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1.
Cell ; 148(1-2): 273-84, 2012 Jan 20.
Artigo em Inglês | MEDLINE | ID: mdl-22265416

RESUMO

Secreted signals, known as morphogens, provide the positional information that organizes gene expression and cellular differentiation in many developing tissues. In the vertebrate neural tube, Sonic Hedgehog (Shh) acts as a morphogen to control the pattern of neuronal subtype specification. Using an in vivo reporter of Shh signaling, mouse genetics, and systems modeling, we show that a spatially and temporally changing gradient of Shh signaling is interpreted by the regulatory logic of a downstream transcriptional network. The design of the network, which links three transcription factors to Shh signaling, is responsible for differential spatial and temporal gene expression. In addition, the network renders cells insensitive to fluctuations in signaling and confers hysteresis--memory of the signal. Our findings reveal that morphogen interpretation is an emergent property of the architecture of a transcriptional network that provides robustness and reliability to tissue patterning.


Assuntos
Redes Reguladoras de Genes , Proteínas Hedgehog/metabolismo , Tubo Neural/metabolismo , Transdução de Sinais , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Proteínas do Olho/genética , Proteínas Hedgehog/genética , Proteína Homeobox Nkx-2.2 , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/metabolismo , Camundongos , Camundongos Transgênicos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Células-Tronco Neurais/metabolismo , Fator de Transcrição 2 de Oligodendrócitos , Fator de Transcrição PAX6 , Fatores de Transcrição Box Pareados/genética , Proteínas Repressoras/genética , Fatores de Transcrição/metabolismo , Proteínas de Peixe-Zebra , Proteína Gli3 com Dedos de Zinco
2.
Development ; 149(10)2022 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-35438131

RESUMO

In many developing and regenerating systems, tissue pattern is established through gradients of informative morphogens, but we know little about how cells interpret these. Using experimental manipulation of early chick embryos, including misexpression of an inducer (VG1 or ACTIVIN) and an inhibitor (BMP4), we test two alternative models for their ability to explain how the site of primitive streak formation is positioned relative to the rest of the embryo. In one model, cells read morphogen concentrations cell-autonomously. In the other, cells sense changes in morphogen status relative to their neighbourhood. We find that only the latter model can account for the experimental results, including some counter-intuitive predictions. This mechanism (which we name the 'neighbourhood watch' model) illuminates the classic 'French Flag Problem' and how positional information is interpreted by a sheet of cells in a large developing system.


Assuntos
Gastrulação , Camadas Germinativas , Animais , Embrião de Galinha , Gástrula
3.
Proc Natl Acad Sci U S A ; 119(10): e2110415119, 2022 03 08.
Artigo em Inglês | MEDLINE | ID: mdl-35238638

RESUMO

SignificanceAmino acids are the building blocks of life and important signaling molecules. Despite their common structure, no universal mechanism for amino acid recognition by cellular receptors is currently known. We discovered a simple motif, which binds amino acids in various receptor proteins from all major life-forms. In humans, this motif is found in subunits of calcium channels that are implicated in pain and neurodevelopmental disorders. Our findings suggest that γ-aminobutyric acid-derived drugs bind to the same motif in human proteins that binds natural ligands in bacterial receptors, thus enabling future improvement of important drugs.


Assuntos
Archaea/química , Proteínas Arqueais/química , Bactérias/química , Proteínas de Bactérias/química , Proteínas de Membrana/química , Motivos de Aminoácidos , Archaea/metabolismo , Proteínas Arqueais/metabolismo , Bactérias/metabolismo , Proteínas de Bactérias/metabolismo , Humanos , Proteínas de Membrana/metabolismo
4.
Breast Cancer Res Treat ; 206(2): 377-385, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38581534

RESUMO

PURPOSE: Although changes in circulating tumour DNA (ctDNA) in breast cancer are well described, the kinetics of their fluctuations has not been described over short timescales. We investigated ctDNA dynamics during alternating cycles of chemotherapy and hormonal treatment in pre-treated patients with oestrogen receptor-positive metastatic breast cancer. METHODS: Patients received alternating, 9-week cycles of eribulin and aromatase inhibitors (AIs). The clinical primary endpoint, progression-free survival (PFS), was monitored at 3, 6 and 9 months; secondary endpoints, clinical benefit rate (CBR), safety and tolerability profiles, were also assessed. Importantly, ctDNA fluctuations were monitored using the Oncomine™ Breast cfDNA assay to test whether biomarkers may change rapidly between chemotherapy and aromatase inhibitor (AI) treatment in the setting of advanced breast cancer, potentially reflecting disease dynamics. RESULTS: The median PFS was 202 days (95% CI: 135-undefined) and 235 days (95% CI: 235-undefined) at 6 and 9 months, respectively, with a 50% CBR at both 6 and 9 months. Dynamic changes in ctDNA were observed in short timescales between chemotherapy and AI treatment and support the clinical benefit (CB) seen in individual patients and, critically, appear informative of acquired resistance in real time. CONCLUSION: Changes in ctDNA can occur rapidly and reflect changes in patients' clinical tumour responses (NCT02681523).


