Jean-François Fonteneau

Measles Virus Vaccine–Infected Tumor Cells Induce Tumor Antigen Cross-Presentation by Human Plasmacytoid Dendritic Cells

Purpose: Plasmacytoid dendritic cells (pDC) are antigen-presenting cells specialized in antiviral response. The measles virus vaccine is proposed as an antitumor agent to target and specifically kill tumor cells without infecting healthy cells. Experimental Design: Here, we investigated, in vitro, the effects of measles virus vaccine–infected tumor cells on the phenotype and functions of human pDC. We studied maturation and tumor antigen cross-presentation by pDC, exposed either to the virus alone, or to measles virus vaccine–infected or UV-irradiated tumor cells. Results: We found that only measles virus vaccine–infected cells induced pDC maturation with a strong production of IFN-α, whereas UV-irradiated tumor cells were unable to activate pDC. This IFN-α production was triggered by the interaction of measles virus vaccine single-stranded RNA (ssRNA) with TLR7. We observed that measles virus vaccine–infected tumor cells were phagocytosed by pDC. Interestingly, we showed cross-presentation of the tumor antigen NYESO-1 to a specific CD8+ T-cell clone when pDC were cocultured with measles virus vaccine–infected tumor cells, whereas pDC were unable to cross-present NYESO-1 after coculture with UV-irradiated tumor cells. Conclusions: Altogether, our results suggest that the use of measles virus vaccine in antitumor virotherapy induces immunogenic tumor cell death, allowing pDC to mature, produce high amounts of IFN-α, and cross-present tumor antigen, thus representing a mode of recruiting these antigen-presenting cells in the immune response. Clin Cancer Res; 19(5); 1147–58. ©2012 AACR.

LFA‐3 co‐stimulates cytokine secretion by cytotoxic T lymphocytes by providing a TCR‐independent activation signal

T cell activation is known to depend not only on efficient antigen recognition and subsequent signaling through TCR, but also on interactions involving multiple adhesion and accessory molecules such as CD28/B7, LFA‐1/ICAM‐1 and LFA‐3/CD2. The present study dissects the role of LFA‐3/CD2 interactions in the activation of melanoma‐specific CD8+ T cell clones. To this end we analyzed the influence of LFA‐3 density on melanoma cells on lysis and cytokine production (TNF, IL‐2, IFN‐γ) by T cells following activation by various amounts of antigenic peptides. Our results indicate that increasing LFA‐3 density on melanoma cells variably affects their lysis susceptibility, but systematically and considerably enhances cytokine production by melanoma‐specific cytotoxic T lymphocyte (CTL) clones. At any stimulatory antigen density, LFA‐3 increased the fraction of responding cells and/or cytokine amounts produced by individual cells, without affecting TCR down‐regulation. These results show that CD2 engagement increases cytokine gene activation essentially by providing to T cells a TCR‐independent co‐activation signal. From a practical point of view, our data demonstrate that the level of LFA‐3 expressed on tumors critically affects cytokine production by specific CTL and thus the efficiency of specific immune reactions mediated by these cells.

MUC1-Specific Cytotoxic T Lymphocytes in Cancer Therapy: Induction and Challenge

MUC1 glycoprotein is often found overexpressed and hypoglycosylated in tumor cells from numerous cancer types. Since its discovery MUC1 has been an attractive target for antitumor immunotherapy. Indeed, in vitro and in vivo experiments have shown T-cell-specific responses against MUC1 in an HLA-restricted and HLA-unrestricted manner, although some animal models have highlighted the possible development of tolerogenic responses against this antigen. These observations permit the development of new T-cell vaccine strategies capable of inducing an MUC1-specific cytotoxic T cell response in cancer patients. Some of these strategies are now being tested in clinical trials against different types of cancer. To date, encouraging clinical responses have been observed with some MUC1 vaccines in phase II/III clinical trials. This paper compiles knowledge regarding MUC1 as a promising tumor antigen for antitumor therapeutic vaccines applicable to numerous cancers. We also summarize the results of MUC1-vaccine-based clinical trials.

Human natural killer cells promote cross-presentation of tumor cell-derived antigens by dendritic cells.

Dendritic cells (DCs) cross‐present antigen (Ag) to initiate T‐cell immunity against most infections and tumors. Natural killer (NK) cells are innate cytolytic lymphocytes that have emerged as key modulators of multiple DC functions. Here, we show that human NK cells promote cross‐presentation of tumor cell‐derived Ag by DC leading to Ag‐specific CD8+ T‐cell activation. Surprisingly, cytotoxic function of NK cells was not required. Instead, we highlight a critical and nonredundant role for IFN‐γ and TNF‐α production by NK cells to enhance cross‐presentation by DC using two different Ag models. Importantly, we observed that NK cells promote cell‐associated Ag cross‐presentation selectively by monocytes‐derived DC (Mo‐DC) and CD34‐derived CD11bnegCD141high DC subsets but not by myeloid CD11b+ DC. Moreover, we demonstrate that triggering NK cell activation by monoclonal antibodies (mAbs)‐coated tumor cells leads to efficient DC cross‐presentation, supporting the concept that NK cells can contribute to therapeutic mAbs efficiency by inducing downstream adaptive immunity. Taken together, our findings point toward a novel role of human NK cells bridging innate and adaptive immunity through selective induction of cell‐associated Ag cross‐presentation by CD141high DC, a process that could be exploited to better harness Ag‐specific cellular immunity in immunotherapy.

