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AIMS: Microtubule inhibitors are widely used in first line cancer therapy, though drug resistance often develops and causes treatment failure. Colchicine binds to tubulins and inhibits tumor growth, but is not approved for cancer therapy due to systemic toxicity. In this study, we aim to improve the therapeutic index of colchicine through structural modification. METHODS: The methoxyl group of the tropolonic ring in colchicine was replaced with amino groups. The cross-resistance of the derivatives with paclitaxel and vincristine was tested. Antitumor effects of target compounds were tested in vivo in A549 and paclitaxel-resistant A549/T xenografts. The interaction of target compounds with tubulins was measured using biological and chemical methods. RESULTS: Methylamino replacement of the tropolonic methoxyl group of colchicine increases, while demethylation loses, selective tubulin binding affinity, G2/M arrest and antiproliferation activity. Methylaminocolchicine is more potent than paclitaxel and vincristine to inhibit tumor growth in vitro and in vivo without showing cross-resistance to paclitaxel. Methylaminocolchicine binds to tubulins in unique patterns and inhibits P-gp with a stable pharmacokinetic profile. CONCLUSION: Methylanimo replacement of the tropolonic methoxyl group of colchicine increases antitumor activity with improved therapeutic index. Methylaminocolchicine represents a new type of mitotic inhibitor with the ability of overcoming paclitaxel and vincristine resistance.
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Antineoplásicos , Neoplasias , Humanos , Paclitaxel/farmacología , Paclitaxel/química , Paclitaxel/uso terapéutico , Colchicina/farmacología , Colchicina/química , Colchicina/metabolismo , Tubulina (Proteína) , Vincristina/farmacología , Vincristina/uso terapéutico , Neoplasias/tratamiento farmacológico , Línea Celular Tumoral , Antineoplásicos/uso terapéuticoRESUMEN
BACKGROUND & AIMS: Agents that induce an immune response against tumors by altering T-cell regulation have increased survival times of patients with advanced-stage tumors, such as melanoma or lung cancer. We aimed to characterize molecular features of immune cells that infiltrate hepatocellular carcinomas (HCCs) to determine whether these types of agents might be effective against liver tumors. METHODS: We analyzed HCC samples from 956 patients. We separated gene expression profiles from tumor, stromal, and immune cells using a non-negative matrix factorization algorithm. We then analyzed the gene expression pattern of inflammatory cells in HCC tumor samples. We correlated expression patterns with the presence of immune cell infiltrates and immune regulatory molecules, determined by pathology and immunohistochemical analyses, in a training set of 228 HCC samples. We validated the correlation in a validation set of 728 tumor samples. Using data from 190 tumors in the Cancer Genome Atlas, we correlated immune cell gene expression profiles with numbers of chromosomal aberrations (based on single-nucleotide polymorphism array) and mutations (exome sequence data). RESULTS: We found approximately 25% of HCCs to have markers of an inflammatory response, with high expression levels of the CD274 molecule (programmed death-ligand 1) and programmed cell death 1, markers of cytolytic activity, and fewer chromosomal aberrations. We called this group of tumors the Immune class. It contained 2 subtypes, characterized by markers of an adaptive T-cell response or exhausted immune response. The exhausted immune response subclass expressed many genes regulated by transforming growth factor beta 1 that mediate immunosuppression. We did not observe any differences in numbers of mutations or expression of tumor antigens between the immune-specific class and other HCCs. CONCLUSIONS: In an analysis of HCC samples from 956 patients, we found almost 25% to express markers of an inflammatory response. We identified 2 subclasses, characterized by adaptive or exhausted immune responses. These findings indicate that some HCCs might be susceptible to therapeutic agents designed to block the regulatory pathways in T cells, such as programmed death-ligand 1, programmed cell death 1, or transforming growth factor beta 1 inhibitors.