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica , Inibidores da Aromatase , Neoplasias da Mama , DNA Tumoral Circulante , Adulto , Idoso , Feminino , Humanos , Pessoa de Meia-Idade , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Inibidores da Aromatase/uso terapêutico , Inibidores da Aromatase/administração & dosagem , Biomarcadores Tumorais/sangue , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Neoplasias da Mama/sangue , Neoplasias da Mama/mortalidade , Neoplasias da Mama/genética , DNA Tumoral Circulante/sangue , DNA Tumoral Circulante/genética , Furanos/uso terapêutico , Furanos/administração & dosagem , Cetonas , Metástase Neoplásica , Policetídeos de Poliéter , Resultado do Tratamento
5.
Clin Chem ; 70(1): 234-249, 2024 01 04.
Artigo em Inglês | MEDLINE | ID: mdl-38175603

RESUMO

BACKGROUND: Programmed death ligand-1 (PD-L1) expression on circulating tumor cells (CTCs) has been suggested to provide prognostic information in non-small cell lung cancer (NSCLC), but consensus relative to treatment outcomes is lacking. We conducted the first comprehensive meta-analysis exploring its potential as a prognostic and predictive marker, and assessed the concordance between PD-L1 + CTCs and paired tumor tissue in NSCLC patients. METHOD: A comprehensive search was applied to PubMed and EMBASE to identify 26 studies that evaluated PD-L1 + CTCs and their association with survival outcomes in 1236 NSCLC patients. RESULTS: The meta-analysis estimated a mean PD-L1 + CTCs detection rate of 61% (95% CI, 49-72). Subgroup analysis based on treatment showed that PD-L1 + CTCs was not significantly associated with better overall survival (OS) in NSCLC patients treated with immune checkpoint inhibitors (ICIs) (Hazard Ratio (HR) = 0.96, 95% CI, 0.35-2.65, P = 0.944), but was predictive of worse OS in those treated with other therapies (HR = 2.11, 95% CI, 1.32-3.36, P = 0.002). Similarly, PD-L1 + CTCs was not significantly associated with superior progressing free survival (PFS) in NSCLCs treated with ICIs (HR = 0.67, 95% CI, 0.41-1.09, P = 0.121), but was significantly associated with shorter PFS in patients treated with other therapies (HR = 1.91, 95% CI, 1.24-2.94, P = 0.001). The overall estimate for the concordance between PD-L1 expression on CTCs and tumor cells was 63% (95% CI, 44-80). CONCLUSION: The average detection rate of PD-L1 + CTCs was comparable to the rate of PD-L1 expression in NSCLC tumors. There was a trend towards better PFS in ICI-treated NSCLC patients with PD-L1 + CTCs. Larger longitudinal studies on the association of PD-L1 + CTCs with clinical outcomes in NSCLC patients treated with ICIs are warranted.


Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Células Neoplásicas Circulantes , Humanos , Antígeno B7-H1 , Carcinoma Pulmonar de Células não Pequenas/diagnóstico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológico
6.
Allergy ; 79(5): 1258-1270, 2024 05.
Artigo em Inglês | MEDLINE | ID: mdl-38108208

RESUMO

BACKGROUND: This is the first report on the effects of abrocitinib, a Janus kinase 1-selective inhibitor, on the expression of skin biomarkers in patients with moderate-to-severe atopic dermatitis (AD). METHODS: JADE MOA (NCT03915496) was a double-blind Phase 2a trial. Adults were randomly assigned 1:1:1 to receive monotherapy with once-daily abrocitinib 200 mg, abrocitinib 100 mg, or placebo for 12 weeks. The primary endpoint was change from baseline in markers of inflammation (matrix metalloproteinase [MMP]-12), epidermal hyperplasia (keratin-16 [KRT16]), T-helper 2 (Th2) immune response (C-C motif chemokine ligand [CCL]17, CCL18, and CCL26), and Th22 immune response (S100 calcium binding protein A8, A9, and A12 [S100A8, S100A9, and S100A12]) in skin through 12 weeks. RESULTS: A total of 46 patients received abrocitinib 200 mg (n = 14), abrocitinib 100 mg (n = 16), or placebo (n = 16). Abrocitinib improved AD clinical signs and reduced itch. Gene expression of MMP-12, KRT16, S100A8, S100A9, and S100A12 was significantly decreased from baseline with abrocitinib 200 mg (at Weeks 2, 4, and 12) and abrocitinib 100 mg (at Weeks 4 and 12) in a dose-dependent manner. Abrocitinib 200 mg resulted in significant decreases from baseline in CCL17 expression at Week 12 and CCL18 expression at Weeks 2, 4, and 12; no significant decreases were observed for CCL26. CONCLUSIONS: Alongside improvements in clinical signs and symptoms of AD, 12 weeks of abrocitinib treatment resulted in downregulation of genes associated with inflammation, epidermal hyperplasia, and Th2 and Th22 immune responses in the skin of patients with moderate-to-severe AD.


