RESUMO
The Oral HIV/AIDS Research Alliance (OHARA) was established in 2006 to provide the capacity to investigate the oral complications associated with HIV/AIDS within the ACTG infrastructure. Its goals were to explore the effects of potent antiretroviral therapy (ART) on the development of opportunistic infections, and variation and resistance of opportunistic pathogens in the context of immune suppression and long-term ART. The objectives of this talk, presented as part of a plenary session at the 7th World Workshop on Oral Health and Disease in AIDS, were to (i) provide an overview of OHARA's most recent research agenda, and how it evolved since OHARA's inception; (ii) describe OHARA's main accomplishments, including examples of research protocols completed and their key findings; and (iii) describe spin-off projects derived from OHARA, lessons learned, and future directions. OHARA has met its central goal and made key contributions to the field in several ways: (i) by developing/updating diagnostic criteria for oral disease endpoints commonly measured in OHARA protocols and in HIV/AIDS research in general and has creating standardized training modules, both for measuring these oral disease endpoints across clinical specialties, and for collecting oral fluid specimens; (ii) by implementing a total of nine protocols, six of which are completed. Three protocols involved domestic research sites, while three involved international research sites (in Africa, India, and South America); (iii) and by developing and validating a number of laboratory assays used in its protocols and in the field of oral HIV/AIDS research.
Assuntos
Pesquisa Biomédica , Candidíase Bucal/imunologia , Infecções por HIV/complicações , Infecções por HIV/imunologia , Infecções por Papillomavirus/imunologia , Sarcoma de Kaposi/virologia , Antirretrovirais/uso terapêutico , Candidíase Bucal/virologia , Infecções por HIV/tratamento farmacológico , Humanos , Infecções por Papillomavirus/virologiaRESUMO
Human herpesviruses (HHVs) and human papillomavirus (HPV) are common in the general population and, in immunocompetent people, are mostly carried asymptomatically. However, once an individual becomes immunocompromised by age, illness or HIV infection these dormant viruses can manifest and produce disease. In HIV-positive patients, there is an increased risk of disease caused by HHVs and HPV infections and cancers caused by the oncoviruses Epstein-Barr Virus, HHV-8 and HPV. This workshop examined four questions regarding the viruses associated with oral cancers and disease in the HIV-positive and -negative populations, the immune response, and biomarkers useful for accurate diagnostics of these infections and their sequalae. Each presenter identified a number of key areas where further research is required.
Assuntos
Coinfecção/complicações , Infecções por Vírus Epstein-Barr/complicações , Infecções por HIV/complicações , Neoplasias Bucais/virologia , Neoplasias Orofaríngeas/virologia , Infecções por Papillomavirus/complicações , Sarcoma de Kaposi/virologia , Biomarcadores , Coinfecção/imunologia , Infecções por Vírus Epstein-Barr/imunologia , Infecções por HIV/imunologia , Herpesvirus Humano 8 , Humanos , Doenças da Boca/virologia , Infecções por Papillomavirus/imunologia , Sarcoma de Kaposi/imunologiaRESUMO
We report that the expression of murine or human mutant p53 proteins in cells with no endogenous p53 proteins confers new or additional phenotypes upon these cells. Mutant p53 proteins expressed in cell lines lacking p53 resulted in either enhanced tumorigenic potential in nude mice ((10)3 cells) or enhanced plating efficiency in agar cell culture (human SAOS-2 cells). Also, mutant human p53 alleles, unlike the wild-type p53 protein, could also enhance the expression of a test gene regulated by the multi-drug resistance enhancer-promoter element. These data demonstrate a gain of function associated with p53 mutations in addition to the loss of function shown previously to be associated with mutations in this tumour suppressor gene.
