Publications by authors named "Ajay P Singh"

93 Publications

The prevalence and clinical relevance of 2R/2R TYMS genotype in patients with gastrointestinal malignancies treated with fluoropyrimidine-based chemotherapy regimens.

Pharmacogenomics J 2021 Feb 19. Epub 2021 Feb 19.

Department of Pathology, The University of South Alabama, Mobile, AL, USA.

Introduction: The prevalence of 2R/2R TYMS genotype is variable but estimated to be around 20-30% in Caucasians. The clinical relevance of TYMS 2R/2R genotype in predicting severe fluoropyrimidine-related adverse events (FrAE) is controversial. Here, we explored the prevalence and clinical relevance of 2R/2R TYMS genotype.

Methods: Between 2011 and 2018, 126 patients were genotyped for TYMS. FrAEs were graded according to CTCAE version 5.0. Fisher's exact test was used for statistical analysis.

Results: The prevalence of TYMS 2R/2R genotype was 24.6%. Among patients with TYMS genotypes (N = 71) that predict decreased TS expression, 2R/2R TYMS genotype was the most common TYMS genotype seen in female (57%) and African American (60%) patients. Among patients with genotypes that predict increased TS expression (N = 55), 12 patients had grade 3-4 FrAEs (22%), while among patients with genotypes that predict decreased TS expression (N = 71), 30 patients had grade 3-4 FrAEs (42%) (p = 0.0219). Compared to patients with genotypes predicting increased TS expression, 17 out of 31 patients (55%) with TYMS 2R/2R genotype had grade 3-4 FrAEs (p = 0.0039) and 15 out 40 patients (38%) with TYMS 2R/3RC and TYMS 3RC/3RC genotype had grade 3-4 FrAEs (p = 0.1108).

Conclusion: The prevalence of TYMS 2R/2R genotype was 24.6%, and it had a unique sex and ethnic distribution. Polymorphism in the promoter region of TYMS gene that predicts decreased TS expression due to 2R/2R variant was associated with grade 3-4 FrAEs. These data suggest that genotyping patients who are not DPD deficient for TYMS might identify patients at risk of severe FrAEs.
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http://dx.doi.org/10.1038/s41397-021-00210-2DOI Listing
February 2021

Characterization of Patients with COVID-19 Admitted to a Community Hospital of East Harlem in New York City.

Cureus 2020 Aug 18;12(8):e9836. Epub 2020 Aug 18.

Department of Medicine, NYC Health + Hospitals / Metropolitan, New York, USA.

Background New York City was the epicenter for the coronavirus disease 2019 (COVID-19) in the United States. Accordingly, the aim of this study was to characterize the population of patients admitted with this condition to a community hospital in East Harlem located in the northeast part of the city. Methods A retrospective review of medical records of patients at least 18 years of age, admitted to the hospital with COVID-19 disease from March 14 to April 30 of 2020. Results Three hundred and seventy-one patients were identified. The majority was comprised of men. Obesity, hypertension, and hyperlipidemia were the most prevalent comorbidities. Most patients were treated with a combination of hydroxychloroquine, azithromycin, zinc, and vitamin C. Twenty-three percent of the patients died from the disease during the study period. Conclusion Morbidity and mortality were substantial in patients with COVID-19 admitted to a community hospital in East Harlem.
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http://dx.doi.org/10.7759/cureus.9836DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7496484PMC
August 2020

Epidermal growth factor receptor-activating mutation(E746_T751>VP) in pancreatic ductal adenocarcinoma responds to erlotinib, followed by epidermal growth factor receptor resistance-mediating mutation (A647T): A case report and literature review.

J Cancer Res Ther 2020 Jul-Sep;16(4):950-954

Division of Interventional Radiology, The University of South Alabama, Mobile, Alabama, USA.

Despite recent advances in treatment with multidrug chemotherapy regimens, outcomes of patients with advanced pancreatic ductal adenocarcinoma (PDAC) remain very poor. Treatment with targeted therapies has shown marginal benefits due to intrinsic or acquired resistance. Actionable mutations, while detected infrequently in patients with PDAC, are becoming increasingly used in personalized medicine. Here, we describe an epidermal growth factor receptor (EGFR)-activating mutation (E746_T751>VP) to erlotinib, a first-generation tyrosine kinase inhibitor (TKI), in a patient with metastatic PDAC. After an initial partial response to erlotinib for 12 months, the patient's disease progressed with emergence of the EGFR A647T mutation. Certainly, the patient also progressed after switching therapy to a third-generation EGFR TKI (osimertinib). This case illustrates the posttreatment evolution of EGFR A647T-mediated resistance to the first- and third-generation TKIs. To our knowledge, this is the first case to report the aforementioned activating and resistance-mediated mutations. In summary, genomic analysis performed in this patient with PDAC on the tumor biopsy and peripheral blood provided tools to understand mechanisms of response and resistance to targeted therapy with EFGR TKIs.
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http://dx.doi.org/10.4103/jcrt.JCRT_729_18DOI Listing
November 2020

Hypoxia alters the release and size distribution of extracellular vesicles in pancreatic cancer cells to support their adaptive survival.

J Cell Biochem 2020 01 12;121(1):828-839. Epub 2019 Aug 12.

Mitchell Cancer Institute, University of South Alabama, Mobile, Alabama.

