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1.
Int J Mol Sci ; 23(7)2022 Apr 04.
Artigo em Inglês | MEDLINE | ID: mdl-35409373

RESUMO

Acute myeloid leukemia (AML) accounts for around 20% of diagnosed childhood leukemia. Cytarabine (CYT) is involved in the AML treatment regimen. AML and CYT showed impairment in spermatogenesis in human and rodents in adulthood. We successfully developed an AML disease model in sexually immature mice. Monocytes and granulocytes were examined in all groups: untreated control, AML alone, CYT alone and AML+CYT (in combination). There was a significant increase in the counts of monocytes and granulocytes in the AML-treated immature mice (AML) compared to the control, and AML cells were demonstrated in the blood vessels of the testes. AML alone and CYT alone impaired the development of spermatogenesis at the adult age of the AML-treated immature mice. The damage was clear in the structure/histology of their seminiferous tubules, and an increase in the apoptotic cells of the seminiferous tubules was demonstrated. Our results demonstrated a significant decrease in the meiotic/post-meiotic cells compared to the control. However, CYT alone (but not AML) significantly increased the count of spermatogonial cells (premeiotic cells) that positively stained with SALL4 and PLZF per tubule compared to the control. Furthermore, AML significantly increased the count of proliferating spermatogonial cells that positively stained with PCNA in the seminiferous tubules compared to the control, whereas CYT significantly decreased the count compared to the control. Our result showed that AML and CYT affected the microenvironment/niche of the germ cells. AML significantly decreased the levels growth factors, such as SCF, GDNF and MCSF) compared to control, whereas CYT significantly increased the levels of MCSF and GDNF compared to control. In addition, AML significantly increased the RNA expression levels of testicular IL-6 (a proinflammatory cytokine), whereas CYT significantly decreased testicular IL-6 levels compared to the control group. Furthermore, AML alone and CYT alone significantly decreased RNA expression levels of testicular IL-10 (anti-inflammatory cytokine) compared to the control group. Our results demonstrate that pediatric AML disease with or without CYT treatment impairs spermatogenesis at adult age (the impairment was more pronounced in AML+CYT) compared to control. Thus, we suggest that special care should be considered for children with AML who are treated with a CYT regimen regarding their future fertility at adult age.


Assuntos
Citarabina , Leucemia Mieloide Aguda , Adulto , Animais , Criança , Citarabina/metabolismo , Citarabina/farmacologia , Citarabina/uso terapêutico , Fator Neurotrófico Derivado de Linhagem de Célula Glial/metabolismo , Humanos , Interleucina-6/metabolismo , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/metabolismo , Masculino , Camundongos , RNA/metabolismo , Túbulos Seminíferos/metabolismo , Espermatogênese , Espermatogônias/metabolismo , Testículo/metabolismo , Microambiente Tumoral
2.
Cell Death Dis ; 13(4): 379, 2022 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-35443722

RESUMO

Venetoclax plus cytarabine therapy is approved for elderly acute myeloid leukemia (AML) patients and needs further improvement. We studied the mechanisms of venetoclax plus cytarabine treatment and searched for a third agent to enhance their effects. Cytarabine induces S phase arrest-mediated DNA damage with activation of DNA replication checkpoint kinase 1 (Chk1) through phosphorylation, while venetoclax induces B cell lymphoma 2 (Bcl-2)-interacting mediator of cell death (Bim)-mediated apoptotic DNA damage. Myeloid cell leukemia-1 (Mcl-1) plays negative roles in both events by sequestering Bim and accelerating Chk1 phosphorylation. Venetoclax releases Bim from Bcl-2 with increased Bim binding to Mcl-1. Artesunate, an antimalaria drug, induces Noxa to replace Bim from Mcl-1 and induces synergistic apoptosis with venetoclax accompanied with Mcl-1 reduction. Silencing Mcl-1 or adding venetoclax/artesunate diminishes the cytarabine resistance pathway p-Chk1. The triple combination exhibits S phase arrest with enhanced DNA damage, improves AML colony formation inhibition, and prolongs survival of two mice xenograft models compared to the venetoclax/cytarabine dual combination. Artesunate serves as a bridge for venetoclax and cytarabine combination by Noxa and Bim-mediated apoptosis and Mcl-1 reduction. We provide a new triple combination for AML treatment by targeting the Noxa/Mcl-1/Bim axis to reverse Mcl-1/p-Chk1 resistance of cytarabine therapy.


