Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 36
Filtrar
1.
Nature ; 594(7863): 418-423, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33953400

RESUMO

Although RAF monomer inhibitors (type I.5, BRAF(V600)) are clinically approved for the treatment of BRAFV600-mutant melanoma, they are ineffective in non-BRAFV600 mutant cells1-3. Belvarafenib is a potent and selective RAF dimer (type II) inhibitor that exhibits clinical activity in patients with BRAFV600E- and NRAS-mutant melanomas. Here we report the first-in-human phase I study investigating the maximum tolerated dose, and assessing the safety and preliminary efficacy of belvarafenib in BRAFV600E- and RAS-mutated advanced solid tumours (NCT02405065, NCT03118817). By generating belvarafenib-resistant NRAS-mutant melanoma cells and analysing circulating tumour DNA from patients treated with belvarafenib, we identified new recurrent mutations in ARAF within the kinase domain. ARAF mutants conferred resistance to belvarafenib in both a dimer- and a kinase activity-dependent manner. Belvarafenib induced ARAF mutant dimers, and dimers containing mutant ARAF were active in the presence of inhibitor. ARAF mutations may serve as a general resistance mechanism for RAF dimer inhibitors as the mutants exhibit reduced sensitivity to a panel of type II RAF inhibitors. The combination of RAF plus MEK inhibition may be used to delay ARAF-driven resistance and suggests a rational combination for clinical use. Together, our findings reveal specific and compensatory functions for the ARAF isoform and implicate ARAF mutations as a driver of resistance to RAF dimer inhibitors.


Assuntos
Resistencia a Medicamentos Antineoplásicos/genética , Melanoma/tratamento farmacológico , Melanoma/genética , Mutação , Proteínas Proto-Oncogênicas A-raf/antagonistas & inibidores , Proteínas Proto-Oncogênicas A-raf/genética , Quinases raf/antagonistas & inibidores , Animais , Linhagem Celular , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Feminino , Humanos , Melanoma/patologia , Camundongos , Multimerização Proteica/efeitos dos fármacos , Proteínas Proto-Oncogênicas A-raf/química , Quinases raf/química
2.
Nature ; 550(7677): 534-538, 2017 10 26.
Artigo em Inglês | MEDLINE | ID: mdl-29045385

RESUMO

The ubiquitin system regulates essential cellular processes in eukaryotes. Ubiquitin is ligated to substrate proteins as monomers or chains and the topology of ubiquitin modifications regulates substrate interactions with specific proteins. Thus ubiquitination directs a variety of substrate fates including proteasomal degradation. Deubiquitinase enzymes cleave ubiquitin from substrates and are implicated in disease; for example, ubiquitin-specific protease-7 (USP7) regulates stability of the p53 tumour suppressor and other proteins critical for tumour cell survival. However, developing selective deubiquitinase inhibitors has been challenging and no co-crystal structures have been solved with small-molecule inhibitors. Here, using nuclear magnetic resonance-based screening and structure-based design, we describe the development of selective USP7 inhibitors GNE-6640 and GNE-6776. These compounds induce tumour cell death and enhance cytotoxicity with chemotherapeutic agents and targeted compounds, including PIM kinase inhibitors. Structural studies reveal that GNE-6640 and GNE-6776 non-covalently target USP7 12 Å distant from the catalytic cysteine. The compounds attenuate ubiquitin binding and thus inhibit USP7 deubiquitinase activity. GNE-6640 and GNE-6776 interact with acidic residues that mediate hydrogen-bond interactions with the ubiquitin Lys48 side chain, suggesting that USP7 preferentially interacts with and cleaves ubiquitin moieties that have free Lys48 side chains. We investigated this idea by engineering di-ubiquitin chains containing differential proximal and distal isotopic labels and measuring USP7 binding by nuclear magnetic resonance. This preferential binding protracted the depolymerization kinetics of Lys48-linked ubiquitin chains relative to Lys63-linked chains. In summary, engineering compounds that inhibit USP7 activity by attenuating ubiquitin binding suggests opportunities for developing other deubiquitinase inhibitors and may be a strategy more broadly applicable to inhibiting proteins that require ubiquitin binding for full functional activity.


Assuntos
Aminopiridinas/química , Aminopiridinas/farmacologia , Indazóis/química , Indazóis/farmacologia , Fenóis/química , Fenóis/farmacologia , Piridinas/química , Piridinas/farmacologia , Peptidase 7 Específica de Ubiquitina/antagonistas & inibidores , Ubiquitina/metabolismo , Animais , Ligação Competitiva , Linhagem Celular Tumoral , Sinergismo Farmacológico , Feminino , Humanos , Camundongos , Camundongos SCID , Modelos Moleculares , Neoplasias/tratamento farmacológico , Neoplasias/enzimologia , Neoplasias/patologia , Ligação Proteica , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Proteínas Proto-Oncogênicas c-pim-1/antagonistas & inibidores , Especificidade por Substrato , Proteína Supressora de Tumor p53/deficiência , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Ubiquitina/química , Peptidase 7 Específica de Ubiquitina/química , Peptidase 7 Específica de Ubiquitina/deficiência , Peptidase 7 Específica de Ubiquitina/metabolismo
3.
Nature ; 520(7547): 307-11, 2015 Apr 16.
Artigo em Inglês | MEDLINE | ID: mdl-25877200

