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
Anogenital and oropharyngeal cancers caused by human papillomavirus (HPV) infections account for 4.5% of all cancer cases worldwide. So far, only the initial infection with selected high-risk types can be prevented by prophylactic vaccination. Already existing persistent HPV infections, however, can currently only be treated by surgical removal of resulting lesions. Therapeutic HPV vaccination, promoting cell-based anti-HPV immunity, would be ideal to eliminate and protect against HPV-induced lesions and tumors. A multitude of vaccination approaches has been tested to date, many of which led to high amounts of HPV-specific T cells in vivo. However, growing evidence suggests that not the induction of systemic but of local immunity is paramount for tackling mucosal infections and tumors. Therefore, recent therapeutic vaccination studies have focused on how to induce tissue-resident T cells in the anogenital and oropharyngeal mucosa. These approaches include direct mucosal vaccinations and influencing the migration of systemic T cells toward the mucosa. The efficacy of these new vaccination approaches is best tested in vivo by utilizing orthotopic tumor models, i.e. HPV-positive tumors being located in the animal's mucosa. In line with this, we here review existing HPV tumor models and describe two novel tumorigenic cell lines for the MHC-humanized mouse model A2.DR1. These were used for the establishment of an HPV16 E6/E7-positive vaginal tumor model, suitable for testing therapeutic vaccines containing HLA-A2-restricted HPV16-derived epitopes. The newly developed MHC-humanized orthotopic HPV16-positive tumor model is likely to improve the translatability of in vivo findings to the clinical setting.
Assuntos
Alphapapillomavirus/patogenicidade , Vacinas Anticâncer/uso terapêutico , Neoplasias Experimentais/prevenção & controle , Infecções por Papillomavirus/prevenção & controle , Vacinas contra Papillomavirus/uso terapêutico , Alphapapillomavirus/imunologia , Animais , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/virologia , Linhagem Celular Tumoral , Interações Hospedeiro-Patógeno , Humanos , Imunidade nas Mucosas , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/virologia , Camundongos Transgênicos , Neoplasias Experimentais/imunologia , Neoplasias Experimentais/virologia , Infecções por Papillomavirus/imunologia , Infecções por Papillomavirus/virologia , VacinaçãoRESUMO
Kaposi's sarcoma (KS), is an AIDS-associated neoplasm caused by the KS herpesvirus (KSHV/ HHV-8). KSHV-induced sarcomagenesis is the consequence of oncogenic viral gene expression as well as host genetic and epigenetic alterations. Although KSHV is found in all KS-lesions, the percentage of KSHV-infected (LANA+) spindle-cells of the lesion is variable, suggesting the existence of KS-spindle cells that have lost KSHV and proliferate autonomously or via paracrine mechanisms. A mouse model of KSHVBac36-driven tumorigenesis allowed us to induce KSHV-episome loss before and after tumor development. Although infected cells that lose the KSHV-episome prior to tumor formation lose their tumorigenicity, explanted tumor cells that lost the KSHV-episome remained tumorigenic. This pointed to the existence of virally-induced irreversible oncogenic alterations occurring during KSHV tumorigenesis supporting the possibility of hit and run viral-sarcomagenesis. RNA-sequencing and CpG-methylation analysis were performed on KSHV-positive and KSHV-negative tumors that developed following KSHV-episome loss from explanted tumor cells. When KSHV-positive cells form KSHV-driven tumors, along with viral-gene upregulation there is a tendency for hypo-methylation in genes from oncogenic and differentiation pathways. In contrast, KSHV-negative tumors formed after KSHV-episome loss, show a tendency towards gene hyper-methylation when compared to KSHV-positive tumors. Regarding occurrence of host-mutations, we found the same set of innate-immunity related mutations undetected in KSHV-infected cells but present in all KSHV-positive tumors occurring en exactly the same position, indicating that pre-existing host mutations that provide an in vivo growth advantage are clonally-selected and contribute to KSHV-tumorigenesis. In addition, KSHV-negative tumors display de novo mutations related to cell proliferation that, together with the PDGFRAD842V and other proposed mechanism, could be responsible for driving tumorigenesis in the absence of KSHV-episomes. KSHV-induced irreversible genetic and epigenetic oncogenic alterations support the possibility of "hit and run" KSHV-sarcomagenesis and point to the existence of selectable KSHV-induced host mutations that may impact AIDS-KS treatment.
