RESUMO
We hypothesized that the combination of oncolytic virotherapy with immune checkpoint modulators would reduce tumor burden by direct cell lysis and stimulate antitumor immunity. In this study, we have generated attenuated Measles virus (MV) vectors encoding antibodies against CTLA-4 and PD-L1 (MV-aCTLA-4 and MV-aPD-L1). We characterized the vectors in terms of growth kinetics, antibody expression, and cytotoxicity in vitro. Immunotherapeutic effects were assessed in a newly established, fully immunocompetent murine model of malignant melanoma, B16-CD20. Analyses of tumor-infiltrating lymphocytes and restimulation experiments indicated a favorable immune profile after MV-mediated checkpoint modulation. Therapeutic benefits in terms of delayed tumor progression and prolonged median overall survival were observed for animals treated with vectors encoding anti-CTLA-4 and anti-PD-L1, respectively. Combining systemic administration of antibodies with MV treatment also improved therapeutic outcome. In vivo oncolytic efficacy against human tumors was studied in melanoma xenografts. MV-aCTLA-4 and MV-aPD-L1 were equally efficient as parental MV in this model, with high rates of complete tumor remission (> 80%). Furthermore, we could demonstrate lysis of tumor cells and transgene expression in primary tissue from melanoma patients. The current results suggest rapid translation of combining immune checkpoint modulation with oncolytic viruses into clinical application.
Assuntos
Antígeno B7-H1/metabolismo , Antígeno CTLA-4/metabolismo , Linfócitos do Interstício Tumoral/metabolismo , Melanoma Experimental/terapia , Vírus Oncolíticos/imunologia , Animais , Vetores Genéticos/administração & dosagem , Vírus do Sarampo/genética , Vírus do Sarampo/imunologia , Vírus do Sarampo/metabolismo , Melanoma Experimental/imunologia , Camundongos , Terapia Viral Oncolítica , Vírus Oncolíticos/genética , Vírus Oncolíticos/metabolismo , Resultado do TratamentoRESUMO
Left ventricular hypertrophy (LVH) correlates with chronic renal failure and is one of the most important causes of cardiac mortality. The understanding of the molecular complexity of the disease will help to find biomarkers that open new perspectives about early diagnosis and therapy. This work describes the identification of mediators during pathogenesis relevant for structural remodeling processes of cardiac tissue in uremic LVH. An established rat model of chronic renal failure allowed whole-genome transcriptome analyses as well as the investigation of differential expressed proteins in uremic LVH. The localization of potential biomarkers encoded by candidate genes was done by immunohistochemical analyses of cardiac tissue of the animal model as well as cardiac sections of LVH diseased patients. In addition, the induction of human cardiac fibroblasts (HCF) and human umbilical vein endothelial cells (HUVEC) with the LVH mediator angiotensin II enabled us to investigate uremic LVH progression in vitro. These results point to alterations of myocardial intercellular and cell-matrix contacts in hypertrophic cardiac tissue. Obviously, structural changes of the extracellular matrix are significantly modulated by beta-catenin associated signaling pathways. Interestingly, intracellular translocation of beta-catenin, alpha-actinin and chondroitin sulfate proteoglycan 6 (CSPG6/SMC3) was observed in the animal model and in LVH patients. Our results show that the parallel investigation of rat and human cardiac tissue as well as human cellular models in vitro represents a promising strategy to identify reliable biomarkers of LVH.
Assuntos
Actinina/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteoglicanas de Sulfatos de Condroitina/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Hipertrofia Ventricular Esquerda/metabolismo , Miocárdio/metabolismo , beta Catenina/metabolismo , Actinina/genética , Angiotensina II/metabolismo , Animais , Proteínas de Ciclo Celular/genética , Células Cultivadas , Proteoglicanas de Sulfatos de Condroitina/genética , Proteínas Cromossômicas não Histona/genética , Modelos Animais de Doenças , Matriz Extracelular/química , Matriz Extracelular/metabolismo , Perfilação da Expressão Gênica , Humanos , Hipertrofia Ventricular Esquerda/patologia , Masculino , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 7 da Matriz/metabolismo , Miocárdio/patologia , Análise de Sequência com Séries de Oligonucleotídeos , Osteonectina/metabolismo , Ratos , Ratos Sprague-Dawley , Remodelação Ventricular/fisiologia , Versicanas/metabolismo , beta Catenina/genéticaRESUMO
BACKGROUND: In contrast to gene-mapping studies of simple Mendelian disorders, genetic analyses of complex traits are far more challenging, and high quality data management systems are often critical to the success of these projects. To minimize the difficulties inherent in complex trait studies, we have developed GeneLink, a Web-accessible, password-protected Sybase database. RESULTS: GeneLink is a powerful tool for complex trait mapping, enabling genotypic data to be easily merged with pedigree and extensive phenotypic data. Specifically designed to facilitate large-scale (multi-center) genetic linkage or association studies, GeneLink securely and efficiently handles large amounts of data and provides additional features to facilitate data analysis by existing software packages and quality control. These include the ability to download chromosome-specific data files containing marker data in map order in various formats appropriate for downstream analyses (e.g., GAS and LINKAGE). Furthermore, an unlimited number of phenotypes (either qualitative or quantitative) can be stored and analyzed. Finally, GeneLink generates several quality assurance reports, including genotyping success rates of specified DNA samples or success and heterozygosity rates for specified markers. CONCLUSIONS: GeneLink has already proven an invaluable tool for complex trait mapping studies and is discussed primarily in the context of our large, multi-center study of hereditary prostate cancer (HPC). GeneLink is freely available at http://research.nhgri.nih.gov/genelink.
