A phase I pilot study evaluating the beneficial effects of black raspberries in patients with Barrett's esophagus.

Black raspberries inhibit a broad range of cancers in preclinical models which has led to clinical evaluations targeting premalignant lesions of the colon, oral cavity and esophagus. A phase I pilot study was conducted in twenty Barrett's esophagus (BE) patients to investigate the effect of lyophilized black raspberries (LBR) on urinary metabolites and markers of lipid peroxidation, DNA damage and tissue markers of cellular proliferation, detoxification, and inflammation. Surveys, biopsies, blood and urine samples were collected before and after 6 months of LBR treatment (32 or 45 g). LBR significantly reduced urinary excretion of 8-epi-prostaglandin F2α, a marker of lipid peroxidation linked to oxidative stress and free radical damage. Urinary levels of the ellagitannin metabolites, urolithin A-glucuronide, urolithin A-sulfate and dimethylellagic acid glucuronide were significantly increased following 12 and 26 weeks of LBR consumption and may prove useful as indicators of compliance in future clinical studies. Immunohistochemical staining of BE biopsies following LBR treatment showed significant increases in mean GST-pi levels, with 55.6% of subjects responding favorably. In summary, LBR significantly decreased urinary lipid peroxidation levels and significantly increased GST-pi, a marker of detoxification, in BE epithelium. Still, LBR may need to be formulated differently, administered at higher concentrations or multiple times a day to increase efficacy.

A novel role of CYP2E1 in human megakaryocyte development.

Cytochrome P450 2E1 (CYP2E1) has active roles in bioconversion and biotransformation in humans. Although predominantly present in hepatocytes, CYP2E1 has also been found in hematopoietic stem cells and subtypes of acute myeloid leukemia with unknown clinical significance except for the metabolism of anti-fungal drugs. In the present study, we demonstrated a novel role of CYP2E1 inducing megakaryocyte development in human hematopoietic stem cells and leukemia bipotent K562 cells. CYP2E1 was induced by phorbol-12-myristate-13-acetate in dose-dependent manner in K562 cells as well as in hematopoietic stem cells by thrombopoietin, and ingenol 3,20-dibenzoate (IDB), respectively. Overexpression of CYP2E1 was positively correlated with megakaryocytes and in megakaryocyte maturation. In addition, plasmid-driven expression of CYP2E1 in K562 cells led to morphological transformation of leukemic blasts to pro- and mature megakaryocytes. In contrast, knockout of CYP2E1 by specific interfering RNA diverted these cells to erythroid differentiation. Finally, treatment of K562 cells by a free radical scavenger, N-acetyl L-lysine significantly inhibited CYP2E1 and megakaryocyte differentiation. In summary, our data demonstrated that activation of CYP2E1 and reactive oxygen species signaling promotes megakaryocyte development.

Unique in vitro and in vivo thrombopoietic activities of ingenol 3,20 dibenzoate, a Ca(++)-independent protein kinase C isoform agonist.

Thrombopoiesis following severe bone marrow injury frequently is delayed, thereby resulting in life-threatening thrombocytopenia for which there are limited treatment options. The reasons for these delays in recovery are not well understood. Protein kinase C (PKC) agonists promote megakaryocyte differentiation in leukemia cell lines and primary cells. However, little is known about the megakaryopoietic effects of PKC agonists on primary CD34+ cells grown in culture or in vivo. Here we present evidence that the novel PKC isoform-selective agonist 3,20 ingenol dibenzoate (IDB) potently stimulates early megakaryopoiesis of human CD34+ cells. In contrast, broad spectrum PKC agonists failed to do so. In vivo, a single intraperitoneal injection of IDB selectively increased platelets in mice without affecting hemoglobin or white counts. Finally, IDB strongly mitigated radiation-induced thrombocytopenia, even when administered 24 hours after irradiation. Our data demonstrate that novel PKC isoform agonists such as IDB may represent a unique therapeutic strategy for accelerating the recovery of platelet counts following severe marrow injury.

Cranberry proanthocyanidins mediate growth arrest of lung cancer cells through modulation of gene expression and rapid induction of apoptosis.

Cranberries are rich in bioactive constituents purported to enhance immune function, improve urinary tract health, reduce cardiovascular disease and more recently, inhibit cancer in preclinical models. However, identification of the cranberry constituents with the strongest cancer inhibitory potential and the mechanism associated with cancer inhibition by cranberries remains to be elucidated. This study investigated the ability of a proanthocyanidin rich cranberry fraction (PAC) to alter gene expression, induce apoptosis and impact the cell cycle machinery of human NCI-H460 lung cancer cells. Lung cancer is the leading cause of cancer-related deaths in the United States and five year survival rates remain poor at 16%. Thus, assessing potential inhibitors of lung cancer-linked signaling pathways is an active area of investigation.

