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1.
J Transl Med ; 22(1): 190, 2024 02 21.
Artigo em Inglês | MEDLINE | ID: mdl-38383458

RESUMO

BACKGROUND: Predictive biomarkers of immune checkpoint inhibitor (ICI) efficacy are currently lacking for non-small cell lung cancer (NSCLC). Here, we describe the results from the Anti-PD-1 Response Prediction DREAM Challenge, a crowdsourced initiative that enabled the assessment of predictive models by using data from two randomized controlled clinical trials (RCTs) of ICIs in first-line metastatic NSCLC. METHODS: Participants developed and trained models using public resources. These were evaluated with data from the CheckMate 026 trial (NCT02041533), according to the model-to-data paradigm to maintain patient confidentiality. The generalizability of the models with the best predictive performance was assessed using data from the CheckMate 227 trial (NCT02477826). Both trials were phase III RCTs with a chemotherapy control arm, which supported the differentiation between predictive and prognostic models. Isolated model containers were evaluated using a bespoke strategy that considered the challenges of handling transcriptome data from clinical trials. RESULTS: A total of 59 teams participated, with 417 models submitted. Multiple predictive models, as opposed to a prognostic model, were generated for predicting overall survival, progression-free survival, and progressive disease status with ICIs. Variables within the models submitted by participants included tumor mutational burden (TMB), programmed death ligand 1 (PD-L1) expression, and gene-expression-based signatures. The best-performing models showed improved predictive power over reference variables, including TMB or PD-L1. CONCLUSIONS: This DREAM Challenge is the first successful attempt to use protected phase III clinical data for a crowdsourced effort towards generating predictive models for ICI clinical outcomes and could serve as a blueprint for similar efforts in other tumor types and disease states, setting a benchmark for future studies aiming to identify biomarkers predictive of ICI efficacy. TRIAL REGISTRATION: CheckMate 026; NCT02041533, registered January 22, 2014. CheckMate 227; NCT02477826, registered June 23, 2015.


Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Humanos , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Inibidores de Checkpoint Imunológico/uso terapêutico , Neoplasias Pulmonares/patologia , Antígeno B7-H1 , Biomarcadores Tumorais
2.
Nat Genet ; 53(7): 942-948, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34183854

RESUMO

The UK Biobank Exome Sequencing Consortium (UKB-ESC) is a private-public partnership between the UK Biobank (UKB) and eight biopharmaceutical companies that will complete the sequencing of exomes for all ~500,000 UKB participants. Here, we describe the early results from ~200,000 UKB participants and the features of this project that enabled its success. The biopharmaceutical industry has increasingly used human genetics to improve success in drug discovery. Recognizing the need for large-scale human genetics data, as well as the unique value of the data access and contribution terms of the UKB, the UKB-ESC was formed. As a result, exome data from 200,643 UKB enrollees are now available. These data include ~10 million exonic variants-a rich resource of rare coding variation that is particularly valuable for drug discovery. The UKB-ESC precompetitive collaboration has further strengthened academic and industry ties and has provided teams with an opportunity to interact with and learn from the wider research community.


Assuntos
Bancos de Espécimes Biológicos , Descoberta de Drogas , Sequenciamento do Exoma , Genética Humana , Pesquisa , Descoberta de Drogas/métodos , Genômica/métodos , Humanos , Reino Unido
3.
Am J Hum Genet ; 108(7): 1350-1355, 2021 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-34115965

RESUMO

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) causes coronavirus disease 2019 (COVID-19), a respiratory illness that can result in hospitalization or death. We used exome sequence data to investigate associations between rare genetic variants and seven COVID-19 outcomes in 586,157 individuals, including 20,952 with COVID-19. After accounting for multiple testing, we did not identify any clear associations with rare variants either exome wide or when specifically focusing on (1) 13 interferon pathway genes in which rare deleterious variants have been reported in individuals with severe COVID-19, (2) 281 genes located in susceptibility loci identified by the COVID-19 Host Genetics Initiative, or (3) 32 additional genes of immunologic relevance and/or therapeutic potential. Our analyses indicate there are no significant associations with rare protein-coding variants with detectable effect sizes at our current sample sizes. Analyses will be updated as additional data become available, and results are publicly available through the Regeneron Genetics Center COVID-19 Results Browser.


