Inhibition of metastasis and angiogenesis in Hep-2 cells by wheatgrass extract - an in vitro and in silico approach.

Metastasis is the major hindrance in the treatment of all cancers, including laryngeal squamous cell carcinoma. Intensive researches are under way to identify the effective natural polyphenols with anti-metastatic ability for cancer treatment. Wheatgrass, an herbal plant has been reported to show anticancer effects. Hence, in this study, we aimed to analyze the anti-metastatic effect of methanol extract of wheatgrass (MEWG). The levels of metastatic marker proteins were determined by western blot. PI3K and AKT levels were determined by real time (RT)-PCR analysis. In silico molecular docking was done to check the interaction of the 14 components (identified by HPLC/GCMS) of MEWG with PI3K and AKT. MEWG effectively decreased the metastatic protein expressions, namely VEGF, MMP-9 and COX-2 and increased TIMP-2. RT-PCR results showed reduced m-RNA levels of both PI3K and AKT when compared to control. Molecular docking studies revealed interaction of most of the identified compounds of the extract with the important residues of PI3K and AKT. These findings indicate that MEWG inhibits metastasis and angiogenesis in Hep-2 cells possibly via PI3K/AKT due to the cumulative effect of polyphenols and other constituent present in extract. The compounds of the extract were also found to be directly involved in inhibition of AKT/PI3K, thus could help to restrain metastasis.

Vitamin and carotenoid intake and risk of head-neck cancer subtypes in the Netherlands Cohort Study.

BACKGROUND: Head and neck cancer (HNC) is the seventh most-common type of cancer worldwide. Evidence regarding the potential protective effect of vitamins and carotenoids on HNC is limited and mostly based on case-control studies. OBJECTIVE: We evaluated the association of intake of dietary vitamins C and E (including supplementation) and the most-common carotenoids (α-carotene, ß-carotene, lutein plus zeaxanthin, lycopene, and ß-cryptoxanthin) and risk of HNC and HNC subtypes in a large prospective study. DESIGN: The Netherlands Cohort Study included 120,852 participants. For efficiency reasons, a case-cohort design was used. At baseline in 1986, participants completed a food-frequency questionnaire. A subcohort was randomly selected from the total cohort. After 20.3 y of follow-up, 3898 subcohort members and 415 HNC cases [131 oral cavity cancer (OCCs), 88 oro-/hypopharyngeal cancer (OHPs), and 193 laryngeal cancer cases] were available for analysis. Rate ratios and 95% CIs for highest (quartile 4) compared with lowest (quartile 1) quartiles of vitamin and carotenoid intake were estimated by using the Cox proportional hazards model. RESULTS: A strong inverse association was shown between vitamin C and HNC overall (multivariable-adjusted rate ratio for quartile 4 compared with quartile 1: 0.39; 95% CI: 0.23, 0.66; P-trend < 0.001), OCC (multivariable-adjusted rate ratio for quartile 4 compared with quartile 1: 0.35; 95% CI: 0.16, 0.77; P-trend < 0.05), and OHPC (multivariable-adjusted rate ratio for quartile 4 compared with quartile 1: 0.29; 95% CI: 0.12, 0.67; P-trend < 0.01). No statistically significant results were shown for vitamin E, α-carotene, ß-carotene, lycopene, and lutein plus zeaxanthin. The association of vitamin E and HNC was modified by alcohol status (P-interaction = 0.003) with lower risks in alcohol abstainers. CONCLUSIONS: With this study, we show an inverse association between intake of vitamin C and the incidence of HNC and HNC-subtypes. Future research is recommended to investigate the underlying mechanisms and to confirm our results, which may be promising for the prevention of HNC.

Green tea prevents down-regulation of gap junction intercellular communication in human keratinocytes treated with PMA.

