RESUMO
Chronic myeloid leukemia (CML) is a model of genomically based diagnosis and management where BCR::ABL1 is successfully targeted by tyrosine kinase inhibitor (TKI) therapy in most patients. The dynamics of BCR::ABL1 transcript decline during therapy is a dependable biomarker of response, relapse, and drug resistance. Missense mutations acquired within the BCR::ABL1 kinase domain that disrupt TKI binding can evolve during therapy and are frequently detected in patients for whom TKI treatment fails. Importantly, specific BCR::ABL1 missense mutations are targetable alterations and direct therapeutic decisions based on the individual mutant TKI sensitivity profile. Nevertheless, BCR::ABL1 mutations are only implicated in approximately half of the cases of acquired resistance. Furthermore, not all patients with a single BCR::ABL1 mutation that is predicted to be sensitive to a specific TKI will experience a response when switched to that TKI. Progression to blast phase heralds independence from BCR::ABL1, and this phase of the disease is notoriously difficult to treat. The independent drivers of resistance and disease progression have long been investigated to both predict progression and to find targets for therapeutic intervention. Recent data reaffirm that drug resistance and disease progression is a mutation-driven process in CML, and somatic variants in genes that are known to drive acute myeloid and lymphoid leukemia have been detected in patients in the advanced phases of CML. Genomic testing over the last few decades for patients with blood cancer has revealed of variety of genomic aberrations that drive disease. Consequently, incorporation of genomic factors into patient management for a range of blood cancers has led to the implementation of high-throughput gene testing to detect clinically actionable variants. Is it time to integrate broader genomic screening into clinical management strategies for patients with CML?
Assuntos
Proteínas de Fusão bcr-abl , Leucemia Mielogênica Crônica BCR-ABL Positiva , Humanos , Proteínas de Fusão bcr-abl/genética , Resistencia a Medicamentos Antineoplásicos/genética , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Leucemia Mielogênica Crônica BCR-ABL Positiva/diagnóstico , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Genômica , Progressão da DoençaRESUMO
The BCR::ABL1 gene fusion initiates chronic myeloid leukemia (CML); however, evidence has accumulated from studies of highly selected cohorts that variants in other cancer-related genes are associated with treatment failure. Nevertheless, the true incidence and impact of additional genetic abnormalities (AGA) at diagnosis of chronic phase (CP)-CML is unknown. We sought to determine whether AGA at diagnosis in a consecutive imatinib-treated cohort of 210 patients enrolled in the TIDEL-II trial influenced outcome despite a highly proactive treatment intervention strategy. Survival outcomes including overall survival, progression-free survival, failure-free survival, and BCR::ABL1 kinase domain mutation acquisition were evaluated. Molecular outcomes were measured at a central laboratory and included major molecular response (MMR, BCR::ABL1 ≤0.1%IS), MR4 (BCR::ABL1 ≤0.01%IS), and MR4.5 (BCR::ABL1 ≤0.0032%IS). AGA included variants in known cancer genes and novel rearrangements involving the formation of the Philadelphia chromosome. Clinical outcomes and molecular response were assessed based on the patient's genetic profile and other baseline factors. AGA were identified in 31% of patients. Potentially pathogenic variants in cancer-related genes were detected in 16% of patients at diagnosis (including gene fusions and deletions) and structural rearrangements involving the Philadelphia chromosome (Ph-associated rearrangements) were detected in 18%. Multivariable analysis demonstrated that the combined genetic abnormalities plus the EUTOS long-term survival clinical risk score were independent predictors of lower molecular response rates and higher treatment failure. Despite a highly proactive treatment intervention strategy, first-line imatinib-treated patients with AGA had poorer response rates. These data provide evidence for the incorporation of genomically-based risk assessment for CML.
