The selective cyclooxygenase (COX)-2 inhibitor, celecoxib, and the vitamin E isoform, γ-tocotrienol, both display potent anticancer activity. However, high dose clinical use of selective COX-2 inhibitors has been limited by gastrointestinal and cardiovascular toxicity, whereas limited absorption and transport of γ-tocotrienol by the body has made it difficult to obtain and sustain therapeutic levels in the blood and target tissues. Studies were conducted to characterize the synergistic anticancer antiproliferative effects of combined low dose celecoxib and γ-tocotrienol treatment on mammary tumor cells in culture. The highly malignant mouse +SA mammary epithelial cells were maintained in culture on serum-free defined control or treatment media. Treatment effects on COX-1, COX-2, Akt, NFκB and prostaglandin E 2 (PGE 2 ) synthesis was assessed following a 3-or 4-day culture period. Treatment with 3-4 μM γ-tocotrienol or 7.5-10 μM celecoxib alone significantly inhibited +SA cell growth in a dose-responsive manner. However, combined treatment with subeffective doses of γ-tocotrienol (0.25 μM) and celecoxib (2.5 μM) resulted in a synergistic antiproliferative effect, as determined by isobologram analysis, and this growth inhibitor effect was associated with a reduction in PGE 2 synthesis, and decrease in COX-2, phospho-Akt (active), and phospho-NFκB (active) levels. These results demonstrate that the synergistic anticancer effects of combined celecoxib and γ-tocotrienol therapy are mediated by COX-2 dependent and independent mechanisms. These findings also suggest that combination therapy with these agents may provide enhanced therapeutic response in breast cancer patients, while avoiding the toxicity associated with high-dose COX-2 inhibitor monotherapy.

Pomegranate extract (PE) inhibits the proliferation of breast cancer cells and stimulates apoptosis in MCF-7 breast cancer cells. While PE is a potent antioxidant, the present studies were conducted to examine the mechanisms of action of PE beyond antioxidation by studying cellular and molecular mechanisms underlying breast tumorigenesis. PE inhibited cell growth by inducing cell cycle arrest in G2 /M followed by the induction of apoptosis. In contrast, antioxidants N-acetylcysteine and Trolox did not affect cell growth at doses containing equivalent antioxidant capacity as PE, suggesting that growth inhibition by PE cannot solely be attributed to its high antioxidant potential. DNA microarray analysis revealed that PE downregulated genes associated with mitosis, chromosome organization, RNA processing, DNA replication and DNA repair, and upregulated genes involved in regulation of apoptosis and cell proliferation. Both microarray and quantitative RT-PCR indicated that PE downregulated important genes involved in DNA double strand break (DSB) repair by homologous recombination (HR), such as MRE11, RAD50, NBS1, RAD51, BRCA1, BRCA2, and BRCC3. Downregulation of HR genes correlated with increased levels of their predicted microRNAs (miRNAs), miR-183 (predicted target RAD50) and miR-24 (predicted target BRCA1), suggesting that PE may regulate miRNAs involved in DNA repair processes. Further, PE treatment increased the frequency of DSBs. These data suggest that PE downregulates HR which sensitizes cells to DSBs, growth inhibition and apoptosis. Because HR represents a novel target for cancer therapy, downregulation of HR by PE may be exploited for sensitization of tumors to anticancer drugs.

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Silver nanoparticles (AgNPs) are widely used in consumer and medical products. However, most AgNP toxicity data are based on in vitro studies. Only a few studies were performed in mammals and no studies systematically assessed cancer risk of AgNPs. In this study, we examined whether oral exposure to polyvinylpyrrolidone (PVP)-coated AgNPs induces DNA damage and permanent genome alterations, and modulates DNA repair gene expression in vivo in mice. We found that AgNPs induced large DNA deletions in developing embryos, irreversible chromosomal damage in bone marrow, and double strand breaks and oxidative DNA damage in peripheral blood and/or bone marrow. DNA Repair RT Profiler PCR Array showed that AgNPs altered expression of 36 of the 84 genes from which 24 genes were downregulated and 12 genes were upregulated. In particular, AgNPs downregulated a significant proportion of base excision repair (BER) genes. We hypothesized that downregulation of BER by AgNPs contributes to oxidative DNA damage and subsequent genomic instability, which predicts that BER defects enhance sensitivity to AgNPs. We tested this hypothesis in mice deficient in MutY homologue (Myh). Myh excises adenine mispaired with 8-oxoguanine to counteract its promutagenic activity and also has a role in cell cycle check points and apoptosis. MYH mutations are common in humans and predispose to colorectal and other types of cancer. Myh deficient mice were hypersensitive to AgNP-induced chromosomal damage. In summary, oral ingestion of AgNPs induces permanent genome alterations and may therefore cause cancer. In addition, BER defects, especially, Myh mutations, enhance sensitivity to AgNPs.