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1.
Zhong Nan Da Xue Xue Bao Yi Xue Ban ; 46(4): 432-437, 2021 Apr 28.
Artículo en Inglés, Chino | MEDLINE | ID: mdl-33967092

RESUMEN

Cardio-facio-cutaneous (CFC) syndrome is an extremely rare autosomal dominant genetic disease due to BRAF and other gene mutations. The main characteristics of the patients are craniofacial deformities, cardiac malformations, skin abnormalities, delay of language and motor development, gastrointestinal dysfunction, intellectual disability, and epilepsy. In this case, the child has a typical CFC syndrome face and developmental delay. The gene results of the second-generation sequencing technology showed that there was a mutation site c.1741A>G (p. Asn581Asp) (heterozygous) in exon 14 of the BRAF (NM_004333.5) gene. The mutation was not observed in the child's parents. The above-mentioned mutation may be a de novo mutation. There is no effective therapy for this disease so far.


Asunto(s)
Anomalías Múltiples , Displasia Ectodérmica , Cardiopatías Congénitas , Niño , Displasia Ectodérmica/genética , Facies , Insuficiencia de Crecimiento , Cardiopatías Congénitas/genética , Humanos , Mutación , Proteínas Proto-Oncogénicas B-raf/genética
2.
Int J Mol Sci ; 22(6)2021 Mar 22.
Artículo en Inglés | MEDLINE | ID: mdl-33810045

RESUMEN

Melanoma is a highly metastatic disease with an increasing rate of incidence worldwide. It is treatment refractory and has poor clinical prognosis; therefore, the development of new therapeutic agents for metastatic melanoma are urgently required. In this study, we created a lung-seeking A375LM5IF4g/Luc BRAFV600E mutant melanoma cell clone and investigated the bioefficacy of a plant sesquiterpene lactone deoxyelephantopin (DET) and its novel semi-synthetic derivative, DETD-35, in suppressing metastatic A375LM5IF4g/Luc melanoma growth in vitro and in a xenograft mouse model. DET and DETD-35 treatment inhibited A375LM5IF4g/Luc cell proliferation, and induced G2/M cell-cycle arrest and apoptosis. Furthermore, A375LM5IF4g/Luc exhibited clonogenic, metastatic and invasive abilities, and several A375LM5IF4g/Luc metastasis markers, N-cadherin, MMP2, vimentin and integrin α4 were significantly suppressed by treatment with either compound. Interestingly, DET- and DETD-35-induced Reactive Oxygen Species (ROS) generation and glutathione (GSH) depletion were found to be upstream events important for the in vitro activities, because exogenous GSH supplementation blunted DET and DETD-35 effects on A375LM5IF4g/Luc cells. DET and DETD-35 also induced mitochondrial DNA mutation, superoxide production, mitochondrial bioenergetics dysfunction, and mitochondrial protein deregulation. Most importantly, DET and DETD-35 inhibited lung metastasis of A375LM5IF4g/Luc in NOD/SCID mice through inhibiting pulmonary vascular permeability and melanoma cell (Mel-A+) proliferation, angiogenesis (VEGF+, CD31+) and EMT (N-cadherin) in the tumor microenvironment in the lungs. These findings indicate that DET and DETD-35 may be useful in the intervention of lung metastatic BRAFV600E mutant melanoma.


Asunto(s)
Antineoplásicos Fitogénicos/aislamiento & purificación , Antineoplásicos Fitogénicos/farmacología , Asteraceae/química , Lactonas/aislamiento & purificación , Lactonas/farmacología , Extractos Vegetales/aislamiento & purificación , Extractos Vegetales/farmacología , Sesquiterpenos/aislamiento & purificación , Sesquiterpenos/farmacología , Animales , Antineoplásicos Fitogénicos/química , Apoptosis/efectos de los fármacos , Apoptosis/genética , Puntos de Control del Ciclo Celular/efectos de los fármacos , Puntos de Control del Ciclo Celular/genética , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Humanos , Inmunohistoquímica , Lactonas/química , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/secundario , Melanoma/patología , Ratones , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Estructura Molecular , Estrés Oxidativo/efectos de los fármacos , Extractos Vegetales/química , Proteínas Proto-Oncogénicas B-raf/genética , Especies Reactivas de Oxígeno/metabolismo , Sesquiterpenos/química , Microambiente Tumoral/efectos de los fármacos , Ensayos Antitumor por Modelo de Xenoinjerto
3.
Sheng Wu Yi Xue Gong Cheng Xue Za Zhi ; 38(1): 191-195, 2021 Feb 25.
Artículo en Chino | MEDLINE | ID: mdl-33899445

RESUMEN

In recent years, with the improvement of the sensitivity of examination equipment and the change of people's living environment and diet, the rate of thyroid cancer has risen rapidly, which has increased nearly five folds in 10 years. The pathogenesis, clinical manifestation, biological behavior, treatment and prognosis of thyroid carcinoma of different pathological types are obviously different. Papillary thyroid carcinoma (PTC) can develop at any age, which accounts for about 90% of thyroid cancer. It progresses slowly and has favourable prognosis, but lymph node metastasis appears easily. Whether PTC is accompanied by lymph node metastasis has an important impact on its prognosis and outcome. The Raf murine sarcoma viral oncogene homolog B(BRAF)gene mutation plays a crucial role in PTC lymph node metastasis. Having an in-depth understanding of the specific role and mechanism of BRAF gene mutation in PTC is expected to provide new ideas for diagnosis and treatment of PTC.