Assuntos
Antineoplásicos , Leucemia Mielogênica Crônica BCR-ABL Positiva , Leucemia Mieloide de Fase Crônica , Humanos , Mesilato de Imatinib/uso terapêutico , Antineoplásicos/uso terapêutico , Cromossomo Filadélfia , Proteínas de Fusão bcr-abl/genética , Proteínas de Fusão bcr-abl/metabolismo , Leucemia Mielogênica Crônica BCR-ABL Positiva/diagnóstico , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mieloide de Fase Crônica/tratamento farmacológico , Inibidores de Proteínas Quinases/uso terapêuticoRESUMO
PURPOSE OF REVIEW: The chronic myeloid leukemia (CML) treatment success story is incomplete as some patients still fail therapy, leading to end-stage disease and death. Here we discuss recent research into CML incidence, the role of comorbidities on survival and detecting patients at risk of failing therapy. RECENT FINDINGS: The incidence of CML has fallen markedly in high social-demographic index (SDI) regions of the world but there is disturbing evidence that this is not the case in low and low-middle SDI countries. Now that CML patients more frequently die from their co-morbid conditions than from CML the Adult Comorbidity Evaluation-27 score can assist in risk assessment at diagnosis. Non-adherence to therapy contributes greatly to treatment failure. A good doctor-patient relationship and social support promote good adherence, but patient age, gender, and financial burden have negative effects, suggesting avenues for intervention. Mutations in cancer-associated genes adversely affect outcome and their detection at diagnosis may guide therapeutic choice and offer non-BCR::ABL1 targeted therapies. A differential gene expression signature to assist risk detection is a highly sought-after diagnostic tool being actively researched on several fronts. Detecting patients at risk of failing therapy is being assisted by recent technological advances enabling highly sensitive genomic and expression analysis of insensitive cells. However, patient lifestyle, adherence to therapy, and comorbidities are critical risk factors that need to be addressed by interventions such as social and financial support.
Assuntos
Proteínas de Fusão bcr-abl , Leucemia Mielogênica Crônica BCR-ABL Positiva , Adulto , Humanos , Proteínas de Fusão bcr-abl/genética , Relações Médico-Paciente , Inibidores de Proteínas Quinases/uso terapêutico , Leucemia Mielogênica Crônica BCR-ABL Positiva/diagnóstico , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Genômica , Fatores de RiscoRESUMO
Mutation detection is increasingly used for the management of hematological malignancies. Prior whole transcriptome and whole exome sequencing studies using total RNA and DNA identified diverse mutation types in cancer-related genes associated with treatment failure in patients with chronic myeloid leukemia. Variants included single-nucleotide variants and small insertions/deletions, plus fusion transcripts and partial or whole gene deletions. The hypothesis that all of these mutation types could be detected by a single cost-effective hybridization capture next-generation sequencing method using total RNA was assessed. A method was developed that targeted 130 genes relevant for myeloid and lymphoid leukemia. Retrospective samples with 121 precharacterized variants were tested using total RNA and/or DNA. Concordance of detection of precharacterized variants using RNA or DNA was 96%, whereas the enhanced sensitivity identified additional variants. Comparison between 24 matched DNA and RNA samples demonstrated 95.3% of 170 variants detectable using DNA were detected using RNA, including all but one variant predicted to activate nonsense-mediated decay. RNA identified an additional 10 variants, including fusion transcripts. Furthermore, the true effect of splice variants on RNA splicing was only evident using RNA. In conclusion, capture sequencing using total RNA alone is suitable for detecting a range of variants relevant in chronic myeloid leukemia and may be more broadly applied to other hematological malignancies where diverse variant types define risk groups.
Assuntos
Neoplasias Hematológicas , Leucemia Mielogênica Crônica BCR-ABL Positiva , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Leucemia Mielogênica Crônica BCR-ABL Positiva/diagnóstico , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Mutação , RNA , Estudos RetrospectivosRESUMO
Biomarkers which better match anticancer drugs with cancer driver genes hold the promise of improved clinical responses and cure rates. We developed a precision medicine platform of rapid high-throughput drug screening (HTS) and patient-derived xenografting (PDX) of primary tumor tissue, and evaluated its potential for treatment identification among 56 consecutively enrolled high-risk pediatric cancer patients, compared with conventional molecular genomics and transcriptomics. Drug hits were seen in the majority of HTS and PDX screens, which identified therapeutic options for 10 patients for whom no targetable molecular lesions could be found. Screens also provided orthogonal proof of drug efficacy suggested by molecular analyses and negative results for some molecular findings. We identified treatment options across the whole testing platform for 70% of patients. Only molecular therapeutic recommendations were provided to treating oncologists and led to a change in therapy in 53% of patients, of whom 29% had clinical benefit. These data indicate that in vitro and in vivo drug screening of tumor cells could increase therapeutic options and improve clinical outcomes for high-risk pediatric cancer patients.
