Exploring the potential of small RNA cargo of extracellular vesicles as a next-generation biomarker for B lymphoblastic leukemia

Longjohn, Modeline Nicholas (2023) Exploring the potential of small RNA cargo of extracellular vesicles as a next-generation biomarker for B lymphoblastic leukemia. Doctoral (PhD) thesis, Memorial University of Newfoundland.

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Abstract

Extracellular vesicles (EV) are novel promising sources of biomarkers in human diseases. Despite the promise that EVs hold, there are currently lots of unknowns, which limit our understanding. Therefore, the work presented in this thesis explored questions about EV s in pediatric B cell acute lymphoblastic leukemia (B-ALL) and compared the effect of collection tube treatment on EVs from selected human biofluids. For B-ALL, we wanted to identify a miRNA signature of pediatric B-ALL, which could be explored in EVs. We performed a reanalysis of rniRNA raw data and metanalysis of published rniRNAs data; to identify differentially expressed miRNAs (DEmiRs). Neither the reanalysis nor the metanalyses revealed a consensus miRNA signature. However, we identified promising miRNAs which could be fmther investigated. Next, we compared EVs from blood plasma of pediatric B-ALL patients and non-cancer donors (NCD) for size, concentration, and small RNA profile differences. We found that B-ALL plasma contains more EVs than NCD plasma. Furthermore, RNA-seq analysis of small RNAs revealed a signature of differentially packaged and exclusively packaged RNAs (including miRNAs, lncRNAs, mRNAs, and tRNAs) that distinguish NCD from B-ALL. Next, we investigated the small RNA cargo packaging patterns in EVs from pediatric B-ALL primary lymphoblasts and immo1talized cell lines. We found that RNA biotypes packaged from cells into EVs include mature and precursor miRNAs and fragments of mRNA, lncRNA and tRNA. We also found that pediatric B-ALL primary lymphoblasts and immo1talized cell lines package multiple RNA biotypes, using putative motifs shared by the selectively packaged RNAs. Finally, we compared EV characteristics (size, shape, concentration) and selected EV miRNA levels between non-Streck (EDTA for plasma) and Streck treated collection tubes for urine and blood sample collection. We found that neither treatment affected EV size and concentration in plasma or urine. However, Streck treatment led to reduced RNA cargo in EVs, compared to non-Streck. Taken together, this work provides new insights into EVs in pediatric B-ALL, and blood and urine sample collection for EV profiling. This is knowledge that can facilitate next generation research for EV based biomarkers and monitoring disease burden.

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