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Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/inmunología , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/inmunología , Factor de Crecimiento Transformador beta1/genética , Inmunidad Adaptativa/genética , Anciano , Antígeno B7-H1/metabolismo , Carcinoma Hepatocelular/química , Carcinoma Hepatocelular/patología , Metilación de ADN , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Tolerancia Inmunológica/genética , Inmunohistoquímica , Inmunofenotipificación , Neoplasias Hepáticas/química , Neoplasias Hepáticas/patología , Linfocitos Infiltrantes de Tumor/inmunología , Linfocitos Infiltrantes de Tumor/metabolismo , Masculino , Persona de Mediana Edad , Mutación , Receptor de Muerte Celular Programada 1/metabolismo , Transducción de Señal , Transcriptoma , Microambiente Tumoral/inmunología , beta Catenina/genéticaRESUMEN
BACKGROUND & AIMS: Fibrolamellar hepatocellular carcinoma (FLC) is a rare primary hepatic cancer that develops in children and young adults without cirrhosis. Little is known about its pathogenesis, and it can be treated only with surgery. We performed an integrative genomic analysis of a large series of patients with FLC to identify associated genetic factors. METHODS: By using 78 clinically annotated FLC samples, we performed whole-transcriptome (n = 58), single-nucleotide polymorphism array (n = 41), and next-generation sequencing (n = 48) analyses; we also assessed the prevalence of the DNAJB1-PRKACA fusion transcript associated with this cancer (n = 73). We performed class discovery using non-negative matrix factorization, and functional annotation using gene-set enrichment analyses, nearest template prediction, ingenuity pathway analyses, and immunohistochemistry. The genomic identification of significant targets in a cancer algorithm was used to identify chromosomal aberrations, MuTect and VarScan2 were used to identify somatic mutations, and the random survival forest was used to determine patient prognoses. Findings were validated in an independent cohort. RESULTS: Unsupervised gene expression clustering showed 3 robust molecular classes of tumors: the proliferation class (51% of samples) had altered expression of genes that regulate proliferation and mammalian target of rapamycin signaling activation; the inflammation class (26% of samples) had altered expression of genes that regulate inflammation and cytokine enriched production; and the unannotated class (23% of samples) had a gene expression signature that was not associated previously with liver tumors. Expression of genes that regulate neuroendocrine function, as well as histologic markers of cholangiocytes and hepatocytes, were detected in all 3 classes. FLCs had few copy number variations; the most frequent were focal amplification at 8q24.3 (in 12.5% of samples), and deletions at 19p13 (in 28% of samples) and 22q13.32 (in 25% of samples). The DNAJB1-PRKACA fusion transcript was detected in 79% of samples. FLC samples also contained mutations in cancer-related genes such as BRCA2 (in 4.2% of samples), which are uncommon in liver neoplasms. However, FLCs did not contain mutations most commonly detected in liver cancers. We identified an 8-gene signature that predicted survival of patients with FLC. CONCLUSIONS: In a genomic analysis of 78 FLC samples, we identified 3 classes based on gene expression profiles. FLCs contain mutations and chromosomal aberrations not previously associated with liver cancer, and almost 80% contain the DNAJB1-PRKACA fusion transcript. By using this information, we identified a gene signature that is associated with patient survival time.
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Carcinoma Hepatocelular/genética , Regulación Neoplásica de la Expresión Génica/genética , Neoplasias Hepáticas/genética , Adolescente , Adulto , Anciano , Proliferación Celular/genética , Niño , Aberraciones Cromosómicas , Análisis por Conglomerados , Subunidades Catalíticas de Proteína Quinasa Dependientes de AMP Cíclico/genética , Variaciones en el Número de Copia de ADN , Femenino , Genoma , Proteínas del Choque Térmico HSP40/genética , Humanos , Inflamación/genética , Masculino , Persona de Mediana Edad , Polimorfismo de Nucleótido Simple , Análisis de Secuencia de ADN , Adulto JovenRESUMEN
BACKGROUND & AIMS: Cholangiocarcinoma, the second most common liver cancer, can be classified as intrahepatic cholangiocarcinoma (ICC) or extrahepatic cholangiocarcinoma. We performed an integrative genomic analysis of ICC samples from a large series of patients. METHODS: We performed a gene expression profile, high-density single-nucleotide polymorphism array, and mutation analyses using formalin-fixed ICC samples from 149 patients. Associations with clinicopathologic traits and patient outcomes were examined for 119 cases. Class discovery was based on a non-negative matrix factorization algorithm and significant copy number variations were identified by Genomic Identification of Significant Targets in Cancer (GISTIC) analysis. Gene set enrichment analysis was used to identify signaling pathways activated in specific molecular classes of tumors, and to analyze their genomic overlap with hepatocellular carcinoma (HCC). RESULTS: We identified 2 main biological classes of ICC. The inflammation class (38% of ICCs) is characterized by activation of inflammatory signaling pathways, overexpression of cytokines, and STAT3 activation. The proliferation class (62%) is characterized by activation of oncogenic signaling pathways (including RAS, mitogen-activated protein kinase, and MET), DNA amplifications at 11q13.2, deletions at 14q22.1, mutations in KRAS and BRAF, and gene expression signatures previously associated with poor outcomes for patients with HCC. Copy number variation-based clustering was able to refine these molecular groups further. We identified high-level amplifications in 5 regions, including 1p13 (9%) and 11q13.2 (4%), and several focal deletions, such as 9p21.3 (18%) and 14q22.1 (12% in coding regions for the SAV1 tumor suppressor). In a complementary approach, we identified a gene expression signature that was associated with reduced survival times of patients with ICC; this signature was enriched in the proliferation class (P < .001). CONCLUSIONS: We used an integrative genomic analysis to identify 2 classes of ICC. The proliferation class has specific copy number alterations, activation of oncogenic pathways, and is associated with worse outcome. Different classes of ICC, based on molecular features, therefore might require different treatment approaches.