Assuntos
Antineoplásicos , Neoplasias , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Criança , Modelos Animais de Doenças , Genômica/métodos , Humanos , Neoplasias/patologia , Medicina de Precisão/métodos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Sphingolipid dysregulation is often associated with insulin resistance, while the enzymes controlling sphingolipid metabolism are emerging as therapeutic targets for improving insulin sensitivity. We report herein that sphingosine kinase 2 (SphK2), a key enzyme in sphingolipid catabolism, plays a critical role in the regulation of hepatic insulin signaling and glucose homeostasis both in vitro and in vivo. Hepatocyte-specific Sphk2 knockout mice exhibit pronounced insulin resistance and glucose intolerance. Likewise, SphK2-deficient hepatocytes are resistant to insulin-induced activation of the phosphoinositide 3-kinase (PI3K)-Akt-FoxO1 pathway and elevated hepatic glucose production. Mechanistically, SphK2 deficiency leads to the accumulation of sphingosine that, in turn, suppresses hepatic insulin signaling by inhibiting PI3K activation in hepatocytes. Either reexpressing functional SphK2 or pharmacologically inhibiting sphingosine production restores insulin sensitivity in SphK2-deficient hepatocytes. In conclusion, the current study provides both experimental findings and mechanistic data showing that SphK2 and sphingosine in the liver are critical regulators of insulin sensitivity and glucose homeostasis.
Assuntos
Glucose/metabolismo , Insulina/metabolismo , Fígado/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Animais , Linhagem Celular Tumoral , Feminino , Hepatócitos/enzimologia , Hepatócitos/metabolismo , Homeostase , Humanos , Fígado/enzimologia , Masculino , Camundongos , Camundongos Knockout , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Esfingolipídeos/metabolismoRESUMO
BACKGROUND: Epithelioid inflammatory myofibroblastic sarcoma (eIMS) is characterised by perinuclear ALK localisation, CD30 expression and early relapse despite crizotinib treatment. We aimed to identify therapies to prevent and/or treat ALK inhibitor resistance. METHODS: Malignant ascites, from an eIMS patient at diagnosis and following multiple relapses, were used to generate matched diagnosis and relapse xenografts. RESULTS: Xenografts were validated by confirmation of RANBP2-ALK rearrangement, perinuclear ALK localisation and CD30 expression. Although brentuximab-vedotin (BV) demonstrated single-agent activity, tumours regrew during BV therapy. BV resistance was associated with reduced CD30 expression and induction of ABCB1. BV resistance was reversed in vitro by tariquidar, but combination BV and tariquidar treatment only briefly slowed xenograft growth compared with BV alone. Combining BV with either crizotinib or ceritinib resulted in marked tumour shrinkage in both xenograft models, and resulted in prolonged tumour-free survival in the diagnosis compared with the relapse xenograft. CONCLUSIONS: CD30 is a therapeutic target in eIMS. BV efficacy is limited by the rapid emergence of resistance. Prolonged survival with combination ALK and CD30-targeted-therapy in the diagnosis model provides the rationale to trial this combination in eIMS patients at diagnosis. This combination could also be considered for other CD30-positive, ALK-rearranged malignancies.
Assuntos
Quinase do Linfoma Anaplásico/antagonistas & inibidores , Quinase do Linfoma Anaplásico/genética , Rearranjo Gênico , Antígeno Ki-1/antagonistas & inibidores , Chaperonas Moleculares/genética , Miofibroblastos/patologia , Complexo de Proteínas Formadoras de Poros Nucleares/genética , Sarcoma/tratamento farmacológico , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/antagonistas & inibidores , Idoso de 80 Anos ou mais , Animais , Brentuximab Vedotin/uso terapêutico , Resistencia a Medicamentos Antineoplásicos , Humanos , Inflamação , Masculino , Camundongos , Sarcoma/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Blast crisis of chronic myeloid leukemia is associated with poor survival and the accumulation of genomic lesions. Using whole-exome and/or RNA sequencing of patients at chronic phase (CP, n = 49), myeloid blast crisis (MBC, n = 19), and lymphoid blast crisis (LBC, n = 20), we found 25 focal gene deletions and 14 fusions in 24 patients in BC. Deletions predominated in LBC (83% of structural variants). Transcriptional analysis identified the upregulation of genes involved in V(D)J recombination, including RAG1/2 and DNTT in LBC. RAG recombination is a reported mediator of IKZF1 deletion. We investigated the extent of RAG-mediated genomic lesions in BC. Molecular hallmarks of RAG activity; DNTT-mediated nucleotide insertions and a RAG-binding motif at structural variants were exclusively found in patients with high RAG expression. Structural variants in 65% of patients in LBC displayed these hallmarks compared with only 5% in MBC. RAG-mediated events included focal deletion and novel fusion of genes associated with hematologic cancer: IKZF1, RUNX1, CDKN2A/B, and RB1. Importantly, 8/8 patients with elevated DNTT at CP diagnosis progressed to LBC by 12 months, potentially enabling early prediction of LBC. This work confirms the central mutagenic role of RAG in LBC and describes potential clinical utility in CML management.