Assuntos
Biomarcadores , Dermatite Atópica , Índice de Gravidade de Doença , Pele , Sulfonamidas , Humanos , Dermatite Atópica/tratamento farmacológico , Feminino , Masculino , Adulto , Pele/patologia , Pele/metabolismo , Pele/efeitos dos fármacos , Sulfonamidas/uso terapêutico , Sulfonamidas/administração & dosagem , Pirimidinas/uso terapêutico , Pirimidinas/administração & dosagem , Pessoa de Meia-Idade , Resultado do Tratamento , Método Duplo-Cego , Adulto Jovem
7.
Clin Chem ; 69(5): 510-518, 2023 04 28.
Artigo em Inglês | MEDLINE | ID: mdl-36747279

RESUMO

BACKGROUND: Somatic copy number alterations (sCNAs) acquired during the evolution of breast cancer provide valuable prognostic and therapeutic information. Here we present a workflow for screening sCNAs using picogram amounts of cell-free DNA (cfDNA) and single circulating tumor cells (CTCs). METHODS: We repurposed the Ion ReproSeq PGS™ preimplantation genetic testing kit to perform shallow whole genome sequencing on 178 cfDNA samples (300 pg) and individual CTCs from 10 MBC patients with metastatic breast cancer (MBC) recovered by CellSearch®/DEPArray™. Results were analyzed using a tailored ichorCNA workflow. RESULTS: sCNAs were detected in cfDNA of 41/105 (39%) patients with MBC and 3/23 (13%) primary breast cancers on follow-up (PBC FU), all of whom subsequently relapsed. In 8 of 10 MBCs, individual CTCs had a higher copy number count than matched cfDNA. The median tumor fraction detected by ichorCNA was 0.34 (range 0.17-0.58) for MBC and 0.36 (range 0.31-0.37) for PBC FU. Patients with detectable tumor fraction (≥ 0.1) and TFx and OncomineTM variants had significantly lower overall survival rates (P values P = 0.002 and P < 0.0001 for the log-rank test, respectively). CONCLUSIONS: The ReproSeq PGS assay is rapid, at approximately $120 per sample, providing both a sCNA profile and estimation of the tumor DNA fraction from limiting cfDNA template (300pg) and individual CTCs. The approach could be used to examine the copy number landscape over time to guide treatment decisions, support future trial designs, and be applied to low volume blood spot samples enabling remote monitoring.


Assuntos
Neoplasias da Mama , Ácidos Nucleicos Livres , Células Neoplásicas Circulantes , Humanos , Feminino , Ácidos Nucleicos Livres/genética , Fluxo de Trabalho , Células Neoplásicas Circulantes/patologia , Neoplasias da Mama/patologia , Sequenciamento Completo do Genoma , Biomarcadores Tumorais/genética
8.
BMC Biol ; 20(1): 111, 2022 05 13.
Artigo em Inglês | MEDLINE | ID: mdl-35549945