Asunto(s)
Carcinoma Papilar , Neoplasias de la Tiroides , Animales , Carcinoma Papilar/genética , Humanos , Metástasis Linfática , Ratones , Mutación , Oncogenes , Proteínas Proto-Oncogénicas B-raf/genética , Cáncer Papilar Tiroideo/genética , Neoplasias de la Tiroides/genética
4.
Nat Commun ; 12(1): 2028, 2021 04 01.
Artículo en Inglés | MEDLINE | ID: mdl-33795686

RESUMEN

Germline mutations in BRAF and other components of the MAPK pathway are associated with the congenital syndromes collectively known as RASopathies. Here, we report the association of Septo-Optic Dysplasia (SOD) including hypopituitarism and Cardio-Facio-Cutaneous (CFC) syndrome in patients harbouring mutations in BRAF. Phosphoproteomic analyses demonstrate that these genetic variants are gain-of-function mutations leading to activation of the MAPK pathway. Activation of the MAPK pathway by conditional expression of the BrafV600E/+ allele, or the knock-in BrafQ241R/+ allele (corresponding to the most frequent human CFC-causing mutation, BRAF p.Q257R), leads to abnormal cell lineage determination and terminal differentiation of hormone-producing cells, causing hypopituitarism. Expression of the BrafV600E/+ allele in embryonic pituitary progenitors leads to an increased expression of cell cycle inhibitors, cell growth arrest and apoptosis, but not tumour formation. Our findings show a critical role of BRAF in hypothalamo-pituitary-axis development both in mouse and human and implicate mutations found in RASopathies as a cause of endocrine deficiencies in humans.


Asunto(s)
Mutación con Ganancia de Función , Hipopituitarismo/genética , Hipotálamo/metabolismo , Hipófisis/metabolismo , Proteínas Proto-Oncogénicas B-raf/genética , Animales , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Células Cultivadas , Niño , Preescolar , Corticotrofos/citología , Corticotrofos/metabolismo , Displasia Ectodérmica/genética , Facies , Insuficiencia de Crecimiento/genética , Células HEK293 , Cardiopatías Congénitas/genética , Humanos , Lactante , Sistema de Señalización de MAP Quinasas/genética , Melanotrofos/citología , Melanotrofos/metabolismo , Ratones Noqueados , Ratones Transgénicos , Proteínas Proto-Oncogénicas B-raf/metabolismo , Secuenciación del Exoma Completo/métodos
5.
Anticancer Res ; 41(4): 1761-1769, 2021 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-33813380

RESUMEN

BACKGROUND: Clinical significance of plasma urothelial carcinoma associated 1 (UCA1) in patients with colorectal cancer (CRC) remains unclear. This study investigated the usefulness of plasma UCA1 as a biomarker in patients with CRC. MATERIALS AND METHODS: UCA1 levels were measured in the plasma and tissue from patients with CRC by quantitative polymerase chain reaction. Relationships between plasma UCA1 and clinicopathological features were examined. RESULTS: Plasma UCA1 levels were significantly lower in patients with CRC than in healthy volunteers. UCA1 expression in B-Raf proto-oncogene serine/threonine kinase (BRAF)-mutant CRC tissue was also lower than that in non-cancerous tissue, although it was higher in CRC with wild-type BRAF. In right-sided CRC, a lower plasma UCA1 level was associated with pT4 and BRAF mutation. In contrast, in left-sided CRC, higher plasma UCA1 was associated with pT4 and pStage 3b-4. CONCLUSION: Plasma UCA1 is a useful biomarker for CRC detection and predicting clinicopathological features, particularly BRAF mutation.


Asunto(s)
Biomarcadores de Tumor/genética , Ácidos Nucleicos Libres de Células/genética , Neoplasias Colorrectales/genética , Mutación , Proteínas Proto-Oncogénicas B-raf/genética , ARN Largo no Codificante/genética , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor/sangre , Estudios de Casos y Controles , Ácidos Nucleicos Libres de Células/sangre , Neoplasias Colorrectales/sangre , Neoplasias Colorrectales/patología , Femenino , Predisposición Genética a la Enfermedad , Humanos , Masculino , Persona de Mediana Edad , Estadificación de Neoplasias , Fenotipo , ARN Largo no Codificante/sangre
6.
Nat Commun ; 12(1): 2288, 2021 04 16.
Artículo en Inglés | MEDLINE | ID: mdl-33863883

RESUMEN

Hypothalamic tanycytes in median eminence (ME) are emerging as a crucial cell population that regulates endocrine output, energy balance and the diffusion of blood-born molecules. Tanycytes have recently been considered as potential somatic stem cells in the adult mammalian brain, but their regenerative and tumorigenic capacities are largely unknown. Here we found that Rax+ tanycytes in ME of mice are largely quiescent but quickly enter the cell cycle upon neural injury for self-renewal and regeneration. Mechanistically, Igf1r signaling in tanycytes is required for tissue repair under injury conditions. Furthermore, Braf oncogenic activation is sufficient to transform Rax+ tanycytes into actively dividing tumor cells that eventually develop into a papillary craniopharyngioma-like tumor. Together, these findings uncover the regenerative and tumorigenic potential of tanycytes. Our study offers insights into the properties of tanycytes, which may help to manipulate tanycyte biology for regulating hypothalamic function and investigate the pathogenesis of clinically relevant tumors.