Assuntos
Crise Blástica/genética , Proteínas de Ligação a DNA/fisiologia , Proteínas de Homeodomínio/fisiologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Proteínas Nucleares/fisiologia , Recombinação Genética , Biologia Computacional , DNA Nucleotidilexotransferase/genética , Proteínas de Ligação a DNA/genética , Deleção de Genes , Proteínas de Homeodomínio/genética , Humanos , Proteínas Nucleares/genéticaRESUMO
Around 10% of acute leukemias harbor a rearrangement of the MLL/KMT2A gene, and the presence of this translocation results in a highly aggressive, therapy-resistant leukemia subtype with survival rates below 50%. There is a high unmet need to identify safer and more potent therapies for MLL-rearranged (MLL-r) leukemia that can be combined with established chemotherapeutics to decrease treatment-related toxicities. The curaxin, CBL0137, has demonstrated nongenotoxic anticancer and chemopotentiating effects in a number of preclinical cancer models and is currently in adult Phase I clinical trials for solid tumors and hematological malignancies. The aim of our study was to investigate whether CBL0137 has potential as a therapeutic and chemopotentiating compound in MLL-r leukemia through a comprehensive analysis of its efficacy in preclinical models of the disease. CBL0137 decreased the viability of a panel of MLL-r leukemia cell lines (n = 12) and xenograft cells derived from patients with MLL-r acute lymphoblastic leukemia (ALL, n = 3) in vitro with submicromolar IC50s. The small molecule drug was well-tolerated in vivo and significantly reduced leukemia burden in a subcutaneous MV4;11 MLL-r acute myeloid leukemia model and in patient-derived xenograft models of MLL-r ALL (n = 5). The in vivo efficacy of standard of care drugs used in remission induction for pediatric ALL was also potentiated by CBL0137. CBL0137 exerted its anticancer effect by trapping Facilitator of Chromatin Transcription (FACT) into chromatin, activating the p53 pathway and inducing an Interferon response. Our findings support further preclinical evaluation of CBL0137 as a new approach for the treatment of MLL-r leukemia.
Assuntos
Antineoplásicos/farmacologia , Carbazóis/farmacologia , Rearranjo Gênico , Histona-Lisina N-Metiltransferase/genética , Proteína de Leucina Linfoide-Mieloide/genética , Animais , Antineoplásicos/uso terapêutico , Apoptose/genética , Carbazóis/uso terapêutico , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/genética , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Proteínas de Grupo de Alta Mobilidade/genética , Humanos , Estimativa de Kaplan-Meier , Leucemia Aguda Bifenotípica/diagnóstico , Leucemia Aguda Bifenotípica/tratamento farmacológico , Leucemia Aguda Bifenotípica/genética , Leucemia Aguda Bifenotípica/mortalidade , Camundongos , Transdução de Sinais/efeitos dos fármacos , Fatores de Elongação da Transcrição/genética , Transcriptoma , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Pediatric high grade gliomas (HGG) are primary brain malignancies that result in significant morbidity and mortality. One of the challenges in their treatment is inter- and intra-tumoral heterogeneity. Precision medicine approaches have the potential to enhance diagnostic, prognostic and/or therapeutic information. In this case study we describe the molecular characterization of a pediatric HGG and the use of an integrated approach based on genomic, in vitro and in vivo testing to identify actionable targets and treatment options. Molecular analysis based on WGS performed on initial and recurrent tumor biopsies revealed mutations in TP53, TSC1 and CIC genes, focal amplification of MYCN, and copy number gains in SMO and c-MET. Transcriptomic analysis identified increased expression of MYCN, and genes involved in sonic hedgehog signaling proteins (SHH, SMO, GLI1, GLI2) and receptor tyrosine kinase pathways (PLK, AURKA, c-MET). HTS revealed no cytotoxic efficacy of SHH pathway inhibitors while sensitivity was observed to the mTOR inhibitor temsirolimus, the ALK inhibitor ceritinib, and the PLK1 inhibitor BI2536. Based on the integrated approach, temsirolimus, ceritinib, BI2536 and standard therapy temozolomide were selected for further in vivo evaluation. Using the PDX animal model (median survival 28 days) we showed significant in vivo activity for mTOR inhibition by temsirolimus and BI2536 (median survival 109 and 115.5 days respectively) while ceritinib and temozolomide had only a moderate effect (43 and 75.5 days median survival respectively). This case study demonstrates that an integrated approach based on genomic, in vitro and in vivo drug efficacy testing in a PDX model may be useful to guide the management of high risk pediatric brain tumor in a clinically meaningful timeframe.
