RESUMEN
MicroRNAs and heterogeneous ribonucleoproteins (hnRNPs) are posttranscriptional gene regulators that bind mRNA in a sequence-specific manner. Here, we report that loss of miR-328 occurs in blast crisis chronic myelogenous leukemia (CML-BC) in a BCR/ABL dose- and kinase-dependent manner through the MAPK-hnRNP E2 pathway. Restoration of miR-328 expression rescues differentiation and impairs survival of leukemic blasts by simultaneously interacting with the translational regulator poly(rC)-binding protein hnRNP E2 and with the mRNA encoding the survival factor PIM1, respectively. The interaction with hnRNP E2 is independent of the microRNA's seed sequence and it leads to release of CEBPA mRNA from hnRNP E2-mediated translational inhibition. Altogether, these data reveal the dual ability of a microRNA to control cell fate both through base pairing with mRNA targets and through a decoy activity that interferes with the function of regulatory proteins.
Asunto(s)
Ribonucleoproteínas Nucleares Heterogéneas/genética , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , MicroARNs/metabolismo , Animales , Crisis Blástica , Proteínas Potenciadoras de Unión a CCAAT/metabolismo , Línea Celular Tumoral , Ribonucleoproteínas Nucleares Heterogéneas/metabolismo , Humanos , Leucemia Mielógena Crónica BCR-ABL Positiva/metabolismo , Ratones , Proteínas Proto-Oncogénicas c-pim-1/metabolismo , Complejo Silenciador Inducido por ARN/metabolismoRESUMEN
Myelofibrosis is a heterogeneous myeloproliferative neoplasm characterized by chronic inflammation, progressive bone marrow failure, and hepatosplenic extramedullary hematopoiesis. Treatments like Janus kinase inhibitor monotherapy (e.g., ruxolitinib) provide significant spleen and symptom relief but demonstrate limited ability to lead to a durable disease modification. There is an urgent unmet medical need for treatments with a novel mechanism of action that can modify the underlying pathophysiology and affect the disease course of myelofibrosis. This review highlights the role of B-cell lymphoma (BCL) protein BCL-extra large (BCL-XL ) in disease pathogenesis and the potential role that navitoclax, a BCL-extra large/BCL-2 inhibitor, may have in myelofibrosis treatment.
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Antineoplásicos , Inhibidores de las Cinasas Janus , Mielofibrosis Primaria , Humanos , Mielofibrosis Primaria/tratamiento farmacológico , Inhibidores de las Cinasas Janus/farmacología , Inhibidores de las Cinasas Janus/uso terapéutico , Janus Quinasa 2 , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Proteínas Proto-Oncogénicas c-bcl-2 , Nitrilos/uso terapéuticoRESUMEN
Patients with relapsed/refractory (R/R) higher-risk myelodysplastic syndromes (MDS) have a dismal median overall survival (OS) after failing hypomethylating agent (HMA) treatment. There is no standard of care for patients after HMA therapy failure; hence, there is a critical need for effective therapeutic strategies. Herein, we present the safety and efficacy of venetoclax + azacitidine in patients with R/R MDS. This phase 1b, open-label, multicenter study enrolled patients ≥18 years. Patients were treated with escalating doses of oral venetoclax: 100, 200, or 400 mg daily for 14 days every 28-day cycle. Azacitidine was administered on Days 1-7 every cycle at 75 mg/m2 /day intravenously/subcutaneously. Responses were assessed per modified 2006 International Working Group (IWG) criteria. Forty-four patients (male 86%, median age 74 years) received venetoclax + azacitidine treatment. Median follow-up was 21.2 months. Hematological adverse events of Grade ≥ 3 included febrile neutropenia (34%), thrombocytopenia (32%), neutropenia (27%), and anemia (18%). Pneumonia (23%) was the most common Grade ≥ 3 infection. Marrow responses were seen including complete remission (CR, n = 3, 7%) and marrow CR (mCR, n = 14, 32%); 36% (16/44) achieved transfusion independence (TI) for RBCs and/or platelets, and 43% (6/14) with mCR achieved hematological improvement (HI). The median time to CR/mCR was 1.2 months, and the median duration of response for CR + mCR was 8.6 months. Median OS was 12.6 months. Venetoclax + azacitidine shows activity in patients with R/R MDS following prior HMA therapy failure and provides clinically meaningful benefits, including HI and TI, and encouraging OS.