RESUMO
BACKGROUND: Pathologists commonly employ the Ki67 immunohistochemistry labelling index (LI) when deciding appropriate therapeutic strategies for patients with breast cancer. However, despite several attempts at standardizing the Ki67 LI, inter-observer and inter-laboratory bias remain problematic. We developed a flow cytometric assay that employed tissue dissociation, enzymatic treatment and a gating process to analyse Ki67 in formalin-fixed paraffin-embedded (FFPE) breast cancer tissue. RESULTS: We demonstrated that mechanical homogenizations combined with thrombin treatment can be used to recover efficiently intact single-cell nuclei from FFPE breast cancer tissue. Ki67 in the recovered cell nuclei retained reactivity against the MIB-1 antibody, which has been widely used in clinical settings. Additionally, since the method did not alter the nucleoskeletal structure of tissues, the nuclei of cancer cells can be enriched in data analysis based on differences in size and complexity of nuclei of lymphocytes and normal mammary cells. In a clinical study using the developed protocol, Ki67 positivity was correlated with the Ki67 LI obtained by hot spot analysis by a pathologist in Japan (rho = 0.756, P < 0.0001). The number of cancer cell nuclei subjected to the analysis in our assay was more than twice the number routinely checked by pathologists in clinical settings. CONCLUSIONS: The findings of this study showed the application of this new flow cytometry method could potentially be used to standardize Ki67 assessments in breast cancer.
Assuntos
Neoplasias da Mama , Citometria de Fluxo , Antígeno Ki-67 , Inclusão em Parafina , Antígeno Ki-67/metabolismo , Antígeno Ki-67/análise , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Humanos , Citometria de Fluxo/métodos , Feminino , Inclusão em Parafina/métodos , Formaldeído , Fixação de Tecidos/métodosRESUMO
OBJECTIVES: Although antibody measurements using lateral flow assay (LFA) kits are convenient, they usually require a specialized reader for quantification. However, a smartphone-based quantification application can be used as a reader for LFA kits. We investigated the quantification ability of the application for SARS-CoV-2-specific antibodies. METHODS: Eight hundred frozen serum samples from 100 healthcare professionals who received a COVID-19 vaccine were analyzed. Images of assayed LFA kits were obtained using a smartphone camera. We determined whether the ratio of color density of the test and control lines of spike protein IgG correlated with chemiluminescent immunoassay-measured titers. RESULTS: Spike protein IgG correlated well with the quantification results of the LFA kits using the application installed on a smartphone (r = 0.886). CONCLUSION: Our results suggest that smartphone-based quantitative analysis of LFA kits enables the quantification of anti-SARS-CoV-2 IgG without special devices, enabling point-of-care assessment of acquired humoral immunity in various settings.
Assuntos
COVID-19 , Smartphone , Humanos , COVID-19/diagnóstico , Vacinas contra COVID-19 , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus , Anticorpos Antivirais , Imunoglobulina GRESUMO
Bacterial aggregation by mixing with polymers is applied as pretreatment to identify pathogens in patients with infectious diseases. However, the detailed interaction between polymers and bacteria has yet to be fully understood. Here, we investigate the interaction between polyallylamine and Escherichia coli by isothermal titration calorimetry. Aggregation was observed at pH 10 and the binding was driven by favorable enthalpic gain such as the electrostatic interaction. Neither aggregation nor the apparent heat of binding was observed at pH 4.0, despite the strong positive charge of polyallylamine. These results suggest that intermolecular repulsive forces of the abundant positive charge of polyallylamine cause an increased loss of conformational entropy by binding. Non-electrostatic interaction plays a critical role for aggregation.
Assuntos
Escherichia coli , Poliaminas , Humanos , Calorimetria , PolímerosRESUMO
Hyper-activation of the MAPK and PI3K-AKT pathways is linked to tumour progression in triple-negative breast cancer (TNBC). However, clinically effective predictive markers for drugs targeted against protein kinases involved in these pathways have not been identified. We investigated the ability of MEK and PI3K catalytic activity to predict sensitivity to trametinib and wortmannin in TNBC. MEK and PI3K activities correlated strongly with each other only in cell lines showing wortmannin-specific sensitivity, as shown by a linear regression curve (R = 0.951). Accordingly, we created a new parameter that distinguishes trametinib and wortmannin sensitivity in vitro and in vivo. Our findings suggest that the catalytic activities of MEK and PI3K might predict the response of TNBC to trametinib and wortmannin.