Assuntos
Ensaios de Seleção de Medicamentos Antitumorais , Genômica , Ensaios de Triagem em Larga Escala , Neoplasias/tratamento farmacológico , Neoplasias/genética , Medicina de Precisão , Fatores Etários , Animais , Biópsia , Criança , Metilação de DNA , Modelos Animais de Doenças , Ensaios de Seleção de Medicamentos Antitumorais/métodos , Genômica/métodos , Humanos , Imageamento por Ressonância Magnética , Masculino , Camundongos , Neoplasias/diagnóstico , Neoplasias/mortalidade , Medicina de Precisão/métodos , Recidiva , Resultado do Tratamento , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Genomic events associated with poor outcome in chronic myeloid leukemia (CML) are poorly understood. We performed whole-exome sequencing, copy-number variation, and/or RNA sequencing for 65 patients to discover mutations at diagnosis and blast crisis (BC). Forty-six patients with chronic-phase disease with the extremes of outcome were studied at diagnosis. Cancer gene variants were detected in 15 (56%) of 27 patients with subsequent BC or poor outcome and in 3 (16%) of 19 optimal responders (P = .007). Frequently mutated genes at diagnosis were ASXL1, IKZF1, and RUNX1 The methyltransferase SETD1B was a novel recurrently mutated gene. A novel class of variant associated with the Philadelphia (Ph) translocation was detected at diagnosis in 11 (24%) of 46 patients comprising fusions and/or rearrangement of genes on the translocated chromosomes, with evidence of fragmentation, inversion, and imperfect sequence reassembly. These were more frequent at diagnosis in patients with poor outcome: 9 (33%) of 27 vs 2 (11%) of 19 optimal responders (P = .07). Thirty-nine patients were tested at BC, and all had cancer gene variants, including ABL1 kinase domain mutations in 58%. However, ABL1 mutations cooccurred with other mutated cancer genes in 89% of cases, and these predated ABL1 mutations in 62% of evaluable patients. Gene fusions not associated with the Ph translocation occurred in 42% of patients at BC and commonly involved fusion partners that were known cancer genes (78%). Genomic analysis revealed numerous relevant variants at diagnosis in patients with poor outcome and all patients at BC. Future refined biomarker testing of specific variants will likely provide prognostic information to facilitate a risk-adapted therapeutic approach.
Assuntos
Biomarcadores Tumorais/genética , Genômica , Leucemia Mielogênica Crônica BCR-ABL Positiva , Proteínas de Neoplasias/genética , Cromossomo Filadélfia , Translocação Genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Intervalo Livre de Doença , Feminino , Seguimentos , Humanos , Leucemia Mielogênica Crônica BCR-ABL Positiva/diagnóstico , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/mortalidade , Masculino , Pessoa de Meia-Idade , Fatores de Risco , Taxa de SobrevidaRESUMO
Sphingosine kinase (SphK) is an important signalling enzyme that catalyses the phosphorylation of sphingosine (Sph) to form sphingosine1phosphate (S1P). The multifunctional lipid, S1P binds to a family of five G protein-coupled receptors (GPCRs). As an intracellular second messenger, S1P activates key signalling cascades responsible for the maintenance of sphingolipid metabolism, and has been implicated in the progression of cancer, and the development of other inflammatory and metabolic diseases. SphK and S1P are critical molecules involved in the regulation of various cellular metabolic processes, such as cell proliferation, survival, apoptosis, adhesion and migration. There is strong evidence supporting the critical roles of SphK and S1P in the progression of diabetes mellitus, including insulin sensitivity and insulin secretion, pancreatic ßcell apoptosis, and the development of diabetic inflammatory state. In this review, we summarise the current state of knowledge for SphK/S1P signalling effects, associated with the development of insulin resistance, pancreatic ßcell death and the vascular complications of diabetes mellitus.
