Determining HER2 Expression and HER2 Glycosylation Changes - Research Paper Example

Determining HER2 Expression and HER2 Glycosylation Changes Objectives: To determine amplification/over-expression of the HER2/neu gene protein and the glycosylation changes connection associated with the biomarkers of breast cancer or cell lines. The significance of study would be to retrospectively experiment the amplification or expression of the HER2 in the secondary breast cancer cell line compared to the primary cell line while exhibiting glycosylation (O-Glycan changes) in malignant cancer cells. The study aims would also illustrate the extra HER2 receptor stimulation of cancer cells while contrasting the validation of HER-2 protein overexpression and amplification in the secondary breast cancer lines compared to the primary cancer line (Kucab, 2009). Introduction In the last years, multiple attempts have been conducted to develop strategies that could actually determine the over-expression of HER 2 positive breast cancer indicators in secondary breast cancer cell line compared to the primary cell line expression and the glycosylation process changes connected. In this regard, breast cancer lines have significantly been used to investigate the cancer pathobiology for new emerging therapies thereby identifying the cancer oncogenesis as a molecular heterogeneous disease (Schwab & Thomson Gale 2008). HER2 over-expression in metastatic breast cancer and O-Glycan changes exhibition has been used in invasive breast cancer in conjunction with the therapy involved for the illness. Therefore, HER-2 issues in metastatic versus primary breast cancer overexpression are associated with the HER-2-positive discovered disorder (Tavani, 2006). The increased prominence of HER-2 overexpression accompanied by glycosylation changes has increased interest in Breast cancer pathobiology researchers and academic learners to focus mainly on the basic theories and explanations for the origin and therapies for this particular condition. Many authors claim that the HER-2 oncogenes have been found to encode a transmembrane tyrosine kinase receptor that is responsible as the central classifier for the targeted therapy and invasive breast cancer disease (Jo & Zeon, 2015). On a broad analysis, the immunohistochemistry, fluorescence and chromogenic in-situ hybridization and the major marketed slide-based HER-2 methods are presented and contrasted broadly against the fundamental background of the HER-2 testing guideline testing (American Society of Clinical Oncology–College of American Pathologists guidelines). Significance of the study The over-expression of the HER2 receptor and glycosylation changes associated with breast cancer cell lines in the primary or secondary expression studies is relevant to the oncogenesis practitioners and other health agencies. This information is helpful for classification and identification of the invasive breast cancer and the targeted therapy for the disease (Jefferis, 2009). This study will guide the breast cancer oncologist and personalized medicine practitioners to design and implement sustainable targeted anti-HER-2 therapy in the management, treatment and prevention of breast cancer among the susceptible or risk population. Further experimentation studies related to this research will help breast cancer agencies such as National Breast Cancer: Foundation (NBCF) and charities working in hand with the health systems to prevent and cure breast cancer. Significantly, it will help in informing the oncologist practitioners in the area of therapeutic or management collaboration among the health professionals in therapy and management discourse. In this regard, this study would forestall duplication of HER-2 overexpression in secondary breast cancer lines and O-glycosylation changes compared to the primary breast cancer lines offering opportunities for exploration of Breast cancer invasiveness and targeted therapies (Jefferis, 2009). Research questions 1. What are the possible strategies that can be employed in determining the amplification or overexpression of HER-2 in secondary breast cancer cell line compared to the primary cancer cell line? 2. What is the role of HER-2 oncogenes in classification of breast cancer invasiveness and target therapy strategies? 3. What are the different mechanisms through which HER-2 oncogenes inhibit O-glycosylation changes applied to the primary and secondary breast cancer cell lines. Statement of the problem Since the breast cancer studies has consistently undergone changes as a result of HER-2 resistance to the current available therapies, Breast cancer management can be well be achieved if oncologists are aware of what is involved in HER-2 amplification or overexpression (Garabagi & Hall, 2012). The research is to be conducted to know the meaning of HER-2 overexpression in the breast cancer cell line and O-glycan inhibition and how it can be determined by experimental and statistical methods. In addition, the research is to uncover the function and signal transduction through which anti-HER-2 therapies opposes the abnormal cell proliferation, as well as the HER-2 oncogenes amplification with the GRBT genes (Browne, 2013). Research objectives 1) To examine the role played by the HER-2 amplification and over-expression in the causes and treatment of breast cancer 2) To explore the relation between the amplification of HER-2 in secondary and primary breast cancer cell line based on the experimentation research 3) To examine the inhibition of O-Glycan changes during the HER-2 oncogenes amplification in the primary and secondary breast cancer cell