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1.
Acad Med ; 98(11S): S14-S23, 2023 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-37556802

RESUMO

PURPOSE: Basic science medical educators (BSME) play a vital role in the training of medical students, yet little is known about the factors that shape their professional identities. This multi-institutional qualitative study investigated factors that support and threaten the professional identity formation (PIF) of these medical educators. METHOD: A qualitative descriptive study was conducted with a purposive sample of 58 BSME from 7 allopathic medical schools in the U.S. In-depth semi-structured interviews of individual BSME were conducted between December 2020 and February 2021 to explore the facilitators and barriers shaping the PIF of BSME. Thematic analysis was conducted. RESULTS: Factors shaping PIF were grouped into 3 broad domains: personal, social, and structural. Interrelated themes described a combination of factors that pushed BSME into teaching (early or positive teaching experiences) and kept them there (satisfaction and rewards of teaching, communities of like-minded people), as well as factors that challenged their PIF (misunderstanding from medical students, clinical, and research faculty, lack of formal training programs, and lack of tenure-track educator positions). The structural environment was reported to be crucial for PIF and determined whether BSME felt that they belonged and were valued. CONCLUSIONS: This study shows that although most BSME derive a sense of fulfillment and meaning from their role as medical educators, they face considerable obstacles during their PIF. Structural change and support are needed to increase recognition, value, promotion, and belonging for BSME to improve the satisfaction and retention of this important group of faculty.


Assuntos
Educação de Graduação em Medicina , Educação Médica , Humanos , Identificação Social , Docentes , Pesquisa Qualitativa
2.
Pharmacol Res Perspect ; 9(3): e00762, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33974344

RESUMO

Expectations for physicians are rapidly changing, as is the environment in which they will practice. In response, preclerkship medical education curricula are adapting to meet these demands, often by reducing the time for foundational sciences. This descriptive study compares preclerkship pharmacology education curricular practices from seven allopathic medical schools across the United States. We compare factors and practices that affect how pharmacology is integrated into the undergraduate medical education curriculum, including teaching techniques, resources, time allocated to pharmacology teaching, and assessment strategies. We use data from seven medical schools in the United States, along with results from a literature survey, to inform the strengths and weaknesses of various approaches and to raise important questions that can guide future research regarding integration of foundational sciences in medical school and health professions' curricula. In this comparative study, we found that there is significant heterogeneity in the number of hours dedicated to pharmacology in the preclerkship curriculum, whereas there was concordance in the use of active learning pedagogies for content delivery. Applying the ICAP (Interactive, Constructive, Active, Passive) Framework for cognitive engagement, our data showed that pharmacology was presented using more highly engaging pedagogies during sessions that are integrated with other foundational sciences. These findings can serve as a model that can be applied beyond pharmacology to other foundational sciences such as genetics, pathology, microbiology, biochemistry, etc.


Assuntos
Currículo , Educação de Graduação em Medicina , Farmacologia Clínica/educação , Faculdades de Medicina , Estados Unidos
3.
Pharmacol Res Perspect ; 9(3): e00773, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33974347

RESUMO

A grounded knowledge of pharmacology is essential for healthcare providers to improve the quality of patients' lives, avoid medical errors, and circumvent potentially dangerous drug-drug interactions. One of the greatest tools to achieve this foundational knowledge of pharmacology is the dedicated pharmacology educators who teach in health sciences programs. Too often, the pharmacology educators responsible for teaching this material are left siloed at their own institutions with little room for dialog and collaboration. As scientists, we know that it is through dialog and collaboration that ideas grow, are refined, and improve. More collaborative work is needed to identify and describe best practices for pharmacology education in health sciences programs. While evidence-based, outcomes-focused studies are the optimum standard for this work, there is also a place for descriptive studies and innovative reports.


Assuntos
Pessoal de Saúde/educação , Práticas Interdisciplinares , Farmacologia Clínica/educação , Currículo , Humanos
4.
Nitric Oxide ; 104-105: 36-43, 2020 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-32891753

