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1.
Cureus ; 16(5): e61106, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38933636

RESUMO

Trisomy 21 often leads to cardiac complications, usually associated with congenital heart disease, such as atrial septal defects, ventricular septal defects, and patent ductus arteriosus. This case describes an unexpected instance of infective endocarditis (IE) in a middle-aged patient with an incidentally discovered patent foramen ovale (PFO). The common risk factors for IE include previous valve surgery, artificial heart valves, pacemakers, prior IE, congenital defects like bicuspid aortic valve, IV drug use, and the congenital defects mentioned earlier.

2.
Int J Mol Sci ; 23(5)2022 Mar 07.
Artigo em Inglês | MEDLINE | ID: mdl-35270029

RESUMO

Aromatase inhibitors (AIs) are standard treatment for estrogen-dependent postmenopausal breast tumors; however, resistance develops leading to tumor relapse and metastasis. We previously demonstrated that glyceollin inhibits proliferation, survival, and migration of hormone-independent letrozole-resistant breast cancer. Since many AI-resistant tumors remain hormone-dependent, identifying distinctions between estrogen-receptor-positive (ER+) and ER-negative (ER-) AI-resistant tumor response to therapy is critical. We hypothesize that treating ER+ letrozole-resistant T47D breast cancer cells (T47DaromLR) with a combination of 10 µM glyceollin and 0.5 µM lapatinib (a dual EGFR/HER2 inhibitor) will decrease cell proliferation through induction of apoptosis. The T47DaromLR cells were found to overexpress HER2 and MAPK while maintaining aromatase and ER levels compared to their letrozole-sensitive (T47Darom) counterparts. In the absence of estrogen stimulation, glyceollin ± lapatinib had no effect on the proliferation of the T47Darom cells, while glyceollin treatment caused 46% reduction in the proliferation of T47DaromLR cells, which was further diminished when combined with lapatinib. While neither agent influenced cell migration, glyceollin and lapatinib reduced S and G2/M phase cell entry and exclusively induced apoptosis by 1.29-fold in the T47DaromLR cells. Taken together, these results suggest that glyceollins and lapatinib may have potential as a novel combination therapeutic approach for hormone-dependent, letrozole-resistant tumors.


Assuntos
Inibidores da Aromatase , Neoplasias da Mama , Apoptose , Aromatase , Inibidores da Aromatase/farmacologia , Inibidores da Aromatase/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos , Estrogênios/farmacologia , Estrogênios/uso terapêutico , Feminino , Humanos , Lapatinib/farmacologia , Lapatinib/uso terapêutico , Letrozol/farmacologia , Recidiva Local de Neoplasia/tratamento farmacológico , Nitrilas/uso terapêutico , Pterocarpanos , Triazóis/farmacologia
3.
J Med Chem ; 64(20): 15141-15169, 2021 10 28.
Artigo em Inglês | MEDLINE | ID: mdl-34643390

RESUMO

Src homology 2 (SH2) domain-containing phosphatase 2 (SHP2) plays a role in receptor tyrosine kinase (RTK), neurofibromin-1 (NF-1), and Kirsten rat sarcoma virus (KRAS) mutant-driven cancers, as well as in RTK-mediated resistance, making the identification of small-molecule therapeutics that interfere with its function of high interest. Our quest to identify potent, orally bioavailable, and safe SHP2 inhibitors led to the discovery of a promising series of pyrazolopyrimidinones that displayed excellent potency but had a suboptimal in vivo pharmacokinetic (PK) profile. Hypothesis-driven scaffold optimization led us to a series of pyrazolopyrazines with excellent PK properties across species but a narrow human Ether-à-go-go-Related Gene (hERG) window. Subsequent optimization of properties led to the discovery of the pyrimidinone series, in which multiple members possessed excellent potency, optimal in vivo PK across species, and no off-target activities including no hERG liability up to 100 µM. Importantly, compound 30 (IACS-15414) potently suppressed the mitogen-activated protein kinase (MAPK) pathway signaling and tumor growth in RTK-activated and KRASmut xenograft models in vivo.