Asunto(s)
Craneofaringioma/patología , Células Ependimogliales/fisiología , Eminencia Media/fisiología , Neoplasias Experimentales/patología , Regeneración , Animales , Carcinogénesis/patología , Autorrenovación de las Células/fisiología , Craneofaringioma/inducido químicamente , Craneofaringioma/genética , Proteínas del Ojo/metabolismo , Femenino , Proteínas de Homeodominio/metabolismo , Eminencia Media/citología , Ratones , Neoplasias Experimentales/inducido químicamente , Neoplasias Experimentales/genética , Proteínas Proto-Oncogénicas B-raf/genética , RNA-Seq , Receptor IGF Tipo 1/metabolismo , Transducción de Señal , Análisis de la Célula Individual , Factores de Transcripción/metabolismo
7.
Zhonghua Bing Li Xue Za Zhi ; 50(3): 229-235, 2021 Mar 08.
Artículo en Chino | MEDLINE | ID: mdl-33677887

RESUMEN

Objective: To investigate the clinicopathological and molecular characteristics of the epithelioid glioblastoma (eGBM) with BRAF V600E mutation. Methods: Sixteen cases of eGBM with BRAF V600E mutation diagnosed at the West China Hospital of Sichuan University, China from 2012 to 2019 were collected. Their clinicopathological and molecular characteristics were analyzed. Results: The range of patients' age was from 7 to 61 years (median 31.5 years). There were 4 males and 12 females, with a male to female ratio of 1∶3. Eleven cases were newly diagnosed eGBM and five cases had a previous history of astrocytomas. Most of the tumors were located in the cerebral hemisphere, often in the frontal lobe, with an average diameter of 4.6 cm (2.0-8.0 cm). The tumors were composed of relatively uniform, closely packed epithelioid cells, some showing discohesion, with distinct cell membrane, eosinophilic cytoplasm, eccentric nuclei, distinct nucleoli and mitotic activity. Palisaded/coagulative necrosis was seen in all cases. Glomerular microvascular proliferation was seen in most of the cases, while mono-or multi-nucleated tumor giant cells were seen in some cases. Focal sarcomatoid area was seen in 2 cases, and focal pleomorphic xanthoastrocytoma (PXA)-like area was seen in 3 cases. Immunohistochemistry showed variable positivity for GFAP, Olig2 and p53. The median Ki-67 index was 30% (10%-50%). Only one case lost ATRX protein expression. Sanger sequencing identified the BRAF V600E mutation in all sixteen patients. Five cases also had mutations in the TERT gene promoter. No IDH1 (R132) or IDH2 (R172) mutation was detected. Surgical resection of the tumors was performed for all patients, and 3 patients also received adjuvant radiotherapy and chemotherapy. Follow-up data were available for 15 patients, with a follow-up time of 1-89 months (median 10 months). Among the 15 patients, 7 patients died of disease and another 5 patients had recurrences. The overall survival time of the patients under 35 years of age was significantly longer than that of the patients aged 35 years or older (P=0.014), but their progression-free survival was not statistically different (P=0.232). Conclusions: eGBM with BRAF V600E mutation is more commonly detected in young women than other the populations (i.e. elderly or male). The epithelioid morphology should include rhabdoid meningioma, anaplastic PXA, atypical teratoid/rhabdoid tumor, metastatic tumors, and melanoma in its differential diagnosis. PXA-like area is observed in some eGBM cases, suggesting a relationship of these two types of tumor. eGBM is a high-grade malignant tumor and most of the cases show recurrences or deaths in a short-period time. The younger patients have a relatively better prognosis than the older ones.


Asunto(s)
Astrocitoma , Neoplasias Encefálicas , Glioblastoma , Adolescente , Adulto , Anciano , Astrocitoma/genética , Neoplasias Encefálicas/genética , Niño , China , Femenino , Glioblastoma/genética , Humanos , Masculino , Persona de Mediana Edad , Mutación , Proteínas Proto-Oncogénicas B-raf/genética , Adulto Joven
8.
J Med Case Rep ; 15(1): 109, 2021 Mar 02.
Artículo en Inglés | MEDLINE | ID: mdl-33653337

RESUMEN

BACKGROUND: Pneumatosis intestinalis (PI) is a rare entity which refers to the presence of gas within the wall of the small bowel or colon which is a radiographic sign. The etiology and clinical presentation are variable. Patients with PI may present either with chronic mild non-specific symptoms or with acute abdominal pain with peritonitis. Some cases of intestinal pneumatosis have been reported as adverse events of new oncological treatments such as targeted therapies that are widely used in multiple tumors. CASE PRESENTATION: A 59-year-old caucasian female with radioactive iodine-refractory metastatic thyroid papillary carcinoma with BRAFV600E mutation was treated with dabrafenib and trametinib as a compassionate use. After 4 months treatment, positron emission tomography-computed tomography (PET-CT) showed PI. At the time of diagnosis, the patient was asymptomatic without signs of peritonitis. The initial treatment was conservative and no specific treatment for PI was needed. Unfortunately, after dabrafenib-trametinib withdrawal, the patient developed tumor progression with significant clinical worsening. CONCLUSIONS: This case report is, in our knowledge, the first description of PI in a patient treated with dabrafenib-trametinib. Conservative treatment is feasible if there are no abdominal symptoms.