RESUMO

It is well established that myoglobin supports mitochondrial respiration through the storage and transport of oxygen as well as through the scavenging of nitric oxide. However, during ischemia/reperfusion (I/R), myoglobin and mitochondria both propagate myocardial injury through the production of oxidants. Nitrite, an endogenous signaling molecule and dietary constituent, mediates potent cardioprotection after I/R and this effect relies on its interaction with both myoglobin and mitochondria. While independent mechanistic studies have demonstrated that nitrite-mediated cardioprotection requires the presence of myoglobin and the post-translational S-nitrosation of critical cysteine residues on mitochondrial complex I, it is unclear whether myoglobin directly catalyzes the S-nitrosation of complex I or whether mitochondrial-dependent nitrite reductase activity contributes to S-nitrosation. Herein, using purified myoglobin and isolated mitochondria, we characterize and directly compare the nitrite reductase activities of mitochondria and myoglobin and assess their contribution to mitochondrial S-nitrosation. We demonstrate that myoglobin is a significantly more efficient nitrite reductase than isolated mitochondria. Further, deoxygenated myoglobin catalyzes the nitrite-dependent S-nitrosation of mitochondrial proteins. This reaction is enhanced in the presence of oxidized (Fe3+) myoglobin and not significantly affected by inhibitors of mitochondrial respiration. Using a Chinese Hamster Ovary cell model stably transfected with human myoglobin, we show that both myoglobin and mitochondrial complex I expression are required for nitrite-dependent attenuation of cell death after anoxia/reoxygenation. These data expand the understanding of myoglobin's role both as a nitrite reductase to a mediator of S-nitrosation and as a regulator of mitochondrial function, and have implications for nitrite-mediated cardioprotection after I/R.


Assuntos
Citoproteção/fisiologia , Mitocôndrias/metabolismo , Mioglobina/metabolismo , Nitrito Redutases/metabolismo , Nitritos/metabolismo , Animais , Células CHO , Hipóxia Celular/fisiologia , Cricetulus , Cisteína/química , Complexo I de Transporte de Elétrons/química , Complexo I de Transporte de Elétrons/metabolismo , Humanos , Proteínas Mitocondriais/química , Proteínas Mitocondriais/metabolismo , Nitrosação
5.
Oncogene ; 39(19): 3791-3802, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32203163

RESUMO

Cyclic nucleotide phosphodiesterases (PDE) break down cyclic nucleotides such as cAMP and cGMP, reducing the signaling of these important intracellular second messengers. Several unique families of phosphodiesterases exist, and certain families are clinically important modulators of vasodilation. In the current work, we have summarized the body of literature that describes an emerging role for the PDE4 subfamily of phosphodiesterases in malignancy. We have systematically investigated PDE4A, PDE4B, PDE4C, and PDE4D isoforms and found evidence associating them with several cancer types including hematologic malignancies and lung cancers, among others. In this review, we compare the evidence examining the functional role of each PDE4 subtype across malignancies, looking for common signaling themes, signaling pathways, and establishing the case for PDE4 subtypes as a potential therapeutic target for cancer treatment.


Assuntos
Neoplasias Hematológicas/genética , Neoplasias Pulmonares/genética , Isoformas de Proteínas/genética , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/genética , Neoplasias Hematológicas/classificação , Neoplasias Hematológicas/patologia , Humanos , Neoplasias Pulmonares/classificação , Neoplasias Pulmonares/patologia , Isoformas de Proteínas/classificação , Transdução de Sinais/genética
6.
Med Sci Educ ; 30(1): 111-115, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-34457648

RESUMO

Facebook is the social media platform used most by both medical educators and students, but there is scant literature investigating effective ways to use the platform across multiple institutions. This multi-institutional study compared student engagement between an official curricular Facebook page and a supplemental Facebook page. While a greater proportion of students were reached by the official course page, greater post engagement was achieved with the supplemental page. We attribute this to the abundance of question-type posts on the supplemental page and show that question-type posts achieve higher engagement with students than statement-type posts, regardless of page type.

7.
J Biol Chem ; 294(18): 7269-7282, 2019 05 03.
Artigo em Inglês | MEDLINE | ID: mdl-30872402

RESUMO

Myoglobin is a monomeric heme protein expressed ubiquitously in skeletal and cardiac muscle and is traditionally considered to function as an oxygen reservoir for mitochondria during hypoxia. It is now well established that low concentrations of myoglobin are aberrantly expressed in a significant proportion of breast cancer tumors. Despite being expressed only at low levels in these tumors, myoglobin is associated with attenuated tumor growth and a better prognosis in breast cancer patients, but the mechanism of this myoglobin-mediated protection against further cancer growth remains unclear. Herein, we report a signaling pathway by which myoglobin regulates mitochondrial dynamics and thereby decreases cell proliferation. We demonstrate in vitro that expression of human myoglobin in MDA-MB-231, MDA-MB-468, and MCF7 breast cancer cells induces mitochondrial hyperfusion by up-regulating mitofusins 1 and 2, the predominant catalysts of mitochondrial fusion. This hyperfusion causes cell cycle arrest and subsequent inhibition of cell proliferation. Mechanistically, increased mitofusin expression was due to myoglobin-dependent free-radical production, leading to the oxidation and degradation of the E3 ubiquitin ligase parkin. We recapitulated this pathway in a murine model in which myoglobin-expressing xenografts exhibited decreased tumor volume with increased mitofusin, markers of cell cycle arrest, and decreased parkin expression. Furthermore, in human triple-negative breast tumor tissues, mitofusin and myoglobin levels were positively correlated. Collectively, these results elucidate a new function for myoglobin as a modulator of mitochondrial dynamics and reveal a novel pathway by which myoglobin decreases breast cancer cell proliferation and tumor growth by up-regulating mitofusin levels.


