Pharmaceutical Research (v.27, #6)

Current Trends in the Chemoprevention of Cancer by Fazlul H. Sarkar (945-949).
is a professor of pathology at Karmanos Cancer Center, Wayne State University with a track-record of cancer research for over 30 years. He received his MS and Ph.D. degrees in biochemistry from two of the most premiere institutions in India in 1974 and 1978, respectively. In 1978, he migrated to the United States for his post-doctoral training in molecular biology at Memorial Sloan Kettering Cancer Center in New York, and he served in different capacities in several other institutions prior to arriving at Wayne State University in 1989. His research is focused on understanding the role of a “master” transcription factor, NF-κB, and the regulation of its upstream and downstream signaling molecules in solid tumors. Moreover, his focused research has also been directed toward elucidating the molecular mechanisms of action of “natural agents” and synthetic small molecules for cancer prevention and therapy. He has done a tremendous amount of work in vitro and in vivo, documenting that several natural agents could be useful for chemopreventive research. Most importantly, his work has led to the discovery of the role of chemopreventive agents in sensitization of cancer cells (reversal of drug resistance) to conventional therapeutics (chemo-radio-therapy). He is one of the pioneers in developing natural agents, such as isoflavones, curcumin and indole compounds like DIM (B-DIM), for clinical use, and his basic science research findings led to the initiation of clinical trials in breast, pancreas and prostate cancers at the Karmanos Cancer Institute. He is a true translational researcher bringing his laboratory research findings into clinical practice. Dr. Sarkar is also involved in several collaborative projects, including breast, lung and pancreatic cancer for both pre-clinical and phase II clinical trials, with other scientists within the institution as well as collaborative work with basic scientists and physician scientists at MD Anderson Cancer Center. He has published over 350 original scientific articles in peer-reviewed journals and written more than 50 review articles and book chapters. He has been continuously funded by NCI, NIH, and the Department of Defense (DoD). Dr. Sarkar has trained numerous pre-doctoral and post-doctoral students throughout the last 20 years at Wayne State University. In addition, Dr. Sarkar has served and is still serving on a number of departmental, university and national committees, and continues to serve both NIH and DoD study sections, including NIH program projects, SPORE grants and Cancer Center Core grants (site visit) for NCI-designated Comprehensive Cancer Centers. He is currently a Senior Editor of the journal Molecular Cancer Therapeutics and a member of the editorial board of many scientific journals.

Cancer Chemoprevention by Natural Products: How Far Have We Come? by Rajendra G. Mehta; Genoveva Murillo; Rajesh Naithani; Xinjian Peng (950-961).
Since ancient times, natural products, herbs and spices have been used for preventing several diseases, including cancer. The term chemoprevention was coined in the late 1970s and referred to the prevention of cancer by selective use of phytochemicals or their analogs. The field utilizes experimental carcinogenesis models to examine the efficacy of chemopreventive agents in a stage-specific manner. The concept of using naturally derived chemicals as potential chemopreventive agents has advanced the field dramatically. Throughout the years, a vast number of chemopreventive agents present in natural products have been evaluated using various experimental models. A number of them have progressed to early clinical trials. More recently, the focus has been directed towards molecular targeting of chemopreventive agents to identify mechanism(s) of action of these newly discovered bioactive compounds. Moreover, it has been recognized that single agents may not always be sufficient to provide chemopreventive efficacy, and, therefore, the new concept of combination chemoprevention by multiple agents or by the consumption of “whole foods” has become an increasingly attractive area of study. Novel technologies, such as nanotechnology, along with a better understanding of cancer stem cells, are certain to continue the advancement of the field of cancer chemoprevention in years to come.
Keywords: chemoprevention; natural products; phytochemicals; signaling

