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1.
Haematologica ; 103(12): 2016-2025, 2018 12.
Artículo en Inglés | MEDLINE | ID: mdl-30049824

RESUMEN

The fusion oncoprotein BCR-ABL1 exhibits aberrant tyrosine kinase activity and it has been proposed that it deregulates signaling networks involving both transcription factors and non-coding microRNAs that result in chronic myeloid leukemia (CML). Previously, microRNA expression profiling showed deregulated expression of miR-150 and miR-155 in CML. In this study, we placed these findings into the broader context of the MYC/miR-150/MYB/miR-155/PU.1 oncogenic network. We propose that up-regulated MYC and miR-155 in CD34+ leukemic stem and progenitor cells, in concert with BCR-ABL1, impair the molecular mechanisms of myeloid differentiation associated with low miR-150 and PU.1 levels. We revealed that MYC directly occupied the -11.7 kb and -0.35 kb regulatory regions in the MIR150 gene. MYC occupancy was markedly increased through BCR-ABL1 activity, causing inhibition of MIR150 gene expression in CML CD34+ and CD34- cells. Furthermore, we found an association between reduced miR-150 levels in CML blast cells and their resistance to tyrosine kinase inhibitors (TKIs). Although TKIs successfully disrupted BCR-ABL1 kinase activity in proliferating CML cells, this treatment did not efficiently target quiescent leukemic stem cells. The study presents new evidence regarding the MYC/miR-150/MYB/miR-155/PU.1 leukemic network established by aberrant BCR-ABL1 activity. The key connecting nodes of this network may serve as potential druggable targets to overcome resistance of CML stem and progenitor cells.


Asunto(s)
Resistencia a Antineoplásicos/genética , Proteínas de Fusión bcr-abl/genética , Genes myc/genética , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , MicroARNs/genética , Adulto , Anciano , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/genética , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Regulación hacia Abajo/efectos de los fármacos , Resistencia a Antineoplásicos/efectos de los fármacos , Femenino , Regulación Leucémica de la Expresión Génica/efectos de los fármacos , Células HL-60 , Humanos , Células K562 , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Leucemia Mielógena Crónica BCR-ABL Positiva/patología , Masculino , Persona de Mediana Edad , Células Madre Neoplásicas/efectos de los fármacos , Células Madre Neoplásicas/metabolismo , Inhibidores de Proteínas Quinasas/farmacología
2.
Leukemia ; 36(7): 1879-1886, 2022 07.
Artículo en Inglés | MEDLINE | ID: mdl-35676453

RESUMEN

Several studies have reported that chronic myeloid leukaemia (CML) patients expressing e14a2 BCR::ABL1 have a faster molecular response to therapy compared to patients expressing e13a2. To explore the reason for this difference we undertook a detailed technical comparison of the commonly used Europe Against Cancer (EAC) BCR::ABL1 reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) assay in European Treatment and Outcome Study (EUTOS) reference laboratories (n = 10). We found the amplification ratio of the e13a2 amplicon was 38% greater than e14a2 (p = 0.015), and the amplification efficiency was 2% greater (P = 0.17). This subtle difference led to measurable transcript-type dependent variation in estimates of residual disease which could be corrected by (i) taking the qPCR amplification efficiency into account, (ii) using alternative RT-qPCR approaches or (iii) droplet digital PCR (ddPCR), a technique which is relatively insensitive to differences in amplification kinetics. In CML patients, higher levels of BCR::ABL1/GUSB were identified at diagnosis for patients expressing e13a2 (n = 67) compared to e14a2 (n = 78) when analysed by RT-qPCR (P = 0.0005) but not ddPCR (P = 0.5). These data indicate that widely used RT-qPCR assays result in subtly different estimates of disease depending on BCR::ABL1 transcript type; these differences are small but may need to be considered for optimal patient management.


