Black Raspberries Research Articles

EP03.01-014 Results From the BEWELL Study: Black Raspberry Nectar for the Prevention of Lung Cancer (NCT04267874)

Lung cancer is a leading cause of cancer deaths. Therefore, prevention strategies are greatly needed, especially in high-risk populations. Dietary interventions can affect disease risk via changes to the microbiome. Black raspberries are a rich source of polyphenols which have attenuated inflammation in preclinical and clinical studies. The BEWELL Study is a placebo-controlled, randomized, cross-over pilot trial that examined the impact of a phytochemical-rich black raspberry (BRB) nectar on markers of chronic inflammation and the microbiome (Trials in Progress, WCLC 2021).

Modification of Diet to Reduce the Stemness and Tumorigenicity of Murine and Human Intestinal Cells.

ScopeBlack raspberries (BRBs) have colorectal cancer (CRC) chemo‐preventative effects. As CRC originates from an intestinal stem cell (ISC) this study has investigated the impact of BRBs on normal and mutant ISCs.Methods and resultsMice with an inducible Apcfl mutation in either the ISC (Lgr5CreERT2 ) or intestinal crypt (AhCre/VillinCreERT2 ) are fed a control or 10% BRB‐supplemented diet. This study uses immunohistochemistry, gene expression analysis, and organoid culture to evaluate the effect of BRBs on intestinal homeostasis. RNAscope is performed for ISC markers on CRC adjacent normal colonic tissue pre and post BRB intervention from patients. 10% BRB diet has no overt effect on murine intestinal homeostasis, despite a reduced stem cell number. Following Apc ISC deletion, BRB diet extends lifespan and reduces tumor area. In the AhCre model, BRB diet attenuates the “crypt‐progenitor” phenotype and reduces ISC marker gene expression. In ex vivo culture BRBs reduce the self‐renewal capacity of murine and human Apc deficient organoids. Finally, the study observes a reduction in ISC marker gene expression in adjacent normal crypts following introduction of BRBs to the human bowel.ConclusionBRBs play a role in CRC chemoprevention by protectively regulating the ISC compartment and further supports the use of BRBs in CRC prevention.

Black raspberry extract inhibits regulatory T-cell activity in a murine model of head and neck squamous cell carcinoma chemoprevention.

Head and neck squamous cell carcinomas (HNSCC) are one of the most diagnosed malignancies globally, with a 5-year survival rate of approximately 40% to 50%. Current therapies are limited to highly invasive surgery, aggressive radiation, and chemotherapies. Recent reports have demonstrated the potential phytochemical properties of black raspberries in inhibiting the progression of various cancers including HNSCCs. However, the effects of black raspberry extracts on immune cells of the tumor microenvironment, specifically regulatory T cells during HNSCC, have not been investigated. We used a mouse model of 4-nitroquinoline-1-oxide (4NQO) chemically induced HNSCC carcinogenesis to determine these effects. C57BL/6 mice were exposed to 4NQO for 16 weeks and regular water for 8 weeks. 4NQO-exposed mice were fed the AIN-76A control mouse diet or the AIN76 diet supplemented with black raspberry extract. At terminal sacrifice, tumor burdens and immune cell recruitment and activity were analyzed in the tumor microenvironment, draining lymph nodes, and spleens. Mice fed the BRB extract-supplemented diet displayed decreased tumor burden compared to mice provided the AIN-76A control diet. Black raspberry extract administration did not affect overall T-cell populations as well as Th1, Th2, or Th17 differentiation in spleens and tumor draining lymph nodes. However, dietary black raspberry extract administration inhibited regulatory T-cell recruitment to HNSCC tumor sites. This was associated with an increased cytotoxic immune response in the tumor microenvironment characterized by increased CD8+ T cells and enhanced Granzyme B production during BRB extract-mediated HNSCC chemoprevention. Interestingly, this enhanced CD8+ T-cell antitumoral response was localized at the tumor sites but not at spleens and draining lymph nodes. Furthermore, we found decreased levels of PD-L1 expression by myeloid populations in draining lymph nodes of black raspberry-administered carcinogen-induced mice. Taken together, our findings demonstrate that black raspberry extract inhibits regulatory T-cell recruitment and promotes cytotoxic CD8 T-cell activity at tumor sites during HNSCC chemoprevention. These results demonstrate the immunomodulatory potential of black raspberry extracts and support the use of black raspberry-derived phytochemicals as a complementary approach to HNSCC chemoprevention and treatment.

