RESUMEN
BACKGROUND: Despite the advances of therapies, multiple myeloma (MM) remains an incurable hematological cancer that most patients experience relapse. Tumor angiogenesis is strongly correlated with cancer relapse. Human leukocyte antigen G (HLA-G) has been known as a molecule to suppress angiogenesis. We aimed to investigate whether soluble HLA-G (sHLA-G) was involved in the relapse of MM. METHODS: We first investigated the dynamics of serum sHLA-G, vascular endothelial growth factor (VEGF) and interleukin 6 (IL-6) in 57 successfully treated MM patients undergoing remission and relapse. The interactions among these angiogenesis-related targets (sHLA-G, VEGF and IL-6) were examined in vitro. Their expression at different oxygen concentrations was investigated using a xenograft animal model by intra-bone marrow and skin grafts with myeloma cells. RESULTS: We found that HLA-G protein degradation augmented angiogenesis. Soluble HLA-G directly inhibited vasculature formation in vitro. Mechanistically, HLA-G expression was regulated by hypoxia-inducible factor-1α (HIF-1α) in MM cells under hypoxia. We thus developed two mouse models of myeloma xenografts in intra-bone marrow (BM) and underneath the skin, and found a strong correlation between HLA-G and HIF-1α expressions in hypoxic BM, but not in oxygenated tissues. Yet when stimulated with IL-6, both HLA-G and HIF-1α could be targeted to ubiquitin-mediated degradation via PARKIN. CONCLUSION: These results highlight the importance of sHLA-G in angiogenesis at different phases of multiple myeloma. The experimental evidence that sHLA-G as an angiogenesis suppressor in MM may be useful for future development of novel therapies to prevent relapse.
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Antígenos HLA-G , Interleucina-6 , Mieloma Múltiple , Neovascularización Patológica , Mieloma Múltiple/sangre , Mieloma Múltiple/metabolismo , Mieloma Múltiple/patología , Humanos , Animales , Neovascularización Patológica/metabolismo , Antígenos HLA-G/sangre , Antígenos HLA-G/metabolismo , Ratones , Interleucina-6/sangre , Interleucina-6/metabolismo , Masculino , Femenino , Factor A de Crecimiento Endotelial Vascular/metabolismo , Factor A de Crecimiento Endotelial Vascular/sangre , Persona de Mediana Edad , Línea Celular Tumoral , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Anciano , Modelos Animales de Enfermedad , AngiogénesisRESUMEN
BACKGROUND: Clinical evidence for the rapidity and effectiveness of fentanyl buccal soluble film (FBSF) in reducing pain intensity of breakthrough cancer pain (BTcP) remains inadequate. This study aimed to evaluate the efficacy of FBSF proportional to the around-the-clock (ATC) opioid regimens in rapidly relieving the intensity of BTcP episodes by determining the percentage of patients requiring further dose titration. METHODS: The study procedure included a dose-finding period followed by a 14-day observation period. Pain intensity was recorded with a Numeric Rating Scale (NRS) at onset and 5, 10, 15, and 30 min after FBSF self-administration. Meaningful pain relief was defined as the final NRS score ≤ 3. Satisfaction survey was conducted for each patient after treatment using the Global Satisfaction Scale. RESULTS: A total of 63 BTcP episodes occurred in 30 cancer patients. Only one patient required rescue medication at first BTcP episode and then achieved meaningful pain relief after titrating FBSF by 200 µg. Most BTcP episodes relieved within 10 min. Of 63 BTcP episodes, 30 (47.6%), 46 (73.0%), and 53 (84.1%) relieved within 5, 10, and 15 min after FBSF administration. Only grade 1/2 adverse events were reported, including somnolence, malaise, and dizziness. Of the 63 BTcP episodes, 82.6% were rated as excellent/good satisfaction with FBSF. CONCLUSION: FBSF can be administrated "on demand" by cancer patients at the onset of BTcP, providing rapid analgesia by achieving meaningful pain relief within 10 min. TRIAL REGISTRATION: This study was retrospectively registered 24 December, 2021 at Clinicaltrial.gov (NCT05209906): https://clinicaltrials.gov/study/NCT05209906 .
