RESUMEN
In Fig. 3c of this Letter, the the effects of CRISPR-Cas9-mediated deletion of NR3C1, TXNIP and CNR2 in patient-derived B-lineage leukaemia cells were shown. For curves depicting NR3C1 (left graph), data s for TXNIP (middle graph) were inadvertently plotted. This figure has been corrected online, and the original Fig. 3c is shown as Supplementary Information to this Amendment for transparency. The error does not affect the conclusions of the Letter. In addition, Source Data files have been added for the Figs. 1-4 and Extended Data Figs. 1-10 of the original Letter.
RESUMEN
B-lymphoid transcription factors, such as PAX5 and IKZF1, are critical for early B-cell development, yet lesions of the genes encoding these transcription factors occur in over 80% of cases of pre-B-cell acute lymphoblastic leukaemia (ALL). The importance of these lesions in ALL has, until now, remained unclear. Here, by combining studies using chromatin immunoprecipitation with sequencing and RNA sequencing, we identify a novel B-lymphoid program for transcriptional repression of glucose and energy supply. Our metabolic analyses revealed that PAX5 and IKZF1 enforce a state of chronic energy deprivation, resulting in constitutive activation of the energy-stress sensor AMPK. Dominant-negative mutants of PAX5 and IKZF1, however, relieved this glucose and energy restriction. In a transgenic pre-B ALL mouse model, the heterozygous deletion of Pax5 increased glucose uptake and ATP levels by more than 25-fold. Reconstitution of PAX5 and IKZF1 in samples from patients with pre-B ALL restored a non-permissive state and induced energy crisis and cell death. A CRISPR/Cas9-based screen of PAX5 and IKZF1 transcriptional targets identified the products of NR3C1 (encoding the glucocorticoid receptor), TXNIP (encoding a glucose-feedback sensor) and CNR2 (encoding a cannabinoid receptor) as central effectors of B-lymphoid restriction of glucose and energy supply. Notably, transport-independent lipophilic methyl-conjugates of pyruvate and tricarboxylic acid cycle metabolites bypassed the gatekeeper function of PAX5 and IKZF1 and readily enabled leukaemic transformation. Conversely, pharmacological TXNIP and CNR2 agonists and a small-molecule AMPK inhibitor strongly synergized with glucocorticoids, identifying TXNIP, CNR2 and AMPK as potential therapeutic targets. Furthermore, our results provide a mechanistic explanation for the empirical finding that glucocorticoids are effective in the treatment of B-lymphoid but not myeloid malignancies. Thus, B-lymphoid transcription factors function as metabolic gatekeepers by limiting the amount of cellular ATP to levels that are insufficient for malignant transformation.
Asunto(s)
Linfocitos B/metabolismo , Metabolismo Energético/genética , Regulación Neoplásica de la Expresión Génica , Glucosa/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras B/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras B/metabolismo , Factores de Transcripción/metabolismo , Quinasas de la Proteína-Quinasa Activada por el AMP , Proteínas Quinasas Activadas por AMP/antagonistas & inhibidores , Adenosina Trifosfato/metabolismo , Animales , Linfocitos B/efectos de los fármacos , Carcinogénesis/genética , Proteínas Portadoras/agonistas , Proteínas Portadoras/metabolismo , Muerte Celular , Inmunoprecipitación de Cromatina , Ciclo del Ácido Cítrico , Modelos Animales de Enfermedad , Femenino , Glucocorticoides/farmacología , Glucocorticoides/uso terapéutico , Humanos , Factor de Transcripción Ikaros/metabolismo , Ratones , Ratones Transgénicos , Factor de Transcripción PAX5/deficiencia , Factor de Transcripción PAX5/genética , Factor de Transcripción PAX5/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras B/tratamiento farmacológico , Leucemia-Linfoma Linfoblástico de Células Precursoras B/patología , Proteínas Serina-Treonina Quinasas/metabolismo , Ácido Pirúvico/metabolismo , Receptor Cannabinoide CB2/agonistas , Receptor Cannabinoide CB2/metabolismo , Receptores de Glucocorticoides/metabolismo , Análisis de Secuencia de ARNRESUMEN
Loss-of-function mutations in hematopoietic transcription factors including PAX5 occur in most cases of B-progenitor acute lymphoblastic leukemia (B-ALL), a disease characterized by the accumulation of undifferentiated lymphoblasts. Although PAX5 mutation is a critical driver of B-ALL development in mice and humans, it remains unclear how its loss contributes to leukemogenesis and whether ongoing PAX5 deficiency is required for B-ALL maintenance. Here we used transgenic RNAi to reversibly suppress endogenous Pax5 expression in the hematopoietic compartment of mice, which cooperates with activated signal transducer and activator of transcription 5 (STAT5) to induce B-ALL. In this model, restoring endogenous Pax5 expression in established B-ALL triggers immunophenotypic maturation and durable disease remission by engaging a transcriptional program reminiscent of normal B-cell differentiation. Notably, even brief Pax5 restoration in B-ALL cells causes rapid cell cycle exit and disables their leukemia-initiating capacity. These and similar findings in human B-ALL cell lines establish that Pax5 hypomorphism promotes B-ALL self-renewal by impairing a differentiation program that can be re-engaged despite the presence of additional oncogenic lesions. Our results establish a causal relationship between the hallmark genetic and phenotypic features of B-ALL and suggest that engaging the latent differentiation potential of B-ALL cells may provide new therapeutic entry points.
