RESUMO
The generation of successful anticancer vaccines relies on the ability to induce efficient and long-lasting immune responses to tumor antigens. In this scenario, dendritic cells (DCs) are essential cellular components in the generation of antitumor immune responses. Thus, delivery of tumor antigens to specific DC populations represents a promising approach to enhance the efficiency of antitumor immunotherapies. In the present study, we employed antibody-antigen conjugates targeting a specific DC C-type lectin receptor. For that purpose, we genetically fused the anti-DEC205 monoclonal antibody to the type 16 human papillomavirus (HPV-16) E7 oncoprotein to create a therapeutic vaccine to treat HPV-associated tumors in syngeneic mouse tumor models. The therapeutic efficacy of the αDEC205-E7 mAb was investigated in three distinct anatomical tumor models (subcutaneous, lingual and intravaginal). The immunization regimen comprised two doses of the αDEC205-E7 mAb coadministered with a DC maturation stimulus (Polyinosinic:polycytidylic acid, poly (I:C)) as an adjuvant. The combined immunotherapy produced robust antitumor effects on both the subcutaneous and orthotopic tumor models, stimulating rapid tumor regression and long-term survival. These outcomes were related to the activation of tumor antigen-specific CD8+ T cells in both systemic compartments and lymphoid tissues. The αDEC205-E7 antibody plus poly (I:C) administration induced long-lasting immunity and controlled tumor relapses. Our results highlight that the delivery of HPV tumor antigens to DCs, particularly via the DEC205 surface receptor, is a promising therapeutic approach, providing new opportunities for the development of alternative immunotherapies for patients with HPV-associated tumors at different anatomical sites.
Assuntos
Antígenos CD/imunologia , Vacinas Anticâncer/administração & dosagem , Células Dendríticas/imunologia , Lectinas Tipo C/imunologia , Antígenos de Histocompatibilidade Menor/imunologia , Neoplasias Experimentais/prevenção & controle , Proteínas E7 de Papillomavirus/imunologia , Infecções por Papillomavirus/prevenção & controle , Receptores de Superfície Celular/imunologia , Adjuvantes Imunológicos , Animais , Linfócitos T CD8-Positivos/imunologia , Vacinas Anticâncer/imunologia , Feminino , Humanos , Memória Imunológica , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias Experimentais/imunologia , Neoplasias Experimentais/virologia , Infecções por Papillomavirus/imunologia , Infecções por Papillomavirus/virologia , Poli I-C/administração & dosagemRESUMO
Cervical cancer is a major public health problem and one of the leading causes of cancer deaths in women. Virtually all cases of cervical cancer, as well as a growing share of anal and head/neck tumors, are associated with human papillomavirus (HPV) infection. Despite the effectiveness, the available prophylactic vaccines do not benefit women with cervical lesions or cancer. Therefore, the search of new immunotherapeutic approaches to treat HPV-induced tumors is still a priority. The present study characterizes a therapeutic antitumor vaccine based on the genetic fusion of the Herpes simplex virus-1 (HSV-1) glycoprotein D (gD) with the E7 oncoprotein from HPV-16 (gDE7). Two subcutaneous doses of gDE7, admixed with poly (I:C), conferred complete and long-lasting therapeutic antitumor protection on mice previously challenged with tumor cells expressing the HPV-16 oncoproteins. The vaccine induced multifunctional E7-specific CD8+ T cells with cytotoxic activity and effector memory phenotype (CD44+ CD62Llow). In addition, gDE7 admixed with poly (I:C) vaccination controlled the expansion of tumor-induced regulatory T cells and myeloid-derived suppressor cells. More importantly, gDE7 activated mouse CD11c+ CD8α+ and human BDCA3+ dendritic cells (DC), specialized in antigen cross-presentation to CD8+ T cells, under in vitro conditions. These results indicated that the activation of a specific DC population, mediated by gD, improved the antigen-specific immune responses and the therapeutic performance induced by antitumor vaccines. These results open perspectives for the clinical testing of gDE7-based vaccines under the concept of active immunization as a tool for the therapeutic control of cancer. Mol Cancer Ther; 16(9); 1922-33. ©2017 AACR.