RESUMO

BACKGROUND: In vertebrate cells, the Golgi functional subunits, mini-stacks, are linked into a tri-dimensional network. How this "ribbon" architecture relates to Golgi functions remains unclear. Are all connections between mini-stacks equal? Is the local structure of the ribbon of functional importance? These are difficult questions to address, without a quantifiable readout of the output of ribbon-embedded mini-stacks. Endothelial cells produce secretory granules, the Weibel-Palade bodies (WPB), whose von Willebrand Factor (VWF) cargo is central to hemostasis. The Golgi apparatus controls WPB size at both mini-stack and ribbon levels. Mini-stack dimensions delimit the size of VWF "boluses" whilst the ribbon architecture allows their linear co-packaging, thereby generating WPBs of different lengths. This Golgi/WPB size relationship suits mathematical analysis. RESULTS: WPB lengths were quantized as multiples of the bolus size and mathematical modeling simulated the effects of different Golgi ribbon organizations on WPB size, to be compared with the ground truth of experimental data. An initial simple model, with the Golgi as a single long ribbon composed of linearly interlinked mini-stacks, was refined to a collection of mini-ribbons and then to a mixture of mini-stack dimers plus long ribbon segments. Complementing these models with cell culture experiments led to novel findings. Firstly, one-bolus sized WPBs are secreted faster than larger secretory granules. Secondly, microtubule depolymerization unlinks the Golgi into equal proportions of mini-stack monomers and dimers. Kinetics of binding/unbinding of mini-stack monomers underpinning the presence of stable dimers was then simulated. Assuming that stable mini-stack dimers and monomers persist within the ribbon resulted in a final model that predicts a "breathing" arrangement of the Golgi, where monomer and dimer mini-stacks within longer structures undergo continuous linking/unlinking, consistent with experimentally observed WPB size distributions. CONCLUSIONS: Hypothetical Golgi organizations were validated against a quantifiable secretory output. The best-fitting Golgi model, accounting for stable mini-stack dimers, is consistent with a highly dynamic ribbon structure, capable of rapid rearrangement. Our modeling exercise therefore predicts that at the fine-grained level the Golgi ribbon is more complex than generally thought. Future experiments will confirm whether such a ribbon organization is endothelial-specific or a general feature of vertebrate cells.


Assuntos
Células Endoteliais , Fator de von Willebrand , Células Cultivadas , Exocitose , Complexo de Golgi , Corpos de Weibel-Palade/fisiologia , Fator de von Willebrand/farmacologia , Fator de von Willebrand/fisiologia
9.
J Allergy Clin Immunol ; 149(4): 1318-1328, 2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-34863853

RESUMO

BACKGROUND: Janus kinase (JAK) inhibitors have shown encouraging results in the treatment of alopecia areata (AA), an autoimmune form of hair loss, in small, uncontrolled studies and case reports. OBJECTIVE: We conducted a biopsy substudy during the randomized, double-blind, placebo-controlled first 24 weeks of a phase 2a clinical trial that evaluated the efficacy and safety of ritlecitinib, an inhibitor of JAK3 and the tyrosine kinase expressed in hepatocellular carcinoma (TEC) kinase family, and brepocitinib, an inhibitor of tyrosine kinase 2 (TYK2)/JAK1 in the treatment of AA. METHODS: Change in biomarkers in lesional scalp biopsy samples between baseline and weeks 12 and 24 was an exploratory end point, and 46 patients participated from the ritlecitinib (n = 18), brepocitinib (n = 16), and placebo (n = 12) groups. Correlations of biomarkers with hair regrowth, measured using the Severity of Alopecia Tool (SALT) score, were also evaluated. CLINICAL TRIAL REGISTRATION: NCT02974868. RESULTS: At week 24, both ritlecitinib and brepocitinib demonstrated improvement exceeding 100% in the lesional scalp transcriptome toward a nonlesional profile. At week 12, the improvements in scalp tissue were greater with brepocitinib than ritlecitinib; however, at week 24, the improvements were greater with ritlecitinib. CONCLUSIONS: For both ritlecitinib and brepocitinib, improvement in the SALT scores was positively associated with expression of TH1 markers and negatively associated with expression of hair keratins. Larger, long-term clinical trials are warranted.


Assuntos
Alopecia em Áreas , Inibidores de Janus Quinases , Alopecia/tratamento farmacológico , Alopecia em Áreas/tratamento farmacológico , Biomarcadores/metabolismo , Humanos , Inibidores de Janus Quinases/uso terapêutico , Inibidores de Proteínas Quinases/uso terapêutico , Couro Cabeludo
10.
Br J Cancer ; 127(10): 1858-1864, 2022 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-36088510

RESUMO

BACKGROUND: We report copy-number profiling by low-pass WGS (LP-WGS) in individual circulating tumour cells (CTCs) for guiding treatment in patients with metastatic breast cancer (MBC), comparing CTC results with mutations detected in circulating tumour DNA (ctDNA) in the same blood samples. METHODS: Across 10 patients with MBC who were progressing at the time of blood sampling and that had >20 CTCs detected by CellSearch®, 63 single cells (50 CTCs and 13 WBCs) and 16 cell pools (8 CTC pools and 8 WBC pools) were recovered from peripheral blood by CellSearch®/DEPArray™ and sequenced with Ampli1 LowPass technology (Menarini Silicon Biosystems). Copy-number aberrations were identified using the MSBiosuite software platform, and results were compared with mutations detected in matched plasma cfDNA analysed by targeted next-generation sequencing using the Oncomine™ Breast cfDNA Assay (Thermo Fisher). RESULTS: LP-WGS data demonstrated copy-number gains/losses in individual CTCs in regions including FGFR1, JAK2 and CDK6 in five patients, ERBB2 amplification in two HER2-negative patients and BRCA loss in two patients. Seven of eight matched plasmas also had mutations in ctDNA in PIK3CA, TP53, ESR1 and KRAS genes with mutant allele frequencies (MAF) ranging from 0.05 to 33.11%. Combining results from paired CTCs and ctDNA, clinically actionable targets were identified in all ten patients. CONCLUSION: This combined analysis of CTCs and ctDNA may offer a new approach for monitoring of disease progression and to direct therapy in patients with advanced MBC, at a time when they are coming towards the end of other treatment options.