Assuntos
Citarabina , Leucemia Mieloide Aguda , Idoso , Animais , Apoptose , Proteínas Reguladoras de Apoptose , Artesunato/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/uso terapêutico , Linhagem Celular Tumoral , Quinase 1 do Ponto de Checagem , Citarabina/farmacologia , Citarabina/uso terapêutico , Humanos , Leucemia Mieloide Aguda/patologia , Camundongos , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Sulfonamidas
3.
PLoS One ; 17(4): e0267508, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35486629

RESUMO

Acute myeloid leukemia (AML) is characterized by blocked differentiation and extensive proliferation of hematopoietic progenitors/precursors. Relapse is often observed after chemotherapy due to the presence of residual leukemic cells, which is also called minimal residual disease (MRD). Subclonal heterogeneity at diagnosis was found to be responsible for MRD after treatment. Patient xenograft mouse models are valuable tools for studying MRD after chemotherapy; however, the contribution of the immune system in these models is usually missing. To evaluate its role in leukemic persistence, we generated an immune-competent AML mouse model of persistence after chemotherapy treatment. We used well-characterized (phenotypically and genetically) subclones of the murine C1498 cell line stably expressing the ZsGreen reporter gene and the WT1 protein, a valuable antigen. Accordingly, these subclones were also selected due to their in vitro aracytidine (Ara-c) sensitivity. A combination of 3 subclones (expressing or not expressing WT1) was found to lead to prolonged mouse survival after Ara-c treatment (as long as 150 days). The presence of residual leukemic cells in the blood and BM of surviving mice indicated their persistence. Thus, a new mouse model that may offer insights into immune contributions to leukemic persistence was developed.


Assuntos
Leucemia Mieloide Aguda , Animais , Citarabina/farmacologia , Citarabina/uso terapêutico , Modelos Animais de Doenças , Progressão da Doença , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/genética , Camundongos , Neoplasia Residual/diagnóstico
4.
Exp Cell Res ; 415(1): 113112, 2022 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-35346671

RESUMO

Chemoresistance contributes to poor survival and high relapse risk in acute myeloid leukemia (AML). As a pro-inflammatory cytokine, interleukin-6 (IL-6) plays a vital role in the chemoresistance of malignancies. However, the underlying mechanisms of chemoresistance in AML have not been widely studied. Lipid metabolism, which contributes to chemoresistance in AML, is enhanced by IL-6 in skeletal muscle cells. We hypothesized that IL-6 promotes the chemoresistance of AML by promoting lipid metabolism. Based on the positive correlation between IL-6 receptor expression and the cellular response to exogenous IL-6, we performed Gene Ontology analysis of a dataset consisting the information of 151 AML patients from The Cancer Genome Atlas. We found that lipid transport-associated genes were upregulated in the high IL-6 receptor expression group. Additionally, IL-6 promoted fatty acid (FA) uptake in both AML cell lines and primary AML cells. Inhibition of FA uptake by sulfo-N-succinimidyl oleate repressed IL-6-induced chemoresistance. Western blotting, quantitative polymerase chain reaction, and chromatin immunoprecipitation assays indicated that IL-6 promoted CD36 expression at both the mRNA and protein levels through stat3 signaling. Knockout of CD36 or stat3 repressed IL-6-induced FA uptake and chemoresistance. Furthermore, in five human AML samples, we validated that compared to CD36-cells, CD36+ primary AML cells were less sensitive to cytosine arabinoside (Ara-c) and that blockade of CD36 re-sensitized CD36+ AML cells to Ara-c. Mice injected with CD36 knockout cells followed by treatment with Ara-c showed markedly decreased leukemia burden and prolonged survival in vivo. Finally, treatment with the CD36 antibody in combination with Ara-c exhibited synergistic effects in vivo. In conclusion, IL-6 promotes chemoresistance in AML through the stat3/CD36-mediated FA uptake. Blockade of CD36 improved the effect of Ara-c, representing a promising strategy for AML therapy.


Assuntos
Interleucina-6 , Leucemia Mieloide Aguda , Animais , Citarabina/metabolismo , Citarabina/farmacologia , Resistencia a Medicamentos Antineoplásicos , Ácidos Graxos , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patologia , Camundongos , Camundongos Knockout , Receptores de Interleucina-6
5.
Exp Hematol ; 110: 20-27, 2022 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-35306047

