Development Refractoriness of MLL-Rearranged Human B Cell Acute Leukemias to Reprogramming into Pluripotency

Induced pluripotent stem cells (iPSCs) are a powerful tool for disease modeling. They are routinely generated from healthy donors and patients from multiple cell types at different developmental stages. However, reprogramming leukemias is an extremely inefficient process. Few studies generated iPSCs from primary chronic myeloid leukemias, but iPSC generation from acute myeloid or lymphoid leukemias (ALL) has not been achieved. We attempted to generate iPSCs from different subtypes of B-ALL to address the developmental impact of leukemic fusion genes. OKSM(L)-expressing mono/polycistronic-, retroviral/lentiviral/episomal-, and Sendai virus vector-based reprogramming strategies failed to render iPSCs in vitro and in vivo. Addition of transcriptomic-epigenetic reprogramming ‘‘boosters’’ also failed to generate iPSCs from B cell blasts and B-ALL lines, and when iPSCs emerged they lacked leukemic fusion genes, demonstrating non-leukemic myeloid origin. Conversely, MLL-AF4-overexpressing hematopoietic stem cells/B progenitors were successfully reprogrammed, indicating that B cell origin and leukemic fusion gene were not reprogramming barriers. Global transcriptome/DNA methylome profiling suggested a developmental/differentiation refractoriness of MLL-rearranged B-ALL to reprogramming into pluripotency

Integrative methylome-transcriptome analysis unravels cancer cell vulnerabilities in infant MLL-rearranged B cell acute lymphoblastic leukemia

B cell acute lymphoblastic leukemia (B-ALL) is the most common childhood cancer. As predicted by its prenatal origin, infant B-ALL (iB-ALL) shows an exceptionally silent DNA mutational landscape, suggesting that alternative epigenetic mechanisms may substantially contribute to its leukemogenesis. Here, we have integrated genome-wide DNA methylome and transcriptome data from 69 patients with de novo MLL-rearranged leukemia (MLLr) and non-MLLr iB-ALL leukemia uniformly treated according to the Interfant-99/06 protocol. iB-ALL methylome signatures display a plethora of common and specific alterations associated with chromatin states related to enhancer and transcriptional control in normal hematopoietic cells. DNA methylation, gene expression, and gene coexpression network analyses segregated MLLr away from non-MLLr iB-ALL and identified a coordinated and enriched expression of the AP-1 complex members FOS and JUN and RUNX factors in MLLr iB-ALL, consistent with the significant enrichment of hypomethylated CpGs in these genes. Integrative methylome-transcriptome analysis identified consistent cancer cell vulnerabilities, revealed a robust iB-ALL–specific gene expression–correlating dmCpG signature, and confirmed an epigenetic control of AP-1 and RUNX members in reshaping the molecular network of MLLr iB-ALL. Finally, pharmacological inhibition or functional ablation of AP-1 dramatically impaired MLLr-leukemic growth in vitro and in vivo using MLLr-iB-ALL patient–derived xenografts, providing rationale for new therapeutic avenues in MLLr-iB-ALL.We thank CERCA/Generalitat de Catalunya (SGR180) and Fundació Josep Carreras-Obra Social la Caixa for their institutional support. Financial support for this work was obtained from the European Research Council (CoG-2014-646903 and PoC-2018-811220 to PM), the Spanish Ministry of Economy and Competitiveness (SAF-2019-108160-R and SAF2016-76758-R to PM and IV, respectively), the Spanish Association against cancer (AECC-CI-2015 and PROYE18061FERN to CB and MFF), the Fundación Uno entre Cienmil (to PM), the Health Institute Carlos III (ISCIII/FEDER, PI17/01028, PI15/00892, PI18/01527 to CB and AFF/MFF, respectively). We also acknowledge the Plan de Ciencia, Tecnología e Innovación from the Asturias Government cofunding 2018–2022/FEDER (IDI/2018/146to MFF). MFF also acknowledges funding from Fundación General CSIC (0348_CIE_6_E). PM also acknowledges financial support from Fundación Leo Messi. JRT and MV are supported by Juan de la Cierva fellowships by the Spanish Ministry of Science and Innovation (FJCI-2015-26965, IJC2018-36825-I, IJCI-2017-3317) and IUOPA-ISPA-FINBA (The IUOPA is supported by the Obra Social Cajastur-Liberbank, Spain). RTR is supported by a fellowship from the AECC scientific foundation. RFP and PSO are supported by the Severo Ochoa program (BP17-114 and BP17-165, respectively).Peer reviewe