Assuntos
Neoplasias da Mama/patologia , Proliferação de Células/fisiologia , Dinâmica Mitocondrial/fisiologia , Mioglobina/fisiologia , Animais , Linhagem Celular Tumoral , Feminino , Fase G1/fisiologia , GTP Fosfo-Hidrolases/metabolismo , Xenoenxertos , Humanos , Camundongos , Proteínas de Transporte da Membrana Mitocondrial/metabolismo , Oxirredução , Fase S/fisiologia , Ubiquitina-Proteína Ligases/metabolismo
8.
Curr Pharm Teach Learn ; 10(8): 1062-1069, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-30314542

RESUMO

BACKGROUND: Traditional interprofessional educational (IPE) exercises are those where learning exists "about, from, and with" trainees in two or more professions in order to prepare health sciences professionals to work on interprofessional teams. One emerging difficulty with IPE is the paucity of health profession students at single institutions, and the geographic and financial constraints of multi-institutional collaboration. INTERPROFESSIONAL EDUCATION ACTIVITY: To circumvent these barriers, we developed a multi-institution telehealth team-based learning (TBL) event between medical and pharmacy students on the topic of pharmacogenomics (PGx). Using a validated pre-post survey design, student attitudes and perceptions were measured before and after an educational intervention designed to simulate interprofessional telehealth collaboration. The survey results showed significant improvement across all areas of student attitudes toward interprofessional collaboration. Also, medical student PGx confidence increased substantially during the exercise even though the only PGx instruction they received was from pharmacy students. DISCUSSION: These data demonstrate that learning exists "about, from, and with" trainees in other professions, even if they do not physically train in the same location. Free tools are available to create virtual interactions between students on different campuses, and telehealth exercises using these tools are a valid way to conduct IPE across different campuses. The instructional experience does not need to be identical for all participants in the IPE event; rather, tailoring the educational experience to each group of students provides opportunities for inter-student teaching.


Assuntos
Educação a Distância/métodos , Relações Interprofissionais , Farmacogenética/educação , Telemedicina/métodos , Humanos , Michigan , Aprendizagem Baseada em Problemas/métodos , Aprendizagem Baseada em Problemas/tendências , Estudantes de Medicina/psicologia , Estudantes de Medicina/estatística & dados numéricos , Estudantes de Farmácia/psicologia , Estudantes de Farmácia/estatística & dados numéricos , Ensino , Interface Usuário-Computador
9.
EXCLI J ; 16: 483-496, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28694752

RESUMO

Ischemia reperfusion (IR) injury is a significant cause of morbidity and mortality in liver transplantation. When oxygen is reintroduced to the liver graft it initiates a cascade of molecular reactions leading to the release of reactive oxygen species (ROS) and pro-inflammatory cytokines. These soluble mediators propagate a sterile immune response to cause significant tissue damage. Ischemic preconditioning (IPC) is one method that reduces hepatocellular injury by altering the immune response and inhibiting the production of ROS. Studies quantifying the effects of IPC in humans have demonstrated an improved liver enzyme panel in patients receiving grafts pretreated with IPC as compared to patients receiving the standard of care. In our review, we explore current literature in the field in order to describe the mechanism through which IPC regulates the production of ROS and improves IR injury.