RESUMO

Cytarabine and other chain-terminating nucleoside analogs that damage replication forks in rapidly proliferating cells are a cornerstone of leukemia chemotherapy, yet the outcomes remain unsatisfactory because of resistance and toxicity. Better understanding of DNA damage repair and downstream effector mechanisms in different disease subtypes can guide combination strategies that sensitize leukemia cells to cytarabine without increasing side effects. We have previously found that mutations in DNMT3A, one of the most commonly mutated genes in acute myeloid leukemia and associated with poor prognosis, predisposed cells to DNA damage and cell killing by cytarabine, cladribine, and other nucleoside analogs, which coincided with PARP1 dysfunction and DNA repair defect (Venugopal K, Feng Y, Nowialis P, et al. Clin Cancer Res 2022;28:756-769). In this article, we first overview DNA repair mechanisms that remove aberrant chain-terminating nucleotides as determinants of sensitivity or resistance to cytarabine and other nucleoside analogs. Next, we discuss PARP inhibition as a rational strategy to increase cytarabine efficacy in cells without DNMT3A mutations, while considering the implications of PARP inhibitor resistance for promoting clonal hematopoiesis. Finally, we examine the utility of p53 potentiators to boost leukemia cell killing by cytarabine in the context of mutant DNMT3A. Systematic profiling of DNA damage repair proficiency has the potential to uncover subtype-specific therapeutic dependencies in AML.


Assuntos
Citarabina , Leucemia Mieloide Aguda , Citarabina/farmacologia , Citarabina/uso terapêutico , Reparo do DNA , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/genética , Mutação , Nucleosídeos/uso terapêutico
6.
Cancer Lett ; 535: 215659, 2022 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-35321842

RESUMO

Adenosine monophosphate activated protein kinase (AMPK) is a master regulator of cell metabolism and is involved in cancer as both a tumor suppressor and a source of resistance to metabolic stress. The role of AMPK in response to chemotherapy has been examined in solid tumor models but remains unclear in acute myeloid leukemia (AML). To determine the role of AMPK in chemotherapy response, AML cell lines were generated lacking AMPK activity. AMPK knock out cells demonstrated significant resistance to cytarabine and doxorubicin both in vitro and in vivo. Mitochondrial mass and function were unchanged in AMPK knockout cells. Mechanistically, AMPK knock out cells demonstrated a diminished DNA damage response with significantly lower γH2AX foci, p53 and p21 induction as well as decreased apoptosis following chemotherapy exposure. Most importantly, TCGA datasets revealed that patients expressing low levels of the PRKAA1 subunit of AMPK had significantly shorter survival. Finally, AML cells were sensitized to chemotherapy with the addition of the AMPK activator AICAR. These data demonstrate that AMPK sensitizes AML cells to chemotherapy and suggests a contribution of the cellular metabolic state to cell fate decisions ultimately affecting therapy response.


Assuntos
Proteínas Quinases Ativadas por AMP , Leucemia Mieloide Aguda , Proteínas Quinases Ativadas por AMP/metabolismo , Monofosfato de Adenosina , Apoptose , Citarabina/farmacologia , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo
7.
Cancer Lett ; 532: 215582, 2022 04 28.
Artigo em Inglês | MEDLINE | ID: mdl-35122876

RESUMO

Interaction between stromal cells and acute myeloid leukemia (AML) cells in bone marrow (BM) is known to contribute importantly to chemoresistance and disease recurrence. Therefore, disruption of a crosstalk between AML cells and BM microenvironment may offer a promising therapeutic strategy for AML treatment. Here, we demonstrate that in a niche-like co-culture system, AML cells took up functional mitochondria from bone marrow stromal cells (BMSCs) and inhibition of such mitochondrial transfer by metformin, the most commonly prescribed drug for type 2 diabetes mellitus, significantly enhanced the chemosensitivity of AML cells co-cultured with BMSCs. The chemo-sensitizing effect of metformin was acted through reducing the mitochondrial transfer and mitochondrial oxidative phosphorylation (OXPHOS) in the recipient AML cells. In addition, metformin potentiated the antitumor efficacy of cytarabine (Ara-C) in vivo in an NCG immunodeficient mouse xenograft model by inhibiting the mitochondrial transfer and OXPHOS activity in the engrafted human AML cells. Altogether, this study identifies a potential application of metformin in sensitizing AML cells to chemotherapy and unveils a novel mechanism by which metformin executes such effect via blocking the mitochondrial transfer from stromal cells to AML cells.


