CD40-mediated amplification of local immunity by epithelial cells is impaired by HPV

The interaction between the transmembrane glycoprotein surface receptor CD40 expressed by skin epithelial cells (ECs) and its T-cell-expressed ligand CD154 was suggested to exacerbate inflammatory skin diseases. However, the full spectrum of CD40-mediated effects by ECs underlying this observation is unknown. Therefore, changes in gene expression after CD40 ligation of ECs were studied by microarrays. CD40-mediated activation for 2 hours stimulated the expression of a coordinated network of immune-involved genes strongly interconnected by IL8 and TNF, whereas after 24 hours anti-proliferative and anti-apoptotic genes were upregulated. CD40 ligation was associated with the production of chemokines and the attraction of lymphocytes and myeloid cells from peripheral blood mononuclear cells (PBMCs). Thus, CD40-mediated activation of ECs resulted in a highly coordinated response of genes required for the local development and sustainment of adaptive immune responses. The importance of this process was confirmed by a study on the effects of human papilloma virus (HPV) infection to the EC's response to CD40 ligation. HPV infection clearly attenuated the magnitude of the response to CD40 ligation and the EC's capacity to attract PBMCs. The fact that HPV attenuates CD40 signaling in ECs indicates the importance of the CD40-CD154 immune pathway in boosting cellular immunity within epithelia

Human papillomavirus downregulates the expression of IFITM1 and RIPK3 to escape from IFNγ- and TNFα-mediated antiproliferative effects and necroptosis

textabstractThe clearance of a high-risk human papillomavirus (hrHPV) infection takes time and requires the local presence of a strong type 1 cytokine T cell response, suggesting that hrHPV has evolved mechanisms to resist this immune attack. Using an unique system for non, newly, and persistent hrHPV infection, we show that hrHPV infection renders keratinocytes (KCs) resistant to the antiproliferative- and necroptosis-inducing effects of IFNγ and TNFα. HrHPV-impaired necroptosis was associated with the upregulation of several methyltransferases, including EZH2,and the downregulation of RIPK3 expression. Restoration of RIPK3 expression using the global histone methyltransferase inhibitor 3-deazaneplanocin increased necroptosis in hrHPV-positive KCs. Simultaneously, hrHPV effectively inhibited IFNγ/TNFα-mediated arrest of cell growth at the S-phase by downregulating

Chemotherapy alters monocyte differentiation to favor generation of cancer-supporting M2 macrophages in the tumor microenvironment

Current therapy of gynecologic malignancies consists of platinum-containing chemotherapy. Resistance to therapy is associated with increased levels of interleukin (IL)-6 and prostaglandin E2 (PGE(2)), 2 inflammatory mediators known to skew differentiation of monocytes to tumor-promoting M2 macrophages. We investigated the impact of cisplatin and carboplatin on 10 different cervical and ovarian cancer cell lines as well as on the ability of the tumor cells to affect the differentiation and function of cocultured monocytes in vitro. Treatment with cisplatin or carboplatin increased the potency of tumor cell lines to induce IL-10-producing M2 macrophages, which displayed increased levels of activated STAT3 due to tumor-produced IL-6 as well as decreased levels of activated STAT1 and STAT6 related to the PGE(2) production of tumor cells. Blockade of canonical NF-κB signaling showed that the effect of the chemotherapy was abrogated, preventing the subsequent increased production of PGE(2) and/or IL-6 by the tumor cell lines. Treatment with the COX-inhibitor indomethacin and/or the clinical monoclonal antibody against interleukin-6 receptor (IL-6R), tocilizumab, prevented M2-differentiation. Importantly, no correlation existed between the production of PGE(2) or IL-6 by cancer cells and their resistance to chemotherapy-induced cell death, indicating that other mechanisms underlie the reported chemoresistance of tumors producing these factors. Our data suggest that a chemotherapy-mediated increase in tumor-promoting M2 macrophages may form an indirect mechanism for chemoresistance. Hence, concomitant therapy with COX inhibitors and/or IL-6R antibodies might increase the clinical effect of platinum-based chemotherapy in otherwise resistant tumor

A beneficial tumor microenvironment in oropharyngeal squamous cell carcinoma is characterized by a high T cell and low IL-17(+) cell frequency

textabstractPatients with HPV-positive oropharyngeal squamous cell carcinomas (OPSCCs) have a better prognosis than patients with non-HPV-induced OPSCC. The role of the immune response in this phenomenon is yet unclear. We studied the number of T cells, regulatory T cells (Tregs), T helper 17 (Th17) cells and IL-17+ non-T cells (mainly granulocytes) in matched HPV-positive and HPV-negative OPSCC cases (n = 162). Furthermore, the production of IFN-γ and IL-17 by tumor-infiltrating T cells was analyzed. The number of tumor-infiltrating T cells and Tregs was higher in HPV-positive than HPV-negative OPSCC (p < 0.0001). In contrast, HPV-negative OPSCC contained significantly higher numbers of IL-17+ non-T cells (p < 0.0001). Although a high number of intra-tumoral T cells showed a trend toward improved survival of all OPSCC patients, their prognostic effect in patients with a low number of intra-tumoral IL-17+ non-T cells was significant with regard to disease-specific (p = 0.033) and disease-free survival (p = 0.012). This suggests that a high frequency of IL-17+ non-T cells was related to a poor immune response, which was further supported by the observation that a high number of T cells was correlated with improved disease-free survival in the HPV-positive OPSCC (p = 0.008). In addition, we detected a minor Th17 cell population. However, T cells obtained from HPV-positive OPSCC produced significantly more IL-17 than those from HPV-negative tumors (p = 0.006). The improved prognosis of HPV-positive OPSCC is thus correlated with higher numbers of tumor-infiltrating T cells, more active Th17 cells and lower numbers of IL-17+ non-T cells

