RESUMEN
DNA double-strand breaks (DSBs) are deleterious lesions that challenge genome integrity. To mitigate this threat, human cells rely on the activity of multiple DNA repair machineries that are tightly regulated throughout the cell cycle1. In interphase, DSBs are mainly repaired by non-homologous end joining and homologous recombination2. However, these pathways are completely inhibited in mitosis3-5, leaving the fate of mitotic DSBs unknown. Here we show that DNA polymerase theta6 (Polθ) repairs mitotic DSBs and thereby maintains genome integrity. In contrast to other DSB repair factors, Polθ function is activated in mitosis upon phosphorylation by Polo-like kinase 1 (PLK1). Phosphorylated Polθ is recruited by a direct interaction with the BRCA1 C-terminal domains of TOPBP1 to mitotic DSBs, where it mediates joining of broken DNA ends. Loss of Polθ leads to defective repair of mitotic DSBs, resulting in a loss of genome integrity. This is further exacerbated in cells that are deficient in homologous recombination, where loss of mitotic DSB repair by Polθ results in cell death. Our results identify mitotic DSB repair as the underlying cause of synthetic lethality between Polθ and homologous recombination. Together, our findings reveal the critical importance of mitotic DSB repair in the maintenance of genome integrity.
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Roturas del ADN de Doble Cadena , Reparación del ADN , ADN Polimerasa Dirigida por ADN , Mitosis , Proteínas Serina-Treonina Quinasas , Humanos , Proteína BRCA1/metabolismo , Proteínas de Ciclo Celular/metabolismo , Muerte Celular/genética , ADN Polimerasa Dirigida por ADN/química , ADN Polimerasa Dirigida por ADN/metabolismo , Recombinación Homóloga/genética , Fosforilación , Proteínas Serina-Treonina Quinasas/metabolismo , Mutaciones Letales Sintéticas , ADN Polimerasa theta , Quinasa Tipo Polo 1RESUMEN
PURPOSE: Mosaic BRCA1 promoter methylation (BRCA1meth) increases the risk of early-onset breast cancer, triple-negative breast cancer and ovarian cancer. As mosaic BRCA1meth are believed to occur de novo, their role in family breast/ovarian cancer has not been assessed. PATIENTS: Blood-derived DNA from 20 unrelated affected cases from families with aggregation of breast/ovarian cancer, but with no germline pathogenic variants in BRCA1/2, PALB2 or RAD51C/D, were screened by methylation-sensitive high-resolution melting. CpG analysis was performed by pyrosequencing on blood and buccal swab. Two probands carried a pathogenic variant in a moderate-penetrance gene (ATM and BARD1), and 8 of 18 others (44%) carried BRCA1meth (vs none of the 20 age-matched controls). Involvement of BRCA1 in tumourigenesis in methylated probands was demonstrated in most tested cases by detection of a loss of heterozygosity and a homologous recombination deficiency signature. Among the eight methylated probands, two had relatives with breast cancer with detectable BRCA1meth in blood, including one with high methylation levels in two non-tumour tissues. CONCLUSIONS: The high prevalence of mosaic BRCA1meth in patients with breast/ovarian cancer with affected relatives, as well as this first description of a family aggregation of mosaic BRCA1meth, shows how this de novo event can contribute to hereditary breast/ovarian cancer pedigrees.
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Neoplasias de la Mama , Neoplasias Ováricas , Humanos , Femenino , Proteína BRCA1/genética , Linaje , Proteína BRCA2/genética , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Metilación , Neoplasias Ováricas/genética , Neoplasias Ováricas/diagnóstico , Mutación de Línea Germinal/genética , Predisposición Genética a la Enfermedad , Metilación de ADN/genéticaRESUMEN
Electrostatic interaction of ampholytic nanocolloidal particles (NPs), which mimic globular proteins, with polyelectrolyte brushes is analyzed within mean-field Poisson-Boltzmann approximation. In accordance with experimental findings, the theory predicts that an electrostatic driving force for the particle uptake by the brush may emerge when the net charge of the particle in the buffer and the charge of the brush are of the same sign. The origin of this driving force is change in the ionization state of weak cationic and anionic groups on the NP surface provoked by interaction with the brush. In experimental systems, the ionic interactions are complemented by excluded-volume, hydrophobic, and other types of interactions that all together control NP uptake by or expulsion from the brush. Here, we focus on the NP-brush ionic interactions. It is demonstrated that deviation between the buffer pH and the NP isoelectric point, considered usually as the key control parameter, does not uniquely determine the insertion free energy patterns. The latter depends also on the proportion of cationic and anionic groups in the NPs and their specific ionization constants as well as on salt concentration in the buffer. The analysis of the free energy landscape proves that a local minimum in the free energy inside the brush appears, provided the NP charge reversal occurs upon insertion into the brush. This minimum corresponds either to a thermodynamically stable or to a metastable state, depending on the pH offset from the IEP and salt concentration, and is separated from the bulk of the solution by a free energy barrier. The latter, being fairly independent of salt concentration in height, may strongly impede the NP absorption kinetically even when it is thermodynamically favorable. Hence, change reversal is a necessary but insufficient condition for the uptake of the NPs by similarly charged polyelectrolyte brushes.