Apigenin: A Promising Molecule for Cancer Prevention by Sanjeev Shukla; Sanjay Gupta (962-978).
Apigenin, a naturally occurring plant flavone, abundantly present in common fruits and vegetables, is recognized as a bioactive flavonoid shown to possess anti-inflammatory, antioxidant and anticancer properties. Epidemiologic studies suggest that a diet rich in flavones is related to a decreased risk of certain cancers, particularly cancers of the breast, digestive tract, skin, prostate and certain hematological malignancies. It has been suggested that apigenin may be protective in other diseases that are affected by oxidative process, such as cardiovascular and neurological disorders, although more research needs to be conducted in this regard. Human clinical trials examining the effect of supplementation of apigenin on disease prevention have not been conducted, although there is considerable potential for apigenin to be developed as a cancer chemopreventive agent.
Keywords: apigenin; apoptosis; cell cycle; chemoprevention; flavonoids

Plant polyphenols are important components of human diet, and a number of them are considered to possess chemopreventive and therapeutic properties against cancer. They are recognized as naturally occurring anti-oxidants but also act as pro-oxidants catalyzing DNA degradation in the presence of metal ions such as copper. The plant polyphenol resveratrol confers resistance to plants against fungal agents and has been implicated as a cancer chemopreventive agent. Of particular interest is the observation that resveratrol has been found to induce apoptosis in cancer cell lines but not in normal cells. Over the last few years, we have shown that resveratrol is capable of causing DNA breakage in cells such as human lymphocytes. Such cellular DNA breakage is inhibited by copper specific chelators but not by iron and zinc chelating agents. Similar results are obtained by using permeabilized cells or with isolated nuclei, indicating that chromatin-bound copper is mobilized in this reaction. It is well established that tissue, cellular and serum copper levels are considerably elevated in various malignancies. Therefore, cancer cells may be more subject to electron transfer between copper ions and resveratrol to generate reactive oxygen species responsible for DNA cleavage. The results are in support of our hypothesis that anti-cancer mechanism of plant polyphenols involves mobilization of endogenous copper and the consequent pro-oxidant action. Such a mechanism better explains the anti-cancer effects of resveratrol, as it accounts for the preferential cytotoxicity towards cancer cells.
Keywords: cancer; comet assay; endogenous copper; pro-oxidant action; resveratrol

Modulating Polo-Like Kinase 1 as a Means for Cancer Chemoprevention by Travis L. Schmit; Mark C. Ledesma; Nihal Ahmad (989-998).
Naturally occurring agents have always been appreciated for their medicinal value for both their chemopreventive and therapeutic effects against cancer. In fact, the majority of the drugs we use today, including the anti-cancer agents, were originally derived from natural compounds, either in their native form or modified to enhance their bioavailability or specificity. It is believed that for maximum effectiveness, it will useful to design novel target-based agents for chemoprevention as well as the treatment of cancer. Recent studies have shown that the serine/threonine kinase polo-like kinase (Plk) 1 is widely overexpressed in a variety of cancers and is being increasingly appreciated as a target for cancer management. Additionally, several chemopreventive agents have been shown to inhibit Plk1 in cancer cells. In this review, we will discuss if Plk1 could also be a target for designing novel strategies for cancer chemoprevention.
Keywords: cancer; chemoprevention; polo-like kinase

Persistent inflammatory tissue damage is causally associated with each stage of carcinogenesis. Inflammation-induced generation of reactive oxygen species, reactive nitrogen species, and other reactive species not only cause DNA damage and subsequently mutations, but also stimulate proliferation of initiated cells and even metastasis and angiogenesis. Induction of cellular cytoprotective enzymes (e.g., heme oxygenase-1, NAD(P)H:quinone oxidoreductase, superoxide dismutase, glutathione-S-transferase, etc.) has been shown to mitigate aforementioned events implicated in inflammation-induced carcinogenesis. A unique feature of genes encoding these cytoprotective enzymes is the presence of a cis-acting element, known as antioxidant response element (ARE) or electrophile response element (EpRE), in their promoter region. A stress-responsive transcription factor, nuclear factor erythroid-2-related factor-2 (Nrf2), initially recognized as a key transcriptional regulator of various cytoprotective enzymes, is known to play a pivotal role in cellular defense against inflammatory injuries. Activation of Nrf2 involves its release from the cytosolic repressor Kelch-like ECH-associated protein-1 (Keap1) and subsequent stabilization and nuclear localization for ARE/EpRE binding. Genetic or pharmacologic inactivation of Nrf2 has been shown to abolish cytoprotective capability and to aggravate experimentally induced inflammatory injuries. Thus, Nrf2-mediated cytoprotective gene induction is an effective strategy for the chemoprevention of inflammation-associated carcinogenesis.
Keywords: anti-inflammation; chemoprevention; chemopreventive agents; inflammation; Keap1; Nrf2; redox signaling