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica , Síndromes Mielodisplásicos , Anciano , Humanos , Masculino , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Azacitidina/uso terapéutico , Leucemia Mieloide Aguda/tratamiento farmacológico , Síndromes Mielodisplásicos/tratamiento farmacológico , Neutropenia/inducido químicamente , Sulfonamidas , Resultado del Tratamiento , FemeninoRESUMEN
As tyrosine kinase inhibitors (TKIs) fail to induce long-term response in blast crisis chronic myelogenous leukemia (CML-BC) and Philadelphia chromosome-positive (Ph(+)) acute lymphoblastic leukemia (ALL), novel therapies targeting leukemia-dysregulated pathways are necessary. Exportin-1 (XPO1), also known as chromosome maintenance protein 1, regulates cell growth and differentiation by controlling the nucleocytoplasmic trafficking of proteins and RNAs, some of which are aberrantly modulated in BCR-ABL1(+) leukemias. Using CD34(+) progenitors from CML, B-ALL, and healthy individuals, we found that XPO1 expression was markedly increased, mostly in a TKI-sensitive manner, in CML-BC and Ph(+) B-ALL. Notably, XPO1 was also elevated in Ph(-) B-ALL. Moreover, the clinically relevant XPO1 inhibitor KPT-330 strongly triggered apoptosis and impaired the clonogenic potential of leukemic, but not normal, CD34(+) progenitors, and increased survival of BCR-ABL1(+) mice, 50% of which remained alive and, mostly, became BCR-ABL1 negative. Moreover, KPT-330 compassionate use in a patient with TKI-resistant CML undergoing disease progression significantly reduced white blood cell count, blast cells, splenomegaly, lactate dehydrogenase levels, and bone pain. Mechanistically, KPT-330 altered the subcellular localization of leukemia-regulated factors including RNA-binding heterogeneous nuclear ribonucleoprotein A1 and the oncogene SET, thereby inducing reactivation of protein phosphatase 2A tumor suppressor and inhibition of BCR-ABL1 in CML-BC cells. Because XPO1 is important for leukemic cell survival, KPT-330 may represent an alternative therapy for TKI-refractory Ph(+) leukemias.
Asunto(s)
Antineoplásicos/farmacología , Regulación Leucémica de la Expresión Génica/efectos de los fármacos , Hidrazinas/farmacología , Carioferinas/antagonistas & inhibidores , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamiento farmacológico , Receptores Citoplasmáticos y Nucleares/antagonistas & inhibidores , Triazoles/farmacología , Adulto , Animales , Antígenos CD34/genética , Antígenos CD34/metabolismo , Apoptosis/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Ensayos Clínicos Fase I como Asunto , Proteínas de Unión al ADN , Evaluación Preclínica de Medicamentos , Proteínas de Fusión bcr-abl/antagonistas & inhibidores , Proteínas de Fusión bcr-abl/genética , Proteínas de Fusión bcr-abl/metabolismo , Chaperonas de Histonas/antagonistas & inhibidores , Chaperonas de Histonas/genética , Chaperonas de Histonas/metabolismo , Humanos , Carioferinas/genética , Carioferinas/metabolismo , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Leucemia Mielógena Crónica BCR-ABL Positiva/patología , Masculino , Ratones , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/patología , Inhibidores de Proteínas Quinasas/farmacología , Transporte de Proteínas , Receptores Citoplasmáticos y Nucleares/genética , Receptores Citoplasmáticos y Nucleares/metabolismo , Ribonucleoproteínas/antagonistas & inhibidores , Ribonucleoproteínas/genética , Ribonucleoproteínas/metabolismo , Factores de Transcripción/antagonistas & inhibidores , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Proteína Exportina 1RESUMEN
