Restriction lift date: 2026-12-31
Development and characterization of high-throughput assay to measure the activity of Rab11, an endosomal recycling regulator linked to breast cancer brain metastasis (BCBM)
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Date
2025
Authors
Parthasarathy, Pavithra
Journal Title
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Publisher
University College Cork
Published Version
Abstract
Approximately 10 - 15% of people with stage IV breast cancer are at a high risk of developing brain metastases, especially those with triple-negative breast cancer and HER2-positve breast cancer. In the past 20 years, significant advances have led to the development of targeted and personalized therapies for breast cancer treatment, however there is still an urgent need to develop new therapies to treat breast cancers that have metastasized and/or acquired drug resistance. The endosomal recycling membrane trafficking pathway is not widely studied as a therapeutic target, but recent studies have identified small GTPases such as Rab11, an endosomal recycling regulator, as a contributor to migration, invasion and tumor progression to the brain, through its regulation of vesicle transport and interaction with integrins and adhesion molecules. Therefore, compounds that modulate the activity of Rab11 may have therapeutic potential for breast cancer patients.The aim of this project is to develop an assay that can measure the intracellular activity of Rab11a, and to use the assay to identify small molecules that modulate its activity. A GST-trap assay was first developed to pull down active Rab11a from lysates of the MCF-7 breast cancer cell line. Next, this assay was modified to allow the measurement of Rab11 activity in intact MCF-7 cells in a multi-well format that can be optimized for automation. We compared the effect of a library of small molecule endosomal recycling inhibitors (ERIs) on Rab11 activity between the pull-down and in-cell assays.
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Keywords
Breast cancer brain metastasis (BCBM) , GST-pull down assay , Endosomal recycling pathway , Endosomal recycling inhibitors (ERIs) , Personalized therapies , Cancer treatment
Citation
Parthasarathy, P. 2025. Development and characterization of high-throughput assay to measure the activity of Rab11, an endosomal recycling regulator linked to breast cancer brain metastasis (BCBM). MRes Thesis, University College Cork.