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Colangiocarcinoma/genética , Colangiocarcinoma/mortalidad , Regulación Neoplásica de la Expresión Génica , Predisposición Genética a la Enfermedad/epidemiología , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/mortalidad , Anciano , Neoplasias de los Conductos Biliares , Conductos Biliares Intrahepáticos , Biopsia con Aguja , Colangiocarcinoma/clasificación , Colangiocarcinoma/patología , Variaciones en el Número de Copia de ADN , Análisis Mutacional de ADN , Bases de Datos Factuales , Progresión de la Enfermedad , Femenino , Perfilación de la Expresión Génica , Humanos , Inmunohistoquímica , Hibridación in Situ , Estimación de Kaplan-Meier , Neoplasias Hepáticas/clasificación , Neoplasias Hepáticas/patología , Masculino , Persona de Mediana Edad , Pronóstico , Estudios Retrospectivos , Análisis de SupervivenciaRESUMEN
Sin3A/B is a master transcriptional scaffold and corepressor that plays an essential role in the regulation of gene transcription and maintenance of chromatin structure, and its inappropriate recruitment has been associated with aberrant gene silencing in cancer. Sin3A/B are highly related, large, multidomian proteins that interact with a wide variety of transcription factors and corepressor components, and we examined whether disruption of the function of a specific domain could lead to epigenetic reprogramming and derepression of specific subsets of genes. To this end, we selected the Sin3A/B-paired amphipathic alpha-helices (PAH2) domain based on its established role in mediating the effects of a relatively small number of transcription factors containing a PAH2-binding motif known as the Sin3 interaction domain (SID). Here, we show that in both human and mouse breast cancer cells, the targeted disruption of Sin3 function by introduction of a SID decoy that interferes with PAH2 binding to SID-containing partner proteins reverted the silencing of genes involved in cell growth and differentiation. In particular, the SID decoy led to epigenetic reprogramming and reexpression of the important breast cancer-associated silenced genes encoding E-cadherin, estrogen receptor alpha, and retinoic acid receptor beta and impaired tumor growth in vivo. Interestingly, the SID decoy was effective in the triple-negative M.D. Anderson-Metastatic Breast-231 (MDA-MB-231) breast cancer cell line, restoring sensitivity to 17beta-estradiol, tamoxifen, and retinoids. Therefore, the development of small molecules that can block interactions between PAH2 and SID-containing proteins offers a targeted epigenetic approach for treating this type of breast cancer that may also have wider therapeutic implications.
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Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Proteínas Represoras/genética , Proteínas Represoras/metabolismo , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Neoplasias de la Mama/patología , Neoplasias de la Mama/terapia , Diferenciación Celular/genética , Línea Celular Tumoral , Ensamble y Desensamble de Cromatina , Cartilla de ADN/genética , Resistencia a Antineoplásicos/genética , Epigénesis Genética , Receptor alfa de Estrógeno/genética , Femenino , Marcación de Gen , Terapia Genética , Humanos , Neoplasias Mamarias Experimentales/genética , Neoplasias Mamarias Experimentales/metabolismo , Neoplasias Mamarias Experimentales/patología , Ratones , Unión Proteica , Estructura Terciaria de Proteína , Receptores de Ácido Retinoico/genética , Proteínas Represoras/antagonistas & inhibidores , Proteínas Represoras/química , Complejo Correpresor Histona Desacetilasa y Sin3RESUMEN
Corepressors play an essential role in nuclear receptor-mediated transcriptional repression. In general, corepressors directly bind to nuclear receptors via CoRNR boxes (L/I-X-X-I/V-I) in the absence of ligand and appear to act as scaffolds to further recruit chromatin remodeling complexes to specific target genes. Here, we describe the identification of the multiple LIM domain protein Ajuba as a unique corepressor for a subset of nuclear hormone receptors. Ajuba contains functional nuclear-receptor interacting motifs and selectively interacts with retinoic acid receptors (RARs) and rexinoid receptor (RXRs) subtypes in a ligand-dependent manner. Simultaneous mutation of these motifs abolishes RAR binding and concomitantly leads to loss of repression on RARE reporter genes. P19 cells depleted of Ajuba are highly sensitized to all-trans retinoic acid (atRA)-induced transcription and differentiation. In the absence of atRA, Ajuba can be readily found at the RARE control elements of RAR endogenous target genes. Stimulation of cells with atRA results in the dissociation of Ajuba from these regions. Moreover, we observed that coexpression of the known Ajuba binding partner Prmt5 (protein arginine methyltransferase-5) inhibited the Ajuba/RAR interaction. The high-affinity Ajuba-RAR/RXR interaction site overlaps the region responsible for Ajuba/Prmt5 binding, and thus binding appears to be mutually exclusive, providing a potential mechanism for these observations. Identification of Ajuba as a unique corepressor for nuclear receptors sheds new light on mechanisms for nuclear receptor-mediated repression and provides a unique target for developing more effective therapeutics to modulate this important pathway.