FTY720 (Fingolimod, Gilenya) is a sphingosine analog used as an immunosuppressant in multiple sclerosis patients. FTY720 is also a potent protein phosphatase 2A (PP2A)-activating drug (PAD). PP2A is a tumor suppressor found inactivated in different types of cancer. We show here that PP2A is inactive in polycythemia vera (PV) and other myeloproliferative neoplasms characterized by the expression of the transforming Jak2(V617F) oncogene. PP2A inactivation occurs in a Jak2(V617F) dose/kinase-dependent manner through the PI-3Kγ-PKC-induced phosphorylation of the PP2A inhibitor SET. Genetic or PAD-mediated PP2A reactivation induces Jak2(V617F) inactivation/downregulation and impairs clonogenic potential of Jak2(V617F) cell lines and PV but not normal CD34(+) progenitors. Likewise, FTY720 decreases leukemic allelic burden, reduces splenomegaly, and significantly increases survival of Jak2(V617F) leukemic mice without adverse effects. Mechanistically, we show that in Jak2(V617F) cells, FTY720 antileukemic activity requires neither FTY720 phosphorylation (FTY720-P) nor SET dimerization or ceramide induction but depends on interaction with SET K209. Moreover, we show that Jak2(V617F) also utilizes an alternative sphingosine kinase-1-mediated pathway to inhibit PP2A and that FTY720-P, acting as a sphingosine-1-phosphate-receptor-1 agonist, elicits signals leading to the Jak2-PI-3Kγ-PKC-SET-mediated PP2A inhibition. Thus, PADs (eg, FTY720) represent suitable therapeutic alternatives for Jak2(V617F) MPNs.
Asunto(s)
Janus Quinasa 2/metabolismo , Leucemia/tratamiento farmacológico , Glicoles de Propileno/farmacología , Proteína Fosfatasa 2/metabolismo , Esfingosina/análogos & derivados , Animales , Línea Celular Transformada , Línea Celular Tumoral , Células Cultivadas , Fosfatidilinositol 3-Quinasa Clase Ib , Proteínas de Unión al ADN , Activación Enzimática/efectos de los fármacos , Clorhidrato de Fingolimod , Chaperonas de Histonas , Humanos , Immunoblotting , Inmunosupresores/farmacología , Janus Quinasa 2/genética , Estimación de Kaplan-Meier , Leucemia/genética , Leucemia/patología , Ratones , Ratones SCID , Mutación , Proteínas Oncogénicas/genética , Proteínas Oncogénicas/metabolismo , Proteína Quinasa C/genética , Proteína Quinasa C/metabolismo , Proteína Fosfatasa 2/genética , Interferencia de ARN , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transducción de Señal/efectos de los fármacos , Esfingosina/farmacología , Resultado del TratamientoRESUMEN
BACKGROUND: Primary analyses of cohort 1a of the REFINE trial showed that addition of navitoclax to ruxolitinib induced a 35% or greater reduction in spleen volume (SVR35) and reduced symptoms in patients with myelofibrosis no longer benefiting from ruxolitinib. Here, we report the exploratory post-hoc biomarker analyses from cohort 1a. METHODS: REFINE is a phase 2, multicentre, open-label trial designed to assess the activity and safety of navitoclax alone or in combination with ruxolitinib in patients with primary or secondary (post-polycythaemia vera or post-essential thrombocythaemia) myelofibrosis. Cohort 1a of the study included patients who had disease progression or suboptimal response on stable ruxolitinib monotherapy. Patients in cohort 1a, who had previously received ruxolitinib for 12 weeks or more, continued their current stable dose, and navitoclax was orally administered at 50 mg per day and escalated weekly to a maximum of 300 mg per day, based on tolerability. The primary activity endpoint was SVR35 at week 24 from baseline. Secondary endpoints were a 50% or greater reduction in total symptom score (TSS50) at week 24 from baseline as measured by the Myelofibrosis Symptom Assessment Form (version 4.0), anaemia response assessed according to International Working Group-Myeloproliferative Neoplasms Research and European LeukemiaNet criteria, and change in grade of bone marrow fibrosis according to the European consensus grading system; and exploratory endpoints included overall survival and changes in inflammatory cytokines. Exploratory analyses investigated potential prognostic biomarkers of the benefit of navitoclax-based combination treatment, including bone marrow fibrosis and variant allele frequency, in patients with a suboptimal response to ruxolitinib. This study is registered with ClinicalTrials.gov (NCT03222609) and is ongoing. FINDINGS: Between Nov 14, 2017, and April 10, 2019, 34 patients in cohort 1a received at least one dose of navitoclax plus ruxolitinib. 