Cranberry proanthocyanidins induce apoptosis and inhibit acid-induced proliferation of human esophageal adenocarcinoma cells.

The occurrence of esophageal adenocarcinoma and its only recognized precursor lesion, Barrett's esophagus, has rapidly increased during the past three decades. The precise reason for the rise remains to be elucidated, but increasing rates have been linked to multiple nutritional factors. Plant-based diets have generally been associated with a reduction of risk for esophageal adenocarcinoma and those of animal origin with risk escalation. Moreover, a number of recent in vitro and limited in vivo investigations have reported that cranberry extracts affect multiple cancer-associated processes in breast, colon, prostate, and other cancer cell lines of epithelial origin. Thus, this study sought to investigate the chemopreventive potential of a cranberry proanthocyanidin rich extract (PAC) in SEG-1 human esophageal adenocarcinoma (EAC) cells. PAC pretreatment significantly inhibited the viability and proliferation of EAC cells in a time- and dose-dependent manner. Moreover, PAC (50 microg/mL) significantly inhibited acid-induced cell proliferation of SEG-1 cells. PAC treatment induced cell cycle arrest at the G1 checkpoint and significantly reduced the percentage of SEG-1 cells in S-phase following 24 and 48 h of exposure. PAC treatment also resulted in significant induction of apoptosis. Thus, PAC modulates cell cycle regulation, aberrant proliferation, and apoptosis, all key biological processes altered during progression to esophageal adenocarcinoma. These findings support that further mechanistic studies are warranted to more fully elucidate the inhibitory potential of PAC against esophageal cancer.

In situ assessment of intraepithelial neoplasia in hamster trachea epithelium using angle-resolved low-coherence interferometry.

Optical spectroscopy was used to evaluate the transformation of nuclear morphology associated with intraepithelial neoplasia in an animal model of carcinogenesis. In this pilot study, we have assessed the capability of angle-resolved low-coherence interferometry (a/LCI) to monitor in situ the neoplastic progression of hamster trachea epithelial tissue. By using the depth resolution made possible by coherence gating, the a/LCI system has been adapted to the unique geometry of the hamster trachea to allow us to extract useful nuclear morphometric information from cells in the epithelial layer without the need for exogenous staining or tissue fixation. Analysis of a/LCI nuclear morphology measurements has identified two important biomarkers of neoplastic transformation in hamster trachea epithelium, the size and the refractive index of epithelial cell nuclei. By comparing the a/LCI measurements of these two biomarkers to pathologic classification, we distinguished nuclear morphology changes for normal tissue, low-grade dysplasia, and high-grade dysplasia. Given its previous usefulness for tracking neoplastic change through nuclear morphometry measurements, the a/LCI technique may prove to be a useful tool in evaluating chemopreventive agents in future studies of hamster trachea epithelium.

Transitioning from preclinical to clinical chemopreventive assessments of lyophilized black raspberries: interim results show berries modulate markers of oxidative stress in Barrett's esophagus patients.

Increased fruit and vegetable consumption is associated with decreased risk of a number of cancers of epithelial origin, including esophageal cancer. Dietary administration of lyophilized black raspberries (LBRs) has significantly inhibited chemically induced oral, esophageal, and colon carcinogenesis in animal models. Likewise, berry extracts added to cell cultures significantly inhibited cancer-associated processes. Positive results in preclinical studies have supported further investigation of berries and berry extracts in high-risk human cohorts, including patients with existing premalignancy or patients at risk for cancer recurrence. We are currently conducting a 6-mo chemopreventive pilot study administering 32 or 45 g (female and male, respectively) of LBRs to patients with Barrett's esophagus (BE), a premalignant esophageal condition in which the normal stratified squamous epithelium changes to a metaplastic columnar-lined epithelium. BE's importance lies in the fact that it confers a 30- to 40-fold increased risk for the development of esophageal adenocarcinoma, a rapidly increasing and extremely deadly malignancy. This is a report on interim findings from 10 patients. To date, the results support that daily consumption of LBRs promotes reductions in the urinary excretion of two markers of oxidative stress, 8-epi-prostaglandin F2alpha (8-Iso-PGF2) and, to a lesser more-variable extent, 8-hydroxy-2'-deoxyguanosine (8-OHdG), among patients with BE.