Assuntos
Diabetes Mellitus/etiologia , Diabetes Mellitus/metabolismo , Transdução de Sinais , Esfingolipídeos/metabolismo , Animais , Complicações do Diabetes/etiologia , Complicações do Diabetes/metabolismo , Ativação Enzimática , Espaço Extracelular/metabolismo , Humanos , Resistência à Insulina , Células Secretoras de Insulina/metabolismo , Espaço Intracelular/metabolismo , Isoenzimas , Lisofosfolipídeos/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Transporte Proteico , Esfingosina/análogos & derivados , Esfingosina/metabolismoRESUMO
We have previously reported that the steroid hormone estrogens stimulate activation of sphingosine kinase 1 (SphK1) and sphingosine-1-phosphate (S1P) receptors in breast cancer cells. Both estrogens and S1P are potent biological modulators of endothelial function in vasculature able to activate multiple effectors, including endothelial nitric oxide synthase (eNOS). In this study we report that treatment of endothelial cells (ECs) with 17ß-estradiol (E2) resulted in a rapid, transient, and dose-dependent increase in SphK activity and increased S1P production. The effect was not reproduced by the inactive E2 analogue 17α-E2. Expression of the dominant-negative mutant SphK1(G82D) or transfection of SphK1-targeted siRNA in ECs caused not only a defect in SphK activation by E2, but also a significant inhibition of E2-induced activation of Akt/eNOS. Furthermore, E2 treatment induced internalization of plasma membrane S1P1 receptor, accompanied with an increase in the amount of cytosolic S1P1. By down-regulating S1P1 receptor expression, the S1P1-specific antisense oligonucleotides significantly inhibited E2-induced activation of Akt/eNOS in ECs. E2-induced EC migration and tube formation were also inhibited by S1P1 down-regulation. Thus, the findings indicate an important role of the SphK1/S1P1 pathway in mediating estrogen signaling and its actions in vasculature.
Assuntos
Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Estrogênios/farmacologia , Receptores de Lisoesfingolipídeo/metabolismo , Animais , Bovinos , Células Cultivadas , Relação Dose-Resposta a Droga , Estradiol/farmacologia , Estrogênios/metabolismo , Humanos , Oligonucleotídeos Antissenso/farmacologia , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Receptores de Lisoesfingolipídeo/antagonistas & inibidores , Receptores de Lisoesfingolipídeo/genética , Relação Estrutura-AtividadeRESUMO
Dipeptidyl peptidase 9 (DPP9) is a ubiquitously expressed member of the DPP4 gene and protease family. Deciphering the biological functions of DPP9 and its roles in pathogenesis has implicated DPP9 in tumor biology, the immune response, apoptosis, intracellular epidermal growth factor-dependent signaling and cell adhesion and migration. We investigated the intracellular distribution of DPP9 chimeric fluorescent proteins and consequent functions of DPP9. We showed that while some DPP9 is associated with mitochondria, the strongest co-localization was with microtubules. Under steady state conditions, DPP9 was not seen at the plasma membrane, but upon stimulation with either phorbol 12-myristate 13-acetate or epidermal growth factor, some DPP9 re-distributed towards the ruffling membrane. DPP9 was seen at the leading edge of the migrating cell and co-localized with the focal adhesion proteins, integrin-ß1 and talin. DPP9 gene silencing and treatment with a DPP8/DPP9 specific inhibitor both reduced cell adhesion and migration. Expression of integrin-ß1 and talin was decreased in DPP9-deficient and DPP9-enzyme-inactive cells. There was a concomitant decrease in the phosphorylation of focal adhesion kinase and paxillin, indicating that DPP9 knockdown or enzyme inhibition suppressed the associated adhesion signaling pathway, causing impaired cell movement. These novel findings provide mechanistic insights into the regulatory role of DPP9 in cell movement, and may thus implicate DPP9 in tissue and tumor growth and metastasis.