RESUMEN
Basal/squamous (Ba/Sq) subtype represents an intrinsic and robust group in the consensus molecular classification of muscle-invasive bladder cancer (MIBC), with poor outcome and controversial chemosensitivity. We aimed to investigate the spectrum of intratumor heterogeneity (ITH) in the Ba/Sq subtype. First, we validated a 29-gene NanoString CodeSet to predict the Ba/Sq subtype for FFPE samples. We identified heterogeneous Ba/Sq tumors in a series of 331 MIBC FFPE samples using dual GATA3/KRT5/6 immunohistochemistry (IHC). Heterogeneous regions with distinct immunostaining patterns were studied separately for gene expression using the 29-gene CodeSet, for mutations by targeted next-generation sequencing, and for copy number alteration (CNA) by microarray hybridization. Among 83 Ba/Sq tumors identified by GATA3/KRT5/6 dual staining, 19 tumors showed heterogeneity at the IHC level. In one third of the 19 cases, regions from the same tumor were classified in different distinct molecular subtypes. The mutational and CNA profiles confirmed the same clonal origin for IHC heterogeneous regions with possible subclonal evolution. Overall, two patterns of intratumoral heterogeneity (ITH) were observed in Ba/Sq tumors: low ITH (regions with distinct immunostaining, but common molecular subtype and shared CNA) or high ITH (regions with distinct immunostaining, molecular subtype, and CNA). These results showed multilayer heterogeneity in Ba/Sq MIBC. In view of personalized medicine, this heterogeneity adds complexity and should be taken into account for sampling procedures used for diagnosis and treatment choice. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
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Biomarcadores de Tumor/genética , Variaciones en el Número de Copia de ADN/genética , Mutación/genética , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/patología , Biomarcadores de Tumor/metabolismo , Perfilación de la Expresión Génica/métodos , Humanos , Inmunohistoquímica/métodos , Medicina de Precisión/métodos , Neoplasias de la Vejiga Urinaria/diagnósticoRESUMEN
SUMMARY: We introduce shallowHRD, a software tool to evaluate tumor homologous recombination deficiency (HRD) based on whole genome sequencing (WGS) at low coverage (shallow WGS or sWGS; â¼1X coverage). The tool, based on mining copy number alterations profile, implements a fast and straightforward procedure that shows 87.5% sensitivity and 90.5% specificity for HRD detection. shallowHRD could be instrumental in predicting response to poly(ADP-ribose) polymerase inhibitors, to which HRD tumors are selectively sensitive. shallowHRD displays efficiency comparable to most state-of-art approaches, is cost-effective, generates low-storable outputs and is also suitable for fixed-formalin paraffin embedded tissues. AVAILABILITY AND IMPLEMENTATION: shallowHRD R script and documentation are available at https://github.com/aeeckhou/shallowHRD. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.
Asunto(s)
Variaciones en el Número de Copia de ADN , Neoplasias , Recombinación Homóloga , Humanos , Neoplasias/genética , Programas Informáticos , Secuenciación Completa del GenomaRESUMEN
Chronic headache is a topical problem of neurology, psychiatry and general practice. The medication-overuse headache (MOH) is one of the leading pathologies in the structure of chronic headache. However, early diagnosis of the MOH is challenging. We analyzed potential proteomic biomarkers of serum and urine in patients with MOH. METHODS: We searched PubMed, Springer, Scopus, Web of Science, ClinicalKey, and Google Scholar databases for English publications over the past 10 years using keywords and their combinations. RESULTS: We found and analyzed seven studies that met the search criteria for the purpose of the review, including 24 serum proteomic biomarkers and 25 urine proteomic biomarkers of MOH. Moreover, the candidate genes and locus of the studied serum (vitamin D-binding protein, lipocalin-type prostaglandin D2 synthase, apolipoprotein E, etc.) and urine proteomic biomarkers (uromodulin, alpha-1-microglobulin, zinc-alpha-2-glycoprotein, etc.) of MOH are presented in this review. CONCLUSIONS: The serum and urine proteomic biomarkers of MOH can potentially help with the identification of patients with MOH development. Due to the relevance of the problem, the authors believe that further investigation of the MOH proteomic biomarkers in different ethnic and racial groups of patients with primary headache is necessary. In addition, it is important to investigate whether medications of different drug classes influence the levels of serum and urine proteomic biomarkers.
