Feasibility of Leukemia-Derived Exosome Enrichment and Co-isolated dsDNA Sequencing in Acute Myeloid Leukemia Patients: A Proof of Concept for New Leukemia Biomarkers Detection

Abstract

Exosomes are extracellular vesicles playing a pivotal role in the intercellular communication. They shuttle different cargoes, including nucleic acids from their cell of origin. For this reason, they have been studied as carriers of tumor markers in different liquid biopsy approaches, in particular for solid tumors. Few data are available concerning exosomes as markers of myeloid neoplasia. To better understand their real potential and the best approach to investigate leukemic exosomes, we present the results of a pilot feasibility study evaluating the application of next-generation sequencing analysis of dsDNA derived from exosomes isolated in 14 adult patients affected by acute myeloid leukemias. In particular, leukemia-derived exosome fractions have been analyzed. The concentration of dsDNA co-extracted with exosomes and the number and types of mutations detected were considered and compared with ones identified in the Bone Marrow (BM) and Peripheral Blood (PB) cells. Exosomal DNA concentration, both considering the cargo and the DNA surrounding the lipid membrane resulted in a linear correlation with leukemic burden. Moreover, exosomal DNA mutation status presented 86.5% of homology with BM and 75% with PB. The results of this pilot study confirmed the feasibility of a leukemia-derived exosome enrichment approach followed by exosomal dsDNA NGS analysis for AML biomarker detection. These data point to the use of liquid biopsy in myeloid neoplasia for the detection of active leukemic cells resident in the BM via a painless procedure.

Utilizing exosomes as sparking clinical biomarkers and therapeutic response in acute myeloid leukemia

“…Bernardi et al. first used the commercially available CE-IVD-based kits for exosome-enrichment methods to investigate leukemic sources and exosomal dsDNA target resequencing for adult AML marker detection; they performed next-generation sequencing analysis of exosome-derived dsDNA isolated from 14 adult patients with AML and identified the optimal amount of exosomal dsDNA as a potential AML biomarker for liquid biopsies; they found exosomal dsDNA can be developed as a tool that can facilitate the monitoring of AML progression and the early diagnosis of relapse after allogeneic hematopoietic stem cell transplantation ( 99 ). In summary, exosomes may offer a novel perspective on AML diagnosis and treatment response ( Table 2 ).…”

Twenty years of evolution of CML therapy: how the treatment goal is moving from disease to patient

“…[50][51][52] Last but not least, the application of dPCR resulted able to performing a precise, sensitive, and accurate quantification of BCR::ABL1 transcripts shuttled by circulating extracellular vesicles, 53,54 similar to what was observed in another type of leukemia. [55][56][57] Considering these encouraging results, it is not surprising that new commercial assays have become available for BCR::ABL1 transcript detection by dPCR 58 and that coordinated multicentric studies are optimizing and standardizing CML MRD monitoring by dPCR. 59,60 These data are constantly confirmed by new evidence and altogether stress the utility of moving to dPCR for minimal residual disease (MRD) monitoring in CML patients, in particular in subjects presenting low levels of BCR::ABL1 transcript and potentially eligible for stopping TKI therapy.…”

“… 50 – 52 Last but not least, the application of dPCR resulted able to performing a precise, sensitive, and accurate quantification of BCR::ABL1 transcripts shuttled by circulating extracellular vesicles, 53 , 54 similar to what was observed in another type of leukemia. 55 – 57 …”