23 (68%) patients were male, with 32 (94%) being White. At data cutoff (May 6, 2021), the median follow-up for survivors was 26·2 months (IQR 21·9-32·3). 33 patients were evaluable for biomarker analyses; 19 (58%) had high molecular risk mutations. Five (31%) of 16 patients had SVR35 at week 24 in the high molecular risk group, as did four (31%) of 13 in the non-high molecular risk group. Four (36%) of 11 patients in the high molecular risk group had TSS50 at week 24 compared with two (25%) of eight in the non-high molecular risk group; seven (39%) of 18 in the high molecular risk group had an improvement in fibrosis by at least one grade compared with five (36%) of 14 in the non-high molecular risk group; and four (28%) of 14 had reductions in variant allele frequency of 20% or greater in the high molecular risk group compared with two (17%) of 12 in the non-high molecular risk group. Patients with improvements in fibrosis of one grade or more and a reduction of 20% of more in variant allele frequency had improved overall survival (median overall survival not reached) compared with those who did not achieve fibrosis improvement or a reduction in variant allele frequency (median overall survival 28·5 months [95% CI 19·6-not estimable] for both), suggesting potential disease modification. Additionally, changes in concentrations of ß-2-microglobulin (week 12: r=0·57; week 24: r=0·57), TIMP metallopeptidase inhibitor 1 (week 12: r=0·47; week 24: r=0·54), TNF receptor type II (r=0·55; week 24: r=0·40), and vascular cell adhesion molecule-1 (r=0·58; week 24: r=0·50) were positively associated with changes in spleen volume. INTERPRETATION: These biomarker analyses reveal clinically meaningful splenic responses independent of high molecular risk mutation status in patients treated with navitoclax plus ruxolitinib who were not benefiting from ruxolitinib monotherapy. Furthermore, the overall survival benefit observed in those with an improvement in fibrosis or a reduction in variant allele frequency is suggestive of disease modification, implying the therapeutic potential of adding navitoclax to ruxolitinib for patients with myelofibrosis who had disease progression or suboptimal response to ruxolitinib monotherapy. FUNDING: AbbVie.
Asunto(s)
Mielofibrosis Primaria , Compuestos de Anilina , Biomarcadores , Progresión de la Enfermedad , Femenino , Fibrosis , Humanos , Masculino , Nitrilos , Mielofibrosis Primaria/tratamiento farmacológico , Mielofibrosis Primaria/genética , Pirazoles , Pirimidinas , SulfonamidasRESUMEN
Persistence of drug-resistant quiescent leukemic stem cells (LSC) and impaired natural killer (NK) cell immune response account for relapse of chronic myelogenous leukemia (CML). Inactivation of protein phosphatase 2A (PP2A) is essential for CML-quiescent LSC survival and NK cell antitumor activity. Here we show that MIR300 has antiproliferative and PP2A-activating functions that are dose dependently differentially induced by CCND2/CDK6 and SET inhibition, respectively. MIR300 is upregulated in CML LSCs and NK cells by bone marrow microenvironment (BMM) signals to induce quiescence and impair immune response, respectively. Conversely, BCR-ABL1 downregulates MIR300 in CML progenitors to prevent growth arrest and PP2A-mediated apoptosis. Quiescent LSCs escape apoptosis by upregulating TUG1 long noncoding RNA that uncouples and limits MIR300 function to cytostasis. Genetic and pharmacologic MIR300 modulation and/or PP2A-activating drug treatment restore NK cell activity, inhibit BMM-induced growth arrest, and selectively trigger LSC apoptosis in vitro and in patient-derived xenografts; hence, the importance of MIR300 and PP2A activity for CML development and therapy.