Assuntos
Biocatálise , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Neoplasias de Mama Triplo Negativas/metabolismo , Androstadienos/farmacologia , Animais , Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Quinases de Proteína Quinase Ativadas por Mitógeno/antagonistas & inibidores , Inibidores de Fosfoinositídeo-3 Quinase , Piridonas/farmacologia , Pirimidinonas/farmacologia , Relação Estrutura-Atividade , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/patologia , Células Tumorais Cultivadas , WortmaninaRESUMO
BACKGROUND: Pathologically low-risk endometrial cancer patients do not receive postoperative treatment; however, 10-15% of these patients show recurrence with poor prognosis. We evaluated the clinical importance of cyclin-dependent kinase 4/6 (CDK4/6) activity, and its significance as a novel biomarker for the prognosis and chemo-sensitivity of endometrioid endometrial carcinoma (EEC). METHODS: Cyclin-dependent kinase 4/6 expression and enzyme activity in 109 tumour samples from patients with EEC were examined with a cell-cycle profiling (C2P) assay. CDK4/6-specific activity (CDK4/6SA) was determined, and its relationship with clinicopathological factors and expression of Ki-67 was analysed. RESULTS: CDK4/6-specific activity was significantly correlated with Ki-67 (P=0.035), but not with any other clinicopathological characteristics. CDK4/6SA was significantly higher (P=0.002) in pathologically low-risk patients (not receiving adjuvant chemotherapy, n=74) than in intermediate- or high-risk patients (receiving adjuvant chemotherapy, n=35). In addition, patients with high CDK4/6SA (>3.0) showed significantly (P=0.024) shorter progression-free survival (PFS) than those with low CDK4/6SA (<3.0). Although Ki-67 expression itself was not a marker for prognosis, the combination of high CDK4/6SA and high Ki-67 expression (>15%) was robustly associated with shorter PFS (P=0.015), and this combination was an independent poor prognostic factor in the low-risk group. Inversely, in the intermediate-/high-risk group, patients with high CDK4/6SA had a tendency of a more favourable prognosis compared with patients with low CDK4/6SA (P=0.063). CONCLUSIONS: CDK4/6-specific activity can be used as a biomarker to predict prognosis and, possibly, chemo-sensitivity. The combination of Ki-67 expression might strengthen the clinical usefulness of CDK4/6SA as a biomarker.
Assuntos
Biomarcadores Tumorais/metabolismo , Carcinoma Endometrioide/enzimologia , Quinase 4 Dependente de Ciclina/metabolismo , Quinase 6 Dependente de Ciclina/metabolismo , Neoplasias do Endométrio/enzimologia , Recidiva Local de Neoplasia/enzimologia , Adulto , Idoso , Carcinoma Endometrioide/tratamento farmacológico , Carcinoma Endometrioide/patologia , Quimioterapia Adjuvante , Neoplasias do Endométrio/tratamento farmacológico , Neoplasias do Endométrio/patologia , Feminino , Humanos , Antígeno Ki-67/metabolismo , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/patologia , Prognóstico , Análise de SobrevidaRESUMO
BACKGROUND: The estimation of recurrence risk remains a critical issue in relation to gastrointestinal stromal tumors (GISTs) treated with adjuvant therapy. The accuracy of the commonly used risk stratifications is not always adequate. METHODS: For this study, data were prospectively collected from 68 patients with GISTs who underwent R0 surgery between 2004 and 2009. The results from this analysis cohort were evaluated using the data obtained from an additional 40 patients in the validation cohort. Cyclin-dependent kinase 1 (CDK1)- and CDK2-specific activities were measured using a non-RI kinase assay system. RESULTS: The specific activities of CDK1 and CDK2, but not their expression, significantly correlated with recurrence. The specific activities of both CDK1 and CDK2 were independently correlated with mitosis and significantly correlated with recurrence-free survival (RFS). In the multivariate analysis, CDK2-specific activity (P = 0.0006), tumor size (P = 0.0347), and KIT deletion mutations (P = 0.0006) were significantly correlated with RFS in the analysis cohort. In the validation cohort, CDK2-specific activity (P = 0.0368) was identified as an independent prognostic factor for tumor recurrences with tumor location (P = 0.0442). CONCLUSION: The results suggest that the specific activities of CDK1 and CDK2 may reflect the proliferative activity of GISTs and that CDK2-specific activity is a good prognostic factor predicting recurrence after macroscopic complete resection of GISTs.
