Author ORCID Identifier

0000-0002-5834-9921

Date of Graduation

8-2020

Document Type

Thesis (MS)

Program Affiliation

Cancer Biology

Degree Name

Masters of Science (MS)

Advisor/Committee Chair

Laura Bover

Committee Member

Stephanie Watowich

Committee Member

Shao-Cong Sun

Committee Member

Jagannadha Sastry

Committee Member

Robert Orlowski

Abstract

Identifying a cancer-specific cellular target is one of the key factors that can pave a “bench to the bedside” path of a target therapy. Bone marrow stromal cell antigen 2 (BST2) is a raft-associated type II transmembrane protein with an unusual topology. Analyses showed a variable degree of BST2 expression in most organs. Two research groups identified different BST2 isoforms of BST2 generated by posttranscriptional modification. In this research, I studied the long BST2 isoform’s distribution in breast tissue and B cell lines using the novel monoclonal antibody (MAb) LA5, an anti-BST2-long MAb. This approach revealed the expression of the long BST2 isoform in the malignant B-cell lines. Moreover, immunohistochemistry staining of ductal breast carcinoma tissues showed the ability of the generated MAb to detect long BST2-positive cancer cells surrounded by negatively stained normal tissue, indicating LA5 specificity for this isoform. In contrast to the existing clone 26F8 that binds to all isoforms, the LA5 MAb specifically recognized malignant cells while distinguishing and excluding surrounding normal cells. The specificity of the generated MAb clearly demonstrates potential diagnostic value in detecting the early infiltrations of malignant breast ductal carcinoma in specimens collected from patients. The other segment of my research entailed exploring the roles of BST2 in breast cancer, using a loss-of-function approach with BST2-targeting short hairpin RNA (shRNA) in the T47D breast cancer cell line. Silencing BST2 was associated with a statistically significant reduction in the viability of T47D (p value < 0.05) but had no impact its proliferation (p value = 0.6). Lastly, the downstream effects of BST2 silencing was evaluated through reverse phase protein array (RPPA) and revealed the statistically significant upregulation of four genes: MAPK1/MAPK3, HES1, FN1 and STAT3 (p value < 0.05). All these genes play different roles in the survival mechanisms that breast cancer cells may initiate when a critical pathway becomes compromised. By examining the adaptive cellular responses to BST2 silencing, this study offers insight into potential gene expression changes in breast cancer cells, with some changes enhancing the survival and proliferation of malignant cells.

Keywords

Monoclonal Antibodies, BST2, Isoforms, affinity, specificity, B cells, T47D, breast cancer, multiple myeloma

Available for download on Monday, July 12, 2021

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