High-efficiency Generation of Multiple Short Noncoding RNA in B-cells and B-cell-derived Extracellular Vesicles

Abstract

Short noncoding (snc)RNAs are important new players in the landscape of biologics with therapeutic potential. Recently, we reported on a new method for the synthesis and delivery of snc RNA in B-cells transfected with plasmid DNA. Here using the same approach, we demonstrate that B-cells can be programmed for the enforced biogenesis and synchronous release of multiple sncRNAs. Our data show that this goal is feasible and that multiple sncRNA are released in the extracellular compartment in amounts comparable to those from B-cells programmed to express and secrete one scnRNA only. Furthermore, we found that the cargo of extracellular vescicles (EVs) isolated from programmed B-cells is remarkably enriched for multiple sncRNA. On average, we found that the content of multiple sncRNAs in EVs is 3.6 copynumber/EV. Collectively, we demonstrate that B-cells can be easily programmed toward the synthesis and release of multiple sncRNAs, including sncRNA-laden EVs, efficiently and specifically.

Extracellular vesicles produced in B cells deliver tumor suppressor miR-335 to breast cancer cells disrupting oncogenic programming in vitro and in vivo

“…At the outset, we reasoned that restoring miR-335 content in LM2 cells would be best achieved by transfecting B cells with a plasmid engineered with two miR-335 precursor stem loops 15 . The general approach and the generation of iEVs in B cells by transfection with plasmid DNA are shown in Fig.…”

“…Recently, we demonstrated that B cells can be reprogrammed for the enforced biogenesis and synchronous release of short noncoding (snc)RNAs 14 . sncRNAs were packaged and enriched as cargo in extracellular vesicles (EVs) induced in B cells (iEVs), with an estimate content of 3.6 copy number/iEV 15 . Here, we demonstrate that iEVs programmed to contain miR-335 cargo deliver and durably restore miR-335 to LM2 cells, modulate target mRNA expression in vitro and in vivo , and greatly reduce the growth of orthotopic LM2 tumors in immune deficient NSG mice.…”

Extracellular vesicles in autoimmune vasculitis - Little dirts light the fire in blood vessels

“…Exosomes derived from Treg cells can suppress Th1 cell proliferation and cytokine production by transferring microRNA Let-7d to Th1 cells [120]. Furthermore, B-cell derived EVs can carry multiple short noncoding RNA molecules that may be involved in a variety of processes in immunity and inflammation [121]. In addition, B cell-derived exosomes express functional integrins that can mediate cell-cell adherence during inflammation [122].…”

Delivery of miR-214 via extracellular vesicles downregulates Xbp1 expression and pro-inflammatory cytokine genes in macrophages

“…miRNAs are involved in the regulation of a variety of cellular processes, including cancer cell growth and metastasis [14][15][16][17] . Recently, this laboratory developed a novel technology wherein B lymphocytes can be programmed with suitable engineered plasmid DNA to enforce the release of extracellular vesicles (iEVs) with a predetermined miRNA payload [18,19] .…”