Asunto(s)
Analgésicos Opioides , Dolor Irruptivo , Fentanilo , Humanos , Fentanilo/uso terapéutico , Fentanilo/administración & dosificación , Femenino , Masculino , Dolor Irruptivo/tratamiento farmacológico , Dolor Irruptivo/etiología , Persona de Mediana Edad , Analgésicos Opioides/uso terapéutico , Analgésicos Opioides/administración & dosificación , Anciano , Administración Bucal , Adulto , Dimensión del Dolor/métodos , Dolor en Cáncer/tratamiento farmacológico , Manejo del Dolor/métodos , Manejo del Dolor/normas , Manejo del Dolor/estadística & datos numéricos , Neoplasias/complicaciones , Neoplasias/tratamiento farmacológico , Anciano de 80 o más AñosRESUMEN
The molecular pathogenesis of extranodal NK/T-cell lymphoma (NKTCL) remains obscured despite the next-generation sequencing (NGS) studies explored on ever larger cohorts in the last decade. We addressed the highly variable mutation frequencies reported among previous studies with comprehensive amplicon coverage and enhanced sequencing depth to achieve higher genomic resolution for novel genetic discovery and comparative mutational profiling of the oncogenesis of NKTCL. Targeted exome sequencing was conducted to interrogate 415 cancer-related genes in a cohort of 36 patients with NKTCL, and a total of 548 single nucleotide variants (SNVs) and 600 Copy number variances (CNVs) were identified. Recurrent amplification of the MCL1 (67%) and PIM1 (56%) genes was detected in a dominant majority of patients in our cohort. Functional mapping of genetic aberrations revealed that an enrichment of mutations in the JAK-STAT signaling pathway, including the cytokine receptor LIFR (copy number loss) upstream of JAK3, STAT3 (activating SNVs), and downstream effectors of MYC, PIM1 and MCL1 (copy number gains). RNA in situ hybridization showed the significant consistence of MCL1 RNA level and copy number of MCL1 gene. We further correlated molecular and clinical parameters with overall survival (OS) of these patients. When correlations were analyzed by univariate followed by multivariate modelling, only copy number loss of LIFR gene and stage (III-IV) were independent prognostic factors of reduced OS. Our findings identified that novel loss of LIFR gene significantly correlated with the adverse clinical outcome of NKTCL patients and provided therapeutic opportunities for this disease through manipulating LIFR.
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Linfoma Extranodal de Células NK-T , Mutación , Humanos , Linfoma Extranodal de Células NK-T/genética , Linfoma Extranodal de Células NK-T/terapia , Linfoma Extranodal de Células NK-T/mortalidad , Masculino , Femenino , Persona de Mediana Edad , Adulto , Anciano , Pronóstico , Variaciones en el Número de Copia de ADNRESUMEN
BCR-ABL, a fusion protein kinase, is a druggable target exclusively expressed in patients with chronic myeloid leukemia (CML). Several anti-leukemia medicines targeting this protein have been developed in recent years. However, therapeutic options are limited for CML patients bearing multiple BCR-ABL1 mutations. Ponatinib (PON), a potent tyrosinase inhibitor, was one of the approved drugs for managing BCR-ABL1 T315I mutant disease. However, treatment of patients with PON reported severe side effects related to cardiovascular events. Asciminib (ASC) was the first allosteric inhibitor approved to target the myristoyl pocket of BCR-ABL protein to inhibit protein activity. The different mechanism of inhibition opens the possibility of co-exposure with both medicines. Reports on cardiovascular side effects due to the combination use of PON + ASC in pre-clinical and clinical studies are minimal. Thus, this study aimed to observe the potential cardiovascular-related side effect after co-exposure to ASC and PON using zebrafish as an animal model. In this study, zebrafish were acutely exposed to both compounds. The cardiovascular physiology parameters and gene expression related to cardiovascular development were evaluated. We demonstrate that combining ASC with PON at no observed effect concentration (NOEC) did not cause any significant change in the cardiac performance parameter in zebrafish. However, a significant increase in nkx2.5 expression level and a substantial decrease in blood flow velocity were recorded, suggesting that combining these compounds at NOEC can cause mild cardiovascular-related side effects.