Assuntos
Bases de Dados Genéticas/tendências , Herança Multifatorial/genética , Alelos , Mapeamento Cromossômico , Primers do DNA/genética , Ligação Genética/genética , Marcadores Genéticos/genética , Predisposição Genética para Doença/genética , Genótipo , Humanos , Desequilíbrio de Ligação/genética , Masculino , Linhagem , Neoplasias da Próstata/genética , Software , Design de SoftwareRESUMO
BACKGROUND: In order to identify predisposition loci to hereditary prostate cancer (HPC), we performed a genome-wide linkage analysis using samples from a genetically homogeneous population, with 13 Finnish multiplex prostate cancer families. METHODS: Altogether 87 DNA samples were genotyped from 13 families. Logarithm-of-odds (LOD) scores were calculated for all autosomes using FASTLINK and GENEHUNTER designating all unaffected men and all women as unknown. RESULTS: The highest LOD scores in the affected-only analyses were found at 11q14, where the two-point LOD score was 2.97 (theta = 0.0 at D11S901), GENEHUNTER heterogeneity LOD (HLOD) of 3.36, and a non-parametric-linkage (NPL) score of 2.67 (P = 0.008). A second positive site was at 3p25-26, with a two-point LOD score of 2.57 (theta = 0.01 at D3S1297), HLOD of 2.15, and NPL score of 2.27 (P = 0.02). CONCLUSIONS: The results suggest two HPC regions in the Finnish population, which have not been reported previously and warrant further study.
Assuntos
Ligação Genética/genética , Genoma Humano , Neoplasias da Próstata/genética , Idoso , Idoso de 80 Anos ou mais , Cromossomos Humanos Par 11/genética , Cromossomos Humanos Par 3/genética , DNA de Neoplasias/química , DNA de Neoplasias/genética , Família , Feminino , Finlândia , Predisposição Genética para Doença , Humanos , Masculino , Repetições de Microssatélites/genética , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Análise de Sequência de DNA , Estatísticas não Paramétricas , Neoplasias Gástricas/genéticaRESUMO
BACKGROUND: Prostate cancer (CaP) is a common disorder with multiple genetic and environmental factors contributing to the disease. CaP susceptibility loci can be identified through genome-wide scans of high-risk families. METHODS: Allele sharing at 405 markers, distributed across the genome, among 50 families with hereditary prostate cancer, ascertained throughout Sweden, was evaluated through linkage analyses. Genotype data were analyzed utilizing multipoint parametric and non-parametric methods. RESULTS: Two regions provided suggestive evidence for linkage: 19p13.3 (marker D19S209, LOD = 2.91, P = 0.0001) and 5q11.2 (marker D5S407, LOD = 2.24, P = 0.0007). Additional regions with moderate evidence for linkage in the complete set of families, or stratified subsets, were observed on chromosome 1, 4, 6, 7, 8, and X. CONCLUSIONS: Our results provide strong confirmatory evidence of linkage at 19q13.3 and 5q11.2. The lack of confirmation of linkage at several loci identified in other genome-wide scans emphasizes the need to combine linkage data between research groups.