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Biomarcadores/sangre , Biomarcadores/orina , Cefaleas Secundarias/sangre , Cefaleas Secundarias/orina , Proteoma , Dolor Crónico/sangre , Dolor Crónico/tratamiento farmacológico , Dolor Crónico/genética , Dolor Crónico/orina , Cefaleas Secundarias/inducido químicamente , Cefaleas Secundarias/genética , HumanosRESUMEN
Chronic pain syndromes are an important medical problem generated by various molecular, genetic, and pathophysiologic mechanisms. Back pain, neuropathic pain, and posttraumatic pain are the most important pathological processes associated with chronic pain in adults. Standard approaches to the treatment of them do not solve the problem of pain chronicity. This is the reason for the search for new personalized strategies for the prevention and treatment of chronic pain. The nitric oxide (NO) system can play one of the key roles in the development of peripheral pain and its chronicity. The purpose of the study is to review publications devoted to changes in the NO system in patients with peripheral chronical pain syndromes. We have carried out a search for the articles published in e-Library, PubMed, Oxford Press, Clinical Case, Springer, Elsevier, and Google Scholar databases. The search was carried out using keywords and their combinations. The role of NO and NO synthases (NOS) isoforms in peripheral pain development and chronicity was demonstrated primarily from animal models to humans. The most studied is the neuronal NOS (nNOS). The role of inducible NOS (iNOS) and endothelial NOS (eNOS) is still under investigation. Associative genetic studies have shown that single nucleotide variants (SNVs) of NOS1, NOS2, and NOS3 genes encoding nNOS, iNOS, and eNOS may be associated with acute and chronic peripheral pain. Prospects for the use of NOS inhibitors to modulate the effect of drugs used to treat peripheral pain syndrome are discussed. Associative genetic studies of SNVs NOS1, NOS2, and NOS3 genes are important for understanding genetic predictors of peripheral pain chronicity and development of new personalized pharmacotherapy strategies.
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Óxido Nítrico Sintasa/metabolismo , Óxido Nítrico/metabolismo , Manejo del Dolor , Dolor/metabolismo , Medicina de Precisión , Animales , Terapia Combinada , Susceptibilidad a Enfermedades , Predisposición Genética a la Enfermedad , Humanos , Dolor/etiología , Polimorfismo de Nucleótido Simple , Medicina de Precisión/métodosRESUMEN
Medullary breast carcinoma (MBC) is a rare subtype of triple-negative breast cancer with specific genomic features within the spectrum of basal-like carcinoma (BLC). In this study of 19 MBCs and 36 non-MBC BLCs, we refined the transcriptomic and genomic knowledge about this entity. Unsupervised and supervised analysis of transcriptomic profiles confirmed that MBC clearly differs from non-MBC BLC, with 92 genes overexpressed and 154 genes underexpressed in MBC compared with non-MBC BLC. Immunity-related pathways are the most differentially represented pathways in MBC compared with non-MBC BLC. The proapoptotic gene BCLG (official name BCL2L14) is by far the most intensely overexpressed gene in MBC. A quantitative RT-PCR validation study conducted in 526 breast tumors corresponding to all molecular subtypes documented the specificity of BCLG overexpression in MBC, which was confirmed at the protein level by immunohistochemistry. We also found that most MBCs belong to the immunomodulatory triple-negative breast cancer subtype. Using pan-genomic analysis, it was found that MBC harbors more losses of heterozygosity than non-MBC BLC. These observations corroborate the notion that MBC remains a distinct entity that could benefit from specific treatment strategies (such as deescalation or targeted therapy) adapted to this rare tumor type.