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Antineoplásicos , Leucemia Mielógena Crónica BCR-ABL Positiva , Piridazinas , Animales , Antineoplásicos/toxicidad , Resistencia a Antineoplásicos/genética , Proteínas de Fusión bcr-abl/genética , Imidazoles , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Monofenol Monooxigenasa , Niacinamida/análogos & derivados , Inhibidores de Proteínas Quinasas/farmacología , Pirazoles , Piridazinas/toxicidad , Pez CebraRESUMEN
Tyrosine kinase inhibitors (TKIs) that target BCR-ABL are the frontline treatments in chronic myeloid leukemia (CML). Growing evidence has shown that TKIs also enhance immunity. Since gamma-delta T (γδT) cells possess the potent anticancer capability, here we investigated the potential involvement of γδT cells in TKI treatments for CML. We characterized γδT cells isolated from chronic-phase CML patients before and during TKI treatments. γδT expression increased significantly in CML patients who achieved major molecular response (MMR) and deep molecular response (DMR). Their Vδ2 subset of γδT also expanded, and increased expression of activating molecules, namely IFN-γ, perforin, and CD107a, as well as γδT cytotoxicity. Mechanistically, TKIs augmented the efflux of isopentenyl pyrophosphate (IPP) from CML cells, which stimulated IFN-γ production and γδT expansion. Notably, the size of the IFN-γ+ naïve γδT population in TKI-treated CML patients was strongly correlated with their rates to reach DMR and with the duration on DMR. Statistical analysis suggests that a cutoff of 7.5% IFN-γ+ naïve subpopulation of γδT in CML patients could serve as a determinant for MR4.0 sustainability. Our results highlight γδT cells as a positive regulator for TKI responses in CML patients.
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Proteínas de Fusión bcr-abl/antagonistas & inhibidores , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Leucemia Mielógena Crónica BCR-ABL Positiva/inmunología , Inhibidores de Proteínas Quinasas/farmacología , Receptores de Antígenos de Linfocitos T gamma-delta/inmunología , Linfocitos T/inmunología , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Proteínas de Fusión bcr-abl/inmunología , Humanos , Células K562 , Leucemia Mielógena Crónica BCR-ABL Positiva/patología , Masculino , Persona de Mediana Edad , Linfocitos T/patologíaRESUMEN
The mechanism exhausting CD8+ T cells is not completely clear against tumors. Literature has demonstrated that cigarette smoking disables the immunological activity, so we propose nicotine is able to exhaust CD8+ T cells. The CD8+ T cells from healthy volunteers with and without cigarette smoking and the capacity of CD8+ T cells against tumor cells were investigated. RNAseq was used to investigate the gene profiling expression in CD8+ T cells. Meanwhile, small RNAseq was also used to search novel microRNAs involved in the exhaustion of CD8+ T cells. The effect of nicotine exhausting CD8+ T cells was investigated in vitro and in the humanized tumor xenografts in vivo. We found that CD8+ T cells were able to reduce cell viability in lung cancer HCC827 and A549 cells, that secreted granzyme B, but CD8+ T cells from the healthy cigarette smokers lost anti-HCC827 effect. Moreover, nicotine suppressed the anti-HCC827 effect of CD8+ T cells. RNAseq revealed lower levels of IL2RB and GZMB in the exhausted CD8+ T cells. We identified that miR-629-5p was increased by nicotine, that targeted IL2RB. Transfection of miR-629-5p mimic reduced IL2RB and GZMB levels. We further validated that nicotine reduced granzyme B levels using a nuclear imaging technique, and demonstrated that nicotine exhausted peripheral blood mononuclear cells against HCC827 growth in the humanized tumor xenografts. This study demonstrated that nicotine exhausted CD8+ T cells against HCC827 cells through increasing miR-629-5p to suppress IL2RB.