Assuntos
Antineoplásicos/farmacologia , Descoberta de Drogas , Inibidores Enzimáticos/farmacologia , Proteína Tirosina Fosfatase não Receptora Tipo 11/antagonistas & inibidores , Administração Oral , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/administração & dosagem , Inibidores Enzimáticos/química , Humanos , Camundongos , Estrutura Molecular , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Proteína Tirosina Fosfatase não Receptora Tipo 11/metabolismo , Relação Estrutura-Atividade
4.
J Org Chem ; 86(18): 13025-13040, 2021 09 17.
Artigo em Inglês | MEDLINE | ID: mdl-34498466

RESUMO

N-Quaternized ketene N,O-acetals are typically an unstable, transient class of compounds most commonly observed as reactive intermediates. In this report, we describe a general synthetic approach to a variety of bench-stable N-quaternized ketene N,O-acetals via treatment of pyridine or aniline bases with acetylenic ethers and an appropriate Brønsted or Lewis acid (triflic acid, triflimide, or scandium(III) triflate). The resulting pyridinium and anilinium salts can be used as reagents or synthetic intermediates in multiple reaction types. For example, N-(1-ethoxyvinyl)pyridinium or anilinium salts can thermally release highly reactive O-ethyl ketenium ions for use in acid catalyst-free electrophilic aromatic substitutions. N-(1-Ethoxyvinyl)-2-halopyridinium salts can be employed in peptide couplings as a derivative of Mukaiyama reagents or react with amines in nucleophilic aromatic substitutions under mild conditions. These preliminary reactions illustrate the broad potential of these currently understudied compounds in organic synthesis.


Assuntos
Acetais , Cetonas , Técnicas de Química Sintética , Etilenos , Indicadores e Reagentes
5.
ACS Chem Biol ; 16(7): 1111-1127, 2021 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-34166593

RESUMO

The interrogation and manipulation of biological systems by small molecules is a powerful approach in chemical biology. Ideal compounds selectively engage a target and mediate a downstream phenotypic response. Although historically small molecule drug discovery has focused on proteins and enzymes, targeting RNA is an attractive therapeutic alternative, as many disease-causing or -associated RNAs have been identified through genome-wide association studies. As the field of RNA chemical biology emerges, the systematic evaluation of target validation and modulation of target-associated pathways is of paramount importance. In this Review, through an examination of case studies, we outline the experimental characterization, including methods and tools, to evaluate comprehensively the impact of small molecules that target RNA on cellular phenotype.


Assuntos
Compostos Orgânicos/farmacologia , RNA/metabolismo , Animais , Linhagem Celular Tumoral , Descoberta de Drogas , Humanos , Splicing de RNA/efeitos dos fármacos , Riboswitch/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/farmacologia
6.
Chem Soc Rev ; 49(19): 7167-7199, 2020 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-32975549

RESUMO

Targeting RNAs with small molecules represents a new frontier in drug discovery and development. The rich structural diversity of folded RNAs offers a nearly unlimited reservoir of targets for small molecules to bind, similar to small molecule occupancy of protein binding pockets, thus creating the potential to modulate human biology. Although the bacterial ribosome has historically been the most well exploited RNA target, advances in RNA sequencing technologies and a growing understanding of RNA structure have led to an explosion of interest in the direct targeting of human pathological RNAs. This review highlights recent advances in this area, with a focus on the design of small molecule probes that selectively engage structures within disease-causing RNAs, with micromolar to nanomolar affinity. Additionally, we explore emerging RNA-target strategies, such as bleomycin A5 conjugates and ribonuclease targeting chimeras (RIBOTACs), that allow for the targeted degradation of RNAs with impressive potency and selectivity. The compounds discussed in this review have proven efficacious in human cell lines, patient-derived cells, and pre-clinical animal models, with one compound currently undergoing a Phase II clinical trial and another that recently garnerd FDA-approval, indicating a bright future for targeted small molecule therapeutics that affect RNA function.