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Carcinoma Medular/genética , Proteínas Proto-Oncogénicas c-bcl-2/genética , Neoplasias de la Mama Triple Negativas/genética , Proteína BRCA2/genética , ADN de Neoplasias/metabolismo , Femenino , Perfilación de la Expresión Génica , Genes Relacionados con las Neoplasias/genética , Humanos , Pérdida de Heterocigocidad/genética , ARN Neoplásico/metabolismo , Estudios Retrospectivos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Ubiquitina-Proteína Ligasas/genéticaRESUMEN
BACKGROUND: The ataxia telangiectasia mutated (ATM) gene is a moderate-risk breast cancer susceptibility gene; germline loss-of-function variants are found in up to 3% of hereditary breast and ovarian cancer (HBOC) families who undergo genetic testing. So far, no clear histopathological and molecular features of breast tumours occurring in ATM deleterious variant carriers have been described, but identification of an ATM-associated tumour signature may help in patient management. METHODS: To characterise hallmarks of ATM-associated tumours, we performed systematic pathology review of tumours from 21 participants from ataxia-telangiectasia families and 18 participants from HBOC families, as well as copy number profiling on a subset of 23 tumours. Morphology of ATM-associated tumours was compared with that of 599 patients with no BRCA1 and BRCA2 mutations from a hospital-based series, as well as with data from The Cancer Genome Atlas. Absolute copy number and loss of heterozygosity (LOH) profiles were obtained from the OncoScan SNP array. In addition, we performed whole-genome sequencing on four tumours from ATM loss-of-function variant carriers with available frozen material. RESULTS: We found that ATM-associated tumours belong mostly to the luminal B subtype, are tetraploid and show LOH at the ATM locus at 11q22-23. Unlike tumours in which BRCA1 or BRCA2 is inactivated, tumours arising in ATM deleterious variant carriers are not associated with increased large-scale genomic instability as measured by the large-scale state transitions signature. Losses at 13q14.11-q14.3, 17p13.2-p12, 21p11.2-p11.1 and 22q11.23 were observed. Somatic alterations at these loci may therefore represent biomarkers for ATM testing and harbour driver mutations in potentially 'druggable' genes that would allow patients to be directed towards tailored therapeutic strategies. CONCLUSIONS: Although ATM is involved in the DNA damage response, ATM-associated tumours are distinct from BRCA1-associated tumours in terms of morphological characteristics and genomic alterations, and they are also distinguishable from sporadic breast tumours, thus opening up the possibility to identify ATM variant carriers outside the ataxia-telangiectasia disorder and direct them towards effective cancer risk management and therapeutic strategies.
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Proteínas de la Ataxia Telangiectasia Mutada/genética , Proteína BRCA1/genética , Neoplasias de la Mama Masculina/genética , Neoplasias de la Mama/genética , Predisposición Genética a la Enfermedad , Adulto , Anciano , Ataxia Telangiectasia/complicaciones , Ataxia Telangiectasia/genética , Ataxia Telangiectasia/patología , Proteína BRCA2/genética , Neoplasias de la Mama/clasificación , Neoplasias de la Mama/complicaciones , Neoplasias de la Mama/patología , Neoplasias de la Mama Masculina/clasificación , Neoplasias de la Mama Masculina/complicaciones , Neoplasias de la Mama Masculina/patología , Daño del ADN/genética , Reparación del ADN/genética , Femenino , Pruebas Genéticas , Genómica , Mutación de Línea Germinal/genética , Humanos , Pérdida de Heterocigocidad/genética , Masculino , Persona de Mediana Edad , Eliminación de Secuencia/genéticaRESUMEN
Disappearance of the Barr body is considered a hallmark of cancer, although whether this corresponds to genetic loss or to epigenetic instability and transcriptional reactivation is unclear. Here we show that breast tumors and cell lines frequently display major epigenetic instability of the inactive X chromosome, with highly abnormal 3D nuclear organization and global perturbations of heterochromatin, including gain of euchromatic marks and aberrant distributions of repressive marks such as H3K27me3 and promoter DNA methylation. Genome-wide profiling of chromatin and transcription reveal modified epigenomic landscapes in cancer cells and a significant degree of aberrant gene activity from the inactive X chromosome, including several genes involved in cancer promotion. We demonstrate that many of these genes are aberrantly reactivated in primary breast tumors, and we further demonstrate that epigenetic instability of the inactive X can lead to perturbed dosage of X-linked factors. Taken together, our study provides the first integrated analysis of the inactive X chromosome in the context of breast cancer and establishes that epigenetic erosion of the inactive X can lead to the disappearance of the Barr body in breast cancer cells. This work offers new insights and opens up the possibility of exploiting the inactive X chromosome as an epigenetic biomarker at the molecular and cytological levels in cancer.