It has been suggested that connexin (Cx) gap junction proteins act as tumor suppressors and green tea has a potential to prevent tumor development, however, the studies on their association with human keratinocytes were rare. We evaluated the effects of a tumor promoter, phorbol-12-myristate-13-acetate (PMA), on the expression of Cxs and gap junction intercellular communication (GJIC) in human keratinocytes (HaCaT cells) and explored the preventive effects of green tea extracts-epicatechin (EC) and epigallocatechin-3-gallate (EGCG). We performed neutral red dye uptake assay to determine the optimal concentrations of PMA, EC, and EGCG for this study and confirmed the expression of Cx mRNAs using RT-PCR. We evaluated GJIC quantitatively using the 'scrape-loading dye transfer (SLDT)' technique after 24-h culture of HaCaT cells treated with agents. To analyze the expression change of Cxs, we also performed Western blot and immunocytochemistry. HaCaT cells were found to express Cx26, Cx30, Cx31, and Cx43, but not Cx29. In 'scrape-loading dye transfer' for functional study for GJIC, EC and EGCG significantly prevented PMA-induced down-regulation of GJIC. Western blot analyses revealed that EC and EGCG prevented down-regulation of Cx26 and Cx43 proteins in HaCaT cells treated with PMA. Immunocytochemistry showed decreased expression and abnormal location of Cx26 and Cx43 in HaCaT cells when treated with PMA, and EC and EGCG inhibited its effect. These results suggest an important role of GJIC played in carcinogenesis involving human keratinocytes and green tea as a useful anticancer diet.

HPV-induced recurrent laryngeal papillomatosis: rationale for adjuvant fatty acid therapy.

The course of human papilloma virus (HPV)-induced recurrent laryngeal papillomatosis (RLP) is variable and unpredictable. Some patients experience spontaneous remission, while others suffer from aggressive growth with dire consequences. Unfortunately, HPV DNA can persist in mucosa after treatment and can be reactivated under immunosuppressive conditions. For this reason, these benign tumors are notoriously recurrent. Better understanding of lipid-driven signaling pathways during tumorigenesis and immune responses in RLP patients can contribute to improve therapeutic approaches in an attempt to obviate this disease. Based on a mountain of evidence in the literature that concerns the immunomodulatory potential of certain FAs, it is clear that there is a rationale for adjuvant FA therapy (concurrent application) in the management of RLP. Of particular importance for immune surveillance is that the Th1 pathway in RLP is down-regulated and it is advocated that conjugated linoleic acid (CLA) and eicosapentaenoic acid (EPA) have the ability to restore the Th1/Th2 balance. Therefore, it is proposed that adjuvant FA therapy with CLA and EPA must be included in the therapeutical regime of RLP, since they are considered excellent anti-viral and anti-tumor agents to improve immune conditions and disease outcome. Immunocompetence plays a pivotal role in the clinical course of RLP and, hence, a new direction with adjuvant FA therapy may be the key to prevent recurrence of this disease.

HPV-induced recurrent laryngeal papillomatosis: fatty acid role-players.

Human papilloma virus-induced recurrent laryngeal papillomatosis is considered a troublesome and dangerous disease, because it can cause airway obstruction. Better understanding of metabolic pathways followed under pathological conditions can contribute to improved therapies by which growth and recurrence may be obviated. Part I of this study presents a clinically relevant total lipid fatty acid profile for papilloma cells, analyzed by gas liquid chromatography and a phosholipid red blood cell profile for RLP patients, analyzed by thin layer chromatography. In the papilloma cells virus interference with delta-6 and delta 5- desaturase activities is prevalent and the n-9 FA metabolic pathway is followed. It is plausible that up-regulated fatty acid synthase and 9 desaturase activities occur, since enhanced saturated fatty acids and monounsaturated fatty acid levels are also prevalent. High saturated fatty acid levels are known for their propensity to interfere with delta-6 and delta-5 desaturase activities and this is reflected in the blood profile of the RLP patients. It is also known that enhanced saturated fatty acid levels can contribute to enhanced cyclooxygenase-2 activity. Furthermore, cumulative oxidative stress with an oxidative burst is responsible for complete exhaustion of exogenous dietary arachidonic acid intake in these patients. The role of linoleic acid needs to be defined. The dietary intakes of lipids and micronutrients in RLP patients and a rationale for adjuvant FA therapy in the management of these patients are discussed in parts II and III of the study.

