RESUMO
Glucocorticoid (GC) resistance is a continuing clinical problem in childhood acute lymphoblastic leukaemia (ALL) but the underlying mechanisms remain unclear. A proteomic approach was used to compare profiles of the B-lineage ALL GC-sensitive cell line, PreB 697, and its GC-resistant sub-line, R3F9, pre- and post-dexamethasone exposure. PAX5, a transcription factor critical to B-cell development was differentially regulated in the PreB 697 compared to the R3F9 cell line in response to GC. PAX5 basal protein expression was less in R3F9 compared to its GC-sensitive parent and confirmed to be lower in other GC-resistant sub-lines of Pre B 697 and was associated with a decreased expression of the PAX5 transcriptional target, CD19. Gene set enrichment analysis showed that increasing GC-resistance was associated with differentiation from preB-II to an immature B-lymphocyte stage. GC-resistant sub-lines were shown to have higher levels of phosphorylated JNK compared to the parent line and JNK inhibition caused re-sensitization to GC. Exploiting this maturation may be key to overcoming GC resistance and targeting signalling pathways linked to the maturation state, such as JNK, may be a novel approach.
Assuntos
Antineoplásicos/farmacologia , Linfócitos B/efeitos dos fármacos , Dexametasona/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , MAP Quinase Quinase 4/antagonistas & inibidores , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Proteínas de Neoplasias/biossíntese , Leucemia-Linfoma Linfoblástico de Células Precursoras B/patologia , Inibidores de Proteínas Quinases/farmacologia , Proteômica/métodos , Apoptose/efeitos dos fármacos , Linfócitos B/patologia , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/fisiologia , Éxons/genética , Regulação Leucêmica da Expressão Gênica/efeitos dos fármacos , Humanos , Reação em Cadeia da Polimerase Multiplex , Mutação , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/genética , Fator de Transcrição PAX5/genética , Fator de Transcrição PAX5/fisiologia , Fosforilação/efeitos dos fármacos , Leucemia-Linfoma Linfoblástico de Células Precursoras B/enzimologia , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Reação em Cadeia da Polimerase em Tempo Real , Espectrometria de Massas em TandemRESUMO
Casitas B-lineage lymphoma (CBL) proteins are RING finger ubiquitin E3 ligases that attenuate the signaling of receptor tyrosine kinases and are mutated in a number of myeloid disorders. In this study, mutational screening of the linker-RING domains of CBL and CBLB was performed by denaturing high performance liquid chromatography in a cohort of diagnostic (n = 180) or relapse (n = 46) samples from children with acute lymphoblastic leukemia. Somatic mutations were identified in three children, giving an overall incidence of 1.7% and involved small deletions affecting the intron/exon boundaries of exon 8, leading to skipping of exon 8 and abolishing E3 ligase function. Mutated primary samples were associated with constitutive activation of the RAS pathway and sensitivity to MEK inhibitors was shown. Thus, mutation of CBL is an alternative route to activate the RAS pathway and may identify children who are candidates for MEK inhibitor clinical trials.