RESUMO

Cell line misidentification, contamination and poor annotation affect scientific reproducibility. Here we outline simple measures to detect or avoid cross-contamination, present a framework for cell line annotation linked to short tandem repeat and single nucleotide polymorphism profiles, and provide a catalogue of synonymous cell lines. This resource will enable our community to eradicate the use of misidentified lines and generate credible cell-based data.


Assuntos
Linhagem Celular/classificação , Linhagem Celular/metabolismo , Curadoria de Dados , Guias como Assunto , Separação Celular , Genótipo , Repetições de Microssatélites/genética , Polimorfismo de Nucleotídeo Único/genética , Controle de Qualidade , Reprodutibilidade dos Testes , Especificidade da Espécie , Terminologia como Assunto
4.
Proc Natl Acad Sci U S A ; 110(21): 8632-7, 2013 May 21.
Artigo em Inglês | MEDLINE | ID: mdl-23657012

RESUMO

Germ-line mutations in PALB2 lead to a familial predisposition to breast and pancreatic cancer or to Fanconi Anemia subtype N. PALB2 performs its tumor suppressor role, at least in part, by supporting homologous recombination-type double strand break repair (HR-DSBR) through physical interactions with BRCA1, BRCA2, and RAD51. To further understand the mechanisms underlying PALB2-mediated DNA repair and tumor suppression functions, we targeted Palb2 in the mouse. Palb2-deficient murine ES cells recapitulated DNA damage defects caused by PALB2 depletion in human cells, and germ-line deletion of Palb2 led to early embryonic lethality. Somatic deletion of Palb2 driven by K14-Cre led to mammary tumor formation with long latency. Codeletion of both Palb2 and Tumor protein 53 (Trp53) accelerated mammary tumor formation. Like BRCA1 and BRCA2 mutant breast cancers, these tumors were defective in RAD51 focus formation, reflecting a defect in Palb2 HR-DSBR function, a strongly suspected contributor to Brca1, Brca2, and Palb2 mammary tumor development. However, unlike the case of Brca1-mutant cells, Trp53bp1 deletion failed to rescue the genomic instability of Palb2- or Brca2-mutant primary lymphocytes. Therefore, Palb2-driven DNA damage control is, in part, distinct from that executed by Brca1 and more similar to that of Brca2. The mechanisms underlying Palb2 mammary tumor suppression functions can now be explored genetically in vivo.


Assuntos
Neoplasias da Mama/metabolismo , Neoplasias Mamárias Experimentais/metabolismo , Síndromes Neoplásicas Hereditárias/metabolismo , Proteínas Nucleares/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Animais , Proteína BRCA1/genética , Proteína BRCA1/metabolismo , Proteína BRCA2/genética , Proteína BRCA2/metabolismo , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Modelos Animais de Doenças , Proteína do Grupo de Complementação N da Anemia de Fanconi , Feminino , Deleção de Genes , Humanos , Neoplasias Mamárias Experimentais/genética , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Mutantes , Síndromes Neoplásicas Hereditárias/genética , Síndromes Neoplásicas Hereditárias/patologia , Proteínas Nucleares/genética , Rad51 Recombinase/genética , Rad51 Recombinase/metabolismo , Proteína Supressora de Tumor p53/genética , Proteínas Supressoras de Tumor/genética
5.
Genome Res ; 22(12): 2315-27, 2012 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-23033341

RESUMO

Lung cancer is a highly heterogeneous disease in terms of both underlying genetic lesions and response to therapeutic treatments. We performed deep whole-genome sequencing and transcriptome sequencing on 19 lung cancer cell lines and three lung tumor/normal pairs. Overall, our data show that cell line models exhibit similar mutation spectra to human tumor samples. Smoker and never-smoker cancer samples exhibit distinguishable patterns of mutations. A number of epigenetic regulators, including KDM6A, ASH1L, SMARCA4, and ATAD2, are frequently altered by mutations or copy number changes. A systematic survey of splice-site mutations identified 106 splice site mutations associated with cancer specific aberrant splicing, including mutations in several known cancer-related genes. RAC1b, an isoform of the RAC1 GTPase that includes one additional exon, was found to be preferentially up-regulated in lung cancer. We further show that its expression is significantly associated with sensitivity to a MAP2K (MEK) inhibitor PD-0325901. Taken together, these data present a comprehensive genomic landscape of a large number of lung cancer samples and further demonstrate that cancer-specific alternative splicing is a widespread phenomenon that has potential utility as therapeutic biomarkers. The detailed characterizations of the lung cancer cell lines also provide genomic context to the vast amount of experimental data gathered for these lines over the decades, and represent highly valuable resources for cancer biology.