Assuntos
MicroRNAs/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/farmacologia , Animais , Sistemas de Liberação de Medicamentos , Humanos , MicroRNAs/química , Conformação de Ácido Nucleico
7.
Cancer Res ; 80(21): 4840-4853, 2020 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-32928921

RESUMO

Src homology 2 domain-containing phosphatase (SHP2) is a phosphatase that mediates signaling downstream of multiple receptor tyrosine kinases (RTK) and is required for full activation of the MAPK pathway. SHP2 inhibition has demonstrated tumor growth inhibition in RTK-activated cancers in preclinical studies. The long-term effectiveness of tyrosine kinase inhibitors such as the EGFR inhibitor (EGFRi), osimertinib, in non-small cell lung cancer (NSCLC) is limited by acquired resistance. Multiple clinically identified mechanisms underlie resistance to osimertinib, including mutations in EGFR that preclude drug binding as well as EGFR-independent activation of the MAPK pathway through alternate RTK (RTK-bypass). It has also been noted that frequently a tumor from a single patient harbors more than one resistance mechanism, and the plasticity between multiple resistance mechanisms could restrict the effectiveness of therapies targeting a single node of the oncogenic signaling network. Here, we report the discovery of IACS-13909, a specific and potent allosteric inhibitor of SHP2, that suppresses signaling through the MAPK pathway. IACS-13909 potently impeded proliferation of tumors harboring a broad spectrum of activated RTKs as the oncogenic driver. In EGFR-mutant osimertinib-resistant NSCLC models with EGFR-dependent and EGFR-independent resistance mechanisms, IACS-13909, administered as a single agent or in combination with osimertinib, potently suppressed tumor cell proliferation in vitro and caused tumor regression in vivo. Together, our findings provide preclinical evidence for using a SHP2 inhibitor as a therapeutic strategy in acquired EGFRi-resistant NSCLC. SIGNIFICANCE: These findings highlight the discovery of IACS-13909 as a potent, selective inhibitor of SHP2 with drug-like properties, and targeting SHP2 may serve as a therapeutic strategy to overcome tumor resistance to osimertinib.


Assuntos
Antineoplásicos/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Neoplasias Experimentais/patologia , Proteína Tirosina Fosfatase não Receptora Tipo 11/antagonistas & inibidores , Acrilamidas/farmacologia , Compostos de Anilina/farmacologia , Animais , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Receptores ErbB/genética , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Camundongos , Mutação , Neoplasias Experimentais/genética , Ensaios Antitumorais Modelo de Xenoenxerto
8.
Am J Pharm Educ ; 83(6): 6959, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31507290

RESUMO

Members from Cohort 13 of the Academic Leadership Fellows Program (ALFP) 2016-2017 were challenged to present a debate on the topic: "In Turbulent Times, Pharmacy Education Leaders Must Take Aggressive Action to Prevent Further Declines in Enrollment" at the American Association of Colleges of Pharmacy INfluence 2017 meeting in Rio Grande, Puerto Rico. This paper is the result of thoughtful insights emerging from this debate. We present a discussion of the question of whether pharmacy education leaders must take aggressive action or strategic approaches to prevent further declines in enrollment. There are many thoughts regarding current declines in enrollment. Some educators contend that a more aggressive approach is needed while others argue that, while aggressive actions might lead to short-term gains, a more viable approach involves strategic actions targeting the underlying causes for decreasing enrollment. This paper explores themes of enrollment challenges, current and future workforce needs, and financial issues for both pharmacy programs and students. In summation, both aggressive actions and a strategic, sustainable approach are urgently needed to address declining enrollment.