Assuntos
COVID-19/diagnóstico , COVID-19/genética , Sequenciamento do Exoma , Exoma/genética , Predisposição Genética para Doença , Hospitalização/estatística & dados numéricos , COVID-19/imunologia , COVID-19/terapia , Feminino , Humanos , Interferons/genética , Masculino , Prognóstico , SARS-CoV-2 , Tamanho da Amostra
5.
Biotechniques ; 69(6): 420-426, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33103912

RESUMO

Although next-generation sequencing assays are routinely carried out using samples from cancer trials, the sequencing data are not always of the required quality. There is a need to evaluate the performance of tissue collection sites and provide feedback about the quality of next-generation sequencing data. This study used a modeling approach based on whole exome sequencing quality control (QC) metrics to evaluate the relative performance of sites participating in the Bristol Myers Squibb Immuno-Oncology clinical trials sample collection. We identified several events for the sample swap. Overall, most sites performed well and few showed poor performance. These findings can increase awareness of sample failure and improve the quality of samples.


Assuntos
Sequenciamento do Exoma , Modelos Teóricos , Manejo de Espécimes , Técnicas de Laboratório Clínico , Humanos , Controle de Qualidade , Sequenciamento do Exoma/normas
6.
ESMO Open ; 5(2)2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32312757

RESUMO

INTRODUCTION: Somatic mutations in STK11 and KEAP1, frequently comutated in non-squamous non-small cell lung cancer (NSQ NSCLC), have been associated with poor response to immune checkpoint blockade (ICB). However, previous reports lack non-ICB controls needed to properly ascertain the predictive nature of those biomarkers. The objective of this study was to evaluate the predictive versus prognostic effect of STK11 or KEAP1 mutations in NSQ NSCLC. METHODS: Patients diagnosed with stage IIIB, IIIC, IVA or IVB NSQ NSCLC from a real-world data cohort from the Flatiron Health Network linked with genetic testing from Foundation Medicine were retrospectively assessed. Real-world, progression-free survival (rwPFS) and overall survival (OS) were calculated from time of initiation of first-line treatment. RESULTS: We analysed clinical and mutational data for 2276 patients including patients treated with anti-programmed death-1 (PD-1)/anti-programmed death ligand 1 (PD-L1) inhibitors at first line (n=574). Mutations in STK11 or KEAP1 were associated with poor outcomes across multiple therapeutic classes and were not specifically associated with poor outcomes in ICB cohorts. There was no observable interaction between STK11 mutations and anti-PD-1/anti-PD-L1 treatment on rwPFS (HR, 1.05; 95% CI 0.76 to 1.44; p=0.785) or OS (HR, 1.13; 95% CI 0.76 to 1.67; p=0.540). Similarly, there was no observable interaction between KEAP1 mutations and treatment on rwPFS (HR, 0.93; 95% CI 0.67 to 1.28; p=0.653) or OS (HR, 0.98; 95% CI 0.66 to 1.45; p=0.913). CONCLUSION: Our results show that STK11-KEAP1 mutations are prognostic, not predictive, biomarkers for anti-PD-1/anti-PD-L1 therapy.


Assuntos
Adenocarcinoma/genética , Proteína 1 Associada a ECH Semelhante a Kelch/genética , Neoplasias Pulmonares/genética , Proteínas Serina-Treonina Quinases/genética , Quinases Proteína-Quinases Ativadas por AMP , Adenocarcinoma/mortalidade , Adenocarcinoma/patologia , Biomarcadores Tumorais , Estudos de Coortes , Feminino , Humanos , Neoplasias Pulmonares/mortalidade , Neoplasias Pulmonares/patologia , Masculino , Mutação , Prognóstico , Análise de Sobrevida
7.
Mol Diagn Ther ; 23(4): 507-520, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31250328