Assuntos
Processamento Alternativo , Regulação Neoplásica da Expressão Gênica , Genoma Humano/genética , Neoplasias Pulmonares/genética , Mutação , Transcriptoma , ATPases Associadas a Diversas Atividades Celulares , Adenosina Trifosfatases/genética , Adenosina Trifosfatases/metabolismo , Linhagem Celular Tumoral , Variações do Número de Cópias de DNA , DNA Helicases/genética , DNA Helicases/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Epigenômica , Éxons , Marcadores Genéticos , Heterozigoto , Histona Desmetilases/genética , Histona Desmetilases/metabolismo , Histona-Lisina N-Metiltransferase , Humanos , Cariotipagem/métodos , Neoplasias Pulmonares/patologia , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Polimorfismo de Nucleotídeo Único , Reprodutibilidade dos Testes , Análise de Sequência de RNA , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Regulação para Cima , Proteínas rac1 de Ligação ao GTP/genética , Proteínas rac1 de Ligação ao GTP/metabolismo
6.
Genome Res ; 21(12): 2181-9, 2011 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-21852388

RESUMO

Retroviral and transposon-based insertional mutagenesis (IM) screens are widely used for cancer gene discovery in mice. Exploiting the full potential of IM screens requires methods for high-throughput sequencing and mapping of transposon and retroviral insertion sites. Current protocols are based on ligation-mediated PCR amplification of junction fragments from restriction endonuclease-digested genomic DNA, resulting in amplification biases due to uneven genomic distribution of restriction enzyme recognition sites. Consequently, sequence coverage cannot be used to assess the clonality of individual insertions. We have developed a novel method, called shear-splink, for the semiquantitative high-throughput analysis of insertional mutations. Shear-splink employs random fragmentation of genomic DNA, which reduces unwanted amplification biases. Additionally, shear-splink enables us to assess clonality of individual insertions by determining the number of unique ligation points (LPs) between the adapter and genomic DNA. This parameter serves as a semiquantitative measure of the relative clonality of individual insertions within heterogeneous tumors. Mixing experiments with clonal cell lines derived from mouse mammary tumor virus (MMTV)-induced tumors showed that shear-splink enables the semiquantitative assessment of the clonality of MMTV insertions. Further, shear-splink analysis of 16 MMTV- and 127 Sleeping Beauty (SB)-induced tumors showed enrichment for cancer-relevant insertions by exclusion of irrelevant background insertions marked by single LPs, thereby facilitating the discovery of candidate cancer genes. To fully exploit the use of the shear-splink method, we set up the Insertional Mutagenesis Database (iMDB), offering a publicly available web-based application to analyze both retroviral- and transposon-based insertional mutagenesis data.


Assuntos
DNA de Neoplasias/genética , Bases de Dados Genéticas , Vírus do Tumor Mamário do Camundongo , Mutagênese Insercional , Infecções por Retroviridae/genética , Infecções Tumorais por Vírus/genética , Animais , Análise Mutacional de DNA/métodos , Camundongos
7.
J Pathol ; 224(1): 10-21, 2011 May.
Artigo em Inglês | MEDLINE | ID: mdl-21404276

RESUMO

PALB2 interacts with BRCA1 and BRCA2 in supercomplexes involved in DNA repair via homologous recombination. Heterozygous germline mutations in PALB2 confer a moderate risk of breast cancer, while biallelic PALB2 mutations are linked to a severe form of Fanconi anaemia characterized by early childhood solid tumours and severe chromosomal instability. In contrast to BRCA1- or BRCA2-associated cancers, breast tumours in heterozygous PALB2 mutation carriers do not show loss of the wild-type allele, suggesting PALB2 might be haploinsufficient for tumour suppression. To study the role of PALB2 in development and tumourigenesis, we have generated Palb2(GT) mouse mutants using a gene trap approach. Whereas Palb2(GT/GT) homozygous mutant embryos died at mid-gestation due to massive apoptosis, Palb2(GT/+) heterozygous mice were viable and did not show any obvious abnormalities. Deletion of p53 alleviated the phenotype of Palb2(GT/GT) embryos, but did not rescue embryonic lethality. In addition, loss of p53 did not significantly collaborate with Palb2 heterozygosity in tumourigenesis in heterozygous or homozygous p53 knockout mice. Tumours arising in Palb2(GT/+) ;p53(+/-) or Palb2(GT/+) ;p53(-/-) compound mutant mice retained the wild-type Palb2 allele and did not display increased genomic instability.