Pancreatic tumors are highly desmoplastic and poorly-vascularized, and therefore must develop adaptive mechanisms to sustain their survival under hypoxic condition. Extracellular vesicles (EV) play vital roles in pancreatic tumor pathobiology by facilitating intercellular communication. Here we studied the effect of hypoxia on the release of EVs and examined their role in adaptive survival of pancreatic cancer (PC) cells. Hypoxia promoted the release of EV in PC cell lines, MiaPaCa and AsPC1, wherein former exhibited a far greater induction. Moreover, a time-dependent, measurable and significant increase was recorded for small EV (SEV) in both the cell lines with only minimal induction observed for medium (MEV) and large EVs (LEV). Similarly, noticeable changes in size distribution of SEV were also recorded with a shift toward smaller average size under extreme hypoxia. Thrombospondin (apoptotic bodies marker) was exclusively detected on LEVs, while Arf6 (microvesicles marker) was mostly present on MEV with some expression in LEV as well. However, CD9 and CD63 (exosome markers) were expressed in both SEV and MEVs with a decreased expression recorded under hypoxia. Among all subfractions, SEV was the most bioactive in promoting the survival of hypoxic PC cells and hypoxia-inducible factor-1α stabilization was involved in heightened EV release under hypoxia and for their potency to promote hypoxic cell survival. Altogether, our findings provide a novel mechanism for the adaptive hypoxic survival of PC cells and should serve as the basis for future investigations on broader functional implications of EV.
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http://dx.doi.org/10.1002/jcb.29328DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6878126PMC
January 2020

Exosomal markers (CD63 and CD9) expression and their prognostic significance using immunohistochemistry in patients with pancreatic ductal adenocarcinoma.

J Gastrointest Oncol 2019 Aug;10(4):695-702

Department of Oncologic Sciences, The University of South Alabama, Mitchell Cancer Institute, Mobile, AL, USA.

Background: Exosomes are important mediators of intercellular communications and play pivotal roles in cancer progression, metastasis and chemoresistance. CD63 and CD9 are widely accepted exosomal markers. In patients with pancreatic ductal adenocarcinoma (PDAC), positive correlation between CD9 expression and overall survival (OS) was reported. CD63 expression was conserved in all patients with no reported prognostic significance. This study explored the prognostic significance of CD63 and CD9 expression using immunohistochemistry (IHC) in patients with PDAC of mixed racial background.

Methods: Between 2012 and 2016, 49 patients with PDAC had available tissues for CD63 and CD9 staining using IHC. Two pathologists independently scored the CD63 and CD9 expression. Staining intensity was graded from 1-3 and staining percentage was estimated in 10% increments. Mean Quick-score (Q-score) (Intensity X Percentage of staining) was calculated.

Results: The mean Q-score for CD63 and CD9 are higher in primary tumor from the pancreas compared to pancreatic tumor from metastatic sites (185 102, P=0.0002) and (48 20, P=0.0418) respectively. We fitted Cox proportion hazard regression models to investigate the impact of the covariates CD63 and CD9 on progression free survival (PFS) and OS. CD63 has significant impact on PFS (P=0.0135) and OS (P=0.003). The higher the CD63 Q-score, the longer the PFS and OS. CD9 doesn't have significant impact on PFS (P=0.5734) or OS (P=0.2682). The mean CD63 and CD9 Q-scores are slightly higher in African American (AA) compared to Caucasians (157 149, P=0.76) and (45 29, P=0.43) respectively.

Conclusions: CD63 and CD9 expression is higher in primary tumor from the pancreas compared to pancreatic tumor from metastatic sites. There is correlation between CD63 expression (but not CD9 in this cohort) and PFS and OS. To our knowledge, this is the first study to show prognostic significance of CD63 expression in patients with PDAC using IHC. A trend of higher expression of CD63 and CD9 among AA compared to Caucasians was also noticed.
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http://dx.doi.org/10.21037/jgo.2018.07.02DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6657322PMC
August 2019

The Prevalence of DPYD*9A(c.85T>C) Genotype and the Genotype-Phenotype Correlation in Patients with Gastrointestinal Malignancies Treated With Fluoropyrimidines: Updated Analysis.

Clin Colorectal Cancer 2019 09 3;18(3):e280-e286. Epub 2019 May 3.

Medical Oncology, The University of South Alabama, Mitchell Cancer Institute, Mobile, AL. Electronic address:

Introduction: The dihydropyrimidine dehydrogenase gene (DPYD)*9A (c.85T>C) genotype is relatively common. The correlation between DPYD*9A genotype and dihydropyrimidine dehydrogenase (DPD) deficiency phenotype is controversial. In a cohort of 28 patients, DPYD*9A was the most commonly diagnosed variant (13 patients [46%]) and there was a noticeable genotype-phenotype correlation. In this study we genotyped a larger cohort of a mixed racial background to explore the prevalence of DPYD*9A variant and to confirm the genotype-phenotype correlation.

Patients And Methods: Between 2011 and 2018, in addition to genotyping for high-risk DPYD variants (DPYD*2A, DPYD*13 and DPYD*9B), genotyping for DPYD*9A variant was performed on 113 patients with gastrointestinal malignancies treated with fluoropyrimidines. Fluoropyrimidines-associated toxicity was graded according to the National Cancer Institute Common Terminology Criteria for Adverse Events (version 5.0). Fisher exact test was used for statistical analysis.

Results: Heterozygous and homozygous DPYD*9A genotypes were identified in 46 (41%) and 11 (10%) patients, respectively. Among patients with DPYD*9A genotypes (n = 57), men and women represented 30 (53%) and 27 (47%) patients, respectively. Caucasian, African American, and other ethnicities represented 29 (50.9%), 26 (45.6%), and 2 (3.5%) patients, respectively. Grade 3/4 toxicities were experienced in 26 patients with DPYD*9A genotype (3 patients had homozygous status) and in 20 patients with wild type DPYD*9A (P = .4405). In patients who received full-dose fluoropyrimidines (n = 85), Grade 3/4 toxicities were experienced in 22 patients with DPYD*9A genotype (2 patients had homozygous status), and in 17 patients with wild type DPYD (P = .8275).

Conclusion: In our updated analysis, the prevalence of heterozygous and homozygous DPYD*9A genotypes were 41% and 10%, respectively. The correlation between DPYD*9A genotype and DPD clinical phenotype was not reproduced. The noticeable correlation that we previously reported is likely because of small sample size and selection bias.
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http://dx.doi.org/10.1016/j.clcc.2019.04.005DOI Listing
September 2019

ETV4: an emerging target in pancreatic cancer.

