Immune evolution from preneoplasia to invasive lung adenocarcinomas and underlying molecular features.

The mechanism by which anti-cancer immunity shapes early carcinogenesis of lung adenocarcinoma (ADC) is unknown. In this study, we characterize the immune contexture of invasive lung ADC and its precursors by transcriptomic immune profiling, T cell receptor (TCR) sequencing and multiplex immunofluorescence (mIF). Our results demonstrate that anti-tumor immunity evolved as a continuum from lung preneoplasia, to preinvasive ADC, minimally-invasive ADC and frankly invasive lung ADC with a gradually less effective and more intensively regulated immune response including down-regulation of immune-activation pathways, up-regulation of immunosuppressive pathways, lower infiltration of cytotoxic T cells (CTLs) and anti-tumor helper T cells (Th), higher infiltration of regulatory T cells (Tregs), decreased T cell clonality, and lower frequencies of top T cell clones in later-stages. Driver mutations, chromosomal copy number aberrations (CNAs) and aberrant DNA methylation may collectively impinge host immune responses and facilitate immune evasion, promoting the outgrowth of fit subclones in preneoplasia into dominant clones in invasive ADC.

miR-543 regulates the epigenetic landscape of myelofibrosis by targeting TET1 and TET2.

Myelofibrosis (MF) is a myeloproliferative neoplasm characterized by cytopenia and extramedullary hematopoiesis, resulting in splenomegaly. Multiple pathological mechanisms (e.g., circulating cytokines and genetic alterations, such as JAKV617F mutation) have been implicated in the etiology of MF, but the molecular mechanism causing resistance to JAK2V617F inhibitor therapy remains unknown. Among MF patients who were treated with the JAK inhibitor ruxolitinib, we compared noncoding RNA profiles of ruxolitinib therapy responders versus nonresponders and found miR-543 was significantly upregulated in nonresponders. We validated these findings by reverse transcription-quantitative PCR. in this same cohort, in 2 additional independent MF patient cohorts from the United States and Romania, and in a JAK2V617F mouse model of MF. Both in vitro and in vivo models were used to determine the underlying molecular mechanism of miR-543 in MF. Here, we demonstrate that miR-543 targets the dioxygenases ten-eleven translocation 1 (TET1) and 2 (TET2) in patients and in vitro, causing increased levels of global 5-methylcytosine, while decreasing the acetylation of histone 3, STAT3, and tumor protein p53. Mechanistically, we found that activation of STAT3 by JAKs epigenetically controls miR-543 expression via binding the promoter region of miR-543. Furthermore, miR-543 upregulation promotes the expression of genes related to drug metabolism, including CYP3A4, which is involved in ruxolitinib metabolism. Our findings suggest miR-543 as a potentially novel biomarker for the prognosis of MF patients with a high risk of treatment resistance and as a potentially new target for the development of new treatment options.

DNA methylation patterns in bladder tumors of African American patients point to distinct alterations in xenobiotic metabolism.

Racial/ethnic disparities have a significant impact on bladder cancer outcomes with African American patients demonstrating inferior survival over European-American patients. We hypothesized that epigenetic difference in methylation of tumor DNA is an underlying cause of this survival health disparity. We analyzed bladder tumors from African American and European-American patients using reduced representation bisulfite sequencing (RRBS) to annotate differentially methylated DNA regions. Liquid chromatography-mass spectrometry (LC-MS/MS) based metabolomics and flux studies were performed to examine metabolic pathways that showed significant association to the discovered DNA methylation patterns. RRBS analysis showed frequent hypermethylated CpG islands in African American patients. Further analysis showed that these hypermethylated CpG islands in patients are commonly located in the promoter regions of xenobiotic enzymes that are involved in bladder cancer progression. On follow-up, LC-MS/MS revealed accumulation of glucuronic acid, S-adenosylhomocysteine, and a decrease in S-adenosylmethionine, corroborating findings from the RRBS and mRNA expression analysis indicating increased glucuronidation and methylation capacities in African American patients. Flux analysis experiments with 13C-labeled glucose in cultured African American bladder cancer cells confirmed these findings. Collectively, our studies revealed robust differences in methylation-related metabolism and expression of enzymes regulating xenobiotic metabolism in African American patients indicate that race/ethnic differences in tumor biology may exist in bladder cancer.

Cancer-associated rs6983267 SNP and its accompanying long noncoding RNA CCAT2 induce myeloid malignancies via unique SNP-specific RNA mutations.