Asunto(s)
Enfisema/diagnóstico por imagen , Imidazoles/efectos adversos , Enfermedades Intestinales/diagnóstico por imagen , Oximas/efectos adversos , Inhibidores de Proteínas Quinasas/efectos adversos , Piridonas/efectos adversos , Pirimidinonas/efectos adversos , Neoplasias de la Base del Cráneo/tratamiento farmacológico , Cáncer Papilar Tiroideo/tratamiento farmacológico , Neoplasias de la Tiroides/patología , Enfisema/inducido químicamente , Femenino , Humanos , Enfermedades Intestinales/inducido químicamente , Radioisótopos de Yodo/uso terapéutico , Persona de Mediana Edad , Tomografía Computarizada por Tomografía de Emisión de Positrones , Proteínas Proto-Oncogénicas B-raf/genética , Neoplasias de la Base del Cráneo/diagnóstico por imagen , Neoplasias de la Base del Cráneo/genética , Neoplasias de la Base del Cráneo/secundario , Cáncer Papilar Tiroideo/diagnóstico por imagen , Cáncer Papilar Tiroideo/genética , Cáncer Papilar Tiroideo/secundario
9.
Methods Mol Biol ; 2265: 265-276, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33704721

RESUMEN

Liquid biopsy has emerged as the next generation target for diagnostics and therapeutic monitoring of many diseases including cancer. Liquid biopsy offers noninvasive analysis of aberrant biomolecular changes (e.g., aberrant protein expression, DNA mutation) which can provide crucial information on disease stages and therapy responses. As a diagnostically important biomarker for melanoma, the detection of the BRAFV600E aberration at the DNA and protein level in liquid biopsies confers an attractive option. This method describes the preparation and operation of an integrated multimolecular sensor (IMMS) for simultaneous detection of the BRAFV600E aberration in both molecular forms from circulating melanoma cells in liquid biopsy. IMMS integrates specific melanoma cell capture, cell release, cell lysis, and electrochemical BRAFV600E detection on a single device. IMMS is demonstrated for a sample-to-answer workflow of plasma spiked with melanoma cells.


Asunto(s)
Técnicas Biosensibles/métodos , Inmunoensayo/métodos , Dispositivos Laboratorio en un Chip , Melanoma/metabolismo , Microfluídica/instrumentación , Microfluídica/métodos , Proteínas Proto-Oncogénicas B-raf/metabolismo , Neoplasias Cutáneas/metabolismo , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Técnicas Biosensibles/instrumentación , Técnicas de Cultivo de Célula/métodos , Humanos , Inmunoensayo/instrumentación , Biopsia Líquida/métodos , Melanoma/genética , Melanoma/patología , Mutación , Células Neoplásicas Circulantes/metabolismo , Células Neoplásicas Circulantes/patología , Proteínas Proto-Oncogénicas B-raf/genética , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/patología
10.
Methods Mol Biol ; 2265: 247-263, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33704720

RESUMEN

In recent years, circulating tumor DNA (ctDNA) has emerged as a promising prognostic and monitoring biomarker of various cancers, including melanoma. However, sensitive methods are required for its preservation, isolation, and detection. Here we describe a sensitive method for plasma ctDNA isolation using a column-based extraction kit, followed by quantification using a single mutational target with a droplet digital PCR system. This sensitive protocol has been successfully used to quantify diverse mutations present in plasma-derived ctDNA from cancer patients. The full procedure, from blood processing to the analysis of results, takes approximately a day of work.


Asunto(s)
ADN Tumoral Circulante/sangre , ADN de Neoplasias/sangre , Melanoma/sangre , Reacción en Cadena de la Polimerasa/métodos , Biomarcadores de Tumor/sangre , Biomarcadores de Tumor/genética , ADN de Neoplasias/genética , ADN de Neoplasias/aislamiento & purificación , Humanos , Melanoma/genética , Melanoma/patología , Plasma/metabolismo , Proteínas Proto-Oncogénicas B-raf/genética
11.
Int J Nanomedicine ; 16: 2323-2335, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33776436

