Breast cancer is a genetic disorder so common that it has claimed the lives of many women all over the world with statistics showing that it’s concentrated on the age of 45 - 55 years. Breast cancer, along with the other category of this disease is the product of the occurrence of a metastasized tumor. Neoplasm forms when genes are expressed abnormally such that the natural outcome of somatic cells is associated. Mutation, deletion and amplification of genes that control cellular growth and differentiation also cause the formation of a neoplasm. Thus, it is very critical to understand the genetic components of tumors because this understanding can be the missing puzzle in finding molecular pathways of malignant tumors. Molecular pathways are essential because they may provide useful information to identify and treat molecular targets for immunotherapy. Several treatments have been developed for breast cancer in the past 40 years. From mastectomy, aggressive treatments were introduced to conserve the breast of the patient. These treatments include adjuvant chemotherapy and hormonal therapy.
The introduction of hormonal therapy during the 1980s has reduced the number of estrogen receptor and progesterone receptor positive cancer. The number of dying women who are suffering from this debilitating disease continues to dwindle as the monoclonal antibody trastuzumab was introduced in the 1990s to treat HER2 positive (human epidermal growth factor receptor) breast cancer. Trastuzumab is the first monoclonal antibody that was approved for immunotherapy that targets oncogenes and has proven to be beneficial for women who are diagnosed with HER2 positive breast cancer.
The HER2 human epidermal receptor is a part of the family of epithelial tyrosine kinase receptors (HER1, HER2, HER3 and HER4) that exists in different combinations. Tyrosine kinase receptors interact with a ligand, an ion that attaches to a metal atom that forms a complex, to facilitate signaling pathways for intercellular communication. Breast cancers that have great excess of HER2 protein are more destructive than other breast cancer. The prognosis and chance of survival is not optimistic as that of patients that does not have an excessive number of HER2 genes.
The precise binding of a ligand to a receptor allows the interaction of a heterodimer, a protein made up of two polypeptide sequence having conflicting composition in the progression, number, or type amino acid deposit, that is responsible in determining which signal pathway is stimulated to control cell growth (3). HER2’s pairing with a ligand in a non-specific sequence has allowed the survival and differentiation of cells. HER2 also promotes ligand binding with heterodimers. As a consequence, the rate of incorporating the ligand-heterodimer complex in intracellular signaling decreases as compared to other heterodimer complexes. Further, HER2 homodimer is inherently active. When the HER2 gene is amplified, HER2 protein increases nearly a hundred times leading to HER2 homodimerization and activation and dysregulation of receptor and this initiates the progression of the tumor in the body.
In a recent trastuzumab-based chemotherapy research, it was shown that the level of amplification of HER2 gene is a good predictor of the sensitivity of a patient to therapy and cancer survival rates. It could link the significance of advanced cancer genome sequencing playing a very crucial role in understanding the nature of HER2 as a potential gene target for cancer treatments.
Mutation in HER2 gene is comparable to the insertion of nine base pairs in exon 20 in the EGFR of HER2 domains. As was previously mentioned, HER2 selectively forms heterodimer complexes with EGF receptors. When mutation occurs, the ATP binding pockets of these receptors increases tyrosine kinase activity. Hence, signal peptides are excessively phosphorylated while neoplastic cells grow in number, its sensitivity or resistance to tyrosine kinase activity decreases. This indicates the need to investigate the insertion in exon 20 of HER2 gene because mutations in HER2 gene can be biomarker for immunotherapy.
SNP has greatly helped oncology with its genetic alterations like insertions and removals and signal transduction study which provided researches the important information into the source and consequence of many varying kinds of cancer, pointing to suitable detection and healing. It is therefore the aim of this study to determine the presence of mutation in the HER2 gene and examine how mutation affects the sensitivity of patients to drugs that target the receptor. This study was preformed by isolating the genomic DNA from human breast cancer cell line. Polymerase chain reaction (PCR) was used to amplify the exon 20 0f the HER2 gene. After purifying the PCR products, the isolated HER2 gene was ligated to pJet 1.2 vector. The pJet/HEX20 was transformed in E. coli.
The plasmids were isolated and analyzed by restriction digestion with HindIII restriction enzyme to verify the presence of the insert. Then it was intended that the DNA would be sequenced and observed for any mutations in the cloned gene.
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