RESUMEN
The Philadelphia chromosome (Ph) encoding the oncogenic BCR-ABL1 kinase defines a subset of acute lymphoblastic leukemia (ALL) with a particularly unfavorable prognosis. ALL cells are derived from B cell precursors in most cases and typically carry rearranged immunoglobulin heavy chain (IGH) variable (V) region genes devoid of somatic mutations. Somatic hypermutation is restricted to mature germinal center B cells and depends on activation-induced cytidine deaminase (AID). Studying AID expression in 108 cases of ALL, we detected AID mRNA in 24 of 28 Ph(+) ALLs as compared with 6 of 80 Ph(-) ALLs. Forced expression of BCR-ABL1 in Ph(-) ALL cells and inhibition of the BCR-ABL1 kinase showed that aberrant expression of AID depends on BCR-ABL1 kinase activity. Consistent with aberrant AID expression in Ph(+) ALL, IGH V region genes and BCL6 were mutated in many Ph(+) but unmutated in most Ph(-) cases. In addition, AID introduced DNA single-strand breaks within the tumor suppressor gene CDKN2B in Ph(+) ALL cells, which was sensitive to BCR-ABL1 kinase inhibition and silencing of AID expression by RNA interference. These findings identify AID as a BCR-ABL1-induced mutator in Ph(+) ALL cells, which may be relevant with respect to the particularly unfavorable prognosis of this leukemia subset.
Asunto(s)
Linfocitos B/inmunología , Citidina Desaminasa/metabolismo , Regulación Neoplásica de la Expresión Génica/inmunología , Cromosoma Filadelfia , Leucemia-Linfoma Linfoblástico de Células Precursoras/inmunología , Proteínas Tirosina Quinasas/genética , Linfocitos B/metabolismo , Secuencia de Bases , Western Blotting , Análisis Mutacional de ADN , Proteínas de Unión al ADN/genética , Citometría de Flujo , Proteínas de Fusión bcr-abl , Genes myc/genética , Humanos , Región Variable de Inmunoglobulina/genética , Datos de Secuencia Molecular , Mutación/genética , Análisis de Secuencia por Matrices de Oligonucleótidos , Oligonucleótidos , Leucemia-Linfoma Linfoblástico de Células Precursoras/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-bcl-6 , Interferencia de ARN , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Alineación de SecuenciaRESUMEN
Bruton's tyrosine kinase (BTK) deficiency results in a differentiation block at the pre-B cell stage. Likewise, acute lymphoblastic leukemia cells are typically arrested at early stages of B cell development. We therefore investigated BTK function in B cell precursor leukemia cells carrying a BCR-ABL1, E2A-PBX1, MLL-AF4, TEL-AML1, or TEL-PDGFRB gene rearrangement. Although somatic mutations of the BTK gene are rare in B cell precursor leukemia cells, we identified kinase-deficient splice variants of BTK throughout all leukemia subtypes. Unlike infant leukemia cells carrying an MLL-AF4 gene rearrangement, where expression of full-length BTK was detectable in only four of eight primary cases, in leukemia cells harboring other fusion genes full-length BTK was typically coexpressed with kinase-deficient variants. As shown by overexpression experiments, kinase-deficient splice variants can act as a dominant-negative BTK in that they suppress BTK-dependent differentiation and pre-B cell receptor responsiveness of the leukemia cells. On the other hand, induced expression of full-length BTK rendered the leukemia cells particularly sensitive to apoptosis. Comparing BTK expression in surviving or preapoptotic leukemia cells after 10-Gy gamma radiation, we observed selective survival of leukemia cells that exhibit expression of dominant-negative BTK forms. These findings indicate that lack of BTK expression or expression of dominant-negative splice variants in B cell precursor leukemia cells can (i) inhibit differentiation beyond the pre-B cell stage and (ii) protect from radiation-induced apoptosis.