The cancer-risk-associated rs6983267 single nucleotide polymorphism (SNP) and the accompanying long noncoding RNA CCAT2 in the highly amplified 8q24.21 region have been implicated in cancer predisposition, although causality has not been established. Here, using allele-specific CCAT2 transgenic mice, we demonstrate that CCAT2 overexpression leads to spontaneous myeloid malignancies. We further identified that CCAT2 is overexpressed in bone marrow and peripheral blood of myelodysplastic/myeloproliferative neoplasms (MDS/MPN) patients. CCAT2 induces global deregulation of gene expression by down-regulating EZH2 in vitro and in vivo in an allele-specific manner. We also identified a novel non-APOBEC, non-ADAR, RNA editing at the SNP locus in MDS/MPN patients and CCAT2-transgenic mice. The RNA transcribed from the SNP locus in malignant hematopoietic cells have different allelic composition from the corresponding genomic DNA, a phenomenon rarely observed in normal cells. Our findings provide fundamental insights into the functional role of rs6983267 SNP and CCAT2 in myeloid malignancies.

In Vivo Functional Platform Targeting Patient-Derived Xenografts Identifies WDR5-Myc Association as a Critical Determinant of Pancreatic Cancer.

Current treatment regimens for pancreatic ductal adenocarcinoma (PDAC) yield poor 5-year survival, emphasizing the critical need to identify druggable targets essential for PDAC maintenance. We developed an unbiased and in vivo target discovery approach to identify molecular vulnerabilities in low-passage and patient-derived PDAC xenografts or genetically engineered mouse model-derived allografts. Focusing on epigenetic regulators, we identified WDR5, a core member of the COMPASS histone H3 Lys4 (H3K4) MLL (1-4) methyltransferase complex, as a top tumor maintenance hit required across multiple human and mouse tumors. Mechanistically, WDR5 functions to sustain proper execution of DNA replication in PDAC cells, as previously suggested by replication stress studies involving MLL1, and c-Myc, also found to interact with WDR5. We indeed demonstrate that interaction with c-Myc is critical for this function. By showing that ATR inhibition mimicked the effects of WDR5 suppression, these data provide rationale to test ATR and WDR5 inhibitors for activity in this disease.

The generation and utilization of a cancer-oriented representation of the human transcriptome by using expressed sequence tags.

Whereas genome sequencing defines the genetic potential of an organism, transcript sequencing defines the utilization of this potential and links the genome with most areas of biology. To exploit the information within the human genome in the fight against cancer, we have deposited some two million expressed sequence tags (ESTs) from human tumors and their corresponding normal tissues in the public databases. The data currently define approximately 23,500 genes, of which only approximately 1,250 are still represented only by ESTs. Examination of the EST coverage of known cancer-related (CR) genes reveals that <1% do not have corresponding ESTs, indicating that the representation of genes associated with commonly studied tumors is high. The careful recording of the origin of all ESTs we have produced has enabled detailed definition of where the genes they represent are expressed in the human body. More than 100,000 ESTs are available for seven tissues, indicating a surprising variability of gene usage that has led to the discovery of a significant number of genes with restricted expression, and that may thus be therapeutically useful. The ESTs also reveal novel nonsynonymous germline variants (although the one-pass nature of the data necessitates careful validation) and many alternatively spliced transcripts. Although widely exploited by the scientific community, vindicating our totally open source policy, the EST data generated still provide extensive information that remains to be systematically explored, and that may further facilitate progress toward both the understanding and treatment of human cancers.

[Chromosome abnormalities caused by computer video display monitors' radiation]. / Alterações cromossômicas causadas pela radiação dos monitores de vídeo de computadores.

OBJECTIVE: Concerns were raised about the potential damaging effects of electromagnetic field (EMF) radiation emissions of ELF (extremely low frequency) and VLF (very low frequency) computer video display monitors (VDM), it was assessed the frequency of structural chromosome abnormalities and investigated the cell cycle kinetics in individuals occupationally exposed to VDM radiation. METHODS: Chromosome aberrations were investigated in 2,000 first cell cycle metaphases obtained after 48-hour cultures of peripheral blood lymphocytes drawn from 10 individuals occupationally exposed to VDM radiation (group E) and 10 controls (group C). Cell cycle kinetics was studied using the mitotic index (MI) and cellular proliferation index (CPI). RESULTS: Statistical analysis showed significantly higher frequencies of anomalous metaphases (E=5.9%; C=3.7%) and anomaly/cell (E=0.066+/-0.026; C= 0.040+/-0.026) in individuals exposed to VDM radiation. The most common cytogenetic alterations seen were chromatid breaks at frequencies of 0.034+/-0.016 in group E and 0.016+/-0.015 in group C. There was no significant difference between MI and CPI frequencies in both groups. CONCLUSIONS: The study findings suggest genotoxic effects of EMF emissions revealed by the higher frequency of chromatid breaks in individuals exposed to VDM radiation. However, there is a need of further studies on EMF genetic effects using other investigation methods.