Effect of High Hydrostatic Pressure and Pulsed Electric Fields Processes on Microbial Safety and Quality of Black/Red Raspberry Juice.

Black and red raspberries are fruits with a high phenolic and vitamin C content but are highly susceptible to deterioration. The effect of high hydrostatic pressure (HHP 400–600 MPa/CUT-10 min) and pulsed electric fields (PEF, frequency 100–500 Hz, pulse number 100, electric field strength from 11.3 to 23.3 kV/cm, and specific energy from 19.7 to 168.4 kJ/L) processes on black/red raspberry juice was studied. The effect on the inactivation of microorganisms and pectin methylesterase (PME) activity, physicochemical parameters (pH, acidity, total soluble solids (°Brix), and water activity (aw)), vitamin C and phenolic compounds content were also determined. Results reveal that all HHP-treatments produced the highest (p < 0.05) log-reduction of molds (log 1.85 to 3.72), and yeast (log 3.19), in comparison with PEF-treatments. Increments in pH, acidity, and TSS values attributed to compounds’ decompartmentalization were found. PME activity was partially inactivated by HHP-treatment at 600 MPa/10 min (22% of inactivation) and PEF-treatment at 200 Hz/168.4 kJ/L (19% of inactivation). Increment in vitamin C and TPC was also observed. The highest increment in TPC (79% of increment) and vitamin C (77% of increment) was observed with PEF at 200 Hz/168.4 kJ/L. The putative effect of HHP and PEF on microbial safety, enzyme inactivation, and phytochemical retention is also discussed in detail. In conclusion, HHP and PEF improve phytochemical compounds’ content, microbial safety, and quality of black/red raspberry juice.

Influența fructelor asupra proprietăților organoleptice ale iaurtului din lapte de capră și vacă

The objectives of this study were to analyze the yogurt processed from a mixture of goat and cow milk (series I) and goat milk (series II) with the addition of fruit puree of black chokeberries (Aronia melanocarpa), raspberries (Rubus idaeus) and strawberries (Fragaria ananassa). From the physico-chemical point of view, sugar content in the goat milk yogurt with strawberies had the lowest values (2.79±0.01 %). At storage, the best evolution of syneresis (1.05 ± 0.04-2.01 ± 0.05 ml/100 ml) and water holding capacity (66.6 ± 0.02-56.1 ± 0.01%) was observed in the mixture of goat/cow milk yogurt with strawberries. From a chromatic point of view, color characteristics with the highest values (85.31 ± 2.75-83.00 ± 3.54) were noticed in the goat/cow milk yogurt with black chokeberries. From an organoleptic point of view, goat/cow milk yogurt with strawberries obtained and kept its highest score, having a refreshing aroma, smooth and viscous gel and a sweet taste.

A Pilot Clinical Study to Investigate the Hypomethylating Properties of Freeze-dried Black Raspberries in Patients with Myelodysplastic Syndrome or Myeloproliferative Neoplasm.

Myelodysplastic syndromes (MDS) and myelodysplastic/myeloproliferative neoplasms (MDS/MPN) are bone marrow disorders characterized by cytopenias and progression to acute myeloid leukemia. Hypomethylating agents (HMAs) are Food and Drug Administration-approved therapies for MDS and MDS/MPN patients. HMAs have improved patients’ survival and quality of life when compared with other therapies. Although HMAs are effective in MDS and MDS/MPN patients, they are associated with significant toxicities that place a large burden on patients. Our goal is to develop a safer and more effective HMA from natural products. We previously reported that black raspberries (BRBs) have hypomethylating effects in the colon, blood, spleen, and bone marrow of mice. In addition, BRBs exert hypomethylating effects in patients with colorectal cancer and familial adenomatous polyposis. In the current study, we conducted a pilot clinical trial to evaluate the hypomethylating effects of BRBs in patients with low-risk MDS or MDS/MPN. Peripheral blood mononuclear cells (PBMCs) were isolated before and after three months of BRB intervention. CD45+ cells were isolated from PBMCs for methylation analysis using a reduced-representation bisulfite sequencing assay. Each patient served as their own matched control, with their measurements assessed before intervention providing a baseline for post-intervention results. Clinically, our data showed that BRBs were well-tolerated with no side effects. When methylation data was combined, BRBs significantly affected methylation levels of 477 promoter regions. Pathway analysis suggests that BRB-induced intragenic hypomethylation drives leukocyte differentiation. A randomized, placebo-controlled clinical trial of BRB use in low-risk MDS or MDS/MPN patients is warranted.