Assuntos
Genes p53 , Proteína Supressora de Tumor p53/fisiologia , Membro 1 da Subfamília B de Cassetes de Ligação de ATP , Animais , Proteínas de Transporte/biossíntese , Proteínas de Transporte/genética , Divisão Celular/genética , Linhagem Celular , Células Clonais/transplante , Regulação da Expressão Gênica/genética , Humanos , Glicoproteínas de Membrana/biossíntese , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Mutação , Neoplasias Experimentais/genética , Fenótipo , Especificidade da Espécie , Células Tumorais Cultivadas , Proteína Supressora de Tumor p53/deficiência , Proteína Supressora de Tumor p53/genéticaRESUMO
BACKGROUND: We conducted the first analysis of viral microRNAs (miRNAs) in lung cancer, with a focus on Epstein-Barr virus (EBV). METHODS: We evaluated viral miRs with a two-channel oligo-array targeting mature, anti-sense miRNAs in 290 cases. In 48 cases, we compared microarray and real-time quantitative PCR (qPCR) expression for three EBV miRNAs. We tested for EBV DNA, RNA, and protein in tumour tissue from six cases with and six cases without strong qPCR-based evidence of EBV miRNAs. RESULTS: The EBV miRNAs strongly differentiated between adenocarcinoma and squamous cell carcinoma using the microarray (P<0.01 for 9 out of 16 EBV miRNAs). However, microarray and qPCR measurements of BART1, BART2, and BHRF1-3 expression were not significantly correlated (P=0.53, 0.94, and 0.47, respectively). Although qPCR provided substantial evidence of EBV miRNAs in 7 out of 48 cases, only 1 of these 7 cases had detectable EBV DNA in tumour tissue. None had detectable EBV RNA or protein by histochemical stains. CONCLUSION: In a comprehensive evaluation of EBV miRNA, DNA, RNA, and protein in lung cancer, we found little evidence of EBV in lung tumour tissue. Discrepancies between microarray- and qPCR-based strategies highlight the difficulty of validating molecular markers of disease. Our results do not support a role of EBV in lung cancer.
Assuntos
Adenocarcinoma/virologia , Carcinoma de Células Escamosas/virologia , Herpesvirus Humano 4/genética , Neoplasias Pulmonares/virologia , MicroRNAs/genética , Adenocarcinoma/complicações , Adenocarcinoma/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Algoritmos , Carcinoma de Células Escamosas/complicações , Carcinoma de Células Escamosas/genética , Estudos de Casos e Controles , DNA Viral/análise , Infecções por Vírus Epstein-Barr/complicações , Infecções por Vírus Epstein-Barr/genética , Infecções por Vírus Epstein-Barr/virologia , Feminino , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Regulação Viral da Expressão Gênica , Herpesvirus Humano 4/isolamento & purificação , Herpesvirus Humano 4/fisiologia , Humanos , Neoplasias Pulmonares/complicações , Neoplasias Pulmonares/genética , Masculino , MicroRNAs/análise , MicroRNAs/fisiologia , Análise em Microsséries , Pessoa de Meia-Idade , RNA Viral/análise , Proteínas Virais/análiseRESUMO
The Oral HIV/AIDS Research Alliance is part of the AIDS Clinical Trials Group, the largest HIV clinical trial organization in the world, and it is funded by the National Institute of Dental and Craniofacial Research, in collaboration with the National Institute of Allergy and Infectious Diseases. The alliance's main objective is to investigate the oral complications associated with HIV/AIDS as the epidemic is evolving-in particular, the effects of potent antiretrovirals on the development of oral mucosal lesions and associated fungal and viral pathogens. Furthermore, oral fluids are being explored for their potential monitoring and diagnostic role with respect to HIV disease and coinfections. This article presents an overview of the alliance, its scientific agenda, and an outline of the novel interventional and noninterventional clinical studies ongoing and developing within the AIDS Clinical Trials Group infrastructure in the United States and internationally.