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Adenocarcinoma del Pulmón/metabolismo , Linfocitos T CD8-positivos/inmunología , Subunidad beta del Receptor de Interleucina-2/metabolismo , MicroARNs/genética , Nicotina/metabolismo , Células A549 , Animales , Línea Celular Tumoral , Fumar Cigarrillos/efectos adversos , Regulación Neoplásica de la Expresión Génica , Granzimas/genética , Granzimas/metabolismo , Humanos , Subunidad beta del Receptor de Interleucina-2/genética , Masculino , Ratones , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
DNA methylation plays several roles in regulating neuronal proliferation, differentiation, and physiological functions. The major de novo methyltransferase, DNMT3, controls the DNA methylation pattern in neurons according to environmental stimulations and behavioral regulations. Previous studies demonstrated that knockout of Dnmt3 induced mouse anxiety; however, controversial results showed that activation of Dnmt3 causes anxiolytic behavior. Thus, an alternative animal model to clarify Dnmt3 on modulating behavior is crucial. Therefore, we aimed to establish a zebrafish (Danio rerio) model to clarify the function of dnmt3 on fish behavior by behavioral endpoint analyses. We evaluated the behaviors of the wild type, dnmt3aa, and dnmt3ab knockout (KO) fish by the novel tank, mirror biting, predator avoidance, social interaction, shoaling, circadian rhythm locomotor activity, color preference, and short-term memory tests. The results indicated that the dnmt3aa KO fish possessed abnormal exploratory behaviors and less fear response to the predator. On the other hand, dnmt3ab KO fish displayed less aggression, fear response to the predator, and interests to interact with their conspecifics, loosen shoaling formation, and dysregulated color preference index ranking. Furthermore, both knockout fishes showed higher locomotion activity during the night cycle, which is a sign of anxiety. However, changes in some neurotransmitter levels were observed in the mutant fishes. Lastly, whole-genome DNA methylation sequencing demonstrates a potential network of Dnmt3a proteins that is responsive to behavioral alterations. To sum up, the results suggested that the dnmt3aa KO or dnmt3ab KO fish display anxiety symptoms, which supported the idea that Dnmt3 modulates the function involved in emotional control, social interaction, and cognition.
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ADN (Citosina-5-)-Metiltransferasas/genética , Animales , Ansiedad/genética , Control de la Conducta/métodos , Conducta Animal/fisiología , ADN/metabolismo , ADN (Citosina-5-)-Metiltransferasas/metabolismo , Metilación de ADN/genética , ADN Metiltransferasa 3A , Metilasas de Modificación del ADN/genética , Metilasas de Modificación del ADN/metabolismo , Modelos Animales de Enfermedad , Femenino , Masculino , Modelos Animales , Neurotransmisores , Pez Cebra/genética , Proteínas de Pez Cebra/genéticaRESUMEN
BAFF supports B-cell survival and homeostasis by activating the NF-κB pathway. While NF-κB is also involved in the priming signal of NLRP3 inflammasome, the role of BAFF in NLRP3 inflammasome regulation is unknown. Here we report BAFF engagement to BAFF receptor elicited both priming and activating signals for NLRP3 inflammasomes in primary B cells and B lymphoma cell lines. This induction of NLRP3 inflammasomes by BAFF led to increased NLRP3 and IL-1ß expression, caspase-1 activation, IL-1ß secretion, and pyroptosis. Mechanistically, BAFF activated NLRP3 inflammasomes by promoting the association of cIAP-TRAF2 with components of NLRP3 inflammasomes, and by inducing Src activity-dependent ROS production and potassium ion efflux. B-cell receptor (BCR) stimulation on the Lyn signaling pathway inhibited BAFF-induced Src activities and attenuated BAFF-induced NLRP3 inflammasome activation. These findings reveal an additional function of BAFF in B-cell homeostasis that is associated with BCR activities.