Assuntos
Proteína Quinase CDC2/metabolismo , Quinase 2 Dependente de Ciclina/metabolismo , Tumores do Estroma Gastrointestinal/enzimologia , Neoplasias Intestinais/enzimologia , Neoplasias Hepáticas/enzimologia , Neoplasias Peritoneais/enzimologia , Neoplasias Gástricas/enzimologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Intervalo Livre de Doença , Feminino , Tumores do Estroma Gastrointestinal/secundário , Tumores do Estroma Gastrointestinal/cirurgia , Genótipo , Humanos , Neoplasias Intestinais/patologia , Neoplasias Intestinais/cirurgia , Intestino Delgado , Neoplasias Hepáticas/secundário , Masculino , Pessoa de Meia-Idade , Mitose , Neoplasias Peritoneais/secundário , Proteínas Proto-Oncogênicas c-kit/genética , Curva ROC , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/genética , Estudos Retrospectivos , Medição de Risco , Deleção de Sequência , Neoplasias Gástricas/patologia , Neoplasias Gástricas/cirurgia , Carga Tumoral , Adulto JovemRESUMO
BACKGROUND: In renal cell carcinoma (RCC), the prediction of metastasis via tumor prognostic markers remains a major problem. The objective of our study was to evaluate the efficacy of cyclin-dependent kinase (CDK)1 and CDK2 activity as a prognostic marker in human RCC. METHODS: Surgical specimens were obtained from 125 patients with RCC without metastasis. Protein expression and kinase activity of CDKs were analyzed using a newly developed assay system named C2P (Sysmex, Kobe, Japan). We then examined the specific activities (SAs) of CDK1 and CDK2 and calculated CDK2SA-CDK1SA ratio in RCC. Also, risk score (RS) was examined. RESULTS: A total of 125 cases were tested, though 34 cases were excluded because of low sample quality (25 cases) and assay failure (9 cases). In total, 91 cases were analyzed. They included 68 male and 23 female patients, ranging in age from 19 to 83 years. At a median follow-up of 36 months (1-109M), tumor with low CDK2SA-CDK1SA ratio showed significantly better 5-year recurrence-free survival than those with high CDK2SA-CDK1SA ratio (88.7% vs. 54.7%, P = 0.00141). Also, RS enabled the classification of RCCs into high-risk and low-risk groups, and patients with tumors classified as low RS showed better recurrence-free survival than patients with tumors with high RS (88.7% vs. 54.7%, P = 0.0141). CONCLUSION: CDK1SA of tumors and the CDK2SA are both associated with recurrence and prognosis. IMPACT: CDK-based risk demonstrated is strongly associated with clinical outcome. CDK-based risk should be an accurate system for predicting recurrence and survival for planning follow-up.
