Membrane-bound HSP70-engineered myeloma cell-derived exosomes stimulate more efficient CD8⁺ CTL- and NK-mediated antitumour immunity than exosomes released from heat-shocked tumour cells expressing cytoplasmic HSP70

Exosomes (EXO) derived from tumour cells have been used to stimulate antitumour immune responses, but only resulting in prophylatic immunity. Tumour-derived heat shock protein 70 (HSP70) molecules are molecular chaperones with a broad repertoire of tumour antigen peptides capable of stimulating dendritic cell (DC) maturation and T-cell immune responses. To enhance EXO-based antitumour immunity, we generated an engineered myeloma cell line J558_HSP expressing endogenous P1A tumour antigen and transgenic form of membrane-bound HSP70 and heat-shocked J558_HS expressing cytoplasmic HSP70, and purified EXO_HSP and EXO_HS from J558_HSP and J558_HS tumour cell culture supernatants by ultracentrifugation. We found that EXO_HSP were able to more efficiently stimulate maturation of DCs with up-regulation of Ia^b, CD40, CD80 and inflammatory cytokines than EXO_HS after overnight incubation of immature bone-marrow-derived DCs (5 × 10⁶ cells) with EXO (100 μg), respectively. We also i.v. immunized BALB/c mice with EXO (30 μg/mouse) and assessed P1A-specific T-cell responses after immunization. We demonstrate that EXO_HSP are able to stimulate type 1 CD4⁺ helper T (Th1) cell responses, and more efficient P1A-specific CD8⁺ cytotoxic T lymphocyte (CTL) responses and antitumour immunity than EXO_HS. In addition, we further elucidate that EXO_HSP-stimulated antitumour immunity is mediated by both P1A-specific CD8⁺ CTL and non-P1A-specific natural killer (NK) responses. Therefore, membrane-bound HSP70-expressing tumour cell-released EXO may represent a more effective EXO-based vaccine in induction of antitumour immunity.