RESUMEN
PURPOSE: Glioblastoma multiforme (GBM) is a devastating disease. Recent studies suggest that the stem cell properties of GBM contribute to the development of therapy resistance. EXPERIMENTAL DESIGN: The expression of Survivin and Ran was evaluated by immunohistochemistry with GBM tissues, and quantitative reverse transcriptase (qRT)-PCR and immunocytochemistry with patient-derived GBM sphere cultures. With a computational structure-based drug design, 11 small-molecule compounds were designed, synthesized, and evaluated as inhibitor candidates for the molecular interaction of Survivin protein. The molecular mechanism of the lead compound, LLP-3, was determined by Western blot, ELISA, in situ proximity ligation assay, and immunocytochemistry. The effects of LLP-3 treatment on GSCs were evaluated both in vitro and in vivo. Quantitative immunohistochemistry was carried out to compare Survivin expression in tissues from 44 newly diagnosed and 31 recurrent post-chemoradiation GBM patients. Lastly, the sensitivities of temozolomide-resistant GBM spheres to LLP-3 were evaluated in vitro. RESULTS: Survivin and Ran were strongly expressed in GBM tissues, particularly in the perivasculature, and also in patient-derived GSC cultures. LLP-3 treatment disrupted the Survivin-Ran protein complex in cancer cells and abolished the growth of patient-derived GBM spheres in vitro and in vivo. This inhibition was dependent on caspase activity and associated with p53 status of cells. Immunohistochemistry showed that Survivin expression is significantly increased in recurrent GBM compared with newly diagnosed tumors, and temozolomide-resistant GBM spheres exhibited high sensitivities to LLP-3 treatment. CONCLUSIONS: Disruption of the Survivin-Ran complex by LLP-3 abolishes survival and growth of GSCs both in vitro and in vivo, indicating an attractive novel therapeutic approach for GBM.
Asunto(s)
Glioblastoma/metabolismo , Proteínas Inhibidoras de la Apoptosis/metabolismo , Células Madre Neoplásicas/metabolismo , Proteína de Unión al GTP ran/metabolismo , Apoptosis , Línea Celular Tumoral , Proliferación Celular , Supervivencia Celular , Resistencia a Antineoplásicos , Glioblastoma/mortalidad , Glioblastoma/cirugía , Humanos , Proteínas Inhibidoras de la Apoptosis/química , Modelos Moleculares , Pronóstico , Unión Proteica/efectos de los fármacos , Conformación Proteica , Multimerización de Proteína , Transducción de Señal , Survivin , Proteína p53 Supresora de Tumor/metabolismo , Ensayos Antitumor por Modelo de Xenoinjerto , Proteína de Unión al GTP ran/químicaRESUMEN
OBJECTIVE: Glioblastomas (GBMs) are lethal cancers that display cellular hierarchies parallel to normal brain. At the apex are GBM stem cells (GSCs), which are relatively resistant to conventional therapy. Interactions with the adjacent perivascular niche are an important driver of malignancy and self-renewal in GSCs. Extracellular matrix (ECM) cues instruct neural stem/progenitor cell-niche interactions, and the objective of our study was to elucidate its composition and contribution to GSC maintenance in the perivascular niche. METHODS: We interrogated human tumor tissue for immunofluorescence analysis and derived GSCs from tumor tissues for functional studies. Bioinformatics analyses were conducted by mining publicly available databases. RESULTS: We find that laminin ECM proteins are localized to the perivascular GBM niche and inform negative patient prognosis. To identify the source of laminins, we characterized cellular elements within the niche and found that laminin α chains were expressed by nonstem tumor cells and tumor-associated endothelial cells (ECs). RNA interference targeting laminin α2 inhibited GSC growth and self-renewal. In co-culture studies of GSCs and ECs, laminin α2 knockdown in ECs resulted in decreased tumor growth. INTERPRETATION: Our studies highlight the contribution of nonstem tumor cell-derived laminin juxtracrine signaling. As laminin α2 has recently been identified as a molecular marker of aggressive ependymoma, we propose that the brain vascular ECM promotes tumor malignancy through maintenance of the GSC compartment, providing not only a molecular fingerprint but also a possible therapeutic target.
Asunto(s)
Neoplasias Encefálicas/patología , Regulación Neoplásica de la Expresión Génica/fisiología , Glioblastoma/patología , Laminina/metabolismo , Células Madre Neoplásicas/fisiología , Antígeno AC133 , Análisis de Varianza , Antígenos CD/metabolismo , Neoplasias Encefálicas/mortalidad , Supervivencia Celular/efectos de los fármacos , Técnicas de Cocultivo , Biología Computacional , Relación Dosis-Respuesta en la Radiación , Células Endoteliales/efectos de los fármacos , Células Endoteliales/metabolismo , Células Endoteliales/patología , Células Endoteliales/efectos de la radiación , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de la radiación , Glioblastoma/mortalidad , Glicoproteínas/metabolismo , Humanos , Estimación de Kaplan-Meier , Laminina/genética , Imagen por Resonancia Magnética , Masculino , Células Madre Neoplásicas/efectos de los fármacos , Células Madre Neoplásicas/efectos de la radiación , Péptidos/metabolismo , Interferencia de ARN/fisiología , ARN Interferente Pequeño/farmacología , Radiación , Análisis de Regresión , Factores de Tiempo , Análisis de Matrices Tisulares , Células Tumorales Cultivadas , Microambiente Tumoral/fisiologíaRESUMEN
Antiplatelet therapy is the cornerstone of management of acute coronary syndromes. Currently used antiplatelet drugs present several limitations that provoke new searches. These limitations include resistance, delay in the onset of action, risk for bleeding, variations in the individual response, and interaction with other medications (i.e. proton pump inhibitors, calcium channel blockers). New concepts and medications have emerged for the effective inhibition of platelets. Prasugrel, AZD6140 (ticagrelor), cangrelor, and SCH 530348 (thrombin receptor antagonist) are among some of the novel agents that survived randomized trials. In this review, we aimed to summarize novel concepts and agents in antiplatelet therapy.