Diet, MicroRNAs and Prostate Cancer by Sharanjot Saini; Shahana Majid; Rajvir Dahiya (1014-1026).
MicroRNAs (miRNAs) constitute an evolutionarily conserved class of small non-coding RNAs that are endogenously expressed with crucial functions in fundamental cellular processes such as cell cycle, apoptosis and differentiation. Disturbance of miRNA expression and function leads to deregulation of basic cellular processes leading to tumorigenesis. A growing body of experimental evidence suggests that human tumors have deregulated expression of microRNAs, which have been proposed as novel oncogenes or tumor suppressors. Recent studies have shown that microRNA expression patterns serve as phenotypic signatures of different cancers and could be used as diagnostic, prognostic and therapeutic tools. A few studies have analyzed global microRNA expression profiles or the functional role of microRNAs in prostate cancer. Here we have reviewed the role of microRNAs in prostate carcinogenesis by summarizing the findings from such studies. In addition, recent evidence indicates that dietary factors play an important role in the process of carcinogenesis through modulation of miRNA expression, though such studies are lacking in regards to prostate cancer. It has been proposed that dietary modulation of miRNA expression may contribute to the cancer-protective effects of dietary components. In this review, we have summarized findings from studies on the effect of dietary agents on miRNA expression and function.
Keywords: diet; microRNAs; prostate cancer

Regulation of microRNAs by Natural Agents: An Emerging Field in Chemoprevention and Chemotherapy Research by Yiwei Li; Dejuan Kong; Zhiwei Wang; Fazlul H. Sarkar (1027-1041).
In recent years, microRNAs have received greater attention in cancer research. These small, non-coding RNAs could inhibit target gene expression by binding to the 3′ untranslated region of target mRNA, resulting in either mRNA degradation or inhibition of translation. miRNAs play important roles in many normal biological processes; however, studies have also shown that aberrant miRNA expression is correlated with the development and progression of cancers. The miRNAs could have oncogenic or tumor suppressor activities. Moreover, some miRNAs could regulate formation of cancer stem cells and epithelial-mesenchymal transition phenotype of cancer cells which are typically drug resistant. Furthermore, miRNAs could be used as biomarkers for diagnosis and prognosis, and thus miRNAs are becoming emerging targets for cancer therapy. Recent studies have shown that natural agents including curcumin, isoflavone, indole-3-carbinol, 3,3′-diindolylmethane, (-)-epigallocatechin-3-gallate, resveratrol, etc. could alter miRNA expression profiles, leading to the inhibition of cancer cell growth, induction of apoptosis, reversal of epithelial-mesenchymal transition, or enhancement of efficacy of conventional cancer therapeutics. These emerging results clearly suggest that specific targeting of miRNAs by natural agents could open newer avenues for complete eradication of tumors by killing the drug-resistant cells to improve survival outcome in patients diagnosed with malignancies.
Keywords: cancer therapy; chemoprevention; microRNA; natural agents

Resveratrol and Resveratrol Analogues—Structure—Activity Relationship by Thomas Szekeres; Monika Fritzer-Szekeres; Philipp Saiko; Walter Jäger (1042-1048).
Resveratrol (3,4′,5-trihydroxy-trans-stilbene) is a compound found in wine and is held responsible for a number of beneficial effects of red wine. Besides the prevention of heart disease and significant anti-inflammatory effects, resveratrol might inhibit tumor cell growth and even play a role in the aging process. We here describe the structure-activity relationship of resveratrol and analogues of resveratrol regarding the free radical scavenging and antitumor effects of this exciting natural compound. In addition, we have synthesized a number of analogues of resveratrol with the aim to further improve the beneficial effects of resveratrol. Our studies were based on the analysis of structural properties, which were responsible for the most important effects of this compound. Striking in vivo effects can be observed with hexahydroxystilbene (M8), the most effective synthetic analogue of resveratrol. We could show that M8 inhibits tumor as well as metastasis growth of human melanoma in two different animal models, alone and in combination with dacarbacine.
Keywords: apoptosis; gallic acid; resveratrol; ribonucleotide reductase; stilbene analogues