Assuntos
Biomarcadores Tumorais/metabolismo , Carcinoma de Células Renais/enzimologia , Quinase 2 Dependente de Ciclina/metabolismo , Quinases Ciclina-Dependentes/metabolismo , Neoplasias Renais/enzimologia , Recidiva Local de Neoplasia/enzimologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Proteína Quinase CDC2 , Carcinoma de Células Renais/patologia , Feminino , Humanos , Neoplasias Renais/patologia , Masculino , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/patologia , Valor Preditivo dos Testes , Prognóstico , Adulto JovemRESUMO
BACKGROUND: Lung cancer is one of the leading causes of cancer death worldwide. Even with complete resection, the prognosis of early-stage non-small cell lung cancer is poor due to local and distant recurrence, and it remains unclear which biomarkers are clinically useful for predicting recurrence or for determining the efficacy of chemotherapy. Recently, several lines of evidence have indicated that the enzymatic activity of cyclin-dependent kinases could be a clinically relevant prognostic marker for some cancers. We investigated whether the specific activity of cyclin-dependent kinases 1 and 2 could predict recurrence or death in early non-small cell lung cancer patients. METHODS: Patients with newly diagnosed, pathologically confirmed non-small cell lung cancer were entered into this blinded cohort study. The activity of cyclin-dependent kinases was determined in 171 samples by the C2P® assay, and the results were subjected to statistical analysis with recurrence or death as a clinical outcome. RESULTS: The Cox proportional hazards model revealed that the activity of cyclin-dependent kinase 1, but not 2, was a predictor of recurrence, independent of sex, age, and stage. By contrast, cyclin-dependent kinase 2 activity was a predictor of death, independent of sex and stage. CONCLUSION: This study suggested the possible clinical use of cyclin-dependent kinase 1 as a predictor of recurrence and cyclin-dependent kinase 2 as a predictor of overall survival in early-stage non-small cell lung cancer. Thus, a combination of activity of cyclin-dependent kinases 1 and 2 is useful in decision-making regarding treatment strategies for non-small cell lung cancer after surgery.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Quinases Ciclina-Dependentes/metabolismo , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Idoso , Idoso de 80 Anos ou mais , Carcinoma Pulmonar de Células não Pequenas/mortalidade , Estudos de Coortes , Feminino , Seguimentos , Humanos , Neoplasias Pulmonares/mortalidade , Masculino , Pessoa de Meia-Idade , Recidiva Local de Neoplasia , Estadiamento de Neoplasias , Prognóstico , Modelos de Riscos Proporcionais , Curva ROCRESUMO
Taxanes are among the drugs most commonly used for preoperative chemotherapy for breast cancer. Taxanes induce mitotic arrest and subsequent apoptosis. The spindle-assembly checkpoint (SAC) is known to be activated during mitosis, along with cyclin-dependent kinase-1 (CDK1), and is required for taxane-induced cell death. We hypothesized that CDK1 activity predicts response to taxane-containing chemotherapy. This study included breast cancer patients who received preoperative chemotherapy- taxane-containing treatment followed by anthracycline-based treatment-and then underwent surgery. Before starting taxane-containing chemotherapy, patients underwent fine-needle aspiration biopsy, and the biopsy samples were incubated in paclitaxel solution to measure CDK activity. Clinical were evaluated after taxane therapy, and pathological resposes were evaluated after completion of all preoperative chemotherapy. Thirty five patients were eligible for analysis of clinical response to taxane-containing therapy. Twenty-six patients had taxane-sensitive and 9 taxane-resistant tumors. Using a cut-off of CDK activity determined by the ROC analysis, patients were classified into SAC function and dysfunction groups. Univariate logistic regression analysis with clinicopathologic parameters showed that only CDK-based SAC functionality was significantly correlated with clinical response (P =0.017). No significant correlation was observed between SAC functionality and pathologic response. CDK-based SAC functionality significantly predicted clinical response (P =.0072, overall agreement = 71.4%), and this is a unique mechanism-based marker for predicting taxane chemosensitivity. Further, large prospective study is needed to determine CDK-based SAC functionality could be developed as a predictive biomarker.
