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In Vitro and In Vivo Inhibition of Human Fanconi Anemia Head and Neck Squamous Carcinoma by a Phytonutrient Combination

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 2015 DOI: 10.3892/ijo.2015.2895

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 PB, a nutrient mixture containing quercetin, curcumin, green tea, cruciferex, and resveratrol, 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 PB: 0, 10, 25, 50, 75 and 100 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% PB. 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 67.6% (p<0.0001) and tumor burden by 63.6% (p<0.0001). PB demonstrated dose-dependent inhibition of cell proliferation, with 27% (p=0.0003) and 48% (p=0.0004) toxicity at 75 and 100 µg/ml, respectively. Zymography revealed MMP-2 and PMA-induced MMP-9 secretion. PB suppressed secretion of both MMPs in a dose-dependent manner, with total block of both at  50 µg/ml.  PB inhibited cell migration (by scratch test) and OHSU-974 invasion through Matrigel in a dose-dependent fashion with total block at 50 µg/ml. H&E staining showed no morphological changes below 50 µg/ml. The results suggest that PB has potential therapeutic use in the treatment of human FA HNSCC.

Running Title: 
Phytonutrients inhibit OHSU-974 growth and invasion

Key Words: 
Fanconi anemia, head and neck squamous carcinoma, phytonutrients, tumor growth, MMPs, Matrigel invasion, cell migration