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Proteínas Co-Represoras/metabolismo , Regulación de la Expresión Génica/fisiología , Proteínas de Homeodominio/metabolismo , Receptores Citoplasmáticos y Nucleares/metabolismo , Transducción de Señal/fisiología , Tretinoina/metabolismo , Secuencias de Aminoácidos/genética , Secuencias de Aminoácidos/fisiología , Animales , Western Blotting , Línea Celular , Inmunoprecipitación de Cromatina , Humanos , Inmunoprecipitación , Proteínas con Dominio LIM , Luciferasas , Ratones , Microscopía Fluorescente , Unión Proteica , Proteína-Arginina N-Metiltransferasas/metabolismo , Receptores de Ácido Retinoico/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Acute myeloid leukemia (AML) patients with FLT3-ITD mutations are associated with poor prognosis. FLT3-ITD inhibitors are developed and result in transient disease remission, but generally resistance develops. We propose that resistance occurs due to apoptosis evasion. We compared the abilities of five clinically used FLT3-ITD inhibitors, namely, midostaurin, crenolanib, gilteritinib, quizartinib, and sorafenib, to induce apoptosis. These drugs inhibit FLT3-ITD and induce apoptosis. Apoptosis induction is associated with GSK3ß activation, Mcl-1 downregulation, and Bim upregulation. Sorafenib-resistant MOLM-13/sor cells have the secondary D835Y mutation and increased Axl signaling pathway with cross-resistance to quizartinib. Gilteritinib and crenolanib inhibit both FLT3-ITD and Axl and induce apoptosis in MOLM-13/sor cells, in which they activate GSK3ß and downregulate Mcl-1. Inactivation of GSK3ß through phosphorylation and inhibitors blocks apoptosis and Mcl-1 reduction. The Axl/GSK3ß/Mcl-1 axis works as a feedback mechanism to attenuate apoptosis of FLT3-ITD inhibition. Homoharringtonine decreases the protein levels of Mcl-1, FLT3-ITD, and Axl. Moreover, it synergistically induces apoptosis with gilteritinib in vitro and prolongs survival of MOLM-13/sor xenografts. The GSK3ß/Mcl-1 axis works as the hub of FLT3-ITD inhibitors and plays a critical role in resistance against FLT3-ITD AML-targeted therapy.
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BACKGROUND & AIMS: Hepatocellular carcinoma (HCC) is a heterogeneous tumor that develops via activation of multiple pathways and molecular alterations. It has been a challenge to identify molecular classes of HCC and design treatment strategies for each specific subtype. MicroRNAs (miRNAs) are involved in HCC pathogenesis, and their expression profiles have been used to classify cancers. We analyzed miRNA expression in human HCC samples to identify molecular subclasses and oncogenic miRNAs. METHODS: We performed miRNA profiling of 89 HCC samples using a ligation-mediated amplification method. Subclasses were identified by unsupervised clustering analysis. We identified molecular features specific for each subclass using expression pattern (Affymetrix U133 2.0; Affymetrix, Santa Clara, CA), DNA change (Affymetrix STY Mapping Array), mutation (CTNNB1), and immunohistochemical (phosphor[p]-protein kinase B, p-insulin growth factor-IR, p-S6, p-epidermal growth factor receptor, ß-catenin) analyses. The roles of selected miRNAs were investigated in cell lines and in an orthotopic model of HCC. RESULTS: We identified 3 main clusters of HCCs: the wingless-type MMTV integration site (32 of 89; 36%), interferon-related (29 of 89; 33%), and proliferation (28 of 89; 31%) subclasses. A subset of patients with tumors in the proliferation subclass (8 of 89; 9%) overexpressed a family of poorly characterized miRNAs from chr19q13.42. Expression of miR-517a and miR-520c (from ch19q13.42) increased proliferation, migration, and invasion of HCC cells in vitro. MiR-517a promoted tumorigenesis and metastatic dissemination in vivo. CONCLUSIONS: We propose miRNA-based classification of 3 subclasses of HCC. Among the proliferation class, miR-517a is an oncogenic miRNA that promotes tumor progression. There is rationale for developing therapies that target miR-517a for patients with HCC.
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Carcinoma Hepatocelular/clasificación , Regulación Neoplásica de la Expresión Génica , Neoplasias Hepáticas/clasificación , MicroARNs/genética , ARN Neoplásico/genética , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Línea Celular Tumoral , Proliferación Celular , Progresión de la Enfermedad , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , MicroARNs/biosíntesis , Reacción en Cadena de la Polimerasa , Células Tumorales CultivadasRESUMEN
All-trans retinoic acid (ATRA)/arsenic trioxide (ATO) combination-based therapy has benefitted newly diagnosed acute promyelocytic leukemia (APL) in short-term studies, but the long-term efficacy and safety remained unclear. From April 2001, we have followed 85 patients administrated ATRA/ATO with a median follow-up of 70 months. Eighty patients (94.1%) entered complete remission (CR). Kaplan-Meier estimates of the 5-year event-free survival (EFS) and overall survival (OS) for all patients were 89.2% +/- 3.4% and 91.7% +/- 3.0%, respectively, and the 5-year relapse-free survival (RFS) and OS for patients who achieved CR (n = 80) were 94.8% +/- 2.5% and 97.4% +/- 1.8%, respectively. Upon ATRA/ATO, prognosis was not influenced by initial white blood cell count, distinct PML-RARalpha types, or FLT3 mutations. The toxicity profile was mild and reversible. No secondary carcinoma was observed, and 24 months after the last dose of ATRA/ATO, patients had urine arsenic concentrations well below the safety limit. These results demonstrate the high efficacy and minimal toxicity of ATRA/ATO treatment for newly diagnosed APL in long-term follow-up, suggesting a potential frontline therapy for de novo APL.