Assuntos
Fármacos Anti-HIV/uso terapêutico , Ensaios Clínicos como Assunto , Pesquisa em Odontologia , Infecções por HIV/complicações , HIV-1 , Doenças da Boca/complicações , Sociedades Odontológicas/organização & administração , Infecções Oportunistas Relacionadas com a AIDS/tratamento farmacológico , Infecções Oportunistas Relacionadas com a AIDS/virologia , Fármacos Anti-HIV/efeitos adversos , Fármacos Anti-HIV/farmacologia , Auditoria Clínica , Infecções por HIV/tratamento farmacológico , Humanos , Cooperação Internacional , Linfonodos/virologia , Doenças da Boca/tratamento farmacológico , Doenças da Boca/virologia , Mucosa Bucal/virologia , Estudos Multicêntricos como Assunto , National Institute of Allergy and Infectious Diseases (U.S.) , National Institute of Dental and Craniofacial Research (U.S.) , Saliva/virologia , Estados Unidos , Carga ViralRESUMO
Kaposi's sarcoma-associated herpesvirus (KSHV/HHV-8) is a novel human lymphotropic herpesvirus linked to several human neoplasms. To date, no animal model for infection by this virus has been described. We have examined the susceptibility of C.B-17 scid/scid mice implanted with human fetal thymus and liver grafts (SCID-hu Thy/Liv mice) to KSHV infection. KSHV virions were inoculated directly into the implants, and viral DNA and mRNA production was assayed using real-time quantitative polymerase chain reaction. This revealed a biphasic infection, with an early phase of lytic replication accompanied and followed by sustained latency. Ultraviolet irradiation of the inoculum abolished all DNA- and mRNA-derived signals, and infection was inhibited by ganciclovir. Viral gene expression was most abundant in CD19(+) B lymphocytes, suggesting that this model faithfully mimics the natural tropism of this virus. Short-term coinfection with HIV-1 did not alter the course of KSHV replication, nor did KSHV alter levels of HIV-1 p24 during the acute phase of the infection. Although no disease was evident in infected animals, SCID-hu Thy/Liv mice should allow the detailed study of KSHV tropism, latency, and drug susceptibility.
Assuntos
Modelos Animais de Doenças , Infecções por Herpesviridae/transmissão , Infecções por Herpesviridae/virologia , Herpesvirus Humano 8 , Animais , Humanos , Camundongos , Camundongos SCIDRESUMO
The replication and transcription activator protein, Rta, is encoded by Orf50 in Kaposi's sarcoma-associated herpesvirus (KSHV) and other known gammaherpesviruses including Epstein-Barr virus (EBV), rhesus rhadinovirus (RRV), herpesvirus saimiri (HVS), and murine herpesvirus 68 (MHV-68). Each Rta/Orf50 homologue of each gammaherpesvirus plays a pivotal role in the initiation of viral lytic gene expression and lytic reactivation from latency. Here we discuss the Rta/Orf50 of KSHV in comparison to the Rta/Orf50s of other gammaherpesviruses in an effort to identify structural motifs, mechanisms of action, and modulating host factors.
Assuntos
Herpesvirus Humano 8/genética , Proteínas Imediatamente Precoces/fisiologia , Transativadores/fisiologia , Proteínas Virais/fisiologia , Animais , Genes Precoces , Herpesvirus Humano 4/genética , Humanos , Proteínas Imediatamente Precoces/química , Proteínas Imediatamente Precoces/genética , Regiões Promotoras Genéticas , Rhadinovirus/genética , Transativadores/química , Transativadores/genética , Ativação Transcricional , Proteínas Virais/química , Proteínas Virais/genética , Ativação ViralAssuntos
Imidazóis/farmacologia , Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Linfoma Folicular/tratamento farmacológico , Inibidores de Fosfoinositídeo-3 Quinase , Inibidores de Proteínas Quinases/farmacologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Quinolinas/farmacologia , Animais , Antineoplásicos/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica , Apoptose/efeitos dos fármacos , Western Blotting , Ácidos Borônicos/farmacologia , Bortezomib , Proliferação de Células/efeitos dos fármacos , Feminino , Fatores de Transcrição Forkhead/fisiologia , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Linfoma Folicular/metabolismo , Linfoma Folicular/patologia , Camundongos , Camundongos Nus , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Pirazinas/farmacologia , RNA Mensageiro/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Serina-Treonina Quinases TOR , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
A three-dimensional model of articular disk dislocations has been developed. Histological examination of 35 temporomandibular joints in near sagittal section reveals sometimes extreme anterior displacement in about 50% and slight posterior displacement of the disk in 6% of the specimens. In the frontal plane medial dislocation prevails in more than 70% of the 32 specimens evaluated, whereas there is only one lateral dislocation. Within the predominantly medio-anterior direction of dislocation there are, however, variations. Excentric forces relative to the center of gravity of the disk always result in dislocations accompanied by rotation in the anterior or posterior part. Where the resulting forces apply on the center of gravity there is a translation in a medio-anterior direction.