Oncoscience 2018 Sep 11;5(9-10):260-261. Epub 2018 Oct 11.

Mitchell Cancer Institute, University of South Alabama, Mobile, AL 36604-1405, USA.

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http://dx.doi.org/10.18632/oncoscience.471DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6231443PMC
September 2018

Germline pharmacogenomics of DPYD*9A (c.85T>C) variant in patients with gastrointestinal malignancies treated with fluoropyrimidines.

J Gastrointest Oncol 2018 Jun;9(3):416-424

Medical Oncology, The University of South Alabama, Mitchell Cancer Institute, Mobile, Alabama, USA.

Background: The correlation between DPYD*9A (c.85T>C) genotype and dihydropyrimidine dehydrogenase (DPD) deficiency clinical phenotype is controversial. Reference laboratories either did not perform DPYD*9A genotyping or have stopped DPYD*9A genotyping and limited genotyping to high-risk variants (DPYD*2A, DPYD*13 and DPYD*9B) only. This study explored DPYD*9A genotype and clinical phenotype correlation in patients with gastrointestinal (GI) malignancies treated with fluoropyrimidines.

Methods: Between 2011 and 2017, 67 patients with GI malignancies were genotyped for DPYD variants. Fluoropyrimidines-associated toxicity was graded according to the National Cancer Institute Common Terminology Criteria for Adverse Events (version 3.0). Fisher's exact test was used for statistical analysis.

Results: DPYD variants were identified in 17 out of 67 (25%) patients. One patient was homozygous for DPYD*9A variant and one patient was double heterozygous for DPYD*9A and DPYD*9B variants. In patients with identified DPYD variants, 13/17 (76%) patients had DPYD*9A variant, 3/17 (18%) patients had DPYD*2A variant and 2/17 (12%) patient had DPYD*9B variant. Only patients genotyped prior to 2015 were genotyped for DPYD*9A variant (N=28). Of those, 13/28 patients (46%) had DPYD*9A variant. Grade 3-4 diarrhea was associated with DPYD*9A variant in patients treated with full dose fluoropyrimidines (P=0.0055).

Conclusions: In our cohort, DPYD*9A variant was the most common diagnosed variant. The correlation between DPYD*9A genotype and DPD deficiency in clinical phenotype was noticeable in patients who received full dose fluoropyrimidines as they all experienced grade 3-4 toxicities (diarrhea).
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http://dx.doi.org/10.21037/jgo.2018.02.03DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6006041PMC
June 2018

Perioperative Communication: Challenges and Opportunities for Anesthesiologists.

J Anaesthesiol Clin Pharmacol 2018 Jan-Mar;34(1):5-6

Department of Anesthesiology, Temple University's Katz College of Medicine, Philadelphia, USA.

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http://dx.doi.org/10.4103/joacp.JOACP_37_18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5885448PMC
April 2018

ETV4 Facilitates Cell-Cycle Progression in Pancreatic Cells through Transcriptional Regulation of Cyclin D1.

Mol Cancer Res 2018 02 8;16(2):187-196. Epub 2017 Nov 8.

Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, Alabama.

The ETS family transcription factor ETV4 is aberrantly expressed in a variety of human tumors and plays an important role in carcinogenesis through upregulation of relevant target gene expression. Here, it is demonstrated that ETV4 is overexpressed in pancreatic cancer tissues as compared with the normal pancreas, and is associated with enhanced growth and rapid cell-cycle progression of pancreatic cancer cells. ETV4 expression was silenced through stable expression of a specific short hairpin RNA (shRNA) in two pancreatic cancer cell lines (ASPC1 and Colo357), while it was ectopically expressed in BXPC3 cells. Silencing of ETV4 in ASPC1 and Colo357 cells reduced the growth by 55.3% and 38.9%, respectively, while forced expression of ETV4 in BXPC3 cells increased the growth by 46.8% in comparison with respective control cells. Furthermore, ETV4-induced cell growth was facilitated by rapid transition of cells from G- to S-phase of the cell cycle. Mechanistic studies revealed that ETV4 directly regulates the expression of , a protein crucial for cell-cycle progression from G- to S-phase. These effects on the growth and cell cycle were reversed by the forced expression of in ETV4-silenced pancreatic cancer cells. Altogether, these data provide the first experimental evidence for a functional role of ETV4 in pancreatic cancer growth and cell-cycle progression. The functional and mechanistic data presented here regarding ETV4 in pancreatic cancer growth and cell-cycle progression suggest that ETV4 could serve as a potential biomarker and novel target for pancreatic cancer therapy. .
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http://dx.doi.org/10.1158/1541-7786.MCR-17-0219DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5805611PMC
February 2018

Racial health disparities in ovarian cancer: not just black and white.

J Ovarian Res 2017 Sep 21;10(1):58. Epub 2017 Sep 21.

Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, 1660 Springhill Avenue, Mobile, AL, 36604-1405, USA.

Ovarian cancer (OC) is the most lethal gynecological malignancy, which disproportionately affects African American (AA) women. Lack of awareness and socioeconomic factors are considered important players in OC racial health disparity, while at the same time, some recent studies have brought focus on the genetic basis of disparity as well. Differential polymorphisms, mutations and expressions of genes have been reported in OC patients of diverse racial and ethnic backgrounds. Combined, it appears that neither genetic nor the socioeconomic factors alone might explain the observed racially disparate health outcomes among OC patients. Rather, a more logical explanation would be the one that takes into consideration the combination and/or the interplay of these factors, perhaps even including some environmental ones. Hence, in this article, we attempt to review the available information on OC racial health disparity, and provide an overview of socioeconomic, environmental and genetic factors, as well as the epigenetic changes that can act as a liaison between the three. A better understanding of these underlying causes will help further research on effective cancer management among diverse patient population and ultimately narrow health disparity gaps.
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http://dx.doi.org/10.1186/s13048-017-0355-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5607508PMC
September 2017

Characterization and quantification of flavonoids and organic acids over fruit development in American cranberry (Vaccinium macrocarpon) cultivars using HPLC and APCI-MS/MS.