Assuntos
Transformação Celular Neoplásica/genética , Desenvolvimento Embrionário/fisiologia , Haploinsuficiência/genética , Proteína Supressora de Tumor p53/deficiência , Proteínas Supressoras de Tumor/deficiência , Animais , Apoptose/genética , Perda do Embrião/genética , Desenvolvimento Embrionário/genética , Proteína do Grupo de Complementação N da Anemia de Fanconi , Feminino , Deleção de Genes , Genes p53 , Instabilidade Genômica/genética , Heterozigoto , Linfoma/genética , Neoplasias Mamárias Experimentais/genética , Camundongos , Camundongos Knockout , Neoplasias do Timo/genética , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/fisiologia
8.
Cell Rep ; 38(6): 110351, 2022 02 08.
Artigo em Inglês | MEDLINE | ID: mdl-35139374

RESUMO

KRAS, which is mutated in ∼30% of all cancers, activates the RAF-MEK-ERK signaling cascade. CRAF is required for growth of KRAS mutant lung tumors, but the requirement for CRAF kinase activity is unknown. Here, we show that subsets of KRAS mutant tumors are dependent on CRAF for growth. Kinase-dead but not dimer-defective CRAF rescues growth inhibition, suggesting that dimerization but not kinase activity is required. Quantitative proteomics demonstrates increased levels of CRAF:ARAF dimers in KRAS mutant cells, and depletion of both CRAF and ARAF rescues the CRAF-loss phenotype. Mechanistically, CRAF depletion causes sustained ERK activation and induction of cell-cycle arrest, while treatment with low-dose MEK or ERK inhibitor rescues the CRAF-loss phenotype. Our studies highlight the role of CRAF in regulating MAPK signal intensity to promote tumorigenesis downstream of mutant KRAS and suggest that disrupting CRAF dimerization or degrading CRAF may have therapeutic benefit.


Assuntos
Carcinogênese/metabolismo , Dimerização , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Animais , Carcinogênese/efeitos dos fármacos , Linhagem Celular Tumoral , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Camundongos , Fosforilação/fisiologia , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas B-raf/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Proteínas ras/genética
9.
PLoS Comput Biol ; 6(1): e1000631, 2010 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-20052266

RESUMO

Tumorigenesis is a multi-step process in which normal cells transform into malignant tumors following the accumulation of genetic mutations that enable them to evade the growth control checkpoints that would normally suppress their growth or result in apoptosis. It is therefore important to identify those combinations of mutations that collaborate in cancer development and progression. DNA copy number alterations (CNAs) are one of the ways in which cancer genes are deregulated in tumor cells. We hypothesized that synergistic interactions between cancer genes might be identified by looking for regions of co-occurring gain and/or loss. To this end we developed a scoring framework to separate truly co-occurring aberrations from passenger mutations and dominant single signals present in the data. The resulting regions of high co-occurrence can be investigated for between-region functional interactions. Analysis of high-resolution DNA copy number data from a panel of 95 hematological tumor cell lines correctly identified co-occurring recombinations at the T-cell receptor and immunoglobulin loci in T- and B-cell malignancies, respectively, showing that we can recover truly co-occurring genomic alterations. In addition, our analysis revealed networks of co-occurring genomic losses and gains that are enriched for cancer genes. These networks are also highly enriched for functional relationships between genes. We further examine sub-networks of these networks, core networks, which contain many known cancer genes. The core network for co-occurring DNA losses we find seems to be independent of the canonical cancer genes within the network. Our findings suggest that large-scale, low-intensity copy number alterations may be an important feature of cancer development or maintenance by affecting gene dosage of a large interconnected network of functionally related genes.


Assuntos
Mapeamento Cromossômico/métodos , Variações do Número de Cópias de DNA/genética , DNA de Neoplasias/genética , Regulação Neoplásica da Expressão Gênica , Genoma Humano/genética , Proteínas de Neoplasias/genética , Transdução de Sinais/genética , Humanos
10.
Cancer Discov ; 11(3): 778-793, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33208393

RESUMO

Hippo pathway dysregulation occurs in multiple cancers through genetic and nongenetic alterations, resulting in translocation of YAP to the nucleus and activation of the TEAD family of transcription factors. Unlike other oncogenic pathways such as RAS, defining tumors that are Hippo pathway-dependent is far more complex due to the lack of hotspot genetic alterations. Here, we developed a machine-learning framework to identify a robust, cancer type-agnostic gene expression signature to quantitate Hippo pathway activity and cross-talk as well as predict YAP/TEAD dependency across cancers. Further, through chemical genetic interaction screens and multiomics analyses, we discover a direct interaction between MAPK signaling and TEAD stability such that knockdown of YAP combined with MEK inhibition results in robust inhibition of tumor cell growth in Hippo dysregulated tumors. This multifaceted approach underscores how computational models combined with experimental studies can inform precision medicine approaches including predictive diagnostics and combination strategies. SIGNIFICANCE: An integrated chemicogenomics strategy was developed to identify a lineage-independent signature for the Hippo pathway in cancers. Evaluating transcriptional profiles using a machine-learning method led to identification of a relationship between YAP/TAZ dependency and MAPK pathway activity. The results help to nominate potential combination therapies with Hippo pathway inhibition.This article is highlighted in the In This Issue feature, p. 521.