EFFECT OF PROCESSING METHODS AND STORAGE TIME ON THE CONTENT OF ASCORBIC ACID IN BERRIES

The article presents the results of a study of the dynamics of the content of ascorbic acid in berries of black and red currants and raspberries during storage in a frozen form and during conservation of berries grated with sugar. As objects of research, we used two varieties of red currant, black currant and raspberry. The ascorbic acid content served as a marker of the berry quality during storage. The analysis of ascorbic acid was carried out by the iodometric method according to the standard method. It was found that immediately after freezing, the loss of ascorbic acid is from 6 to 20%, which is significantly less than when grinding with sugar (65–97%). The decrease in the content of ascorbic acid in berries during storage occurs unevenly and after 6 months the concentration stabilizes. When storing frozen berries, additional losses of vitamin C in red and black currants averaged 83%, and in raspberries – 66%. During storage of berry puree, additional losses were 32% for red and black currants, and 89% for raspberries. The best preservation of ascorbic acid was observed when stored frozen – in experiments with black currants, while storing berry puree – in experiments with red currants.

Chemical Composition of Volatile Extracts From Blackberries, Black Raspberries, and Blueberries and Their Apoptotic Effect on A549 Non-Small-Cell Lung Cancer Cells

ObjectivesBerry volatiles are responsible for the flavor and aroma of berries. Berry volatiles has been recently reported to have an anti-inflammatory effect. However, there is still limited information available on the health-promoting activities of berry volatiles. The objectives of this study were to evaluate the chemical composition of volatile extracts from blackberries, black raspberries, and blueberries and to investigate the anti-proliferative effect of three berry volatile extracts on A549 non-small-cell lung cancer cells and their mechanism. MethodsThe chemical composition of berry volatile extracts (BVEs) from blackberries (BK), black raspberries (BR), and blueberries (BU) were identified by using gas chromatography-mass spectrometry. A549 non-small cell lung cancer cells were treated with different dilutions (2-, 4-, and 8-fold dilutions) of three berry volatile extracts for 12, 24, and 48 h. The proliferation of A549 was measured by using an MTS assay. The induction of apoptosis and changes in the cell cycle by berry volatile extracts were analyzed by using a cell death detection ELISA kit and flow cytometry. All statistical tests were analyzed by using one-way ANOVA, followed by Tukey’s multiple comparisons. The significant difference was defined at P < 0.05. ResultsTotal volatiles in BK, BR, and BU volatile extracts were 1588.5 ± 170.9,3184.9 ± 75.3, and 3107.5 ± 256.4 μg/L, respectively. Among them, monoterpene was the most abundant compound in all three BVEs. All BVEs with two-fold dilution significantly induced apoptosis after 24 h (p < 0.05) and inhibited the proliferation of A549 after 48 h treatment (p < 0.05). In the cell cycle, the proportion of cells in G0/G1 interphase increased with two-fold diluted BVE treatments (64.9–73.1%) compared to control (60.3%) after 48 h. ConclusionsThese findings showed that berry volatile extracts from blackberries, black raspberries, and blueberries have the anti-proliferative effect on non-small lung cancer cells through apoptosis and possibly cell cycle arrest. This suggests that berry volatiles from blackberry, black raspberry, and blueberry may have a potential impact on lung cancer. Funding SourcesArkansas Biosciences Institute.

Protocatechuic Acid, a Gut Bacterial Metabolite of Black Raspberries, Inhibits Adenoma Development and Alters Gut Microbiome Profiles in Apc Min/+ Mice.