Asunto(s)
Leucemia Mielógena Crónica BCR-ABL Positiva , MicroARNs , Humanos , Células Asesinas Naturales , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , MicroARNs/genética , Células Madre Neoplásicas , Inhibidores de Proteínas Quinasas/metabolismo , Proteína Fosfatasa 2/genética , Microambiente Tumoral/genéticaRESUMEN
BACKGROUND: Activating mutations [internal tandem duplication (ITD)] or overexpression of the FMS-like tyrosine kinase receptor-3 (FLT3) gene are associated with poor outcome in acute myeloid leukemia (AML) patients, underscoring the need for novel therapeutic approaches. The natural product silvestrol has potent antitumor activity in several malignancies, but its therapeutic impact on distinct molecular high-risk AML subsets remains to be fully investigated. We examined here the preclinical activity of silvestrol in FLT3-ITD and FLT3 wild-type (wt) AML. METHODS: Silvestrol in vitro anti-leukemic activity was examined by colorimetric cell viability assay, colony-forming and flow cytometry assays assessing growth inhibition and apoptosis, respectively. Pharmacological activity of silvestrol on FLT3 mRNA translation, mRNA and protein expression was determined by RNA-immunoprecipitation, qRT-PCR and immunoblot analyses, respectively. Silvestrol in vivo efficacy was investigated using MV4-11 leukemia-engrafted mice. RESULTS: Silvestrol shows antileukemia activity at nanomolar concentrations both in FLT3-wt overexpressing (THP-1) and FLT3-ITD (MV4-11) expressing AML cell lines (IC50 = 3.8 and 2.7 nM, respectively) and patients' primary blasts [IC50 = ~12 nM (FLT3-wt) and ~5 nM (FLT3-ITD)]. Silvestrol increased apoptosis (~4fold, P = 0.0001), and inhibited colony-formation (100%, P < 0.0001) in primary blasts. Silvestrol efficiently inhibited FLT3 translation reducing FLT3 protein expression by 80-90% and decreased miR-155 levels (~60%), a frequently co-regulated onco-miR in FLT3-ITD-positive AML. The median survival of silvestrol-treated vs vehicle-treated mice was 63 vs 29 days post-engraftment, respectively (P < 0.0001). CONCLUSIONS: Silvestrol exhibits significant in vivo and in vitro antileukemic activities in AML through a novel mechanism resulting in inhibition of FLT3 and miR-155 expression. These encouraging results warrant a rapid translation of silvestrol for clinical testing in AML.
