Date of Award

Fall 11-19-2025

Embargo Period

12-12-2025

Document Type

Thesis - MUSC Only

Degree Name

Master of Biomedical Science

Department

Biochemistry and Molecular Biology

College

College of Graduate Studies

First Advisor

Philip Howe

Second Advisor

Saverio Gentile

Third Advisor

Lalima K Madan

Fourth Advisor

Ozgur Sahin

Abstract

In comparison to other subtypes, triple-negative breast cancer (TNBC) has limited treatment options due to its heterogeneous nature and lack of therapeutic targets compared to non-TNBC. Accounting for 15-20% of all breast cancers, TNBC is characterized by its lack of expression of estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2). The standard of care is neoadjuvant chemotherapy, followed by surgery if applicable and adjuvant chemotherapy. The effectiveness of this treatment course is limited due to its severe side-effects such as cardiotoxicity and chemoresistance. The development of resistance to chemotherapy is more common in TNBC than in other subtypes and its underlying mechanism is still elusive.

We aim to define the mechanisms through which the E3 ubiquitin ligase ARIH-1 impacts chemoresponse in TNBC, as breast cancer (BC) patient data show higher ARIH-1 transcript levels correlate with worse survival. We have previously identified a novel role for ARIH-1 in the epithelial to mesenchymal transition (EMT), tumor initiation, and metastatic progression of TNBC. EMT, a process that is aberrantly activated in cancer, is associated with increased cell migration, invasion, and the acquisition of cancer stem cell (CSC) traits that enhance chemoresistance. Based on these findings, we looked at the impact of ARIH-1 in response to chemotherapy.

We assessed ARIH-1 expression in the Doxorubicin-resistant cell line of our TNBC MDA-MB-231 cells model, showing ARIH-1 is upregulated in these cells at the transcript and protein level. To test the impact of ARIH-1 on cancer cell viability, we generated MDA-MB-231 TNBC cells overexpressing (OE) or silencing (KD) ARIH-1. We show that ARIH-1 silencing increases response to chemotherapy treatment with Doxorubicin, a DNA-damaging anthracycline. To assess the biochemical mechanism linking ARIH-1 to chemoresistance, we utilized miniTurboID proximity labeling method to identify and test novel targets of ARIH-1 potentially related to chemoresistance or DNA damage response pathways. Here, we identified topoisomerase II-alpha (TOP2A), an enzyme involved in DNA topology regulation. Cycloheximide chase assay successfully demonstrated TOP2A stability is regulated by ARIH-1-mediated proteasomal degradation and silencing ARIH-1 improves TOP2A’s DNA decatenation ability when compared to control. Together, we validate this target as a candidate substrate of ARIH-1 that may be impacting chemotherapy sensitivity.

Rights

Copyright is held by the author. All rights reserved.

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