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Neoplasias de la Mama/genética , Cromosomas Humanos X/genética , Epigénesis Genética/genética , Genes Ligados a X/genética , Inactivación del Cromosoma X/genética , Antígenos de Neoplasias/metabolismo , Biomarcadores de Tumor/genética , Línea Celular Tumoral , Núcleo Celular/patología , ADN Helicasas/metabolismo , Metilación de ADN/genética , Femenino , Histona Desacetilasas/metabolismo , Histona Demetilasas/genética , Histona Demetilasas/metabolismo , Histonas/genética , Humanos , Proteínas de Neoplasias/metabolismo , Proteínas Nucleares/metabolismo , Regiones Promotoras Genéticas/genética , ARN Largo no Codificante/genética , Proteínas Represoras/metabolismo , Cromatina Sexual/genética , Transcripción Genética/genética , Transducina/metabolismo , Proteínas Supresoras de Tumor , Ubiquitina-Proteína Ligasas/genética , Ubiquitina-Proteína Ligasas/metabolismo , Proteína Nuclear Ligada al Cromosoma XRESUMEN
BACKGROUND: The introduction of complex antiretroviral therapy has resulted in signifi cant decrease in the mortality rate of HIV positive patients, but it still remains unacceptably high, especially in some groups of patients. AIM: To investigate the death rate in patients with HIV/AIDS, lethality and mortality in co-infection, and the most common causes and predictors of fatal outcome, focused on early diagnosis and appropriate therapy. MATERIALS AND METHODS: The study included 53 deceased patients with HIV/AIDS, monitored at the Clinic of Infectious Diseases in St George University Hospital, Plovdiv between 01.01.2010 and 31.12.2014. The methods of research included clinical analysis, laboratory tests, microbiological and serological tests (HCV, HBV, toxoplasmosis), ELISA, PCR. Statistical analysis was performed by descriptive statistics, the Student's t-test, the method of Van der Ward, and regression analysis (logistic regression). RESULTS: During the study period 316 patients with HIV/AIDS were monitored, 53 of them with lethal outcome. Lethality was 16.7% for the whole group; in intravenous drug users - 13.8%; in co-infected patients: HIV/M. tuberculosis - 46%, in HIV/HCV - 17.8%. Lethality and mortality in HIV(+) patients with co-infections in populations of diff erent age, gender, duration since starting ÑÐRТ and degree of immunodefi ciency (according to CD4, VL) was compared with the lethality and mortality in patients with these conditions from the general population. CONCLUSIONS: Fatal outcome in patients with HIV/AIDS was most commonly associated with co-infections HIV/M. tuberculosis and HIV/HCV. Predictors of a fatal outcome are pulmonary tuberculosis, advanced immunodefi ciency with VL> 500 000 c/µL and CD4 <100/mm3, absence or non-systemic antiretroviral therapy.
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Infecciones por VIH/mortalidad , Adulto , Bulgaria/epidemiología , Coinfección/mortalidad , Femenino , Infecciones por VIH/transmisión , Humanos , Masculino , Factores de TiempoRESUMEN
Therapeutic strategies targeting Homologous Recombination Deficiency (HRD) in breast cancer requires patient stratification. The LST (Large-scale State Transitions) genomic signature previously validated for triple-negative breast carcinomas (TNBC) was evaluated as biomarker of HRD in luminal (hormone receptor positive) and HER2-overexpressing (HER2+) tumors. The LST genomic signature related to the number of large-scale chromosomal breakpoints in SNP-array tumor profile was applied to identify HRD in in-house and TCGA sets of breast tumors, in which the status of BRCA1/2 and other genes was also investigated. In the in-house dataset, HRD was predicted in 5% (20/385) of sporadic tumors luminal or HER2+ by the LST genomic signature and the inactivation of BRCA1, BRCA2 or RAD51C confirmed this prediction in 75% (12/16) of the tested cases. In 14% (6/43) of tumors occurring in BRCA1/2 mutant carriers, the corresponding wild-type allele was retained emphasizing the importance of determining the tumor status. In the TCGA luminal and HER2+ subtypes HRD incidence was estimated at 5% (18/329, 95%CI: 5-8%) and 2% (1/59, 95%CI: 2-9%), respectively. In TNBC cisplatin-based neo-adjuvant clinical trials, HRD is shown to be a necessary condition for cisplatin sensitivity. This analysis demonstrates the high performance of the LST genomic signature for HRD detection in breast cancers, which suggests its potential as a biomarker for genetic testing and patient stratification for clinical trials evaluating platinum salts and PARP inhibitors.
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Biomarcadores de Tumor/genética , Neoplasias de la Mama/genética , Carcinoma/genética , Reparación del ADN por Recombinación/genética , Transcriptoma/genética , Neoplasias de la Mama/patología , Carcinoma/patología , Rotura Cromosómica , Femenino , Genes BRCA2 , Humanos , Análisis de Secuencia por Matrices de Oligonucleótidos , Receptor ErbB-2/genéticaRESUMEN
The genetic cause of some familial nonsyndromic renal cell carcinomas (RCC) defined by at least two affected first-degree relatives is unknown. By combining whole-exome sequencing and tumor profiling in a family prone to cases of RCC, we identified a germline BAP1 mutation c.277A>G (p.Thr93Ala) as the probable genetic basis of RCC predisposition. This mutation segregated with all four RCC-affected relatives. Furthermore, BAP1 was found to be inactivated in RCC-affected individuals from this family. No BAP1 mutations were identified in 32 familial cases presenting with only RCC. We then screened for germline BAP1 deleterious mutations in familial aggregations of cancers within the spectrum of the recently described BAP1-associated tumor predisposition syndrome, including uveal melanoma, malignant pleural mesothelioma, and cutaneous melanoma. Among the 11 families that included individuals identified as carrying germline deleterious BAP1 mutations, 6 families presented with 9 RCC-affected individuals, demonstrating a significantly increased risk for RCC. This strongly argues that RCC belongs to the BAP1 syndrome and that BAP1 is a RCC-predisposition gene.