Plant Sci 2017 Sep 8;262:91-102. Epub 2017 Jun 8.

Department of Plant Biology, Rutgers University, New Brunswick, NJ, USA; Philip E. Marucci Center for Blueberry and Cranberry Research and Extension, Rutgers University, Chatsworth, NJ, USA. Electronic address:

Cranberry flavonoids, including anthocyanins, flavonol glycosides and proanthocyanidins, and organic acids were characterized and quantified by HPLC and LC-MS/MS during fruit development and ripening in eight cranberry cultivars. Anthocyanin biosynthesis initiated at early fruit development and reached highest level in mature fruit, with significant differences between cultivars. Major flavonol glycosides, including the most abundant quercetin-3-galactoside and myricetin-3-galactoside, showed consistent concentrations during the season with moderate fluctuation, and were at similar levels in mature fruits of the eight cultivars. Proanthocyanidins declined during fruit development and then increased slightly in later maturation stages. Levels of various proanthocyanidin oligomers/polymers with different degree-of-polymerization were highly correlated within a cultivar during fruit development. Cultivars with coancestry exhibited similar levels (high/low) of anthocyanins or proanthocyanidins, indicating genetic effects on biosynthesis of such flavonoids. All cultivars showed similar levels of malic and citric acids, and declining levels of quinic acid during fruit development. Benzoic acid was extremely low early in the season and increased sharply during fruit ripening. Levels of quinic and citric acids were significantly different among cultivars in the mature fruit. Concentrations of proanthocyanidins, anthocyanins, quinic acid and benzoic acid have a strong developmental association in developing ovaries.
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http://dx.doi.org/10.1016/j.plantsci.2017.06.004DOI Listing
September 2017

Exosomal Markers (CD63 and CD9) Expression Pattern Using Immunohistochemistry in Resected Malignant and Nonmalignant Pancreatic Specimens.

Pancreas 2017 07;46(6):782-788

From the *Medical Oncology and †Department of Oncologic Sciences, Mitchell Cancer Institute, and Departments of ‡Pathology and §Surgical Oncology, The University of South Alabama, Mobile, AL.

Objectives: Exosomes are important mediators in intercellular communications and play a role in cancer progression and metastasis. Exosomal membranes are enriched in endosome-specific tetraspanins (CD9 and CD63). Here, we explored the expression of CD63 and CD9 utilizing immunohistochemistry in malignant and nonmalignant cells in 29 resected pancreatic specimens (RPSs) of mixed racial background.

Methods: The pathologic tissues (PTs) and adjacent normal tissues (ANTs) in each RPS were stained for CD63 and CD9. Two pathologists independently scored the expression of CD63 and CD9. Staining intensity was graded from 1 to 3. Staining percentage was estimated in 10% increments. An average Quick score (Q score) (intensity × percentage of staining) was calculated. Unpaired t test was used for statistical analysis.

Results: The mean multiplicative Q score for CD63 and CD9 expression is higher in PTs (209 and 72) compared with ANTs (154 and 24) (P = 0.0041, P = 0.0018), respectively. The Mean Q score for CD63 and CD9 expression is higher in the malignant PTs (231 and 85) compared with ANTs (129 and 25) (P < 0.0001 and P < 0.0124).

Conclusions: Exosomal markers (CD63 and CD9) expression assessment using immunohistochemistry is feasible in RPS. The expression of CD63 and CD9 is higher in PTs and malignant PTs compared with their ANTs.
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http://dx.doi.org/10.1097/MPA.0000000000000847DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5494969PMC
July 2017

MicroRNAs in gynecological cancers: Small molecules with big implications.

Cancer Lett 2017 10 24;407:123-138. Epub 2017 May 24.

Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, AL 36604, USA; Department of Biochemistry and Molecular Biology, College of Medicine, University of South Alabama, Mobile, AL 36688, USA. Electronic address:

Gynecological cancers (GCs) are often diagnosed at advanced stages, limiting the efficacy of available therapeutic options. Thus, there remains an urgent and unmet need for innovative research for the efficient clinical management of GC patients. Research over past several years has revealed the enormous promise of miRNAs. These small non-coding RNAs can aid in the diagnosis, prognosis and therapy of all major GCs, viz., ovarian cancers, cervical cancers and endometrial cancers. Mechanistic details of the miRNAs-mediated regulation of multiple biological functions are under constant investigation, and a number of miRNAs are now believed to influence growth, proliferation, invasion, metastasis, chemoresistance and the relapse of different GCs. Modulation of tumor microenvironment by miRNAs can possibly explain some of their reported biological effects. miRNA signatures have been proposed as biomarkers for the early detection of GCs, even the various subtypes of individual GCs. miRNA signatures are also being pursued as predictors of response to therapies. This review catalogs the knowledge gained from collective studies, so as to assess the progress made so far. It is time to ponder over the knowledge gained, so that more meaningful pre-clinical and translational studies can be designed to better realize the potential that miRNAs have to offer.
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http://dx.doi.org/10.1016/j.canlet.2017.05.011DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5601032PMC
October 2017

Emerging evidence for the role of differential tumor microenvironment in breast cancer racial disparity: a closer look at the surroundings.

Carcinogenesis 2017 08;38(8):757-765

Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, AL 36604, USA.