Asunto(s)
Factor Activador de Células B/metabolismo , Linfocitos B/metabolismo , Inflamasomas/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , HumanosRESUMEN
BACKGROUND: HER3 mediates drug resistance against epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors (TKIs), resulting in tumor relapse in lung cancers. Previously, we demonstrated that EGFR induces HER3 overexpression, which facilitates the formation of cancer stem-like tumorspheres. However, the cellular mechanism through which EGFR regulates HER3 expression remains unclear. We hypothesized that EGFR downstream of STAT3 participates in HER3 expression because STAT3 contributes to cancer stemness and survival of EGFR-TKI resistant cancers. METHODS: First, RNAseq was used to uncover potential genes involved in the formation of lung cancer HCC827-derived stem-like tumorspheres. EGFR-positive lung cancer cell lines, including HCC827, A549, and H1975, were individually treated with a panel containing 172 therapeutic agents targeting stem cell-associated genes to search for potential agents that could be applied against EGFR-positive lung cancers. In addition, gene knockdown and RNAseq were used to investigate molecular mechanisms through which STAT3 regulates tumor progression and the survival in lung cancer. RESULTS: BBI608, a STAT3 inhibitor, was a potential therapeutic agent that reduced the cell viability of EGFR-positive lung cancer cell lines. Notably, the inhibitory effects of BBI608 were similar with those associated with YM155, an ILF3 inhibitor. Both compounds reduced G9a-mediated HER3 expression. We also demonstrated that STAT3 upregulated G9a to silence miR-145-5p, which exacerbated HER3 expression in this study. CONCLUSIONS: The present study revealed that BBI608 could eradicate EGFR-positive lung cancers and demonstrated that STAT3 enhanced the expression of HER3 through miR-145-5p repression by G9a, indicating that STAT3 is a reliable therapeutic target against EGFR-TKI-resistant lung cancers.
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Antígenos de Histocompatibilidad/metabolismo , N-Metiltransferasa de Histona-Lisina/metabolismo , Neoplasias Pulmonares/metabolismo , Inhibidores de Proteínas Quinasas/farmacología , Receptor ErbB-3/metabolismo , Factor de Transcripción STAT3/metabolismo , Células A549 , Animales , Benzofuranos/farmacología , Movimiento Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Resistencia a Antineoplásicos/efectos de los fármacos , Receptores ErbB/antagonistas & inhibidores , Receptores ErbB/metabolismo , Técnicas de Silenciamiento del Gen , Antígenos de Histocompatibilidad/genética , N-Metiltransferasa de Histona-Lisina/genética , Humanos , Imidazoles/farmacología , Neoplasias Pulmonares/patología , Masculino , Ratones , Ratones Endogámicos NOD , Ratones SCID , MicroARNs/genética , MicroARNs/metabolismo , Naftoquinonas/farmacología , Proteínas del Factor Nuclear 90/antagonistas & inhibidores , Proteínas del Factor Nuclear 90/genética , Inhibidores de Proteínas Quinasas/efectos adversos , Receptor ErbB-3/antagonistas & inhibidores , Factor de Transcripción STAT3/antagonistas & inhibidores , Factor de Transcripción STAT3/genética , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
BACKGROUND: Cancer stem cells are capable of undergoing cell division after surviving cancer therapies, leading to tumor progression and recurrence. Inhibitory agents against cancer stem cells may be therapeutically used for efficiently eradicating tumors. Therefore, the aim of this study was to identify the relevant driver genes that maintain cancer stemness in epidermal growth factor receptor (EGFR)-positive colorectal cancer (CRC) cells and to discover effective therapeutic agents against these genes. METHODS: In this study, EGFR-positive cancer stem-like cells (CSLCs) derived from HCT116 and HT29 cells were used as study models for in vitro inductions. To identify the differential genes that maintain CSLCs, RNAseq analysis was conducted followed by bioinformatics analysis. Moreover, a panel containing 172 therapeutic agents targeting the various pathways of stem cells was used to identify effective therapeutics against CSLCs. RESULTS: RNAseq analysis revealed that 654 and 840 genes were significantly upregulated and downregulated, respectively, in the HCT116 CSLCs. Among these genes, notably, platelet-derived growth factor A (PDGFA) and signal transducer and activator of transcription 3 (STAT3) were relevant according to the cancer pathway analyzed using NetworkAnalyst. Furthermore, therapeutic screening revealed that the agents targeting STAT3 and Wnt signaling pathways were efficient in reducing the cell viabilities of both HCT116 and HT29 cells. Consequently, we discovered that STAT3 inhibition using homoharringtonine and STAT3 knockdown significantly reduced the formation and survival of HT29-derived tumorspheres. We also observed that STAT3 phosphorylation was regulated by epidermal growth factor (EGF) to induce PDGFA and Wnt signaling cascades. CONCLUSIONS: We identified the potential genes involved in tumorsphere formation and survival in selective EGFR-positive CRCs. The results reveal that the EGF-STAT3 signaling pathway promotes and maintains CRC stemness. In addition, a crosstalk between STAT3 and Wnt activates the Wnt/ß-catenin signaling pathway, which is also responsible for cancer stemness. Thus, STAT3 is a putative therapeutic target for CRC treatment.