Assuntos
Vacinas Anticâncer/imunologia , Células Dendríticas/imunologia , Neoplasias/etiologia , Neoplasias/patologia , Papillomaviridae/imunologia , Infecções por Papillomavirus/complicações , Infecções por Papillomavirus/imunologia , Proteínas do Envelope Viral/imunologia , Animais , Apresentação de Antígeno/imunologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Linhagem Celular Tumoral , Apresentação Cruzada/imunologia , Células Dendríticas/metabolismo , Feminino , Humanos , Imunização , Memória Imunológica , Camundongos , Camundongos Knockout , Neoplasias/terapia , Proteínas E7 de Papillomavirus/imunologia , Poli I-C , Especificidade do Receptor de Antígeno de Linfócitos T , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismoRESUMO
One important goal of cancer immunotherapy is to prevent and treat tumor metastasis. We have previously reported the significant antitumor effect induced by the immunization with our human papillomavirus therapeutic protein-based vaccine (LALF32-51-E7) without adjuvant and admixed with clinically relevant adjuvants in the subcutaneous TC-1 tumor challenge model. In the present study, we evaluated the efficacy of the above mentioned vaccine formulations in controlling the hematogenous spread of TC-1 tumor cells using a more tumourigenic clone named TC-1* and other intravenous injection site less stressful than the tail vein. We generated a lung metastasis model by injecting TC-1* cells into the retro-orbital venous sinus and this is the first study describing it. Also, this is the first study that demonstrates the efficacy of the immunization with LALF32-51-E7 without adjuvant and admixed with VSSP or Al(OH)3 in controlling metastatic tumors increasing the survival of the mice. Our TC-1 lung metastasis model can be used to test the efficacy of other immunotherapeutic strategies based on E6/E7 antigens.
Assuntos
Imunoterapia , Neoplasias Pulmonares/secundário , Neoplasias Pulmonares/terapia , Proteínas E7 de Papillomavirus/imunologia , Vacinas contra Papillomavirus/uso terapêutico , Neoplasias do Colo do Útero/terapia , Animais , Feminino , Vetores Genéticos , Humanos , Neoplasias Pulmonares/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Proteolipídeos , Linfócitos T Citotóxicos , Células Tumorais Cultivadas , Neoplasias do Colo do Útero/imunologia , Neoplasias do Colo do Útero/patologiaRESUMO
A recombinant strain of Lactococcus lactis displaying a cell-surface anchored fibronectin binding protein A (FnBPA) from Staphylococcus aureus (LL-FnBPA) had been shown to be more efficient in delivering plasmid than its wild-type counterpart both in vitro and in vivo, and have the ability to orientate the immune response toward a Th2 profile in a context of a DNA vaccination. The aim of this work was to test whether this LL-FnBPA strain could shape the immune response after mucosal administration in mice. For this, we used a mouse model of human papilloma virus (HPV)-induced cancer and a L. lactis strain displaying at its cell surface both HPV-16-E7 antigen (LL-E7) and FnBPA (LL-E7+FnBPA). Our results revealed a more efficient systemic Th1 immune response with recombinant LL-E7+FnBPA. Furthermore, mice vaccinated with LL-E7+FnBPA were better protected when challenged with HPV-16-induced tumors. Altogether, the results suggest that FnBPA displays adjuvant properties when used in the context of mucosal delivery using L. lactis as a live vector.