10.
Acad Forensic Pathol ; 7(1): 99-111, 2017 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31239962

RESUMO

From 2000 to 2014, drug overdose deaths increased 137% in the United States, and 61% of these deaths included some form of opiate. The vast majority of opiate-related drug fatalities include multiple drugs, although there is scant data quantitatively describing the exact drugs that contribute to deaths due to multiple drugs. In the present study, we sought to quantitatively identify the drugs that occur with opiates in accidental multidrug-related fatalities. We retrospectively explored fatal drug trends in four Michigan counties, with a focus on profiling drugs present concurrently with opiates. Blood and urine toxicology reports for mixed drug fatalities (N=180) were analyzed using frequent item analysis approaches to identify common analyte trends in opiate-related fatalities. Within our cohort, the most prevalent serum analytes included caffeine (n=147), morphine (n=90), alprazolam (n=69), gabapentin (n=46), and tetrahydrocannabinol (n=44). In 100% of cases where gabapentin was present (n=46), an opiate was also present in the serum or urine. The average gabapentin serum concentration was 13.56 µg/mL (SEM =0.33 µg/mL), with a range of 0.5-88.7 µg/mL. Gabapentin was found at very high frequency in accidental mixed drug fatalities. Gabapentin concentrations were generally within the normal therapeutic range (2-20 µg/mL). It is unknown whether a synergistic effect with opioids may contribute to central respiratory depression. Further research is warranted to determine any contributory role of gabapentin in these deaths. Confirmed interactions could have broad implications for future reporting by forensic pathologists as well as prescribing practices by clinicians.

11.
Cell Rep ; 15(8): 1700-14, 2016 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-27184852

RESUMO

Inhalation of environmental antigens such as allergens does not always induce inflammation in the respiratory tract. While antigen-presenting cells (APCs), including dendritic cells and macrophages, take up inhaled antigens, the cell-intrinsic molecular mechanisms that prevent an inflammatory response during this process, such as activation of the transcription factor NF-κB, are not well understood. Here, we show that the nuclear receptor PPARγ plays a critical role in blocking NF-κB activation in response to inhaled antigens to preserve immune tolerance. Tolerance induction promoted mitochondrial respiration, generation of H2O2, and suppression of NF-κB activation in WT, but not PPARγ-deficient, APCs. Forced restoration of H2O2 in PPARγ-deficient cells suppressed IκBα degradation and NF-κB activation. Conversely, scavenging reactive oxygen species from mitochondria promoted IκBα degradation with loss of regulatory and promotion of inflammatory T cell responses in vivo. Thus, communication between PPARγ and the mitochondria maintains immune quiescence in the airways.


Assuntos
Células Apresentadoras de Antígenos/imunologia , Peróxido de Hidrogênio/metabolismo , Pulmão/citologia , Mitocôndrias/metabolismo , NF-kappa B/metabolismo , Animais , Antígeno CD11c/metabolismo , Proliferação de Células , Citocinas/genética , Células Dendríticas/metabolismo , Ácidos Graxos/metabolismo , Regulação da Expressão Gênica , Tolerância Imunológica , Mediadores da Inflamação/metabolismo , Camundongos Endogâmicos C57BL , PPAR gama/deficiência , PPAR gama/metabolismo , Regiões Promotoras Genéticas/genética , Espécies Reativas de Oxigênio/metabolismo , Linfócitos T/citologia
12.
J Invest Surg ; 28(5): 292-300, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26374984

RESUMO

Liver ischemia reperfusion injury is mediated by a complex system of signaling cascades and inflammatory response resulting in organ damage. Selectins are a group of cell adhesion glycoproteins that play a key role in the initial immunological response. L-selectins, found on leukocytes, initiate the original adhesion and rolling phase of leukocyte extravasation upon liver sinusoidal endothelial cells (LSECs). P-selectins, found on platelets and tissue-specific endothelial cells, further increases leukocyte-endothelial adhesion and rolling. P-selectin-ligand binding also initiates intracellular signals that produce adhesion molecules to start firm adhesion and increase local chemokine production. L-selectin-ligand binding on the leukocytes increases adhesion molecule expression and chemokines, but also initiate changes in intracellular structural actin. E-selectin expression occurs with the presence of TNF-α and/or IL-1ß. E-selectin-ligand binding decreases leukocyte rolling velocity and increases adhesion molecules. Together, these glycoproteins transition the leukocyte response from original margination and rolling to firm adhesion and eventually migration.


Assuntos
Hepatopatias/metabolismo , Traumatismo por Reperfusão/metabolismo , Selectinas/metabolismo , Animais , Humanos , Hepatopatias/prevenção & controle , Traumatismo por Reperfusão/prevenção & controle
13.
Arch Toxicol ; 89(5): 651-7, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25566829

RESUMO

Ischemia/reperfusion (IR) injury occurs when oxygen is rapidly reintroduced into ischemic tissue, resulting in cell death and necrotic tissue damage. This is a major concern during liver transplantation procedures since there is an inevitable interruption and subsequent restoration of circulation. IR injury in liver tissue is initiated through reactive oxygen species (ROS), which are generated by hepatocytes during IR insult. Although these ROS are thought to play a protective roll since they are known to activate several pathways involved in the hypoxic response, they also trigger a localized sterile immune response that results in the recruitment of Kupffer cells and neutrophils to the site of IR insult. These immune cells generate larger quantities of ROS that trigger apoptosis and oncotic necrosis in liver tissue. In this review, we will summarize what is currently known about the response of liver tissue to IR insult at the molecular level.