Administration of black raspberries (BRBs) and their anthocyanin metabolites, including protocatechuic acid (PCA), has been demonstrated to exert chemopreventive effects against colorectal cancer through alteration of innate immune cell trafficking, modulation of metabolic and inflammatory pathways, etc. Previous research has shown that the gut microbiome is important in the effectiveness of chemoprevention of colorectal cancer. This study aimed to assess the potency of PCA versus BRB dietary administration for colorectal cancer prevention using an ApcMin/+ mouse model and determine how bacterial profiles change in response to PCA and BRBs. A control AIN-76A diet supplemented with 5% BRBs, 500 ppm PCA, or 1,000 ppm PCA was administered to ApcMin/+ mice. Changes in incidence, polyp number, and polyp size regarding adenomas of the small intestine and colon were assessed after completion of the diet regimen. There were significant decreases in adenoma development by dietary administration of PCA and BRBs in the small intestine and the 5% BRB-supplemented diet in the colon. Pro-inflammatory bacterial profiles were replaced with anti-inflammatory bacteria in all treatments, with the greatest effects in the 5% BRB and 500 ppm PCA-supplemented diets accompanied by decreased COX-2 and prostaglandin E2 levels in colonic mucosa. We further showed that 500 ppm PCA, but not 1,000 ppm PCA, increased IFN-γ and SMAD4 levels in primary cultured human natural killer cells. These results suggest that both BRBs and a lower dose PCA can benefit colorectal cancer patients by inhibiting the growth and proliferation of adenomas and promoting a more favorable gut microbiome condition.

The New Face of Berries: A Review of Their Antiviral Proprieties.

Due to rising consumer preference for natural remedies, the search for natural antiviral agents has accelerated considerably in recent years. Among the natural sources of compounds with potential antiviral proprieties, berries are interesting candidates, due to their association with health-promoting properties, including antioxidant, antimutagenic, anticancer, antimicrobial, anti-inflammatory, and neuroprotective properties. The past two decades have witnessed a flurry of new findings. Studies suggest promising antiviral proprieties against enveloped and non-enveloped viruses, particularly of cranberries, blueberries, blackcurrants, black raspberries, and pomegranates. The aim of this review is to assemble these findings, to list the implied mechanisms of action, and thereby point out promising subjects for research in this field, in the hope that compounds obtainable from natural sources such as berries may be used someday to treat, or even prevent, viral infections.

Potential of dietary supplementation with berries to enhance immunity in humans

One of the most prominent concerns that has arisen from global events such as the Coronavirus Disease 2019 (COVID-19) pandemic is how the immune system might be enhanced to provide greater protection against malignant conditions and diseases. Dietary modification is one of the major fields of research, with special focus on how diet interacts with immunity through impacts on factors such as the gut microbiome, inflammation, and nutritional imbalance. This review focuses on current research regarding the use of bioactives derived from berries as a dietary supplement to improve immunity, with discussions of relevant clinical studies. Major bioactive, metabolic compounds of focus- flavonoids, anthocyanins, alkaloids, dietary fiber, and stilbenes- have demonstrated biochemical merits in modulating immunity. In addition, blueberries, goji berries, black raspberries, and cranberries, which have been extensively researched and recently gained interest for their effects on the immune system in animal and cell models, may also hold promise in providing similar benefits to humans, though the precise immunological effects have yet to be clearly determined. Overall, the field of berry research as it relates to diet and immunity shows potential, but more clinical studies will be necessary for a full understanding of the mechanisms of berry immunomodulation.

Dietary supplementation with black raspberries prolongs survival in ApcMin/+ mice.

A diet supplemented with freeze-dried black raspberries (BRBs) has been demonstrated to modulate various biochemical and physiological pathways in both colorectal cancer (CRC) patients and ApcMin/+ mice, which are utilized to model CRC. These changes have been previously shown to exert beneficial chemopreventive effects against CRC, with outcomes such as reduction of adenoma development and inflammation. This study aimed to assess whether these effects manifest in a meaningful change in survival rates by comparing these rates between ApcMin/+ mice administered a 5% BRB-supplemented diet or a control AIN-76A diet. Percent survival over days elapsed was assessed in order to determine a median length of survival for each group of mice. Significant increases in survival rates with consumption of the BRB diet versus the control diet were demonstrated in both male and female mouse study groups. Male and female control groups were also compared in order to reduce confounding due to the sex of the mice; the difference in survival rates between male and female mice was not significant (p = 0.07, *p < 0.05), as male mice lived for a median of 143 days and females for 194 days. The results of this study suggest that administration of a BRB-supplemented diet may potentially prolong the lifespan and increase survival rates of colorectal cancer patients.

A High-Throughput Metabolic Microarray Assay Reveals Antibacterial Effects of Black and Red Raspberries and Blackberries against Helicobacter pylori Infection.