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Carcinoma de Células Renales/genética , Mutación de Línea Germinal , Neoplasias Renales/genética , Mutación Missense , Proteínas Supresoras de Tumor/genética , Ubiquitina Tiolesterasa/genética , Adulto , Secuencia de Bases , Carcinoma de Células Renales/enzimología , Carcinoma de Células Renales/patología , Exoma , Femenino , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Humanos , Neoplasias Renales/enzimología , Neoplasias Renales/patología , Masculino , Persona de Mediana Edad , Linaje , Polimorfismo de Nucleótido Simple , Análisis de Secuencia de ADNRESUMEN
There is a well-established association between aging and the onset of metastasis. Although the mechanisms through which age impinges upon the malignant phenotype remain uncharacterized, the role of a senescent microenvironment has been emphasized. We reported previously that human epithelial cells that undergo telomere-driven chromosome instability (T-CIN) display global microRNA (miR) deregulation and develop migration and invasion capacities. Here, we show that post-crisis cells are not able to form tumors unless a senescent microenvironment is provided. The characterization of cell lines established from such tumors revealed that these cells have acquired cell autonomous tumorigenicity, giving rise to heterogeneous tumors. Further experiments demonstrate that explanted cells, while displaying differences in cell differentiation markers, are all endowed of enhanced stem cell properties including self-renewal and multilineage differentiation capacity. Treatments of T-CIN+ cells with senescence-conditioned media induce sphere formation exclusively in cells with senescence-associated tumorigenicity, a capacity that depends on miR-145 repression. These results indicate that the senescent microenvironment, while promoting further transdifferentiations in cells with genome instability, is able to propel the progression of premalignant cells towards a malignant, cell stem-like state.
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Envejecimiento/genética , Transformación Celular Neoplásica/genética , MicroARNs/biosíntesis , Células Madre Neoplásicas/patología , Microambiente Tumoral/genética , Envejecimiento/patología , Diferenciación Celular/genética , Línea Celular Tumoral , Movimiento Celular/genética , Proliferación Celular/genética , Senescencia Celular/genética , Inestabilidad Cromosómica/genética , Células Epiteliales/metabolismo , Células Epiteliales/patología , Regulación Neoplásica de la Expresión Génica , Humanos , MicroARNs/genética , Invasividad Neoplásica/genética , Metástasis de la Neoplasia , Telómero/genéticaRESUMEN
The treatment of epithelial ovarian cancer (EOC) is narrowly focused despite the heterogeneity of this disease in which outcomes remain poor. To stratify EOC patients for targeted therapy, we developed an approach integrating expression and genomic analyses including the BRCAness status. Gene expression and genomic profiling were used to identify genes recurrently (>5%) amplified and overexpressed in 105 EOC. The LST (Large-scale State Transition) genomic signature of BRCAness was applied to define molecular subgroups of EOC. Amplified/overexpressed genes clustered mainly in 3q, 8q, 19p and 19q. These changes were generally found mutually exclusive. In the 85 patients for which the genomic signature could be determined, genomic BRCAness was found in 52 cases (61.1%) and non-BRCAness in 33 (38.8%). A striking mutual exclusivity was observed between BRCAness and amplification/overexpression data. Whereas 3q and 8q alterations were preferentially observed in BRCAness EOC, most alterations on chromosome 19 were in non-BRCAness cases. CCNE1 (19q12) and BRD4 (19p13.1) amplification/overexpression was found in 19/33 (57.5%) of non-BRCAness cases. Such disequilibrium was also found in the TCGA EOC data set used for validation. Potential target genes are frequently amplified/overexpressed in non-BRCAness EOC. We report that BRD4, already identified as a target in several tumor models, is a new potential target in high grade non-BRCAness ovarian carcinoma.