RESUMO

INTRODUCTION: Tumor mutational burden (TMB) has emerged as a clinically relevant biomarker that may be associated with immune checkpoint inhibitor efficacy. Standardization of TMB measurement is essential for implementing diagnostic tools to guide treatment. OBJECTIVE: Here we describe the in-depth evaluation of bioinformatic TMB analysis by whole exome sequencing (WES) in formalin-fixed, paraffin-embedded samples from a phase III clinical trial. METHODS: In the CheckMate 026 clinical trial, TMB was retrospectively assessed in 312 patients with non-small-cell lung cancer (58% of the intent-to-treat population) who received first-line nivolumab treatment or standard-of-care chemotherapy. We examined the sensitivity of TMB assessment to bioinformatic filtering methods and assessed concordance between TMB data derived by WES and the FoundationOne® CDx assay. RESULTS: TMB scores comprising synonymous, indel, frameshift, and nonsense mutations (all mutations) were 3.1-fold higher than data including missense mutations only, but values were highly correlated (Spearman's r = 0.99). Scores from CheckMate 026 samples including missense mutations only were similar to those generated from data in The Cancer Genome Atlas, but those including all mutations were generally higher. Using databases for germline subtraction (instead of matched controls) showed a trend for race-dependent increases in TMB scores. WES and FoundationOne CDx outputs were highly correlated (Spearman's r = 0.90). CONCLUSIONS: Parameter variation can impact TMB calculations, highlighting the need for standardization. Encouragingly, differences between assays could be accounted for by empirical calibration, suggesting that reliable TMB assessment across assays, platforms, and centers is achievable.


Assuntos
Biomarcadores Tumorais , Carcinoma Pulmonar de Células não Pequenas/genética , Biologia Computacional , Neoplasias Pulmonares/genética , Mutação , Carcinoma Pulmonar de Células não Pequenas/mortalidade , Carcinoma Pulmonar de Células não Pequenas/patologia , Biologia Computacional/métodos , Estudos de Associação Genética , Predisposição Genética para Doença , Humanos , Neoplasias Pulmonares/patologia , Prognóstico , Reprodutibilidade dos Testes , Sequenciamento do Exoma , Fluxo de Trabalho
8.
J Clin Oncol ; 37(12): 992-1000, 2019 04 20.
Artigo em Inglês | MEDLINE | ID: mdl-30785829

RESUMO

PURPOSE: CheckMate 568 is an open-label phase II trial that evaluated the efficacy and safety of nivolumab plus low-dose ipilimumab as first-line treatment of advanced/metastatic non-small-cell lung cancer (NSCLC). We assessed the association of efficacy with programmed death ligand 1 (PD-L1) expression and tumor mutational burden (TMB). PATIENTS AND METHODS: Two hundred eighty-eight patients with previously untreated, recurrent stage IIIB/IV NSCLC received nivolumab 3 mg/kg every 2 weeks plus ipilimumab 1 mg/kg every 6 weeks. The primary end point was objective response rate (ORR) in patients with 1% or more and less than 1% tumor PD-L1 expression. Efficacy on the basis of TMB (FoundationOne CDx assay) was a secondary end point. RESULTS: Of treated patients with tumor available for testing, 252 patients (88%) of 288 were evaluable for PD-L1 expression and 98 patients (82%) of 120 for TMB. ORR was 30% overall and 41% and 15% in patients with 1% or greater and less than 1% tumor PD-L1 expression, respectively. ORR increased with higher TMB, plateauing at 10 or more mutations/megabase (mut/Mb). Regardless of PD-L1 expression, ORRs were higher in patients with TMB of 10 or more mut/Mb (n = 48: PD-L1, ≥ 1%, 48%; PD-L1, < 1%, 47%) versus TMB of fewer than 10 mut/Mb (n = 50: PD-L1, ≥ 1%, 18%; PD-L1, < 1%, 5%), and progression-free survival was longer in patients with TMB of 10 or more mut/Mb versus TMB of fewer than 10 mut/Mb (median, 7.1 v 2.6 months). Grade 3 to 4 treatment-related adverse events occurred in 29% of patients. CONCLUSION: Nivolumab plus low-dose ipilimumab was effective and tolerable as a first-line treatment of advanced/metastatic NSCLC. TMB of 10 or more mut/Mb was associated with improved response and prolonged progression-free survival in both tumor PD-L1 expression 1% or greater and less than 1% subgroups and was thus identified as a potentially relevant cutoff in the assessment of TMB as a biomarker for first-line nivolumab plus ipilimumab.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Antígeno B7-H1/biossíntese , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológico , Mutação , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígeno B7-H1/imunologia , Biomarcadores Tumorais/biossíntese , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/imunologia , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/imunologia , Feminino , Humanos , Ipilimumab/administração & dosagem , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/imunologia , Masculino , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/tratamento farmacológico , Estadiamento de Neoplasias , Nivolumabe/administração & dosagem , Resultado do Tratamento
10.
Cancer Discov ; 8(7): 822-835, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-29773717

