SPIO Enhance the Cross-Presentation and Migration of DCs and Anionic SPIO Influence the Nanoadjuvant Effects Related to Interleukin-1ß.

Superparamagnetic iron oxide nanoparticles (SPIO) have been synthesized and explored for use as carriers of various nanoadjuvants via loading into dendritic cells (DCs). In our study, homogeneous and superparamagnetic nanoparticles are susceptible to internalization by DCs and SPIO-pulsed DCs showed excellent biocompatibility and capacity for ovalbumin (OVA) cross-presentation. Herein, we found that SPIO-loaded DCs can promote the maturation and migration of DCs in vitro. SPIO coated with 3-aminopropyltrimethoxysilane (APTS) and meso-2,3-dimercaptosuccinic acid (DMSA), which present positive and negative charges, respectively, were prepared. We aimed to investigate whether the surface charge of SPIO can affect the antigen cross-presentation of the DCs. Additionally, the formation of interleukin-1ß (IL-1ß) was examined after treatment with oppositely charged SPIO to identify the nanoadjuvants mechanism. In conclusion, our results suggest that SPIO are biocompatible and can induce the migration of DCs into secondary lymph nodes. SPIO coated with APTS (SPIO/A+) exhibited excellent adjuvant potentials for the promotion of antigen cross-presentation and T cell activation and surpassed that of DMSA-coated nanoparticles (SPIO/D-). This process may be related to the secretion of IL-1ß. Our study provides insights into the predictive modification of nanoadjuvants, which will be valuable in DC vaccine design and could lead to the creation of new adjuvants for applications in vaccines for humans.

Vx3-Functionalized Alumina Nanoparticles Assisted Enrichment of Ubiquitinated Proteins from Cancer Cells for Enhanced Cancer Immunotherapy.

A simple and effective strategy was developed to enrich ubiquitinated proteins (UPs) from cancer cell lysate using the α-Al2O3 nanoparticles covalently linked with ubiquitin binding protein (Vx3) (denoted as α-Al2O3-Vx3) via a chemical linker. The functionalized α-Al2O3-Vx3 showed long-term stability and high efficiency for the enrichment of UPs from cancer cell lysates. Flow cytometry analysis results indicated dendritic cells (DCs) could more effectively phagocytize the covalently linked α-Al2O3-Vx3-UPs than the physical mixture of α-Al2O3 and Vx3-UPs (α-Al2O3/Vx3-UPs). Laser confocal microscopy images revealed that α-Al2O3-Vx3-UPs localized within the autophagosome of DCs, which then cross-presented α-Al2O3-Vx3-UPs to CD8+ T cells in an autophagosome-related cross-presentation pathway. Furthermore, α-Al2O3-Vx3-UPs enhanced more potent antitumor immune response and antitumor efficacy than α-Al2O3/cell lysate or α-Al2O3/Vx3-UPs. This work highlights the potential of using the Vx3 covalently linked α-Al2O3 as a simple and effective platform to enrich UPs from cancer cells for the development of highly efficient therapeutic cancer vaccines.

Ubiquitinated Proteins Isolated From Tumor Cells Are Efficient Substrates for Antigen Cross-Presentation.

We have previously shown that inhibition of the proteasome causes defective ribosomal products to be shunted into autophagosomes and subsequently released from tumor cells as defective ribosomal products in Blebs (DRibbles). These DRibbles serve as an excellent source of antigens for cross-priming of tumor-specific T cells. Here, we examine the role of ubiquitinated proteins (Ub-proteins) in this pathway. Using purified Ub-proteins from tumor cells that express endogenous tumor-associated antigen or exogenous viral antigen, we tested the ability of these proteins to stimulate antigen-specific T-cell responses, by activation of monocyte-derived dendritic cells generated from human peripheral blood mononuclear cells. Compared with total cell lysates, we found that purified Ub-proteins from both a gp100-specific melanoma cell line and from a lung cancer cell line expressing cytomegalovirus pp65 antigen produced a significantly higher level of IFN-γ in gp100- or pp65-specific T cells, respectively. In addition, Ub-proteins from an allogeneic tumor cell line could be used to stimulate tumor-infiltrating lymphocytes isolated and expanded from non-small cell lung cancer patients. These results establish that Ub-proteins provide a relevant source of antigens for cross-priming of antitumor immune responses in a variety of settings, including endogenous melanoma and exogenous viral antigen presentation, as well as antigen-specific tumor-infiltrating lymphocytes. Thus, ubiquitin can be used as an affinity tag to enrich for unknown tumor-specific antigens from tumor cell lysates to stimulate tumor-specific T cells ex vivo or to be used as vaccines to target short-lived proteins.

The vitamin E analogue α-TEA stimulates tumor autophagy and enhances antigen cross-presentation.

The semisynthetic vitamin E derivative alpha-tocopheryloxyacetic acid (α-TEA) induces tumor cell apoptosis and may offer a simple adjuvant supplement for cancer therapy if its mechanisms can be better understood. Here we report that α-TEA also triggers tumor cell autophagy and that it improves cross-presentation of tumor antigens to the immune system. α-TEA stimulated both apoptosis and autophagy in murine mammary and lung cancer cells and inhibition of caspase-dependent apoptosis enhanced α-TEA-induced autophagy. Cell exposure to α-TEA generated double-membrane-bound vesicles indicative of autophagosomes, which efficiently cross-primed antigen-specific CD8(+) T cells. Notably, vaccination with dendritic cells pulsed with α-TEA-generated autophagosomes reduced lung metastases and increased the survival of tumor-bearing mice. Taken together, our findings suggest that both autophagy and apoptosis signaling programs are activated during α-TEA-induced tumor cell killing. We suggest that the ability of α-TEA to stimulate autophagy and enhance cross-priming of CD8(+) T cells might be exploited as an adjuvant strategy to improve stimulation of antitumor immune responses.

Alpha-alumina nanoparticles induce efficient autophagy-dependent cross-presentation and potent antitumour response.

Therapeutic cancer vaccination is an attractive strategy because it induces T cells of the immune system to recognize and kill tumour cells in cancer patients. However, it remains difficult to generate large numbers of T cells that can recognize the antigens on cancer cells using conventional vaccine carrier systems. Here we show that α-Al(2)O(3) nanoparticles can act as an antigen carrier to reduce the amount of antigen required to activate T cells in vitro and in vivo. We found that α-Al(2)O(3) nanoparticles delivered antigens to autophagosomes in dendritic cells, which then presented the antigens to T cells through autophagy. Immunization of mice with α-Al(2)O(3) nanoparticles that are conjugated to either a model tumour antigen or autophagosomes derived from tumour cells resulted in tumour regression. These results suggest that α-Al(2)O(3) nanoparticles may be a promising adjuvant in the development of therapeutic cancer vaccines.