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Arsenicales/efectos adversos , Arsenicales/uso terapéutico , Leucemia Promielocítica Aguda/diagnóstico , Leucemia Promielocítica Aguda/tratamiento farmacológico , Óxidos/efectos adversos , Óxidos/uso terapéutico , Tretinoina/efectos adversos , Tretinoina/uso terapéutico , Acuaporinas/genética , Acuaporinas/metabolismo , Trióxido de Arsénico , Regulación Leucémica de la Expresión Génica/efectos de los fármacos , Leucemia Promielocítica Aguda/genética , Leucemia Promielocítica Aguda/metabolismo , Proteínas de Fusión Oncogénica/genética , Proteínas de Fusión Oncogénica/metabolismo , Pronóstico , Tasa de Supervivencia , Factores de TiempoRESUMEN
SIN3A, a scaffold protein has regulatory functions in tumor biology. Through its Paired amphipathic helix (PAH2) domain, SIN3A interacts with PHF12 (PF1), a protein with SIN3 interaction domain (SID) that forms a complex with MRG15 and KDM5A/B. These components are often overexpressed in cancer. In the present study, we evaluated the role of SIN3A and its interacting partner PF1 in mediating inhibition of tumor growth and invasion in triple negative breast cancer (TNBC). We found profound inhibition of invasion, migration, and induction of cellular senescence by specific disruption of the PF1/SIN3A PAH2 domain interaction in TNBC cells expressing PF1-SID transcript or peptide treatment. Genome-wide transcriptomic analysis by RNA-seq revealed that PF1-SID downregulates several gene sets and pathways linked to invasion and migration. Integrin α6 (ITGA6) and integrin ß1 (ITGB1) and their downstream target proteins were downregulated in PF1-SID cells. We further determined increased presence of SIN3A and transcriptional repressor, KLF9, on promoters of ITGA6 and ITGB1 in PF1-SID cells. Knockdown of KLF9 leads to re-expression of ITGA6 and ITGB1 and restoration of the invasive phenotype, functionally linking KLF9 to this process. Overall, these data demonstrate that specific disruption of PF1/SIN3A, inhibits tumor growth, migration, and invasion. Also, PF1-SID not only inhibits tumor growth by senescence induction and reduced proliferation, but it also targets cancer stem cell gene expression and blocks mammosphere formation. Overall, these data demonstrate a mechanism whereby invasion and metastasis of TNBC can be suppressed by inhibiting SIN3A-PF1 interaction and enhancing KLF9 mediated suppression of ITGA6 and ITGB1.
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Retinoids are essential in balancing proliferation, differentiation and apoptosis, and they exert their effects through retinoic acid receptors (RARs) and retinoid X receptors (RXRs). RARß is a tumor-suppressor gene silenced by epigenetic mechanisms such as DNA methylation in breast, cervical and non-small cell lung cancers. An increased expression of RARß has been associated with improved breast cancer-specific survival. The PAH2 domain of the scaffold protein SIN3A interacts with the specific Sin3 Interaction Domain (SID) of several transcription factors, such as MAD1, bringing chromatin-modifying proteins such as histone deacetylases, and it targets chromatin for specific modifications. Previously, we have established that blocking the PAH2-mediated Sin3A interaction with SID-containing proteins using SID peptides or small molecule inhibitors (SMI) increased RARß expression and induced retinoic acid metabolism in breast cancer cells, both in in vitro and in vivo models. Here, we report studies designed to understand the mechanistic basis of RARß induction and function. Using human breast cancer cells transfected with MAD1 SID or treated with the MAD SID peptide, we observed a dissociation of MAD1, RARα and RARß from Sin3A in a coimmunoprecipitation assay. This was associated with increased RARα and RARß expression and function by a luciferase assay, which was enhanced by the addition of AM580, a specific RARα agonist; EMSA showed that MAD1 binds to E-Box, similar to MYC, on the RARß promoter, which showed a reduced enrichment of Sin3A and HDAC1 by ChIP and was required for the AM580-enhanced RARß activation in MAD1/SID cells. These data suggest that the Sin3A/HDAC1/2 complex co-operates with the classical repressors in regulating RARß expression. These data suggest that SIN3A/MAD1 acts as a second RARß repressor and may be involved in fine-tuning retinoid sensitivity.
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Neoplasias de la Mama , Proteínas de Ciclo Celular , Receptores de Ácido Retinoico , Complejo Correpresor Histona Desacetilasa y Sin3 , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Proteínas de Ciclo Celular/genética , Cromatina , Femenino , Humanos , Receptores de Ácido Retinoico/metabolismo , Proteínas Represoras/metabolismo , Complejo Correpresor Histona Desacetilasa y Sin3/genéticaRESUMEN
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.