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Neoplasias Colorrectales/genética , Receptores ErbB/genética , Células Madre Neoplásicas/patología , Factor de Transcripción STAT3/genética , Neoplasias Colorrectales/diagnóstico , Neoplasias Colorrectales/patología , Detección Precoz del Cáncer , Factor de Crecimiento Epidérmico/genética , Regulación Neoplásica de la Expresión Génica , Células HT29 , Humanos , Fosforilación , Factor de Crecimiento Derivado de Plaquetas/genética , Análisis de Secuencia de ARN , Vía de Señalización WntRESUMEN
The epidermal growth factor (EGF) receptor (EGFR) overexpressed in many cancers, including lung and head and neck cancers, and is involved in cancer cell progression and survival. PD-L1, increases in tumor cells to evade and inhibit CD8+ T cells, is a clinical immunotherapeutic target. This study investigated the molecular mechanism of EGF on regulating PD-L1 in EGFR-positive cancers and determined potential agents to reduce PD-L1 expression. RNA sequencing (RNAseq) and bioinformatics analysis were performed to determine potential driver genes that regulate PD-L1 in tumor cells-derived tumorspheres which mimicking cancer stem cells. Then, the specific inhibitors targeting EGFR were applied to reduce the expression of PD-L1 in vitro and in vivo. We validated that EGF could induce PD-L1 expression in the selected EGFR-positive cancers. RNAseq results revealed that STAT1 increased as a driver gene in KOSC-3-derived tumorspheres; these data were analyzed using PANTHER followed by NetworkAnalyst. The blockade of EGFR by afatinib resulted in decreased STAT1 and IRF-1 levels, both are transcriptional factors of PD-L1, and disabled the IFNr-STAT1-mediated PD-L1 axis in vitro and in vivo. Moreover, STAT1 knockdown significantly reduced EGF-mediated PD-L1 expression, and ruxolitinib, a JAK1/JAK2 inhibitor, significantly inhibited STAT1 phosphorylation to reduce the IFNr-mediated PD-L1 axis. These results indicate that EGF exacerbates PD-L1 by increasing the protein levels of STAT1 to enforce the IFNr-JAK1/2-mediated signaling axis in selected EGFR-positive cancers. The inhibition of EGFR by afatinib significantly reduced PD-L1 and may be a potential strategy for enhancing immunotherapeutic efficacy.
