secretion from vascular endothelial cells, _i.e._, pro-angiogenesis MMP-2 secretion was decreased but anti-angiogenesis TIMP-2 secretion was increased. This study also indicated that the anti-angiogenesis action of TIIA might be independent of the cytotoxic effect, because there was no sign of apoptosis or necrosis when HUVECs were treated with up to 20 \(\upmu\)M of TIIA [216]. The anti-angiogenesis activity of TIIA was also evaluated using cancer cell models. TIIA at low micromolar concentrations could inhibit the 2-dimension (2-D) and 3-dimension (3-D) migration of MDA-MB-435 cells (now a confirmed melanoma cell line), measured by wound-healing and transwell migration assays, respectively [217, 218]. TIIA also inhibited the tube formation of newborn cattle aortic endothelial cells (NCAECs) stimulated by co-incubation with MDA-MB-435 cells [217]. TIIA-induced inhibition of angiogenesis was accompanied by down-regulation of pro-angiogenesis factors such as VEGF, HIF-1\(\upalpha\) and c-Myc [217, 218]. Similarly, TIIA-induced inhibition of VEGF, HIF-1\(\upalpha\) and c-Myc was also reported in other cancer cells [219, 220, 221, 222, 223]. Moreover, an _in vivo_ study showed that TIIA (0.5, 1 and 2 mg/kg, _i.v._ injection) decreased serum VEGF levels and microvessel density (MVD) in C26 colon cancer xenografts in mice, in a dose-dependent manner [223]. In addition to TIIA, other tanshinones also possess potential anti-angiogenesis activities. [PERSON] _et al._[129] reported that TI suppressed HUVECs growth with IC\({}_{50}\) of \(\sim\)2.5 \(\upmu\)M and inhibited HUVECs migration and tube formation, in concentration-dependent manner. _In vivo_ administration of TI (150 and 200 mg/kg of TI in corn oil, oral gavage) decreased MVD in DU 145 prostate cancer and H1299 lung cancer xenograft tumors [129, 130]. [PERSON] _et al._[110] reported that, in a concentration range from 1-50 \(\upmu\)M, TI inhibited TNF-\(\upalpha\)-induced production of VEGF from HUVECs and MDA-MB-231 breast cancer cells, as well as VEGF-stimulated tube formation of HUVECs. CT was reported to suppress cell growth and bFGF-induced invasion and tube formation of bovine aortic endothelial cells (BAECs) in a concentration-dependent manner [224].