M.W. Roomi, T. Kalinovsky, N.W. Roomi, A. Niedzwiecki and M. Rath
Dr. Rath Research Institute, 1260 Memorex Drive, Santa Clara, CA 95050
International Journal of Oncology 2012; 41(6), 1996-2004

Abstract
Head and neck squamous cell carcinoma (HNSCC) and acute myeloid leukemia are the major causes of mortality and morbidity in Fanconi anemia (FA) patients. The objective of this study was to investigate the antineoplastic activity of NM, a novel antineoplastic nutrient mixture (containing lysine, proline, ascorbic acid and green tea extract) on human FA HNSCC in vitro and in vivo. Human FA HNSCC cell line OHSU-974 (Fanconi Anemia Research Fund) was cultured in RPMI medium supplemented with 20% FBS and antibiotics. At near confluence, cells were treated in triplicate with different concentrations of NM: 0, 50, 100, 250, 500 and 1000 µg/ml.

Cells were also treated with PMA to induce MMP-9 activity. Cell proliferation was detected by MTT assay, secretion of MMPs by gelatinase zymography, invasion through Matrigel, migration by scratch test and morphology by H&E staining. In vivo, athymic male nude mice (n=12) were inoculated with 3x106 OHSU-974 cells subcutaneously and randomly divided into two groups: group A was fed a regular diet and group B a regular diet supplemented with 1% NM. Four weeks later, the mice were sacrificed and their tumors were excised, weighed and processed for histology. NM inhibited the growth of OHSU-974 tumor by 47% and tumor burden by 50%. At lower concentrations, NM demonstrated no effect on proliferation, but at 1000 µg/ml showed 40% toxicity. Zymography revealed MMP-2 and PMA-induced MMP-9 secretion. NM suppressed secretion of both MMPs in a dose-dependent manner; with virtual inhibition at 500 µg/ml.  NM inhibited OHSU-974 invasion through Matrigel in a dose-dependent fashion with total block at 1000 µg/ml. H&E staining showed no morphological changes below 500 µg/ml. The results suggest that NM has potential therapeutic use in the treatment of human FA HNSCC.

Key words: 
Fanconi anemia, head and neck squamous carcinoma, nutrient mixture, tumor growth, MMPs, Matrigel invasion, cell migration

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http://www.spandidos-publications.com/10.3892/ijo.2012.1639