M.W. Roomi, N. Roomi, V. Ivanov, T. Kalinovsky, A. Niedzwiecki, M. Rath
Oncology Reports 2005, 14(4): 807-815
Degradation of extracellular matrix (ECM) is a hallmark of tumor invasion, metastasis and angiogenesis. Based on Rath multitargeted approach to cancer by using natural substances to control ECM stability and enhancing its strength we developed a novel formulation (NM) of lysine, proline, ascorbic acid and green tea extract that has shown significant anti-cancer activity against a number of cancer cell lines. The aim of the present study was to determine whether NM exhibits anti-angiogenic and antimetastatic effects using in vitro and in vivo experimental models.
Angiogenesis was measured using chorioallantoic membrane (CAM) assay in chick embryos and bFGF-induced vessel growth in C57BL/6J female mice. To determine the in vivo effect of NM on tumor xenograft growth, male nude mice were inoculated with 3x106 MNNG-HOS cells. Control mice were fed a mouse chow diet, while the test group was fed a mouse chow diet supplemented with 0.5% NM for four weeks. In vitro studies on cell proliferation (MTT assay), MMP secretion (zymography) and Matrigel invasion were conducted on human osteosarcoma U2OS, maintained in McCoy medium, supplemented with 10% FBS, penicillin and streptomycin in 24-well tissue culture plates and tested with NM at 0, 10, 50, 100, 500, and 1000 µg/ml in triplicate at each dose.
NM at 250 µg/ml caused significant (P<0.05) reduction in bFGF-induced angiogenesis in CAM. NM inhibited tumor growth of osteosarcoma MNNG-HOS cell xenografts in nude mice by 53%; furthermore, tumors in NM-treated mice were less vascular and expressed lower levels of VEGF and MMP-9 immunohistochemically than did tumors in the control group. In addition, NM exhibited dose-dependent inhibition of osteosarcoma U2OS cell proliferation (up to 60% at 1000 µg/ml), MMP-2 and -9 expression (with virtual total inhibition at 500 µg/ml NM), and invasion through Matrigel (with total inhibition at 100 µg/ml NM). Moreover, NM decreased U2OS cell expression of VEGF, angiopoietin-2, bFGF, PDGF and TGF beta-1.
These results with our earlier findings suggest that NM is a relatively non-toxic formulation that inhibits growth, invasion, metastasis, and angiogenesis of tumor cells.