Assuntos
Neoplasias da Mama , Ácidos Nucleicos Livres , DNA Tumoral Circulante , Células Neoplásicas Circulantes , Humanos , Feminino , Neoplasias da Mama/patologia , Células Neoplásicas Circulantes/patologia , DNA Tumoral Circulante/genética , Ácidos Nucleicos Livres/genética , Mutação , Biomarcadores Tumorais/genética
11.
Development ; 146(23)2019 12 04.
Artigo em Inglês | MEDLINE | ID: mdl-31784457

RESUMO

Cell division, movement and differentiation contribute to pattern formation in developing tissues. This is the case in the vertebrate neural tube, in which neurons differentiate in a characteristic pattern from a highly dynamic proliferating pseudostratified epithelium. To investigate how progenitor proliferation and differentiation affect cell arrangement and growth of the neural tube, we used experimental measurements to develop a mechanical model of the apical surface of the neuroepithelium that incorporates the effect of interkinetic nuclear movement and spatially varying rates of neuronal differentiation. Simulations predict that tissue growth and the shape of lineage-related clones of cells differ with the rate of differentiation. Growth is isotropic in regions of high differentiation, but dorsoventrally biased in regions of low differentiation. This is consistent with experimental observations. The absence of directional signalling in the simulations indicates that global mechanical constraints are sufficient to explain the observed differences in anisotropy. This provides insight into how the tissue growth rate affects cell dynamics and growth anisotropy and opens up possibilities to study the coupling between mechanics, pattern formation and growth in the neural tube.


Assuntos
Diferenciação Celular/fisiologia , Células-Tronco Neurais/metabolismo , Tubo Neural/embriologia , Neurogênese/fisiologia , Neurônios/metabolismo , Transdução de Sinais/fisiologia , Animais , Epitélio/embriologia , Camundongos , Células-Tronco Neurais/citologia , Tubo Neural/citologia , Neurônios/citologia
12.
Prev Med ; 154: 106884, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34780853

RESUMO

Cardiovascular disease (CVD) events are highly preventable through appropriate treatment and disproportionally affect socioeconomically disadvantaged individuals. This study quantified the relationship of socioeconomic factors to dispensing and persistent use of lipid- and blood pressure-lowering medication following hospital admission for a major CVD event (myocardial infarction, ischaemic stroke/transient ischaemic attack). Data from 8285 people with such events aged ≥45 years from the Australian 45 and Up Study with linked medication data were used to estimate relative risks (RRs) for combined lipid- and blood pressure-lowering dispensing at three-months following hospital discharge and for 12-month persistent use, in relation to education, income, and level of medication subsidisation. Overall, 56% were dispensed guideline-recommended medications at three months and 37% persistently used them across 12 months. After adjusting for demographic factors, type of CVD and history of CVD hospitalisation, RRs for lowest (no educational qualifications) compared to highest education level (university degree) were 1.14 (95% CI: 1.06, 1.22) for medication dispensing and 1.15 (1.02, 1.29) for persistent medication use; 1.14 (1.06, 1.22) and 1.17 (1.04, 1.32) respectively for lowest (<$20,000) versus highest (≥$70,000) household pre-tax income; and 1.25 (1.17, 1.33) and 1.28 (1.15, 1.43) respectively for those receiving highest versus lowest subsidisation. There was little to no evidence of a relationship of income and education to medication use after adjustment for medication subsidisation. While preventive medication use is sub-optimal, subsidisation is substantially associated with increased use and accounts for most of the relationship with socioeconomic position, suggesting subsidy schemes are working in the intended direction.