Assuntos
Transformação Celular Viral , Metilação de DNA , Regulação Neoplásica da Expressão Gênica , Regulação Viral da Expressão Gênica , Herpesvirus Humano 8 , Neoplasias Experimentais , Plasmídeos , Sarcoma de Kaposi , Animais , Linhagem Celular , Herpesvirus Humano 8/genética , Herpesvirus Humano 8/metabolismo , Sequenciamento de Nucleotídeos em Larga Escala , Camundongos , Neoplasias Experimentais/genética , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Neoplasias Experimentais/virologia , Plasmídeos/genética , Plasmídeos/metabolismo , Sarcoma de Kaposi/genética , Sarcoma de Kaposi/metabolismo , Sarcoma de Kaposi/patologia , Sarcoma de Kaposi/virologiaRESUMO
BACKGROUND: In vivo targeting of human papillomavirus (HPV) derived antigens to dendritic cells might constitute an efficient immunotherapeutic strategy against cervical cancer. In previous works, we have shown that the extra domain A from murine fibronectin (mEDA) can be used to target antigens to toll-like receptor 4 (TLR4) expressing dendritic cells and induce strong antigen-specific immune responses. In the present study, we have produced a bivalent therapeutic vaccine candidate consisting of the human EDA (hEDA) fused to E7 proteins from HPV16 and HPV18 (hEDA-HPVE7-16/18) and evaluate its potential as a therapeutic vaccine against cervical cancer. MATERIALS AND METHODS: Recombinant fusion proteins containing HPV E7 proteins from HPV16 and HPV18 virus subtypes fused to hEDA were produced and tested in vitro on their capacity to bind TLR4 and induce the production of tumor necrosis factor-α or interleukin (IL)-12 by human monocytes and dendritic cells. The immunogenicity and potential therapeutic activity of the vaccine in combination with cisplatin or with the TLR3 agonist molecules polyinosinic-polycytidylic acid (Poly IC) or Poly ICLC was evaluated in mice bearing subcutaneous or genital orthotopic HPV16 TC-1 tumors. RESULTS: hEDA-HPVE7-16/18 prototype vaccine binds human TLR4 and stimulate TLR4-dependent signaling pathways and IL-12 production by human monocyte-derived dendritic cell. Vaccination with hEDA-HPVE7-16/18 induced strong HPVE7-specific Cytotoxic T lymphocyte (CTL) responses and eliminated established tumors in the TC-1-based tumor model. The antitumor efficacy was significantly improved by combining the fusion protein with cisplatin or with the TLR-3 ligand Poly IC and especially with the stabilized analog Poly ICLC. Moreover, hEDA-HPVE7-16/18+Poly ICLC induced full tumor regression in 100% of mice bearing orthotopic genital HPV tumors. CONCLUSION: Our results suggest that this therapeutic vaccine formulation may be an effective treatment for cervical tumors that do not respond to current therapies.
Assuntos
Vacinas Anticâncer/administração & dosagem , Proteínas de Ligação a DNA/imunologia , Fibronectinas/imunologia , Neoplasias Experimentais/prevenção & controle , Proteínas Oncogênicas Virais/imunologia , Proteínas E7 de Papillomavirus/imunologia , Infecções por Papillomavirus/prevenção & controle , Animais , Vacinas Anticâncer/imunologia , Células Dendríticas/imunologia , Feminino , Papillomavirus Humano 16/genética , Papillomavirus Humano 16/imunologia , Papillomavirus Humano 18/genética , Papillomavirus Humano 18/imunologia , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias Experimentais/imunologia , Neoplasias Experimentais/virologia , Infecções por Papillomavirus/imunologia , Infecções por Papillomavirus/virologia , Linfócitos T Citotóxicos/imunologiaRESUMO
BACKGROUND: Envisioned as a similar process to tumorigenesis in terms of biological behaviors and molecular basis, embryogenesis necessitates an immune surveillance system to eliminate erratically transformed cells. Our previous study demonstrated that fetal macrophage-like phagocytes triggered Th2-skewed immunity following endocytosing prenatally administered ovalbumin to facilitate postnatal allergic airway responses, highlighting the critical role fetal phagocytes played in dealing with antigens present in developing fetuses and shaping subsequent immune responses. It prompted us to examine whether fetuses could mount Th1 tumoricidal immunity against tumorigenesis following in utero exposure to tumor antigens. METHODS: Gestational day 14 murine fetuses underwent in utero injection of Th1-promoting human papilloma virus (HPV) E7 peptides. Postnatally, recipients were examined for immunological consequences and the resistance to TC-1 tumorigenesis. RESULTS: Fetal exposure to HPV E7 did not cause tolerance but rather immunization in the recipients, characterized by proinflammatory Th1 polarization of their lymphocytes. Fetal macrophage-like phagocytes were responsible for taking up HPV E7 and triggering HPV E7-specific T-cell cytotoxicity and humoral immunity that rendered recipients resistant to TC-1 tumorigenesis in postnatal life. Adoptive transfer of HPV E7-loaded fetal phagocytes also elicited Th1 immunity with rapid expansion of HPV E7-specific cytotoxic CD8+ T-cell clones in response to TC-1 cell challenge so as to protect the recipients from TC-1 tumorigenesis, but failed to completely eliminate pre-existing TC-1 cells despite perceptible attenuation of local TC-1 tumor growth. CONCLUSIONS: Our study revealed that Th2-biasing fetus was not immune-privileged to foreign peptides, but competent to mount Th1 cytotoxic immunity and generate immunoglobulins against tumorigenesis following in utero exposure to Th1-promoting oncoantigen. It shed light on the role of fetal macrophage-like phagocytes in bridging toward tumor antigen-specific cellular and humoral immunity potentially as an immune surveillance system to eliminate transformed cells that might be egressing during embryogenesis and leftover until postnatal life.