RESUMEN

Background: Colon cancer (CRC) was a malignant tumor and there were about 25% of patients with tumor metastasis at diagnosis stage. Chemotherapeutic agents for metastatic CRC patients were with great side effects and the clinical treatment results of advanced CRC were still not satisfactory. Human epidermal growth factor receptor 2 (HER2) is overexpressed in some CRC patients and is an effective target for CRC patient treatment. Anti-HER2 therapy had a beneficial role in the treatment of HER2-positive metastatic CRC with fewer side effects. CRC patients with BRAF mutations were resistant to HER2 antibodies treatment. Therefore, there was an urgent need to develop new therapeutic agents. Methods: HER2 targeted nanoparticles (TPLNP) drug delivery system loading triptolide (TPL) were prepared and identified. The effects of TPLNP and free TPL on cell viability, targeting and cell cycle progression on HT29 (BRAF mutation) with HER2 overexpression, were evaluated by Cell Counting Kit-8 (CCK8), Fluorescence Activating Cell Sorter (FACS) and immunofluorescence methods, respectively. The anti-tumor efficacies of TPLNP were evaluated in subcutaneous xenograft model of colon cancer and the survival rate, tumor volume, liver and kidney indexes of tumor-bearing mice were measured. Results: TPLNP was small in nanosize (73.4±5.2nm) with narrow size distribution (PDI=0.15±0.02) and favorable zeta potential (pH=9.6, zeta potential: -57.3±6.69mV; pH=7.0, zeta potential: -28.7±5.1mV; pH=5.6, zeta potential: -21.1±4.73mV). Comparing with free TPL treatment group, TPLNP developed stranger colon cancer-killing efficiency in a dose- and time-dependent manner detected with CCK8 method; achieved good in vitro colon cancer targeting detected with flow cytometry and immunofluorescence experiments; enhanced more HT29-HER2 apoptosis and induced more cell cycle arrested in G1-S phase detected with FACS in vitro. As for in vivo antitumor response, TPLNP remarkably inhibited the growth of colon cancer in the colon cancer xenograft model, significantly improved the survival rate and did not exhibit significant liver and kidney toxicity in contrast with free TPL in vivo. Conclusion: TPLNP was effectively against colon cancer with HER2 overexpression and BRAF mutation in pre-clinical models. In summary, the TPLNP appeared to be a promising treatment option for CRC in clinical application based on improved efficacy and the favorable safety profile.


Asunto(s)
Diterpenos/farmacología , Sistemas de Liberación de Medicamentos , Mutación/genética , Nanopartículas/química , Fenantrenos/farmacología , Proteínas Proto-Oncogénicas B-raf/genética , Receptor ErbB-2/metabolismo , Animales , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Carcinogénesis/efectos de los fármacos , Ciclo Celular/efectos de los fármacos , Muerte Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Neoplasias del Colon/tratamiento farmacológico , Neoplasias del Colon/genética , Diterpenos/toxicidad , Compuestos Epoxi/farmacología , Compuestos Epoxi/toxicidad , Femenino , Células HT29 , Humanos , Concentración 50 Inhibidora , Riñón/efectos de los fármacos , Riñón/patología , Hígado/efectos de los fármacos , Hígado/patología , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Nanomedicina , Fenantrenos/toxicidad , Transducción de Señal/efectos de los fármacos
12.
BMC Cancer ; 21(1): 287, 2021 Mar 17.
Artículo en Inglés | MEDLINE | ID: mdl-33731038

RESUMEN

BACKGROUND: While molecularly targeted therapies and immune checkpoint inhibitors have improved the prognosis of advanced melanoma, biomarkers are required to monitor drug responses. Circulating tumor cells (CTCs) are released from primary and/or metastatic tumors into the peripheral blood. We examined whether CTCs have potential as biomarkers by checking the number of CTCs, as well as the BRAF genotype of individual CTCs, in melanoma patients undergoing BRAF/MEK inhibitor treatment. METHODS: CTCs were isolated from peripheral blood using a high-density dielectrophoretic microwell array, followed by labeling with melanoma-specific markers (MART-1 and/or gp100) and a leukocyte marker (CD45). The numbers of CTCs were analyzed in fifteen patients with stage 0-III melanoma. Furthermore, changes in CTC numbers were assessed in five patients with stage IV melanoma at four time points during BRAF/MEK inhibitor treatment, and the BRAF genotype was analyzed in CTCs isolated from one patient. RESULTS: We examined CTCs in patients with stage 0-III (five samples per stage: stage 0-I, stage II, and stage III), and detected CTCs even in patients with early disease (stage 0 and I). Interestingly, recurrence occurred in the lymph nodes of one stage I patient 2 years after the detection of a high number of CTCs in the patient's blood. The total number of CTCs in four of five patients with stage IV melanoma fluctuated in response to BRAF/MEK inhibitor treatment, suggesting that CTC number has potential for use as a drug response marker in advanced disease patients. Interestingly, one of those patients had CTCs harboring seven different BRAF genotypes, and the mutated CTCs disappeared upon BRAF/MEK inhibitor treatment, except for those harboring BRAFA598V. CONCLUSIONS: CTCs are present even in the early stage of melanoma, and the number of CTCs seems to reflect patients' responses to BRAF/MEK inhibitor treatment. Furthermore, genetic heterogeneity of BRAF may contribute to resistance to BRAF/MEK inhibitors. Our findings demonstrate the usefulness of CTC analysis for monitoring responses to targeted therapies in melanoma patients, and for understanding the mechanism of drug resistance.