Although increased awareness leading to early detection and prevention, as well as advancements in treatment strategies, have resulted in superior clinical outcomes, African American women with breast cancer continue to have greater mortality rates, compared to Caucasian American counterparts. Moreover, African American women are more likely to have breast cancer at a younger age and be diagnosed with aggressive tumor sub-types. Such racial disparities can be attributed to socioeconomic differences, but it is increasingly being recognized that these disparities may indeed be due to certain genetic and other non-genetic biological differences. Tumor microenvironment, which provides a favorable niche for the growth of tumor cells, is comprised of several types of stromal cells and the various proteins secreted as a consequence of bi-directional tumor-stromal cross-talk. Emerging evidence suggests inherent biological differences in the tumor microenvironment of breast cancer patients from different racial backgrounds. Tumor microenvironment components, affected by the genetic make-up of the tumor cells as well as other non-tumor-associated factors, may also render patients more susceptible to the development of aggressive tumors and faster progression of disease resulting in early onset, thus adversely affecting patients' survival. This review provides an overview of breast cancer racial disparity and discusses the existence of race-associated differential tumor microenvironment and its underlying genetic and non-genetic causal factors. A better understanding of these aspects would help further research on effective cancer management and improved approaches for reducing the racial disparities gaps in breast cancer patients.
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http://dx.doi.org/10.1093/carcin/bgx037DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5862302PMC
August 2017

Resistin potentiates chemoresistance and stemness of breast cancer cells: Implications for racially disparate therapeutic outcomes.

Cancer Lett 2017 06 14;396:21-29. Epub 2017 Mar 14.

Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, AL, USA; Department of Biochemistry and Molecular Biology, College of Medicine, University of South Alabama, Mobile, AL, USA. Electronic address:

Breast cancer (BC) continues to be the most frequently diagnosed cancer in American women, which disproportionately affects women of African-American (AA) descent. Previously, we reported greater serum levels of resistin in AA BC patients relative to Caucasian-American (CA) patients, and established its role in growth and aggressiveness of breast tumor cells. Here we have investigated the role of resistin in BC-chemoresistance. MDA-MB-231 and MDA-MB-468 BC cells of CA and AA origin, respectively, were incubated with resistin prior to doxorubicin treatment. Our data suggest that resistin conferred chemoresistance to both BC cell lines; however, the effect on AA cells was more profound. Furthermore, the resistin-induced doxorubicin-resistance was shown to occur due to suppression of apoptosis. Resistin treatment also affected the stemness of BC cells, as suggested by reduced cell surface expression of CD24, induced expression of CD44 and ALDH1, and increased capability of cells to form mammospheres. Mechanistic studies revealed that resistin-induced chemoresistance, apoptosis and stemness of BC cells were mediated through STAT3 activation. Taken together, our findings provide novel insight into the role of resistin in BC biology, and strengthen its role in racially disparate clinical outcomes.
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http://dx.doi.org/10.1016/j.canlet.2017.03.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5437742PMC
June 2017

Exosomes confer chemoresistance to pancreatic cancer cells by promoting ROS detoxification and miR-155-mediated suppression of key gemcitabine-metabolising enzyme, DCK.

Br J Cancer 2017 Feb 2;116(5):609-619. Epub 2017 Feb 2.

Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, AL, USA.

Background: Chemoresistance is a significant clinical problem in pancreatic cancer (PC) and underlying molecular mechanisms still remain to be completely understood. Here we report a novel exosome-mediated mechanism of drug-induced acquired chemoresistance in PC cells.

Methods: Differential ultracentrifugation was performed to isolate extracellular vesicles (EVs) based on their size from vehicle- or gemcitabine-treated PC cells. Extracellular vesicles size and subtypes were determined by dynamic light scattering and marker profiling, respectively. Gene expression was examined by qRT-PCR and/or immunoblot analyses, and direct targeting of DCK by miR-155 was confirmed by dual-luciferase 3'-UTR reporter assay. Flow cytometry was performed to examine the apoptosis indices and reactive oxygen species (ROS) levels in PC cells using specific dyes. Cell viability was determined using the WST-1 assay.

Results: Conditioned media (CM) from gemcitabine-treated PC cells (Gem-CM) provided significant chemoprotection to subsequent gemcitabine toxicity and most of the chemoresistance conferred by Gem-CM resulted from its EVs fraction. Sub-fractionation grouped EVs into distinct subtypes based on size distribution and marker profiles, and exosome (Gem-Exo) was the only sub-fraction that imparted chemoresistance. Gene expression analyses demonstrated upregulation of SOD2 and CAT (ROS-detoxifying genes), and downregulation of DCK (gemcitabine-metabolising gene) in Gem-Exo-treated cells. SOD/CAT upregulation resulted, at least in part, from exosome-mediated transfer of their transcripts and they suppressed basal and gemcitabine-induced ROS production, and partly promoted chemoresistance. DCK downregulation occurred through exosome-delivered miR-155 and either the functional suppression of miR-155 or restoration of DCK led to marked abrogation of Gem-Exo-mediated chemoresistance.

Conclusions: Together, these findings establish a novel role of exosomes in mediating the acquired chemoresistance of PC.
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http://dx.doi.org/10.1038/bjc.2017.18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5344296PMC
February 2017

Biological basis of cancer health disparities: resources and challenges for research.

Am J Cancer Res 2017 1;7(1):1-12. Epub 2017 Jan 1.

Department of Oncologic Sciences, Mitchell Cancer Institute, University of South AlabamaMobile, Alabama 36604, USA; Department of Biochemistry and Molecular Biology, College of Medicine, University of South AlabamaMobile, Alabama 36688, USA.