Assuntos
Quimioinformática/métodos , Biologia Computacional/métodos , Genômica/métodos , Via de Sinalização Hippo , Sistema de Sinalização das MAP Quinases , Aprendizado de Máquina , Transdução de Sinais , Humanos
11.
BMC Cancer ; 10: 455, 2010 Aug 24.
Artigo em Inglês | MEDLINE | ID: mdl-20735817

RESUMO

BACKGROUND: Genomic gains and losses are a result of genomic instability in many types of cancers. BRCA1- and BRCA2-mutated breast cancers are associated with increased amounts of chromosomal aberrations, presumably due their functions in genome repair. Some of these genomic aberrations may harbor genes whose absence or overexpression may give rise to cellular growth advantage. So far, it has not been easy to identify the driver genes underlying gains and losses. A powerful approach to identify these driver genes could be a cross-species comparison of array comparative genomic hybridization (aCGH) data from cognate mouse and human tumors. Orthologous regions of mouse and human tumors that are commonly gained or lost might represent essential genomic regions selected for gain or loss during tumor development. METHODS: To identify genomic regions that are associated with BRCA1- and BRCA2-mutated breast cancers we compared aCGH data from 130 mouse Brca1Δ/Δ;p53Δ/Δ, Brca2Δ/Δ;p53Δ/Δ and p53Δ/Δ mammary tumor groups with 103 human BRCA1-mutated, BRCA2-mutated and non-hereditary breast cancers. RESULTS: Our genome-wide cross-species analysis yielded a complete collection of loci and genes that are commonly gained or lost in mouse and human breast cancer. Principal common CNAs were the well known MYC-associated gain and RB1/INTS6-associated loss that occurred in all mouse and human tumor groups, and the AURKA-associated gain occurred in BRCA2-related tumors from both species. However, there were also important differences between tumor profiles of both species, such as the prominent gain on chromosome 10 in mouse Brca2Δ/Δ;p53Δ/Δ tumors and the PIK3CA associated 3q gain in human BRCA1-mutated tumors, which occurred in tumors from one species but not in tumors from the other species. This disparity in recurrent aberrations in mouse and human tumors might be due to differences in tumor cell type or genomic organization between both species. CONCLUSIONS: The selection of the oncogenome during mouse and human breast tumor development is markedly different, apart from the MYC gain and RB1-associated loss. These differences should be kept in mind when using mouse models for preclinical studies.


Assuntos
Proteína BRCA1/genética , Proteína BRCA2/genética , Biomarcadores Tumorais/genética , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Perfilação da Expressão Gênica , Mutação/genética , Animais , Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/classificação , Aberrações Cromossômicas , Hibridização Genômica Comparativa , Feminino , Instabilidade Genômica , Humanos , Camundongos , Análise de Sequência com Séries de Oligonucleotídeos , RNA Mensageiro/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Especificidade da Espécie
12.
BMC Cancer ; 10: 662, 2010 Dec 02.
Artigo em Inglês | MEDLINE | ID: mdl-21126340

RESUMO

BACKGROUND: Accurate staging of colorectal cancer (CRC) with clinicopathological parameters is important for predicting prognosis and guiding treatment but provides no information about organ site of metastases. Patterns of genomic aberrations in primary colorectal tumors may reveal a chromosomal signature for organ specific metastases. METHODS: Array Comparative Genomic Hybridization (aCGH) was employed to asses DNA copy number changes in primary colorectal tumors of three distinctive patient groups. This included formalin-fixed, paraffin-embedded tissue of patients who developed liver metastases (LM; n = 36), metastases (PM; n = 37) and a group that remained metastases-free (M0; n = 25).A novel statistical method for identifying recurrent copy number changes, KC-SMART, was used to find specific locations of genomic aberrations specific for various groups. We created a classifier for organ specific metastases based on the aCGH data using Prediction Analysis for Microarrays (PAM). RESULTS: Specifically in the tumors of primary CRC patients who subsequently developed liver metastasis, KC-SMART analysis identified genomic aberrations on chromosome 20q. LM-PAM, a shrunken centroids classifier for liver metastases occurrence, was able to distinguish the LM group from the other groups (M0&PM) with 80% accuracy (78% sensitivity and 86% specificity). The classification is predominantly based on chromosome 20q aberrations. CONCLUSION: Liver specific CRC metastases may be predicted with a high accuracy based on specific genomic aberrations in the primary CRC tumor. The ability to predict the site of metastases is important for improvement of personalized patient management.