Asunto(s)
Melanoma/terapia , Células Neoplásicas Circulantes , Inhibidores de Proteínas Quinasas/uso terapéutico , Proteínas Proto-Oncogénicas B-raf/antagonistas & inhibidores , Neoplasias Cutáneas/terapia , Anciano , Anciano de 80 o más Años , Quimioterapia Adyuvante/métodos , Resistencia a Antineoplásicos/genética , Estudios de Factibilidad , Femenino , Heterogeneidad Genética , Humanos , Masculino , Melanoma/sangre , Melanoma/diagnóstico , Melanoma/genética , Persona de Mediana Edad , Terapia Molecular Dirigida/métodos , Estadificación de Neoplasias , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas B-raf/genética , Neoplasias Cutáneas/sangre , Neoplasias Cutáneas/genética
13.
Nat Commun ; 12(1): 1536, 2021 03 09.
Artículo en Inglés | MEDLINE | ID: mdl-33750776

RESUMEN

Hyperactivation of the MAPK signaling pathway motivates the clinical use of MAPK inhibitors for BRAF-mutant melanomas. Heterogeneity in differentiation state due to epigenetic plasticity, however, results in cell-to-cell variability in the state of MAPK dependency, diminishing the efficacy of MAPK inhibitors. To identify key regulators of such variability, we screen 276 epigenetic-modifying compounds, individually or combined with MAPK inhibitors, across genetically diverse and isogenic populations of melanoma cells. Following single-cell analysis and multivariate modeling, we identify three classes of epigenetic inhibitors that target distinct epigenetic states associated with either one of the lysine-specific histone demethylases Kdm1a or Kdm4b, or BET bromodomain proteins. While melanocytes remain insensitive, the anti-tumor efficacy of each inhibitor is predicted based on melanoma cells' differentiation state and MAPK activity. Our systems pharmacology approach highlights a path toward identifying actionable epigenetic factors that extend the BRAF oncogene addiction paradigm on the basis of tumor cell differentiation state.


Asunto(s)
Diferenciación Celular/efectos de los fármacos , Epigenómica/métodos , Melanoma/metabolismo , Dependencia del Oncogén , Animales , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Epigénesis Genética/efectos de los fármacos , Femenino , Histona Demetilasas/metabolismo , Humanos , Histona Demetilasas con Dominio de Jumonji/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Melanocitos/metabolismo , Melanoma/genética , Ratones , Ratones Desnudos , Mutación , Dependencia del Oncogén/genética , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas B-raf/genética , Transducción de Señal/efectos de los fármacos , Ensayos Antitumor por Modelo de Xenoinjerto
14.
Anticancer Res ; 41(3): 1387-1399, 2021 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-33788730

RESUMEN

BACKGROUND/AIM: Despite clinical benefit from treatment with dabrafenib and trametinib in melanoma patients with BRAF mutations, half relapse within months and one-third are unresponsive to treatment. We evaluated the anticancer potential of metformin in combination with trametinib plus paclitaxel, against four melanoma cell lines. MATERIALS AND METHODS: Metformin with trametinib and paclitaxel was tested for effects on cell viability, signaling molecules in MAPK and mTOR pathways, factors involved in epithelial-mesenchymal transition (EMT), and cell motility. RESULTS: The combination of metformin with trametinib and paclitaxel showed differential growth inhibitory effects; synergistic effects were observed in a cell line in which metformin suppresses ERK activity, whereas the combination showed antagonistic effects in a cell line with metformin-induced ERK activation. Trametinib or paclitaxel increased the expression of EMT regulators and melanoma cell motility, which were suppressed by combining metformin with trametinib and paclitaxel. CONCLUSION: The combined treatment of metformin with trametinib and paclitaxel showed divergent effects on melanoma cell viability. Metformin might be useful as a potential adjuvant against cell proliferation and metastatic activity in melanoma patients.


Asunto(s)
Melanoma/patología , Metformina/farmacología , Paclitaxel/farmacología , Piridonas/farmacología , Pirimidinonas/farmacología , Neoplasias Cutáneas/patología , Antineoplásicos/farmacología , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Sinergismo Farmacológico , Transición Epitelial-Mesenquimal/efectos de los fármacos , Humanos , Melanoma/genética , Melanoma/metabolismo , Mutación , Proteínas Proto-Oncogénicas B-raf/genética , Transducción de Señal/efectos de los fármacos , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/metabolismo
15.
Medicine (Baltimore) ; 100(13): e25346, 2021 Apr 02.
Artículo en Inglés | MEDLINE | ID: mdl-33787635

RESUMEN

RATIONALE: Glioblastoma is the most lethal and common malignant brain tumor but rare in patients with neurofibromatosis type 1. The clinical findings and pathological findings with gene signatures in female patients have not been well clarified. PATIENT CONCERNS: A 51-year-old female patient complained of headache and left limb weakness lasting for 20 days. The patient underwent a cesarean section 20 years ago and hysterectomy 1 year ago because of uterine leiomyomas. Multiple café-au-lait spots and neurofibromas were found over patient's chest, neck, back, and arms. The myodynamia of left distant and proximate epipodite were grade 0 and grade 1 respectively. The myodynamia of lower left limb was grade 3. DIAGNOSES: Magnetic resonance imaging revealed a malignant lesion which was most likely a glioblastoma in the right temporo-parietal lobe, approximately 5.6 × 5.9 × 6.9 cm in size with a rounded boundary. INTERVENTIONS: A right temporo-parietal craniotomy was performed to resect the space-occupying lesion for gross total removal. Then, the patient received concurrent chemoradiotherapy. Histological examination confirmed a glioblastoma without v-RAF murine sarcoma viral oncogene homolog B1 gene, isocitrate dehydrogenase 1 gene, and telomerase reverse transcriptase gene promoter mutations. OUTCOMES: After surgery, the headache was relieved and the muscular strength of left limbs did improve. After receiving the standard treatment regimen, the patient was alive at 13 months follow-up. LESSONS: This is the first reported glioblastoma in female neurofibromatosis type 1 patient without v-RAF murine sarcoma viral oncogene homolog B1 gene, isocitrate dehydrogenase 1 gene, and telomerase reverse transcriptase gene promoter mutations. Tumors in adult patients with these signatures were less aggressive with well-circumscribed border and had long-term survivals which strengthened the evidence that these patients may comprise a unique subset in glioblastoma.