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Antígeno B7-H1/metabolismo , Factor de Crecimiento Epidérmico/metabolismo , Receptores ErbB/metabolismo , Regulación Neoplásica de la Expresión Génica , Interferón gamma/metabolismo , Neoplasias/genética , Neoplasias/metabolismo , Factor de Transcripción STAT1/genética , Afatinib/farmacología , Animales , Antígeno B7-H1/antagonistas & inhibidores , Biomarcadores , Línea Celular Tumoral , Receptores ErbB/antagonistas & inhibidores , Humanos , Inmunofenotipificación , Masculino , Ratones , Inhibidores de Proteínas Quinasas/farmacología , Factor de Transcripción STAT1/metabolismoRESUMEN
OBJECTIVES: YM155, an inhibitor of interleukin enhancer-binding factor 3 (ILF3), significantly suppresses cancer stemness property, implying that ILF3 contributes to cell survival of cancer stem cells. However, the molecular function of ILF3 inhibiting cancer stemness remains unclear. This study aimed to uncover the potential function of ILF3 involving in cell survival of epidermal growth factor receptor (EGFR)-positive lung stem-like cancer, and to investigate the potential role to improve the efficacy of anti-EGFR therapeutics. MATERIALS AND METHODS: The association of EGFR and ILF3 in expression and regulations was first investigated in this study. Lung cancer A549 cells with deprivation of ILF3 were created by the gene-knockdown method and then RNAseq was applied to identify the putative genes regulated by ILF3. Meanwhile, HCC827- and A549-derived cancer stem-like cells were used to investigate the role of ILF3 in the formation of cancer stem-like tumorspheres. RESULTS: We found that EGFR induced ILF3 expression, and YM155 reduced EGFR expression. The knockdown of ILF3 reduced not only EGFR expression in mRNA and protein levels, but also cell proliferation in vitro and in vivo, demonstrating that ILF3 may play an important role in contributing to cancer cell survival. Moreover, the knockdown and inhibition of ILF3 by shRNA and YM155, respectively, reduced the formation and survival of HCC827- and A549-derived tumorspheres through inhibiting ErbB3 (HER3) expression, and synergized the therapeutic efficacy of afatinib, a tyrosine kinase inhibitor, against EGFR-positive A549 lung cells. CONCLUSION: This study demonstrated that ILF3 plays an oncogenic like role in maintaining the EGFR-mediated cellular pathway, and can be a therapeutic target to improve the therapeutic efficacy of afatinib. Our results suggested that YM155, an ILF3 inhibitor, has the potential for utilization in cancer therapy against EGFR-positive lung cancers.
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Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Imidazoles/farmacología , Neoplasias Pulmonares/metabolismo , Naftoquinonas/farmacología , Células Madre Neoplásicas/metabolismo , Proteínas del Factor Nuclear 90/metabolismo , Células A549 , Afatinib/administración & dosificación , Animales , Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , Sinergismo Farmacológico , Receptores ErbB/antagonistas & inhibidores , Receptores ErbB/metabolismo , Humanos , Imidazoles/administración & dosificación , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/patología , Masculino , Ratones , Ratones Endogámicos NOD , Ratones SCID , Terapia Molecular Dirigida , Naftoquinonas/administración & dosificación , Células Madre Neoplásicas/efectos de los fármacos , Células Madre Neoplásicas/patología , Proteínas del Factor Nuclear 90/antagonistas & inhibidores , Fosforilación , Inhibidores de Proteínas Quinasas/administración & dosificación , Distribución Aleatoria , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Chronic myeloid leukemia (CML) is a hematological malignancy characterized by the presence of t(9;22) chromosomal translocation that results in BCR-ABL fusion gene. ABL tyrosine kinase inhibitors (TKIs), such as imatinib, nilotinib, and dasatinib, are currently the front-line treatment options for CML. Recently, natural killer (NK) cell activation and expansion have been shown to be associated with optimal treatment responses for CML. To investigate the effects and mechanisms of these TKIs on NK cells, here we characterized activating and inhibitory NK receptors in CD3-CD16+CD56dim NK cells isolated from CML patients in chronic phase (CP). The expressions of activating NK receptors, such as NKG2D, natural cytotoxicity receptor (NCR) and DNAM-1, rebounded after successful TKI treatments for CML. In contrast, among the three surveyed inhibitory receptors (NKG2A, KIR2DL1, and KIR3DL1), only the expression of NKG2A was reverted and suppressed to a very low level by dasatinib, and not by imatinib or nilotinib. CML patients treated with dasatinib indeed expressed fewer NKG2A+ NK cells, which send negative signals for induction of NK cytotoxicity. For these dasatinib-treated patients, the duration to reach major molecular response (MMR) was shorter, and significantly correlated with individual's NKG2A+ NK cell number. This clinical relevance to NKG2A was not observed in treatments with imatinib or nilotinib. In line with dasatinib-specific down-regulation of NKG2A, NK cytotoxicity evaluated by the killing assay was also significantly higher in patients treated with dasatinib than in those treated with imatinib or nilotinib. The lower NK cytotoxicity from imatinib or nilotinib treatments could be reverted by NKG2A blockade using anti-NKG2A antibody. Further in vitro experiments revealed mechanistically that dasatinib could inactivate p38 mitogen-activated protein kinase (MAPK), and consequently affect nuclear import of GATA-3 and GATA-3 transcriptional activities for NKG2A. Our results highlight the dual effects of dasatinib in direct inhibition of ABL kinase and in immunomodulation through NKG2A down-regulation, contributing to accelerated molecular responses (MR) in CML.