Assuntos
Diabetes Mellitus Tipo 2 , Leucemia Mieloide Aguda , Metformina , Animais , Citarabina/farmacologia , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/patologia , Metformina/farmacologia , Metformina/uso terapêutico , Camundongos , Camundongos Nus , Camundongos SCID , Mitocôndrias , Células Estromais/patologia , Microambiente Tumoral
8.
Crit Rev Oncol Hematol ; 171: 103607, 2022 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-35101585

RESUMO

Therapy-related acute myeloid leukemia (t-AML), defined as AML arising from prior cytotoxic, radiation, or immunosuppressive therapy for an unrelated disease, accounts for 7 %-8 % of AML cases and primarily occurs in elderly patients. t-AML is associated with an increased probability of adverse cytogenetics and shortened survival compared with de novo AML. Factors predicting poorer prognosis in t-AML include older age, unfavorable karyotype, presence of certain mutations, poor performance status, and poor bone marrow reserve. Few clinical studies have focused specifically on patients with t-AML, and the choice of induction therapy for t-AML is thus typically based on subset analyses of larger studies or on extrapolation. In patients deemed fit, t-AML treatment can involve CPX-351 (liposomal daunorubicin and cytarabine) or conventional chemotherapy, ideally followed by hematopoietic cell transplantation. Patients who are not candidates for intensive therapy may benefit from lower-intensity therapies. Additional agents and combination regimens are being evaluated in clinical studies.


Assuntos
Antineoplásicos , Transplante de Células-Tronco Hematopoéticas , Leucemia Mieloide Aguda , Idoso , Antineoplásicos/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Citarabina/farmacologia , Citarabina/uso terapêutico , Humanos , Leucemia Mieloide Aguda/etiologia , Leucemia Mieloide Aguda/genética , Mutação , Prognóstico
9.
Biotechniques ; 72(3): 81-84, 2022 03.
Artigo em Inglês | MEDLINE | ID: mdl-35119307

RESUMO

Acute myeloid leukemia patients with FMS-like tyrosine kinase 3-internal tandem duplications and mixed lineage leukemia-protein AF9 fusion proteins suffer from poor clinical outcomes. The MOLM-13 acute myeloid leukemia cell line harbors both of these abnormalities and is used in CRISPR experiments to identify disease drivers. However, experimental observations may be biased or inconclusive in the absence of experimentally validated positive control genes. We validated sgRNAs for knockdown of TP53 for cell proliferation and for DCK knockdown and CDA upregulation for cytarabine resistance control genes in MOLM-13 cells. We have provided a detailed CRISPR protocol applicable to both gene knockdown or activation experiments and downstream leukemic phenotype analyses. Inclusion of these controls in CRISPR experiments will enhance the capacity to identify novel myeloid leukemia drivers in MOLM-13 cells.


Assuntos
Citarabina , Leucemia Mieloide Aguda , Linhagem Celular , Linhagem Celular Tumoral , Proliferação de Células/genética , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/genética , Citarabina/farmacologia , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/genética
10.
Biochem Pharmacol ; 198: 114948, 2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-35192847

RESUMO

Acute myeloid leukemia (AML) is a malignant proliferative disease of myeloid hematopoietic origin and cannot be treated appropriately at present. This is due to the fact that leukemia cells are not sensitive to some of the traditional chemotherapy drugs. Or some chemotherapeutic drugs are too toxic to normal cells, affecting their wide clinical application. In this study, we identified BAM15 as a novel mitochondrial uncoupling agent by screening a library of small molecule compounds that inhibit AML cell activity. BAM15 significantly inhibited proliferation and promoted apoptosis in AML cells while at the same time being less cytotoxic to normal cells. The mechanism may be related to the disturbance of the ROS production balance. In vivo investigations revealed that BAM15 effectively suppressed AML progression and prolonged the survival time of mice. In addition, we found that BAM15 can be used in combination with cytarabine to enhance its anti-cancer activity and inhibit the activity of primary cells in AML. Therefore, we identified BAM15 as a potential drug candidate for the treatment of AML.


Assuntos
Leucemia Mieloide Aguda , Animais , Apoptose , Linhagem Celular Tumoral , Proliferação de Células , Citarabina/farmacologia , Citarabina/uso terapêutico , Diaminas , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/patologia , Camundongos , Oxidiazóis , Pirazinas/farmacologia , Espécies Reativas de Oxigênio
11.
J Healthc Eng ; 2022: 2842066, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35126914

RESUMO

PURPOSE: This study aims to determine the influence of targeting araC-resistant acute myeloid leukemia by dual inhibition cyclin-dependent protein kinase (CDK9) and B-cell lymphoma-2 (Bcl-2). METHOD: The c-Myc inhibitor 10058-F4 and the CDK9 inhibitor AZD4573 were used to determine the cell cycle arrest and apoptosis. RESULTS: 10058-F4 reduces c-Myc protein levels and suppresses HepG2 cell proliferation, possibly by upregulating cyclin-dependent kinase (CDK) inhibitors, p21WAF1, and reducing intracellular alpha-fetal protein (AFP) levels. CONCLUSION: The combination of AZD4573 and 10058-F4 has a synergistic anti-araC-resistant AML activity, providing a solid database for the aforementioned scientific hypothesis.