Bitter Melon: Antagonist to Cancer by Pratibha Nerurkar; Ratna B. Ray (1049-1053).
The incidence of cancer is increasing worldwide, in spite of substantial progress in the development of anti-cancer therapies. One approach to control cancer could be its prevention by diet, which inhibits one or more neoplastic events and reduces cancer risk. Dietary compounds offer great potential in the fight against cancer by inhibiting the carcinogenesis process through the regulation of cell homeostasis and cell-death machineries. For centuries, Ayurveda (Indian traditional medicine) has recommended the use of bitter melon (Momordica charantia) as a functional food to prevent and treat diabetes and associated complications. It is noteworthy to mention that bitter melon extract has no-to-low side effects in animals as well as in humans. The anti-tumor activity of bitter melon has recently begun to emerge. This review focuses on recent advancements in cancer chemopreventive and anti-cancer efficacy of bitter melon and its active constituents. Several groups of investigators have reported that treatment of bitter-melon-related products in a number of cancer cell lines induces cell cycle arrest and apoptosis without affecting normal cell growth. Therefore, the effect of bitter melon should be beneficial for health, and use of the non-modified dietary product is cost effective.
Keywords: bitter melon; cell proliferation; chemoprevention; therapeutics

Nanochemoprevention by Bioactive Food Components: A Perspective by Imtiaz A. Siddiqui; Hasan Mukhtar (1054-1060).
Chemoprevention through the use of bioactive food components is a practical approach for cancer control. Despite abundant efficacy data under preclinical settings, this strategy has resulted in limited success for human cancer control. Amongst many reasons, inefficient systemic delivery and bioavailability of promising chemopreventive agents are considered to significantly contribute to such a disconnect. We recently introduced a novel concept in which we utilized nanotechnology for enhancing the outcome of chemoprevention (Cancer Res. 2009; 69:1712–6) and termed it nanochemoprevention. To establish the proof-of-principle of nanotechnology for cancer management, we determined the efficacy of a well-known chemopreventive agent epigallocatechin-3-gallate (EGCG) encapsulated in polylactic acid (PLA)-polyethylene glycol (PEG) nanoparticles in preclinical settings and observed that, compared to non-encapsulated EGCG, nano-EGCG retained its biological efficacy with over 10-fold dose advantage both in cell culture system and in vivo settings in athymic nude mice implanted with human prostate cancer cells. This study laid the foundation of nanochemoprevention by bioactive food components. Since oral consumption is the most desirable and acceptable form of delivery of bioactive food components, it will be important to develop nanoparticles containing bioactive food components that are suitable for oral consumption for which experiments are underway in this laboratory.
Keywords: bioactive food components; chemoprevention; nanochemoprevention; nanotechnology

Humans typically consume “natural agents” that are believed to be chemoprotective and are known to decrease inflammation biomarker NF-κB in vitro; however, no intervention studies in humans have been done to date. This commentary documents the in vivo results as a powerful example for supporting the superiority of a complex mixture of natural agents. Human volunteers consumed two 500 mg capsules (BID) containing a mixture of natural agents for a period of 2 weeks, and blood samples were collected pre- and post-intervention. The purified lymphocytes were subjected to ex-vivo exposure to TNF-α or kept as untreated control. The mean NF-κB DNA binding activity was increased upon TNF-α treatment in pre-intervention samples; however, TNF-α was unable to induce NF-κB in post-intervention samples, suggesting that the mixture of four important natural agents could be useful to protect humans against oxidative stress.
Keywords: curcumin; epigallocatechin-3-gallate (EGCG); isoflavone; NF-κB; TNF-α; resveratrol

