RESUMO
Cancer is the leading cause of death in dogs, yet there are no established screening paradigms for early detection. Liquid biopsy methods that interrogate cancer-derived genomic alterations in cell-free DNA in blood are being adopted for multi-cancer early detection in human medicine and are now available for veterinary use. The CANcer Detection in Dogs (CANDiD) study is an international, multi-center clinical study designed to validate the performance of a novel multi-cancer early detection "liquid biopsy" test developed for noninvasive detection and characterization of cancer in dogs using next-generation sequencing (NGS) of blood-derived DNA; study results are reported here. In total, 1,358 cancer-diagnosed and presumably cancer-free dogs were enrolled in the study, representing the range of breeds, weights, ages, and cancer types seen in routine clinical practice; 1,100 subjects met inclusion criteria for analysis and were used in the validation of the test. Overall, the liquid biopsy test demonstrated a 54.7% (95% CI: 49.3-60.0%) sensitivity and a 98.5% (95% CI: 97.0-99.3%) specificity. For three of the most aggressive canine cancers (lymphoma, hemangiosarcoma, osteosarcoma), the detection rate was 85.4% (95% CI: 78.4-90.9%); and for eight of the most common canine cancers (lymphoma, hemangiosarcoma, osteosarcoma, soft tissue sarcoma, mast cell tumor, mammary gland carcinoma, anal sac adenocarcinoma, malignant melanoma), the detection rate was 61.9% (95% CI: 55.3-68.1%). The test detected cancer signal in patients representing 30 distinct cancer types and provided a Cancer Signal Origin prediction for a subset of patients with hematological malignancies. Furthermore, the test accurately detected cancer signal in four presumably cancer-free subjects before the onset of clinical signs, further supporting the utility of liquid biopsy as an early detection test. Taken together, these findings demonstrate that NGS-based liquid biopsy can offer a novel option for noninvasive multi-cancer detection in dogs.
Assuntos
Hemangiossarcoma , Osteossarcoma , Animais , Biomarcadores Tumorais/genética , Cães , Detecção Precoce de Câncer , Testes Hematológicos , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Biópsia LíquidaRESUMO
This proof-of-concept study demonstrates that blood-based liquid biopsy using next generation sequencing of cell-free DNA can non-invasively detect multiple classes of genomic alterations in dogs with cancer, including alterations that originate from spatially separated tumor sites. Eleven dogs with a variety of confirmed cancer diagnoses (including localized and disseminated disease) who were scheduled for surgical resection, and five presumably cancer-free dogs, were enrolled. Blood was collected from each subject, and multiple spatially separated tumor tissue samples were collected during surgery from 9 of the cancer subjects. All samples were analyzed using an advanced prototype of a novel liquid biopsy test designed to non-invasively interrogate multiple classes of genomic alterations for the detection, characterization, and management of cancer in dogs. In five of the nine cancer patients with matched tumor and plasma samples, pre-surgical liquid biopsy testing identified genomic alterations, including single nucleotide variants and copy number variants, that matched alterations independently detected in corresponding tumor tissue samples. Importantly, the pre-surgical liquid biopsy test detected alterations observed in spatially separated tissue samples from the same subject, demonstrating the potential of blood-based testing for comprehensive genomic profiling of heterogeneous tumors. Among the three patients with post-surgical blood samples, genomic alterations remained detectable in one patient with incomplete tumor resection, suggesting utility for non-invasive detection of minimal residual disease following curative-intent treatment. Liquid biopsy allows for non-invasive profiling of cancer-associated genomic alterations with a simple blood draw and has potential to overcome the limitations of tissue-based testing posed by tissue-level genomic heterogeneity.
RESUMO
Lung macrophages mature after birth, placing newborn infants, particularly those born preterm, within a unique window of susceptibility to disease. We hypothesized that in preterm infants, lung macrophage immaturity contributes to the development of bronchopulmonary dysplasia (BPD), the most common serious complication of prematurity. By measuring changes in lung macrophage gene expression in preterm patients at risk of BPD, we show here that patients eventually developing BPD had higher inflammatory mediator expression even on the first day of life. Surprisingly, the ex vivo response to LPS was similar across all samples. Our analysis did however uncover macrophage signature genes whose expression increased in the first week of life specifically in patients resilient to disease. We propose that these changes describe the dynamics of human lung macrophage differentiation. Our study therefore provides new mechanistic insight into both neonatal lung disease and human developmental immunology.
