Mevyn Nizard

PD-1–Expressing Tumor-Infiltrating T Cells Are a Favorable Prognostic Biomarker in HPV-Associated Head and Neck Cancer

Head and neck cancers positive for human papillomavirus (HPV) have a more favorable clinical outcome than HPV-negative cancers, but it is unknown why this is the case. We hypothesized that prognosis was affected by intrinsic features of HPV-infected tumor cells or differences in host immune response. In this study, we focused on a comparison of regulatory Foxp3(+) T cells and programmed death-1 (PD-1)(+) T cells in the microenvironment of tumors that were positive or negative for HPV, in two groups that were matched for various clinical and biologic parameters. HPV-positive head and neck cancers were more heavily infiltrated by regulatory T cells and PD-1(+) T cells and the levels of PD-1(+) cells were positively correlated with a favorable clinical outcome. In explaining this paradoxical result, we showed that these PD-1(+) T cells expressed activation markers and were functional after blockade of the PD-1-PD-L1 axis in vitro. Approximately 50% of PD-1(+) tumor-infiltrating T cells lacked Tim-3 expression and may indeed represent activated T cells. In mice, administration of a cancer vaccine increased PD-1 on T cells with concomitant tumor regression. In this setting, PD-1 blockade synergized with vaccine in eliciting antitumor efficacy. Our findings prompt a need to revisit the significance of PD-1-infiltrating T cells in cancer, where we suggest that PD-1 detection may reflect a previous immune response against tumors that might be reactivated by PD-1/PD-L1 blockade.

VEGF-A modulates expression of inhibitory checkpoints on CD8+ T cells in tumors

VEGF-A production in the tumor microenvironment enhances expression of PD-1 and other inhibitory checkpoints involved with CD8+ T cell exhaustion, which can be reversed with anti-VEGF/VEGFR treatment.

Comprehensive analysis of current approaches to inhibit regulatory T cells in cancer

CD4+CD25+Foxp3+ regulatory T cells (Treg) have emerged as a dominant T cell population inhibiting anti-tumor effector T cells. Initial strategies used for Treg-depletion (cyclophosphamide, anti-CD25 mAb…) also targeted activated T cells, as they share many phenotypic markers. Current, ameliorated approaches to inhibit Treg aim to either block their function or their migration to lymph nodes and the tumor microenvironment. Various drugs originally developed for other therapeutic indications (anti-angiogenic molecules, tyrosine kinase inhibitors,etc) have recently been discovered to inhibit Treg. These approaches are expected to be rapidly translated to clinical applications for therapeutic use in combination with immunomodulators.

Induction of resident memory T cells enhances the efficacy of cancer vaccine

Tissue-resident memory T cells (Trm) represent a new subset of long-lived memory T cells that remain in tissue and do not recirculate. Although they are considered as early immune effectors in infectious diseases, their role in cancer immunosurveillance remains unknown. In a preclinical model of head and neck cancer, we show that intranasal vaccination with a mucosal vector, the B subunit of Shiga toxin, induces local Trm and inhibits tumour growth. As Trm do not recirculate, we demonstrate their crucial role in the efficacy of cancer vaccine with parabiosis experiments. Blockade of TFGβ decreases the induction of Trm after mucosal vaccine immunization, resulting in the lower efficacy of cancer vaccine. In order to extrapolate this role of Trm in humans, we show that the number of Trm correlates with a better overall survival in lung cancer in multivariate analysis. The induction of Trm may represent a new surrogate biomarker for the efficacy of cancer vaccine. This study also argues for the development of vaccine strategies designed to elicit them.

Synergy of Radiotherapy and a Cancer Vaccine for the Treatment of HPV-Associated Head and Neck Cancer

