Angiogenesis is a key step for tumour growth and metastasis, and anti-angiogenesis has been proposed as an important strategy for cancer therapy. Tryptanthrin is a weakly basic alkaloid isolated from the dried roots of medicinal indigo plants and has been shown to possess anti-tumour activities on various cancer cell types. This study aims to investigate the in vitro and in vivo anti-angiogenic activities of tryptanthrin and to unravel its underlying molecular action mechanisms. Our results show that tryptanthrin inhibited the in vitro proliferation, migration, and tube formation of the human microvascular endothelial cells (HMEC-1) in a concentration-dependent manner and significantly suppressed angiogenesis in Matrigel plugs in mice. Mechanistic studies indicated that tryptanthrin reduced the expression of several pro-angiogenic factors (Ang-1, PDGFB and MMP2). Tryptanthrin was also found to suppress the VEGFR2-mediated ERK1/2 signalling pathway in HMEC-1 cells and molecular docking simulation indicated that tryptanthrin could bound to the ATP-binding site of VEGFR2. Collectively, the present study demonstrated that tryptanthrin exhibited both in vitro and in vivo anti-angiogenic activities by targeting the VEGFR2-mediated ERK1/2 signalling pathway and might have therapeutic potential for the treatment of angiogenesis-related diseases.
References
[1]
Folkman J (1971) Tumor angiogenesis: therapeutic implications. N Engl J Med 285: 1182–1186.
[2]
Folkman J (2002) Role of angiogenesis in tumor growth and metastasis. Semin Oncol 29: 15–18.
[3]
Shojaei F (2012) Anti-angiogenesis therapy in cancer: Current challenges and future perspectives. Cancer Lett 320: 130–137.
[4]
Eggert A, Ikegaki N, Kwiatkowski J, Zhao HQ, Brodeur GM, et al. (2000) High-level expression of angiogenic factors is associated with advanced tumor stage in human neuroblastomas. Clin Cancer Res 6: 1900–1908.
[5]
Karkkainen MJ, Petrova TV (2000) Vascular endothelial growth factor receptors in the regulation of angiogenesis and lymphangiogenesis. Oncogene 19: 5598–5605.
[6]
Olsson AK, Dimberg A, Kreuger J, Claesson-Welsh L (2006) VEGF receptor signalling - in control of vascular function. Nat Rev Mol Cell Biol 7: 359–371.
[7]
Holmes K, Roberts OL, Thomas AM, Cross MJ (2007) Vascular endothelial growth factor receptor-2: structure, function, intracellular signalling and therapeutic inhibition. Cell Signal 19: 2003–2012.
[8]
Koch S, Tugues S, Li X, Gualandi L, Claesson-Welsh L (2011) Signal transduction by vascular endothelial growth factor receptors. Biochem J 437: 169–183.
[9]
Zachary I, Gliki G (2001) Signaling transduction mechanisms mediating biological actions of the vascular endothelial growth factor family. Cardiovasc Res 49: 568–581.
[10]
Rousseau S, Houle F, Kotanides H, Witte L, Waltenberger J, et al. (2000) Vascular endothelial growth factor (VEGF)-driven actin-based motility is mediated by VEGFR2 and requires concerted activation of stress-activated protein kinase 2 (SAPK2/p38) and geldanamycin-sensitive phosphorylation of focal adhesion kinase. J Biol Chem 275: 10661–10672.
[11]
Thakker GD, Hajjar DP, Muller WA, Rosengart TK (1999) The role of phosphatidylinositol 3-kinase in vascular endothelial growth factor signaling. J Biol Chem 274: 10002–10007.
[12]
Pedram A, Razandi M, Levin ER (1998) Extracellular signal-regulated protein kinase/Jun kinase cross-talk underlies vascular endothelial cell growth factor-induced endothelial cell proliferation. J Biol Chem 273: 26722–22678.
[13]
Takahashi S (2011) Vascular Endothelial Growth Factor (VEGF), VEGF Receptors and Their Inhibitors for Antiangiogenic Tumor Therapy. Biol Pharm Bull 34: 1785–1788.