Assuntos
Antineoplásicos , Leucemia Mieloide Aguda , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Apoptose , Linhagem Celular Tumoral , Quinase 9 Dependente de Ciclina , Citarabina/farmacologia , Citarabina/uso terapêutico , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico
12.
Int J Biol Macromol ; 199: 150-161, 2022 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-34973988

RESUMO

Anticancer drug cytarabine, has been widely used for treating haematological malignancies while it has minimal activity against solid tumours, which demands continuous infusion leading to high dose cytarabine toxicity. In this study, folate conjugated chitosan nanoparticles (FCCNP) were used for targeted delivery of cytarabine in breast adenocarcinoma cell lines by making use of the overexpressed folate receptors on the surface of MCF-7. Folate was conjugated to chitosan using carbodiimide. FCCNPs show spherical morphology with a size of<50 nm. Zeta potential of + 45.2 mV and PDI of 0.98 from DLS measurement confirms a stable monodisperse nanoformulation. Cytotoxicity was studied in folate receptor positive, MCF-7 and folate receptor negative, A-549 cell lines. Increased cellular uptake of the drug incorporated nanoparticles was confirmed in MCF-7 cells with fluorophore, squaraine 650 compared to A-549 cells. The relative fold of expression of genes involved in apoptosis such as bax, cyt c and cas 9 were upregulated. The present in vitro study confirms improved cytotoxicity of cytarabine folate conjugated chitosan nanoparticles in MCF-7 cells.


Assuntos
Neoplasias da Mama , Quitosana , Nanopartículas , Neoplasias da Mama/patologia , Sobrevivência Celular , Quitosana/uso terapêutico , Citarabina/farmacologia , Portadores de Fármacos/uso terapêutico , Sistemas de Liberação de Medicamentos , Feminino , Ácido Fólico , Humanos , Células MCF-7
13.
Biochem Biophys Res Commun ; 587: 78-84, 2022 01 08.
Artigo em Inglês | MEDLINE | ID: mdl-34872003

RESUMO

An interaction between acute myeloid leukemia (AML) cells and endothelial cells in the bone marrow seems to play a critical role in chemosensitivity on leukemia treatment. The endothelial niche reportedly enhances the paracrine action of the soluble secretory proteins responsible for chemoresistance in a vascular endothelial growth factor A (VEGF-A)/VEGF receptor 2 (VEGFR-2) signaling pathway-dependent manner. To further investigate the contribution of VEGF-A/VEGFR-2 signaling to the chemoresistance of AML cells, a biochemical assay system in which the AML cells were cocultured with human endothelial EA.hy926 cells in a monolayer was developed. By coculture with EA.hy926 cells, this study revealed that the AML cells resisted apoptosis induced by the anticancer drug cytarabine. SU4312, a VEGFR-2 inhibitor, attenuated VEGFR-2 phosphorylation and VEGF-A/VEGFR-2 signaling-dependent endothelial cell migration; thus, this inhibitor was observed to block VEGF-A/VEGFR-2 signaling. Interestingly, this inhibitor did not reverse the chemoresistance. When VEGFR-2 was knocked out in EA.hy926 cells using the CRISPR-Cas9 system, the cytarabine-induced apoptosis of AML cells did not significantly change compared with that of wild-type cells. Thus, coculture-induced chemoresistance appears to be independent of VEGF-A/VEGFR-2 signaling. When the transwell, a coculturing device, separated the AML cells from the EA.hy926 cells in a monolayer, the coculture-induced chemoresistance was inhibited. Given that the migration of VEGF-A/VEGFR-2 signaling-dependent endothelial cells is necessary for the endothelial niche formation in the bone marrow, VEGF-A/VEGFR-2 signaling contributes to chemoresistance by mediating the niche formation process, but not to the chemoresistance of AML cells in the niche.