Helicobacter pylori infection is commonly treated with a combination of antibiotics and proton pump inhibitors. However, since H. pylori is becoming increasingly resistant to standard antibiotic regimens, novel treatment strategies are needed. Previous studies have demonstrated that black and red berries may have antibacterial properties. Therefore, we analyzed the antibacterial effects of black and red raspberries and blackberries on H. pylori. Freeze-dried powders and organic extracts from black and red raspberries and blackberries were prepared, and high-performance liquid chromatography was used to measure the concentrations of anthocyanins, which are considered the major active ingredients. To monitor antibiotic effects of the berry preparations on H. pylori, a high-throughput metabolic growth assay based on the Biolog system was developed and validated with the antibiotic metronidazole. Biocompatibility was analyzed using human gastric organoids. All berry preparations tested had significant bactericidal effects in vitro, with MIC90 values ranging from 0.49 to 4.17%. Antimicrobial activity was higher for extracts than powders and appeared to be independent of the anthocyanin concentration. Importantly, human gastric epithelial cell viability was not negatively impacted by black raspberry extract applied at the concentration required for complete bacterial growth inhibition. Our data suggest that black and red raspberry and blackberry extracts may have potential applications in the treatment and prevention of H. pylori infection but differ widely in their MICs. Moreover, we demonstrate that the Biolog metabolic assay is suitable for high-throughput antimicrobial susceptibility screening of H. pylori.

Review of Functional and Pharmacological Activities of Berries.

Functional plant-based foods (such as fruits, vegetables, and berries) can improve health, have a preventive effect, and diminish the risk of different chronic diseases during in vivo and in vitro studies. Berries contain many phytochemicals, fibers, vitamins, and minerals. The primary phytochemicals in berry fruits are phenolic compounds including flavonoids (anthocyanins, flavonols, flavones, flavanols, flavanones, and isoflavonoids), tannins, and phenolic acids. Since berries have a high concentration of polyphenols, it is possible to use them for treating various diseases pharmacologically by acting on oxidative stress and inflammation, which are often the leading causes of diabetes, neurological, cardiovascular diseases, and cancer. This review examines commonly consumed berries: blackberries, blackcurrants, blueberries, cranberries, raspberries, black raspberries, and strawberries and their polyphenols as potential medicinal foods (due to the presence of pharmacologically active compounds) in the treatment of diabetes, cardiovascular problems, and other diseases. Moreover, much attention is paid to the bioavailability of active berry components. Hence, this comprehensive review shows that berries and their bioactive compounds possess medicinal properties and have therapeutic potential. Nevertheless, future clinical trials are required to study and improve the bioavailability of berries’ phenolic compounds and extend the evidence that the active compounds of berries can be used as medicinal foods against various diseases.

Black Raspberry Supplementation Alters the Gut Microbiome and Improves Alpha Diversity in a Mouse Model of Inflammation-Associated Colorectal Cancer

ObjectivesAnti-inflammatory bioactives in black raspberries (BRB) have been shown to have protective effects on the colon epithelium and may influence gut microbiome. The goal of this study was to determine the effects of dietary intervention with BRB on the dynamic composition of the gut microbiome composition in mice. MethodsUsing a 2 × 2 factorial design, C57BL/6J male mice were fed the standard AIN93G diet or the total Western diet (TWD) for 16 weeks with or without 10% (w/w) whole, freeze-dried BRB powder. The azoxymethane + dextran sodium sulfate model of inflammation-associated colorectal cancer was employed to assess the dynamic response of the gut microbiome to basal diet and BRB treatment prior to, during, and after active colitis and at the study end. Microbiome composition was determined using 16s rRNA sequencing followed by diversity analyses (alpha and beta) and identification of discriminating taxa by with linear discriminant analyses by effect size (lefse). ResultsAlpha diversity was markedly reduced during colitis for mice consuming either AIN93G or TWD, with some improvement noted by the recovery phase. Of note, consumption of BRB for two weeks significantly increased alpha diversity measures, and BRB improved alpha diversity in mice fed the AIN93G diet during colitis. Alternatively, BRB appeared less effective in mice fed TWD. Beta diversity was also significantly affected with notable clustering of microbiomes by BRB treatment during and after colitis. Consumption of BRB affected the relative abundance of several key taxa over the course of colitis and recovery from gut injury, including Erysipelotrichaceae, Bifidobacteriaceae, Streptococcaceae, Rikenellaceae, Ruminococcaceae and Akkermansiaceae, among others. ConclusionsDietary supplementation with BRB shifted the composition of the gut microbiome during colitis and recovery from gut injury, though the effects were inconsistent with respect to the basal diet consumed. Funding SourcesUSDA NIFA AFRI grant no. 2018-67017-27,516 and 2014-67017-21755.