Assuntos
Luxações Articulares/patologia , Transtornos da Articulação Temporomandibular/patologia , Idoso , HumanosRESUMO
Cell cycle progression during cytomegalovirus infection was investigated by fluorescence-activated cell sorter (FACS) analysis of the DNA content in growth-arrested as well as serum-stimulated human fibroblasts. Virus-infected cells maintained in either low (0.2%) or high (10%) serum failed to progress into S phase and failed to divide. DNA content analysis in the presence of G1/S (hydroxyurea and mimosine) and G2/M (nocodazole and colcemid) inhibitors demonstrated that upon virus infection of quiescent (G0) cells, the cell cycle did not progress beyond the G1/S border even after serum stimulation. Proteins which normally indicate G1/S transition (proliferating cell nuclear antigen [PCNA]) or G2/M transition (cyclin B1) were elevated by virus infection. PCNA levels were induced in infected cells and exhibited a punctate pattern of nuclear staining instead of the diffuse pattern observed in mock-infected cells. Cyclin B1 was induced in infected cells which exhibited a G1/S DNA content by FACS analysis, suggesting that expression of this key cell cycle function was dramatically altered by viral functions. These data demonstrate that contrary to expectations, cytomegalovirus inhibits normal cell cycle progression. The host cell is blocked prior to S phase to provide a favorable environment for viral replication.
Assuntos
Infecções por Citomegalovirus/patologia , Citomegalovirus , Fase G1 , Fase S , Fibroblastos/patologia , Fibroblastos/virologia , Citometria de Fluxo , Humanos , Antígeno Nuclear de Célula em Proliferação/análiseRESUMO
Kaposi's sarcoma (KS) is an angiogenic tumor of mixed cellularity most commonly found in homosexual men infected with HIV. Both molecular and epidemiologic evidence has linked a newly described herpesvirus to this disease. This virus, Kaposi's sarcoma-associated herpesvirus (KSHV), encodes a number of cellular homologues, including two genes that share remarkable similarity to the human chemokine macrophage inhibitory factor-1 alpha. Recently, studies have begun to shed light on the roles these viral chemokines (vMIP-I and vMIP-II) may play in the complex pathogenesis of KS. The vMIP peptides may contribute to the formation of new blood vessels (neovascularization), inhibit infection by certain strains of HIV-1 and modify the cellular immune response.
Assuntos
Quimiocinas/química , Herpesvirus Humano 8/química , Sarcoma de Kaposi/fisiopatologia , Proteínas Virais/química , Síndrome da Imunodeficiência Adquirida/complicações , Sequência de Aminoácidos , Antivirais/química , Quimiocinas/farmacologia , Infecções por HIV/virologia , HIV-1/metabolismo , Humanos , Fatores Inibidores da Migração de Macrófagos/química , Dados de Sequência Molecular , Neovascularização Fisiológica/fisiologia , Proteínas Virais/farmacologiaRESUMO
Similar to that of other herpesviruses, Kaposi's sarcoma-associated herpesvirus (KSHV/HHV-8) lytic replication destroys the host cell, while the virus can persist in a latent state in synchrony with the host. During latency only a few genes are transcribed, and the question becomes one of what determines latent versus lytic gene expression. Here we undertake a detailed analysis of the latency-associated nuclear antigen (LANA [orf73]) promoter (LANAp). We characterized a minimal region that is necessary and sufficient to maintain high-level transcription in all tissues tested, including primary endothelial cells and B cells, which are the suspected natural host for KSHV. We show that in transient-transfection assays LANAp mimics the expression pattern observed for the authentic promoter in the context of the KSHV episome. Unlike other KSHV promoters tested thus far, LANAp is not affected by tetradecanoyl phorbol acetate or viral lytic cycle functions. It is, however, subject to control by LANA itself and cellular regulatory factors, such as p53. This is in contrast to the K14/vGCR (orf74) promoter, which overlaps LANAp and directs transcription on the opposite strand. We isolated a minimal cis-regulatory region sufficient for K14/vGCR promoter activity and show that it, too, mimics the regulation observed for the authentic viral promoter. In particular, we demonstrate that its activity is absolutely dependent on the immediate-early transactivator orf50, the KSHV homolog of the Epstein-Barr virus Rta transactivator.