Asunto(s)
Biomarcadores de Tumor/genética , Neoplasias Encefálicas/diagnóstico , Glioblastoma/diagnóstico , Neurofibromatosis 1/diagnóstico , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patología , Neoplasias Encefálicas/cirugía , Craneotomía , Femenino , Glioblastoma/genética , Glioblastoma/patología , Glioblastoma/cirugía , Humanos , Isocitrato Deshidrogenasa/genética , Imagen por Resonancia Magnética , Persona de Mediana Edad , Mutación , Neurofibromatosis 1/complicaciones , Neurofibromatosis 1/genética , Lóbulo Parietal/diagnóstico por imagen , Lóbulo Parietal/patología , Lóbulo Parietal/cirugía , Regiones Promotoras Genéticas/genética , Proteínas Proto-Oncogénicas B-raf/genética , Telomerasa/genética , Lóbulo Temporal/diagnóstico por imagen , Lóbulo Temporal/patología , Lóbulo Temporal/cirugía , Transcriptoma/genética
16.
Methods Mol Biol ; 2279: 109-126, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33683689

RESUMEN

Driver mutations in non-small cell lung cancer (NSCLC) have a relevant significance for clinical management. EGFR mutations are the most important predictive biomarkers for NSCLC, although KRAS and BRAF mutations can also be prognostic and predictive biomarkers, respectively. PCR-based approaches followed by sequencing are useful for EGFR, KRAS, and BRAF mutational analysis. Herein, all steps for a PCR-based technique, from DNA isolation from tumor tissue sections to DNA sequencing for genetic analysis of EGFR, KRAS, and BRAF hotspot regions are described.


Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/genética , Neoplasias Pulmonares/genética , Reacción en Cadena de la Polimerasa , Proteínas Proto-Oncogénicas B-raf/genética , Proteínas Proto-Oncogénicas p21(ras)/genética , Carcinoma de Pulmón de Células no Pequeñas/patología , Análisis Mutacional de ADN , Receptores ErbB/genética , Humanos , Neoplasias Pulmonares/patología
17.
Methods Mol Biol ; 2265: 277-286, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33704722

RESUMEN

Molecular testing of tumor biopsies allows for the identification of the key mutations driving a patient's cancer. However, this is limited to singular nodes and may not accurately reflect cancer heterogeneity. Circulating tumor cell (CTC) analyses offer a noninvasive method of interrogating the genomic profile of patient-derived tumor material. To date, molecular analysis of CTCs has relied on the characterization of bulk or pooled CTC lysates, limiting the detection of minor tumorigenic CTC subclones. Here, we show a workflow enabling BRAFV600E/NRASQ61R mutation detection from single cultured melanoma cells by combining micromanipulation and genomic material amplification methods. This workflow can be directly integrated into circulating tumor cell analysis applications.


Asunto(s)
GTP Fosfohidrolasas/genética , Melanoma/genética , Proteínas de la Membrana/genética , Mutación Missense , Células Neoplásicas Circulantes , Proteínas Proto-Oncogénicas B-raf/genética , Análisis de la Célula Individual , Sustitución de Aminoácidos , Línea Celular Tumoral , Humanos , Melanoma/patología
18.
Endocr Pract ; 27(3): 206-211, 2021 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-33655886

RESUMEN

OBJECTIVE: To determine the association between pathologic features and molecular classes (BRAF-like, RAS-like, and non-BRAF-like non-RAS-like [NBNR]). METHODS: Retrospective review of a merged database containing 676 patients, 84% (571/676) were assigned to a molecular class from publicly accessible sequenced data of thyroid neoplasms. RESULTS: The merged cohort included 571 neoplasms: 353 (62%) BRAF-like, 172 (30%) RAS-like, and 46 (8.1%) NBNR. Lymph node metastasis (any N1 disease) was present in 166/337 (49%) of BRAF-like, 23/164 (14%) of RAS-like, and 0/46 (0%) of NBNR and are significantly different (P < .001). Gross extra-thyroidal extension was observed in 27 patients, including 24/331 (7%) of BRAF-like, 2/160 (1%) of RAS-like, and 1/46 (2%) of NBNR (P = .01). N1B lymph node metastases or T4 disease was present in 74/333 (22%) of BRAF-like, 10/160 (6%) of RAS-like, and 1/46 (2%) of NBNR (P < .0001). Distant metastasis was present in 4/151 (2.6%) of BRAF-like, 2/50 (4%) of RAS-like and 0/46 for NBNR (P = .627). Angioinvasion was present in 0/81 (0%) of BRAF-like, 3/53 (6%) of RAS-like, and 3/46 (7%) of NBNR (P = .08); and multifocality was present in 27/81 (33%) of BRAF-like, 9/53 (17%) of RAS-like, and 1/46 (2%) for NBNR (P = .0001). CONCLUSION: Pathological features of metastasis, gross extra-thyroidal extension, and multifocality were more prevalent in BRAF-like samples compared to RAS-like and NBNR. A trend towards increased frequency of angioinvasion in RAS-like and NBNR cancers compared to BRAF-like samples was observed. Further studies are needed to evaluate if preoperative knowledge of molecular mutations in thyroid tumors aids in decision-making regarding extent of surgery.