RESUMO

KRAS is the most common oncogenic driver in lung adenocarcinoma (LUAC). We previously reported that STK11/LKB1 (KL) or TP53 (KP) comutations define distinct subgroups of KRAS-mutant LUAC. Here, we examine the efficacy of PD-1 inhibitors in these subgroups. Objective response rates to PD-1 blockade differed significantly among KL (7.4%), KP (35.7%), and K-only (28.6%) subgroups (P < 0.001) in the Stand Up To Cancer (SU2C) cohort (174 patients) with KRAS-mutant LUAC and in patients treated with nivolumab in the CheckMate-057 phase III trial (0% vs. 57.1% vs. 18.2%; P = 0.047). In the SU2C cohort, KL LUAC exhibited shorter progression-free (P < 0.001) and overall (P = 0.0015) survival compared with KRASMUT;STK11/LKB1WT LUAC. Among 924 LUACs, STK11/LKB1 alterations were the only marker significantly associated with PD-L1 negativity in TMBIntermediate/High LUAC. The impact of STK11/LKB1 alterations on clinical outcomes with PD-1/PD-L1 inhibitors extended to PD-L1-positive non-small cell lung cancer. In Kras-mutant murine LUAC models, Stk11/Lkb1 loss promoted PD-1/PD-L1 inhibitor resistance, suggesting a causal role. Our results identify STK11/LKB1 alterations as a major driver of primary resistance to PD-1 blockade in KRAS-mutant LUAC.Significance: This work identifies STK11/LKB1 alterations as the most prevalent genomic driver of primary resistance to PD-1 axis inhibitors in KRAS-mutant lung adenocarcinoma. Genomic profiling may enhance the predictive utility of PD-L1 expression and tumor mutation burden and facilitate establishment of personalized combination immunotherapy approaches for genomically defined LUAC subsets. Cancer Discov; 8(7); 822-35. ©2018 AACR.See related commentary by Etxeberria et al., p. 794This article is highlighted in the In This Issue feature, p. 781.


Assuntos
Adenocarcinoma de Pulmão/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/genética , Neoplasias Pulmonares/tratamento farmacológico , Mutação , Nivolumabe/uso terapêutico , Proteínas Serina-Treonina Quinases/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Quinases Proteína-Quinases Ativadas por AMP , Adenocarcinoma de Pulmão/genética , Adenocarcinoma de Pulmão/metabolismo , Adenocarcinoma de Pulmão/terapia , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Antineoplásicos Imunológicos/farmacologia , Antineoplásicos Imunológicos/uso terapêutico , Modelos Animais de Doenças , Humanos , Imunoterapia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/terapia , Masculino , Camundongos , Pessoa de Meia-Idade , Nivolumabe/farmacologia , Prognóstico , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Intervalo Livre de Progressão
11.
Cancer Cell ; 33(5): 853-861.e4, 2018 05 14.
Artigo em Inglês | MEDLINE | ID: mdl-29731394

RESUMO

Durable responses and encouraging survival have been demonstrated with immune checkpoint inhibitors in small-cell lung cancer (SCLC), but predictive markers are unknown. We used whole exome sequencing to evaluate the impact of tumor mutational burden on efficacy of nivolumab monotherapy or combined with ipilimumab in patients with SCLC from the nonrandomized or randomized cohorts of CheckMate 032. Patients received nivolumab (3 mg/kg every 2 weeks) or nivolumab plus ipilimumab (1 mg/kg plus 3 mg/kg every 3 weeks for four cycles, followed by nivolumab 3 mg/kg every 2 weeks). Efficacy of nivolumab ± ipilimumab was enhanced in patients with high tumor mutational burden. Nivolumab plus ipilimumab appeared to provide a greater clinical benefit than nivolumab monotherapy in the high tumor mutational burden tertile.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Ipilimumab/administração & dosagem , Neoplasias Pulmonares/tratamento farmacológico , Mutação , Nivolumabe/administração & dosagem , Carcinoma de Pequenas Células do Pulmão/tratamento farmacológico , Adulto , Idoso , Idoso de 80 Anos ou mais , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Relação Dose-Resposta a Droga , Esquema de Medicação , Feminino , Humanos , Ipilimumab/farmacologia , Neoplasias Pulmonares/genética , Masculino , Pessoa de Meia-Idade , Nivolumabe/farmacologia , Carcinoma de Pequenas Células do Pulmão/genética , Resultado do Tratamento , Carga Tumoral/efeitos dos fármacos , Sequenciamento do Exoma
12.
N Engl J Med ; 378(22): 2093-2104, 2018 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-29658845