A Spliced Antigenic Peptide Comprising a Single Spliced Amino Acid Is Produced in the Proteasome by Reverse Splicing of a Longer Peptide Fragment followed by Trimming

Peptide splicing is a novel mechanism of production of peptides relying on the proteasome and involving the linkage of fragments originally distant in the parental protein. Peptides produced by splicing can be presented on class I molecules of the MHC and recognized by CTLs. In this study, we describe a new antigenic peptide, which is presented by HLA-A3 and comprises two noncontiguous fragments of the melanoma differentiation Ag gp100PMEL17 spliced together in the reverse order to that in which they appear in the parental protein. Contrary to the previously described spliced peptides, which are produced by the association of fragments of 3–6 aa, the peptide described in this work results from the ultimate association of an 8-aa fragment with a single arginine residue. As described before, peptide splicing takes place in the proteasome by transpeptidation involving an acyl-enzyme intermediate linking one of the peptide fragment to a catalytic subunit of the proteasome. Interestingly, we observe that the peptide causing the nucleophilic attack on the acyl-enzyme intermediate must be at least 3 aa long to give rise to a spliced peptide. The spliced peptide produced from this reaction therefore bears an extended C terminus that needs to be further trimmed to produce the final antigenic peptide. We show that the proteasome is able to perform the final trimming step required to produce the antigenic peptide described in this work.

Attenuated measles virus used as an oncolytic virus activates myeloid and plasmacytoid dendritic cells.

Attenuated measles viruses (MV) are assessed in clinical trials for their capacity to preferentially infect and kill tumor cells. We recently showed that MV-infected tumor cells are able to activate tumor antigen cross-presentation by myeloid and plasmacytoid dendritic cells. Thus, MV-based antitumor virotherapy may stimulate antitumor immune response.

Dendritic cell preparation for immunotherapeutic interventions.

Much effort has been made over the last decade to use dendritic cells (DCs) in vaccines to induce specific antitumor immune responses. However, the great hope provided by in vitro and in vivo preclinical investigations was not translated to the clinic in terms of clinical efficacy. Thus, one of the challenges resides in optimizing DC-based therapy to give maximum clinical efficacy while using manufacturing processes that enable quality control and scale-up of consistent products. In this article, we review DC biology and the DC-based clinical trials performed to date and focus on the DC maturation status compatible with the goals of cancer immunotherapy. We also highlight the different approaches used in these clinical studies, such as the DC types or subtypes used and their preparation. Finally, we discuss the immunological and clinical outcomes in treated patients, with emphasis on the strategies that could be used to improve DC-based vaccination.

Natural Oncolytic Activity of Live-Attenuated Measles Virus against Human Lung and Colorectal Adenocarcinomas

Lung and colorectal cancers are responsible for approximately 2 million deaths each year worldwide. Despite continual improvements, clinical management of these diseases remains challenging and development of novel therapies with increased efficacy is critical to address these major public health issues. Oncolytic viruses have shown promising results against cancers that are resistant to conventional anticancer therapies. Vaccine strains of measles virus (MV) exhibit such natural antitumor properties by preferentially targeting cancer cells. We tested the ability of live-attenuated Schwarz strain of MV to specifically infect tumor cells derived from human lung and colorectal adenocarcinomas and demonstrated that live-attenuated MV exhibits oncolytic properties against these two aggressive neoplasms. We also showed that Schwarz MV was able to prevent uncontrollable growth of large, established lung and colorectal adenocarcinoma xenografts in nude mice. Moreover, MV oncolysis is associated with in vivo activation of caspase-3 in colorectal cancer model, as shown by immunohistochemical staining. Our results provide new arguments for the use of MV as an antitumor therapy against aggressive human malignancies.

Vaccination with epigenetically treated mesothelioma cells induces immunisation and blocks tumour growth

Malignant mesothelioma (MM) is an aggressive tumour associated with poor outcome in patients. Current treatments for MM are of limited efficacy. Our recent findings suggest that epigenetic drugs may induce both cytotoxicity and an immune response against MM cells. Thus, we used a mouse model of MM (AK7) to analyse how epigenetic drugs could modulate MM development in vivo. The treatment of tumour-bearing mice with an epigenetic drug already tested in clinical MM treatments (SAHA/Vorinostat) reduced the tumour mass and induced a moderate lymphocytic infiltration. However, the treatment did not stop tumour development. In order to show the potential effect of this epigenetic drug on tumour immunogenicity, in addition to cell cytotoxicity, we immunised mice either with AK7 cells pre-treated with SAHA, or with one of two cytotoxic drugs (curcumin or selenite), prior to transplantation of live AK7 cells. A specific immune response was observed only in mice immunised with AK7 cells pre-treated with the epigenetic drug (SAHA) and the tumour growth was arrested. An increase in the proportion of CD3+ CD8+ lymphocytes occurred in the peritoneal cavity. We also observed large conglomerates of immune cells in the omentum with clusters of CD8+ T cells, together with lymphocytes directed against residual AK7 cells in the interlobular connective tissue of the pancreas. Our data demonstrate that epigenetic drugs, such as SAHA, can stimulate tumour immunogenicity and improve the recognition of aggressive MM cells by the immune system in vivo.

Antitumor Virotherapy by Attenuated Measles Virus (MV)

Antitumor virotherapy consists of the use of replication-competent viruses to infect and kill tumor cells preferentially, without damaging healthy cells. Vaccine-attenuated strains of measles virus (MV) are good candidates for this approach. Attenuated MV uses the CD46 molecule as a major entry receptor into cells. This molecule negatively regulates the complement system and is frequently overexpressed by cancer cells to escape lysis by the complement system. MV exhibits oncolytic properties in many cancer types in vitro, and in mouse models. Phase I clinical trials using MV are currently underway. Here, we review the state of this therapeutic approach, with a focus on the effects of MV on the antitumor immune response.