Asunto(s)
Proteínas de Fusión bcr-abl , Leucemia Mielógena Crónica BCR-ABL Positiva , Proteínas de Fusión bcr-abl/genética , Humanos , Mesilato de Imatinib , Leucemia Mielógena Crónica BCR-ABL Positiva/diagnóstico , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Neoplasia Residual/genética , Reacción en Cadena en Tiempo Real de la Polimerasa
3.
Front Oncol ; 11: 744373, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-34616685

RESUMEN

Somatic mutations are a common molecular mechanism through which chronic myeloid leukemia (CML) cells acquire resistance to tyrosine kinase inhibitors (TKIs) therapy. While most of the mutations in the kinase domain of BCR-ABL1 can be successfully managed, the recurrent somatic mutations in other genes may be therapeutically challenging. Despite the major clinical relevance of mutation-associated resistance in CML, the mechanisms underlying mutation acquisition in TKI-treated leukemic cells are not well understood. This work demonstrated de novo acquisition of mutations on isolated single-cell sorted CML clones growing in the presence of imatinib. The acquisition of mutations was associated with the significantly increased expression of the LIG1 and PARP1 genes involved in the error-prone alternative nonhomologous end-joining pathway, leading to genomic instability, and increased expression of the UNG, FEN and POLD3 genes involved in the base-excision repair (long patch) pathway, allowing point mutagenesis. This work showed in vitro and in vivo that de novo acquisition of resistance-associated mutations in oncogenes is the prevalent method of somatic mutation development in CML under TKIs treatment.

4.
Leukemia ; 34(8): 2113-2124, 2020 08.
Artículo en Inglés | MEDLINE | ID: mdl-32472084

RESUMEN

This work investigated patient-specific genomic BCR-ABL1 fusions as markers of measurable residual disease (MRD) in chronic myeloid leukaemia, with a focus on relevance to treatment-free remission (TFR) after achievement of deep molecular response (DMR) on tyrosine kinase inhibitor (TKI) therapy. DNA and mRNA BCR-ABL1 measurements by qPCR were compared in 2189 samples (129 patients) and by digital PCR in 1279 sample (62 patients). A high correlation was found at levels of disease above MR4, but there was a poor correlation for samples during DMR. A combination of DNA and RNA MRD measurements resulted in a better prediction of molecular relapse-free survival (MRFS) after TKI stop (n = 17) or scheduled interruption (n = 25). At 18 months after treatment cessation, patients with stopped or interrupted TKI therapy who were DNA negative/RNA negative during DMR maintenance (green group) had an MRFS of 80% and 100%, respectively, compared with those who were DNA positive/RNA negative (MRFS = 57% and 67%, respectively; yellow group) or DNA positive/RNA positive (MRFS = 20% for both cohorts; red group). Thus, we propose a "traffic light" stratification as a TFR predictor based on DNA and mRNA BCR-ABL1 measurements during DMR maintenance before TKI cessation.


Asunto(s)
Proteínas de Fusión bcr-abl/genética , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Reacción en Cadena de la Polimerasa/métodos , Inhibidores de Proteínas Quinasas/uso terapéutico , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Adulto , Anciano , Femenino , Humanos , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Leucemia Mielógena Crónica BCR-ABL Positiva/mortalidad , Masculino , Persona de Mediana Edad , Neoplasia Residual , ARN Mensajero/análisis , Inducción de Remisión , Privación de Tratamiento
5.
PLoS One ; 13(11): e0206620, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-30408048

RESUMEN

The increasing interest in exploring the human genome and identifying genetic risk factors contributing to the susceptibility to and outcome of diseases has supported the rapid development of genome-wide techniques. However, the large amount of obtained data requires extensive bioinformatics analysis. In this work, we established an approach combining amplified fragment length polymorphism (AFLP), AFLP in silico and next generation sequencing (NGS) methods to map the malignant genome of patients with chronic myeloid leukemia. We compared the unique DNA fingerprints of patients generated by the AFLP technique approach with those of healthy donors to identify AFLP markers associated with the disease and/or the response to treatment with imatinib, a tyrosine kinase inhibitor. Among the statistically significant AFLP markers selected for NGS analysis and virtual fingerprinting, we identified the sequences of three fragments in the region of DNA repeat element OldhAT1, LINE L1M7, LTR MER90, and satellite ALR/Alpha among repetitive elements, which may indicate a role of these non-coding repetitive sequences in hematological malignancy. SNPs leading to the presence/absence of these fragments were confirmed by Sanger sequencing. When evaluating the results of AFLP analysis for some fragments, we faced the frequently discussed size homoplasy, resulting in co-migration of non-identical AFLP fragments that may originate from an insertion/deletion, SNP, somatic mutation anywhere in the genome, or combination thereof. The AFLP-AFLP in silico-NGS procedure represents a smart alternative to microarrays and relatively expensive and bioinformatically challenging whole-genome sequencing to detect the association of variable regions of the human genome with diseases.