Assuntos
Antimetabólitos Antineoplásicos/farmacologia , Citarabina/farmacologia , Resistencia a Medicamentos Antineoplásicos/genética , Fator A de Crescimento do Endotélio Vascular/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética , Inibidores da Angiogênese/farmacologia , Ciclo Celular/efeitos dos fármacos , Ciclo Celular/genética , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Técnicas de Cocultura , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Regulação Leucêmica da Expressão Gênica , Técnicas de Inativação de Genes , Células HL-60 , Humanos , Indóis/farmacologia , Células Jurkat , Células K562 , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patologia , Modelos Biológicos , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismo , Neovascularização Patológica/patologia , Neovascularização Patológica/prevenção & controle , Fosforilação , Transdução de Sinais , Células U937 , Fator A de Crescimento do Endotélio Vascular/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/deficiência
14.
Virchows Arch ; 480(3): 655-666, 2022 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-34738194

RESUMO

SAMHD1 is a deoxynucleoside triphosphate triphosphohydrolase (dNTPase) that restricts viral replication in infected cells and limits the sensitivity to cytarabine by hydrolysing its active metabolite, as recently shown in acute myeloid leukemia. Cytarabine is an essential component in the Nordic mantle cell lymphoma protocols (MCL2 and MCL3) for induction and high-dose chemotherapy treatment before autologous stem cell transplantation for younger patients with mantle cell lymphoma (MCL). We here investigated the expression of SAMHD1 in a population-based cohort of MCL (N = 150). SAMHD1 was highly variably expressed in MCL (range, 0.4% to 100% of positive tumor cells). Cases with blastoid/pleomorphic morphology had higher SAMHD1 expression (P = 0.028) and SAMHD1 was also correlated to tumor cell proliferation (P = 0.016). SAMHD1 expression showed moderate correlation to the expression of the transcriptional regulator SOX11 (P = 0.036) but genetic silencing of SOX11 and SAMHD1 by siRNA in MCL cell lines did not suggest mutual regulation. We hypothesized that expression of SAMHD1 could predict short time to progression in patients treated with Cytarabine as part of high-dose chemotherapy. Despite the correlation with known biological adverse prognostic factors, neither low or high SAMHD1 expression correlated to PFS or OS in patients treated according to the Nordic MCL2 or MCL3 protocols (N = 158).


Assuntos
Transplante de Células-Tronco Hematopoéticas , Linfoma de Célula do Manto , Adulto , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Citarabina/farmacologia , Citarabina/uso terapêutico , Transplante de Células-Tronco Hematopoéticas/métodos , Humanos , Linfoma de Célula do Manto/tratamento farmacológico , Linfoma de Célula do Manto/genética , Linfoma de Célula do Manto/patologia , Proteína 1 com Domínio SAM e Domínio HD/genética , Transplante Autólogo
15.
Exp Hematol ; 105: 39-49, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34767916

RESUMO

Acute myeloid leukemia (AML) remains a clinical challenge. Venetoclax is an effective Bcl-2 selective inhibitor approved by the U.S. Food and Drug Administration (FDA) for treatment of AML in patients who are 75 years and older or who have comorbidities. However, resistance to venetoclax limits its clinical efficacy. Mcl-1 has been identified as one determinant of resistance to venetoclax treatment. In this study, we investigate the Mcl-1 inhibitor S63845 in combination with venetoclax in AML cells. We found that S63845 synergizes with venetoclax in AML cell lines and primary patient samples. Bak/Bax double knockdown and treatment with the pan-caspase inhibitor Z-VAD-FMK revealed that the combination induces intrinsic apoptosis in AML cells. Inhibition of Mcl-1 using another Mcl-1 selective inhibitor, AZD5991, also synergistically enhanced apoptosis induced by venetoclax in a caspase-dependent manner. Importantly, S63845 in combination with venetoclax can effectively combat AML cells with acquired resistance to the standard chemotherapy drug cytarabine. In light of these facts, the combined inhibition of Mcl-1 and Bcl-2 shows promise against AML cells, including relapse/refractory AML.


Assuntos
Antineoplásicos/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Citarabina/farmacologia , Resistencia a Medicamentos Antineoplásicos , Leucemia Mieloide Aguda/tratamento farmacológico , Pirimidinas/farmacologia , Sulfonamidas/farmacologia , Tiofenos/farmacologia , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Sinergismo Farmacológico , Humanos , Terapia de Alvo Molecular , Proteína de Sequência 1 de Leucemia de Células Mieloides/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores
16.
FASEB J ; 36(1): e22094, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34888943