There is growing interest in the association of radiotherapy and immunotherapy for the treatment of solid tumors. Here, we report an extremely effective combination of local irradiation (IR) and Shiga Toxin B (STxB)–based human papillomavirus (HPV) vaccination for the treatment of HPV-associated head and neck squamous cell carcinoma (HNSCC). The efficacy of the irradiation and vaccine association was tested using a model of HNSCC obtained by grafting TC-1/luciferase cells at a submucosal site of the inner lip of immunocompetent mice. Irradiation and the STxB-E7 vaccine acted synergistically with both single and fractionated irradiation schemes, resulting in complete tumor clearance in the majority of the treated mice. A dose threshold of 7.5 Gy was required to elicit the dramatic antitumor response. The combined treatment induced high levels of tumor-infiltrating, antigen-specific CD8+ T cells, which were required to trigger the antitumor activity. Treatment with STxB-E7 and irradiation induced CD8+ T-cell memory, which was sufficient to exert complete antitumor responses in both local recurrences and distant metastases. We also report for the first time that a combination therapy based on local irradiation and vaccination induces an increased pericyte coverage (as shown by αSMA and NG2 staining) and ICAM-1 expression on vessels. This was associated with enhanced intratumor vascular permeability that correlated with the antitumor response, suggesting that the combination therapy could also act through an increased accessibility for immune cells. The combination strategy proposed here offers a promising approach that could potentially be transferred into early-phase clinical trials. Mol Cancer Ther; 14(6); 1336–45. ©2015 AACR.

Mucosal vaccines: Novel strategies and applications for the control of pathogens and tumors at mucosal sites

The mucosal immune system displays several adaptations reflecting the exposure to the external environment. The efficient induction of mucosal immune responses also requires specific approaches, such as the use of appropriate administration routes and specific adjuvants and/or delivery systems. In contrast to vaccines delivered via parenteral routes, experimental, and clinical evidences demonstrated that mucosal vaccines can efficiently induce local immune responses to pathogens or tumors located at mucosal sites as well as systemic response. At least in part, such features can be explained by the compartmentalization of mucosal B and T cell populations that play important roles in the modulation of local immune responses. In the present review, we discuss molecular and cellular features of the mucosal immune system as well as novel immunization approaches that may lead to the development of innovative and efficient vaccines targeting pathogens and tumors at different mucosal sites.

Resident Memory T Cells as Surrogate Markers of the Efficacy of Cancer Vaccines

Cancer vaccine boost via the cervicovaginal rather than the intramuscular route of immunization appears to be crucial to induce genital CD8+ T cells and tumor regression. This clinical activity is correlated with the ability of the mucosal boost to elicit resident memory T cells in the genital tract. Clin Cancer Res; 22(3); 530–2. ©2015 AACR. See related article by Sun et al., p. 657

Immunotherapy of HPV-associated head and neck cancer: Critical parameters

Various arguments support the development of a vaccine targeting human papillomavirus (HPV) for the treatment of HPV-associated head and neck cancer. However, the mucosal localization of this tumor, the HPV-driven downregulation of MHC Class I molecules and various other immunosuppressive mechanisms must be carefully considered to improve the clinical efficacy of such an immunotherapeutic strategy.

Abstract 2830: Mucosal imprinting of vaccine induced-CD8+T cells is crucial to inhibit mucosal tumors.