Assuntos
Isquemia/metabolismo , Fígado/irrigação sanguínea , Traumatismo por Reperfusão/metabolismo , Humanos , Isquemia/etiologia , Células de Kupffer/fisiologia , Fígado/metabolismo , Fígado/patologia , Transplante de Fígado/efeitos adversos , Necrose , Espécies Reativas de Oxigênio/metabolismo , Traumatismo por Reperfusão/etiologia
14.
Free Radic Biol Med ; 70: 45-53, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24556414

RESUMO

Though it is well accepted that adipose tissue is central in the regulation of glycemic homeostasis, the molecular mechanisms governing adipocyte glucose uptake remain unclear. Recent studies demonstrate that mitochondrial dynamics (fission and fusion) regulate lipid accumulation and differentiation in adipocytes. However, the role of mitochondrial dynamics in glucose homeostasis has not been explored. The nitric oxide oxidation products nitrite and nitrate are endogenous signaling molecules and dietary constituents that have recently been shown to modulate glucose metabolism, prevent weight gain, and reverse the development of metabolic syndrome in mice. Although the mechanism of this protection is unclear, the mitochondrion is a known subcellular target for nitrite signaling. Thus, we hypothesize that nitrite modulates mitochondrial dynamics and function to regulate glucose uptake in adipocytes. Herein, we demonstrate that nitrite significantly increases glucose uptake in differentiated murine adipocytes through a mechanism dependent on mitochondrial fusion. Specifically, nitrite promotes mitochondrial fusion by increasing the profusion protein mitofusin 1 while concomitantly activating protein kinase A (PKA), which phosphorylates and inhibits the profission protein dynamin-related protein 1 (Drp1). Functionally, this signaling augments cellular respiration, fatty acid oxidation, mitochondrial oxidant production, and glucose uptake. Importantly, inhibition of PKA or Drp1 significantly attenuates nitrite-induced mitochondrial respiration and glucose uptake. These findings demonstrate that mitochondria play an essential metabolic role in adipocytes, show a novel role for both nitrite and mitochondrial fusion in regulating adipocyte glucose homeostasis, and have implications for the potential therapeutic use of nitrite and mitochondrial modulators in glycemic regulation.


Assuntos
Adipócitos/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Glucose/metabolismo , Nitritos/metabolismo , Células 3T3-L1 , Animais , Diferenciação Celular/genética , Respiração Celular/genética , Dinaminas/metabolismo , Radicais Livres/metabolismo , Metabolismo dos Lipídeos , Camundongos , Mitocôndrias/metabolismo , Dinâmica Mitocondrial/genética , Fosforilação
15.
Cardiovasc Res ; 101(1): 57-68, 2014 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-24081164

RESUMO

AIMS: Nitrite (NO2(-)), a dietary constituent and nitric oxide (NO) oxidation product, mediates cardioprotection after ischaemia/reperfusion (I/R) in a number of animal models when administered during ischaemia or as a pre-conditioning agent hours to days prior to the ischaemic episode. When present during ischaemia, the reduction of nitrite to bioactive NO by deoxygenated haem proteins accounts for its protective effects. However, the mechanism of nitrite-induced pre-conditioning, a normoxic response which does not appear to require reduction of nitrite to NO, remains unexplored. METHODS AND RESULTS: Using a model of hypoxia/reoxygenation (H/R) in cultured rat H9c2 cardiomyocytes, we demonstrate that a transient (30 min) normoxic nitrite treatment significantly attenuates cell death after a hypoxic episode initiated 1 h later. Mechanistically, this protection depends on the activation of protein kinase A, which phosphorylates and inhibits dynamin-related protein 1, the predominant regulator of mitochondrial fission. This results morphologically, in the promotion of mitochondrial fusion and functionally in the augmentation of mitochondrial membrane potential and superoxide production. We identify AMP kinase (AMPK) as a downstream target of the mitochondrial reactive oxygen species (ROS) generated and show that its oxidation and subsequent phosphorylation are essential for cytoprotection, as scavenging of ROS prevents AMPK activation and inhibits nitrite-mediated protection after H/R. The protein kinase A-dependent protection mediated by nitrite is reproduced in an intact isolated rat heart model of I/R. CONCLUSIONS: These data are the first to demonstrate nitrite-dependent normoxic modulation of both mitochondrial morphology and function and reveal a novel signalling pathway responsible for nitrite-mediated cardioprotection.