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Citarabina , Leucemia Mieloide Aguda , Anciano , Animales , Apoptosis , Proteínas Reguladoras de la Apoptosis , Artesunato/farmacología , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Compuestos Bicíclicos Heterocíclicos con Puentes/uso terapéutico , Línea Celular Tumoral , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1) , Citarabina/farmacología , Citarabina/uso terapéutico , Humanos , Leucemia Mieloide Aguda/patología , Ratones , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , SulfonamidasRESUMEN
Fibrolamellar carcinoma is a rare primary malignant liver neoplasm that usually affects adolescents and young adults with no underlying liver disease. Morphologically, the tumor cells resemble oncocytic hepatocytes arranged in cords with a stroma of lamellated collagen fibers. Immunohistochemical studies have found that fibrolamellar carcinomas express markers associated with both biliary (CK7 and epithelial membrane antigen) and hepatocytic (heppar-1and glypican-3) differentiation, as well as markers associated with hepatic progenitor cells (CK19 and EpCAM) and stem cells (CD133 and CD44). Genetic studies show fewer alterations compared with classic hepatocellular carcinoma. Pooled data from comparative genomic hybridization studies show that fibrolamellar carcinomas have fewer and less frequent genomic alterations when compared with classic hepatocellular carcinoma, cholangiocarcinoma, and hepatoblastoma. Of the alterations seen in fibrolamellar carcinoma, the most frequent are gains in 1q and 8q (also frequently seen in other hepatic tumors) and loss of 18q. Fibrolamellar carcinoma also has less frequent methylation of tumor suppressor promoters compared with hepatocellular carcinoma and minimal alterations in mitochondrial DNA. Fibrolamellar carcinoma is associated with better survival than hepatocellular carcinoma and cholangiocarcinoma, presumably due to the young age of the patients and the lack of cirrhosis. These features make more aggressive surgical therapy possible. There is currently very little information on the effectiveness of chemotherapy for fibrolamellar carcinoma.
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Biomarcadores de Tumor/genética , Neoplasias Hepáticas/genética , Adolescente , Adulto , Biomarcadores de Tumor/análisis , Carcinoma Hepatocelular/química , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/mortalidad , Carcinoma Hepatocelular/patología , Carcinoma Hepatocelular/terapia , Niño , Epigénesis Genética , Femenino , Regulación Neoplásica de la Expresión Génica , Predisposición Genética a la Enfermedad , Humanos , Inmunohistoquímica , Neoplasias Hepáticas/química , Neoplasias Hepáticas/mortalidad , Neoplasias Hepáticas/patología , Neoplasias Hepáticas/terapia , Masculino , Persona de Mediana Edad , Fenotipo , Pronóstico , Adulto JovenRESUMEN
PURPOSE OF REVIEW: Since the 1970s, the concept of differentiation therapy has been viewed as a promising and revolutionary approach for the treatment of acute myeloid leukemia (AML) and other cancers. However, the successful clinical application of differentiation therapy has only been realized since the late 1980s and only in one subtype of AML, acute promyelocytic leukemia (APL). The use of all-trans-retinoic acid (ATRA) and arsenic trioxide, both of which induce degradation of the progressive multifocal leukoencephalopathy/retinoic acid receptor alpha oncoprotein, in combination with chemotherapy is currently the accepted treatment of APL, presenting a potential paradigm for differentiation therapy in clinical oncology. RECENT FINDINGS: We have begun to understand why ATRA fails to induce differentiation in AML. The underlying reasons identified thus far are associated with an inability to target the removal of leukemogenic fusion proteins, aberrant epigenetic regulation of genes involved in the ATRA signaling pathway and the presence of factors that interfere with proper retinoic acid receptor alpha function. SUMMARY: Here, we examine the reasons why the exquisite sensitivity of APL to ATRA-based differentiation therapy has not been extended to other of AML subtypes. Current differentiation-based combinatorial approaches to target AML will also be analyzed. Finally, we will evaluate the potential of novel strategies, high-throughput screening, and functional genomics to uncover new differentiation-based therapies for AML.
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Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Arsenicales/uso terapéutico , Leucemia Mieloide Aguda/tratamiento farmacológico , Óxidos/uso terapéutico , Tretinoina/uso terapéutico , Trióxido de Arsénico , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Receptores de Ácido Retinoico/metabolismo , Receptor alfa de Ácido Retinoico , Sensibilidad y Especificidad , Tretinoina/metabolismoRESUMEN
BACKGROUND: As a higher proportion of adults live beyond 85 years, their cancer burden is expected to increase. While trends among the oldest old are established for major epithelial cancers (breast, prostate, lung, and colorectal cancers), they are less studied for minor cancers. This study describes age trends of cancer mortality, with emphasis on individuals aged 85+ years. RESULTS: Overall cancer mortality peaked at 85 years old and decreased or stabilized for all countries except the USA, France, and Japan, in which mortality continued to increase after age 85 years. For most countries, cancers of the oesophagus, stomach, liver, and larynx have a similar flat trend patterns across all ages. Bladder and kidney cancers as well as non-Hodgkin lymphoma, multiple myeloma, and leukemia showed a decreasing pattern after 85 years for UK, Germany, Italy and Poland. Lung cancer peaked at 80 years, although the age-specific peak among women did not follow the same pattern among all countries. Breast and prostate cancers increased after 85 years. CONCLUSION: Mortality stabilized or decreased after age 85, particularly for non-hormonal cancers. Whether this reflects a true biological levelling of mortality rates, or lower validity of cancer registration among the oldest old, remains open to discussion. METHODS: Completed death data were obtained from the World Health Organization (WHO) for eight countries (2000 to 2014). Age-specific mortality rates were calculated for each 5-year age group above age 64. Joinpoint regression models were used to identify significant changes in mortality trends by age.