Assuntos
Educação em Farmácia/tendências , Faculdades de Farmácia/tendências , Humanos , Liderança , Assistência Farmacêutica/tendências , Farmácia/tendências , Estudantes de Farmácia , Estados Unidos
9.
Front Med (Lausanne) ; 6: 122, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31214592

RESUMO

Experimental therapeutic oncology agents are often combined to circumvent tumor resistance to individual agents. However, most combination trials fail to demonstrate sufficient safety and efficacy to advance to a later phase. This study collected survey data on phase 1 combination therapy trials identified from ClinicalTrials.gov between January 1, 2003 and November 30, 2017 to assess trial design and the progress of combinations toward regulatory approval. Online surveys (N = 289, 23 questions total) were emailed to Principal Investigators (PIs) of early-phase National Cancer Institute and/or industry trials; 263 emails (91%) were received and 113 surveys completed (43%). Among phase 1 combination trials, 24.9% (95%CI: 15.3%, 34.4%) progressed to phase 2 or further; 18.7% (95%CI: 5.90%, 31.4%) progressed to phase 3 or regulatory approval; and 12.4% (95%CI: 0.00%, 25.5%) achieved regulatory approval. Observations of "clinical promise" in phase 1 combination studies were associated with higher rates of advancement past each milestone toward regulatory approval (cumulative OR = 11.9; p = 0.0002). Phase 1 combination study designs were concordant with Clinical Trial Design Task Force (CTD-TF) Recommendations 79.6% of the time (95%CI: 72.2%, 87.1%). Most discordances occurred where no plausible pharmacokinetic or pharmacodynamic interactions were expected. Investigator-defined "clinical promise" of a combination is associated with progress toward regulatory approval. Although concordance between study designs of phase 1 combination trials and CTD-TF Recommendations was relatively high, it may be beneficial to raise awareness about the best study design to use when no plausible pharmacokinetic or pharmacodynamic interactions are expected.

10.
Molecules ; 23(2)2018 Feb 14.
Artigo em Inglês | MEDLINE | ID: mdl-29443883

RESUMO

N-Substituted pyridinium salts constitute one of the most valuable reagent classes in organic synthesis, due to their versatility and ease of use. Herein we report a preliminary synthesis and detailed structural analysis of several N-(1-ethoxyvinyl)pyridinium triflates, an unusual class of pyridinium salts with potentially broad use as a reagent in organic synthesis. Treatment of pyridines with trifluoromethane sulfonic acid and ethoxyacetylene generates stable, isolable adducts which have been extensively characterized, due to their novelty. Three-dimensional structural stability is perpetuated by an array of C-H•••O hydrogen bonds involving oxygen atoms from the -SO3 groups of the triflate anion, and hydrogen atoms from the aromatic ring and vinyl group of the pyridinium cation. Predictions from density functional theory calculations of the energy landscape for rotation about the exocyclic C-N bond of 2-chloro-1-(1-ethoxyvinyl)pyridine-1-ium trifluoromethanesulfonate (7) and 1-(1-ethoxyvinyl)pyridine-1-ium trifluoromethanesulfonate (16) are also reported. Notably, the predicted global energy minimum of 7 was nearly identical to that found within the crystal structure.


Assuntos
Mesilatos/química , Piridinas/química , Compostos de Piridínio/química , Ligação de Hidrogênio , Mesilatos/síntese química , Modelos Moleculares , Estrutura Molecular , Oxigênio/química , Piridinas/síntese química , Compostos de Piridínio/síntese química , Sais/química
11.
Int J Environ Res Public Health ; 13(1): ijerph13010010, 2015 Dec 22.
Artigo em Inglês | MEDLINE | ID: mdl-26703648

RESUMO

Although aromatase inhibitors are standard endocrine therapy for postmenopausal women with early-stage metastatic estrogen-dependent breast cancer, they are limited by the development of drug resistance. A better understanding of this process is critical towards designing novel strategies for disease management. Previously, we demonstrated a global proteomic signature of letrozole-resistance associated with hormone-independence, enhanced cell motility and implications of epithelial mesenchymal transition (EMT). Letrozole-resistant breast cancer cells (LTLT-Ca) were treated with a novel phytoalexin, glyceollin I, and exhibited morphological characteristics synonymous with an epithelial phenotype and decreased proliferation. Letrozole-resistance increased Zinc Finger E-Box Binding Homeobox 1 (ZEB1) expression (4.51-fold), while glyceollin I treatment caused a -3.39-fold reduction. Immunofluorescence analyses resulted of glyceollin I-induced increase and decrease in E-cadherin and ZEB1, respectively. In vivo studies performed in ovariectomized, female nude mice indicated that glyceollin treated tumors stained weakly for ZEB1 and N-cadherin and strongly for E-cadherin. Compared to letrozole-sensitive cells, LTLT-Ca cells displayed enhanced motility, however in the presence of glyceollin I, exhibited a 68% and 83% decrease in invasion and migration, respectively. These effects of glyceollin I were mediated in part by inhibition of ZEB1, thus indicating therapeutic potential of glyceollin I in targeting EMT in letrozole resistant breast cancer.