Assuntos
Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Dinaminas/metabolismo , Precondicionamento Isquêmico Miocárdico , Dinâmica Mitocondrial , Nitritos/metabolismo , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Linhagem Celular , Citoproteção , Hipóxia/metabolismo , Masculino , Fosforilação , Ratos , Ratos Sprague-Dawley , Superóxidos/metabolismo
16.
Clin Cancer Res ; 19(2): 380-92, 2013 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-23213056

RESUMO

PURPOSE: Strategies to inhibit the EGF receptor (EGFR) using the tyrosine kinase inhibitor erlotinib have been associated with limited clinical efficacy in head and neck squamous cell carcinoma (HNSCC). Co-activation of alternative kinases may contribute to erlotinib resistance. EXPERIMENTAL DESIGN: We generated HNSCC cells expressing dominant-active c-Src (DA-Src) to determine the contribution of c-Src activation to erlotinib response. RESULTS: Expression of DA-Src conferred resistance to erlotinib in vitro and in vivo compared with vector-transfected control cells. Phospho-Met was strongly upregulated by DA-Src, and DA-Src cells did not produce hepatocyte growth factor (HGF). Knockdown of c-Met enhanced sensitivity to erlotinib in DA-Src cells in vitro, as did combining a c-Met or c-Src inhibitor with erlotinib. Inhibiting EGFR resulted in minimal reduction of phospho-Met in DA-Src cells, whereas complete phospho-Met inhibition was achieved by inhibiting c-Src. A c-Met inhibitor significantly sensitized DA-Src tumors to erlotinib in vivo, resulting in reduced Ki67 labeling and increased apoptosis. In parental cells, knockdown of endogenous c-Src enhanced sensitivity to erlotinib, whereas treatment with HGF to directly induce phospho-Met resulted in erlotinib resistance. The level of endogenous phospho-c-Src in HNSCC cell lines was also significantly correlated with erlotinib resistance. CONCLUSIONS: Ligand-independent activation of c-Met contributes specifically to erlotinib resistance, not cetuximab resistance, in HNSCC with activated c-Src, where c-Met activation is more dependent on c-Src than on EGFR, providing an alternate survival pathway. Addition of a c-Met or c-Src inhibitor to erlotinib may increase efficacy of EGFR inhibition in patients with activated c-Src.


Assuntos
Neoplasias de Cabeça e Pescoço/genética , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-met/metabolismo , Proteínas Proto-Oncogênicas pp60(c-src)/genética , Quinazolinas/farmacologia , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/genética , Ativação Enzimática , Cloridrato de Erlotinib , Expressão Gênica , Técnicas de Silenciamento de Genes , Neoplasias de Cabeça e Pescoço/tratamento farmacológico , Neoplasias de Cabeça e Pescoço/metabolismo , Humanos , Fosforilação , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Proto-Oncogênicas c-met/genética , Quinazolinas/uso terapêutico , Ativação Transcricional
17.
Cancer Biol Ther ; 13(10): 935-45, 2012 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-22785204

RESUMO

The epidermal growth factor receptor (EGFR) is widely expressed in head and neck squamous cell carcinomas (HNSCC) and can activate many growth and survival pathways within tumor cells. Despite ubiquitous EGFR expression, therapies targeting the receptor are only modestly effective in the treatment of HNSCC. A consistent mechanism of resistance to EGFR targeting agents has not yet been identified in HNSCC likely due, in part, to the paucity of preclinical models. We assessed the in vitro and in vivo responses of a panel of 10 genotypically validated HNSCC cell lines to the EGFR inhibitors erlotinib and cetuximab to determine their validity as models of resistance to these agents. We defined a narrow range of response to erlotinib in HNSCC cells in vitro and found a positive correlation between EGFR protein expression and erlotinib response. We observed cross-sensitivity in one HNSCC cell line, 686LN, between erlotinib and cetuximab in vivo. We attempted to generate models of cetuximab resistance in HNSCC cell line-derived xenografts and heterotopic tumorgrafts generated directly from primary patient tumors. While all 10 HNSCC cell line xenografts tested were sensitive to cetuximab in vivo, heterotopic patient tumorgrafts varied in response to cetuximab indicating that these models may be more representative of clinical responses. These studies demonstrate the limitations of using HNSCC cell lines to reflect the heterogeneous clinical responses to erlotinib and cetuximab, and suggest that different approaches including heterotopic tumorgrafts may prove more valuable to elucidate mechanisms of clinical resistance to EGFR inhibitors in HNSCC.