RESUMO

BACKGROUND: Nivolumab plus ipilimumab showed promising efficacy for the treatment of non-small-cell lung cancer (NSCLC) in a phase 1 trial, and tumor mutational burden has emerged as a potential biomarker of benefit. In this part of an open-label, multipart, phase 3 trial, we examined progression-free survival with nivolumab plus ipilimumab versus chemotherapy among patients with a high tumor mutational burden (≥10 mutations per megabase). METHODS: We enrolled patients with stage IV or recurrent NSCLC that was not previously treated with chemotherapy. Those with a level of tumor programmed death ligand 1 (PD-L1) expression of at least 1% were randomly assigned, in a 1:1:1 ratio, to receive nivolumab plus ipilimumab, nivolumab monotherapy, or chemotherapy; those with a tumor PD-L1 expression level of less than 1% were randomly assigned, in a 1:1:1 ratio, to receive nivolumab plus ipilimumab, nivolumab plus chemotherapy, or chemotherapy. Tumor mutational burden was determined by the FoundationOne CDx assay. RESULTS: Progression-free survival among patients with a high tumor mutational burden was significantly longer with nivolumab plus ipilimumab than with chemotherapy. The 1-year progression-free survival rate was 42.6% with nivolumab plus ipilimumab versus 13.2% with chemotherapy, and the median progression-free survival was 7.2 months (95% confidence interval [CI], 5.5 to 13.2) versus 5.5 months (95% CI, 4.4 to 5.8) (hazard ratio for disease progression or death, 0.58; 97.5% CI, 0.41 to 0.81; P<0.001). The objective response rate was 45.3% with nivolumab plus ipilimumab and 26.9% with chemotherapy. The benefit of nivolumab plus ipilimumab over chemotherapy was broadly consistent within subgroups, including patients with a PD-L1 expression level of at least 1% and those with a level of less than 1%. The rate of grade 3 or 4 treatment-related adverse events was 31.2% with nivolumab plus ipilimumab and 36.1% with chemotherapy. ical; CheckMate 227 ClinicalTrials.gov number, NCT02477826 .). CONCLUSIONS: Progression-free survival was significantly longer with first-line nivolumab plus ipilimumab than with chemotherapy among patients with NSCLC and a high tumor mutational burden, irrespective of PD-L1 expression level. The results validate the benefit of nivolumab plus ipilimumab in NSCLC and the role of tumor mutational burden as a biomarker for patient selection. (Funded by Bristol-Myers Squibb and Ono Pharmaceut


Assuntos
Anticorpos Monoclonais/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Ipilimumab/administração & dosagem , Neoplasias Pulmonares/tratamento farmacológico , Mutação , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticorpos Monoclonais/efeitos adversos , Antineoplásicos Imunológicos/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Antígeno B7-H1/genética , Antígeno B7-H1/metabolismo , Biomarcadores Tumorais , Carcinoma Pulmonar de Células não Pequenas/genética , Intervalo Livre de Doença , Feminino , Humanos , Ipilimumab/efeitos adversos , Estimativa de Kaplan-Meier , Neoplasias Pulmonares/genética , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Nivolumabe
13.
PLoS One ; 12(7): e0179726, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28749946

RESUMO

Therapeutic options for the treatment of an increasing variety of cancers have been expanded by the introduction of a new class of drugs, commonly referred to as checkpoint blocking agents, that target the host immune system to positively modulate anti-tumor immune response. Although efficacy of these agents has been linked to a pre-existing level of tumor immune infiltrate, it remains unclear why some patients exhibit deep and durable responses to these agents while others do not benefit. To examine the influence of tumor genetics on tumor immune state, we interrogated the relationship between somatic mutation and copy number alteration with infiltration levels of 7 immune cell types across 40 tumor cohorts in The Cancer Genome Atlas. Levels of cytotoxic T, regulatory T, total T, natural killer, and B cells, as well as monocytes and M2 macrophages, were estimated using a novel set of transcriptional signatures that were designed to resist interference from the cellular heterogeneity of tumors. Tumor mutational load and estimates of tumor purity were included in our association models to adjust for biases in multi-modal genomic data. Copy number alterations, mutations summarized at the gene level, and position-specific mutations were evaluated for association with tumor immune infiltration. We observed a strong relationship between copy number loss of a large region of chromosome 9p and decreased lymphocyte estimates in melanoma, pancreatic, and head/neck cancers. Mutations in the oncogenes PIK3CA, FGFR3, and RAS/RAF family members, as well as the tumor suppressor TP53, were linked to changes in immune infiltration, usually in restricted tumor types. Associations of specific WNT/beta-catenin pathway genetic changes with immune state were limited, but we noted a link between 9p loss and the expression of the WNT receptor FZD3, suggesting that there are interactions between 9p alteration and WNT pathways. Finally, two different cell death regulators, CASP8 and DIDO1, were often mutated in head/neck tumors that had higher lymphocyte infiltrates. In summary, our study supports the relevance of tumor genetics to questions of efficacy and resistance in checkpoint blockade therapies. It also highlights the need to assess genome-wide influences during exploration of any specific tumor pathway hypothesized to be relevant to therapeutic response. Some of the observed genetic links to immune state, like 9p loss, may influence response to cancer immune therapies. Others, like mutations in cell death pathways, may help guide combination therapeutic approaches.