Assuntos
Dipeptidil Peptidases e Tripeptidil Peptidases/metabolismo , Proteína-Tirosina Quinases de Adesão Focal/metabolismo , Paxilina/metabolismo , Adesão Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Movimento Celular/efeitos dos fármacos , Colágeno/farmacologia , Dipeptidil Peptidases e Tripeptidil Peptidases/antagonistas & inibidores , Regulação para Baixo/efeitos dos fármacos , Família de Proteínas EGF/farmacologia , Fibronectinas/farmacologia , Adesões Focais/efeitos dos fármacos , Adesões Focais/metabolismo , Humanos , Integrina beta1/metabolismo , Microtúbulos/efeitos dos fármacos , Microtúbulos/metabolismo , Fosforilação/efeitos dos fármacos , Frações Subcelulares/efeitos dos fármacos , Frações Subcelulares/metabolismo , Talina/metabolismo , Acetato de Tetradecanoilforbol/farmacologiaRESUMO
BACKGROUND: Technological advances including high-throughput sequencing have identified numerous tumor-specific genetic changes in pediatric and adolescent cancers that can be exploited as targets for novel therapies. SCOPE OF REVIEW: This review provides a detailed overview of recent advances in the application of target-specific therapies for childhood cancers, either as single agents or in combination with other therapies. The review summarizes preclinical evidence on which clinical trials are based, early phase clinical trial results, and the incorporation of predictive biomarkers into clinical practice, according to cancer type. MAJOR CONCLUSIONS: There is growing evidence that molecularly targeted therapies can valuably add to the arsenal available for treating childhood cancers, particularly when used in combination with other therapies. Nonetheless the introduction of molecularly targeted agents into practice remains challenging, due to the use of unselected populations in some clinical trials, inadequate methods to evaluate efficacy, and the need for improved preclinical models to both evaluate dosing and safety of combination therapies. GENERAL SIGNIFICANCE: The increasing recognition of the heterogeneity of molecular causes of cancer favors the continued development of molecularly targeted agents, and their transfer to pediatric and adolescent populations.
RESUMO
FTY720, a sphingosine analog, is a novel immunosuppressant currently undergoing multiple clinical trials for the prevention of organ transplant rejection and treatment of various autoimmune diseases. Recent studies indicate an additional cytotoxic effect of FTY720 and its preclinical efficacy in a variety of cancer models, yet the underlying mechanisms remain unclear. We demonstrate here for the first time that FTY720 exhibits a potent, dose- and time-dependent cytotoxic effect in human ovarian cancer cells, even in the cells that are resistant to cisplatin, a commonly prescribed chemotherapeutic drug for treatment of ovarian cancer. In contrast to the previously reported cytotoxicity of FTY720 in many other cancer cell types, FTY720 kills ovarian cancer cells independent of caspase 3 activity and induces cellular swelling and cytoplasmic vacuolization with evident features of necrotic cell death. Furthermore, the presence of autophagic hallmarks, including an increased number of autophagosomes and the formation and accumulation of LC3-II, are observed in FTY720-treated cells before cell death. FTY720 treatment enhances autophagic flux as reflected in the increased LC3 turnover and p62 degradation. Notably, blockade of autophagy by either specific chemical inhibitors or siRNAs targeting Beclin 1 or LC3 resulted in aggravated necrotic cell death in response to FTY720, suggesting that FTY720-induced autophagy plays a self-protective role against its own cytotoxic effect. Thus, our findings not only demonstrate a new death pathway underlying the cytotoxic effect of FTY720, but also suggest that targeting autophagy could augment the anticancer potency, providing the framework for further development of FTY720 as a new chemotherapeutic agent for ovarian cancer treatment.
Assuntos
Autofagia/efeitos dos fármacos , Citoproteção/efeitos dos fármacos , Neoplasias Ovarianas/patologia , Propilenoglicóis/farmacologia , Esfingosina/análogos & derivados , Apoptose/efeitos dos fármacos , Caspases/metabolismo , Linhagem Celular Tumoral , Cisplatino/farmacologia , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Cloridrato de Fingolimode , Humanos , Concentração Inibidora 50 , Lisofosfolipídeos/metabolismo , Necrose , Neoplasias Ovarianas/enzimologia , Neoplasias Ovarianas/ultraestrutura , Transdução de Sinais/efeitos dos fármacos , Esfingosina/metabolismo , Esfingosina/farmacologiaRESUMO
Epithelial-mesenchymal transition (EMT), crucial during embryogenesis for new tissue and organ formation, is also considered to be a prerequisite to cancer metastasis. We report here that the protein tyrosine phosphatase Pez is expressed transiently in discrete locations in developing brain, heart, pharyngeal arches, and somites in zebrafish embryos. We also find that Pez knock-down results in defects in these organs, indicating a crucial role in organogenesis. Overexpression of Pez in epithelial MDCK cells causes EMT, with a drastic change in cell morphology and function that is accompanied by changes in gene expression typical of EMT. Transfection of Pez induced TGFbeta signaling, critical in developmental EMT with a likely role also in oncogenic EMT. In zebrafish, TGFbeta3 is co- expressed with Pez in a number of tissues and its expression was lost from these tissues when Pez expression was knocked down. Together, our data suggest Pez plays a crucial role in organogenesis by inducing TGFbeta and EMT.