Effect of Dietary Polyphenon E and EGCG on Lung Tumorigenesis in A/J Mice by Qi Zhang; Huijing Fu; Jing Pan; Jun He; Seto Ryota; Yukihiko Hara; Yian Wang; Ronald A Lubet; Ming You (1066-1071).
To compare the chemopreventive efficacy of Polyphenon E (Poly E), (−)-epigallocatechin-3-gallate (EGCG) and Polyphenon E without EGCG (Poly E-EGCG) on the development of benzo(a)pyrene (B(a)P)-induced lung tumors in A/J mice.Female A/J mice were given a single intraperitoneal injection of B(a)P (100 mg/kg body weight). One week after B(a)P injection, animals received AIN-76A purified powder diet containing 0.975% (wt/wt) EGCG, 0.525% (wt/wt) Poly E-EGCG or 1.5% (wt/wt) Poly E for 24 weeks or control diet with no additives.Poly E treatment significantly decreased tumor multiplicity by 52% and tumor load by 64%, while EGCG and Poly E-EGCG did not significantly inhibit lung tumor multiplicity. EGCG was more stable in a complex mixture (Poly E) than as a pure compound.EGCG was ineffective when administered by diet likely due to its instability. Thus, EGCG’s efficacy on mice lung tumorigenesis requires the presence of other tea catechins.
Keywords: chemoprevention; degradation; EGCG; lung tumorigenesis; polyphenon E

Diallyl Trisulfide-Induced G2/M Phase Cell Cycle Arrest in DU145 Cells Is Associated with Delayed Nuclear Translocation of Cyclin-Dependent Kinase 1 by Anna Herman-Antosiewicz; Young-Ae Kim; Su-Hyeong Kim; Dong Xiao; Shivendra V. Singh (1072-1079).
The present study was undertaken to gain insight into the molecular mechanism of G2/M phase cell cycle arrest resulting from treatment of DU145 cells with diallyl trisulfide (DATS), a promising cancer chemopreventive constituent of garlic.Cell cycle distribution was determined by flow cytometry. Immunoblotting was performed to determine protein expression. Overexpression of wild-type or mutant Cdc25C was achieved by transient transfection. Nuclear and cytoplasmic localization of cyclin B1 and cyclin-dependent kinase 1 (cdk1) was studied by immunoblotting.Exposure of DU145 human prostate cancer cells to DATS resulted in concentration- and time-dependent accumulation of G2/M phase cells, which correlated with down-regulation as well as increased S216 phosphorylation of Cdc25C. Ectopic expression of wild-type or redox-insensitive mutants (C330S and C330S/C377S) or S216A mutant of Cdc25C failed to confer protection against DATS-induced G2/M phase arrest. The DATS-mediated G2/M phase cell cycle arrest was also independent of reduced complex formation between cdk1 and cyclin B1, but correlated with delayed nuclear translocation of cdk1.The present study indicates that the DATS-mediated G2/M phase cell cycle arrest in DU145 cells results from differential kinetics of nuclear localization of cdk1 and cyclin B1.
Keywords: Cdc25C; cdk1; cell cycle; cyclin B1; diallyl trisulfide

Suppression of the Inflammatory Cascade is Implicated in Resveratrol Chemoprevention of Experimental Hepatocarcinogenesis by Anupam Bishayee; Abhijeet Waghray; Kendra F. Barnes; Thomas Mbimba; Deepak Bhatia; Malay Chatterjee; Altaf S. Darvesh (1080-1091).
Resveratrol, present in grapes and red wine, has been found to prevent diethylnitrosamine (DENA)-initiated rat liver tumorigenesis, though the chemopreventive mechanisms are not completely elucidated. The current study was designed to explore whether the antiinflammatory properties of resveratrol play a role in its antihepatocarcinogenic action.Liver samples were harvested from a 20-week chemopreventive study in which resveratrol (50, 100 and 300 mg/kg) was shown to inhibit DENA-induced hepatocyte nodules in Sprague-Dawley rats in a dose-responsive manner. Hepatic preneoplastic and inflammatory markers, namely heat shock protein (HSP70), cyclooxygenase-2 (COX-2) and nuclear factor-κB (NF-κB), were studied using immunohistochemical as well as Western blot techniques.Resveratrol dose-dependently suppressed DENA-induced increased expressions of hepatic HSP70 and COX-2. Resveratrol also attenuated the DENA-mediated translocation of NF-κB p65 from the cytosol to the nucleus with stabilization of inhibitory κB.The present findings indicate that resveratrol exerts chemoprevention of hepatocarcinogenesis possibly through antiinflammatory effects during DENA-evoked rat liver carcinogenesis by suppressing elevated levels of HSP70, COX-2 as well as NF-κB. These beneficial effects combined with an excellent safety profile encourage the development of resveratrol for chemoprevention and intervention of human HCC that remains a devastating disease.
Keywords: chemoprevention; hepatocarcinogenesis; inflammation; NF-κB; resveratrol