Assuntos
Estudo de Associação Genômica Ampla , Neoplasias/genética , Neoplasias/imunologia , Biomarcadores Tumorais/genética , Linfócitos T CD8-Positivos/imunologia , Cromossomos Humanos Par 9/genética , Dosagem de Genes , Neoplasias de Cabeça e Pescoço/genética , Humanos , Mutação/genética , Proteínas de Neoplasias/genética , Análise de Sequência de RNA , Proteína Supressora de Tumor p53/genética
14.
Am J Hum Genet ; 100(5): 737-750, 2017 May 04.
Artigo em Inglês | MEDLINE | ID: mdl-28457472

RESUMO

Keratolytic winter erythema (KWE) is a rare autosomal-dominant skin disorder characterized by recurrent episodes of palmoplantar erythema and epidermal peeling. KWE was previously mapped to 8p23.1-p22 (KWE critical region) in South African families. Using targeted resequencing of the KWE critical region in five South African families and SNP array and whole-genome sequencing in two Norwegian families, we identified two overlapping tandem duplications of 7.67 kb (South Africans) and 15.93 kb (Norwegians). The duplications segregated with the disease and were located upstream of CTSB, a gene encoding cathepsin B, a cysteine protease involved in keratinocyte homeostasis. Included in the 2.62 kb overlapping region of these duplications is an enhancer element that is active in epidermal keratinocytes. The activity of this enhancer correlated with CTSB expression in normal differentiating keratinocytes and other cell lines, but not with FDFT1 or NEIL2 expression. Gene expression (qPCR) analysis and immunohistochemistry of the palmar epidermis demonstrated significantly increased expression of CTSB, as well as stronger staining of cathepsin B in the stratum granulosum of affected individuals than in that of control individuals. Analysis of higher-order chromatin structure data and RNA polymerase II ChIA-PET data from MCF-7 cells did not suggest remote effects of the enhancer. In conclusion, KWE in South African and Norwegian families is caused by tandem duplications in a non-coding genomic region containing an active enhancer element for CTSB, resulting in upregulation of this gene in affected individuals.


Assuntos
Catepsina B/metabolismo , Elementos Facilitadores Genéticos , Eritema/genética , Duplicação Gênica , Regulação da Expressão Gênica , Ceratose/genética , Dermatopatias Genéticas/genética , Estudos de Casos e Controles , Catepsina B/genética , Mapeamento Cromossômico , Cromossomos Humanos Par 8/genética , Variações do Número de Cópias de DNA , DNA Glicosilases/genética , DNA Glicosilases/metabolismo , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/genética , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/metabolismo , Epiderme/metabolismo , Epigenômica , Eritema/epidemiologia , Feminino , Marcadores Genéticos , Humanos , Queratinócitos/metabolismo , Ceratose/epidemiologia , Células MCF-7 , Masculino , Noruega/epidemiologia , Linhagem , Dermatopatias Genéticas/epidemiologia , África do Sul/epidemiologia
15.
Nat Commun ; 6: 8666, 2015 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-26487268

RESUMO

Ciliopathies are a large group of clinically and genetically heterogeneous disorders caused by defects in primary cilia. Here we identified mutations in TRAF3IP1 (TNF Receptor-Associated Factor Interacting Protein 1) in eight patients from five families with nephronophthisis (NPH) and retinal degeneration, two of the most common manifestations of ciliopathies. TRAF3IP1 encodes IFT54, a subunit of the IFT-B complex required for ciliogenesis. The identified mutations result in mild ciliary defects in patients but also reveal an unexpected role of IFT54 as a negative regulator of microtubule stability via MAP4 (microtubule-associated protein 4). Microtubule defects are associated with altered epithelialization/polarity in renal cells and with pronephric cysts and microphthalmia in zebrafish embryos. Our findings highlight the regulation of cytoplasmic microtubule dynamics as a role of the IFT54 protein beyond the cilium, contributing to the development of NPH-related ciliopathies.