Asunto(s)
Diferenciación Celular/genética , Factor de Transcripción PAX5/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/patología , Células Precursoras de Linfocitos B/citología , Animales , Línea Celular Tumoral , Células Cultivadas , Regulación Neoplásica de la Expresión Génica , Técnicas de Silenciamiento del Gen , Genes myc/genética , Humanos , Ratones , Ratones Transgénicos , Factor de Transcripción PAX5/metabolismo , Factor de Transcripción STAT5/metabolismo , Transducción de SeñalRESUMEN
B cell and plasma cell fates are controlled by different transcriptional networks, as exemplified by the mutually exclusive expression and cross-antagonism of the B cell identity factor Pax5 and the plasma cell regulator Blimp1. It has been postulated that repression of Pax5 by Blimp1 is essential for plasma cell development. Here, we challenged this hypothesis by analyzing the IghPax5/+ mouse, which expressed a Pax5 minigene from the immunoglobulin heavy-chain locus. Despite high Pax5 expression, plasma cells efficiently developed in young IghPax5/+ mice at steady state and upon immunization, while their number moderately declined in older mice. Although Pax5 significantly deregulated the plasma cell expression program, key plasma cell regulators were normally expressed in IghPax5/+ plasma cells. While IgM and IgA secretion by IghPax5/+ plasma cells was normal, IgG secretion was modestly decreased. Hence, Pax5 repression is not essential for robust plasma cell development and antibody secretion, although it is required for optimal IgG production and accumulation of long-lived plasma cells.
Asunto(s)
Linfocitos B/inmunología , Diferenciación Celular/genética , Factor de Transcripción PAX5/metabolismo , Células Plasmáticas/inmunología , Traslado Adoptivo , Animales , Diferenciación Celular/inmunología , Inmunización , Cambio de Clase de Inmunoglobulina/genética , Inmunoglobulina G/biosíntesis , Cadenas Pesadas de Inmunoglobulina/genética , Cadenas Pesadas de Inmunoglobulina/metabolismo , Inmunoglobulina M/biosíntesis , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Factor de Transcripción PAX5/genética , Factor 1 de Unión al Dominio 1 de Regulación Positiva/metabolismo , Transducción de Señal/genética , Transducción de Señal/inmunología , Linfocitos T/inmunologíaRESUMEN
Tumors are composed of phenotypically heterogeneous cancer cells that often resemble various differentiation states of their lineage of origin. Within this hierarchy, it is thought that an immature subpopulation of tumor-propagating cancer stem cells (CSCs) differentiates into non-tumorigenic progeny, providing a rationale for therapeutic strategies that specifically eradicate CSCs or induce their differentiation. The clinical success of these approaches depends on CSC differentiation being unidirectional rather than reversible, yet this question remains unresolved even in prototypically hierarchical malignancies, such as acute myeloid leukemia (AML). Here, we show in murine and human models of AML that, upon perturbation of endogenous expression of the lineage-determining transcription factor PU.1 or withdrawal of established differentiation therapies, some mature leukemia cells can de-differentiate and reacquire clonogenic and leukemogenic properties. Our results reveal plasticity of CSC maturation in AML, highlighting the need to therapeutically eradicate cancer cells across a range of differentiation states.
Asunto(s)
Diferenciación Celular/fisiología , Transdiferenciación Celular/fisiología , Leucemia Mieloide Aguda/patología , Células Madre Neoplásicas/fisiología , Proteínas Proto-Oncogénicas/metabolismo , Transactivadores/metabolismo , Animales , Carcinogénesis , Plasticidad de la Célula , Células Cultivadas , Humanos , Leucemia Mieloide Aguda/metabolismo , Ratones , Proteínas Proto-Oncogénicas/genética , Transactivadores/genética , Tretinoina/metabolismoRESUMEN
Genetic alterations disrupting the transcription factor IKZF1 (encoding IKAROS) are associated with poor outcome in B lineage acute lymphoblastic leukemia (B-ALL) and occur in >70% of the high-risk BCR-ABL1+ (Ph+) and Ph-like disease subtypes. To examine IKAROS function in this context, we have developed novel mouse models allowing reversible RNAi-based control of Ikaros expression in established B-ALL in vivo. Notably, leukemias driven by combined BCR-ABL1 expression and Ikaros suppression rapidly regress when endogenous Ikaros is restored, causing sustained disease remission or ablation. Comparison of transcriptional profiles accompanying dynamic Ikaros perturbation in murine B-ALL in vivo with two independent human B-ALL cohorts identified nine evolutionarily conserved IKAROS-repressed genes. Notably, high expression of six of these genes is associated with inferior event-free survival in both patient cohorts. Among them are EMP1, which was recently implicated in B-ALL proliferation and prednisolone resistance, and the novel target CTNND1, encoding P120-catenin. We demonstrate that elevated Ctnnd1 expression contributes to maintenance of murine B-ALL cells with compromised Ikaros function. These results suggest that IKZF1 alterations in B-ALL leads to induction of multiple genes associated with proliferation and treatment resistance, identifying potential new therapeutic targets for high-risk disease.