Assuntos
Anticorpos Monoclonais/farmacologia , Antineoplásicos/farmacologia , Carcinoma de Células Escamosas/tratamento farmacológico , Receptores ErbB/antagonistas & inibidores , Neoplasias de Cabeça e Pescoço/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Quinazolinas/farmacologia , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais Humanizados , Antineoplásicos/administração & dosagem , Carcinoma de Células Escamosas/metabolismo , Linhagem Celular Tumoral , Cetuximab , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Receptores ErbB/metabolismo , Cloridrato de Erlotinib , Neoplasias de Cabeça e Pescoço/metabolismo , Humanos , Concentração Inibidora 50 , Camundongos , Camundongos Nus , Inibidores de Proteínas Quinases/administração & dosagem , Quinazolinas/administração & dosagem , Carcinoma de Células Escamosas de Cabeça e Pescoço , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
18.
Clin Cancer Res ; 17(18): 5935-44, 2011 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-21791633

RESUMO

PURPOSE: Acquired resistance to cetuximab, a chimeric epidermal growth factor receptor (EGFR)-targeting monoclonal antibody, is a widespread problem in the treatment of solid tumors. The paucity of preclinical models has limited investigations to determine the mechanism of acquired therapeutic resistance, thereby limiting the development of effective treatments. The purpose of this study was to generate cetuximab-resistant tumors in vivo to characterize mechanisms of acquired resistance. EXPERIMENTAL DESIGN: We generated cetuximab-resistant clones from a cetuximab-sensitive bladder cancer cell line in vivo by exposing cetuximab-sensitive xenografts to increasing concentrations of cetuximab, followed by validation of the resistant phenotype in vivo and in vitro using invasion assays. A candidate-based approach was used to examine the role of HER2 on mediating cetuximab resistance both in vitro and in vivo. RESULTS: We generated a novel model of cetuximab resistance, and, for the first time in the context of EGFR-inhibitor resistance, we identified increased phosphorylation of a C-terminal fragment of HER2 (611-CTF) in cetuximab-resistant cells. Afatinib (BIBW-2992), an irreversible kinase inhibitor targeting EGFR and HER2, successfully inhibited growth of the cetuximab-resistant cells in vitro. When afatinib was combined with cetuximab in vivo, we observed an additive growth inhibitory effect in cetuximab-resistant xenografts. CONCLUSIONS: These data suggest that the use of dual EGFR-HER2 kinase inhibitors can enhance responses to cetuximab, perhaps in part due to downregulation of 611-CTF. This study conducted in a novel in vivo model provides a mechanistic rationale for ongoing phase I clinical trials using this combination treatment modality.


Assuntos
Anticorpos Monoclonais/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Receptores ErbB/antagonistas & inibidores , Inibidores de Proteínas Quinases/uso terapêutico , Receptor ErbB-2/antagonistas & inibidores , Animais , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais Humanizados , Linhagem Celular Tumoral , Cetuximab , Modelos Animais de Doenças , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Sinergismo Farmacológico , Inativação Gênica , Humanos , Camundongos , Camundongos Nus , Neoplasias/tratamento farmacológico , Neoplasias/genética , Neoplasias/metabolismo , Fosforilação , Inibidores de Proteínas Quinases/farmacologia , RNA Interferente Pequeno/genética , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
19.
J Proteome Res ; 9(6): 3073-82, 2010 Jun 04.
Artigo em Inglês | MEDLINE | ID: mdl-20426488