Assuntos
Antineoplásicos/metabolismo , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/fisiopatologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Nitrilas/metabolismo , Pterocarpanos/metabolismo , Triazóis/metabolismo , Animais , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral/efeitos dos fármacos , Movimento Celular , Feminino , Humanos , Letrozol , Camundongos , Camundongos Nus , Nitrilas/uso terapêutico , Fatores de Transcrição/metabolismo , Triazóis/uso terapêutico
12.
Science ; 348(6240): 1217, 2015 Jun 12.
Artigo em Inglês | MEDLINE | ID: mdl-26068842

RESUMO

Reid and Nicchitta propose that most cellular translation is carried out by a noncycling pool of endoplasmic reticulum (ER)-associated ribosomes. However, proximity-specific ribosome profiling data place an upper bound of about 7 to 16% on the fraction of cytosolic protein translation carried out by ribosomes accessible to ER-tethered biotin ligases. Moreover, yeast pulse-labeling experiments argue against there being a static population of ER-associated ribosomes.


Assuntos
Células/metabolismo , Retículo Endoplasmático/metabolismo , Mitocôndrias/metabolismo , Biossíntese de Proteínas , Ribossomos/metabolismo , Humanos
13.
Appl Acoust ; 102: 1-11, 2015 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-27053775

RESUMO

Knowledge of the auditory and non-auditory effects of noise has increased dramatically over the past decade, but indoor noise exposure measurement methods have not advanced appreciably, despite the introduction of applicable new technologies. This study evaluated various conventional and smart devices for exposure assessment in the National Children's Study. Three devices were tested: a sound level meter (SLM), a dosimeter, and a smart device with a noise measurement application installed. Instrument performance was evaluated in a series of semi-controlled tests in office environments over 96-hour periods, followed by measurements made continuously in two rooms (a child's bedroom and a most used room) in nine participating homes over a 7-day period with subsequent computation of a range of noise metrics. The SLMs and dosimeters yielded similar A-weighted average noise levels. Levels measured by the smart devices often differed substantially (showing both positive and negative bias, depending on the metric) from those measured via SLM and dosimeter, and demonstrated attenuation in some frequency bands in spectral analysis compared to SLM results. Virtually all measurements exceeded the Environmental Protection Agency's 45 dBA day-night limit for indoor residential exposures. The measurement protocol developed here can be employed in homes, demonstrates the possibility of measuring long-term noise exposures in homes with technologies beyond traditional SLMs, and highlights potential pitfalls associated with measurements made by smart devices.

14.
Science ; 346(6210): 748-51, 2014 Nov 07.
Artigo em Inglês | MEDLINE | ID: mdl-25378625

RESUMO

Nearly all mitochondrial proteins are nuclear-encoded and are targeted to their mitochondrial destination from the cytosol. Here, we used proximity-specific ribosome profiling to comprehensively measure translation at the mitochondrial surface in yeast. Most inner-membrane proteins were cotranslationally targeted to mitochondria, reminiscent of proteins entering the endoplasmic reticulum (ER). Comparison between mitochondrial and ER localization demonstrated that the vast majority of proteins were targeted to a specific organelle. A prominent exception was the fumarate reductase Osm1, known to reside in mitochondria. We identified a conserved ER isoform of Osm1, which contributes to the oxidative protein-folding capacity of the organelle. This dual localization was enabled by alternative translation initiation sites encoding distinct targeting signals. These findings highlight the exquisite in vivo specificity of organellar targeting mechanisms.