Assuntos
Isquemia Encefálica , Doenças Cardiovasculares , Acidente Vascular Cerebral , Austrália , Doenças Cardiovasculares/tratamento farmacológico , Doenças Cardiovasculares/prevenção & controle , Estudos de Coortes , Humanos , Lipídeos , Estudos Prospectivos , Fatores de Risco , Fatores Socioeconômicos
13.
Breast Cancer Res Treat ; 188(2): 465-476, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-34097174

RESUMO

PURPOSE: There is growing interest in the application of circulating tumour DNA (ctDNA) as a sensitive tool for monitoring tumour evolution and guiding targeted therapy in patients with cancer. However, robust comparisons of different platform technologies are still required. Here we compared the InVisionSeq™ ctDNA Assay with the Oncomine™ Breast cfDNA Assay to assess their concordance and feasibility for the detection of mutations in plasma at low (< 0.5%) variant allele fraction (VAF). METHODS: Ninety-six plasma samples from 50 patients with estrogen receptor (ER)-positive metastatic breast cancer (mBC) were profiled using the InVision Assay. Results were compared to the Oncomine assay in 30 samples from 26 patients, where there was sufficient material and variants were covered by both assays. Longitudinal samples were analysed for 8 patients with endocrine resistance. RESULTS: We detected alterations in 59/96 samples from 34/50 patients analysed with the InVision assay, most frequently affecting ESR1, PIK3CA and TP53. Complete or partial concordance was found in 28/30 samples analysed by both assays, and VAF values were highly correlated. Excellent concordance was found for most genes, and most discordant calls occurred at VAF < 1%. In longitudinal samples from progressing patients with endocrine resistance, we detected consistent alterations in sequential samples, most commonly in ESR1 and PIK3CA. CONCLUSION: This study shows that both ultra-deep next-generation sequencing (NGS) technologies can detect genomic alternations even at low VAFs in plasma samples of mBC patients. The strong agreement of the technologies indicates sufficient reproducibility for clinical use as prognosic and predictive biomarker.


Assuntos
Neoplasias da Mama , DNA Tumoral Circulante , Biomarcadores Tumorais/genética , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , DNA Tumoral Circulante/genética , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Reprodutibilidade dos Testes
14.
J Theor Biol ; 528: 110838, 2021 11 07.
Artigo em Inglês | MEDLINE | ID: mdl-34303702

RESUMO

Cancer cells obtain mutations which rely on the production of diffusible growth factors to confer a fitness benefit. These mutations can be considered cooperative, and studied as public goods games within the framework of evolutionary game theory. The population structure, benefit function and update rule all influence the evolutionary success of cooperators. We model the evolution of cooperation in epithelial cells using the Voronoi tessellation model. Unlike traditional evolutionary graph theory, this allows us to implement global updating, for which birth and death events are spatially decoupled. We compare, for a sigmoid benefit function, the conditions for cooperation to be favoured and/or beneficial for well-mixed and structured populations. We find that when population structure is combined with global updating, cooperation is more successful than if there were local updating or the population were well-mixed. Interestingly, the qualitative behaviour for the well-mixed population and the Voronoi tessellation model is remarkably similar, but the latter case requires significantly lower incentives to ensure cooperation.


Assuntos
Comportamento Cooperativo , Teoria dos Jogos , Evolução Biológica , Contagem de Células
15.
J Am Acad Dermatol ; 85(2): 379-387, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-33757798

RESUMO

BACKGROUND: Alopecia areata (AA) is an autoimmune form of hair loss with limited treatments. OBJECTIVE: To evaluate the efficacy and safety of the Janus kinase inhibitors ritlecitinib and brepocitinib in patients who have AA with ≥ 50% scalp hair loss. METHODS: Patients were randomized to once-daily ritlecitinib, brepocitinib, or placebo. The primary efficacy endpoint was a 24-week change from baseline in the Severity of Alopecia Tool (SALT) score. The key secondary efficacy endpoint was the proportion of patients achieving 30% improvement in SALT score (SALT30). RESULTS: The ritlecitinib, brepocitinib, and placebo groups included 48, 47, and 47 patients, respectively. At week 24, least-squares mean difference from placebo in SALT score change from baseline was 31.1 (95% confidence interval [CI], 18.8-43.5) for ritlecitinib and 49.2 (95% CI, 36.6-61.7) for brepocitinib (P < .0001 for both comparisons with placebo). SALT30 was achieved by 50% (90% CI, 38%-62%) of patients receiving ritlecitinib, 64% (90% CI, 51%-75%) receiving brepocitinib, and 2% (90% CI, 0%-9%) receiving placebo. Two patients experienced a serious adverse event (rhabdomyolysis) in the brepocitinib group only. LIMITATIONS: Only a single-dosage regimen of each study drug was included. CONCLUSION: Treatment with ritlecitinib or brepocitinib for 24 weeks was efficacious and generally well tolerated.


