RII Track-4:NSF: ExoPRIME Technology and Mass Spectroscopy Proteomic Analysis of Immunocaptured Exosomes for Liquid Biopsy

Extracellular vesicles (EVs) are nanosized particles secreted by the cells that contain protein and nucleic acid and are a promising tool for liquid biopsy. Proteomics analysis of circulating EVs is an emerging method for precise and non-invasive early diagnosis of pathological conditions. Selective isolation of EV subpopulations that express the same surface antibody is critical for the precise identification of disease-specific proteomics biomarkers.

This proposed research will enable the specificity assessment of technology for direct purification and proteomic analysis of subpopulations of EVs that express the same surface marker, and therefore originate from diseased tissue. The results from this work will also improve our understanding of biomarkers-specific signatures of cancer-originating EVs for early diagnostics via a liquid biopsy approach.

The fellowship program will train the PI and a graduate student from Louisiana Tech University in both technical aspects of sample processing for mass spectrometry as well as computational approaches for high throughput proteomics data analysis. This will enable the PI to modernize and increase the rigor of the proteomics courses at Louisiana Tech University.

This Research Infrastructure Improvement Track-4 EPSCoR Research Fellows (RII Track-4) project would provide a fellowship to an Assistant Professor and a graduate student at Louisiana Tech University. This work would be conducted in collaboration with researchers at the University of Houston. Selective immunopurification of EVs from complex biological fluids using disease-specific surface markers provides a non-invasive method for early disease diagnosis.

In this project, the PI and one graduate student will visit the Mass Spectrometry Facility at the University of Houston to perform bottom-up mass spectrometry proteomic analysis of astrocytes and tumor marker specific (EpCAM, EGFRvIII, and HSP-27) glioblastoma-secreted EVs utilizing Bruker's timsTOF Pro mass spectrometry system. The PI has demonstrated the feasibility of a tool for immunocapture of EVs.

At the core of the ExoPRIME approach is an antibody-functionalized stainless steel microscopic pin (3mm×130µm) for the purification of EVs expressing a specific surface marker. The main objectives of this research study are to identify glioblastoma-specific proteomics biomarkers and assess the feasibility of the ExoPRIME tool for clinical diagnostics at the point of care.

This project will achieve the following objectives:1) assess the proteomics profile of glioblastoma and healthy astrocytes-derived EVs purified using a commercial immunocapture approach; and 2) perform proteomics analysis of glioblastoma-derived EpCAM, EGFRvIII, and HSP-27 specific EVs purified using the ExoPRIME technology. Expertise gained during this fellowship will also enable the PI to train and educate students at Louisiana Tech University in cutting-edge proteomics analysis techniques.

This research has potential to lead to a comprehensive understanding of the proteomics profile of tumor-specific EVs which opens the door to technology development for biomarker discovery and early disease diagnostics.

This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.