Food groups and laryngeal cancer risk: a case-control study from Italy and Switzerland.

Besides tobacco and alcohol, diet has been thought to be associated with laryngeal cancer risk. We thus analyzed the role of various food groups, as well as specific seasoning fats, in a case-control study conducted in Northern Italy and the Swiss Canton of Vaud from 1992 to 2000. Our study included 527 incident, histologically confirmed cases and 1,297 frequency-matched controls, selected among patients admitted to the same hospitals as cases for acute, nonneoplastic conditions, unrelated to smoking, alcohol consumption and long-term modifications of diet. The subjects' usual diet was investigated through a validated food frequency questionnaire, including 78 foods and beverages. Odds ratios (OR) and 95% confidence intervals (CI) were estimated using unconditional multiple logistic regression models. After adjustment for major confounding factors, a significant trend of increasing risk was observed for eggs (OR = 1.7 for the highest compared to the lowest quintile), red meat (OR = 3.1), processed meat (OR = 1.7), fish (OR = 1.6) and sugars (OR = 1.6). Significant inverse associations were observed for pulses (OR = 0.7), raw vegetables (OR = 0.2), cooked vegetables (OR = 0.3), citrus fruit (OR = 0.6) and other fruit (OR = 0.5). In regard to seasoning fats, a significant reduction of cancer risk was observed for olive oil (OR = 0.4) and specific seed oils (OR = 0.6), while mixed seed oils were directly associated with laryngeal cancer risk (OR = 2.2). Our study suggests that increasing vegetables and fruit, decreasing meat consumption and perhaps substituting olive oil or specific seed oils for other types of seasoning lipids might help reduce laryngeal cancer risk.

Plant foods and risk of laryngeal cancer: A case-control study in Uruguay.

In order to examine the relationships between plant foods, defined as the grouping of vegetables, fruits, tubers and legumes, with the risk of developing laryngeal cancer, a case-control study was conducted in Uruguay between 1998-1999. The study included 148 cases with histologically verified squamous cell carcinoma of the larynx, which were frequency matched on age, residence and urban/rural status with 444 hospitalized controls, afflicted by non-neoplastic conditions. Both series of patients were face-to-face interviewed in the hospitals shortly after admittance using a detailed questionnaire. This questionnaire included 62 queries on food items, representative of the usual diet of the Uruguayan population. Food items and food groups were adjusted for tobacco smoking, alcohol drinking and total energy intake. High consumption of plant foods was associated with an OR of 0.42 (95% CI 0.21-0.84). Among subgroups of plant foods, fruits and raw vegetables were associated with a strong reduction in risk (OR for the highest quartile of raw vegetables 0.29, 95% CI 0.15-0.56). Also, legumes were associated with a protective effect (OR 0.62, 95% CI 0.33-1.19). Among individual food items, tomatoes and oranges were associated with the stronger protective effects (OR for tomato intake 0.32, 95% CI 0. 17-0.58). The joint effect of heavy smoking and the low intake of vegetables and fruits displayed an increased risk of 19.2 (95% CI 5. 7-64.9).

Biochemoprevention for dysplastic lesions of the upper aerodigestive tract.