RESUMO
Increasing ability of early breast cancer (BC) diagnosis leading to more early stage detection, better survival, and low relapse marks one of the milestones achieved over the decades. Foregoing poses a challenge for clinicians regarding optimal treatment, in which over- and under-treatment should be avoided. Classical prognostic and predictive factors fall short for individualized adjuvant therapy selection in this patient group. The key to better characterization may be found in the biology underlying individual tumors. We hypothesized that markers related to cellular proliferation and apoptosis and the balance between these two processes in tumor development will be predictive for clinical outcome. Our study population (N = 822) consisted of all early stage BC patients primarily treated with surgery in our center between 1985 and 1996. Sections of available tumor tissue (87 %, 714/822) were immunohistochemically stained for expression of p53, active-caspase-3, and Ki67. In 43 % (304/714) and 18 % (126/714) of this cohort, respectively, a biochemical C2P(®) risk prediction and caspase-3 assay were performed. Expression data of the mentioned markers, single, or combined, were analyzed. Results showed that both the single and combined markers, whether of apoptotic or proliferative origin had associations with clinical outcome. An additive effect was seen for the hazard ratios when data on p53, active caspase-3, and Ki67 status were combined. The assembled prognostic apoptotic-proliferative subtype showed significant association for both the overall survival (p = 0.024) and relapse-free period (p = 0.001) in the multivariate analyses of grade I breast tumors. Combined markers of tumor cell apoptosis and proliferation represent tumor aggressiveness. The apoptotic-proliferative subtypes that we present in this study represent a clinical prognostic profile with solid underlying biological rationale and pose a promising method for accurate identification of grade I BC patients in need of an aggressive therapeutic approach, thus contributing to precision medicine in BC disease.
Assuntos
Apoptose/fisiologia , Biomarcadores Tumorais/análise , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Neoplasias da Mama/cirurgia , Adulto , Idoso , Neoplasias da Mama/mortalidade , Caspase 3/análise , Caspase 3/metabolismo , Proliferação de Células , Detecção Precoce de Câncer , Feminino , Humanos , Antígeno Ki-67/análise , Antígeno Ki-67/metabolismo , Pessoa de Meia-Idade , Valor Preditivo dos Testes , Prognóstico , Estudos Retrospectivos , Resultado do Tratamento , Proteína Supressora de Tumor p53/análise , Proteína Supressora de Tumor p53/metabolismoRESUMO
BACKGROUND: Muscle oxygenation correlates with systemic oxygen uptake (V(O2)) in normal subjects; however, whether this relationship exists in COPD patients remains unclear. The purpose of this study was to investigate the influence of skeletal muscle oxygenation on V(O2) during exercise in patients with COPD. METHODS: Eight subjects performed an incremental cycle ergometer exercise test. We measured ventilation and pulmonary gas exchange with a metabolic measurement system. We also continuously monitored S(pO2), and measured tissue oxygen saturation (S(tO2)) in the vastus lateralis with continuous-wave near-infrared spectroscopy. We calculated the muscle oxygen extraction rate (MOER) based on S(pO2) and S(tO2). In addition, we calculated Pearson correlation coefficients to examine the relationships between the V(O2) obtained during exercise testing and the mean values of S(pO2), S(tO2), heart rate (HR), and MOER for each 30-second interval of the tests. Finally, we analyzed the relationships between the peak V(O2) and the slopes of HR/V(O2), S(pO2)/V(O2), S(tO2)/V(O2), and MOER/V(O2). RESULTS: With the increasing exercise intensity, many subjects showed a gradual decrease in S(tO2) and S(pO2), but a gradual increase in HR and MOER. V(O2) was negatively correlated with S(tO2) and S(pO2), and was positively correlated with HR and MOER. However, peak V(O2) was not correlated with any of the slopes. CONCLUSIONS: V(O2) is highly influenced by oxygen utilization in exercising muscles, as well as by blood oxygenation levels and cardiac function. However, the impact of skeletal muscle utilization during exercise on peak V(O2) varied greatly among the subjects.