Assuntos
Adesinas Bacterianas/imunologia , Vacinas Anticâncer/imunologia , Lactococcus lactis , Proteínas E7 de Papillomavirus/imunologia , Infecções por Papillomavirus/prevenção & controle , Animais , Células CACO-2 , Linhagem Celular Tumoral , Técnicas de Visualização da Superfície Celular , Feminino , Papillomavirus Humano 16 , Humanos , Imunidade Celular , Imunidade Humoral , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Plasmídeos , Staphylococcus aureus , Linfócitos T Citotóxicos/imunologia , Células Th1/imunologiaRESUMO
The human papillomavirus (HPV)16 E6 and E7 correlation with chemokine ligand (CCL)20 expression and Langerhans cells (LCs) in cervical lesions was investigated. We enrolled 43 patients with surgically treated cervical lesions from the Department of Gynecology in our hospital, and 20 controls without cervical lesions. Subjects were divided by pathology: HPV16(-) and HPV16(+) normal cervical groups (N = 10 each), and HPV16(+) cervical intraepithelial neoplasia (CIN), cervical invasive carcinoma (N = 15 each), and in situ carcinoma (N = 13) groups. E6, E7, the LC surface marker CD1a, and CCL20 were analyzed by immunohistochemistry. E6 and E7 in HPV16-type lesions were correlated with CCL20 and LCs. The average high power field cell numbers of CD1a+ LCs in the HPV(-) and HPV(+) normal cervix groups, and the CINI-II, CINIII in situ and cervical carcinoma groups were 22.89 ± 4.84, 13.7 ± 2.26, 9.2 ± 1.68, 5.9 ± 1.59, and 5.5 ± 1.58, respectively. Significant between-group differences existed except between cervical carcinoma and CINIII groups (P < 0.05). CCL20+ rates in each group were 70, 60, 60, 15.38, and 13.33%, respectively. E6/E7-positive expression rates in each group were 20/20, 66.7/66.7, 76.9/69.2, and 86.67/73.3%, respectively. CCL20 was positively correlated with CD1a (r = 0.649), and negatively correlated with E7 (r = -0.946) and E6 (r = -0.949). CD1a was negatively correlated with E6 (r = -0.632) and E7 (r = -0.632). Downregulation of CCL20 leading to LC decline is a key factor in cervical lesions. High-risk HPV-type lesions might inhibit the chemokine CCL20 through E6 and E7 to escape the immune response.
Assuntos
Carcinoma de Células Escamosas/genética , Quimiocina CCL20/genética , Papillomavirus Humano 16/genética , Proteínas Oncogênicas Virais/genética , Proteínas E7 de Papillomavirus/genética , Infecções por Papillomavirus/genética , Proteínas Repressoras/genética , Displasia do Colo do Útero/genética , Neoplasias do Colo do Útero/genética , Adulto , Antígenos CD1/genética , Antígenos CD1/imunologia , Carcinoma de Células Escamosas/imunologia , Carcinoma de Células Escamosas/patologia , Carcinoma de Células Escamosas/virologia , Estudos de Casos e Controles , Colo do Útero/imunologia , Colo do Útero/patologia , Colo do Útero/virologia , Quimiocina CCL20/imunologia , Feminino , Regulação da Expressão Gênica , Papillomavirus Humano 16/imunologia , Papillomavirus Humano 16/patogenicidade , Humanos , Evasão da Resposta Imune , Células de Langerhans/imunologia , Células de Langerhans/patologia , Células de Langerhans/virologia , Pessoa de Meia-Idade , Proteínas Oncogênicas Virais/imunologia , Proteínas E7 de Papillomavirus/imunologia , Infecções por Papillomavirus/imunologia , Infecções por Papillomavirus/patologia , Infecções por Papillomavirus/virologia , Proteínas Repressoras/imunologia , Transdução de Sinais , Neoplasias do Colo do Útero/imunologia , Neoplasias do Colo do Útero/patologia , Neoplasias do Colo do Útero/virologia , Displasia do Colo do Útero/imunologia , Displasia do Colo do Útero/patologia , Displasia do Colo do Útero/virologiaRESUMO
Cervical cancer is a common type of cancer among women worldwide and infection with high-risk human papillomavirus (HPVs) types represents the major risk factor for the etiopathogenesis of the disease. HPV-16 is the most frequently identified HPV type in cervical lesions and expression of E6 and E7 oncoproteins is required for the uncontrolled cellular proliferation. In the present study we report the design and experimental testing of a recombinant multi-epitope protein containing immunogenic epitopes of HPV-16 E6 and E7. Tumor preventive assays, based on the engraftment of TC-1 cells in mice, showed that the E6E7 multi-epitope protein induced a full preventive anti-tumor protection in wild-type mice, as well as in mice deficient in expression of CD4+ T cells and TLR4 receptor. Nonetheless, no anti-tumor protection was observed in mice deficient in CD8+ T cells. Also, the vaccine promoted high activation of E6/E7-specific T cells and in a therapeutic-approach, E6E7 protein conferred full anti-tumor protection in mice. These results show a potential use of this E6E7 multi-epitope antigen as a new and promising antigen for the development of a therapeutic vaccine against tumors induced by HPV.
Assuntos
Alphapapillomavirus/imunologia , Vacinas Anticâncer/imunologia , Epitopos/imunologia , Proteínas Oncogênicas Virais/imunologia , Proteínas E7 de Papillomavirus/imunologia , Proteínas Repressoras/imunologia , Neoplasias do Colo do Útero/prevenção & controle , Sequência de Aminoácidos , Animais , Linhagem Celular , Epitopos/química , Feminino , Interferon gama/biossíntese , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Dados de Sequência MolecularRESUMO
Recently, Bacillus subtilis spores were shown to be endowed with strong adjuvant capacity when co-administered with purified antigenic proteins. In the present study we assessed whether spores possess adjuvant properties when combined with DNA vaccines. We showed that B. subtilis spores promoted the activation of dendritic cells in vitro and induced migration of pro-inflammatory cells after parenteral administration to mice. Likewise, co-administration of spores with a DNA vaccine encoding the human papillomavirus type 16 (HPV-16) E7 protein enhanced the activation of antigen-specific CD8(+) T cell responses in vivo. Mice immunized with the DNA vaccine admixed with spores presented a protective immunity increase to previously implanted tumor cells, capable of expressing HPV-16 oncoproteins. Finally, we observed that the adjuvant effect can vary accordingly to the number of co-administered spores which may be ascribed with the ability to induce. Collectively, the present results demonstrate for the first time that B. subtilis spores can also confer adjuvant effects to DNA vaccines.
Assuntos
Adjuvantes Imunológicos/administração & dosagem , Bacillus subtilis/imunologia , Esporos Bacterianos/imunologia , Vacinas de DNA/imunologia , Animais , Bacillus subtilis/fisiologia , Linfócitos T CD8-Positivos/imunologia , Linhagem Celular Tumoral , Citocinas/imunologia , Células Dendríticas/imunologia , Ensaio de Imunoadsorção Enzimática , Humanos , Interferon gama/imunologia , Masculino , Camundongos Endogâmicos C57BL , Proteínas E7 de Papillomavirus/genética , Proteínas E7 de Papillomavirus/imunologia , Vacinas de DNA/administração & dosagemRESUMO
Although current polyvalent vaccines can prevent development of cervical cancer, they cannot be used to treat patients who already have the disease. Adenovirus expressing calreticulin-E7 (Ad-CRT-E7) has shown promising results in the cervical cancer murine model. We also demonstrated that immunization with Lactococcus lactis encoding HPV-16 E7 (Ll-E7) anchored to its surface induces significant HPV-16 E7-specific immune response. Here, we assessed the combination of both approaches in the treatment of a cervical cancer animal model. Intranasal preimmunization of Ll-E7, followed by a single Ad-CRT/E7 application, induced â¼80% of tumor suppression in comparison with controls. Mice treated with a combination of Ll-E7 and Ad-CRT/E7 resulted in a 70% survival rate 300 days post-treatment, whereas 100% of the mice in the control groups died by 50 days. Significant CD8+ cytotoxic T-lymphocytes infiltration was detected in the tumors of mice treated with Ll-E7+Ad-CRT/E7. Tumors with regression showed a greater number of positive cells for in situ TUNEL staining than controls. Our results suggest that preimmunization with Ll-E7 enhances the Ad-CRT/E7-mediated antitumor effect. This treatment provides an enormous advantage over repeated applications of Ad-CRT/E7 by maintaining the effectiveness of the three-dose application of Ad-CRT/E7, but avoiding the high systemic toxicities associated with such repeat treatments.
Assuntos
Antineoplásicos/administração & dosagem , Imunoterapia/métodos , Proteínas E7 de Papillomavirus/administração & dosagem , Proteínas E7 de Papillomavirus/imunologia , Vacinas contra Papillomavirus/imunologia , Neoplasias do Colo do Útero/terapia , Vacinas Sintéticas/imunologia , Adenoviridae/genética , Animais , Linfócitos T CD8-Positivos/imunologia , Calreticulina/administração & dosagem , Técnicas de Visualização da Superfície Celular/métodos , Modelos Animais de Doenças , Portadores de Fármacos/administração & dosagem , Feminino , Vetores Genéticos , Lactococcus lactis/genética , Camundongos Endogâmicos C57BL , Proteínas E7 de Papillomavirus/genética , Vacinas contra Papillomavirus/administração & dosagem , Vacinas contra Papillomavirus/genética , Proteínas Recombinantes de Fusão/administração & dosagem , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Análise de Sobrevida , Resultado do Tratamento , Neoplasias do Colo do Útero/patologia , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/genéticaRESUMO
Despite recent advances in the development of novel therapies, esophageal carcinoma remains an aggressive cancer associated with a poor prognosis. The lack of a high throughput, reproducible syngeneic animal model that replicates human disease is partly responsible for the paucity of novel therapeutic approaches. In this report, we present the first successful syngeneic, orthotopic model for esophageal cancer. This model was used to test an established adenoviral-based tumor vaccine. We utilized a murine esophageal cancer cell line established from the ED-L2-cyclin D1;p53 mouse that was transduced to express a viral tumor antigen, the Human Papilloma Virus (HPV) E7 protein. The tumor was established in its natural microenvironment at the gastroesophageal junction. Tumor growth was consistent and reproducible. An adenoviral vaccine to E7 (Ad.E7) induced an E7-specific population of functionally active CD8 T cells that trafficked into the tumors and retained cytotoxicity. Ad.E7 vaccination reduced local tumor growth and prolonged overall survival. These findings suggest that orthotopic tumor growth is a reasonable preclinical model to validate novel therapies.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Vacinas Anticâncer , Carcinoma/terapia , Neoplasias Esofágicas/terapia , Imunoterapia/métodos , Proteínas E7 de Papillomavirus/metabolismo , Adenoviridae/genética , Animais , Carcinoma/imunologia , Processos de Crescimento Celular , Linhagem Celular Tumoral , Modelos Animais de Doenças , Neoplasias Esofágicas/imunologia , Feminino , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Proteínas E7 de Papillomavirus/genética , Proteínas E7 de Papillomavirus/imunologiaRESUMO
HPV L1-based virus-like particles vaccines (VLPs) efficiently induce temporary prophylactic activity through the induction of neutralizing antibodies; however, VLPs that can provide prophylactic as well as therapeutic properties for longer periods of time are needed. For this purpose, we generated a novel HPV 16 L1-based chimeric virus-like particle (cVLP) produced in plants that contains a string of T-cell epitopes from HPV 16 E6 and E7 fused to its C-terminus. In the present study, we analyzed the persistence of specific IgG antibodies with neutralizing activity induced by immunization with these cVLPs, as well as their therapeutic potential in a tumor model of C57BL/6 mice. We observed that these cVLPs induced persistent IgG antibodies for over 12 months, with reactivity and neutralizing activity for VLPs composed of only the HPV-16 L1 protein. Efficient protection for long periods of time and inhibition of tumor growth induced by TC-1 tumor cells expressing HPV-16 E6/E7 oncoproteins, as well as significant tumor reduction (57 %), were observed in mice immunized with these cVLPs. Finally, we discuss the possibility that chimeric particles of the type described in this work may be the basis for developing HPV prophylactic and therapeutic vaccines with high efficacy.