An Isoform Of Human Mammaglobin And Its Expression In Breast Cancer Cells

Defining a specific tumor marker of mammary carcinoma will be greatly beneficial to its diagnosis and metastases/mircrometastases monitoring. Human mammaglobin (hMAM), a 93-amino-acid glycoprotein, was expressed exclusively to mammary tissue, and over-expressed at a 10-fold level in primary breast cancer tissues and cell lines. hMAM was found as a complex with BU101 in a head to tail orientation. There is a growing body of evidences shows that hMAM is not only a potential marker for breast cancer due to its specific expression profile and secretary nature, but also a target in the immunotherapy and vaccine design. Though certain achievements have been made in the studies of hMAM as tumor marker and associated antigen, some researchers hold different views in its application. Since alterative splicing occurs frequently in breast cancer, we doubted whether there existed other forms of hMAM in breast cancer cell line MCF-7, which was reported hMAM negative. In this study, we isolated total RNA of the cell, and used reverse transcription and twice PCR to amplify the objective cDNA, investigated its bioinformatic features and expression in breast cancer cells.The aim of this study is to explore whether there exists an isoform of hMAM in MCF-7 cells, to study the possible cause of its formation, to identify its functional alteration in comparison with the wild type, and to investigate its expression in cultured breast cancer cells and peripheral disseminated cells, so as to imply the isoform as the complimentarity of mammary tumor marker ----- hMAM (wild type as we called). The project was studied as the following 3 sections.1. Gene cloning of hMAM isoformUsing a pair of specific primers designed according to the published hMAM cDNA in GenBank, a fragment of 270 bp cDNA was achieved with reverse transcription and twice PCR amplification from breast cancer cells MCF-7, which was hMAM negative as reported. The sequencing result of 3 independent experiments indicated that the obtained sequence had a deletion of 9 continual nucleotides (236-244,GTGTTTATG) compared with that of hMAM cDNA. We defined this cDNA as an isoform of hMAM. It encodes a product of 90 amino acids, with a deletion of 3 continual amino acids (79-81), valine (V), phenylalaine...