Last few decades have witnessed remarkable progress in our understanding of cancer initiation and progression leading to refinement of prevention and treatment approaches. Although these advances have improved the survival of cancer patients in general, certain racial/ethnic groups have benefited only partially. Footprints of cancer-associated racial disparities are very much evident in cancers of the prostate, breast, cervical, colorectal, endometrium, liver and lung. These health inequalities are mostly attributed to socioeconomic differences among races, but there is a growing realization that these may actually be due to inherent biological differences as well. Indeed, significant data now exist to support the biological basis of racial disparities in cancer, warranting basic research investigations, using appropriate tools and model systems. In this article, we have aimed to succinctly review the literature supporting the biological bases of racial disparities in cancer, along with available resources, databases and model systems that will be of interest to researchers. Moreover, we have highlighted the specific areas that need attention in terms of development of resources and/or tools, and discuss the opportunities and challenges in basic biological research in cancer health disparities.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5250676PMC
January 2017

Epigenetic basis of cancer health disparities: Looking beyond genetic differences.

Biochim Biophys Acta Rev Cancer 2017 Aug 17;1868(1):16-28. Epub 2017 Jan 17.

Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, AL, USA; Department of Biochemistry and Molecular Biology, University of South Alabama, Mobile, AL, USA. Electronic address:

Despite efforts at various levels, racial health disparities still exist in cancer patients. These inequalities in incidence and/or clinical outcome can only be explained by a multitude of factors, with genetic basis being one of them. Several investigations have provided convincing evidence to support epigenetic regulation of cancer-associated genes, which results in the differential transcriptome and proteome, and may be linked to a pre-disposition of individuals of certain race/ethnicity to early or more aggressive cancers. Recent technological advancements and the ability to quickly analyze whole genome have aided in these efforts, and owing to their relatively easy detection, methylation events are much well-characterized, than the acetylation events, across human populations. The early trend of investigating a pre-determined set of genes for differential epigenetic regulation is paving way for more unbiased screening. This review summarizes our current understanding of the epigenetic events that have been tied to the racial differences in cancer incidence and mortality. A better understanding of the epigenetics of racial diversity holds promise for the design and execution of novel strategies targeting the human epigenome for reducing the disparity gaps.
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http://dx.doi.org/10.1016/j.bbcan.2017.01.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5513784PMC
August 2017

Racial disparities in prostate cancer: a molecular perspective.

Front Biosci (Landmark Ed) 2017 01 1;22:772-782. Epub 2017 Jan 1.

Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, Alabama, USA,Department of Biochemistry and Molecular Biology, College of Medicine, University of South Alabama, Mobile, Alabama, USA.

Prostate cancer incidence and mortality rates are remarkably higher in African-American men as compared to their European-Americans counterparts. Despite these recognitions, precise causes underlying such prevalent racial disparities remain poorly understood. Although socioeconomic factors could account for such differences up to a certain extent, it is now being increasingly realized that such disparity has a molecular basis. Indeed, several differences, including genetic polymorphism, gene mutations, epigenetic modifications, miRNAs alterations, . have been reported in malignant prostate tissues from patients of diverse racial backgrounds. Here, we attempt to provide a molecular perspective on prostate cancer racial disparities by gathering available information on these associated factors and discussing their potential significance in disproportionate incidence and clinical outcomes.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5242333PMC
http://dx.doi.org/10.2741/4515DOI Listing
January 2017

Comparative analysis of the relative potential of silver, Zinc-oxide and titanium-dioxide nanoparticles against UVB-induced DNA damage for the prevention of skin carcinogenesis.

Cancer Lett 2016 12 28;383(1):53-61. Epub 2016 Sep 28.

Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, AL 36604, USA; Department of Biochemistry and Molecular Biology, College of Medicine, University of South Alabama, Mobile, AL 36688, USA. Electronic address:

Sunscreen formulations containing UVB filters, such as Zinc-oxide (ZnO) and titanium-dioxide (TiO) nanoparticles (NPs) have been developed to limit the exposure of human skin to UV-radiations. Unfortunately, these UVB protective agents have failed in controlling the skin cancer incidence. We recently demonstrated that silver nanoparticles (Ag-NPs) could serve as novel protective agents against UVB-radiations. Here our goal was to perform comparative analysis of direct and indirect UVB-protection efficacy of ZnO-, TiO- and Ag-NPs. Sun-protection-factor calculated based on their UVB-reflective/absorption abilities was the highest for TiO-NPs followed by Ag- and ZnO-NPs. This was further confirmed by studying indirect protection of UVB radiation-induced death of HaCaT cells. However, only Ag-NPs were active in protecting HaCaT cells against direct UVB-induced DNA-damage by repairing bulky-DNA lesions through nucleotide-excision-repair mechanism. Moreover, Ag-NPs were also effective in protecting HaCaT cells from UVB-induced oxidative DNA damage by enhancing SOD/CAT/GPx activity. In contrast, ZnO- and TiO-NPs not only failed in providing any direct protection from DNA-damage, but rather enhanced oxidative DNA-damage by increasing ROS production. Together, these findings raise concerns about safety of ZnO- and TiO-NPs and establish superior protective efficacy of Ag-NPs.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5086276PMC
http://dx.doi.org/10.1016/j.canlet.2016.09.026DOI Listing
December 2016

Urinary Clearance of Cranberry Flavonol Glycosides in Humans.

J Agric Food Chem 2016 Oct 18;64(42):7931-7939. Epub 2016 Oct 18.

Department of Plant Biology and Pathology, Rutgers University , New Brunswick, New Jersey 08901, United States.

Cranberry is reported to have health benefits, including prevention of urinary tract infections and other chronic diseases, due to the high content of polyphenols, including flavonols and flavan-3-ols. The aim of this study was to determine the clearance of flavonol glycosides and flavan-3-ols and/or their metabolites in human urine. Ten healthy women volunteers ingested 240 mL of cranberry juice containing flavonol glycosides. Urine samples were collected at 0, 90, 225, and 360 min postingestion. While flavan-3-ols were not detected, five flavonol glycosides common in cranberry were identified. Quercetin-3-galactoside, the most abundant cranberry flavonol, exhibited the highest peak urine concentration (C) of 1315 pg/mg creatinine, followed by quercetin-3-rhamnoside, quercetin-3-arabinoside, myricetin-3-arabinoside, and myricetin-3-galactoside. Quercetin-3-arabinoside showed delayed clearance, C at 237 min (T), relative to other flavonols (90-151 min). Both aglycone and the conjugated sugar moiety structure mediate the flavonol's bioavailability. Interindividual variation for bioavailability and clearance is also apparent. Metabolites, e.g. glucoronides, were not detected.
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http://dx.doi.org/10.1021/acs.jafc.6b03611DOI Listing
October 2016

Honokiol suppresses pancreatic tumor growth, metastasis and desmoplasia by interfering with tumor-stromal cross-talk.