The purpose of this study was to determine the chemopreventive mechanism of dietary grape seed proanthocyanidins (GSPs) against ultraviolet (UV) radiation-induced skin tumor development in mice.Six-to-seven-week-old SKH-1 hairless mice were subjected to photocarcinogenesis protocol, and exposed to UVB radiation (180 mJ/cm2) three times/week for 24 weeks. Mice were fed a standard AIN76A control diet with or without supplementation with grape seed proanthocyanidins (GSPs; 0.2% or 0.5%, w/w). At the termination of the experiment, mice were sacrificed, and skin and skin tumor samples were harvested and subjected to the analysis of biomarkers related to inflammation using immunostaining, western blot analysis, ELISA and real-time PCR.Dietary GSPs inhibited UVB-induced infiltration of proinflammatory leukocytes and the levels of myeloperoxidase, cyclooxygenase-2 (COX-2), prostaglandin (PG) E2, cyclin D1 and proliferating cell nuclear antigen (PCNA) in the skin and skin tumors compared to non-GSPs-treated UVB irradiated mouse skin and skin tumors. GSPs also significantly inhibited the levels of proinflammatory cytokines, tumor necrosis factor-α (P < 0.01), IL-1β (P < 0.001) and IL-6 (P < 0.001), in UVB-exposed skin and skin tumors.The results from this study clearly suggest that dietary GSPs inhibit photocarcinogenesis in mice through the inhibition of UVB-induced inflammation and mediators of inflammation in mouse skin.
Keywords: chemoprevention; COX-2; grape seed proanthocyanidins; skin cancer; ultraviolet radiation

Melanoma is an aggressive neoplasm with a propensity for metastases and resistance to therapy. Previously, we showed that (-)-epigallocatechin-3-gallate (EGCG), the major polyphenolic antioxidant present in green tea, resulted in a significant decrease in the viability and growth of melanoma and induction of apoptosis via modulation of the cki-cdk-cyclin network and Bcl2 family proteins. Epigenetic regulation of gene transcription by histone deacetylase (HDAC) inhibitors is gaining momentum as a novel cancer therapy. SAHA-suberoylanilidine hydroxamic acid Zolinza® (vorinostat) is the first HDAC inhibitor approved by the U.S. FDA. In this study, we determined if vorinostat alone or in combination with EGCG imparts anti-proliferative effects against human melanoma cells.Employing human melanoma cell lines A-375, Hs-294T and G-361, we determined the effect of vorinostat and/or EGCG on 1) growth/viability and colony formation, 2) apoptosis, and 3) the critical molecules involved in cell cycle and apoptosis regulation.Our data demonstrated that the anti-proliferative effects of vorinostat were greater than or similar to those of EGCG among the cell lines tested. Furthermore, relative to monotherapy, the combination treatment resulted in significantly greater inhibition of cell proliferation, increased apoptosis, activation of p21, p27 and caspases (3, 7 and 9) and Bax as well as down-regulation of cdk2, cdk4, cyclin A, NF-κB protein p65/RelA and Bcl2 protein and transcript.Our preclinical findings suggest that combination therapy with EGCG and vorinostat may be beneficial for the management of human melanoma.
Keywords: chemoprevention; EGCG; green tea; melanoma; vorinostat