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Antineoplásicos/farmacología , Regulación Leucémica de la Expresión Génica/efectos de los fármacos , Subunidad gamma Común de Receptores de Interleucina/fisiología , Leucemia Mieloide Aguda/tratamiento farmacológico , MicroARNs/antagonistas & inhibidores , Triterpenos/farmacología , Tirosina Quinasa 3 Similar a fms/antagonistas & inhibidores , Animales , Apoptosis/efectos de los fármacos , Crisis Blástica/tratamiento farmacológico , Crisis Blástica/metabolismo , Crisis Blástica/patología , Western Blotting , Proliferación Celular/efectos de los fármacos , Citometría de Flujo , Humanos , Técnicas In Vitro , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patología , Ratones , Ratones Endogámicos NOD , Ratones SCID , MicroARNs/genética , ARN Mensajero/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Células Tumorales Cultivadas , Tirosina Quinasa 3 Similar a fms/genética , Tirosina Quinasa 3 Similar a fms/metabolismoRESUMEN
The success of tyrosine kinase inhibitors (TKIs) in treating chronic myeloid leukemia (CML) depends on the requirement for BCR-ABL1 kinase activity in CML progenitors. However, CML quiescent HSCs are TKI resistant and represent a BCR-ABL1 kinase-independent disease reservoir. Here we have shown that persistence of leukemic HSCs in BM requires inhibition of the tumor suppressor protein phosphatase 2A (PP2A) and expression--but not activity--of the BCR-ABL1 oncogene. Examination of HSCs from CML patients and healthy individuals revealed that PP2A activity was suppressed in CML compared with normal HSCs. TKI-resistant CML quiescent HSCs showed increased levels of BCR-ABL1, but very low kinase activity. BCR-ABL1 expression, but not kinase function, was required for recruitment of JAK2, activation of a JAK2/ß-catenin survival/self-renewal pathway, and inhibition of PP2A. PP2A-activating drugs (PADs) markedly reduced survival and self-renewal of CML quiescent HSCs, but not normal quiescent HSCs, through BCR-ABL1 kinase-independent and PP2A-mediated inhibition of JAK2 and ß-catenin. This led to suppression of human leukemic, but not normal, HSC/progenitor survival in BM xenografts and interference with long-term maintenance of BCR-ABL1-positive HSCs in serial transplantation assays. Targeting the JAK2/PP2A/ß-catenin network in quiescent HSCs with PADs (e.g., FTY720) has the potential to treat TKI-refractory CML and relieve lifelong patient dependence on TKIs.
Asunto(s)
Antineoplásicos/farmacología , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Células Madre Neoplásicas/efectos de los fármacos , Inhibidores de Proteínas Quinasas/farmacología , Proteína Fosfatasa 2/metabolismo , Animales , Apoptosis , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Resistencia a Antineoplásicos , Activadores de Enzimas/farmacología , Clorhidrato de Fingolimod , Proteínas de Fusión bcr-abl/metabolismo , Células Madre Hematopoyéticas/efectos de los fármacos , Células Madre Hematopoyéticas/enzimología , Humanos , Janus Quinasa 2/metabolismo , Células K562 , Ratones , Ratones Transgénicos , Células Madre Neoplásicas/enzimología , Glicoles de Propileno/farmacología , Esfingosina/análogos & derivados , Esfingosina/farmacología , Vía de Señalización Wnt , Ensayos Antitumor por Modelo de Xenoinjerto , beta Catenina/metabolismoRESUMEN
The use of first- and second-generation tyrosine kinase inhibitors (TKIs) significantly improves prognosis for patients with early chronic phase chronic myeloid leukemia (CML) and efficiently counteracts leukemia in most patients with CML bearing a disease characterized by the expression of BCR-ABL1 mutants. However, the so-called 'tinib' TKIs (e.g. imatinib, nilotinib, dasatinib, and bosutinib) are both ineffective in patients who undergo blastic transformation and unable to eradicate CML at the stem cell level. This raises a few important questions. Is BCR-ABL1 expression and/or activity essential for blastic transformation? Is blastic transformation the result of genetic or epigenetic events that occur at the stem cell level which only become apparent in the granulocyte-macrophage progenitor (GMP) cell pool, or does it arise directly at the GMP level? As altered mRNA metabolism contributes to the phenotype of blast crisis CML progenitors (decreased translation of tumor suppressor genes and transcription factors essential for terminal differentiation and increased translation of anti-apoptotic genes), one attractive concept is to restore levels of these essential molecules to their normal levels. In this review, we discuss the mechanisms by which mRNA processing, translation, and degradation are deregulated in BCR-ABL1 myeloid blast crisis CML progenitors, and present encouraging results from studies with pharmacologic inhibitors which support their inclusion in the clinic.