Assuntos
Aberrações Cromossômicas , Cromossomos Humanos Par 20 , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Dosagem de Genes , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/secundário , Adulto , Idoso , Distribuição de Qui-Quadrado , Neoplasias Colorretais/mortalidade , Neoplasias Colorretais/terapia , Hibridização Genômica Comparativa , Bases de Dados Genéticas , Feminino , Fixadores , Formaldeído , Perfilação da Expressão Gênica/métodos , Predisposição Genética para Doença , Humanos , Neoplasias Hepáticas/mortalidade , Neoplasias Hepáticas/terapia , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Países Baixos , Análise de Sequência com Séries de Oligonucleotídeos , Inclusão em Parafina , Fenótipo , Valor Preditivo dos Testes , Análise de Sobrevida , Fatores de Tempo , Fixação de Tecidos/métodos , Resultado do Tratamento
13.
Nucleic Acids Res ; 36(2): e13, 2008 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-18187509

RESUMO

Tumor formation is in part driven by DNA copy number alterations (CNAs), which can be measured using microarray-based Comparative Genomic Hybridization (aCGH). Multiexperiment analysis of aCGH data from tumors allows discovery of recurrent CNAs that are potentially causal to cancer development. Until now, multiexperiment aCGH data analysis has been dependent on discretization of measurement data to a gain, loss or no-change state. Valuable biological information is lost when a heterogeneous system such as a solid tumor is reduced to these states. We have developed a new approach which inputs nondiscretized aCGH data to identify regions that are significantly aberrant across an entire tumor set. Our method is based on kernel regression and accounts for the strength of a probe's signal, its local genomic environment and the signal distribution across multiple tumors. In an analysis of 89 human breast tumors, our method showed enrichment for known cancer genes in the detected regions and identified aberrations that are strongly associated with breast cancer subtypes and clinical parameters. Furthermore, we identified 18 recurrent aberrant regions in a new dataset of 19 p53-deficient mouse mammary tumors. These regions, combined with gene expression microarray data, point to known cancer genes and novel candidate cancer genes.


Assuntos
Aberrações Cromossômicas , Genes Neoplásicos , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Animais , Neoplasias da Mama/genética , Perfilação da Expressão Gênica , Humanos , Neoplasias Mamárias Experimentais/genética , Camundongos , Análise de Regressão
14.
Cancer Discov ; 10(2): 232-253, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31699795

RESUMO

With only a fraction of patients responding to cancer immunotherapy, a better understanding of the entire tumor microenvironment is needed. Using single-cell transcriptomics, we chart the fibroblastic landscape during pancreatic ductal adenocarcinoma (PDAC) progression in animal models. We identify a population of carcinoma-associated fibroblasts (CAF) that are programmed by TGFß and express the leucine-rich repeat containing 15 (LRRC15) protein. These LRRC15+ CAFs surround tumor islets and are absent from normal pancreatic tissue. The presence of LRRC15+ CAFs in human patients was confirmed in >80,000 single cells from 22 patients with PDAC as well as by using IHC on samples from 70 patients. Furthermore, immunotherapy clinical trials comprising more than 600 patients across six cancer types revealed elevated levels of the LRRC15+ CAF signature correlated with poor response to anti-PD-L1 therapy. This work has important implications for targeting nonimmune elements of the tumor microenvironment to boost responses of patients with cancer to immune checkpoint blockade therapy. SIGNIFICANCE: This study describes the single-cell landscape of CAFs in pancreatic cancer during in vivo tumor evolution. A TGFß-driven, LRRC15+ CAF lineage is associated with poor outcome in immunotherapy trial data comprising multiple solid-tumor entities and represents a target for combinatorial therapy.This article is highlighted in the In This Issue feature, p. 161.


Assuntos
Fibroblastos Associados a Câncer/imunologia , Carcinoma Ductal Pancreático/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/genética , Inibidores de Checkpoint Imunológico/farmacologia , Proteínas de Membrana/metabolismo , Miofibroblastos/imunologia , Neoplasias Pancreáticas/tratamento farmacológico , Animais , Antígeno B7-H1/antagonistas & inibidores , Antígeno B7-H1/imunologia , Fibroblastos Associados a Câncer/efeitos dos fármacos , Fibroblastos Associados a Câncer/metabolismo , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/imunologia , Linhagem Celular Tumoral , Linhagem da Célula/genética , Linhagem da Célula/imunologia , Ensaios Clínicos como Assunto , Biologia Computacional , Modelos Animais de Doenças , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/imunologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/imunologia , Humanos , Inibidores de Checkpoint Imunológico/uso terapêutico , Camundongos , Miofibroblastos/efeitos dos fármacos , Miofibroblastos/metabolismo , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/imunologia , RNA-Seq , Análise de Célula Única , Fator de Crescimento Transformador beta/metabolismo , Resultado do Tratamento , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/genética , Microambiente Tumoral/imunologia
15.
Cancer Res ; 79(19): 4828-4839, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31416841