Assuntos
Exercício Físico/fisiologia , Músculo Esquelético/metabolismo , Consumo de Oxigênio , Oxigênio/metabolismo , Doença Pulmonar Obstrutiva Crônica/metabolismo , Idoso , Idoso de 80 Anos ou mais , Teste de Esforço , Frequência Cardíaca , Humanos , Masculino , Pessoa de Meia-Idade , Oxiemoglobinas/metabolismo , Doença Pulmonar Obstrutiva Crônica/fisiopatologia , Troca Gasosa Pulmonar , Ventilação Pulmonar , Espectroscopia de Luz Próxima ao InfravermelhoRESUMO
Aberrant promoter methylation of genes is a common molecular event in breast cancer. Thus, DNA methylation analysis is expected to be a new tool for cancer diagnosis. In this article, we have established a new, high-performance DNA methylation assay, the one-step methylation-specific polymerase chain reaction (OS-MSP) assay, which is optimized for analyzing gene methylation in serum DNA. The OS-MSP assay is designed to detect aberrant promoter methylation of GSTP1, RASSF1A, and RARß2 genes in serum DNA. Moreover, two quality control markers were designed for monitoring the bisulfite conversion efficiency and measuring the DNA content in the serum. Serum samples were collected from patients with primary (n = 101, stages I-III) and metastatic breast cancers (n = 58) as well as from healthy controls (n = 87). If methylation of at least one of the three genes was observed, the OS-MSP assay was considered positive. The sensitivity of this assay was significantly higher than that of the assay involving conventional tumor markers (CEA and/or CA15-3) for stages I (24 vs. 8%) and II (26 vs. 8%) breast cancer and similar to that of the assay involving the conventional tumor markers for stage III (18 vs. 19%) and metastatic breast cancers (55 vs. 59%). The results of the OS-MSP assay and those of the assay involving CEA and/or CA15-3 seemed to compensate for each other because sensitivity of these assays increased to 78% when used in combination for metastatic breast cancer. In conclusion, we have developed a new OS-MSP assay with improved sensitivity and convenience; thus, this assay is more suitable for detecting aberrant promoter methylation in serum DNA. Moreover, the combination of the OS-MSP assay and the assay involving CEA and/or CA15-3 is promising for enhancing the sensitivity of diagnosis of metastatic breast cancer.
Assuntos
Biomarcadores Tumorais/sangue , Neoplasias da Mama/diagnóstico , Carcinoma Ductal de Mama/diagnóstico , DNA/sangue , Glutationa S-Transferase pi/sangue , Receptores do Ácido Retinoico/sangue , Proteínas Supressoras de Tumor/sangue , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/genética , Neoplasias da Mama/sangue , Neoplasias da Mama/patologia , Antígeno Carcinoembrionário/sangue , Carcinoma Ductal de Mama/sangue , Carcinoma Ductal de Mama/secundário , Estudos de Casos e Controles , DNA/genética , Metilação de DNA , Feminino , Estudos de Associação Genética , Glutationa S-Transferase pi/genética , Humanos , Metástase Linfática , Pessoa de Meia-Idade , Técnicas de Diagnóstico Molecular , Mucina-1/sangue , Estadiamento de Neoplasias , Reação em Cadeia da Polimerase/métodos , Regiões Promotoras Genéticas , Receptores do Ácido Retinoico/genética , Sensibilidade e Especificidade , Análise de Sequência de DNA , Proteínas Supressoras de Tumor/genéticaRESUMO
It is necessary to evaluate the degree of severity and make a diagnosis of exacerbation factor for acute exacerbation of COPD patients. The highest frequency exacerbation factors are respiratory infection and aerial pollution, so clinical examinations and treatments should be done in consideration of these exacerbation factors. In case of respiratory failure, it is preferable to treat a patient in the hospital. Furthermore, it is important to evaluate the type of respiratory failure, in other words, with or without of hypercapnia. When patients have respiratory failure with hypercapnia, ventilatory assisit therapy is necessary in addition to oxygen therapy. Moreover, it is very important to prevent exacerbation, because patients of COPD exacerbation have poor prognosis and QOL.