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Cromosomas Humanos Par 19/genética , Ciclina E/genética , Neoplasias Glandulares y Epiteliales/genética , Neoplasias Glandulares y Epiteliales/patología , Proteínas Nucleares/genética , Proteínas Oncogénicas/genética , Neoplasias Ováricas/genética , Neoplasias Ováricas/patología , Factores de Transcripción/genética , Carcinoma Epitelial de Ovario , Proteínas de Ciclo Celular , Cromosomas Humanos Par 3/genética , Cromosomas Humanos Par 8/genética , Femenino , Amplificación de Genes , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Humanos , Persona de Mediana EdadRESUMEN
Metaplastic breast carcinoma is a rare and aggressive histologic type of breast cancer, preferentially displaying a triple-negative phenotype. We sought to define the transcriptomic heterogeneity of metaplastic breast cancers on the basis of current gene expression microarray-based classifiers, and to determine whether these tumors display gene copy number profiles consistent with those of BRCA1-associated breast cancers. Twenty-eight consecutive triple-negative metaplastic breast carcinomas were reviewed, and the metaplastic component present in each frozen specimen was defined (ie, spindle cell, squamous, chondroid metaplasia). RNA and DNA extracted from frozen sections with tumor cell content >60% were subjected to gene expression (Illumina HumanHT-12 v4) and copy number profiling (Affymetrix SNP 6.0), respectively. Using the best practice PAM50/claudin-low microarray-based classifier, all metaplastic breast carcinomas with spindle cell metaplasia were of claudin-low subtype, whereas those with squamous or chondroid metaplasia were preferentially of basal-like subtype. Triple-negative breast cancer subtyping using a dedicated website (http://cbc.mc.vanderbilt.edu/tnbc/) revealed that all metaplastic breast carcinomas with chondroid metaplasia were of mesenchymal-like subtype, spindle cell carcinomas preferentially of unstable or mesenchymal stem-like subtype, and those with squamous metaplasia were of multiple subtypes. None of the cases was classified as immunomodulatory or luminal androgen receptor subtype. Integrative clustering, combining gene expression and gene copy number data, revealed that metaplastic breast carcinomas with spindle cell and chondroid metaplasia were preferentially classified as of integrative clusters 4 and 9, respectively, whereas those with squamous metaplasia were classified into six different clusters. Eight of the 26 metaplastic breast cancers subjected to SNP6 analysis were classified as BRCA1-like. The diversity of histologic features of metaplastic breast carcinomas is reflected at the transcriptomic level, and an association between molecular subtypes and histology was observed. BRCA1-like genomic profiles were found only in a subset (31%) of metaplastic breast cancers, and were not associated with a specific molecular or histologic subtype.
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Carcinoma/genética , Carcinoma/patología , Neoplasias de la Mama Triple Negativas/genética , Neoplasias de la Mama Triple Negativas/patología , Biomarcadores de Tumor/análisis , Femenino , Perfilación de la Expresión Génica/métodos , Humanos , Inmunohistoquímica , Metaplasia/genética , Metaplasia/patología , Persona de Mediana Edad , Análisis de Secuencia por Matrices de OligonucleótidosRESUMEN
MOTIVATION: Because of its low cost, amplicon sequencing, also known as ultra-deep targeted sequencing, is now becoming widely used in oncology for detection of actionable mutations, i.e. mutations influencing cell sensitivity to targeted therapies. Amplicon sequencing is based on the polymerase chain reaction amplification of the regions of interest, a process that considerably distorts the information on copy numbers initially present in the tumor DNA. Therefore, additional experiments such as single nucleotide polymorphism (SNP) or comparative genomic hybridization (CGH) arrays often complement amplicon sequencing in clinics to identify copy number status of genes whose amplification or deletion has direct consequences on the efficacy of a particular cancer treatment. So far, there has been no proven method to extract the information on gene copy number aberrations based solely on amplicon sequencing. RESULTS: Here we present ONCOCNV, a method that includes a multifactor normalization and annotation technique enabling the detection of large copy number changes from amplicon sequencing data. We validated our approach on high and low amplicon density datasets and demonstrated that ONCOCNV can achieve a precision comparable with that of array CGH techniques in detecting copy number aberrations. Thus, ONCOCNV applied on amplicon sequencing data would make the use of additional array CGH or SNP array experiments unnecessary.