Assuntos
Alopecia em Áreas/tratamento farmacológico , Inibidores de Janus Quinases/uso terapêutico , Inibidores de Proteínas Quinases/uso terapêutico , Pirazóis/uso terapêutico , Pirimidinas/uso terapêutico , Pirróis/uso terapêutico , Adulto , Método Duplo-Cego , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Fatores de Tempo , Resultado do Tratamento , Adulto Jovem
16.
Br J Cancer ; 123(8): 1271-1279, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32719550

RESUMO

BACKGROUND: Gastroesophageal adenocarcinoma (GOA) has poor clinical outcomes and lacks reliable blood markers. Here we present circulating tumour DNA (ctDNA) as an emerging biomarker. METHODS: Forty patients (17 palliative and 23 curative) were followed by serial plasma monitoring. Primary tumour DNA was analysed by targeted next-generation sequencing to identify somatic single-nucleotide variants (SNVs), and Nanostring nCounter® to detect copy number alterations (CNAs). Patient-specific SNVs and CNA amplifications (CNAamp) were analysed in plasma using digital droplet PCR and quantitative PCR, respectively. RESULTS: Thirty-five patients (13 palliative, 22 curative) had ≥1 SNVs and/or CNAamp detected in primary tumour DNA suitable for tracking in plasma. Eighteen of 35 patients (nine palliative, nine curative) had ≥1 ctDNA-positive plasma sample. Detection of postoperative ctDNA predicted short RFS (190 vs 934 days, HR = 3.7, p = 0.028) and subsequent relapse (PPV for relapse 0.83). High ctDNA levels (>60.5 copies/ml) at diagnosis of metastatic disease predicted poor OS (90 vs 372 days, HR = 11.7 p < 0.001). CONCLUSION: Sensitive ctDNA detection allows disease monitoring and prediction of short OS in metastatic patients. Presence of ctDNA postoperatively predicts relapse and defines a 'molecular relapse' before overt clinical disease. This lead time defines a potential therapeutic window for additional anticancer therapy.


Assuntos
Adenocarcinoma/genética , DNA Tumoral Circulante/sangue , Neoplasias Esofágicas/genética , Recidiva Local de Neoplasia/genética , Neoplasias Gástricas/genética , Adenocarcinoma/mortalidade , Idoso , Idoso de 80 Anos ou mais , Variações do Número de Cópias de DNA , Neoplasias Esofágicas/mortalidade , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Estudos Longitudinais , Pessoa de Meia-Idade , Prognóstico , Neoplasias Gástricas/mortalidade
17.
Breast Cancer Res ; 21(1): 149, 2019 12 19.
Artigo em Inglês | MEDLINE | ID: mdl-31856868

RESUMO

BACKGROUND: Breast cancer (BC) is the most common cancer in women, and despite the introduction of new screening programmes, therapies and monitoring technologies, there is still a need to develop more useful tests for monitoring treatment response and to inform clinical decision making. The purpose of this study was to compare circulating cell-free DNA (cfDNA) and circulating tumour cells (CTCs) with conventional breast cancer blood biomarkers (CA15-3 and alkaline phosphatase (AP)) as predictors of response to treatment and prognosis in patients with metastatic breast cancer (MBC). METHODS: One hundred ninety-four female patients with radiologically confirmed MBC were recruited to the study. Total cfDNA levels were determined by qPCR and compared with CELLSEARCH® CTC counts and CA15-3 and alkaline phosphatase (AP) values. Blood biomarker data were compared with conventional tumour markers, treatment(s) and response as assessed by RECIST and survival. Non-parametric statistical hypothesis tests were used to examine differences, correlation analysis and linear regression to determine correlation and to describe its effects, logistic regression and receiver operating characteristic curve (ROC curve) to estimate the strength of the relationship between biomarkers and clinical outcomes and value normalization against standard deviation to make biomarker values comparable. Kaplan-Meier estimator and Cox regression models were used to assess survival. Univariate and multivariate models were performed where appropriate. RESULTS: Multivariate analysis showed that both the amount of total cfDNA (p value = 0.024, HR = 1.199, CI = 1.024-1.405) and the number of CTCs (p value = 0.001, HR = 1.243, CI = 1.088-1.421) are predictors of overall survival (OS), whereas total cfDNA levels is the sole predictor for progression-free survival (PFS) (p value = 0.042, HR = 1.193, CI = 1.007-1.415) and disease response when comparing response to non-response to treatment (HR = 15.917, HR = 12.481 for univariate and multivariate analysis, respectively). Lastly, combined analysis of CTCs and cfDNA is more informative than the combination of two conventional biomarkers (CA15-3 and AP) for prediction of OS. CONCLUSION: Measurement of total cfDNA levels, which is a simpler and less expensive biomarker than CTC counts, is associated with PFS, OS and response in MBC, suggesting potential clinical application of a cheap and simple blood-based test.