Assuntos
Hospitalização , Unidades de Terapia Intensiva , Doença Pulmonar Obstrutiva Crônica/terapia , Humanos , Doença Pulmonar Obstrutiva Crônica/tratamento farmacológico , Doença Pulmonar Obstrutiva Crônica/fisiopatologiaRESUMO
OBJECTIVE: Ovarian clear cell carcinoma (CCC) carries a poor prognosis because of its insensitivity to chemotherapy. We previously found an association between reduced proliferation of CCC and chemoresistance; here we investigated the mechanism of the reduced proliferation. METHODS: We assessed cell cycle function by measuring the activity of cyclin-dependent kinases (CDKs) and the protein expression of cyclins, the CDK inhibitors, and p53 in 22 ovarian cancer cell lines and 60 human ovarian cancer specimens. We examined the cellular location of p27, p27 phosphorylated at threonine 157 (p27(Thr157)), and CDK2 protein by confocal microscopy and western blotting. We tested the effect of the inhibitor of phosphatidylinositol-3-kinase (PI3K) and small interfering RNA against p27 (si-p27) in two CCC cell lines (RMG-I, SMOV-2). RESULTS: CCC cells had lower CDK2 activity and higher p27 expression than serous adenocarcinoma (SA) cells. Low CDK2 activity correlated with high p27 protein expression. p27(Thr157) sequestered CDK2 in the cytoplasm, but PI3K inhibitor or si-p27 maintained CDK2 in the nucleus and restored its activity. In human specimens, CDK2 was mostly in the cytoplasm and was spatially associated with p27; CDK2 activity was lower in the CCC than in the SA specimens. si-p27 enhanced the cytotoxic effect of cisplatin, doxorubicin, and gemcitabine in both RMG-I cells and SMOV-2 cells. CONCLUSIONS: Reduced CDK2 activity via the cytoplasmic sequestration of CDK2 by p27(Thr157) may contribute to suppression of CCC proliferation. A prospective study is needed to determine whether the cytoplasmic sequestration of CDK2 results in the chemoresistance of CCC.
Assuntos
Adenocarcinoma de Células Claras/metabolismo , Adenocarcinoma de Células Claras/patologia , Quinase 2 Dependente de Ciclina/metabolismo , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/patologia , Adenocarcinoma de Células Claras/tratamento farmacológico , Processos de Crescimento Celular/efeitos dos fármacos , Processos de Crescimento Celular/fisiologia , Linhagem Celular Tumoral , Cisplatino/farmacologia , Inibidor de Quinase Dependente de Ciclina p27/deficiência , Inibidor de Quinase Dependente de Ciclina p27/genética , Citoplasma/metabolismo , Desoxicitidina/análogos & derivados , Desoxicitidina/farmacologia , Doxorrubicina/farmacologia , Ativação Enzimática/efeitos dos fármacos , Feminino , Técnicas de Silenciamento de Genes , Humanos , Neoplasias Ovarianas/tratamento farmacológico , Fosfatidilinositol 3-Quinases/metabolismo , Inibidores de Fosfoinositídeo-3 Quinase , Fosforilação , GencitabinaRESUMO
OBJECTIVE: The cell cycle profile test is suggested to be an independent prognostic indicator for breast cancer patients. To further clarify the prognostic value, we applied this to breast cancer patients treated with postoperative 5-fluorouracil-based chemotherapy. METHODS: A total of 153 breast cancer patients, who were treated with postoperative 5-fluorouracil-based chemotherapies, were randomly selected. Specific activities of cyclin-dependent kinases 1 and 2 in the tumor samples were analyzed. Patients were divided into three categories (low, intermediate or high risk) based on cell cycle profile analysis. RESULTS: The proportions of the cell cycle profile categories were 39% for low risk, 10% for intermediate risk and 45% for high risk, respectively. Although the cell cycle profile test did not show a significant predictive power for relapse-free survival (high vs. low risk; P = 0.052), the cell cycle profile categories were significant prognostic factors in a subgroup of 98 patients with fewer than three involved nodes (high vs. low risk, P = 0.004). Multivariate analyses also indicated that a cell cycle profile parameter (high vs. low risk) was an independent prognostic indicator from the number of involved nodes and clinical stage in this subgroup (hazard ratio = 2.46, P = 0.01). Interestingly, the prognostic power of the cell cycle profile test was significant in 75 patients treated with oral 5-fluorouracil derivatives alone (hazard ratio = 6.29 for high vs. low risk, P = 0.02). CONCLUSIONS: These findings suggest that the cell cycle profile test is useful for predicting a higher risk of relapse in patients treated with postoperative 5-fluorouracil-based chemotherapy.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Ciclo Celular , Fluoruracila/administração & dosagem , Linfonodos/patologia , Mastectomia , Adulto , Idoso , Antimetabólitos Antineoplásicos/administração & dosagem , Biomarcadores Tumorais/análise , Neoplasias da Mama/química , Neoplasias da Mama/cirurgia , Quimioterapia Adjuvante , Intervalo Livre de Doença , Feminino , Humanos , Metástase Linfática , Mastectomia/métodos , Computação Matemática , Pessoa de Meia-Idade , Análise Multivariada , Estadiamento de Neoplasias , Razão de Chances , Valor Preditivo dos Testes , Prognóstico , Modelos de Riscos ProporcionaisRESUMO
Patients with severe COPD may develop hypercapnic respiratory failure requiring mechanical ventilation and are at risk of becoming ventilator-dependent. We describe a patient with uncontrollable hypercapnic respiratory failure on mechanical ventilation whose respiratory condition and life were improved after bronchoscopic lung volume reduction. Further discussion of the possibility of bronchoscopic lung volume reduction in patients who require mechanical ventilation due to hypercapnic respiratory failure is warranted.
