EXOSOME PROTEOMICS AND POTENCY TESTING – PAVING THE WAY TO THERAPY Exosomes are emerging as superstars amidst the myriad of other soluble and trophic factors that mesenchymal stem cells (MSCs) secrete; and rightfully so, as these mediators of intercellular communication carry the inherent immunomodulatory capacity and therapeutic vigour of MSCs. Previously, the identity, purity and safety of ENDEXOME EXOTM , nanovesicles isolated from our proprietary immortalised MSC lines, have been reported using various techniques including Nano Flow Cytometry. We showed >97% of an intact EV population with a size of 50-200 nm, transmembrane identity markers such as CD63 and CD81; identity markers CD73 and CD105 establishing mesenchymal origin. Given the functional heterogeneity of Extracellular vesicles, it becomes imperative to evaluate their potency prior to clinical administration. In this study, we aim to validate potency of our ENDEXOME EXOTM using elaborate adaptations of established functional assays and MSC-specific assays. Next, we take a comprehensive approach to characterise and analyse the surfaceome (total number surface membrane proteins on exosomes) of all our ENDEXOME EXOTM with an aim to comprehensively characterise their distinct components including but not limited to integral membrane proteins (IMPs), peripherally associated membrane proteins (PMPs), RNA-binding Proteins (RBPs), RNA nucleoproteins (RNPs) in addition to surface-localised microRNAs (miRNAs) bound to the outer exosome surface, using mass spectrometry based protein identification and comprehensive proteomics profiling. This instils confidence towards the potential and practicality of using our novel exosomal entities in therapeutic modalities for disease treatment. (#80)
Devashree Vakil
1
,
Kuldip Sidhu
1
,
David Greening
2
- CK Cell Technologies, Norwest, NSW, Australia
- Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, Bundoora, VIC, Australia
EXOSOME PROTEOMICS AND POTENCY TESTING – PAVING THE WAY TO THERAPY Exosomes are emerging as superstars amidst the myriad of soluble trophic factors that mesenchymal stem cells (MSCs) secrete; and rightfully so, as they inherit the immunomodulation and therapeutic vigour of MSCs. The identity, purity and safety of ENDEXOME EXOTM , nano-sized extracellular vesicles (EVs) isolated from our proprietary immortalised MSC lines, have been established using Nano Flow Cytometry; with >97% of an intact population 50-200 nm in size and expressing transmembrane identity markers CD63 and CD81; mesenchymal origin identity markers CD73 and CD105. Given the functional heterogeneity of EVs, it becomes imperative to evaluate their potency prior to clinical administration. Next, we validate potency of our ENDEXOME EXOTM using adaptations of the mixed lymphocyte functional assays and MSC-specific CD73 assay. We also characterise and analyse the ENDEXOME EXOTM surfaceome (total surface membrane proteins on exosomes) aiming to comprehensively characterise their distinct components including but not limited to integral membrane proteins (IMPs), peripherally associated membrane proteins (PMPs), RNA-binding Proteins (RBPs), RNA nucleoproteins (RNPs) in addition to surface-localised microRNAs (miRNAs) bound to the outer exosome surface, using mass spectrometry based protein identification and comprehensive proteomics profiling. This instils confidence towards the potential and practicality of using our novel exosomal entities in therapeutic modalities for disease treatment. (95480)
Devashree Vakil 1 , Kuldip Sidhu 1 , David Greening 2
- CK Cell Technologies, Norwest, NSW, Australia
- Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, Bundoora, VIC, Australia