Daidzein Effect on Hormone Refractory Prostate Cancer In Vitro and In Vivo Compared to Genistein and Soy Extract: Potentiation of Radiotherapy by Vinita Singh-Gupta; Hao Zhang; Christopher K. Yunker; Zahra Ahmad; Danielle Zwier; Fazlul H. Sarkar; Gilda G. Hillman (1115-1127).
Genistein, the major bioactive isoflavone of soybeans, acts as a radiosensitizer for prostate cancer (PCa) both in vitro and in vivo. However, pure genistein promoted increased metastasis to lymph nodes. A mixture of soy isoflavones (genistein, daidzein, glycitein) did not cause increased metastasis, but potentiated radiotherapy. We tested whether daidzein could negate genistein-induced metastasis.Mice bearing PC-3 prostate tumors were treated with daidzein, genistein or both, and with tumor irradiation. Primary tumors and metastases were evaluated. The effects of each isoflavone and soy were compared in vitro using PC-3 (AR−) and C4-2B (AR+) androgen-independent PCa cell lines.Daidzein did not increase metastasis to lymph nodes and acted as a radiosensitizer for prostate tumors. Daidzein inhibited cell growth and enhanced radiation in vitro but at doses higher than genistein or soy. Daidzein caused milder effects on inhibition of expression and/or activities of APE1/Ref-1, HIF-1α and NF-κB in PC-3 and C4-2B cells.Daidzein could be the component of soy that protects against genistein-induced metastasis. Daidzein inhibited cell growth and synergized with radiation, affecting APE1/Ref-1, NF-κB and HIF-1α, but at lower levels than genistein and soy, in AR+ and AR- PCa cells, suggesting it is an AR-independent mechanism.
Keywords: androgen receptor; prostate cancer; radiation; soy isoflavones

In Vitro and In Vivo Effects of Water Extract of White Cocoa Tea (Camellia ptilophylla) Against Human Prostate Cancer by Li Peng; Naghma Khan; Farrukh Afaq; Chuangxing Ye; Hasan Mukhtar (1128-1137).
We evaluated the chemotherapeutic effect of water extract of white cocoa tea (WCTE) against human prostate cancer (PCa) in vitro and in vivo.Cell viability and cell cycle distribution were determined by MTT assay and flow cytometry, respectively. Western blotting was performed to determine changes in levels of various proteins. Effect of WCTE was determined in athymic nude mice implanted with PC-3 cells.Treatment with WCTE (100–150 µg/ml) inhibited cell proliferation, which correlated with G2/M phase arrest in PC-3 cells. WCTE treatment to PC-3 cells resulted in (1) induction of WAF1/p21 and KIP1/p27, (2) decrease in cyclins D1, D2 and E, (3) decrease in cyclin-dependent kinase (cdk) 2, 4 and 6, (4) induction of Bax and down-regulation of Bcl-2, (5) decrease in procaspase-3, -8, (6) inhibition of nuclear translocation and phosphorylation of NF-κB and activation of IKKα, and (7) inhibition of phosphorylation and degradation of IκBα. Oral administration of WCTE (0.1 and 0.2%, wt/vol) to athymic nude mice resulted in greater than 50% inhibition of tumor growth. There was a decrease in expressions of cyclin D1, Bcl-2 and p-NF-κB and an increase in WAF1/p21 and Bax in tumor tissues of mice.WCTE can be a useful chemotherapeutic agent against human PCa.
Keywords: apoptosis; cell cycle arrest; prostate cancer; white cocoa tea