Although many human cancers are located in mucosal sites, most cancer vaccines are tested against subcutaneous tumors in preclinical models. The utility of preferentially inducing an anti-tumor immune response in the mucosal anatomic site of tumors has never been addressed. We therefore wondered whether mucosa-specific homing instructions to the immune system might influence mucosal tumor outgrowth. For this purpose, we set up original orthotopic models of head and neck and lung cancers monitored by magnetic resonance imaging or luciferase based in vivo optical imaging and vaccine based on a non replicative delivery system, the B subunit of Shiga toxin (STxB) as mucosal vector which has previously been shown to target antigen to dendritic cells. We showed that the growth of orthotopic head and neck or lung cancers expressing the E7 protein from HPV16 was only inhibited, when a cancer vaccine was delivered by the intranasal (i.n) mucosal and not the intramuscular (i.m) route. This anti-tumor effect was dependent on mucosal CD8+T cells as : i) Only a vaccine composed of STxB coupled to an E7 derived polypeptide (STxB-E7), but not the free E7 polypeptide elicited mucosal CD8+T cells. This mucosal induction of anti-E7 CD8+T cells, but not the systemic (spleen) specific anti-E7 CD8+T cells correlated with mucosal tumor protection. ii) A greater mucosal tumor infiltration of CD8+T cells was detected 7 days after tumor graft in mice that had been previously intranasally immunized with STxB-E7, than in mice vaccinated by the i.m. route. iii) CD8+T cell-depleted mice vaccinated with STxB-E7 by the i.n. route died before 20 days, whereas mice survived more than 6 months without CD8 depletion. As control, both routes of vaccine administration controlled the growth of subcutaneous tumors and elicited anti-E7 specific CD8+T cells in the spleen. To explain this finding, we demonstrated that only i.n. vaccination elicited mucosal specific CD8+T cells expressing the mucosal integrin CD49a. Blockade of CD49a decreased intratumoral CD8+T cell infiltration and the efficacy of cancer vaccine on mucosal tumor. We then showed that after intranasal vaccination, only dendritic cell from lung parenchyma, but not from spleen induced the expression of CD49a on co-cultured specific CD8+T cells. Tumor-infiltrating lymphocytes from human mucosal lung cancer also expressed CD49a at higher levels than TIL from non mucosal tumors, supporting the relevance and possible extrapolation of these results in humans. We thus identified a link between the route of vaccination and the induction of a mucosal homing program on induced CD8+T cells controlling their trafficking with a direct application on the efficacy of cancer vaccine to control mucosal tumors. Citation Format: Federico Sandoval, Mevyn Nizard, Magali Terme, Cecile Badoual, Michel-Francis Bureau, Olivier Clement, Elie Marcheteau, Alain Gey, Estelle Dransart, Francoise Quintin-Colonna, Gwenhael Autret, Tzyy-Choou Wu, Wolf H. Fridman, Ludger Johannes, Eric Tartour. Mucosal imprinting of vaccine induced-CD8+T cells is crucial to inhibit mucosal tumors. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2830. doi:10.1158/1538-7445.AM2013-2830

Abstract 5388: Local mucosal CD8+T cell response is required to inhibit the growth of orthotopic head and neck and lung cancers

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL Tumors may be located at mucosal or non-mucosal sites. However, the clinical benefit of preferentially inducing an anti-tumor immune response in the anatomic site of tumors has never been addressed. We set up orthotopic models of head and neck and lung cancers to compare the systemic and local anti-tumor immune response after administration of a cancer vaccine by mucosal and systemic routes. We also selected a non replicative delivery system, the B subunit of Shiga toxin (STxB), as a mucosal vector able to target dendritic cells. We show that intranasal immunization of mice with STxB based vaccine is the best route to elicit polyfunctional specific CD8+T cells in cervical and mediastinal lymph node than the use of non vectorized antigen or the intramuscular route. In line with these results, nasal mucosal administration of a model tumor antigen (E7 polypeptide from HPV 16) targeted to dendritic cells by STxB is more efficient to inhibit the growth of established orthotopic head and neck and lung cancers expressing the E7 antigen, than the administration of non vectorized antigen or the use of intramuscular route. A higher infiltration of CD8+T cells was detected 7 days after tumor graft, when mice were previously intranasally immunized with STxB-E7, than in mice vaccinated by the intramuscular route. Specific anti-E7 CD8+T cell tumor infiltration, was only observed after nasal immunization. Indeed, depletion of CD8+T cells inhibited the clinical efficiency of tumor vaccine demonstrating their role in tumor protection. In contrast, both routes completely controlled the growth of a subcutaneously E7 expressing tumor, which correlated with a similar induction of anti-E7 CD8+T cells in the spleen. Analysis of Integrin and chemokine receptor expression on tetramere positive cells showed that intranasal immunization induced higher levels of CD103 on T cells in bronchoalveolar lavage than intramuscular immunization with the same vaccine. This study emphasizes the need to elicit a potent anti-tumor response at the anatomic site of tumor and not just in the systemic compartment to induce tumor regression. This was achieved by i) administration of the vaccine by the intranasal route which was efficient in inducing CD8+T cells response at both locoregional and systemic sites allowing the control of both mucosal and non mucosal tumors. ii) The targeting of antigen to dendritic cells by STxB. This study is relevant to humans, as 30% of head and neck cancers express HPV16. Our results support the development of STxB-E7 vaccine administered by the i.n. route for the treatment of these HPV associated head and neck cancers. More generally, this study provides direct evidence for the compartmentalization of tumor immunity, a critical finding for the design of better cancer vaccines. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5388. doi:1538-7445.AM2012-5388