RESUMEN
BACKGROUND & AIMS: The advent of targeted therapies in hepatocellular carcinoma (HCC) has underscored the importance of pathway characterization to identify novel molecular targets for treatment. We evaluated mTOR signaling in human HCC, as well as the antitumoral effect of a dual-level blockade of the mTOR pathway. METHODS: The mTOR pathway was assessed using integrated data from mutation analysis (direct sequencing), DNA copy number changes (SNP-array), messenger RNA levels (quantitative reverse-transcription polymerase chain reaction and gene expression microarray), and protein activation (immunostaining) in 351 human samples [HCC (n = 314) and nontumoral tissue (n = 37)]. Effects of dual blockade of mTOR signaling using a rapamycin analogue (everolimus) and an epidermal/vascular endothelial growth factor receptor inhibitor (AEE788) were evaluated in liver cancer cell lines and in a xenograft model. RESULTS: Aberrant mTOR signaling (p-RPS6) was present in half of the cases, associated with insulin-like growth factor pathway activation, epidermal growth factor up-regulation, and PTEN dysregulation. PTEN and PI3KCA-B mutations were rare events. Chromosomal gains in RICTOR (25% of patients) and positive p-RPS6 staining correlated with recurrence. RICTOR-specific siRNA down-regulation reduced tumor cell viability in vitro. Blockage of mTOR signaling with everolimus in vitro and in a xenograft model decelerated tumor growth and increased survival. This effect was enhanced in vivo after epidermal growth factor blockade. CONCLUSIONS: MTOR signaling has a critical role in the pathogenesis of HCC, with evidence for the role of RICTOR in hepato-oncogenesis. MTOR blockade with everolimus is effective in vivo. These findings establish a rationale for targeting the mTOR pathway in clinical trials in HCC.
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Carcinoma Hepatocelular/genética , Regulación Neoplásica de la Expresión Génica , Neoplasias Hepáticas/genética , Proteínas Quinasas/genética , ARN Neoplásico/genética , Adulto , Anciano , Anciano de 80 o más Años , Animales , Apoptosis , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patología , Línea Celular Tumoral , Femenino , Humanos , Immunoblotting , Inmunohistoquímica , Etiquetado Corte-Fin in Situ , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , Neoplasias Hepáticas Experimentales/genética , Neoplasias Hepáticas Experimentales/metabolismo , Neoplasias Hepáticas Experimentales/patología , Masculino , Ratones , Ratones Desnudos , Persona de Mediana Edad , Fosfatidilinositol 3-Quinasas , Proteínas Quinasas/biosíntesis , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transducción de Señal/genética , Serina-Treonina Quinasas TORRESUMEN
The mechanism of apoptosis induced by treatment with As(2)O(3) alone or in combination with buthionine sulfoximine (BSO) was studied in NB4, U937, Namalwa, and Jurkat cells. As(2)O(3) at concentrations <2 micromol/L induced apoptosis in NB4 cells and Namalwa cells but not in U937 and Jurkat cells. As(2)O(3)-induced apoptosis in NB4 cells and Namalwa cells correlated with increase of H(2)O(2) and caspase activation without activation of c-Jun NH(2)-terminal kinase (JNK). BSO (10 micromol/L) depleted the reduced form of intracellular glutathione without inducing apoptosis but synergized with 1 micromol/L As(2)O(3) to induce apoptosis in all four cell lines. This synergy correlated with JNK activation. Treatment with As(2)O(3) plus BSO, but not with As(2)O(3) alone, increased the levels of death receptor (DR) 5 protein and caspase-8 cleavage. The JNK inhibitor SP600125 inhibited the increase in DR5 protein and attenuated apoptosis induced by treatment with As(2)O(3) plus BSO. These observations suggest that a DR-mediated pathway activated by JNK is involved in apoptosis induced by treatment with As(2)O(3) plus BSO.
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Apoptosis/efectos de los fármacos , Arsenicales/farmacología , Butionina Sulfoximina/farmacología , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Óxidos/farmacología , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Clorometilcetonas de Aminoácidos/farmacología , Antracenos/farmacología , Trióxido de Arsénico , Western Blotting , Caspasa 3/metabolismo , Inhibidores de Caspasas , Proliferación Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Sinergismo Farmacológico , Activación Enzimática/efectos de los fármacos , Glutatión/metabolismo , Humanos , Peróxido de Hidrógeno/metabolismo , Proteínas Quinasas JNK Activadas por Mitógenos/antagonistas & inhibidores , Células Jurkat , Leucemia/metabolismo , Leucemia/patología , Linfoma/metabolismo , Linfoma/patología , Factores de Tiempo , Células U937RESUMEN
Arsenic trioxide (ATO) has been found to be an effective treatment for acute promyelocytic leukemia patients and is being tested for treating other hematologic malignancies. We have previously shown that AML1/MDS1/EVI1 (AME), a fusion gene generated by a t(3;21)(q26;q22) translocation found in patients with chronic myelogenous leukemia during blast phase, myelodysplastic syndrome, or acute myelogenous leukemia (AML), impairs hematopoiesis and eventually induces an AML in mice. Both fusion partners of AME, AML1 and MDS1/EVI1, encode transcription factors and are also targets of a variety of genetic abnormalities in human hematologic malignancies. In addition, aberrant expression of ectopic viral integration site 1 (EVI1) has also been found in solid tumors, such as ovarian and colon cancers. In this study, we examined whether ATO could target AME and related oncoproteins. We found that ATO used at therapeutic levels degrades AME. The ATO treatment induces differentiation and apoptosis in AME leukemic cells in vitro as well as reduces tumor load and increases the survival of mice transplanted with these cells. We further found that ATO targets AME via both myelodysplastic syndrome 1 (MDS1) and EVI1 moieties and degrades EVI1 via the ubiquitin-proteasome pathway and MDS1 in a proteasome-independent manner. Our results suggest that ATO could be used as a part of targeted therapy for AME-, AML1/MDS1-, MDS1/EVI1-, and EVI1-positive human cancers.