Assuntos
Regulação Viral da Expressão Gênica , Herpesvirus Humano 8/genética , Proteínas Nucleares/genética , Regiões Promotoras Genéticas , Receptores de Quimiocinas/genética , Proteínas Virais/genética , Latência Viral , Animais , Antígenos Virais , Linhagem Celular , Herpesvirus Humano 8/fisiologia , Humanos , Camundongos , Proteínas Nucleares/metabolismo , Fases de Leitura Aberta/genética , Plasmídeos/genética , Receptores de Quimiocinas/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sarcoma de Kaposi/virologia , Transcrição Gênica , Transfecção , Proteína Supressora de Tumor p53/metabolismo , Proteínas Virais/metabolismo , Ativação ViralRESUMO
Prolonged bed rest and immobilization inevitably lead to complications. Such complications are much easier to prevent than to treat. Musculoskeletal complications include loss of muscle strength and endurance, contractures and soft tissue changes, disuse osteoporosis, and degenerative joint disease. Cardiovascular complications include an increased heart rate, decreased cardiac reserve, orthostatic hypotension, and venous thromboembolism.
Assuntos
Repouso em Cama/efeitos adversos , Doenças Ósseas/etiologia , Cardiopatias/etiologia , Imobilização/efeitos adversos , Doenças Musculares/etiologia , Doenças Vasculares/etiologia , HumanosRESUMO
Prolonged immobilization affects almost every organ system. Respiratory complications include decreased ventilation, atelectasis, and pneumonia. Decreased basal metabolic rate, increased diuresis, natriuresis, and nitrogen and calcium depletion affect metabolism. Genitourinary problems include renal stones and more frequent urinary tract infections. Glucose intolerance, anorexia, constipation, and pressure sores might develop. Central nervous system changes could affect balance and coordination and lead to increasing dependence on caregivers.
Assuntos
Repouso em Cama/efeitos adversos , Imobilização/efeitos adversos , Doenças do Sistema Endócrino/etiologia , Humanos , Pneumopatias/etiologia , Úlcera por Pressão/etiologiaRESUMO
The commonly used term for solvent abuse, glue sniffing, generally encompasses a variety of substances, including spray paint, thinners, nail varnish remover, gasoline, marking pens, and lighter fluids. Inhaled vapours eventually reach the lipids in the nervous system, where they can be stored for long periods. In three cases of glue-sniffing-related neurotoxicity, the peripheral nervous system was affected in two cases and predominantly the central nervous system in the third. Unfortunately follow up is difficult with this patient population and symptoms are often complicated by alcohol abuse.