Asunto(s)
Carcinoma Papilar Folicular , Neoplasias de la Tiroides , Humanos , Metástasis Linfática , Mutación , Proteínas Proto-Oncogénicas B-raf/genética , Estudios Retrospectivos , Neoplasias de la Tiroides/epidemiología , Neoplasias de la Tiroides/genética
19.
Nat Commun ; 12(1): 920, 2021 02 10.
Artículo en Inglés | MEDLINE | ID: mdl-33568647

RESUMEN

Over 300 BRAF missense mutations have been identified in patients, yet currently approved drugs target V600 mutants alone. Moreover, acquired resistance inevitably emerges, primarily due to RAF lesions that prevent inhibition of BRAF V600 with current treatments. Therefore, there is a need for new therapies that target other mechanisms of activated BRAF. In this study, we use the Proteolysis Targeting Chimera (PROTAC) technology, which promotes ubiquitination and degradation of neo-substrates, to address the limitations of BRAF inhibitor-based therapies. Using vemurafenib-based PROTACs, we achieve low  nanomolar degradation of all classes of BRAF mutants, but spare degradation of WT RAF family members. Our lead PROTAC outperforms vemurafenib in inhibiting cancer cell growth and shows in vivo efficacy in a Class 2 BRAF xenograft model. Mechanistic studies reveal that BRAFWT is spared due to weak ternary complex formation in cells owing to its quiescent inactivated conformation, and activation of BRAFWT sensitizes it to degradation. This study highlights the degree of selectivity achievable with degradation-based approaches by targeting mutant BRAF-driven cancers while sparing BRAFWT, providing an anti-tumor drug modality that expands the therapeutic window.


Asunto(s)
Antineoplásicos/administración & dosificación , Neoplasias/tratamiento farmacológico , Inhibidores de Proteínas Quinasas/administración & dosificación , Proteínas Proto-Oncogénicas B-raf/genética , Proteínas Proto-Oncogénicas B-raf/metabolismo , Vemurafenib/administración & dosificación , Animales , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Femenino , Humanos , Ratones , Ratones Desnudos , Terapia Molecular Dirigida , Mutación , Neoplasias/enzimología , Neoplasias/genética , Neoplasias/fisiopatología , Proteolisis/efectos de los fármacos , Proteínas Proto-Oncogénicas B-raf/antagonistas & inhibidores , Ubiquitinación/efectos de los fármacos , Ensayos Antitumor por Modelo de Xenoinjerto
20.
Medicine (Baltimore) ; 100(6): e24458, 2021 Feb 12.
Artículo en Inglés | MEDLINE | ID: mdl-33578538

RESUMEN

RATIONALE: The rare BRAF L597Q (c.T1790A) point mutation has been previously reported in childhood acute lymphoblastic leukemia. We present the first rare case of occult papillary thyroid carcinoma with BRAF L597Q mutation in a Tibetan patient. PATIENT CONCERNS: A 57-year-old male patient presented with a protruding mass on the left forehead for 2 years and numbness in the right limb for 3 weeks. DIAGNOSES: The patient had a double mutation of BRAF L597Q and V600E in 2 separate lesions at thyroid and brain, the immunohistochemical staining showed that the cytokeratin (CK), thyroglobulin (Tg) and thyroid transforming factor-1 (TTF-1) were immunoreactive. All the findings supported the diagnosis of solitary brain metastasis of occult papillary thyroid carcinoma. INTERVENTIONS: The patient underwent left frontal lobe metastasis (thyroid cancer) resection that involved craniectomy and artificial skull repair. OUTCOMES: During the 24-month follow-up, no postoperative complications or recurrence and metastasis were found. LESSONS: This is the first case of solitary brain metastasis of occult papillary thyroid carcinoma with double mutation of BRAF L597Q and V600E in 2 separate lesions reported in the literature. Our study extends the disease spectrum of occult papillary thyroid carcinoma and suggests that the BRAF L597Q mutation might play a specific role in inducing the solitary brain metastasis of occult papillary thyroid carcinoma in a Chinese Tibetan patient, but the detailed molecular mechanism remains to be confirmed by a large number of functional experiments and clinical research.


Asunto(s)
Neoplasias Encefálicas/secundario , Mutación Puntual/genética , Proteínas Proto-Oncogénicas B-raf/genética , Cáncer Papilar Tiroideo/genética , Neoplasias de la Tiroides/genética , Neoplasias Encefálicas/diagnóstico por imagen , Neoplasias Encefálicas/genética , Humanos , Imagen por Resonancia Magnética , Masculino , Persona de Mediana Edad , Neuroimagen , Cáncer Papilar Tiroideo/patología , Neoplasias de la Tiroides/patología , Tomografía Computarizada por Rayos X
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