Engineered Tumor-Derived Exosomes: Novel Tools for CD8+ T Cells Induction

Background and Aim: In spite of recent advances in immunotherapyof cancer, there have been limitations in cancer treatment and patientsurvival due to lack of antigen recognition and immunosuppressivetumor microenvironment. Although tumor-derived exosomes (TEX)are involved in the transmission of suppressive signals between tumorand immune cells, they contain tumor-specific antigens and severalheat shock proteins which are necessary for the induction of immuneresponse. In the present study, we used modified TEX with Let-7i andmiR-142 to target dendritic cells and CD8+ T cells as well as tumormicroenvironment.Methods: Mouse mammalian breast cancer cell line; 4T1, was cultured.Differential ultracentrifugation was used for isolation and purificationof exosome population from exosome free medium. Morphology, sizedistribution and protein expression of exosomes were evaluated byscanning electron microscopy (SEM), dynamic light scattering (DLS),western blot and flow cytometry. Bone marrow-derived DC wasdifferentiated from bone marrow progenitors and miRNAs inoculatedinto exosomes through electroporation. Delivery of miRNAs wasconfirmed by real-time PCR. Immature DC was incubated with LPS, TEXand manipulated TEX as control positive, control negative and test groupsrespectively. T cells were isolated from lymph node and co-culturedwith mature DCs. Proliferation, phenotype and cytokine release wereassessed using CFSE, flow cytometry by PE-conjugated anti- CD3 andFITC conjugated anti-CD8 and ELISA for IFNg and Granzyme-B.Results: Morphology and size of isolated exosomes evaluated usingelectron microscopy and dynamic light scattering. Exosomes werepositive for CD81, CD63 and TSG101 using western blotting and flowcytometry. The morphology of DCs changed and branched projectionson mDCs were observed during incubation with modified TEX likeLPS group but not TEX. DC-modified TEX in comparison with DC-TEX,expressed a higher percentage of CD11c, MHCII, CD80, and CD40,as activatory surface molecules. Higher proliferation index and CD8+population with a dominant increased expression of IFNg and GranzymeB were determined in the test group.Conclusion: We found that Let-7i could efficiently increase DC maturation also a combination of Let-7i with miR-142 have notably enhanced theeffect on either DC maturation or CTL induction and cytokine release.This outcome indicates that modified TEX may act as a cell-free vaccinein the future of cancer treatment.

Keywords:

Tumor-derived exosomes , miRNA , Dendritic cells , Cytotoxic T cells