Alteraçöes cromossômicas causadas pela radiaçäo dos monitores de vídeo de computadores / Chromosome abnormalities caused by computer video display monitors' radiation

OBJETIVO: Em decorrência dos questionamentos sobre o efeito deletério das radiaçöes emitidas pelo campo eletromagnético (CEM) dos tipos ELF (extremely low frequency) e VLF (very low frequency) transmitidas pelos monitores de vídeo dos computadores (CRT), foi avaliada a freqüência de anomalias cromossômicas estruturais e a cinética do ciclo celular em indivíduos expostos por seu trabalho à radiaçäo dos CRT. MÉTODOS: A pesquisa de aberraçöes cromossômicas foi realizada em 2.000 metáfases de primeira divisäo celular obtidas de culturas de 48h de linfócitos de sangue venoso periférico de dez indivíduos expostos ao CRT (grupo E) e de dez controles (grupo C). A cinética do ciclo celular foi pesquisada pelos índices mitótico (IM) e de proliferaçäo celular (IPC). RESULTADOS: A análise estatística evidenciou freqüências significativamente maiores de metáfases com anomalias cromossômicas (E=5,9por cento; C=3,7por cento) e anomalias/célula (E=0,066ñ0,026; C=0,040ñ0,026) nos indivíduos expostos aos CRTs. As alteraçöes citogenéticas mais comuns foram as quebras cromatídicas, com freqüência de 0,034ñ0,016 no grupo E e de 0,016ñ0,015 no grupo C. As freqüências de IM e IPC näo apresentaram diferenças significantes entre os grupos avaliados. CONCLUSOES: Os resultados sugerem um efeito genotóxico do CEM emitido pelos CRTs devido à freqüência mais elevada de quebras cromatídicas, enfatizando a necessidade de haver um número maior de estudos com diferentes técnicas que vise a investigar a açäo do CEM sobre o material genético

Elementos Transponíveis e Doenças Humanas: da Instabilidade Genônica à Carcinogênese / Transposable Elements and Human Diseases: from Genomic Instability to Carcinogenesis

Os elementos transponíveis säo sequências de DNA que podem se mover no genoma e por isso säo a principal fonte de mutações espontâneas. Dez a 15 por cento do DNA humano säo compostos pelos transposons Alu e Line-1. O grande número desses elementos, no genoma, propicia ampla oportunidade para recombinaçäo homóloga desigual, ocasionando deleções ou duplicações gênicas e mesmo alterações mais complexas do material genético. Säo citados na literatura diversos casos de doenças causadas pela inserçäo de Alu e Line-1 em diversos genes, tanto na linhagem germinativa como somática. Também tem sido encontrada uma correlaçäo direta entre a hipometilaçäo de sequências regulatórias de Line-1 e a presença de produtos de transcriçäo, ou de proteína por ele produzida, em teratocarcinomas, tumores da linhagem germinativa e em alguns cânceres de cérebro e mama. Tal correlaçäo sugere que esses elementos estäo ativos e podem estar envolvidos na iniciaçäo e na manutençäo do estado maligno, principalmente porque a desmetilaçäo do DNA genômico parece ser um evento precoce na progressäo tumoral

Relevância da genética na etiologia do autismo / Relevance of genetics in autism etiology

O autismo constitui uma das orincipais patologias integrantes dos transtornos invasivos do desenvolvimento. É caracterizado por atraso e desvio do desenvolvimento social, de comunicaçäo e cognitivo e se manifesta precocemente. Atualmente, é reconhecida a participaçäo de fatores genéticos em sua etiologia. Esse distúrbio näo segue padröes clássicos de herança mendeliana e apresenta grande heterogeneidade genética. Entre as síndromes gênicas observa-se forte associaçäo com a síndrome do cromossomo X frágil (gene FMR-1) com a esclerose tuberosa. Um número crescente de trabalhos têm relatado a ocorrência de anomalias cromossômicas em autismo, incluindo cromossomos extranumerários derivados do 15, envolvendo a regiäo 15q11-q13 e o cromossomo 7. Na regiäo 15q11-q13 encontram-se genes cujo produtos atuam no sistema nervoso central e que säo subunidades de recptores de neurotransmissores ou de canais de íons (UBE3A, GABRA5, GABRG3, CHRNA7, I ITO). Algumas regiöes do cromossomo 7 também contêm genes (EN-2 e HOXA) importantes para o desenvolvimento (homeogenes), com atuaçäo na formaçäo do sistema nervoso central. Os cromossomos 7, 15 e X, portanto, parecem conter genes relacionados com a etiologia do autismo, cuja avaliaçäo genética pode gerar conhecimentos sobre os mecanismos biológicos envolvidos neste grave distúrbio