Assuntos
Biomarcadores/análise , Displasia Broncopulmonar/patologia , Regulação da Expressão Gênica , Redes Reguladoras de Genes , Macrófagos/imunologia , Pneumonia/patologia , Transcriptoma , Displasia Broncopulmonar/genética , Displasia Broncopulmonar/imunologia , Idade Gestacional , Humanos , Recém-Nascido , Recém-Nascido Prematuro , Macrófagos/metabolismo , Macrófagos/patologia , Pneumonia/genética , Pneumonia/imunologiaRESUMO
Distinct macrophage subsets populate the developing embryo and fetus in distinct waves. However little is known about the functional differences between in utero macrophage populations or how they might contribute to fetal and neonatal immunity. Here we tested the innate immune response of mouse macrophages derived from the embryonic yolk sac and from fetal liver. When isolated from liver or lung, CD11bHI fetal liver derived macrophages responded to the TLR4 agonist LPS by expressing and releasing inflammatory cytokines. However F4/80HI macrophages from the yolk sac did not respond to LPS treatment. While differences in TLR4 expression did not appear to explain these data, F4/80HI macrophages had much lower NLRP3 inflammasome expression compared to CD11bHI macrophages. Gene expression profiling also demonstrated LPS-induced expression of inflammatory genes in CD11bHI macrophages, but not in F4/80HI cells. Genes expressed in LPS-treated CD11bHI macrophages were more likely to contain predicted NF-κB binding sites in their promoter regions. Our data show that CD11bHI macrophages derived from fetal liver are the major pro-inflammatory cells in the developing fetus. These findings could have important implications in better understanding the fetal inflammatory response and the unique features of neonatal immunity.
Assuntos
Feto/imunologia , Ativação de Macrófagos/imunologia , Macrófagos/imunologia , Animais , Citocinas/metabolismo , Feto/citologia , Perfilação da Expressão Gênica , Imunidade Inata , Inflamassomos/metabolismo , Inflamação , Lipopolissacarídeos/farmacologia , Fígado/citologia , Fígado/embriologia , Fígado/imunologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Especificidade de Órgãos , Receptor 4 Toll-Like/metabolismo , Saco Vitelino/citologia , Saco Vitelino/imunologiaRESUMO
Identification of novel biomarkers and targets in renal cell carcinoma (RCC) remains a priority and one cellular compartment that is a rich potential source of such molecules is the plasma membrane. A shotgun proteomic analysis of cell surface proteins enriched by cell surface biotinylation and avidin affinity chromatography was explored using the UMRC2- renal cancer cell line, which lacks von Hippel-Lindau (VHL) tumour suppressor gene function, to determine whether proteins of interest could be detected. Of the 814 proteins identified ~22% were plasma membrane or membrane-associated, including several with known associations with cancer. This included ß-dystroglycan, the transmembrane subunit of the DAG1 gene product. VHL-dependent changes in the form of ß-dystroglycan were detected in UMRC2-/+VHL transfectants. Deglycosylation experiments showed that this was due to differential sialylation. Analysis of normal kidney cortex and conventional RCC tissues showed that a similar change also occurred in vivo. Investigation of the expression of genes involved in glycosylation in UMRC2-/+VHL cells using a focussed microarray highlighted a number of enzymes involved in sialylation; upregulation of bifunctional UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE) was validated in UMRC2- cells compared with their +VHL counterparts and also found in conventional RCC tissue. These results implicate VHL in the regulation of glycosylation and raise interesting questions regarding the extent and importance of such changes in RCC.
Assuntos
Distroglicanas/genética , Distroglicanas/metabolismo , Regulação Neoplásica da Expressão Gênica , Glicoproteínas/genética , Neoplasias Renais/metabolismo , Proteína Supressora de Tumor Von Hippel-Lindau/metabolismo , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Western Blotting , Membrana Celular/metabolismo , Cromatografia Líquida , Cromatografia Gasosa-Espectrometria de Massas , Perfilação da Expressão Gênica , Glicoproteínas/metabolismo , Glicosilação , Humanos , Neoplasias Renais/genética , Análise de Sequência com Séries de Oligonucleotídeos , Células Tumorais Cultivadas , Proteína Supressora de Tumor Von Hippel-Lindau/genéticaRESUMO
The bracken fern Pteridium aquilinum is a plant known to be carcinogenic to animals. Epidemiological studies have shown an association between bracken fern exposure and gastric cancer development in humans. The biological effects of exposure to this plant within the gastric carcinogenesis process are not fully understood. In the present work, effects in the gastric mucosa of mice treated with Pteridium aquilinum were evaluated, as well as molecular mechanisms underlying the synergistic role with Helicobacter pylori infection. Our results showed that exposure to Pteridium aquilinum induces histomorphological modifications including increased expression of acidic glycoconjugates in the gastric mucosa. The transcriptome analysis of gastric mucosa showed that upon exposure to Pteridium aquilinum several glycosyltransferase genes were differently expressed, including Galntl4, C1galt1 and St3gal2, that are mainly involved in the biosynthesis of simple mucin-type carbohydrate antigens. Concomitant treatment with Pteridium aquilinum and infection with Helicobacter pylori also resulted in differently expressed glycosyltransferase genes underlying the biosynthesis of terminal sialylated Lewis antigens, including Sialyl-Lewis(x). These results disclose the molecular basis for the altered pattern of glycan structures observed in the mice gastric mucosa. The gene transcription alterations and the induced glycophenotypic changes observed in the gastric mucosa contribute for the understanding of the molecular mechanisms underlying the role of Pteridium aquilinum in the gastric carcinogenesis process.