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Updated: May 29, 2025
Desthiobiotin-Streptavidin-Affinity Mediated Purification of RNA-Interacting Proteins in Mesothelioma Cells
Published on: April 25, 2018
Exploring RNA cargo in extracellular vesicles for pleural mesothelioma detection
Agnieszka Kraft1,2,3,4, Michaela B Kirschner1,4, Vanessa Orlowski1,4
1Department of Thoracic Surgery, University Hospital Zurich, Raemistrasse 100, 8091, Zurich, Switzerland.
Background:
Pleural Mesothelioma (PM) is a highly aggressive cancer, for which effective early detection remains a challenge due to limited screening options and low sensitivity of biomarkers discovered so far. While extracellular vesicles (EVs) have emerged as promising candidates for blood-based biomarkers, their role in PM has not been studied yet. In this study, we characterized the transcriptomic profile of EVs secreted by PM primary cells and explored their potential as a biomarker source for PM detection.
Methods:
We collected cell culture supernatant from early-passage PM cell cultures derived from the pleural effusion of 4 PM patients. EVs were isolated from the supernatant using Qiagen exoEasy Maxi kit. RNA isolation from EVs was done using the mirVana PARIS kit. Finally, single-end RNA sequencing was done with Illumina Novaseq 6000.
Results:
We identified a range of RNA species expressed in EVs secreted by PM cells, including protein-coding RNA (80%), long non-coding RNA (13%), pseudogenes (4.5%), and short non-coding RNA (1.6%). We detected a subset of genes associated with the previously identified epithelioid (32 genes) and sarcomatoid molecular components (36 genes) in PM-EVs. To investigate whether these markers could serve as biomarkers for PM detection in blood, we compared the RNA content of PM-EVs with the cargo of EVs isolated from the plasma of healthy donors (publicly available data). Majority of upregulated genes in PM-EVs were protein-coding and long non-coding RNAs. Interestingly, 25 of them were the sarcomatoid and epithelioid marker genes. Finally, functional analysis revealed that the PM-EV RNA cargo was associated with Epithelial-Mesenchymal transition, glycolysis, and hypoxia.
Conclusions:
This is the first study to characterize the transcriptomic profile of EVs secreted by PM primary cell cultures, demonstrating their potential as biomarker source for early detection. Further investigation of the functional role of PM-EVs will provide new insights into disease biology and therapeutic avenues.