Assuntos
Mitocôndrias/metabolismo , Proteínas Mitocondriais/metabolismo , Iniciação Traducional da Cadeia Peptídica , Ribossomos/metabolismo , Saccharomyces cerevisiae/metabolismo , Animais , Retículo Endoplasmático/metabolismo , Proteínas Mitocondriais/biossíntese , Proteínas Mitocondriais/química , Dobramento de Proteína , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Succinato Desidrogenase/metabolismo
15.
Science ; 346(6210): 1257521, 2014 Nov 07.
Artigo em Inglês | MEDLINE | ID: mdl-25378630

RESUMO

Localized protein synthesis is a fundamental mechanism for creating distinct subcellular environments. Here we developed a generalizable proximity-specific ribosome profiling strategy that enables global analysis of translation in defined subcellular locations. We applied this approach to the endoplasmic reticulum (ER) in yeast and mammals. We observed the large majority of secretory proteins to be cotranslationally translocated, including substrates capable of posttranslational insertion in vitro. Distinct translocon complexes engaged nascent chains at different points during synthesis. Whereas most proteins engaged the ER immediately after or even before signal sequence (SS) emergence, a class of Sec66-dependent proteins entered with a looped SS conformation. Finally, we observed rapid ribosome exchange into the cytosol after translation termination. These data provide insights into how distinct translocation mechanisms act in concert to promote efficient cotranslational recruitment.


Assuntos
Células/metabolismo , Retículo Endoplasmático/metabolismo , Mitocôndrias/metabolismo , Biossíntese de Proteínas , Ribossomos/metabolismo , Biotinilação , Carbono-Nitrogênio Ligases/genética , Carbono-Nitrogênio Ligases/metabolismo , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Células HEK293 , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Fosfoproteínas Fosfatases/genética , Fosfoproteínas Fosfatases/metabolismo , Sinais Direcionadores de Proteínas , Transporte Proteico , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Ribossomos/genética , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo
16.
Mol Cell Proteomics ; 12(9): 2440-55, 2013 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-23704778

RESUMO

Aromatase inhibitors, such as letrozole, have become the first-line treatment for postmenopausal women with estrogen-dependent breast cancer. However, acquired resistance remains a major clinical obstacle. Previous studies demonstrated constitutive activation of the MAPK signaling, overexpression of HER2, and down-regulation of aromatase and ERα in letrozole-resistant breast cancer cells. Given the complex signaling network involved in letrozole-refractory breast cancer and the lack of effective treatment for hormone resistance, further investigation of aromatase inhibitor resistance by a novel systems biology approach may reveal previously unconsidered molecular changes that could be utilized as therapeutic targets. This study was undertaken to characterize for the first time global proteomic alterations occurring in a letrozole-resistant cell line. A quantitative proteomic analysis of the whole cell lysates of LTLT-Ca (resistant) versus AC-1 cells (sensitive) was performed to identify significant protein expression changes. A total of 1743 proteins were identified and quantified, of which 411 were significantly up-regulated and 452 significantly down-regulated (p < 0.05, fold change > 1.20). Bioinformatics analysis revealed that acquired letrozole resistance is associated with a hormone-independent, more aggressive phenotype. LTLT-Ca cells exhibited 84% and 138% increase in migration and invasion compared with the control cells. The ROCK inhibitor partially abrogated the enhanced migration and invasion of the letrozole-resistant cells. Flow cytometric analyses also demonstrated an increase in vimentin and twist expression in letrozole-resistance cells, suggesting an onset of epithelial to mesenchymal transition (EMT). Moreover, targeted gene expression arrays confirmed a 28-fold and sixfold up-regulation of EGFR and HER2, respectively, whereas ERα and pS2 were dramatically reduced by 28-fold and 1100-fold, respectively. Taken together, our study revealed global proteomic signatures of a letrozole-resistant cell line associated with hormone independence, enhanced cell motility, EMT and the potential values of several altered proteins as novel prognostic markers or therapeutic targets for letrozole resistant breast cancer.