RESUMO

Mutations in KEAP1 and NFE2L2 (encoding the protein Nrf2) are prevalent in both adeno and squamous subtypes of non-small cell lung cancer, as well as additional tumor indications. The consequence of these mutations is stabilized Nrf2 and chronic induction of a battery of Nrf2 target genes. We show that knockdown of Nrf2 caused modest growth inhibition of cells growing in two-dimension, which was more pronounced in cell lines expressing mutant KEAP1. In contrast, Nrf2 knockdown caused almost complete regression of established KEAP1-mutant tumors in mice, with little effect on wild-type (WT) KEAP1 tumors. The strong dependency on Nrf2 could be recapitulated in certain anchorage-independent growth environments and was not prevented by excess extracellular glutathione. A CRISPR screen was used to investigate the mechanism(s) underlying this dependence. We identified alternative pathways critical for Nrf2-dependent growth in KEAP1-mutant cell lines, including the redox proteins thioredoxin and peroxiredoxin, as well as the growth factor receptors IGF1R and ERBB3. IGF1R inhibition was effective in KEAP1-mutant cells compared with WT, especially under conditions of anchorage-independent growth. These results point to addiction of KEAP1-mutant tumor cells to Nrf2 and suggest that inhibition of Nrf2 or discrete druggable Nrf2 target genes such as IGF1R could be an effective therapeutic strategy for disabling these tumors. SIGNIFICANCE: This study identifies pathways activated by Nrf2 that are important for the proliferation and tumorigenicity of KEAP1-mutant non-small cell lung cancer.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Transdução de Sinais/fisiologia , Animais , Carcinoma Pulmonar de Células não Pequenas/genética , Linhagem Celular Tumoral , Proliferação de Células/fisiologia , Xenoenxertos , Humanos , Proteína 1 Associada a ECH Semelhante a Kelch/genética , Neoplasias Pulmonares/genética , Camundongos , Mutação , Fator 2 Relacionado a NF-E2/metabolismo , Receptor ErbB-3/metabolismo , Receptor IGF Tipo 1/metabolismo
16.
J Cell Biol ; 217(2): 763-777, 2018 02 05.
Artigo em Inglês | MEDLINE | ID: mdl-29229751

RESUMO

Epithelial-to-mesenchymal transition is implicated in metastasis, where carcinoma cells lose sessile epithelial traits and acquire mesenchymal migratory potential. The mesenchymal state is also associated with cancer stem cells and resistance to chemotherapy. It might therefore be therapeutically beneficial to promote epithelial identity in cancer. Because large-scale cell identity shifts are often orchestrated on an epigenetic level, we screened for candidate epigenetic factors and identified the histone methyltransferase SUV420H2 (KMT5C) as favoring the mesenchymal identity in pancreatic cancer cell lines. Through its repressive mark H4K20me3, SUV420H2 silences several key drivers of the epithelial state. Its knockdown elicited mesenchymal-to-epithelial transition on a molecular and functional level, and cells displayed decreased stemness and increased drug sensitivity. An analysis of human pancreatic cancer biopsies was concordant with these findings, because high levels of SUV420H2 correlated with a loss of epithelial characteristics in progressively invasive cancer. Together, these data indicate that SUV420H2 is an upstream epigenetic regulator of epithelial/mesenchymal state control.


Assuntos
Transição Epitelial-Mesenquimal , Histona-Lisina N-Metiltransferase/metabolismo , Neoplasias Pancreáticas/metabolismo , Linhagem Celular Tumoral , Epigênese Genética/genética , Regulação Neoplásica da Expressão Gênica/genética , Histona-Lisina N-Metiltransferase/genética , Humanos , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia
17.
PLoS One ; 13(6): e0199264, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29912950

RESUMO

Mutant KRAS represents one of the most frequently observed oncogenes in NSCLC, yet no therapies are approved for tumors that express activated KRAS variants. While there is strong rationale for the use of MEK inhibitors to treat tumors with activated RAS/MAPK signaling, these have proven ineffective clinically. We therefore implemented a CRISPR screening approach to identify novel agents to sensitize KRAS mutant NSCLC cells to MEK inhibitor treatment. This approach identified multiple components of the canonical RAS/MAPK pathway consistent with previous studies. In addition, we identified MAPK7 as a novel, strong hit and validated this finding using multiple orthogonal approaches including knockdown and pharmacological inhibition. We show that MAPK7 inhibition attenuates the re-activation of MAPK signaling occurring following long-term MEK inhibition, thereby illustrating that MAPK7 mediates pathway reactivation in the face of MEK inhibition. Finally, genetic knockdown of MAPK7 combined with the MEK inhibitor cobimetinib in a mutant KRAS NSCLC xenograft model to mediate improved tumor growth inhibition. These data highlight that MAPK7 represents a promising target for combination treatment with MEK inhibition in KRAS mutant NSCLC.