[14]
Hurwitz HI, Fehrenbacher L, Hainsworth JD, Heim W, Berlin J, et al. (2005) Bevacizumab in combination with fluorouracil and leucovorin: an active regimen for first-line metastatic colorectal cancer. J Clin Oncol 23: 3502–3508.
[15]
Hurwitz H, Fehrenbacher L, Novotny W, Cartwright T, Hainsworth J, et al. (2004) Bevacizumab plus irinotecan, fluorouracil, and leucovorin for metastatic colorectal cancer. N Engl J Med 350: 2335–2342.
[16]
Kankanala J, Latham AM, Johnson AP, Homer-Vanniasinkam S, Fishwick CWG, et al. (2012) A combinatorial in silico and cellular approach to identify a new class of compounds that target VEGFR2 receptor tyrosine kinase activity and angiogenesis. Br J Pharmacol 166: 737–748.
[17]
Tong Y, Zhang X, Tian F, Yi Y, Xu Q, et al. (2005) Philinopside A, a novel marine-derived compound possessing dual anti-angiogenic and anti-tumor effects. Int J Cancer 114: 843–853.
[18]
He MF, Huang YH, Wu LW, Ge W, Shaw PC, et al. (2010) Triptolide functions as a potent angiogenesis inhibitor. Int J Cancer 126: 266–278.
[19]
Lai L, Liu J, Zhai D, Lin Q, He L, et al. (2012) Plumbagin inhibits tumour angiogenesis and tumour growth through the Ras signalling pathway following activation of the VEGF receptor-2. Br J Pharmacol 165: 1084–1096.
[20]
Mu X, Shi W, Sun L, Li H, Jiang Z, et al. (2012) Pristimerin, a triterpenoid, inhibits tumor angiogenesis by targeting VEGFR2 activation. Molecules 17: 6854–6868.
[21]
Honda G, Tabata M, Tsuda M (1979) The antimicrobial specificity of tryptanthrin. Planta Med 37: 172–174.
[22]
Lin YK, Leu YL, Huang TH, Wu YH, Chung PJ, et al. (2009) Anti-inflammatory effects of the extract of indigo naturalis in human neutrophils. J Ethnopharmacol 125: 51–58.
[23]
Recio MC, Cerda-Nicolas M, Potterat O, Hamburger M, Rios JL (2006) Anti-inflammatory and antiallergic activity in vivo of lipophilic Isatis tinctoria extracts and tryptanthrin. Planta Medica 72: 539–546.
[24]
Kimoto T, Hino K, Koya-Miyata S, Yamamoto Y, Takeuchi M, et al. (2001) Cell differentiation and apoptosis of monocytic and promyelocytic leukemia cells (U-937 and HL-60) by tryptanthrin, an active ingredient of Polygonum tinctorium Lour. Pathol Int 51: 315–325.
[25]
Yu ST, Chen TM, Tseng SY, Chen YH (2007) Tryptanthrin inhibits MDR1 and reverses doxorubicin resistance in breast cancer cells. Biochem Biophys Res Commun 358: 79–84.
[26]
Chan HL, Yip HY, Mak NK, Leung KN (2009) Modulatory effects and action mechanisms of tryptanthrin on murine myeloid leukemia cells. Cell Mol Immunol 6: 335–342.
[27]
Liao XM, Leung KN (2013) Tryptanthrin induces growth inhibition and neuronal differentiation in the human neuroblastoma LA-N-1 cells. Chem Biol Interact 203: 512–521.
[28]
Chan JYW, Koon JCM, Liu XZ, Detmar M, Yu BA, et al. (2011) Polyphyllin D, a steroidal saponin from Paris polyphylla, inhibits endothelial cell functions in vitro and angiogenesis in zebrafish embryos in vivo. J Ethnopharmacol 137: 64–69.
[29]
Chan YK, Kwok HH, Chan LS, Leung KS, Shi J, et al. (2012) An indirubin derivative, E804, exhibits potent angiosuppressive activity. Biochem Pharmacol 83: 598–607.
[30]
Trott O, Olson AJ (2010) AutoDock Vina: improving the speed and accuracy of docking with a new scoring function, efficient optimization, and multithreading. J Comput Chem 31: 455–461.