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Arsenicales/farmacología , Subunidad alfa 2 del Factor de Unión al Sitio Principal/metabolismo , Proteínas de Fusión Oncogénica/metabolismo , Óxidos/farmacología , Proteínas Represoras/metabolismo , Animales , Apoptosis/efectos de los fármacos , Trióxido de Arsénico , Western Blotting , Diferenciación Celular/efectos de los fármacos , Línea Celular Tumoral , Subunidad alfa 2 del Factor de Unión al Sitio Principal/genética , Relación Dosis-Respuesta a Droga , Regulación hacia Abajo/efectos de los fármacos , Citometría de Flujo , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Inhibidores de Crecimiento/farmacología , Humanos , Leucemia Experimental/genética , Leucemia Experimental/patología , Leucemia Experimental/prevención & control , Masculino , Ratones , Ratones Endogámicos BALB C , Células 3T3 NIH , Proteínas de Fusión Oncogénica/genética , Complejo de la Endopetidasa Proteasomal/metabolismo , Proteínas Represoras/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transducción de Señal/efectos de los fármacos , Análisis de Supervivencia , Transfección , Ubiquitina/metabolismoRESUMEN
Acute myeloid leukemia (AML) with Fms-related tyrosine kinase 3 internal tandem duplication (FLT3-ITD) mutation is notoriously hard to treat. We identified two drugs that together form an effective combination therapy against FLT3-ITD AML. One of the drugs, Sorafenib, an inhibitor of FLT3-ITD and other kinase activity, produces an impressive but short-lived remission in FLT3-ITD AML patients. The second, arsenic trioxide (ATO), at therapeutically achievable concentrations, reduces the level of FLT3-ITD and Mcl-1 proteins, and induces apoptosis in leukemic cell lines and in primary cells expressing FLT3-ITD. We linked this relative sensitivity to ATO to low levels of reduced glutathione. While producing proapoptotic effects, ATO treatment also has an unwanted effect whereby it causes the accumulation of the phosphorylated (inactive) form of glycogen synthase kinase 3ß (GSK3ß), a kinase necessary for apoptosis. When ATO is combined with Sorafenib, GSK3ß is activated, Mcl-1 is further reduced, and proapoptotic proteins Bak and Bax are activated. Mice xenografted with FLT3-ITD MOLM13 cell line treated with the Sorafenib/ATO combination have significantly improved survival. This combination has potential to improve the therapeutic outcome of FLT3-ITD-targeted therapy of AML patients. Mol Cancer Ther; 17(9); 1871-80. ©2018 AACR.
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Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Leucemia Mieloide/tratamiento farmacológico , Mutaciones Letales Sintéticas/efectos de los fármacos , Ensayos Antitumor por Modelo de Xenoinjerto , Tirosina Quinasa 3 Similar a fms/genética , Enfermedad Aguda , Animales , Apoptosis/efectos de los fármacos , Trióxido de Arsénico/administración & dosificación , Línea Celular Tumoral , Humanos , Estimación de Kaplan-Meier , Leucemia Mieloide/genética , Leucemia Mieloide/patología , Ratones Endogámicos NOD , Ratones Noqueados , Ratones SCID , Sorafenib/administración & dosificación , Células THP-1 , Secuencias Repetidas en Tándem/genéticaRESUMEN
BACKGROUND: Tumorigenesis is associated with changes in gene expression and involves many pathways. Dysregulated genes include "housekeeping" genes that are often used for normalization for quantitative real-time RT-PCR (qPCR), which may lead to unreliable results. This study assessed eight stages of hepatitis C virus (HCV) induced hepatocellular carcinoma (HCC) to search for appropriate genes for normalization. RESULTS: Gene expression profiles using microarrays revealed differential expression of most "housekeeping" genes during the course of HCV-HCC, including glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and beta-actin (ACTB), genes frequently used for normalization. QPCR reactions confirmed the regulation of these genes. Using them for normalization had strong effects on the extent of differential expressed genes, leading to misinterpretation of the results. CONCLUSION: As shown here in the case of HCV-induced HCC, the most constantly expressed gene is the arginine/serine-rich splicing factor 4 (SFRS4). The utilization of at least two genes for normalization is robust and advantageous, because they can compensate for slight differences of their expression when not co-regulated. The combination of ribosomal protein large 41 (RPL41) and SFRS4 used for normalization led to very similar results as SFRS4 alone and is a very good choice for reference in this disease as shown on four differentially expressed genes.