Intratumoral HPV16-specific T cells constitute a type I–oriented tumor microenvironment to improve survival in HPV16-driven oropharyngeal cancer

Purpose: Human papillomavirus (HPV)–associated oropharyngeal squamous cell cancer (OPSCC) has a much better prognosis than HPV-negative OPSCC, and this is linked to dense tumor immune infiltration. As the viral antigens may trigger potent immunity, we studied the relationship between the presence of intratumoral HPV-specific T-cell responses, the immune contexture in the tumor microenvironment, and clinical outcome. Experimental Design: To this purpose, an in-depth analysis of tumor-infiltrating immune cells in a prospective cohort of 97 patients with HPV16-positive and HPV16-negative OPSCC was performed using functional T-cell assays, mass cytometry (CyTOF), flow cytometry, and fluorescent immunostaining of tumor tissues. Key findings were validated in a cohort of 75 patients with HPV16-positive OPSCC present in the publicly available The Cancer Genome Atlas database. Results: In 64% of the HPV16-positive tumors, type I HPV16-specific T cells were present. Their presence was not only strongly related to a better overall survival, a smaller tumor size, and less lymph node metastases but also to a type I–oriented tumor microenvironment, including high numbers of activated CD161þ T cells, CD103þ tissue-resident T cells, dendritic cells (DC), and DC-like macrophages. Conclusions: The viral antigens trigger a tumor-specific T-cell response that shapes a favorable immune contexture for the response to standard therapy. Hence, reinforcement of HPV16-specific T-cell reactivity is expected to boost this process

An Unexpectedly Large Polyclonal Repertoire of HPV-Specific T Cells Is Poised for Action in Patients with Cervical Cancer

The diversity and extent of the local tumor-specific T-cell response in a given individual is largely unknown. We have performed an in-depth study of the local T-cell repertoire in a selected group of patients with cervical cancer, by systematic analyses of the proportion, breadth, and polarization of human papillomavirus (HPV) E6/E7-specific T cells within the total population of tumor-infiltrating lymphocytes (TIL) and tumor-draining lymph node cells (TDLNC). Isolated T cells were stimulated with sets of overlapping E6 and E7 peptides and analyzed by multiparameter flow cytometry with respect to activation, cytokine production, and T-cell receptor Vbeta usage. HPV-specific CD4+ and CD8+ T-cell responses were detected in TIL and TDLNC and their relative contribution varied between <1% and 66% of all T cells. In general, these HPV-specific responses were surprisingly broad, aimed at multiple E6 and E7 epitopes and involved multiple dominant and subdominant T-cell receptor Vbetas per single peptide-epitope. In most patients, only few IFNgamma-producing T cells were found and the amount of IFNgamma produced was low, suggesting that these are poised T cells, rendered functionally inactive within the tumor environment. Importantly, stimulation of the TIL and TDLNC with cognate antigen in the presence of commonly used Toll-like receptor ligands significantly enhanced the effector T-cell function. In conclusion, our study suggests that within a given patient with HPV-specific immunity many different tumor-specific CD4+ and CD8+ T cells are locally present and poised for action. This vast existing local T-cell population is awaiting proper stimulation and can be exploited for the immunotherapy of cance

A phase 1/2 study combining gemcitabine, Pegintron and p53 SLP vaccine in patients with platinum-resistant ovarian cancer

Purpose: Preclinical tumor models show that chemotherapy has immune modulatory properties which can be exploited in the context of immunotherapy. The purpose of this study was to determine the feasibility and immunogenicity of combinations of such an immunomodulatory chemotherapeutic agent with immunotherapy, p53 synthetic long peptide (SLP) vaccine and Pegintron (IFN-alpha) in patients with platinum-resistant p53-positive epithelial ovarian cancer (EOC). Experimental design: This is a phase 1/2 trial in which patients sequential 6 cycles of gemcitabine (1000 mg/kg(2) iv; n = 3), gemcitabine with Pegintron before and after the first gemcitabine cycle (Pegintron 1 mu g/kg sc; n = 6), and gemcitabine and Pegintron combined with p53 SLP vaccine (0.3 mg/peptide, 9 peptides; n = 6). At baseline, 22 days after the 2nd and 6th cycle, blood was collected for immunomonitoring. Toxicity, CA-125, and radiologic response were evaluated after 3 and 6 cycles of chemotherapy. Results: None of the patients enrolled experienced dose-limiting toxicity. Predominant grade 3/4 toxicities were nausea/vomiting and dyspnea. Grade 1/2 toxicities consisted of fatigue (78%) and Pegintron-related flu-like symptoms (72%). Gemcitabine reduced myeloid-derived suppressor cells (p = 0.0005) and increased immune-supportive M1 macrophages (p = 0.04). Combination of gemcitabine and Pegintron stimulated higher frequencies of circulating proliferating CD4+ and CD8+ T-cells but not regulatory T-cells. All vaccinated patients showed strong vaccine-induced p53-specific T-cell responses. Conclusion: Combination of gemcitabine, the immune modulator Pegintron and therapeutic peptide vaccination is a viable approach in the development of combined chemo-immunotherapeutic regimens to treat cancer

Intratumoral HPV16-Specific T Cells Constitute a Type I–Oriented Tumor Microenvironment to Improve Survival in HPV16-Driven Oropharyngeal Cancer

Transplantation and autoimmunit