Assuntos
Células Epiteliais/citologia , Mesoderma/citologia , Organogênese , Proteínas Tirosina Fosfatases/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/embriologia , Peixe-Zebra/metabolismo , Animais , Núcleo Celular/metabolismo , Cães , Embrião não Mamífero/citologia , Embrião não Mamífero/metabolismo , Desenvolvimento Embrionário/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Humanos , Mesoderma/efeitos dos fármacos , Mesoderma/metabolismo , Oligonucleotídeos Antissenso/farmacologia , Organogênese/efeitos dos fármacos , Fenótipo , Transporte Proteico/efeitos dos fármacos , Proteínas Tirosina Fosfatases/genética , Proteínas Tirosina Fosfatases não Receptoras/genética , Proteínas Tirosina Fosfatases não Receptoras/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteína Smad4/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta1/genética , Fator de Crescimento Transformador beta1/metabolismo , Fator de Crescimento Transformador beta2/genética , Fator de Crescimento Transformador beta2/metabolismo , Fator de Crescimento Transformador beta3/genética , Fator de Crescimento Transformador beta3/metabolismo , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/genéticaRESUMO
The transactivation of enhanced growth factor receptor (EGFR) by G protein-coupled receptor (GPCR) ligands is recognized as an important signaling mechanism in the regulation of complex biological processes, such as cancer development. Estrogen (E2), which is a steroid hormone that is intimately implicated in breast cancer, has also been suggested to function via EGFR transactivation. In this study, we demonstrate that E2-induced EGFR transactivation in human breast cancer cells is driven via a novel signaling system controlled by the lipid kinase sphingosine kinase-1 (SphK1). We show that E2 stimulates SphK1 activation and the release of sphingosine 1-phosphate (S1P), by which E2 is capable of activating the S1P receptor Edg-3, resulting in the EGFR transactivation in a matrix metalloprotease-dependent manner. Thus, these findings reveal a key role for SphK1 in the coupling of the signals between three membrane-spanning events induced by E2, S1P, and EGF. They also suggest a new signal transduction model across three individual ligand-receptor systems, i.e., "criss-cross" transactivation.
Assuntos
Receptores ErbB/metabolismo , Estrogênios/farmacologia , Fosfotransferases (Aceptor do Grupo Álcool)/fisiologia , Receptores de Lisoesfingolipídeo/metabolismo , Transdução de Sinais , Linhagem Celular Tumoral , Receptores ErbB/efeitos dos fármacos , Feminino , Humanos , Modelos Biológicos , Ativação TranscricionalRESUMO
A number of recent reviews in the field have described many of the known growth factors and signalling pathways that may be involved in regulating epithelial-mesenchymal transitions. This perspective will focus on some aspects of posttranslational regulation of cell-cell adhesion that are less well understood and their potential role in initiating epithelial-mesenchymal transitions. In addition, a potential novel intermediate in the signalling pathway of epithelial-mesenchymal transition will also be described.
Assuntos
Adesão Celular , Células Epiteliais/citologia , Epitélio/embriologia , Regulação da Expressão Gênica no Desenvolvimento , Mesoderma/metabolismo , Junções Aderentes , Animais , Caderinas/metabolismo , Comunicação Celular , Linhagem Celular , Cães , Regulação para Baixo , Humanos , Proteínas Tirosina Fosfatases/metabolismo , Transdução de Sinais , Transcrição Gênica , CicatrizaçãoRESUMO
Cell-cell adhesion regulates processes important in embryonal development, normal physiology, and cancer progression. It is regulated by various mechanisms including tyrosine phosphorylation. We have previously shown that the protein tyrosine phosphatase Pez is concentrated at intercellular junctions in confluent, quiescent monolayers but is nuclear in cells lacking cell-cell contacts. We show here with an epithelial cell model that Pez localizes to the adherens junctions in confluent monolayers. A truncation mutant lacking the catalytic domain acts as a dominant negative mutant to upregulate tyrosine phosphorylation at adherens junctions. We identified beta-catenin, a component of adherens junctions, as a substrate of Pez by a "substrate trapping" approach and by in vitro dephosphorylation with recombinant Pez. Consistent with this, ectopic expression of the dominant negative mutant caused an increase in tyrosine phosphorylation of beta-catenin, demonstrating that Pez regulates the level of tyrosine phosphorylation of adherens junction proteins, including beta-catenin. Increased tyrosine phosphorylation of adherens junction proteins has been shown to decrease cell-cell adhesion, promoting cell migration as a result. Accordingly, the dominant negative Pez mutant enhanced cell motility in an in vitro "wound" assay. This suggests that Pez is also a regulator of cell motility, most likely through its action on cell-cell adhesion.