Assuntos
Doenças do Sistema Nervoso Periférico/induzido quimicamente , Solventes/efeitos adversos , Transtornos Relacionados ao Uso de Substâncias/complicações , Transtornos Relacionados ao Uso de Substâncias/etiologia , Adolescente , Adulto , Assistência ao Convalescente , Alcoolismo/epidemiologia , Biópsia , Comorbidade , Eletromiografia , Feminino , Marcha , Humanos , Masculino , Condução Nervosa , Transtornos Relacionados ao Uso de Substâncias/diagnóstico , Transtornos Relacionados ao Uso de Substâncias/epidemiologia , Transtornos Relacionados ao Uso de Substâncias/reabilitação , Tomografia Computadorizada por Raios XRESUMO
Kaposi's sarcoma (KS) is a complex proliferative lesion long suspected of being dependent on exogenous paracrine signaling molecules to stimulate its proliferative, angiogenic, and inflammatory components. In particular, both clinical and experimental observations have pointed to a potential role for inflammatory cytokines as permissive factors for KS development, but KS pathogenesis is also critically dependent on infection by an exogenous herpesvirus, the KS-associated herpesvirus (KSHV). To examine the possible links between inflammatory cytokines and KSHV replication, we tested for the ability of such cytokines to induce lytic viral reactivation in the latently infected BCBL-1 cell line. Interferon-gamma consistently activated KSHV replication, whereas tumor necrosis factor, interleukin-1, interleukin-2, interleukin-6, granulocyte-macrophage colony stimulating factor, and basic fibroblast growth factor did not. Glucocorticoids also failed to induce lytic KSHV growth in these cells, but ionomycin, a calcium ionophore, induced replication and strongly augmented the known inductive effects of phorbol esters. Interferon-alpha had a dose-dependent inhibitory effect on KSHV induction by ionomycin. The identification of interferon-gamma as an activator and interferon-alpha as an inhibitor of KSHV induction in vitro correlates well with in vivo observations and demonstrates for the first time that inflammatory cytokines can directly modulate KSHV replication.
Assuntos
Citocinas/farmacologia , Herpesvirus Humano 8/fisiologia , Ativação Viral/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos , Citometria de Fluxo , Humanos , Ionomicina/farmacologia , Ionóforos/farmacologia , Ésteres de Forbol/farmacologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células Tumorais Cultivadas , Latência ViralRESUMO
Kaposi's sarcoma-associated herpesvirus (KSHV), also called human herpesvirus 8 (HHV-8), is the likely etiological agent of Kaposi's sarcoma and primary effusion lymphoma. Common to these malignancies is that tumor cells are latently infected with KSHV. Viral gene expression is limited to a few genes, one of which is the latency-associated nuclear antigen (LANA), the product of ORF73. Examination of the primary sequence of LANA reveals some structural features reminiscent of transcription factors, leading us to hypothesize that LANA may regulate viral and cellular transcription during latency. In reporter gene-based transient transfection assays, we found that LANA can have either positive or negative effects on gene expression. While expression of a reporter gene from several synthetic promoters was increased in the presence of LANA, expression from the human immunodeficiency virus (HIV) long terminal repeat (LTR)-and from NF-kappaB-dependent reporter genes-was reduced by LANA expression. In addition, the promoter of KSHV ORF73 itself is activated up to 5.5-fold by LANA. This autoregulation may be important in tumorigenesis, because two other genes (v-cyclin and v-FLIP) with likely roles in cell growth and survival are also controlled by this element. To identify cellular genes influenced by LANA, we employed cDNA array-based expression profiling. Six known genes (and nine expressed sequence tags) were found to be upregulated in LANA-expressing cell lines. One of these, Staf-50, is known to inhibit expression from the HIV LTR; most of the other known genes are interferon inducible, although the interferon genes themselves were not induced by LANA. These data demonstrate that LANA expression has effects on cellular and viral gene expression. We suggest that, whether direct or indirect in origin, these effects may play important roles in the pathobiology of KSHV infection.