Assuntos
Proteínas de Transporte/genética , Doenças Renais Císticas/genética , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Microtúbulos/metabolismo , Mutação , Degeneração Retiniana/genética , Proteínas de Peixe-Zebra/genética , Animais , Western Blotting , Proteínas de Transporte/metabolismo , Polaridade Celular/genética , Dicroísmo Circular , Embrião não Mamífero , Feminino , Imunofluorescência , Técnicas de Inativação de Genes , Células HEK293 , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Imunoprecipitação , Doenças Renais Císticas/metabolismo , Masculino , Microftalmia/genética , Linhagem , Degeneração Retiniana/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Peixe-Zebra , Proteínas de Peixe-Zebra/metabolismo
16.
Biomark Res ; 2: 17, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25285214

RESUMO

BACKGROUND: Exact sample annotation in expression microarray datasets is essential for any type of pharmacogenomics research. RESULTS: Candidate markers were explored through the application of Hartigans' dip test statistics to a publically available human whole genome microarray dataset. The marker performance was tested on 188 serial samples from 53 donors and of variable tissue origin from five public microarray datasets. A qualified transcript marker panel consisting of three probe sets for human leukocyte antigens HLA-DQA1 (2 probe sets) and HLA-DRB4 identified sample donor identifier inconsistencies in six of the 188 test samples. About 3% of the test samples require root-cause analysis due to unresolvable inaccuracies. CONCLUSIONS: The transcript marker panel consisting of HLA-DQA1 and HLA-DRB4 represents a robust, tissue-independent composite marker to assist control donor annotation concordance at the transcript level. Allele-selectivity of HLA genes renders them good candidates for "fingerprinting" with donor specific expression pattern.

17.
Arthritis Rheumatol ; 66(12): 3486-95, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25047945

RESUMO

OBJECTIVE: Macrophage activation syndrome (MAS), a life-threatening complication of systemic juvenile idiopathic arthritis (JIA), resembles familial hemophagocytic lymphohistiocytosis (HLH), a constellation of autosomal-recessive immune disorders resulting from deficiency in cytolytic pathway proteins. We undertook this study to test our hypothesis that MAS predisposition in systemic JIA could be attributed to rare gene sequence variants affecting the cytotolytic pathway. METHODS: Whole-exome sequencing was used in 14 patients with systemic JIA and MAS and in their parents to identify protein-altering single-nucleotide polymorphisms/indels in known HLH-associated genes. To discover new candidate genes, the entire whole-exome sequencing data were filtered to identify protein-altering, rare recessive homozygous, compound heterozygous, and de novo variants with the potential to affect the cytolytic pathway. RESULTS: Heterozygous protein-altering rare variants in the known genes (LYST,MUNC13-4, and STXBP2) were found in 5 of 14 patients with systemic JIA and MAS (35.7%). This was in contrast to only 4 variants in 4 of 29 patients with systemic JIA without MAS (13.8%). Homozygosity and compound heterozygosity analysis applied to the entire whole-exome sequencing data in systemic JIA/MAS revealed 3 recessive pairs in 3 genes and compound heterozygotes in 73 genes. We also identified 20 heterozygous rare protein-altering variants that occurred in at least 2 patients. Many of the identified genes encoded proteins with a role in actin and microtubule reorganization and vesicle-mediated transport. "Cellular assembly and organization" was the top cellular function category based on Ingenuity Pathways Analysis (P < 3.10 × 10(-5) ). CONCLUSION: Whole-exome sequencing performed in patients with systemic JIA and MAS identified rare protein-altering variants in known HLH-associated genes as well as in new candidate genes.