RESUMO

Modifications in sphingolipid (SL) metabolism and mitochondrial bioenergetics are key factors implicated in cancer cell response to chemotherapy, including chemotherapy resistance. In the present work, we utilized acute myeloid leukemia (AML) cell lines, selected to be refractory to various chemotherapeutics, to explore the interplay between SL metabolism and mitochondrial biology supportive of multidrug resistance (MDR). In agreement with previous findings in cytarabine or daunorubicin resistant AML cells, relative to chemosensitive wildtype controls, HL-60 cells refractory to vincristine (HL60/VCR) presented with alterations in SL enzyme expression and lipidome composition. Such changes were typified by upregulated expression of various ceramide detoxifying enzymes, as well as corresponding shifts in ceramide, glucosylceramide, and sphingomyelin (SM) molecular species. With respect to mitochondria, despite consistent increases in both basal respiration and maximal respiratory capacity, direct interrogation of the oxidative phosphorylation (OXPHOS) system revealed intrinsic deficiencies in HL60/VCR, as well as across multiple MDR model systems. Based on the apparent requirement for augmented SL and mitochondrial flux to support the MDR phenotype, we explored a combinatorial therapeutic paradigm designed to target each pathway. Remarkably, despite minimal cytotoxicity in peripheral blood mononuclear cells (PBMC), co-targeting SL metabolism, and respiratory complex I (CI) induced synergistic cytotoxicity consistently across multiple MDR leukemia models. Together, these data underscore the intimate connection between cellular sphingolipids and mitochondrial metabolism and suggest that pharmacological intervention across both pathways may represent a novel treatment strategy against MDR.


Assuntos
Resistência a Múltiplos Medicamentos , Resistencia a Medicamentos Antineoplásicos , Leucemia/metabolismo , Mitocôndrias/metabolismo , Fosforilação Oxidativa , Esfingolipídeos/metabolismo , Citarabina/farmacologia , Daunorrubicina/farmacologia , Células HL-60 , Humanos , Leucemia/patologia , Mitocôndrias/patologia , Vincristina/farmacologia
17.
Nat Commun ; 12(1): 6936, 2021 11 26.
Artigo em Inglês | MEDLINE | ID: mdl-34836965

RESUMO

Chemoresistance posts a major hurdle for treatment of acute leukemia. There is increasing evidence that prolonged and intensive chemotherapy often fails to eradicate leukemic stem cells, which are protected by the bone marrow niche and can induce relapse. Thus, new therapeutic approaches to overcome chemoresistance are urgently needed. By conducting an ex vivo small molecule screen, here we have identified Quinacrine (QC) as a sensitizer for Cytarabine (AraC) in treating acute lymphoblastic leukemia (ALL). We show that QC enhances AraC-mediated killing of ALL cells, and subsequently abrogates AraC resistance both in vitro and in an ALL-xenograft model. However, while combo AraC+QC treatment prolongs the survival of primary transplanted recipients, the combination exhibits limited efficacy in secondary transplanted recipients, consistent with the survival of niche-protected leukemia stem cells. Introduction of Cdc42 Activity Specific Inhibitor, CASIN, enhances the eradication of ALL leukemia stem cells by AraC+QC and prolongs the survival of both primary and secondary transplanted recipients without affecting normal long-term human hematopoiesis. Together, our findings identify a small-molecule regimen that sensitizes AraC-mediated leukemia eradication and provide a potential therapeutic approach for better ALL treatment.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Carbazóis/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Quinacrina/farmacologia , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Carbazóis/uso terapêutico , Linhagem Celular Tumoral , Citarabina/farmacologia , Citarabina/uso terapêutico , Resistencia a Medicamentos Antineoplásicos/genética , Feminino , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologia , Cultura Primária de Células , Quinacrina/uso terapêutico , Ensaios Antitumorais Modelo de Xenoenxerto
18.
Int J Mol Sci ; 22(22)2021 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-34830054

RESUMO

Chimeric RNAs are often associated with chromosomal rearrangements in cancer. In addition, they are also widely detected in normal tissues, contributing to transcriptomic complexity. Despite their prevalence, little is known about the characteristics and functions of chimeric RNAs. Here, we examine the genetic structure and biological roles of CLEC12A-MIR223HG, a novel chimeric transcript produced by the fusion of the cell surface receptor CLEC12A and the miRNA-223 host gene (MIR223HG), first identified in chronic myeloid leukemia (CML) patients. Surprisingly, we observed that CLEC12A-MIR223HG is not just expressed in CML, but also in a variety of normal tissues and cell lines. CLEC12A-MIR223HG expression is elevated in pro-monocytic cells resistant to chemotherapy and during monocyte-to-macrophage differentiation. We observed that CLEC12A-MIR223HG is a product of trans-splicing rather than a chromosomal rearrangement and that transcriptional activation of CLEC12A with the CRISPR/Cas9 Synergistic Activation Mediator (SAM) system increases CLEC12A-MIR223HG expression. CLEC12A-MIR223HG translates into a chimeric protein, which largely resembles CLEC12A but harbours an altered C-type lectin domain altering key disulphide bonds. These alterations result in differences in post-translational modifications, cellular localization, and protein-protein interactions. Taken together, our observations support a possible involvement of CLEC12A-MIR223HG in the regulation of CLEC12A function. Our workflow also serves as a template to study other uncharacterized chimeric RNAs.