Assuntos
Antígenos Virais/fisiologia , Regulação da Expressão Gênica , Herpesvirus Humano 8/fisiologia , Proteínas Nucleares/fisiologia , Animais , Northern Blotting , Células COS , Proteínas de Ligação a DNA/genética , Humanos , Antígenos de Histocompatibilidade Menor , Regiões Promotoras Genéticas , RNA Mensageiro/análise , Proteínas Repressoras/genética , Fator de Transcrição STAT1 , Sequências Repetidas Terminais , Transativadores/genética , Fatores de Transcrição/genética , Proteínas com Motivo TripartidoRESUMO
The p53 protein is an attractive target for immunotherapy, because mutations in the p53 gene are the most common genetic alterations found in human tumors. These mutations result in high levels of p53 protein in the tumor cell, whereas the expression level of wild-type p53 in nonmalignant tissue is usually much lower. Several canarypox virus recombinants expressing human or murine p53 in wild-type or mutant form were constructed. Immunization with these viruses protected BALB/c mice from a challenge with an isogenic and highly tumorigenic mouse fibroblast tumor cell line expressing high levels of mutant p53. The tumor protection was equally effective regardless of whether wild-type or mutant p53 was used for the immunization, indicating that the immunologic response was not dependent on any particular p53 mutation and that immunization with this live virus vaccine works effectively against mutant p53 protein expressed in a tumor cell. In tumors escaping immunologic rejection, the expression of the p53 protein was commonly down-regulated.
Assuntos
Neoplasias Experimentais/prevenção & controle , Proteína Supressora de Tumor p53/imunologia , Vacinas Sintéticas/farmacologia , Sequência de Aminoácidos , Animais , Anticorpos Antineoplásicos/biossíntese , Avipoxvirus/genética , Avipoxvirus/imunologia , Regulação para Baixo , Feminino , Expressão Gênica , Genes p53 , Vetores Genéticos , Humanos , Imunização , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Mutação , Neoplasias Experimentais/imunologia , Células Tumorais Cultivadas , Proteína Supressora de Tumor p53/genética , Vacinas Virais/farmacologiaRESUMO
Infection with Kaposi's sarcoma-associated herpesvirus (KSHV) is closely associated with Kaposi's sarcoma (KS) and primary effusion lymphoma, with viral genomes present in a latent state in the majority of tumor cells. Here we describe a cluster of latently expressed viral genes whose mRNAs are generated from a common promoter. Two mRNAs in this region encode the latency-associated nuclear antigen, the product of open reading frame 73 (ORF73). The larger RNA, of 5.8 kb, is an unspliced transcript that includes ORF72 and -71 at its 3' end; it initiates at nucleotides (nt) 127880 to 127886 from a promoter lacking recognizable TATA elements. A less abundant mRNA, of 5.4 kb, is a variant of this transcript, in which 336 nt of 5' noncoding information has been removed by RNA splicing. A third, more abundant RNA is generated from the same promoter region via splicing from the common splice donor at nt 127813 to an acceptor 5' to ORF72; this transcript is the presumed mRNA for ORF72, which encodes the viral cyclin D homolog. All three RNAs are 3' coterminal. In situ hybridization analysis with probes that can detect all three transcripts shows that the RNAs are detectable in a large fraction of BCBL-1 cells prior to lytic induction and in >70% of KS spindle cells in primary KS tumors. This confirms that these transcripts are indeed latent RNAs and suggests a role for their products in viral persistence and/or KSHV-associated proliferation.
Assuntos
Genes Virais , Herpesvirus Humano 8/genética , Família Multigênica , Processamento Alternativo , Sequência de Bases , Primers do DNA , DNA Viral , Genes Reporter , Hibridização In Situ , Fases de Leitura Aberta , Regiões Promotoras Genéticas , RNA Mensageiro/genética , Transcrição GênicaRESUMO
A major barrier to the design of immunotherapeutics and vaccines for cancer is the idiosyncratic nature of many tumor antigens and the possibility that T cells may be tolerant of broadly distributed antigens. We have devised an experimental strategy that exploits species differences in protein sequences to circumvent tolerance of high-affinity T cells. HLA transgenic mice were used to obtain cytotoxic T lymphocytes specific for peptides from the human p53 tumor-suppressor molecule presented in association with HLA-A2.1. Although such p53-specific cytotoxic T cells did not recognize nontransformed human cells, they were able to lyse a wide variety of human tumor cells lines, thus confirming the existence of broadly distributed determinants that may serve as targets for immunotherapy.