[31]
Zhou X, Wang Y, Hu T, Or PM, Wong J, et al. (2013) Enzyme kinetic and molecular docking studies for the inhibitions of miltirone on major human cytochrome P450 isozymes. Phytomedicine 20: 367–374.
[32]
Hanahan D, Folkman J (1996) Patterns and emerging mechanisms of the angiogenic switch during tumorigenesis. Cell 86: 353–364.
[33]
Kaneko T, Nagata I, Miyamoto S, Kubo H, Kikuchi H, et al. (1992) Effects of nicardipine on tube formation of bovine vascular endothelial cells in vitro. Stroke 23: 1637–1642.
[34]
Lee K, Jeong KW, Lee Y, Song JY, Kim MS, et al. (2010) Pharmacophore modeling and virtual screening studies for new VEGFR-2 kinase inhibitors. Eur J Med Chem 45: 5420–5427.
[35]
Zhu X, Zhang X, Ma G, Yan J, Wang H, et al. (2011) Transport characteristics of tryptanthrin and its inhibitory effect on P-gp and MRP2 in Caco-2 cells. J Pharm Pharm Sci 14: 325–335.
[36]
Ades EW, Candal FJ, Swerlick RA, George VG, Summers S, et al. (1992) HMEC-1: establishment of an immortalized human microvascular endothelial cell line. J Invest Dermatol 99: 683–690.
[37]
Nanobashvili J, Jozkowicz A, Neumayer C, Fügl A, Sporn E, et al. (2003) Comparison of angiogenic potential of human microvascular endothelial cells and human umbilical vein endothelial cells. Eur Surg 35: 214–218.
[38]
Folkman J, D’Amore PA (1996) Blood vessel formation: what is its molecular basis? Cell 87: 1153–1155.
[39]
Cattaneo MG, Pola S, Deho V, Sanguini AM, Vicentini LM (2003) Alprostadil suppresses angiogenesis in vitro and in vivo in the murine Matrigel plug assay. Br J Pharmacol 138: 377–385.
[40]
Suri C, Jones PF, Patan S, Bartunkova S, Maisonpierre PC, et al. (1996) Requisite role of angiopoietin-1, a ligand for the TIE2 receptor, during embryonic angiogenesis. Cell 87: 1171–1180.
[41]
Maisonpierre PC, Suri C, Jones PF, Bartunkova S, Wiegand SJ, et al. (1997) Angiopoietin-2, a natural antagonist for Tie2 that disrupts in vivo angiogenesis. Science 277: 55–60.
[42]
Bar RS, Boes M, Booth BA, Dake BL, Henley S, et al. (1989) The effects of platelet-derived growth factor in cultured microvessel endothelial cells. Endocrinology 124: 1841–1848.
[43]
Risau W, Drexler H, Mironov V, Smits A, Siegbahn A, et al. (1992) Platelet-derived growth factor is angiogenic in vivo. Growth Factors 7: 261–266.
[44]
Carmeliet P (2000) Mechanisms of angiogenesis and arteriogenesis. Nat Med 6: 389–395.
[45]
Schrenk D, Riebniger D, Till M, Vetter S, Fiedler HP (1997) Tryptanthrins: A novel class of agonists of the aryl hydrocarbon receptor. Biochem Pharmacol 54: 165–171.
[46]
Paz K, Zhu Z (2005) Development of angiogenesis inhibitors to vascular endothelial growth factor receptor 2. Current status and future perspective. Front Biosci 10: 1415–1439.
[47]
Yue GG, Fan JT, Lee JK, Zeng GZ, Ho TW, et al. (2011) Cyclopeptide RA-V inhibits angiogenesis by down-regulating ERK1/2 phosphorylation in HUVEC and HMEC-1 endothelial cells. Br J Pharmacol 164: 1883–1898.
[48]
McTigue MA, Wickersham JA, Pinko C, Showalter RE, Parast CV, et al. (1999) Crystal structure of the kinase domain of human vascular endothelial growth factor receptor 2: a key enzyme in angiogenesis. Structure 7: 319–330.
[49]
Noble ME, Endicott JA, Johnson LN (2004) Protein kinase inhibitors: insights into drug design from structure. Science 303: 1800–1805.