Carcinogenesis 2016 11;37(11):1052-1061

Department of Oncologic Sciences, Mitchell Cancer Institute.

The poor clinical outcome of pancreatic cancer (PC) is largely attributed to its aggressive nature and refractoriness to currently available therapeutic modalities. We previously reported antitumor efficacy of honokiol (HNK), a phytochemical isolated from various parts of Magnolia plant, against PC cells in short-term in vitro growth assays. Here, we report that HNK reduces plating efficiency and anchorage-independent growth of PC cells and suppresses their migration and invasiveness. Furthermore, significant inhibition of pancreatic tumor growth by HNK is observed in orthotopic mouse model along with complete-blockage of distant metastases. Histological examination suggests reduced desmoplasia in tumors from HNK-treated mice, later confirmed by immunohistochemical analyses of myofibroblast and extracellular matrix marker proteins (α-SMA and collagen I, respectively). At the molecular level, HNK treatment leads to decreased expression of sonic hedgehog (SHH) and CXCR4, two established mediators of bidirectional tumor-stromal cross-talk, both in vitro and in vivo . We also show that the conditioned media (CM) from HNK-treated PC cells have little growth-inducing effect on pancreatic stellate cells (PSCs) that could be regained by the addition of exogenous recombinant SHH. Moreover, pretreatment of CM of vehicle-treated PC cells with SHH-neutralizing antibody abolishes their growth-inducing potential on PSCs. Likewise, HNK-treated PC cells respond poorly to CM from PSCs due to decreased CXCR4 expression. Lastly, we show that the transfection of PC cells with constitutively active IKKβ mutant reverses the suppressive effect of HNK on nuclear factor-kappaB activation and partially restores CXCR4 and SHH expression. Taken together, these findings suggest that HNK interferes with tumor-stromal cross-talk via downregulation of CXCR4 and SHH and decreases pancreatic tumor growth and metastasis.
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http://dx.doi.org/10.1093/carcin/bgw096DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5091041PMC
November 2016

Deep sequencing and in silico analyses identify MYB-regulated gene networks and signaling pathways in pancreatic cancer.

Sci Rep 2016 06 29;6:28446. Epub 2016 Jun 29.

Department of Oncologic Sciences, Mitchell Cancer Institute, University of South Alabama, Mobile, AL 36604, USA.

We have recently demonstrated that the transcription factor MYB can modulate several cancer-associated phenotypes in pancreatic cancer. In order to understand the molecular basis of these MYB-associated changes, we conducted deep-sequencing of transcriptome of MYB-overexpressing and -silenced pancreatic cancer cells, followed by in silico pathway analysis. We identified significant modulation of 774 genes upon MYB-silencing (p < 0.05) that were assigned to 25 gene networks by in silico analysis. Further analyses placed genes in our RNA sequencing-generated dataset to several canonical signalling pathways, such as cell-cycle control, DNA-damage and -repair responses, p53 and HIF1α. Importantly, we observed downregulation of the pancreatic adenocarcinoma signaling pathway in MYB-silenced pancreatic cancer cells exhibiting suppression of EGFR and NF-κB. Decreased expression of EGFR and RELA was validated by both qPCR and immunoblotting and they were both shown to be under direct transcriptional control of MYB. These observations were further confirmed in a converse approach wherein MYB was overexpressed ectopically in a MYB-null pancreatic cancer cell line. Our findings thus suggest that MYB potentially regulates growth and genomic stability of pancreatic cancer cells via targeting complex gene networks and signaling pathways. Further in-depth functional studies are warranted to fully understand MYB signaling in pancreatic cancer.
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http://dx.doi.org/10.1038/srep28446DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4926062PMC
June 2016

Exploiting Nanotechnology for the Development of MicroRNA-Based Cancer Therapeutics.

J Biomed Nanotechnol 2016 Jan;12(1):28-42

MicroRNAs (miRNAs/miRs) represent a novel class of small non-coding RNAs that post-transcriptionally regulate gene expression by base pairing with complementary sequences in the 3' untranslated region (UTR) of target mRNAs. Functional studies suggest that miRNAs control almost every biological process, and their aberrant expression leads to a disease state, such as cancer. Differential expression of miRNAs in cancerous versus normal cells have generated enormous interest for the development of miRNA-based cancer cell-targeted therapeutics. Depending on the miRNA function and expression in cancer, two types of miRNA-based therapeutic strategies can be utilized that either restore or inhibit miRNA function through exogenous delivery of miRNAs mimics or inhibitors (anti-miRs). However, hydrophilic nature of miRNA mimics/anti-miRs, sensitivity to nuclease degradation in serum, poor penetration and reduced uptake by the tumor cells are chief hurdles in accomplishing their efficient in vivo delivery. To overcome these barriers, several nanotechnology-based systems are being developed and tested for delivery efficacy. This review summarizes the importance of miRNAs-based therapeutics in cancer, associated translational challenges and novel nanotechnology-assisted delivery systems that hold potential for next-generation miRNA-based cancer therapeutics.
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http://dx.doi.org/10.1166/jbn.2016.2172DOI Listing
January 2016

MYB Promotes Desmoplasia in Pancreatic Cancer through Direct Transcriptional Up-regulation and Cooperative Action of Sonic Hedgehog and Adrenomedullin.

J Biol Chem 2016 07 31;291(31):16263-70. Epub 2016 May 31.