Asunto(s)
Análisis del Polimorfismo de Longitud de Fragmentos Amplificados/métodos , ADN de Neoplasias/genética , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Secuencias Repetitivas de Ácidos Nucleicos , Análisis de Secuencia de ADN/métodos , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Antineoplásicos/uso terapéutico , Secuencia de Bases , Estudios de Casos y Controles , Estudios de Cohortes , Biología Computacional/métodos , Simulación por Computador , Dermatoglifia del ADN/métodos , Femenino , Genoma Humano , Humanos , Mesilato de Imatinib/uso terapéutico , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Masculino , Persona de Mediana Edad , Inhibidores de Proteínas Quinasas/uso terapéutico , Adulto Joven
7.
Curr Hematol Malig Rep ; 8(1): 28-36, 2013 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-23296408

RESUMEN

The efficacy of therapeutic modalities in chronic myeloid leukemia (CML) depends on both genetic and epigenetic mechanisms. This review focuses on epigenetic mechanisms involved in the pathogenesis of CML and in resistance of tumor cells to tyrosine kinase inhibitors leading to the leukemic clone escape and propagation. Regulatory events at the levels of gene regulation by transcription factors and microRNAs are discussed in the context of CML pathogenesis and therapeutic modalities.


Asunto(s)
Epigenómica , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , MicroARNs/fisiología , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Metilación de ADN , Resistencia a Antineoplásicos , Proteínas de Fusión bcr-abl/antagonistas & inhibidores , Proteínas de Fusión bcr-abl/genética , Proteínas de Fusión bcr-abl/uso terapéutico , Regulación Leucémica de la Expresión Génica/efectos de los fármacos , Humanos , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Leucemia Mielógena Crónica BCR-ABL Positiva/patología , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/uso terapéutico
8.
Environ Toxicol Pharmacol ; 36(3): 989-96, 2013 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-24095716

RESUMEN

Benzo[a]pyrene (BaP) is a human carcinogen requiring metabolic activation prior to reaction with DNA. Cytochrome P450 (CYP) 1A1 is the most important hepatic and intestinal enzyme in both BaP activation and detoxification. CYP1A2 is also capable of oxidizing BaP, but to a lesser extent. The induction of CYP1A1/2 by BaP and/or ß-naphthoflavone in liver and small intestine of rats was investigated. Both BaP and ß-naphthoflavone induced CYP1A expression and increased enzyme activities in both organs. Moreover, the induction of CYP1A enzyme activities resulted in an increase in formation of BaP-DNA adducts detected by (32)P-postlabeling in rat liver and in the distal part of small intestine in vivo. The increases in CYP1A enzyme activity were also associated with bioactivation of BaP and elevated BaP-DNA adduct levels in ex vivo incubations of microsomes of both organs with DNA and BaP. These findings indicate a stimulating effect of both compounds on BaP-induced carcinogenesis.


Asunto(s)
Benzo(a)pireno/toxicidad , Citocromo P-450 CYP1A1/metabolismo , Aductos de ADN/metabolismo , Contaminantes Ambientales/toxicidad , Animales , Western Blotting , Cromatografía en Capa Delgada , Citocromo P-450 CYP1A2/biosíntesis , Citocromo P-450 CYP1A2/genética , Activación Enzimática/efectos de los fármacos , Intestino Delgado/efectos de los fármacos , Intestino Delgado/enzimología , Intestino Delgado/metabolismo , Isoenzimas/efectos de los fármacos , Isoenzimas/metabolismo , Hígado/efectos de los fármacos , Hígado/enzimología , Hígado/metabolismo , Masculino , Microsomas Hepáticos/efectos de los fármacos , Microsomas Hepáticos/enzimología , Microsomas Hepáticos/metabolismo , Ratas , Ratas Wistar , beta-naftoflavona/farmacología
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