Comparison of anthocyanins, polyphenols, and antioxidant capacities among raspberry, blackberry, and Japanese wild Rubus species

The breeding of raspberries (Rubus idaeus L.) and blackberries (R. fruticosus Agg.) has shifted in focus toward traits associated with the sensory qualities and potential health benefits of the fruit, and Rubus wild species have attracted a great deal of attention as new sources of genetic diversity. In this study, the anthocyanin and polyphenol contents and compositions, and three types of antioxidant capacities were compared in mature fruits including raspberries, blackberries, and Japanese wild Rubus species. ‘Blackcap’ black raspberry showed the significantly highest values of total anthocyanin content and total polyphenol content among 12 Rubus species or cultivars. Cyanidin-3-xylosyl-rutinoside was contained only in black raspberries, and was characteristic anthocyanin in Rubus species. R. croceacanthus H. Lév., a Japanese wild species that is adapted to warm regions, had a deep red fruit color, and had the highest total anthocyanin and total polyphenol contents among the four Japanese wild species examined. As for the antioxidant capacities, the highest values were observed in ‘Blackcap’ among 12 Rubus species. Furthermore, a high positive correlation was shown between total anthocyanin or total polyphenol content and antioxidant capacities. In particular, black raspberries and blackberries contained high concentrations of rutin. Thus, rutin might contribute to the high antioxidant capacities in these cultivars.

P40.03 Report on a Phytochemical-rich Dietary Intervention Trial to Prevent Lung Cancer: Implementation in a High-Risk Lung Screening Clinic

The BeWell Trial, Black Raspberry Beverage for the Prevention of Lung Cancer, is currently underway at The Ohio State University (OSU) James Cancer Hospital. The study is a placebo-controlled, randomized, cross-over pilot study to assess the impact of a novel black raspberry food product on the microbiome and immune functions in a high risk cohort for lung cancer. Our central hypothesis is that phytochemical–rich black raspberries (BRB) will increase the diversity of the gut microbiome, increase the abundance of specific desirable microbes, such as Akkermansia muciniphila, and inhibit cancer by indirect effects on host inflammatory cascades, perhaps via impact on the microbiota. This report will describe the trial progress, including challenges in accrual/retention, compliance, safety, and additional insights on conducting BRB and cancer trials. BeWell Study participants are recruited from the OSU Lung Screening Clinic, pulmonary clinics, and social media (e.g. Facebook). Inclusion criteria include: age 55-77 years, 30+ pack years smoking history, and smoked within the last 15 years. Exclusion criteria include: oral corticosteroid use and uncontrolled gastrointestinal or metabolic disorders. Participants are asked to follow a low-phenolic diet and we obtain a 24-hour urine sample, stool sample, blood draw and weight/vitals at each clinic visit. They are randomized to receive the BRB or placebo beverage, (blocked by gender and smoking status) for 4-weeks (170 ml, 2X/day), have a 2-week washout period, and then cross-over to 4 weeks of BRB or placebo (10 weeks total). The crossover design greatly minimizes variation, which is particularly beneficial in microbiome studies. Participants also complete diet compliance, medical history, food frequency and physical activity questionnaires. Of the accrual target of 42 patients, 32 have been fully consented, and 2 have fully completed the 10-week study. Thirteen individuals are former smokers (6 females and 7 males), with 7 current smokers (all female). Their ages range from 58 to 76 years. The dropout rate is 37%. Participants mostly dropped out before the first study visit. Nearly half of dropouts occurred during the holidays. We have approached 244 eligible participants who declined (13% consent rate). The reasons given for non-participation include: not interested, too busy, hard to get to appointments, dislike home urine/stool tests, live too far and work conflicts. One adverse event was reported event (diarrhea). Current accrual rates suggest study completion in December, 2020. The accrual and retention process for this study of high risk smokers has been challenging, with 13% of individuals agreeing to enroll and high dropout rate (37%). As a result, we extended our estimated recruiting period from 6 to 12 months, increased the compensation level and increased the number of targeted advertisements on social media, with A/B testing to optimize text and images. Objective measures of various recruitment and advertising success are being captured. We suggest that future trials with similar groups consider time of year and season when recruiting, ensure adequate reimbursement and offer any possible incentives or conveniences to increase participation and retention, thus increasing the success of similar future trials.