Assuntos
Ligação Genética/genética , Genoma Humano , Neoplasias da Próstata/genética , Idoso , Alelos , Cromossomos Humanos Par 19/genética , Cromossomos Humanos Par 5/genética , DNA de Neoplasias/química , DNA de Neoplasias/genética , Família , Feminino , Predisposição Genética para Doença , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Análise de Sequência de DNA , Estatísticas não Paramétricas , SuéciaRESUMO
BACKGROUND: Although the subject of intensive study, the genetic influences responsible for familial clustering of prostate cancer remain largely unidentified. Genome-wide scans for linkage in prostate cancer families can be used to systematically search for genes capable of affecting risk for the disease. METHODS: All available family members from 188 families, each having at least three first-degree relatives affected with prostate cancer, were genotyped at 406 markers distributed across the genome at average intervals of less than 10 cM. Genotype data was analyzed using primarily a non-parametric, multipoint approach, although parametric analyses were performed as well. RESULTS: The strongest evidence for linkage was observed at D4S1615, at 4q21 (LOD of 2.8, P = 0.0002). Two other regions had LOD scores over 2.0: at 9q34 (marker D9S1826, LOD = 2.17, P = 0.0008) and at 2q23 (marker D2S151, LOD = 2.03, P = 0.001). An additional 12 regions had LOD scores over 1.0, including markers at 1q24-25 and 7q22 having scores >1.6. Stratifying the linkage results by age of diagnosis indicated that the linkages to chromosomes 2 and 4 were strongest in families with early and late ages of diagnosis, respectively. CONCLUSIONS: Our data implicate several new loci as harboring prostate cancer susceptibility genes, and provide confirmatory evidence of linkage at several loci identified previously in other genome-wide scans, including the three regions (4q21, 9q34, and 2q23) with strongest evidence for prostate cancer linkage. These data also emphasize the need to combine linkage data from large numbers of prostate cancer families in efforts to effectively address the extensive heterogeneity that characterizes genetic aspects of this disease.
Assuntos
Ligação Genética/genética , Genoma Humano , Neoplasias da Próstata/genética , Idoso , DNA de Neoplasias/química , DNA de Neoplasias/genética , Família , Marcadores Genéticos/genética , Predisposição Genética para Doença , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Análise de Sequência de DNA , Estatísticas não ParamétricasRESUMO
BACKGROUND: Previous linkage studies have suggested prostate cancer susceptibility genes located on chromosomes 1, 20, and X. Several putative prostate cancer candidate genes have also been identified including RNASEL, MSR1, and ELAC2. Presently, these linkage regions and candidate genes appear to explain only a small proportion of hereditary prostate cancer cases suggesting the need for additional whole genome analyses. METHODS: A genome-wide mode-of-inheritance-free linkage scan, using 405 genetic markers, was conducted on 175 pedigrees, the majority containing three or more affected individuals diagnosed with prostate cancer. Stratified linkage analyses were performed based on previously established criteria. RESULTS: Results based on the entire set of 175 pedigrees showed strong suggestive evidence for linkage on chromosome 17q (LOD = 2.36), with strongest evidence coming from the subset of pedigrees with four or more affected individuals (LOD = 3.27). Race specific analyses revealed strong suggestive evidence for linkage in our African-American pedigrees on chromosome 22q (LOD = 2.35). CONCLUSIONS: Genome-wide analysis of a large set of prostate cancer families indicates new areas of the genome that may harbor prostate cancer susceptibility genes. Specifically, our linkage results suggest that there is a prostate cancer susceptibility gene on chromosome 17 that is independent of ELAC2. Further research including combined analyses of independent genome-wide scan data may clarify the most important regions for future investigation.
Assuntos
Cromossomos Humanos Par 17/genética , Genes BRCA1 , Ligação Genética/genética , Genoma Humano , Neoplasias da Próstata/genética , Idade de Início , Idoso , População Negra , DNA de Neoplasias/química , DNA de Neoplasias/genética , Família , Predisposição Genética para Doença , Humanos , Masculino , Repetições de Microssatélites/genética , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Análise de Sequência de DNA , População BrancaRESUMO
Over the past 20 years, the incidence of cutaneous malignant melanoma (CMM) has increased dramatically worldwide. A positive family history of the disease is among the most established risk factors for CMM; it is estimated that 10% of CMM cases result from an inherited predisposition. Although mutations in two genes, CDKN2A and CDK4, have been shown to confer an increased risk of CMM, they account for only 20%-25% of families with multiple cases of CMM. Therefore, to localize additional loci involved in melanoma susceptibility, we have performed a genomewide scan for linkage in 49 Australian pedigrees containing at least three CMM cases, in which CDKN2A and CDK4 involvement has been excluded. The highest two-point parametric LOD score (1.82; recombination fraction [theta] 0.2) was obtained at D1S2726, which maps to the short arm of chromosome 1 (1p22). A parametric LOD score of 4.65 (theta=0) and a nonparametric LOD score of 4.19 were found at D1S2779 in nine families selected for early age at onset. Additional typing yielded seven adjacent markers with LOD scores >3 in this subset, with the highest parametric LOD score, 4.95 (theta=0) (nonparametric LOD score 5.37), at D1S2776. Analysis of 33 additional multiplex families with CMM from several continents provided further evidence for linkage to the 1p22 region, again strongest in families with the earliest mean age at diagnosis. A nonparametric ordered sequential analysis was used, based on the average age at diagnosis in each family. The highest LOD score, 6.43, was obtained at D1S2779 and occurred when the 15 families with the earliest ages at onset were included. These data provide significant evidence of a novel susceptibility gene for CMM located within chromosome band 1p22.