From the Departments of Oncologic Sciences and Departments of Biochemistry and Molecular Biology and

Extensive desmoplasia is a prominent pathological characteristic of pancreatic cancer (PC) that not only impacts tumor development, but therapeutic outcome as well. Recently, we demonstrated a novel role of MYB, an oncogenic transcription factor, in PC growth and metastasis. Here we studied its effect on pancreatic tumor histopathology and associated molecular and biological mechanisms. Tumor-xenografts derived from orthotopic-inoculation of MYB-overexpressing PC cells exhibited far-greater desmoplasia in histological analyses compared with those derived from MYB-silenced PC cells. These findings were further confirmed by immunostaining of tumor-xenograft sections with collagen-I, fibronectin (major extracellular-matrix proteins), and α-SMA (well-characterized marker of myofibroblasts or activated pancreatic stellate cells (PSCs)). Likewise, MYB-overexpressing PC cells provided significantly greater growth benefit to PSCs in a co-culture system as compared with the MYB-silenced cells. Interrogation of deep-sequencing data from MYB-overexpressing versus -silenced PC cells identified Sonic-hedgehog (SHH) and Adrenomedullin (ADM) as two differentially-expressed genes among others, which encode for secretory ligands involved in tumor-stromal cross-talk. In-silico analyses predicted putative MYB-binding sites in SHH and ADM promoters, which was later confirmed by chromatin-immunoprecipitation. A cooperative role of SHH and ADM in growth promotion of PSCs was confirmed in co-culture by using their specific-inhibitors and exogenous recombinant-proteins. Importantly, while SHH acted exclusively in a paracrine fashion on PSCs and influenced the growth of PC cells only indirectly, ADM could directly impact the growth of both PC cells and PSCs. In summary, we identified MYB as novel regulator of pancreatic tumor desmoplasia, which is suggestive of its diverse roles in PC pathobiology.
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http://dx.doi.org/10.1074/jbc.M116.732651DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4965574PMC
July 2016

Influence of Degree-of-Polymerization and Linkage on the Quantification of Proanthocyanidins using 4-Dimethylaminocinnamaldehyde (DMAC) Assay.

J Agric Food Chem 2016 Mar 9;64(11):2190-9. Epub 2016 Mar 9.

Department of Plant Biology and Pathology, Rutgers University , New Brunswick, New Jersey 08901, United States.

Proanthocyanidins (PACs) are naturally occurring flavonoids possessing health beneficial bioactivities. Their quantification often utilizes the 4-dimethylaminocinnamaldehyde (DMAC) spectrophotometric assay with the assumption that molar absorption coefficients (MACs) are similar across the various PAC species. To assess the validity of this assumption, individual PAC monomers and oligomers were examined for their absorbance response with DMAC. Our results have shown that PAC dimers and trimers with interflavan linkage variations exhibited differential absorbance response. Absence of A-type linkage between the terminal and second units in PAC molecule not only impacts absorbance intensity at 640 nm but also elicits a prominent secondary 440 nm absorbance peak. Cranberry (A-type) and cocoa (B-type) oligomeric PACs exhibited differential absorbance (MACs) relationship with degree-of-polymerization. Thus, PAC structural variations have considerable impact on the resulting MAC. The use of DMAC assay in PAC quantification, especially in comparing across specific oligomers and compositions, should not assume MACs are similar.
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http://dx.doi.org/10.1021/acs.jafc.5b05408DOI Listing
March 2016

Cranberry Flavonoids Modulate Cariogenic Properties of Mixed-Species Biofilm through Exopolysaccharides-Matrix Disruption.

PLoS One 2015 29;10(12):e0145844. Epub 2015 Dec 29.

Biofilm Research Labs, Department of Orthodontics and Divisions of Pediatric Dentistry & Community Oral Health, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.

The exopolysaccharides (EPS) produced by Streptococcus mutans-derived glucosyltransferases (Gtfs) are essential virulence factors associated with the initiation of cariogenic biofilms. EPS forms the core of the biofilm matrix-scaffold, providing mechanical stability while facilitating the creation of localized acidic microenvironments. Cranberry flavonoids, such as A-type proanthocyanidins (PACs) and myricetin, have been shown to inhibit the activity of Gtfs and EPS-mediated bacterial adhesion without killing the organisms. Here, we investigated whether a combination of cranberry flavonoids disrupts EPS accumulation and S. mutans survival using a mixed-species biofilm model under cariogenic conditions. We also assessed the impact of cranberry flavonoids on mechanical stability and the in situ pH at the biofilm-apatite interface. Topical application of an optimized combination of PACs oligomers (100-300 μM) with myricetin (2 mM) twice daily was used to simulate treatment regimen experienced clinically. Treatments with cranberry flavonoids effectively reduced the insoluble EPS content (>80% reduction vs. vehicle-control; p<0.001), while hindering S. mutans outgrowth within mixed-species biofilms. As a result, the 3D architecture of cranberry-treated biofilms was severely compromised, showing a defective EPS-matrix and failure to develop microcolonies on the saliva-coated hydroxyapatite (sHA) surface. Furthermore, topical applications of cranberry flavonoids significantly weaken the mechanical stability of the biofilms; nearly 90% of the biofilm was removed from sHA surface after exposure to a shear stress of 0.449 N/m2 (vs. 36% removal in vehicle-treated biofilms). Importantly, in situ pH measurements in cranberry-treated biofilms showed significantly higher pH values (5.2 ± 0.1) at the biofilm-apatite interface vs. vehicle-treated biofilms (4.6 ± 0.1). Altogether, the data provide important insights on how cranberry flavonoids treatments modulate virulence properties by disrupting the biochemical and ecological changes associated with cariogenic biofilm development, which could lead to new alternative or adjunctive antibiofilm/anticaries chemotherapeutic formulations.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0145844PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4699891PMC
July 2016