Assuntos
Broncoscopia/métodos , Hipercapnia/cirurgia , Pneumonectomia/métodos , Enfisema Pulmonar/cirurgia , Idoso , Humanos , Masculino , Doença Pulmonar Obstrutiva Crônica/cirurgia , Respiração Artificial , Insuficiência Respiratória/cirurgia , Índice de Gravidade de Doença , Resultado do TratamentoRESUMO
Massive hemoptysis, an extremely life-threatening condition with a high mortality, requires prompt diagnosis. Pheochromocytoma describes various clinical manifestations attributable to the excessive secretion of catecholamine by the tumor, however the disease is not typically included the differential diagnoses of massive hemoptysis. We describe a case of a 33-year-old man with acute respiratory failure due to cardiovascular crisis and massive hemoptysis as a presenting symptom of pheochromocytoma. Failure to recognize massive hemoptysis as a presenting symptom of pheochromocytoma could miss the diagnosis and a risk a catastrophic outcome. Therefore, massive hemoptysis should be considered as a possible presenting symptom of pheochromocytoma, especially in patients with severe paroxysmal hypertension.
Assuntos
Neoplasias das Glândulas Suprarrenais/diagnóstico , Hemoptise/diagnóstico , Feocromocitoma/diagnóstico , Síndrome do Desconforto Respiratório/diagnóstico , Adulto , Diagnóstico Diferencial , Humanos , MasculinoRESUMO
INTRODUCTION: Paclitaxel is used widely in the treatment of breast cancer. Not all tumors respond to this drug, however, and the characteristics that distinguish resistant tumors from sensitive tumors are not well defined. Activation of the spindle assembly checkpoint is required for paclitaxel-induced cell death. We hypothesized that cyclin-dependent kinase (CDK) 1 activity and CDK2 activity in cancer cells, which reflect the activation state of the spindle assembly checkpoint and the growth state, respectively, predict sensitivity to paclitaxel. METHODS: Cell viability assays and DNA and chromatin morphology analyses were performed in human breast cancer cell lines to evaluate sensitivity to paclitaxel and the cell cycle response to paclitaxel. We then examined the specific activities of CDK1 and CDK2 in these cell lines and in xenograft models of human breast cancer before and after paclitaxel treatment. Protein expression and kinase activity of CDKs and cyclins were analyzed using a newly developed assay system. RESULTS: In the cell lines, biological response to paclitaxel in vitro did not accurately predict sensitivity to paclitaxel in vivo. Among the breast cancer xenograft tumors, however, tumors with significantly increased CDK1 specific activity after paclitaxel treatment were sensitive to paclitaxel in vivo, whereas tumors without such an increase were resistant to paclitaxel in vivo. Baseline CDK2 specific activity was higher in tumors that were sensitive to paclitaxel than in tumors that were resistant to paclitaxel. CONCLUSIONS: The change in CDK1 specific activity of xenograft tumors after paclitaxel treatment and the CDK2 specific activity before paclitaxel treatment are both associated with the drug sensitivity in vivo. Analysis of cyclin-dependent kinase activity in the clinical setting could be a powerful approach for predicting paclitaxel sensitivity.