Assuntos
Biomarcadores Tumorais/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Proteína Quinase 7 Ativada por Mitógeno/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Células A549 , Animais , Carcinoma Pulmonar de Células não Pequenas/patologia , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/genética , Humanos , Sistema de Sinalização das MAP Quinases/genética , Camundongos , Ensaios Antitumorais Modelo de Xenoenxerto
18.
Cancer Cell ; 34(4): 611-625.e7, 2018 10 08.
Artigo em Inglês | MEDLINE | ID: mdl-30300582

RESUMO

Targeting KRAS mutant tumors through inhibition of individual downstream pathways has had limited clinical success. Here we report that RAF inhibitors exhibit little efficacy in KRAS mutant tumors. In combination drug screens, MEK and PI3K inhibitors synergized with pan-RAF inhibitors through an RAS-GTP-dependent mechanism. Broad cell line profiling with RAF/MEK inhibitor combinations revealed synergistic efficacy in KRAS mutant and wild-type tumors, with KRASG13D mutants exhibiting greater synergy versus KRASG12 mutant tumors. Mechanistic studies demonstrate that MEK inhibition induced RAS-GTP levels, RAF dimerization and RAF kinase activity resulting in MEK phosphorylation in synergistic tumor lines regardless of KRAS status. Taken together, our studies uncover a strategy to rewire KRAS mutant tumors to confer sensitivity to RAF kinase inhibition.


Assuntos
Fosfatidilinositol 3-Quinases/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas p21(ras)/efeitos dos fármacos , Linhagem Celular Tumoral , Guanosina Trifosfato/metabolismo , Humanos , Mutação/efeitos dos fármacos , Mutação/genética , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas B-raf/efeitos dos fármacos , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteínas ras/efeitos dos fármacos , Proteínas ras/genética
19.
NPJ Precis Oncol ; 2(1): 7, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29872725

RESUMO

KRAS- and BRAF-mutant tumors are often dependent on MAPK signaling for proliferation and survival and thus sensitive to MAPK pathway inhibitors. However, clinical studies have shown that MEK inhibitors are not uniformly effective in these cancers indicating that mutational status of these oncogenes does not accurately capture MAPK pathway activity. A number of transcripts are regulated by this pathway and are recurrently identified in genome-based MAPK transcriptional signatures. To test whether the transcriptional output of only 10 of these targets could quantify MAPK pathway activity with potential predictive or prognostic clinical utility, we created a MAPK Pathway Activity Score (MPAS) derived from aggregated gene expression. In vitro, MPAS predicted sensitivity to MAPK inhibitors in multiple cell lines, comparable to or better than larger genome-based statistical models. Bridging in vitro studies and clinical samples, median MPAS from a given tumor type correlated with cobimetinib (MEK inhibitor) sensitivity of cancer cell lines originating from the same tissue type. Retrospective analyses of clinical datasets showed that MPAS was associated with the sensitivity of melanomas to vemurafenib (HR: 0.596) and negatively prognostic of overall or progression-free survival in both adjuvant and metastatic CRC (HR: 1.5 and 1.4), adrenal cancer (HR: 1.7), and HER2+ breast cancer (HR: 1.6). MPAS thus demonstrates potential clinical utility that warrants further exploration.

20.
Sci Signal ; 11(547)2018 09 11.
Artigo em Inglês | MEDLINE | ID: mdl-30206136

RESUMO

The Hippo signaling pathway regulates organ size and plays critical roles in maintaining tissue growth, homeostasis, and regeneration. Dysregulated in a wide spectrum of cancers, in mammals, this pathway is regulated by two key effectors, YAP and TAZ, that may functionally overlap. We found that TAZ promoted liver inflammation and tumor development. The expression of TAZ, but not YAP, in human liver tumors positively correlated with the expression of proinflammatory cytokines. Hyperactivated TAZ induced substantial myeloid cell infiltration into the liver and the secretion of proinflammatory cytokines through a TEAD-dependent mechanism. Furthermore, tumors with hyperactivated YAP and TAZ had distinct transcriptional signatures, which included the increased expression of inflammatory cytokines in TAZ-driven tumors. Our study elucidated a previously uncharacterized link between TAZ activity and inflammatory responses that influence tumor development in the liver.


Assuntos
Proteínas de Ligação a DNA/genética , Inflamação/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Neoplasias Hepáticas/genética , Fígado/metabolismo , Proteínas Nucleares/genética , Proteínas Serina-Treonina Quinases/genética , Fatores de Transcrição/genética , Animais , Proteínas de Ciclo Celular , Citocinas/genética , Citocinas/metabolismo , Proteínas de Ligação a DNA/metabolismo , Perfilação da Expressão Gênica/métodos , Via de Sinalização Hippo , Humanos , Inflamação/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Fígado/patologia , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Camundongos Endogâmicos C57BL , Mutação , Proteínas Nucleares/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais/genética , Fatores de Transcrição de Domínio TEA , Transativadores , Fatores de Transcrição/metabolismo , Proteínas com Motivo de Ligação a PDZ com Coativador Transcricional , Transplante Heterólogo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA