Cervical mucus prorperties stratifv risk for preterm birth

Thesis (M. Eng.)--Massachusetts Institute of Technology, Dept. of Biological Engineering, 2012.Cataloged from PDF version of thesis.Includes bibliographical references (p. 46-52).Preterm birth impacts 15 million babies every year, leading to morbidity, mortality, significant health care costs, and lifelong consequences. The causes of preterm birth are unknown, resulting in ineffective treatment, but it is correlated with ascension of vaginal bacteria through the cervix, which is normally protected by a dense mucus plug during pregnancy. This mucus plug, consisting of a tight meshwork of glycoproteins called mucins, should prevent pathogens from accessing the sterile uterine environment. Cervical mucus from women at high risk and low risk for preterm birth was collected and compared. The aim of this study was to discover differences that will lead to clues about why preterm birth occurs, and ultimately what can be done about it in terms of prevention and intervention. Using rheological techniques and a translocation assay, we found that cervical mucus from women at high risk is more translucent and more elastic under both elongational and shear stress, than cervical mucus in normal pregnancies. These properties more closely resemble mucus during ovulation, when spermatozoa can most easily penetrate the barrier, than mucus in normal pregnancy. Furthermore, high risk mucus is more permeable to beads of comparable size to viruses, suggesting the barrier is weakened and foreign particles may harmfully traverse it to cause intrauterine infection. The techniques in this paper have not been previously used to study cervical mucus in the context of preterm labor, but their results may have important implications. If these mucus properties in women indeed permit increased bacterial infection through the cervix, then they can be used to stratify patients, allowing for more personalized prenatal care to lower the rate of preterm birth.by Grace Yao.M.Eng

Metabolomics approach for increasing CHO cell specific productivity

Chinese hamster ovary cells are the most commonly used expression system in the production of monoclonal antibody therapeutic drugs. The biomanufacturing industry has made significant advances in increasing protein titers of these cell cultures by over 100-fold since the 1980s to gram-per-liter ranges, and much of this progress has been made via increasing cell density and viability. However, even next generation processes are approaching the limits of how high cell densities can be reached with available technologies. On the other hand, the specific productivity (qP) of the cell lines, though much higher now than at the advent of biologics production, has not been improved to the same degree, and advances on this front are needed to attain higher titers in shorter times. In this work, a library of twelve cell lines, having a wide range of qPs but all derived from the same parental cell line and expressing one of two different antibodies, was investigated using an untargeted metabolomics approach. Spent medium samples were collected from each fed-batch culture at two time points. BioCAn (Biologically Consistent Annotation), a recently developed automated annotation tool, was used to determine the most likely identities of features detected in LC-MS data from these cell lines. A correlation analysis was then performed to find annotated features that were significantly associated with either cell growth (37 features), qP (32 features), or both (56 features). Interestingly, all features associated with cell growth showed a negative correlation, while all features associated with qP showed a positive correlation. To investigate whether metabolites positively correlated with qP reflect endogenous metabolic activity beneficial for productivity, several metabolites were added to the culture medium at varying concentrations. We found that supplementing the medium with one or more select metabolites could improve qP without negatively impacting cell growth. We next evaluated whether these metabolites could be used as biomarkers to identify clones with potential for high productivity, as current screening methods can falsely eliminate clones due to sub-optimal culture media or process conditions. Together, these studies demonstrate opportunities for using untargeted metabolomics to achieve higher titer in biologics production processes. Further, the identification of biomarkers has potential to shorten cell line development timelines, which is on the critical path to biologics manufacturing

Biologically consistent annotation of CHO cell culture metabolomics data

Metabolomics represents the effort to understand the role of metabolites in a biological system. Unfortunately, unambiguous metabolite identification represents a major bottleneck in liquid chromatography-mass spectrometry (LC-MS) based untargeted metabolomics. A widely used approach is to search spectral (MS/MS) libraries in reference databases for matching metabolites; however, this approach is limited by incomplete coverage. An alternative approach is to match detected features to candidate chemical structures based on their mass and computationally predicted fragmentation pattern. Both approaches often return too many possible matches; moreover, the results from different annotation tools rarely agree. This presentation describes a novel annotation tool that combines search results from several MS/MS libraries and computational fragmentation tools, and evaluates these results based on the content of a metabolic model. This captures the relevant biological context to determine the most likely identity for a given LC-MS data feature. This workflow, termed Biologically Consistent Annotation (BioCAn), improves on other publicly available annotation tools, achieving superior accuracy and sensitivity, while reducing the false discovery rate. The utility of this tool for investigating metabolic inefficiencies in cell culture processes is demonstrated by identifying novel CHO cell metabolites associated with enhanced or reduced cell growth and monoclonal antibody production. The function of these metabolites was evaluated in shake flask and controlled bioreactor experiments

Zebrafish as a model to study live mucus physiology

Dysfunctional mucus barriers can result in important pulmonary and gastrointestinal conditions, but model systems to study the underlying causes are largely missing. We identified and characterized five mucin homologues in zebrafish, and demonstrated a strategy for fluorescence labeling of one selected mucin. These tools can be used for in vivo experiments and in pharmacological and genetic screens to study the dynamics and mechanisms of mucosal physiology.National Institute of Environmental Health Sciences (Grant P30-ES002109)Johnson & Johnson. Corporate Office of Science and TechnologyNational Institutes of Health (U.S.) (Grant CA106416)Kathy and Curt Marble Cancer Research FundDavid H. Koch Institute for Integrative Cancer Research at MIT (Zebrafish Core Facility

