Hong, Lisa
(2025)
Nitroalkenes Exploit Dependence on Autophagy-Lysosome Pathway in PARPi-Resistant Triple Negative Breast Cancer.
Doctoral Dissertation, University of Pittsburgh.
(Unpublished)
Abstract
Triple negative breast cancer (TNBC) currently lacks targeted therapy despite being the most aggressive breast subtype. However, patients with a mutation in the BRCA1/2 genes can receive treatment that exploits a vulnerability in DNA double-strand break (DSB) repair. Loss of BRCA1/2 function critically sensitizes tumors to poly-ADP ribose polymerase inhibitor (PARPi) treatment through prolonged DNA damage. Unfortunately, resistance to PARPi is an insurmountable problem for patients. Mechanisms conferring insensitivity to therapy include increased DNA damage repair and autophagy.
Electrophilic nitroalkenes (NFA) have emerged as potent anticancer treatments and sensitize TNBC cells to PARPi and other DNA-damaging therapies. NFAs post-translationally modify reactive protein cysteine thiols through a Michael addition reaction. To better understand the mechanism of action of NFA in cancer, we applied click chemistry pulldown approaches that identified NFA targets in the autophagy pathway, a PARPi resistance mechanism in TNBC. Thus, we hypothesize that NFAs sensitize PARPi-resistant TNBC cells to PARPi. To test this hypothesis, we generated three PARPi-resistant TNBC cell lines. RNA-seq analysis and proteomic approaches revealed upregulation in autophagy and lysosomal pathways in PARPi-resistant TNBC cell lines. Using click-chemistry pulldown approaches, we confirmed the autophagy regulator SQSTM1/p62 as an NFA target. p62 has redox-sensitive cysteine residues, Cys105 and Cys113, needed for its oligomerization during autophagy. We confirmed Cys105 and Cys113 as NFA targets, and NFA treatment phenocopied Cys105.113Ala mutants, including impaired p62 oligomerization, degradation, and inhibition of autophagy. Importantly, like chloroquine (autophagy inhibitor), treatment of PARPi-resistant TNBC with NFA resensitized TNBC to PARPi therapy by inhibiting autophagy and decreasing lysosomal biomass.
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Details
| Item Type: |
University of Pittsburgh ETD
|
| Status: |
Unpublished |
| Creators/Authors: |
|
| ETD Committee: |
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| Date: |
13 February 2025 |
| Date Type: |
Publication |
| Defense Date: |
12 November 2024 |
| Approval Date: |
13 February 2025 |
| Submission Date: |
16 December 2024 |
| Access Restriction: |
2 year -- Restrict access to University of Pittsburgh for a period of 2 years. |
| Number of Pages: |
197 |
| Institution: |
University of Pittsburgh |
| Schools and Programs: |
School of Medicine > Molecular Pharmacology |
| Degree: |
PhD - Doctor of Philosophy |
| Thesis Type: |
Doctoral Dissertation |
| Refereed: |
Yes |
| Uncontrolled Keywords: |
breast cancer, triple negative breast cancer, PARP, PARPi, TNBC, nitroalkene, resistance |
| Date Deposited: |
13 Feb 2025 18:18 |
| Last Modified: |
13 Feb 2025 18:18 |
| URI: |
http://d-scholarship.pitt.edu/id/eprint/47291 |
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