Multiple Berry Types Prevent N-nitrosomethylbenzylamine-Induced Esophageal Cancer in Rats by Gary D. Stoner; Li-Shu Wang; Claire Seguin; Claudio Rocha; Kristen Stoner; Steven Chiu; A. Douglas Kinghorn (1138-1145).
The present study compared the ability of different berry types to prevent chemically-induced tumorigenesis in the rat esophagus. We also determined if berries influence the levels of inflammatory cytokines in the serum of carcinogen-treated rats.Rats were treated with the carcinogen N-nitrosomethylbenzylamine (NMBA) for 5 weeks, then placed on diets containing 5% of either black or red raspberries, strawberries, blueberries, noni, açaí or wolfberry until the end of the study. The effects of the berries on tumor incidence, multiplicity and size were determined, as well as their effects on the levels of selected inflammatory cytokines in serum.All berry types were about equally effective in inhibiting NMBA-induced tumorigenesis in the rat esophagus. They also reduced the levels of the serum cytokines, interleukin 5 (IL-5) and GRO/KC, the rat homologue for human interleukin-8 (IL-8), and this was associated with increased serum antioxidant capacity.Seven berry types were about equally capable of inhibiting tumor progression in the rat esophagus in spite of known differences in levels of anthocyanins and ellagitannins. Serum levels of IL-5 and GRO/KC (IL-8) may be predictive of the inhibitory effect of chemopreventive agents on rat esophageal carcinogenesis.
Keywords: berries; chemoprevention; esophagus; rat

Structure-Activity Studies on Therapeutic Potential of Thymoquinone Analogs in Pancreatic Cancer by Sanjeev Banerjee; Asfar S. Azmi; Subhash Padhye; Marjit W. Singh; Jubaraj B. Baruah; Philip A. Philip; Fazlul H. Sarkar; Ramzi M. Mohammad (1146-1158).
Pancreatic cancer (PC) is one of the deadliest of all tumors. Previously, we were the first to show that Thymoquinone (TQ) derived from black seed (Nigella sativa) oil has anti-tumor activity against PC. However, the concentration of TQ required was considered to be high to show this efficacy. Therefore, novel analogs of TQ with lower IC50 are highly desirable.We have synthesized a series of 27 new analogs of TQ by modifications at the carbonyl sites or the benzenoid sites using single pot synthesis and tested their biological activity in PC cells.Among these compounds, TQ-2G, TQ-4A1 and TQ-5A1 (patent pending) were found to be more potent than TQ in terms of inhibition of cell growth, induction of apoptosis and modulation of transcription factor-NF-κB. We also found that our novel analogs were able to sensitize gemcitabine and oxaliplatin-induced apoptosis in MiaPaCa-2 (gemcitabine resistant) PC cells, which was associated with down-regulation of Bcl-2, Bcl-xL, survivin, XIAP, COX-2 and the associated Prostaglandin E2.From our results, we conclude that three of our novel TQ analogs warrant further investigation against PC, especially in combination with conventional chemotherapeutic agents.
Keywords: apoptosis; pancreatic cancer; thymoquinone; thymoquinone analogs

FoxM1 is a Novel Target of a Natural Agent in Pancreatic Cancer by Zhiwei Wang; Aamir Ahmad; Sanjeev Banerjee; Asfar Azmi; Dejuan Kong; Yiwei Li; Fazlul H. Sarkar (1159-1168).
Pancreatic cancer remains the fourth most common cause of cancer-related death in the United States. Therefore, novel strategies for the prevention and/or treatment are urgently needed. Genistein has been found to be responsible for lowering the rate of pancreatic cancer. However, the molecular mechanisms by which genistein elicits its effects on pancreatic cancer cells has not been fully elucidated. Therefore, the purpose of the current study was to elucidate the anti-cancer mechanism(s) of genistein.Multiple molecular techniques, such as Real-time RT-PCR, Western blot analysis, invasion assay, immunofluorescence assay, gene transfection, MTT assay, and Histone/DNA ELISA, were used.We found that genistein inhibited cell growth accompanied by induction of apoptosis with concomitant attenuation of FoxM1 and its downstream genes, such as survivin, cdc25a, MMP-9, and VEGF, resulting in the inhibition of pancreatic cancer cell invasion. We also found that down-regulation of FoxM1 by siRNA prior to genistein treatment resulted in enhanced cell growth inhibition and induction of apoptosis.This is the first report showing the molecular role of FoxM1 in mediating the biological effects of genistein in pancreatic cancer cells, suggesting that FoxM1 could be a novel target for the treatment of pancreatic cancer.
Keywords: FoxM1; genistein; invasion; pancreatic cancer; proliferation

AAPS Connection (1169-1170).