Assuntos
Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Movimento Celular/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Estrogênios/metabolismo , Nitrilas/farmacologia , Proteômica/métodos , Triazóis/farmacologia , Citoesqueleto de Actina/efeitos dos fármacos , Citoesqueleto de Actina/metabolismo , Amidas/farmacologia , Western Blotting , Neoplasias da Mama/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Forma Celular/efeitos dos fármacos , Forma Celular/genética , Resistencia a Medicamentos Antineoplásicos/genética , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Estimativa de Kaplan-Meier , Letrozol , Mesoderma/efeitos dos fármacos , Mesoderma/metabolismo , Mesoderma/patologia , Invasividade Neoplásica , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Piridinas/farmacologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Vimentina/metabolismo , Quinases Associadas a rho/metabolismo , Proteína rhoA de Ligação ao GTP/metabolismo
17.
J Health Care Poor Underserved ; 24(1 Suppl): 93-103, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-23395946

RESUMO

Diabetic patients taking metformin have lower incidence of breast cancer than those taking other anti-diabetic medications. Additionally, triple negative breast cancer (TNBC), a form of breast cancer disproportionately afflicting premenopausal African American women, shows atypical susceptibility to metformin's antiproliferative effect. The mechanisms involved in metformin's function in TNBC has not yet been fully elucidated. Therefore, we sought to identify pathways regulated by metformin in using the MDA-MB-468 TNBC cell model. Metformin dose-dependently caused apoptosis, decreased cell viability, and induced cell morphology/chromatin condensation consistent with the permanent proliferative arrest. Furthermore, gene expression arrays revealed that metformin caused expression of stress markers DDIT3, CYP1A1,and GDF-15 and a concomitant reduction in PTGS1 expression. Our findings show that metformin may affect the viability and proliferative capacity of TNBC by inducing an antiproliferative gene signature, and that metformin may be effective in the treatment/prevention of TNBC.


Assuntos
Neoplasias da Mama/genética , Senescência Celular/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica , Metformina/farmacologia , Técnicas de Cultura de Células , Feminino , Humanos
18.
Cell ; 151(5): 1042-54, 2012 Nov 21.
Artigo em Inglês | MEDLINE | ID: mdl-23178123

RESUMO

The conserved transcriptional regulator heat shock factor 1 (Hsf1) is a key sensor of proteotoxic and other stress in the eukaryotic cytosol. We surveyed Hsf1 activity in a genome-wide loss-of-function library in Saccaromyces cerevisiae as well as ~78,000 double mutants and found Hsf1 activity to be modulated by highly diverse stresses. These included disruption of a ribosome-bound complex we named the Ribosome Quality Control Complex (RQC) comprising the Ltn1 E3 ubiquitin ligase, two highly conserved but poorly characterized proteins (Tae2 and Rqc1), and Cdc48 and its cofactors. Electron microscopy and biochemical analyses revealed that the RQC forms a stable complex with 60S ribosomal subunits containing stalled polypeptides and triggers their degradation. A negative feedback loop regulates the RQC, and Hsf1 senses an RQC-mediated translation-stress signal distinctly from other stresses. Our work reveals the range of stresses Hsf1 monitors and elucidates a conserved cotranslational protein quality control mechanism.


Assuntos
Complexos Multiproteicos/metabolismo , Biossíntese de Proteínas , Ribossomos/metabolismo , Saccharomyces cerevisiae/metabolismo , Adenosina Trifosfatases/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Choque Térmico/genética , Peptídeos/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas de Ligação a RNA , Saccharomyces cerevisiae/citologia , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Estresse Fisiológico , Fatores de Transcrição/genética , Ubiquitina-Proteína Ligases/metabolismo , Proteína com Valosina
19.
Elife ; 1: e00048, 2012 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-23066505