Assuntos
Fusão Gênica , Lectinas Tipo C/genética , Leucemia Mieloide/genética , MicroRNAs/genética , Proteínas Mutantes Quiméricas/genética , Receptores Mitogênicos/genética , Trans-Splicing , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Diferenciação Celular/genética , Linhagem Celular , Citarabina/farmacologia , Humanos , Lectinas Tipo C/metabolismo , Leucemia Mieloide/metabolismo , MicroRNAs/metabolismo , Proteínas Mutantes Quiméricas/metabolismo , Receptores Mitogênicos/metabolismo , Ativação Transcricional
19.
Zhongguo Shi Yan Xue Ye Xue Za Zhi ; 29(5): 1403-1410, 2021 Oct.
Artigo em Chinês | MEDLINE | ID: mdl-34627417

RESUMO

OBJECTIVE: To establish cytarabine-resistant acute lymphoblastic leukemia (ALL) cell lines and investigate its possible resistant mechanism. METHODS: Low-concentration cytarabine (Ara-C) continuously induced and cultured Jurkat and Nalm-6 cells to construct cytarabine-resistant cell lines Jurkat/Ara-C and Nalm-6/Ara-C. The cell viability was detected by CCK-8 assay, and the distribution of cell cycle was detected by flow cytometry. Real-time fluorescence quantitative PCR was used to detect the mRNA expression levels of multidrug resistant gene and Ara-C metabolic enzymes. The expression levels of cyclin were detected by Western blot. RESULTS: Jurkat/Ara-C and Nalm-6/Ara-C drug-resistant cell lines were successfully established, the resistance index of which was 1 973.908±161.163 and 7 231.643± 1 190.624, respectively. Drug-resistant cell lines had no cross-resistance to commonly used chemotherapeutic drugs, such as doxorubicin. Flow cytometry showed that the ratio of G0/G1 phase in Jurkat/Ara-C cells increased but G2/M phase decreased (P<0.05), while the cell cycle distribution of Nalm-6/Ara-C cells did not change in comparison with Nalm-6 cells. The results of real-time quantitative PCR showed that the expression of deoxycytidine kinase (DCK) and cytidine deaminase (CDA) were significantly down-regulated in drug-resistant cells (P<0.05), MRP was up-regulated in Nalm-6/Ara-C cells (P<0.05), while MDR1, LRP and BCRP did not increase in comparison with parental cells. Western blot analysis revealed that cyclinB1 was significantly under-expressed in drug-resistant cells (P<0.05), while cyclinD1 did not change, when compared with parental cells. CONCLUSION: Cytarabine-resistant ALL cell lines are successfully established by using low concentration continuous induction method, and its drug-resistant mechanism may be related to the deficiencies of DCK and cyclinB1.


Assuntos
Citarabina , Leucemia-Linfoma Linfoblástico de Células Precursoras , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Linhagem Celular , Citarabina/farmacologia , Resistencia a Medicamentos Antineoplásicos , Humanos , Proteínas de Neoplasias
20.
J Microencapsul ; 38(7-8): 546-558, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34632926

RESUMO

AIM: To synthesise cytarabine-loaded SLNs modified with the RGD peptide as a ligand, suitable for effective cancer therapy. METHODS: SLNs were synthesised by the high shear, hot homogenisation technique. A 2 level 3 factor analysis was used in optimisation. Particle size, zeta potential, poly-dispersion index and surface morphology were measured. Drug encapsulation, drug release, release kinetics, nanoparticle stability and chemical structure were determined. LIVE/DEAD® Fluorescence Assay was used to qualify cytotoxicity and Tryphan Blue assay to quantify. RESULTS: Cyt-SLNs exhibited a size of 161 ± 2.25 nm, a PDI of 0.49 ± 0.15 and a zeta potential of -19.8 mV. Entrapment fell at 88.87 ± 0.02% and release at 83.5 ± 0.95%. The in vitro release kinetics pointed towards a diffusion-based drug release mechanism. SLNs remained stable for 60 d. Cytotoxicity studies revealed that conjugation of the ligand with the RDG peptide resulted in a significant decrease in cell viability in both cell lines. CONCLUSION: Overall, the study suggests that RGD-SLN-cyt can be used for effective cancer therapy.


Assuntos
Lipídeos , Nanopartículas , Citarabina/farmacologia , Portadores de Fármacos , Lipossomos , Oligopeptídeos , Tamanho da Partícula
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