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Antineoplásicos/uso terapéutico , Proteínas de Fusión bcr-abl/metabolismo , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , ARN Mensajero/metabolismo , Proteínas de Fusión bcr-abl/genética , Regulación Leucémica de la Expresión Génica , Humanos , MicroARNs/genética , MicroARNs/metabolismo , Proteínas de Unión al ARN/genética , Proteínas de Unión al ARN/metabolismoRESUMEN
The kinase inhibitors imatinib mesylate and dasatinib are the preferred treatment for Philadelphia chromosome-positive (Ph+) leukemias, and they are highly successful in the chronic phase of chronic myeloid leukemia (CML). However, they are not efficient in Ph+ B-cell acute lymphoblastic leukemia (B-ALL). Ph+ leukemia cells are highly resistant to apoptosis, and evidence from cell lines and primary cells suggest Bcl-xL as a critical mediator of resistance to apoptosis: however, this concept has never been rigorously tested in an animal model. To clarify the role of Bcl-xL in Ph+ B-ALL, we generated 2 mouse models. In the first model, Ph+ B-ALL and loss of Bcl-xL expression are coinduced; in the second model, leukemia is induced with expression of Bcl-xL protein well above the levels found in wild-type lymphoblasts. Deletion of Bcl-xL did not inhibit leukemogenesis or affect apoptosis, but increased cellular proliferation. Consistent with this result, overexpression of Bcl-xL led to decreased cellular proliferation. These models reveal an unexpected role for Bcl-xL in cell-cycle entry and the proliferation of tumor cells.
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Ciclo Celular , Leucemia de Células B/metabolismo , Cromosoma Filadelfia , Leucemia-Linfoma Linfoblástico de Células Precursoras/metabolismo , Proteína bcl-X/biosíntesis , Animales , Ciclo Celular/genética , Modelos Animales de Enfermedad , Regulación Leucémica de la Expresión Génica/genética , Humanos , Leucemia de Células B/genética , Leucemia de Células B/patología , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Leucemia Mielógena Crónica BCR-ABL Positiva/metabolismo , Leucemia Mielógena Crónica BCR-ABL Positiva/patología , Ratones , Ratones Noqueados , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/patología , Proteína bcl-X/genéticaRESUMEN
We describe the development of a novel method for real-time in situ characterization of polycyclic aromatic hydrocarbons (PAHs) in submerged freshwater sediments. Laser-induced fluorescence (LIF) spectroscopy, a mature technique for PAH characterization in terrestrial sediments, was adapted for shipboard use. A cone penetrometer-type apparatus was designed for probe penetration at a constant rate (1 cm/s) to a depth of 3 m. A field-portable LIF system was used for in situ measurements in which the output of a pulsed excimer laser was transmitted by optical fiber to a sapphire window (6.4-mm o.d.) in the probe wall; fluorescent emission was collected by a separate optical fiber for transmission to the spectrometer on deck. Four wavelengths (340, 390, 440, 490 nm) were selected via optical delay lines, and multiple-wavelength waveforms were created. These multiple-wavelength waveforms contain information on the fluorescence frequency, intensity, and emission decay rate. Field testing was conducted at 10 sites in Milwaukee Harbor (total PAH concentrations ranged from approximately 10 to 650 microg/g); conventional sediment core samples were collected concurrently. The core samples were analyzed by EPA methods 3545 (pressurized fluid extraction, PFE) and 8270C (gas chromatography-mass spectrometry, GC-MS) for PAHs. A partial least-squares regression (PLSR) model wasthen created based on laboratory LIF measurements and PFE-GC-MS of the core samples. The PLSR model was applied to the in situ field test data, and 13 of the 16 EPA-regulated PAHs were quantified with a relative error of <30% overall (the remaining three PAHs were found at levels insufficient to quantify). We additionally describe preliminary source apportionment relationships that were revealed by the PLSR model for the in situ LIF measurements.