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Dosificación de Gen , Genes Relacionados con las Neoplasias , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Análisis de Secuencia de ADN/métodos , Hibridación Genómica Comparativa , ADN de Neoplasias/química , Exoma , Femenino , Humanos , Masculino , Reacción en Cadena de la Polimerasa , Polimorfismo de Nucleótido SimpleRESUMEN
INTRODUCTION: Pure invasive micropapillary carcinoma (IMPC) is a special type of breast carcinoma characterised by clusters of cells presenting polarity abnormalities. The biological alterations underlying this pattern remain unknown. METHODS: Pangenomic analysis (n=39), TP53 (n=43) and PIK3CA (n=41) sequencing in a series of IMPCs were performed. A subset of cases was also analysed with whole-exome sequencing (n=4) and RNA sequencing (n=6). Copy number variation profiles were compared with those of oestrogen receptors and grade-matched invasive ductal carcinomas (IDCs) of no special type. RESULTS: Unsupervised analysis of genomic data distinguished two IMPC subsets: one (Sawtooth/8/16) exhibited a significant increase in 16p gains (71%), and the other (Firestorm/Amplifier) was characterised by a high frequency of 8q (35%), 17q (20% to 46%) and 20q (23% to 30%) amplifications and 17p loss (74%). TP53 mutations (10%) were more frequently identified in the amplifier subset, and PIK3CA mutations (4%) were detected in both subsets. Compared to IDC, IMPC exhibited specific loss of the 6q16-q22 region (45%), which is associated with downregulation of FOXO3 and SEC63 gene expression. SEC63 and FOXO3 missense mutations were identified in one case each (2%). Whole-exome sequencing combined with RNA sequencing of IMPC allowed us to identify somatic mutations in genes involved in polarity, DNAH9 and FMN2 (8% and 2%, respectively) or ciliogenesis, BBS12 and BBS9 (2% each) or genes coding for endoplasmic reticulum protein, HSP90B1 and SPTLC3 (2% each) and cytoskeleton, UBR4 and PTPN21 (2% each), regardless of the genomic subset. The intracellular biological function of the mutated genes identified by gene ontology analysis suggests a driving role in the clinicopathological characteristics of IMPC. CONCLUSION: In our comprehensive molecular analysis of IMPC, we identified numerous genomic alterations without any recurrent fusion genes. Recurrent somatic mutations of genes participating in cellular polarity and shape suggest that they, together with other biological alterations (such as epigenetic modifications and stromal alterations), could contribute to the morphological pattern of IMPC. Though none of the individual abnormalities demonstrated specificity for IMPC, whether their combination in IMPC may have a cumulative effect that drives the abnormal polarity of IMPC needs to be examined further with in vitro experiments.
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Neoplasias de la Mama/genética , Carcinoma Ductal de Mama/genética , Polaridad Celular/genética , Invasividad Neoplásica/genética , Dineínas Axonemales/genética , Secuencia de Bases , Mama/patología , Neoplasias de la Mama/patología , Proteínas de Unión a Calmodulina/genética , Carcinoma Ductal de Mama/patología , Chaperoninas , Fosfatidilinositol 3-Quinasa Clase I , Proteínas del Citoesqueleto/genética , Variaciones en el Número de Copia de ADN , Exoma/genética , Femenino , Proteína Forkhead Box O3 , Factores de Transcripción Forkhead/biosíntesis , Factores de Transcripción Forkhead/genética , Forminas , Amplificación de Genes/genética , Chaperoninas del Grupo II/genética , Humanos , Glicoproteínas de Membrana/genética , Proteínas de la Membrana/biosíntesis , Proteínas de la Membrana/genética , Proteínas de Microfilamentos/biosíntesis , Chaperonas Moleculares , Mutación Missense , Proteínas de Neoplasias/genética , Proteínas Nucleares/biosíntesis , Fosfatidilinositol 3-Quinasas/genética , Proteínas Tirosina Fosfatasas no Receptoras/genética , Proteínas de Unión al ARN , Receptor ErbB-2/biosíntesis , Receptores de Estrógenos/biosíntesis , Estudios Retrospectivos , Análisis de Secuencia de ADN , Análisis de Secuencia de ARN , Eliminación de Secuencia/genética , Serina C-Palmitoiltransferasa/genética , Proteína p53 Supresora de Tumor/genética , Ubiquitina-Proteína LigasasRESUMEN
BRCA2 is the major high-penetrance predisposition gene for luminal (estrogen receptor [ER] positive) breast cancers. However, many BRCA2 mutant carriers lack family history of breast/ovarian cancers and do not benefit from genetic testing. Specific genomic features associated with BRCA2 inactivation in tumors could help identify patients for whom a genetic test for BRCA2 may be proposed. A series of ER-positive invasive ductal carcinomas (IDCs) including 30 carriers of BRCA2 mutations and 215 control cases was studied by single-nucleotide polymorphism (SNP) arrays. Cases and controls were stratified by grade and HER2 status. Independently, 7 BRCA2 and 51 control cases were used for validation. Absolute copy number and Loss of heterozygosity (LOH) profiles were obtained from SNP arrays by the genome alteration print (GAP) method. BRCA2 tumors were observed to display a discriminatively greater number of chromosomal breaks calculated after filtering out and smoothing <3 Mb variations. This argues for a BRCA2-associated genomic instability responsible for long-segment aberrations. Co-occurrence of two genomic features-LOH of 13q13 and 14q32-was found to predict BRCA2 status with 90% of sensitivity and 87% of specificity in discovery series of high-grade HER2-negative IDCs and 100% of sensitivity and 88% of specificity in an independent series of 58 IDCs. Estimated positive predictive value was 17.2% (confidence interval: 6.7-33.5) in the whole series. In conclusion, the simplified BRCA2 classifier based on the co-occurrence of LOH at 13q13 and 14q32 could provide an indication to test for BRCA2 mutation in patients with ER-positive IDC.