Treatment of life-threatening Epstein-Barr virus (EBV)–associated tumors remains a great challenge, especially for patients with relapsed or refractory disease. Here, we found that exosomes derived from phosphoantigen-expanded Vδ2-T cells (Vδ2-T-Exos) contained death-inducing ligands (FasL and TRAIL), an activating receptor for natural killer (NK) cells (NKG2D), immunostimulatory ligands (CD80 and CD86), and antigen-presenting molecules (MHC class I and II). Vδ2-T-Exos targeted and efficiently killed EBV-associated tumor cells through FasL and TRAIL pathways and promoted EBV antigen–specific CD4 and CD8 T cell expansion. Administration of Vδ2-T-Exos effectively controlled EBV-associated tumors in Rag2−/−γc−/− and humanized mice. Because expanding Vδ2-T cells and preparing autologous Vδ2-T-Exos from cancer patients ex vivo in large scale is challenging, we explored the antitumor activity of allogeneic Vδ2-T-Exos in humanized mouse cancer models. Here, we found that allogeneic Vδ2-T-Exos had more effective antitumor activity than autologous Vδ2-T-Exos in humanized mice; the allogeneic Vδ2-T-Exos increased the infiltration of T cells into tumor tissues and induced more robust CD4 and CD8 T cell–mediated antitumor immunity. Compared with exosomes derived from NK cells (NK-Exos) with direct cytotoxic antitumor activity or dendritic cells (DC-Exos) that induced T cell antitumor responses, Vδ2-T-Exos directly killed tumor cells and induced T cell–mediated antitumor response, thus resulting in more effective control of EBV-associated tumors. This study provided proof of concept for the strategy of using Vδ2-T-Exos, especially allogeneic Vδ2-T-Exos, to treat EBV-associated tumors.

Targeting cancer stem cells (CSCs) has been proposed as a new strategy to eradicate malignancies, including hepatocellular carcinoma (HCC). However, the mechanisms by which CSCs sustain their self-renewal and chemoresistance remain elusive. Nanog is a master transcriptional regulator of stemness, especially in CSCs. Its expression is tightly regulated by the ubiquitin-proteasome system in embryonic stem cells (ESCs). Whether the suppression of Nanog ubiquitination contributes to its over-expression in CSCs has not been explored. In addition, the role of receptor for activated C kinase 1 (RACK1), an adaptor protein implicated in HCC growth, in liver CSC-like traits remains to be determined. Methods : In vitro and in vivo assays were performed to investigate the role of RACK1 in liver CSC-like phenotype and murine ESC function. How RACK1 regulates Nanog expression was explored by immunoblotting and immunohistochemistry. The interaction of RACK1 with Nanog and the consequent effects on Nanog ubiquitination and stemness were then analyzed. Results : RACK1 promotes self-renewal and chemoresistance of human liver CSCs and maintains murine ESC function. Consistently, RACK1 enhances the expression of Nanog in human HCC cells and murine ESCs. The protein levels of RACK1 in clinical HCC tissues positively correlate with those of Nanog. Further exploration indicates that RACK1 directly binds to Nanog, which prevents its recruitment of E3 ubiquitin ligase FBXW8 and ubiquitin-dependent degradation. The interaction with Nanog is essential for RACK1 to promote stemness. Conclusions : Our data provide novel insights into the regulation of Nanog protein levels, as well the key role of RACK1 to enhance self-renewal and chemoresistance of CSCs in human HCC.

CD4+ T cells play predominant roles in protective immunity against blood-stage Plasmodium infection, both for IFN-γ-dependent effector mechanisms and providing B cell helper signals. Neddylation, an ubiquitination-like process triggered by covalent conjugation of NEDD8 to specific targets, has emerged as a potential regulator of T cell activities to TCR engagement. However, its contribution to T cell-mediated immunity to blood-stage malaria remains unclear. Here using an experimental model induced by Plasmodium yoelii 17XNL, and conditional knockout mice with T cell-specific deficiency of crucial components of neddylation pathway, we demonstrate activation of neddylation in T cells during blood-stage Plasmodium infection is essential for parasite control and host survival. Mechanistically, we show that apart from promoting CD4+ T cell activation, proliferation, and development of protective T helper 1 (Th1) cell response as suggested previously, neddylation is also required for supporting CD4+ T cell survival, mainly through B-cell lymphoma-2 (Bcl-2) mediated suppression of the mitochondria-dependent apoptosis. Furthermore, we provide evidence that neddylation contributes to follicular helper T (Tfh) cell differentiation, probably via augmenting the ubiquitin ligase Itch activity and proteasomal degradation of FoxO1, thereby facilitating germinal center (GC) formation and parasite-specific antibody production. This study identifies neddylation as a positive regulator of anti-Plasmodium immunity and provides insight into an involvement of such pathway in host resistance to infectious diseases.

Influenza epidemics and pandemics are constant threats to global public health. Although strategies including vaccines and antiviral drugs have achieved great advances in controlling influenza virus infection, the efficacy of these strategies is limited by the highly frequent mutations in the viral genome and the emergence of drug-resistant strains. Our previous study indicated that boosting the immunity of human Vγ9Vδ2-T cells with the phosphoantigen pamidronate could be a therapeutic strategy to treat seasonal and avian influenza virus infections. However, one notable drawback of γδ-T cell-based immunotherapy is the rapid exhaustion of proliferation and effector responses due to repeated treatments with phosphoantigens. Here, we found that the expression of CD137 was inducible in Vγ9Vδ2-T cells following antigenic stimulation. CD137 + Vγ9Vδ2-T cells displayed more potent antiviral activity against influenza virus than their CD137 − counterparts in vitro and in Rag2-/γc-/mice. We further demonstrated that CD137 costimulation was essential for Vγ9Vδ2-T cell activation, proliferation, survival and effector functions. In humanized mice reconstituted with human peripheral blood mononuclear cells, CD137 costimulation with a recombinant human CD137L protein boosted the therapeutic effects of pamidronate against influenza virus. Our study provides a novel strategy of targeting CD137 to improve the efficacy of Vγ9Vδ2-T cell-based immunotherapy.