RESUMO

Epidermal growth factor receptor (EGFR) is overexpressed in up to 90% of head and neck cancer (HNC), where increased expression levels of EGFR correlate with poor prognosis. To date, EGFR expression levels have not predicted the clinical response to the EGFR-targeting therapies. Elucidation of the molecular mechanisms underlying anti-EGFR-induced antitumor effects may shed some light on the mechanisms of HNC resistance to EGFR-targeting therapeutics and provide novel targets for improving the treatment of HNC. Here, we conducted a quantitative proteomics analysis to determine the molecular networks regulated by EGFR levels in HNC by specifically knocking-down EGFR and employing stable isotope labeling with amino acids in cell culture (SILAC). Following data normalization to minimize systematic errors and Western blotting validation, 12 proteins (e.g., p21, stratifin, and maspin) and 24 proteins (e.g., cdc2 and MTA2) were found to be significantly upregulated or downregulated by EGFR knockdown, respectively. Bioinformatic analysis revealed that these proteins were mainly involved in long-chain fatty acid biosynthesis and beta-oxidation, cholesterol biosynthesis, cell proliferation, DNA replication, and apoptosis. Cell cycle analysis confirmed that G(2)/M phase progression was significantly inhibited by EGFR knockdown, a hypothesis generated from network modeling. Further investigation of these molecular networks may not only enhance our understanding of the antitumor mechanisms of EGFR targeting but also improve patient selection and provide novel targets for better therapeutics.


Assuntos
Receptores ErbB/genética , Receptores ErbB/metabolismo , Neoplasias de Cabeça e Pescoço/genética , Neoplasias de Cabeça e Pescoço/metabolismo , Proteômica/métodos , Western Blotting , Proteínas de Ciclo Celular/química , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Colesterol/metabolismo , Ácidos Graxos/metabolismo , Técnicas de Silenciamento de Genes , Neoplasias de Cabeça e Pescoço/enzimologia , Humanos , Marcação por Isótopo , Redes e Vias Metabólicas , Proteoma/metabolismo , RNA Interferente Pequeno/genética , Reprodutibilidade dos Testes , Transdução de Sinais
20.
Clin Cancer Res ; 15(11): 3740-50, 2009 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-19470725

RESUMO

PURPOSE: We determined hepatocyte growth factor (HGF) and c-Met expression and signaling in human head and neck squamous cell carcinoma (HNSCC) cells and primary tissues and tested the ability of c-Met tyrosine kinase inhibitors (TKI) to block HGF-induced biological signaling. EXPERIMENTAL DESIGN: Expression and signaling were determined using immunoblotting, ELISA, and immunohistochemistry. Biological end points included wound healing, cell proliferation, and invasion. c-Met TKIs were tested for their ability to block HGF-induced signaling and biological effects in vitro and in xenografts established in nude mice. RESULTS: c-Met was expressed and functional in HNSCC cells. HGF was secreted by HNSCC tumor-derived fibroblasts, but not by HNSCC cells. Activation of c-Met promoted phosphorylation of AKT and mitogen-activated protein kinase as well as release of the inflammatory cytokine interleukin-8. Cell growth and wound healing were also stimulated by HGF. c-Met TKIs blocked HGF-induced signaling, interleukin-8 release, and wound healing. Enhanced invasion of HNSCC cells induced by the presence of tumor-derived fibroblasts was completely blocked with a HGF-neutralizing antibody. PF-2341066, a c-Met TKI, caused a 50% inhibition of HNSCC tumor growth in vivo with decreased proliferation and increased apoptosis within the tumors. In HNSCC tumor tissues, both HGF and c-Met protein were increased compared with expression in normal mucosa. CONCLUSIONS: These results show that HGF acts mainly as a paracrine factor in HNSCC cells, the HGF/c-Met pathway is frequently up-regulated and functional in HNSCC, and a clinically relevant c-Met TKI shows antitumor activity in vivo. Blocking the HGF/c-Met pathway may be clinically useful for the treatment of HNSCC.


Assuntos
Carcinoma de Células Escamosas/metabolismo , Neoplasias de Cabeça e Pescoço/metabolismo , Fator de Crescimento de Hepatócito/metabolismo , Proteínas Proto-Oncogênicas c-met/metabolismo , Transdução de Sinais/fisiologia , Animais , Western Blotting , Carcinoma de Células Escamosas/patologia , Carcinoma de Células Escamosas/fisiopatologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Crizotinibe , Relação Dose-Resposta a Droga , Neoplasias de Cabeça e Pescoço/patologia , Neoplasias de Cabeça e Pescoço/fisiopatologia , Fator de Crescimento de Hepatócito/farmacologia , Humanos , Imuno-Histoquímica , Indóis/farmacologia , Camundongos , Camundongos Nus , Transplante de Neoplasias , Comunicação Parácrina/fisiologia , Piperazinas/farmacologia , Piperidinas/farmacologia , Proteínas Proto-Oncogênicas c-met/antagonistas & inibidores , Pirazóis , Piridinas/farmacologia , Transdução de Sinais/efeitos dos fármacos , Estresse Mecânico , Sulfonamidas/farmacologia , Transplante Heterólogo , Carga Tumoral/efeitos dos fármacos
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