α2 Integrin-Dependent Suppression of Pancreatic Adenocarcinoma Cell Invasion Involves Ectodomain Regulation of Kallikrein-Related Peptidase-5

Previous reports demonstrate that the α2-integrin (α2) mediates pancreatic ductal adenocarcinoma (PDAC) cell interactions with collagens. We found that while well-differentiated cells use α2 exclusively to adhere and migrate on collagenI, poorly differentiated PDAC cells demonstrate reduced reliance on, or complete loss of, α2. Since well-differentiated PDAC lines exhibit reduced in vitro invasion and α2-blockade suppressed invasion of well-differentiated lines exclusively, we hypothesized that α2 may suppress the malignant phenotype in PDAC. Accordingly, ectopic expression of α2 retarded in vitro invasion and maintenance on collagenI exacerbated this effect. Affymetrix profiling revealed that kallikrein-related peptidase-5 (KLK5) was specifically upregulated by α2, and reduced α2 and KLK5 expression was observed in poorly differentiated PDAC cells in situ. Accordingly, well-differentiated PDAC lines express KLK5, and KLK5 blockade increased the invasion of KLK5-positive lines. The α2-cytoplasmic domain was dispensable for these effects, demonstrating that the α2-ectodomain and KLK5 coordinately regulate a less invasive phenotype in PDAC

Differential and Joint Effects of Metformin and Statins on Overall Survival of Elderly Patients with Pancreatic Adenocarcinoma: A Large Population-Based Study.

Background: Published evidence indicates that individual use of metformin and statin is associated with reduced cancer mortality. However, their differential and joint effects on pancreatic cancer survival are inconclusive.Methods: We identified a large population-based cohort of 12,572 patients ages 65 years or older with primary pancreatic ductal adenocarcinoma (PDAC) diagnosed between 2008 and 2011 from the Surveillance, Epidemiology, and End Results (SEER)-Medicare-linked database. Exposure to metformin and statins was ascertained from Medicare Prescription Drug Event files. Cox proportional hazards models with time-varying covariates adjusted for propensity scores were used to assess the association while controlling for potential confounders.Results: Of 12,572 PDAC patients, 950 (7.56%) had used metformin alone, 4,506 (35.84%) had used statin alone, and 2,445 (19.45%) were dual users. Statin use was significantly associated with improved overall survival [HR, 0.94; 95% confidence interval (CI), 0.90-0.98], and survival was more pronounced in postdiagnosis statin users (HR, 0.69; 95% CI, 0.56-0.86). Metformin use was not significantly associated with overall survival (HR, 1.01; 95% CI, 0.94-1.09). No beneficial effect was observed for dual users (HR, 1.00; 95% CI, 0.95-1.05).Conclusions: Our findings suggest potential benefits of statins on improving survival among elderly PDAC patients; further prospective studies are warranted to corroborate the putative benefit of statin therapy in pancreatic cancer.Impact: Although more studies are needed to confirm our findings, our data add to the body of evidence on potential anticancer effects of statins. Cancer Epidemiol Biomarkers Prev; 26(8); 1225-32. ©2017 AACR

Natural Experiment Examining Impact of Aggressive Screening and Treatment on Prostate Cancer Mortality in Two Fixed Cohorts from Seattle Area and Connecticut

To determine whether the more intensive screening and treatment for prostate cancer in the Seattle≠Puget Sound area in 1987≠90 led to lower mortality from prostate cancer than in Connecticut

Longitudinal Associations of Leisure-Time Physical Activity and Cancer Mortality in the Third National Health and Nutrition Examination Survey (1986–2006)

Longitudinal associations between leisure-time physical activity (LTPA) and overall cancer mortality were evaluated within the Third National Health and Nutrition Examination Survey (NHANES III; 1988–2006; n = 15,535). Mortality status was ascertained using the National Death Index. Self-reported LTPA was divided into inactive, regular low-to-moderate and vigorous activity. A frequency-weighted metabolic equivalents (METS/week) variable was also computed. Hazard ratios (HRs) and 95% confidence intervals (CI) were calculated for overall cancer mortality in the whole sample, by body mass index categories and insulin resistance (IR) status. Nonsignificant protective associations were observed for regular low-to-moderate and vigorous activity, and for the highest quartile of METS/week (HRs range: 0.66–0.95). Individuals without IR engaging in regular vigorous activity had a 48% decreased risk of cancer mortality (HR: 0.52; 95% CI: 0.28–0.98) in multivariate analyses. Conversely, nonsignificant positive associations were observed in people with IR. In conclusion, regular vigorous activity may reduce risk of cancer mortality among persons with normal insulin-glucose metabolism in this national sample

A Sm(II)-mediated cascade approach to Dibenzoindolo[3,2-b]carbazoles:synthesis and evaluation

Previously unstudied dibenzoindolo[3,2-b]carbazoles have been prepared by two-directional, phase tag-assisted synthesis utilizing a connective-Pummerer cyclization and a SmI2-mediated tag cleavage-cyclization cascade. The use of a phase tag allows us to exploit unstable intermediates that would otherwise need to be avoided. The novel materials were characterized by X-ray, cyclic voltammetry, UV-vis spectroscopy, TGA, and DSC. Preliminary studies on the performance of OFET devices are also described