RESUMO

The unfolded protein response (UPR) monitors the protein folding capacity of the endoplasmic reticulum (ER). In all organisms analyzed to date, the UPR drives transcriptional programs that allow cells to cope with ER stress. The non-conventional splicing of Hac1 (yeasts) and XBP1 (metazoans) mRNA, encoding orthologous UPR transcription activators, is conserved and dependent on Ire1, an ER membrane-resident kinase/endoribonuclease. We found that the fission yeast Schizosaccharomyces pombe lacks both a Hac1/XBP1 ortholog and a UPR-dependent-transcriptional-program. Instead, Ire1 initiates the selective decay of a subset of ER-localized-mRNAs that is required to survive ER stress. We identified Bip1 mRNA, encoding a major ER-chaperone, as the sole mRNA cleaved upon Ire1 activation that escapes decay. Instead, truncation of its 3' UTR, including loss of its polyA tail, stabilized Bip1 mRNA, resulting in increased Bip1 translation. Thus, S. pombe uses a universally conserved stress-sensing machinery in novel ways to maintain homeostasis in the ER.DOI:http://dx.doi.org/10.7554/eLife.00048.001.


Assuntos
Proteínas Fúngicas/genética , Regulação Fúngica da Expressão Gênica , Proteínas de Choque Térmico HSP70/genética , Proteínas Serina-Treonina Quinases/genética , RNA Mensageiro/genética , Schizosaccharomyces/genética , Resposta a Proteínas não Dobradas , Regiões 3' não Traduzidas , Retículo Endoplasmático/metabolismo , Proteínas Fúngicas/metabolismo , Proteínas de Choque Térmico HSP70/metabolismo , Homeostase/genética , Biossíntese de Proteínas , Dobramento de Proteína , Proteínas Serina-Treonina Quinases/metabolismo , Splicing de RNA , Estabilidade de RNA , RNA Mensageiro/metabolismo , Schizosaccharomyces/metabolismo , Transdução de Sinais , Transcrição Gênica
20.
Endocrinology ; 152(6): 2517-26, 2011 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-21505052

RESUMO

Phosphorylation of estrogen receptor α (ERα) is important for receptor function, although the role of specific ERα phosphorylation sites in ERα-mediated transcription remains to be fully evaluated. Transcriptional activation by ERα involves dynamic, coordinate interactions with coregulators at promoter enhancer elements to effect gene expression. To determine whether ERα phosphorylation affects recruitment of unique protein complexes at gene-specific promoters, changes in ERα Ser118 phosphorylation were assessed for effects on receptor and coregulator recruitment and transcription of ERα-regulated genes. Chromatin immunoprecipitation assays to measure promoter association found a 17ß-estradiol (E2)-dependent recruitment of ERα at 150 min to ERα-regulated promoters, whereas ERα phosphorylated at Ser118 was dissociated from promoters after E2 treatment. Mutation of Ser118 to alanine (S118A) altered unliganded and ligand-induced association of ERα and p160 coregulators with ERα target promoters when compared with wild-type (WT)-ERα transfection. S118A and WT-ERα exhibited a similar level of recruitment to the estrogen response element-driven pS2 promoter and induced pS2 mRNA after E2 treatment. Although WT-ERα was recruited to c-myc and cyclin D1 promoters after E2 treatment and induced mRNA expression, S118A exhibited reduced interaction with c-myc and cyclin D1 promoters, and E2 did not induce c-myc and cyclin D1 mRNA. In addition, S118A resulted in increased recruitment of steroid receptor coactivator-1, glucocorticoid receptor interacting protein-1, and activated in breast cancer-1 to pS2, c-myc, and cyclin D1 irrespective of the presence of E2. Together, these data indicate that site specific phosphorylation of ERα directs gene-specific recruitment of ERα and transcriptional coregulators to ERα target gene promoters.


Assuntos
Receptor alfa de Estrogênio/química , Receptor alfa de Estrogênio/metabolismo , Regiões Promotoras Genéticas , Serina/metabolismo , Ativação Transcricional , Motivos de Aminoácidos , Linhagem Celular , Ciclina D1/genética , Estradiol/metabolismo , Receptor alfa de Estrogênio/genética , Estrogênios/metabolismo , Humanos , Fosforilação , Ligação Proteica , Proteínas Proto-Oncogênicas c-myb/genética , Serina/genética , Especificidade da Espécie , Transcrição Gênica
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