Assuntos
Biomarcadores Tumorais , Neoplasias da Mama/genética , Neoplasias da Mama/mortalidade , DNA Tumoral Circulante , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias da Mama/diagnóstico , Neoplasias da Mama/terapia , Tomada de Decisão Clínica , Gerenciamento Clínico , Feminino , Humanos , Estimativa de Kaplan-Meier , Biópsia Líquida , Imageamento por Ressonância Magnética , Pessoa de Meia-Idade , Células Neoplásicas Circulantes/metabolismo , Células Neoplásicas Circulantes/patologia , Razão de Chances , Prognóstico , Tomografia Computadorizada por Raios X
18.
J Theor Biol ; 481: 44-53, 2019 11 21.
Artigo em Inglês | MEDLINE | ID: mdl-31059715

RESUMO

A ring oscillator is a system in which one species regulates the next, which regulates the next and so on until the last species regulates the first. In addition, the number of the regulations which are negative, and so result in a reduction in the regulated species, is odd, making the overall feedback in the loop negative. In ring oscillators, the probability of oscillations is maximised if the degradation rates of the species are equal. When there is more than one loop in the regulatory network, the dynamics can be more complicated. Here, a systematic way of organising the characteristic equation of ODE models of regulatory networks is provided. This facilitates the identification of Hopf bifurcations. It is shown that the probability of oscillations in non-ring systems is maximised for unequal degradation rates. For example, when there is a ring and a second ring employing a subset of the genes in the first ring, then the probability of oscillations is maximised when the species in the sub-ring degrade more slowly than those outside, for a negative feedback subring. When the sub-ring forms a positive feedback loop, the optimal degradation rates are larger for the species in the sub-ring, provided the positive feedback is not too strong. By contrast, optimal degradation rates are smaller for the species in the sub-ring, when the positive feedback is very strong. Adding a positive feedback loop to a repressilator increases the probability of oscillations, provided the positive feedback is not too strong, whereas adding a negative feedback loop decreases the probability of oscillations. The work is illustrated with numerical simulations of example systems: an autoregulatory gene model in which transcription is downregulated by the protein dimer and three-species and four-species gene regulatory network examples.


Assuntos
Relógios Biológicos , Simulação por Computador , Redes Reguladoras de Genes , Modelos Genéticos
19.
Phys Rev Lett ; 120(12): 128102, 2018 Mar 23.
Artigo em Inglês | MEDLINE | ID: mdl-29694079

RESUMO

Cell state determination is the outcome of intrinsically stochastic biochemical reactions. Transitions between such states are studied as noise-driven escape problems in the chemical species space. Escape can occur via multiple possible multidimensional paths, with probabilities depending nonlocally on the noise. Here we characterize the escape from an oscillatory biochemical state by minimizing the Freidlin-Wentzell action, deriving from it the stochastic spiral exit path from the limit cycle. We also use the minimized action to infer the escape time probability density function.


Assuntos
Relógios Biológicos , Modelos Biológicos , Algoritmos , Fenômenos Fisiológicos Celulares , Processos Estocásticos
20.
J Biol Chem ; 291(39): 20402-16, 2016 09 23.
Artigo em Inglês | MEDLINE | ID: mdl-27489103

RESUMO

CaVß subunits interact with the voltage-gated calcium channel CaV2.2 on a site in the intracellular loop between domains I and II (the I-II loop). This interaction influences the biophysical properties of the channel and leads to an increase in its trafficking to the plasma membrane. We have shown previously that a mutant CaV2.2 channel that is unable to bind CaVß subunits (CaV2.2 W391A) was rapidly degraded (Waithe, D., Ferron, L., Page, K. M., Chaggar, K., and Dolphin, A. C. (2011) J. Biol. Chem. 286, 9598-9611). Here we show that, in the absence of CaVß subunits, a construct consisting of the I-II loop of CaV2.2 was directly ubiquitinated and degraded by the proteasome system. Ubiquitination could be prevented by mutation of all 12 lysine residues in the I-II loop to arginines. Including a palmitoylation motif at the N terminus of CaV2.2 I-II loop was insufficient to target it to the plasma membrane in the absence of CaVß subunits even when proteasomal degradation was inhibited with MG132 or ubiquitination was prevented by the lysine-to-arginine mutations. In the presence of CaVß subunit, the palmitoylated CaV2.2 I-II loop was protected from degradation, although oligoubiquitination could still occur, and was efficiently trafficked to the plasma membrane. We propose that targeting to the plasma membrane requires a conformational change in the I-II loop that is induced by binding of the CaVß subunit.


Assuntos
Canais de Cálcio Tipo N/metabolismo , Membrana Celular/metabolismo , Lipoilação/fisiologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteólise , Ubiquitinação/fisiologia , Substituição de Aminoácidos , Animais , Canais de Cálcio Tipo N/genética , Linhagem Celular , Membrana Celular/genética , Mutação de Sentido Incorreto , Complexo de Endopeptidases do Proteassoma/genética , Estrutura Secundária de Proteína , Coelhos , Ratos
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