Evaluation of Anti-cancer Effect of Different Concentrations of Black Raspberries Extract on Oral Squamous Cell Carcinoma Cell Line (In Vitro Study)

Background: Black Raspberry has a wide range of pharmacological and therapeutic aspects including anti-inflammatory and antioxidant activities. Moreover, the effect of Black Raspberry has been studied in many cancers such as human cervical, esophageal and colorectal cancer and showed cytotoxic and anti-proliferative effects against these types of cancer. Aim: To identify the possible anti-cancer effect of Black Raspberry on Oral Squamous Cell Carcinoma cell line. Material and methods: Human tongue squamous cell carcinoma cell line was divided into 3 groups: one control group, second group treated with Black Raspberry, and the third group treated with Doxorubicin, each of the two treatments was applied for 24, 48, 72 hours, respectively. Then, cellular viability was measured using Microculture Tetrazolium assay, cell cycle analysis was done using Flow Cytometry and eventually, apoptotic activity was evaluated using Enzyme-linked Immunosorbent assay. Results: Black Raspberry succeeded to decrease the percentage of viable and proliferating cells with increasing time. On the other hand, it increased the percentage of apoptotic cells and levels of caspase-3. Conclusion: Black Raspberry has a potential cytotoxic effect on oral squamous cell carcinoma cell line in a dose and time dependent manner, and exerts this action through induction of apoptosis, and its action is comparable to Doxorubicin action.

Black Raspberries Suppress Colorectal Cancer by Enhancing Smad4 Expression in Colonic Epithelium and Natural Killer Cells.

Innate immune cells in the tumor microenvironment have been proposed to control the transition from benign to malignant stages. In many cancers, increased infiltration of natural killer (NK) cells associates with good prognosis. Although the mechanisms that enable NK cells to restrain colorectal cancer (CRC) are unclear, the current study suggests the involvement of Smad4. We found suppressed Smad4 expression in circulating NK cells of untreated metastatic CRC patients. Moreover, NK cell-specific Smad4 deletion promoted colon adenomas in DSS-treated ApcMin/+ mice and adenocarcinomas in AOM/DSS-treated mice. Other studies have shown that Smad4 loss or weak expression in colonic epithelium associates with poor survival in CRC patients. Therefore, targeting Smad4 in both colonic epithelium and NK cells could provide an excellent opportunity to manage CRC. Toward this end, we showed that dietary intervention with black raspberries (BRBs) increased Smad4 expression in colonic epithelium in patients with FAP or CRC and in the two CRC mouse models. Also, benzoate metabolites of BRBs, such as hippurate, upregulated Smad4 and Gzmb expression that might enhance the cytotoxicity of primary human NK cells. Of note, increased levels of hippurate is a metabolomic marker of a healthy gut microbiota in humans, and hippurate also has antitumor effects. In conclusion, our study suggests a new mechanism for the action of benzoate metabolites derived from plant-based foods. This mechanism could be exploited clinically to upregulate Smad4 in colonic epithelium and NK cells, thereby delaying CRC progression.

Mapping of the Waxy Bloom Gene in ‘Black Jewel’ in a Parental Linkage Map of ‘Black Jewel’ × ‘Glen Ample’ (Rubus) Interspecific Population

Black and red raspberries (Rubus occidentalis L. and Rubus idaeus L.) are the prominent members of the genus Rubus (Rosaceae family). Breeding programs coupled with the low costs of high-throughput sequencing have led to a reservoir of data that have improved our understanding of various characteristics of Rubus and facilitated the mapping of different traits. Gene B controls the waxy bloom, a clearly visible epicuticular wax on canes. The potential effects of this trait on resistance/susceptibility to cane diseases in conjunction with other morphological factors are not fully studied. Previous studies suggested that gene H, which controls cane pubescence, is closely associated with gene B. Here, we used tunable genotyping-by-sequencing technology to identify the de novo SNPs of R. occidentalis and R. idaeus using an interspecific population that segregates for the waxy bloom phenotype. We created linkage maps of both species and mapped the identified SNPs to the seven chromosomes (Ro01–Ro07) of Rubus. Importantly, we report, for the first time, the mapping of gene B to chromosome 2 of R. occidentalis using a genetic map consisting of 443 markers spanning 479.76 cM. We observed the poor transferability of R. idaeus SSRs to R. occidentalis and discrepancies in their previously reported chromosome locations.