Caracterización genómica y transcriptómica de la leucemia linfoblástica aguda pro-b del lactante con t(4;11)(q21q23)/MLL-AF4+

RESUMEN: La leucemia linfoblástica aguda pro-b es el cáncer más prevalente en la infancia. Este tumor es especialmente letal en infantes con la translocación t(4;11), que produce el gen de fusión MLL-AF4 y que está presente en aproximadamente el ochenta por ciento de los casos. La traslocación MLL-AF4 es un evento clonal que podrían ser el evento iniciador de la leucemia sin la ayuda de eventos secundarios, sin embargo, ha sido muy difícil generar un modelo adecuado que recopile los síntomas desarrollados por los pacientes. Esto, unido al poco número de casos, hace especialmente difícil la caracterización molecular de esta enfermedad. En esta tesis, hemos llevado a cabo un gran esfuerzo de caracterización genómica y transcripcional completa en la colección de pacientes más grande estudiada hasta la fecha que han sido sometidos además a un protocolo estandarizado de tratamiento. Los resultados muestran la menor tasa de mutación por megabase encontrada en cualquier otro tipo tumoral siendo los genes RAS los únicos que muestran mutaciones recurrentes. A pesar de su recurrencia, las mutaciones en los genes RAS se pierden frecuentemente en las muestras en recaída a pesar de la gran acumulación de mutaciones en estas muestras lo que evidencia la presencia de inestabilidad genética. Los estudios transcriptómicos han permitido identificar nuevas rutas moleculares potencialmente implicadas en la patología de la enfermedad y, junto con los análisis del repertorio de variantes del receptor de célula B, confirman que la leucemia se origina en una célula inmadura pro-B antes de la reorganización del receptor BCR. Por último, hemos comprobado que aproximadamente la mitad de los pacientes muestran expresión del gen de fusión recíproco AF4-MLL lo que se asocia con la sobreexpresión de los genes HOXA y con una mayor supervivencia de los pacientes. Todos estos resultados han ayudado a una mejor comprensión sobre el origen y las rutas moleculares implicadas en la leucemia infantil pro-B con reorganizaciones en el gen MLL lo que podría con el tiempo contribuir a mejorar el diagnóstico, manejo y tratamiento de los pacientes de esta trágica enfermedad

Integrative methylome-transcriptome analysis unravels cancer cell vulnerabilities in infant MLL-rearranged B cell acute lymphoblastic leukemia

B cell acute lymphoblastic leukemia (B-ALL) is the most common childhood cancer. As predicted by its prenatal origin, infant B-ALL (iB-ALL) shows an exceptionally silent DNA mutational landscape, suggesting that alternative epigenetic mechanisms may substantially contribute to its leukemogenesis. Here, we have integrated genome-wide DNA methylome and transcriptome data from 69 patients with de novo MLL-rearranged leukemia (MLLr) and non-MLLr iB-ALL leukemia uniformly treated according to the Interfant-99/06 protocol. iB-ALL methylome signatures display a plethora of common and specific alterations associated with chromatin states related to enhancer and transcriptional control in normal hematopoietic cells. DNA methylation, gene expression, and gene coexpression network analyses segregated MLLr away from non-MLLr iB-ALL and identified a coordinated and enriched expression of the AP-1 complex members FOS and JUN and RUNX factors in MLLr iB-ALL, consistent with the significant enrichment of hypomethylated CpGs in these genes. Integrative methylome-transcriptome analysis identified consistent cancer cell vulnerabilities, revealed a robust iB-ALL–specific gene expression–correlating dmCpG signature, and confirmed an epigenetic control of AP-1 and RUNX members in reshaping the molecular network of MLLr iB-ALL. Finally, pharmacological inhibition or functional ablation of AP-1 dramatically impaired MLLr-leukemic growth in vitro and in vivo using MLLr-iB-ALL patient–derived xenografts, providing rationale for new therapeutic avenues in MLLr-iB-ALL