Engineered exosomes emerging from muscle cells break the tolerance towards tumor self-antigens

Researchers at the National Center for Global Health, Italy recently described a novel biotechnological platform for the production of unrestricted cytotoxic T lymphocyte (CTL) vaccines. It relies on in vivo engineering of exosomes, i.e., nanovesicles constitutively released by all cells, with full-length antigens of choice upon fusion with an exosome-anchoring protein referred to as Nefmut. They are produced upon intramuscular injection of a DNA vector and, when uploaded with a viral tumor antigen, were found to elicit an immune response inhibiting the tumor growth in a model of transplantable tumors. However, for a possible application in cancer immunotherapy, a number of key issues remained unmet. Among these, the researchers have now investigated: (i) whether the immunogenic stimulus induced by the engineered exosomes can break immune tolerance, and (ii) their effectiveness when applied in human system.

As a model of immune tolerance, the researchers considered mice transgenic for the expression of activated rat HER2/neu which spontaneously develop adenocarcinomas in all mammary glands. When these mice were injected with a DNA vector expressing the product of fusion between Nefmut and the extracellular domain of HER2/neu, antigen-specific CD8+ T lymphocytes became readily detectable. This immune response associated with a HER2-directed CTL activity and a significant delay in tumor development. On the other hand, through cross-priming experiments, they demonstrated the effectiveness of the engineered exosomes emerging from transfected human primary muscle cells in inducing antigen-specific CTLs. They propose their CTL vaccine platform as part of new immunotherapy strategies against tumors expressing self-antigens, i.e., products highly expressed in oncologic lesions but tolerated by the immune system.

Nef-specific CTL activity elicited by human DCs co-cultivated with
muscle cells expressing Nefmut-based DNA vectors


a Scheme of the cross-priming assays. SKMC were transfected and, 48 h later, put in co-culture with iDCs which, after additional 24 h, were isolated and matured. Autologous PBLs were then added to mDCs, and the co-cultivation was carried out for 7 days. Afterwards, the PBL stimulation was repeated, and after additional 7 days, CD8+ T lymphocytes were isolated and tested in CTL assays through co-cultivation with syngeneic target cells. b Western blot analysis of cell lysates from either parental or Nefmut stably transfected MCF-7 cells. Filters were incubated with either anti-Nef or anti-β-actin Abs. Arrow signs the relevant protein product. Molecular markers are given in kDa. c CTL assay carried out by co-cultivating primed CD8+ T lymphocytes with MCF-7 cells expressing or not Nefmut in a 10:1 cell ratio. Results were presented as mean values +SD calculated from triplicate conditions of three independent experiments. *p < 0.05. Mean value of background conditions (i.e., co-culture of naïve CD8+ T lymphocytes with MCF-7), 11.9 ± 5

These researchers have established a novel, exosome-based method to produce unrestricted CTL vaccines. This strategy is effective in breaking the tolerance towards tumor self-antigens. The method is also useful to elicit antigen-specific CTL immunity in humans. These findings open the way towards the use of this antitumor strategy in clinic.

Anticoli S, Aricò E, Arenaccio C, Manfredi F, Chiozzini C, Olivetta E, Ferrantelli F, Lattanzi L, D’Urso MT, Proietti E, Federico M. (2017) Engineered exosomes emerging from muscle cells break immune tolerance to HER2 in transgenic mice and induce antigen-specific CTLs upon challenge by human dendritic cells. J Mol Med (Berl) [Epub ahead of print]. [abstract]

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