OBJECTIVES: To evaluate the efficacy and secondarily the toxic effects of biochemopreventive therapy (high-dose isotretinoin [13-cis-retinoic acid], alpha-tocopherol, and interferon alfa) in the reversal of advanced premalignant lesions of the upper aerodigestive tract and to correlate the therapeutic events with modulation of biomarkers. DESIGN: Prospective, nonrandomized chemoprevention trial. SETTING: Tertiary cancer care referral center and ambulatory care. PARTICIPANTS: Thirty-six patients with advanced premalignant lesions of the upper aerodigestive tract, without cancer during the 2 years before the intervention, with evaluable lesions, and without retinoid therapy for 3 months before the trial. INTERVENTION: Administration of oral isotretinoin (100 mg/m2 per day), oral alpha-tocopherol (1200 IU/d), and subcutaneous interferon alfa (3 megaunits per square meter twice weekly) for 12 months, with serial biopsies and clinical examination at 0, 6, 12, and 18 months from study start. MAIN OUTCOME MEASURES: Clinical and histologic responses to the intervention. RESULTS: Of the 36 patients, evaluation was possible in 30 for response at 6 months and in 21 at 12 months. At 6 months, there were 10 pathologic complete responses and 7 partial responses; at 12 months, 7 complete and 3 partial responses. A striking difference in response was observed in favor of laryngeal lesions (9/19 [47%] complete response rate at 6 months and 7/14 [50%] at 12 months vs 1/11 [9%] and 0/7 [0%], respectively, for oral lesions). Toxic effects were acceptable and did not exceed grade 3. CONCLUSION: Biochemoprevention is a promising biologic approach for laryngeal dysplasia and needs to be investigated further.

Inhibition by beta-carotene of upper respiratory tumorigenesis in hamsters receiving diethylnitrosamine followed by cigarette smoke exposure.

In recent intervention studies, beta-carotene failed to reduce or even increased the incidence of lung cancers in smokers. In the present investigation, the modifying effects of beta-carotene at various doses on the development of upper respiratory tract tumors were investigated in Syrian hamsters treated with diethylnitrosamine (DEN) and cigarette smoke. A total of 120 male 5-week-old hamsters were divided into 4 groups, each consisting of 30 animals. After a single subcutaneous (s.c.) injection of 100 mg/kg DEN, hamsters in groups 1-4 were respectively administered diets supplemented with beta-carotene at doses of 0.5%, 0.05%, 0.005% or 0% during experimental weeks 1 to 13, and simultaneously exposed to cigarette smoke. The duration of cigarette smoke exposure was 9 min twice a day, 5 days a week. Because of a marked reduction of body weight in group 1, the highest dose of beta-carotene was changed to 0.25% after 10 days. In all groups, epithelial hyperplasias and/or papillomas were induced in the larynx and trachea. However, the incidence and multiplicity of papillomas in group 1 were significantly (P < 0.05) lower than the group 4 values. Moreover, the beta-carotene treatments significantly (P < 0.05 or 0.01) reduced both the incidence and multiplicity of hyperplasias in a dose-dependent manner. The levels of retinol and beta-carotene in the serum, and the retinol level in the liver, were also elevated with dose dependence. Our results thus indicate that beta-carotene inhibits tumorigenesis, even at the high dose of 0.25%, under the present experimental conditions.

Positive impact of retinyl palmitate in leukoplakia of the larynx.

Laryngeal leukoplakia can be a premalignant precursor of squamous cell carcinoma, is often tobacco-related and can usually be readily monitored by indirect laryngoscopy. One of the main motivations for using retinyl palmitate in patients with persistent leukoplakia was to avoid general anesthesia for elderly patients, who are considered to be high-risk patients when direct larynoscopy is required for possible tissue biopsy. Our study was the first to investigate the effectiveness and toxicity of high-dose retinyl palmitate in the treatment of laryngeal leukoplakia. Treatment was divided into two phases. In the first phase, all patients underwent induction therapy with 300,000 IU/day of retinyl palmitate for the 1st week, which was then adjusted up to 1,500,000 IU/day in the 5th week in patients with resistant lesions. Patients whose lesions progressed during this period were withdrawn from the study. In the second phase, patients whose lesions responded to treatment or remained stable were assigned to a maintenance therapy of 150,000 IU/day. Complete remission was observed in 15 of 20 patients (75% of cases). Partial response was seen in the remaining 5 patients, with 3 of the patients relapsing. The median duration of treatment and follow-up was 18 months (range, 12-24 months). These results indicate that retinyl palmitate has substantial activity in laryngeal leukoplakias. Since only minor side effects were seen, the medication is an excellent candidate as a preventive agent for laryngeal cancer.