Assuntos
Artrite Juvenil/genética , Linfo-Histiocitose Hemofagocítica/genética , Síndrome de Ativação Macrofágica/genética , Adolescente , Artrite Juvenil/complicações , Criança , Pré-Escolar , Exoma , Feminino , Predisposição Genética para Doença , Genótipo , Humanos , Lactente , Síndrome de Ativação Macrofágica/complicações , Masculino , Mutação , Pais , Polimorfismo de Nucleotídeo Único , Análise de Sequência de DNA
18.
J Am Soc Nephrol ; 24(8): 1216-22, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23687361

RESUMO

LMX1B encodes a homeodomain-containing transcription factor that is essential during development. Mutations in LMX1B cause nail-patella syndrome, characterized by dysplasia of the patellae, nails, and elbows and FSGS with specific ultrastructural lesions of the glomerular basement membrane (GBM). By linkage analysis and exome sequencing, we unexpectedly identified an LMX1B mutation segregating with disease in a pedigree of five patients with autosomal dominant FSGS but without either extrarenal features or ultrastructural abnormalities of the GBM suggestive of nail-patella-like renal disease. Subsequently, we screened 73 additional unrelated families with FSGS and found mutations involving the same amino acid (R246) in 2 families. An LMX1B in silico homology model suggested that the mutated residue plays an important role in strengthening the interaction between the LMX1B homeodomain and DNA; both identified mutations would be expected to diminish such interactions. In summary, these results suggest that isolated FSGS could result from mutations in genes that are also involved in syndromic forms of FSGS. This highlights the need to include these genes in all diagnostic approaches to FSGS that involve next-generation sequencing.


Assuntos
Glomerulosclerose Segmentar e Focal/genética , Proteínas com Homeodomínio LIM/genética , Síndrome da Unha-Patela/genética , Fatores de Transcrição/genética , Adolescente , Adulto , Criança , Feminino , Genes Dominantes , Humanos , Masculino , Pessoa de Meia-Idade , Mutação , Linhagem , Análise de Sequência de DNA , Adulto Jovem
19.
Bioinformatics ; 29(8): 1083-5, 2013 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-23428642

RESUMO

SUMMARY: For heterogeneous tissues, measurements of gene expression through mRNA-Seq data are confounded by relative proportions of cell types involved. In this note, we introduce an efficient pipeline: DeconRNASeq, an R package for deconvolution of heterogeneous tissues based on mRNA-Seq data. It adopts a globally optimized non-negative decomposition algorithm through quadratic programming for estimating the mixing proportions of distinctive tissue types in next-generation sequencing data. We demonstrated the feasibility and validity of DeconRNASeq across a range of mixing levels and sources using mRNA-Seq data mixed in silico at known concentrations. We validated our computational approach for various benchmark data, with high correlation between our predicted cell proportions and the real fractions of tissues. Our study provides a rigorous, quantitative and high-resolution tool as a prerequisite to use mRNA-Seq data. The modularity of package design allows an easy deployment of custom analytical pipelines for data from other high-throughput platforms. AVAILABILITY: DeconRNASeq is written in R, and is freely available at http://bioconductor.org/packages. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.


Assuntos
Perfilação da Expressão Gênica/métodos , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Análise de Sequência de RNA/métodos , Software , Algoritmos , Simulação por Computador , Interpretação Estatística de Dados , Modelos Lineares , RNA Mensageiro/metabolismo
20.
Mol Imaging ; 12(1): 2-7, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23348786

RESUMO

Signaling pathways are the fundamental grammar of cellular communication, yet few frameworks are available to analyze molecular imaging probes in the context of signaling pathways. Such a framework would aid in the design and selection of imaging probes for measuring specific signaling pathways and, vice versa, help illuminate which pathways are being assayed by a given probe. RAMP (Researching imaging Agents through Molecular Pathways) is a bioinformatics framework for connecting signaling pathways and imaging probes using a controlled vocabulary of the imaging targets. RAMP contains signaling pathway data from MetaCore, the Kyoto Encyclopedia of Genes and Genomes, and the Gene Ontology project; imaging probe data from the Molecular Imaging and Contrast Agent Database (MICAD); and tissue protein expression data from The Human Protein Atlas. The RAMP search tool is available at . Examples are presented to demonstrate the utility of RAMP for pathway-based searches of molecular imaging probes.


Assuntos
Biologia Computacional/métodos , Meios de Contraste/química , Meios de Contraste/metabolismo , Imagem Molecular , Transdução de Sinais , Software , Bases de Dados Factuais , Humanos , Internet , Modelos Biológicos , Proteínas/análise , Proteínas/metabolismo
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