Assuntos
Imunidade Adaptativa/imunologia , Antígenos de Neoplasias/imunologia , Feto/imunologia , Neoplasias Experimentais/imunologia , Papillomaviridae/imunologia , Proteínas E7 de Papillomavirus/imunologia , Infecções por Papillomavirus/complicações , Animais , Feminino , Feto/virologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias Experimentais/prevenção & controle , Neoplasias Experimentais/virologia , Infecções por Papillomavirus/virologia , GravidezRESUMO
Viruses are associated with up to 15% of human cancer. MicroRNAs (miRNAs) encoded by numerous oncogenic viruses including Kaposi's sarcoma-associated herpesvirus (KSHV) play significant roles in regulating the proliferation and survival of virus-induced cancer cells, hence representing attractive therapeutic targets. Here, we report that specific inhibition of viral miRNAs by carbon dots (Cdots)-mediated delivery of locked nucleic acid (LNA)-based suppressors inhibit the proliferation of KSHV-associated primary effusion lymphoma (PEL) cells. Specifically, a combination of Cdots-LNAs to knock down the levels of KSHV miR-K12-1, miR-K12-4, and miR-K12-11 induces apoptosis and inhibits proliferation of PEL cells. Significantly, these Cdots-LNAs effectively inhibit the initiation of PEL and regress established PEL in a xenograft mouse model. These results demonstrate the feasibility of using Cdots to deliver miRNA suppressors for targeting viral cancers. Our study with viral miRNAs as targets may provide the scientific basis for using antisense drugs for human cancers associated with oncogenic viruses.
Assuntos
Antineoplásicos/farmacologia , Carbono/química , Herpesvirus Humano 8/química , Linfoma/tratamento farmacológico , Oligonucleotídeos/farmacologia , RNA Viral/antagonistas & inibidores , Animais , Antineoplásicos/química , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Linfoma/patologia , Linfoma/virologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos NOD , Camundongos SCID , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/patologia , Neoplasias Experimentais/virologia , Oligonucleotídeos/química , Tamanho da Partícula , Pontos Quânticos/química , Ratos , Propriedades de SuperfícieRESUMO
Cytomegalovirus (CMV) has been implicated in glioblastoma (GBM); however, a mechanistic connection in vivo has not been established. The purpose of this study is to characterize the effects of murine CMV (MCMV) on GBM growth in murine models. Syngeneic GBM models were established in mice perinatally infected with MCMV. We found that tumor growth was markedly enhanced in MCMV+ mice, with a significant reduction in overall survival compared with that of controls (P < 0.001). We observed increased angiogenesis and tumor blood flow in MCMV+ mice. MCMV reactivation was observed in intratumoral perivascular pericytes and tumor cells in mouse and human GBM specimens, and pericyte coverage of tumor vasculature was strikingly augmented in MCMV+ mice. We identified PDGF-D as a CMV-induced factor essential for pericyte recruitment, angiogenesis, and tumor growth. The antiviral drug cidofovir improved survival in MCMV+ mice, inhibiting MCMV reactivation, PDGF-D expression, pericyte recruitment, and tumor angiogenesis. These data show that MCMV potentiates GBM growth in vivo by increased pericyte recruitment and angiogenesis due to alterations in the secretome of CMV-infected cells. Our model provides evidence for a role of CMV in GBM growth and supports the application of antiviral approaches for GBM therapy.
Assuntos
Infecções por Citomegalovirus , Citomegalovirus/metabolismo , Glioblastoma , Neoplasias Experimentais , Neovascularização Patológica , Pericitos , Animais , Linhagem Celular Tumoral , Infecções por Citomegalovirus/metabolismo , Infecções por Citomegalovirus/patologia , Glioblastoma/irrigação sanguínea , Glioblastoma/metabolismo , Glioblastoma/patologia , Glioblastoma/virologia , Humanos , Linfocinas/metabolismo , Camundongos , Células NIH 3T3 , Proteínas de Neoplasias/metabolismo , Neoplasias Experimentais/irrigação sanguínea , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Neoplasias Experimentais/virologia , Neovascularização Patológica/metabolismo , Neovascularização Patológica/patologia , Neovascularização Patológica/virologia , Pericitos/metabolismo , Pericitos/patologia , Fator de Crescimento Derivado de Plaquetas/metabolismoRESUMO
Stem cells are maintained by transcriptional programs that promote self-renewal and repress differentiation. Here, we found that the transcription factor c-Myb was essential for generating and maintaining stem cells in the CD8+ T cell memory compartment. Following viral infection, CD8+ T cells lacking Myb underwent terminal differentiation and generated fewer stem cell-like central memory cells than did Myb-sufficient T cells. c-Myb acted both as a transcriptional activator of Tcf7 (which encodes the transcription factor Tcf1) to enhance memory development and as a repressor of Zeb2 (which encodes the transcription factor Zeb2) to hinder effector differentiation. Domain-mutagenesis experiments revealed that the transactivation domain of c-Myb was necessary for restraining differentiation, whereas its negative regulatory domain was critical for cell survival. Myb overexpression enhanced CD8+ T cell memory formation, polyfunctionality and recall responses that promoted curative antitumor immunity after adoptive transfer. These findings identify c-Myb as a pivotal regulator of CD8+ T cell stemness and highlight its therapeutic potential.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Memória Imunológica/imunologia , Neoplasias Experimentais/imunologia , Proteínas Proto-Oncogênicas c-myb/imunologia , Células-Tronco/imunologia , Animais , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/virologia , Diferenciação Celular/imunologia , Linhagem Celular Tumoral , Células HEK293 , Humanos , Memória Imunológica/genética , Coriomeningite Linfocítica/imunologia , Coriomeningite Linfocítica/metabolismo , Coriomeningite Linfocítica/virologia , Vírus da Coriomeningite Linfocítica/imunologia , Vírus da Coriomeningite Linfocítica/fisiologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/virologia , Proteínas Proto-Oncogênicas c-myb/genética , Proteínas Proto-Oncogênicas c-myb/metabolismo , Células-Tronco/metabolismo , Células-Tronco/virologia , Fator 1 de Transcrição de Linfócitos T/genética , Fator 1 de Transcrição de Linfócitos T/imunologia , Fator 1 de Transcrição de Linfócitos T/metabolismoRESUMO
Circular RNAs (circRNAs) are a novel class of non-coding RNA molecules in eukaryotic organisms that have potentially important roles in gene regulation. Nevertheless, whether viruses can encode circRNA is still uncertain. To examine whether large genome DNA viruses can generate circRNA during the infection of human cells, we performed RNA sequencing of ribosomal RNA-depleted total RNA from Epstein-Barr virus (EBV)-infected cell lines, including SNU-719, AGS-EBV, C666-1 and Akata. We identified an EBV-encoded circRNA, ebv_circ_RPMS1, that consists of the head-to-tail splicing of exons 2-4 from the RPMS1 gene. Furthermore, we demonstrated that ebv_circ_RPMS1 was localized in both cytoplasm and nuclei. Given that circRNAs shape gene expression by titrating microRNAs, regulating transcription and/or interfering with splicing, we identified a novel viral regulator of host and/or viral gene expression.
Assuntos
Herpesvirus Humano 4/genética , RNA não Traduzido/genética , RNA Viral/genética , Animais , Linhagem Celular , Feminino , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Neoplasias Experimentais/virologiaRESUMO
PURPOSE: Over 95% of human anal cancers are etiologically associated with high-risk HPVs, with HPV type 16 (HPV16) the genotype most commonly found. Activating mutations in the catalytic subunit of Phosphatidylinositol (3,4,5)-trisphosphate kinase (PI3K), encoded by the Pik3ca gene, are detected in approximately 20% of human anal cancers.Experimental Design: We asked if common activating mutations in Pik3ca contribute to anal carcinogenesis using an established mouse model for anal carcinogenesis in which mice are topically treated with the chemical carcinogen 7,12-Dimethylbenz(a)anthracene (DMBA). Mice expressing in their anal epithelium one of two activating mutations in Pik3ca genes, Pik3caH1047R or Pik3caE545K , were monitored for anal carcinogenesis in the presence or absence of transgenes expressing the HPV16 E6 and E7 oncogenes. RESULTS: Both mutant forms of Pik3ca increased susceptibility to anal carcinogenesis in the absence of HPV16 oncogenes, and cooperated with HPV16 oncogenes to induce the highest level and earliest onset of anal cancers. The combination of HPV16 oncogenes and Pik3ca mutations led to anal cancers even in the absence of treatment with DMBA. We further observed that the investigational mTOR1/2 dual inhibitor, TAK-228, significantly reduced the size of anal cancer-derived tumor spheroids in vitro and reduced the growth rates of anal cancer-derived tumor grafts in vivo. CONCLUSIONS: These data demonstrate that activating mutations in Pik3ca drive anal carcinogenesis together with HPV16 oncogenes, and that the PI3K/mTOR pathway is a relevant target for therapeutic intervention.
Assuntos
Neoplasias do Ânus/genética , Carcinogênese/genética , Classe I de Fosfatidilinositol 3-Quinases/genética , Papillomavirus Humano 16/patogenicidade , Neoplasias Experimentais/genética , 9,10-Dimetil-1,2-benzantraceno/toxicidade , Canal Anal/efeitos dos fármacos , Canal Anal/patologia , Animais , Neoplasias do Ânus/induzido quimicamente , Neoplasias do Ânus/tratamento farmacológico , Neoplasias do Ânus/virologia , Benzoxazóis/administração & dosagem , Carcinogênese/efeitos dos fármacos , Carcinógenos/toxicidade , Classe I de Fosfatidilinositol 3-Quinases/metabolismo , Feminino , Mutação com Ganho de Função , Humanos , Camundongos , Camundongos Transgênicos , Neoplasias Experimentais/induzido quimicamente , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/virologia , Cultura Primária de Células , Pirimidinas/administração & dosagem , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Serina-Treonina Quinases TOR/antagonistas & inibidores , Serina-Treonina Quinases TOR/metabolismo , Células Tumorais CultivadasRESUMO
Mouse models have been instrumental in establishing fundamental principles of cancer initiation and progression and continue to be invaluable in the discovery and further development of cancer therapies. Nevertheless, important aspects of human disease are imperfectly approximated in mouse models, notably the involvement of endogenous retroviruses (ERVs). Replication-defective ERVs, present in both humans and mice, may affect tumor development and antitumor immunity through mechanisms not involving infection. Here, we revealed an adverse effect of murine ERVs with restored infectivity on the behavior of mouse cancer models. In contrast to human cancer, where infectious ERVs have never been detected, we found that ERV infectivity was frequently restored in transplantable, as well as genetic, mouse cancer models. Such replication-competent, ERV-derived retroviruses were responsible for unusually high expression of retroviral nucleic acids and proteins in mouse cancers. Infectious ERV-derived retroviruses produced by mouse cancer cells could directly infect tumor-infiltrating host immune cells and fundamentally modified the host's immune defenses to cancer, as well as the outcome of immunotherapy. Therefore, infectious retroviruses, variably arising in mouse cancer models, but not in human cancer, have the potential to confound many immunologic studies and should be considered as a variable, if not altogether avoided. Cancer Immunol Res; 6(11); 1292-300. ©2018 AACR.
Assuntos
Retrovirus Endógenos/patogenicidade , Neoplasias Experimentais/imunologia , Neoplasias Experimentais/virologia , Animais , Linhagem Celular Tumoral , Feminino , Vírus da Leucemia Murina/genética , Vírus da Leucemia Murina/patogenicidade , Linfócitos do Interstício Tumoral/patologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Camundongos Transgênicos , Neoplasias Experimentais/patologia , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética , Proteínas Proto-Oncogênicas B-raf/genética , Infecções por Retroviridae/virologia , Tropismo Viral/fisiologiaRESUMO
High numbers of mast cells populate the stroma of many types of neoplasms, including human papilloma virus-induced benign and malignant tumors in man and mouse. Equipped with numerous pattern recognition receptors and capable of executing important pro-inflammatory responses, mast cells are considered innate sentinels that significantly impact tumor biology. Mast cells were reported to promote human papilloma virus (HPV)-induced epithelial hyperproliferation and neo-angiogenesis in an HPV-driven mouse model of skin cancer. We analyzed HPV-induced epithelial hyperplasia and squamous cell carcinoma formation, as well as growth of tumors inoculated into the dermis, in mice lacking skin mast cells. Unexpectedly, the absence of mast cells had no effect on HPV-induced epithelial growth or angiogenesis, on growth kinetics of inoculated tumors, or on the immunological tumor micro-milieu. Thus, the conspicuous recruitment of mast cells into tumor tissues cannot necessarily be equated with important mast cell functions in tumor growth.
Assuntos
Proliferação de Células , Transformação Celular Viral/imunologia , Mastócitos , Neoplasias Experimentais , Neovascularização Patológica , Papillomaviridae/imunologia , Animais , Linhagem Celular , Células Epiteliais/imunologia , Células Epiteliais/patologia , Células Epiteliais/virologia , Mastócitos/imunologia , Mastócitos/patologia , Camundongos , Camundongos Transgênicos , Transplante de Neoplasias , Neoplasias Experimentais/irrigação sanguínea , Neoplasias Experimentais/imunologia , Neoplasias Experimentais/patologia , Neoplasias Experimentais/virologia , Neovascularização Patológica/imunologia , Neovascularização Patológica/patologia , Neovascularização Patológica/virologiaRESUMO
Mouse papillomavirus has shown broad tissue tropism in nude mice. Previous studies have tested cutaneous infections in different immunocompromised and immunocompetent mouse strains. In the current study, we examined mucosal infection in several immunocompetent and immunocompromised mouse strains. Viral DNA was monitored periodically by Q-PCR of lavage samples. Immunohistochemistry and in situ hybridization were used to determine viral capsid protein and viral DNA respectively. All athymic nude mouse strains showed active infections at both cutaneous and mucosal sites. Interestingly, NOD/SCID mice, which have a deficiency in T, B, and NK cells, showed minimal disease at cutaneous sites but developed persistent infection at the mucosal sites including those of the anogenital region and the oral cavity. Three strains of immunocompetent mice supported mucosal infections. Infections of the lower genital tract in heterozygous (immunocompetent) mice of the NU/J strain progressed to high grade dysplasia and to carcinoma in situ. Anti-MmuPV1 neutralizing antibodies were detected in the sera of all immunocompetent animals. Our findings demonstrate that the mucosae may be the preferred sites for this virus in mice. The mouse model is expected to be a valuable model for the study of mucosal papillomavirus disease, progression, and host immune control.
Assuntos
Doenças da Boca/virologia , Mucosa/virologia , Infecções por Papillomavirus/imunologia , Animais , Anticorpos Neutralizantes/imunologia , DNA Viral/análise , Modelos Animais de Doenças , Feminino , Heterozigoto , Homozigoto , Interferon-alfa/genética , Camundongos Pelados , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos Mutantes , Doenças da Boca/imunologia , Doenças da Boca/patologia , Mucosa/patologia , Neoplasias Experimentais/virologia , Papillomaviridae/genética , Papillomaviridae/patogenicidade , Infecções por Papillomavirus/patologia , Dermatopatias Infecciosas/virologiaRESUMO
Glioblastoma is a highly lethal brain cancer that frequently recurs in proximity to the original resection cavity. We explored the use of oncolytic virus therapy against glioblastoma with Zika virus (ZIKV), a flavivirus that induces cell death and differentiation of neural precursor cells in the developing fetus. ZIKV preferentially infected and killed glioblastoma stem cells (GSCs) relative to differentiated tumor progeny or normal neuronal cells. The effects against GSCs were not a general property of neurotropic flaviviruses, as West Nile virus indiscriminately killed both tumor and normal neural cells. ZIKV potently depleted patient-derived GSCs grown in culture and in organoids. Moreover, mice with glioblastoma survived substantially longer and at greater rates when the tumor was inoculated with a mouse-adapted strain of ZIKV. Our results suggest that ZIKV is an oncolytic virus that can preferentially target GSCs; thus, genetically modified strains that further optimize safety could have therapeutic efficacy for adult glioblastoma patients.
Assuntos
Neoplasias Encefálicas/virologia , Glioblastoma/virologia , Células-Tronco Neoplásicas/virologia , Terapia Viral Oncolítica/métodos , Zika virus/fisiologia , Animais , Antineoplásicos Alquilantes/uso terapêutico , Neoplasias Encefálicas/terapia , Linhagem Celular Tumoral , Proliferação de Células , Chlorocebus aethiops , Terapia Combinada , Dacarbazina/análogos & derivados , Dacarbazina/uso terapêutico , Feminino , Imunofluorescência , Glioblastoma/terapia , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias Experimentais/terapia , Neoplasias Experimentais/virologia , Temozolomida , Células VeroRESUMO
Second mitochondrial activator of caspase (Smac)-mimetic compounds and oncolytic viruses were developed to kill cancer cells directly. However, Smac-mimetic compound and oncolytic virus therapies also modulate host immune responses in ways we hypothesized would complement one another in promoting anticancer T-cell immunity. We show that Smac-mimetic compound and oncolytic virus therapies synergize in driving CD8+ T-cell responses toward tumors through distinct activities. Smac-mimetic compound treatment with LCL161 reinvigorates exhausted CD8+ T cells within immunosuppressed tumors by targeting tumor-associated macrophages for M1-like polarization. Oncolytic virus treatment with vesicular stomatitis virus (VSVΔM51) promotes CD8+ T-cell accumulation within tumors and CD8+ T-cell activation within the tumor-draining lymph node. When combined, LCL161 and VSVΔM51 therapy engenders CD8+ T-cell-mediated tumor control in several aggressive mouse models of cancer. Smac-mimetic compound and oncolytic virus therapies are both in clinical development and their combination therapy represents a promising approach for promoting anticancer T-cell immunity.Oncolytic viruses (OV) and second mitochondrial activator of caspase (Smac)-mimetic compounds (SMC) synergistically kill cancer cells directly. Here, the authors show that SMC and OV therapies combination also synergize in vivo by promoting anticancer immunity through an increase in CD8+ T-cell response.
Assuntos
Materiais Biomiméticos/farmacologia , Linfócitos T CD8-Positivos/efeitos dos fármacos , Neoplasias Experimentais/terapia , Terapia Viral Oncolítica/métodos , Animais , Antineoplásicos/farmacologia , Proteínas Reguladoras de Apoptose , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/virologia , Linhagem Celular Tumoral , Terapia Combinada , Feminino , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Proteínas Mitocondriais/metabolismo , Neoplasias Experimentais/imunologia , Neoplasias Experimentais/virologia , Vírus Oncolíticos/imunologia , Vírus Oncolíticos/fisiologia , Tiazóis/farmacologia , Resultado do Tratamento , Vírus da Estomatite Vesicular Indiana/imunologia , Vírus da Estomatite Vesicular Indiana/fisiologiaRESUMO
Oncolytic virotherapy is being developed as a promising platform for cancer therapy due to its ability to lyse cancer cells in a tumor-specific manner. Vaccinia virus has been used as a live vaccine in the smallpox eradication program and now is being potential in cancer therapy with a great safety profile. Vaccinia strain Guang9 (VG9) is an attenuated Chinese vaccinia virus and its oncolytic efficacy has been evaluated in our previous study. To improve the tumor selectivity and oncolytic efficacy, we here developed a thymidine kinase (TK)-deleted vaccinia virus based on Guang9 strain. The viral replication, marker gene expression and cytotoxicity in various cell lines were evaluated; antitumor effects in vivo were assessed in multiple tumor models. In vitro, the TK-deleted vaccinia virus replicated rapidly, but the cytotoxicity varied in different cell lines. It was notably attenuated in normal cells and resting cells in vitro, while tumor-selectively replicated in vivo. Significant antitumor effects were observed both in murine melanoma tumor model and human hepatoma tumor model. It significantly inhibited the growth of subcutaneously implanted tumors and prolonged the survival of tumor-bearing mice. Collectively, TK-deleted vaccinia strain Guang9 is a promising constructive virus vector for tumor-directed gene therapy and will be a potential therapeutic strategy in cancer treatment.
Assuntos
Neoplasias Experimentais/terapia , Terapia Viral Oncolítica/métodos , Vírus Oncolíticos/genética , Timidina Quinase/genética , Vaccinia virus/genética , Ensaios Antitumorais Modelo de Xenoenxerto , Células A549 , Animais , Linhagem Celular , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Chlorocebus aethiops , Feminino , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Nus , Células NIH 3T3 , Neoplasias Experimentais/genética , Neoplasias Experimentais/virologia , Vírus Oncolíticos/fisiologia , Timidina Quinase/deficiência , Resultado do Tratamento , Vaccinia virus/fisiologia , Células VeroRESUMO
Background: Glioblastoma multiforme and high-risk neuroblastoma are cancers with poor outcome. Immunotherapy in the form of neurotropic oncolytic viruses is a promising therapeutic approach for these malignancies. Here we evaluate the oncolytic capacity of the neurovirulent and partly IFNß-resistant Semliki Forest virus (SFV)-4 in glioblastoma multiformes and neuroblastomas. To reduce neurovirulence we constructed SFV4miRT, which is attenuated in normal central nervous system (CNS) cells through insertion of microRNA target sequences for miR124, miR125, miR134.Methods: Oncolytic activity of SFV4miRT was examined in mouse neuroblastoma and glioblastoma multiforme cell lines and in patient-derived human glioblastoma cell cultures (HGCC). In vivo neurovirulence and therapeutic efficacy was evaluated in two syngeneic orthotopic glioma models (CT-2A, GL261) and a syngeneic subcutaneous neuroblastoma model (NXS2). The role of IFNß in inhibiting therapeutic efficacy was investigated.Results: The introduction of miRNA target sequences reduced neurovirulence of SFV4 in terms of attenuated replication in mouse CNS cells and ability to cause encephalitis when administered intravenously. A single intravenous injection of SFV4miRT prolonged survival and cured four of eight mice (50%) with NXS2 and three of 11 mice (27%) with CT-2A, but not for GL261 tumor-bearing mice. In vivo therapeutic efficacy in different tumor models inversely correlated to secretion of IFNß by respective cells upon SFV4 infection in vitro Similarly, killing efficacy of HGCC lines inversely correlated to IFNß response and interferon-α/ß receptor-1 expression.Conclusions: SFV4miRT has reduced neurovirulence, while retaining its oncolytic capacity. SFV4miRT is an excellent candidate for treatment of glioblastoma multiforme and neuroblastoma with low IFN-ß secretion. Clin Cancer Res; 23(6); 1519-30. ©2016 AACR.
Assuntos
Glioblastoma/terapia , Neoplasias Experimentais/terapia , Neuroblastoma/terapia , Vírus Oncolíticos/genética , Animais , Sistema Nervoso Central/virologia , Modelos Animais de Doenças , Glioblastoma/genética , Glioblastoma/virologia , Humanos , Interferon beta/genética , Camundongos , MicroRNAs/genética , MicroRNAs/uso terapêutico , Neoplasias Experimentais/genética , Neoplasias Experimentais/virologia , Neuroblastoma/virologia , Terapia Viral Oncolítica/efeitos adversos , Vírus da Floresta de Semliki/genéticaRESUMO
Oncolytic virus (OV) therapy has emerged as a novel tool in our therapeutic arsenals for fighting cancer. As a live biologic agent, OV has the ability to target and selectively amplify at the tumor sites. We have reported that a vaccinia-based OV (Pexa-Vec) has shown good efficacy in preclinical models and in clinical trials. To give an additional tool to clinicians to allow both treatment of the tumor and improved visualization of tumor margins, we developed new viral-based platforms with 2 specific gene reporters. METHODS: We incorporated the human sodium iodide symporter (hNIS) and the human somatostatin receptor 2 (hSSR2) in the vaccinia-based OV and tested viral constructs for their abilities to track and treat tumor development in vivo. RESULTS: Early and high-level expression of hNIS is detrimental to the recombinant virus, leading to the aggregation of hNIS protein and early cell death. Putting hNIS under a late synthetic promoter allowed a higher functional expression of the protein and much stronger 123I or 99Tc uptake. In vivo, the hNIS-containing virus infected and amplified in the tumor site, showing a better efficacy than the parental virus. The hNIS expression at the tumor site allowed for the imaging of viral infection and tumor regression. Similarly, hSSR2-containing OV vaccinia infected and lysed cancer cells. CONCLUSION: When tumor-bearing mice were given hNIS- and hSSR2-containing OV, 99Tc and 111In signals coalesced at the tumor, highlighting the power of using these viruses for tumor diagnosis and treatment.
Assuntos
Neoplasias Experimentais/diagnóstico por imagem , Neoplasias Experimentais/terapia , Terapia Viral Oncolítica/métodos , Receptores de Somatostatina/genética , Simportadores/genética , Vaccinia virus/fisiologia , Animais , Linhagem Celular Tumoral , Feminino , Genes Reporter/genética , Humanos , Camundongos , Camundongos Nus , Neoplasias Experimentais/virologia , Vírus Oncolíticos/fisiologia , Tomografia por Emissão de Pósitrons/métodos , Recombinação Genética/genética , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Nanomedicina Teranóstica/métodos , Resultado do Tratamento , Regulação para Cima/genéticaRESUMO
Two human papillomavirus (HPV) viral oncoproteins, E6 and E7 represent ideal targets for development of a therapeutic HPV vaccine. It is important to reduce the rate of HPV-associated malignancies through improvement of vaccine modalities. In this study, we used a short amphipathic peptide carrier, Pep-1, for delivery of the full-length HPV16 E7 protein into mammalian cells and evaluated immune responses and protective effects of different formulations in C57BL/6 tumor mice model. Our results showed that the complexes of E7/Pep-1 protein form stable nanoparticles through noncovalent binding with an average size of 120 to 250 nm. The efficient delivery of E7 protein by Pep-1 at molar ratio of 1:20 was detected in HEK-293T cell line for 1 h and 3 h post-transfection. Immunization with E7/Pep-1 nanoparticles at a ratio of 1:20 induced a higher Th1 cellular immune response with the predominant IgG2a and IFN-γ levels than those induced by E7 protein in a murine tumor model. These data suggest that Pep-1 peptide would indicate promising applications for improvement of HPV therapeutic vaccines. © 2016 IUBMB Life, 68(6):459-467, 2016.
Assuntos
Cisteamina/análogos & derivados , Nanopartículas/química , Proteínas E7 de Papillomavirus/administração & dosagem , Vacinas contra Papillomavirus/administração & dosagem , Peptídeos/química , Células Th1/imunologia , Animais , Cisteamina/administração & dosagem , Cisteamina/química , Citocinas/metabolismo , Sistemas de Liberação de Medicamentos/métodos , Feminino , Células HEK293 , Humanos , Imunidade Celular/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Nanopartículas/administração & dosagem , Neoplasias Experimentais/prevenção & controle , Neoplasias Experimentais/virologia , Proteínas E7 de Papillomavirus/imunologia , Vacinas contra Papillomavirus/imunologia , Vacinas contra Papillomavirus/farmacologia , Tamanho da Partícula , Peptídeos/administração & dosagem , Células Th1/efeitos dos fármacosRESUMO
UNLABELLED: Clinical development of a mesogenic strain of Newcastle disease virus (NDV) as an oncolytic agent for cancer therapy has been hampered by its select agent status due to its pathogenicity in avian species. Using reverse genetics, we have generated a lead candidate oncolytic NDV based on the mesogenic NDV-73T strain that is no longer classified as a select agent for clinical development. This recombinant NDV has a modification at the fusion protein (F) cleavage site to reduce the efficiency of F protein cleavage and an insertion of a 198-nucleotide sequence into the HN-L intergenic region, resulting in reduced viral gene expression and replication in avian cells but not in mammalian cells. In mammalian cells, except for viral polymerase (L) gene expression, viral gene expression is not negatively impacted or increased by the HN-L intergenic insertion. Furthermore, the virus can be engineered to express a foreign gene while still retaining the ability to grow to high titers in cell culture. The recombinant NDV selectively replicates in and kills tumor cells and is able to drive potent tumor growth inhibition following intratumoral or intravenous administration in a mouse tumor model. The candidate is well positioned for clinical development as an oncolytic virus. IMPORTANCE: Avian paramyxovirus type 1, NDV, has been an attractive oncolytic agent for cancer virotherapy. However, this virus can cause epidemic disease in poultry, and concerns about the potential environmental and economic impact of an NDV outbreak have precluded its clinical development. Here we describe generation and characterization of a highly potent oncolytic NDV variant that is unlikely to cause Newcastle disease in its avian host, representing an essential step toward moving NDV forward as an oncolytic agent. Several attenuation mechanisms have been genetically engineered into the recombinant NDV that reduce chicken pathogenicity to a level that is acceptable worldwide without impacting viral production in cell culture. The selective tumor replication of this recombinant NDV, both in vitro and in vivo, along with efficient tumor cell killing makes it an attractive oncolytic virus candidate that may provide clinical benefit to patients.