M.W. Roomi, N.W. Roomi, M. Rath and A. Niedzwiecki
Dr. Rath Research Institute, Santa Clara, Ca
Presented at:
49th Annual Meeting of the Society of Toxicology, Salt Lake City, Utah, March 7-11, 2010.
Published in:
The Toxicologists, Abstract #2003, page 426
Abstract
Introduction:
Cancers of the breast, cervix and ovary are the most prevalent cancers in women worldwide. Proteases play a key role in tumor cell invasion and metastasis by digesting the basement membrane and ECM components. Two families of proteases: serine (u-PA) and MMPs (-2 and –9) are necessary for invasion and metastatic potential. Strong clinical and experimental evidence show that elevated levels of u-PA and MMPs are associated with tumor growth, cancer progression, metastasis and shortened patient survival. MMP activities are regulated by specific tissue inhibitors of metalloproteinases TIMPS).
Objective:
A nutrient mixture (NM) containing lysine, proline, ascorbic acid and green tea extract showed anticancer activity against a number of cancer cell lines. We investigated the effect of NM on activity of u-PA, MMPs and TIMPs in human breast, cervix, and ovarian cancer cell lines.
Material and Methods:
Human breast cancer (MDA-MB-231 and MCF-7), cervical cancer (Hela) and ovarian (SKOV3) cancer cell lines (ATCC) were cultured in MEM media with 10% FBS and antibiotics in 24-well tissue culture plates. At near confluence, the cells were treated with NM at 0, 50, 100, 250 , 500 and 1000 µg/ml in triplicate at each concentration. U-PA activity was carried out by fibrin zymography, MMPs by gelatinase zymography and TIMPs by reverse zymography.
Results:
U-PA activity was detected in both breast cancer cell lines, showing two bands corresponding to 55 and 33 kD. NM exerted dose response inhibition and significant reduction in u-PA activity at 250 µg/ml. However, no bands corresponding to u-PA were detected for Hela and SKOV3 cell lines. On gelatinase zymography, MDA-MB-231 and MCF-7 showed one band corresponding to MMP-9, Hela showed two bands, an intense band corresponding to MMP-2 and a faint band corresponding to MMP-9, and SKOV3 showed only a band corresponding to MMP-2. NM inhibited their expression in all cell lines at 100 µg/ml and blocked expression at 250 µg/ml. activity of TIMPs was up regulated in all cancer cell lines in a dose–dependent manner. Minimum activity was expressed at 50 and maximum at 1000 µg/ml. Analysis of correlation revealed a positive correlation between u-PA and MMPs and a negative correlation between u-PA /MMPs and TIMPs.
Conclusions:
These findings suggest that NM could potentially be developed as a new anticancer agent that inhibits u-PA and MMPs and increases TIMPs.
Comment
Cancers of the breast, cervix and ovary are the most prevalent cancers in women worldwide Strong clinical and experimental evidence show that elevated levels of u-PA and MMPs are associated with tumor growth, cancer progression, metastasis and shortened patient survival. TIMPs regulate MMPs. We found in vitro that a micronutrient mixture (NM) containing lysine, proline, ascorbic acid, green tea extract and other micronutrients inhibited u-PA and MMPs in human breast cancer (MDA-MB-231 and MCF-7 cell lines) in a dose-dependent response. Cervical cancer (Hela) and ovarian (SKOV3) cancer cell lines did not express u-PA but did express MMPs, which were inhibited in a dose-dependent response by NM. In addition, all cell lines tested exhibited increased TIMPs with NM treatment. These findings are significant as they suggest that NM, a non-toxic mixture of micronutrients, could potentially be developed as a new anticancer agent that inhibits u-PA and MMPs and increases TIMPs.
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SOT_upa_mmps_2010.pdf