lines Methodology and Approaches Statistical method Since this research would be designed to fit in an oncogene experimental studies, both analytical and experimental approaches will be necessary for achieving the research objectives and results outcome. However, this study will experimentation of the patients identified in the feasibility study of the patients diagnosed with the primary breast cancer stages I-III, at ……………………Clinic between…………….. and ………….., for who ABO data will be available (n=120), and a second sub-cohort (n= 150) for whom all clinical details, except ABO status, will be available. The data captured in this study will incorporate the date of birth, gender living/diseased breast cancer condition, cancer diagnosis date, and family history of the breast cancer, date of death and co-morbidities documentation at the time of death. In addition, last menses dates as recorded in the medical record or the approximate age of menopause of the population sample to be experimented (EE Ugiagbe & Obaseki, 2012). Conversely, the HER-2/neu status (hormone receptor status), blood type, the tumor grade and stage, the breast cancer site, morphology, treatment as well as the regional lymph node involvement will be among the essential components that will be used in data collection. Demographic data will be abstracted from ………….s’ EMR. The reference population or sample and ABO data for the individuals with the breast cancer will be summarized from the ………………… Clinic/……………….Joint Venture Laboratory Blood Bank database and the EMR. The population sample will be used to determine the blood group antigens that will be drawn from the …………………. Epidemiologic Study Area (..ESA). Data combined from the EMR will be captured in the warehouse during the period of study. All the tumors status data such as the date of diagnosis, site and grade, age at breast cancer diagnosis, morphology and treatment will be obtained from the Regional Cancer, Registry Database for the known ABO status index cohort. The additional sub-cohort data of the breast cancer sample population except the ABO status would be obtained. Menopausal status data will either be determined electrically or manually (Browne, 2013). The HER2/neu results will be abstracted from the EMR. Over-expression of the HER-2 receptor protein cell membrane breast cancer is associated with altered glycosylation increased tumor cell growth, increased metastatic potential, adhesiveness and the disease aggressiveness. The clone to be used for the study will be a polyclonal (HER-2/neu Hercep Test Kit). The study criteria to be used in evaluating and scoring the HER-2/neu status (amplification and O-glycosylation) during the time frame of the study to be conducted. In the study to be conducted out, both cytoplasmic and membranous overexpression rates would be compared in various tests during the scoring of the primary and secondary breast tumors (Al-Rubeai, 2011). Laboratory methods and approaches Immunohistochemical HER-2 detection: Tissue fixation, antigen retrieval, slide storage and incubation time will be carefully be monitored such as the study outcome to remain unaltered or influenced. HER-2 antibody variation in 26 sample population will be analyzed. Remarkably, the total number of differently type of antibodies will be reported as either monoclonal or polyclonal antibodies will be recorded and analyzed. The antibodies will be organized in relation to the scoring categories for their publication and separated into cytoplasmic and membranous over-expression (Alhamdani & Hoheisel, 2011). Incubation times of the variation of antibodies to be studied will range significantly from 60 min to overnight. The antibodies will be diluted in the ratio between 1:20 and 1:250 though the differences in variation will not be used in the result for explaining the HER-2 stain due to inconsistency in the correlation over-expression and the incubation period. Cytoplasmic HER2 and Fish methods would also used in the study in determining the number of HER-2 oncogenes that overexpressed during the study period (Al-Rubeai, 2011). Scoring procedure HER-2 publication will be used to analyze the amplification in the secondary breast cancer tumors based on the genomic features such as FISH, RT_PCR, and the Northern blotting. 0 = Negative no staining cases O+ Negative barely visible membrane staining case 2+ Weak Positive Weak Positive membrane staining 3+ Positive Strong Positive membrane staining Statistical analysis The difference between the two independent groups of the breast sample population will be determined by the Mann-Whitney U test. However, the differences between the experimentation variables or relationships such as the relationship between overexpression in secondary breast cancer lines, secondary breast cancer lines, and the glycosylation changes will be determined by one-way ANOVA-Bonferroni comparison. In addition, the ROC curves will be used to estimate logistic aggressions in the study considering the conditions of the tumors (malignant/nonmalignant) comparing secondary/primary breast cancer cell lines as dependent variables (Browne, 2013). HER-2 overexpression and the O-glycan changes will be considered as an independent variable. Similarly, the study specificity and sensitivity patterns will be used for examining the CDC evaluated. In addition, the Two-sided Fisher’s exact will be used to examine the relationships that exist between the categorical variables. Conversely, a significance testing of the study correlation will be performed with the Spearman rank correlation analysis method. 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