Asunto(s)
Citotoxicidad Inmunológica , Dasatinib/uso terapéutico , Regulación Leucémica de la Expresión Génica/efectos de los fármacos , Células Asesinas Naturales/efectos de los fármacos , Células Asesinas Naturales/fisiología , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Leucemia Mielógena Crónica BCR-ABL Positiva/etiología , Subfamília C de Receptores Similares a Lectina de Células NK/genética , Adulto , Anciano , Anciano de 80 o más Años , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Biomarcadores , Línea Celular Tumoral , Dasatinib/farmacología , Femenino , Expresión Génica , Humanos , Inmunofenotipificación , Leucemia Mielógena Crónica BCR-ABL Positiva/diagnóstico , Leucemia Mielógena Crónica BCR-ABL Positiva/metabolismo , Masculino , Persona de Mediana Edad , Subfamília C de Receptores Similares a Lectina de Células NK/metabolismo , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/uso terapéutico , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
Germline variations at JAK2, TERT, HBS1L-MYB and MECOM have been found to associate with myeloproliferative neoplasms (MPNs) in European populations. Whether these germline variations are associated with MPNs in Taiwanese population is obscure. Here we aimed to evaluate the association of five germline variations (JAK2 46/1 haplotype tagged by rs12343867, JAK2 intron 8 rs12339666, TERT rs2736100, HBS1L-MYB rs9376092 and MECOM rs2201862) and the risk of MPNs in Taiwanese population. A total of 178 MPN patients (109 essential thrombocythemia, 54 polycythemia vera and 15 primary myelofibrosis) were enrolled into this study. The information of 17033 control subjects was obtained from Taiwan Biobank database. The JAK2 46/1 haplotype, JAK2 rs12339666 and TERT rs2736100 were significantly associated with Taiwanese MPNs (P = 3.6×10-19, 1.9×10-19 and 3.1×10-6, respectively), and JAK2V617F-positive MPNs (n=121) (P = 5.6×10-21, 4.4×10-21 and 8.6×10-7, respectively). In JAK2V617F-negative cases (n=55), only the JAK2 46/1 haplotype and JAK2 rs12339666 remained statistically significant (P= 0.009 and 0.007, respectively). When stratified by disease subtypes, the JAK2 46/1 haplotype and JAK2 rs12339666 were significantly associated with all three MPN subtypes, but TERT rs2736100 was only associated with essential thrombocythemia and polycythemia vera. We did not find any association of these five SNPs with CALR mutations in our cohort. Furthermore, the risk alleles of MECOM rs2201862 and HBS1L-MYB rs9376092 were demonstrated to be negatively associated with the risk of developing polycythemia vera. In conclusion, germline variations at JAK2 (both the 46/1 haplotype and rs12339666) and TERT rs2736100 were associated with MPNs in Taiwanese population.
RESUMEN
Cancer stem cell survival is the leading factor for tumor recurrence after tumor-suppressive treatments. Therefore, specific and efficient inhibitors of cancer stemness must be discovered for reducing tumor recurrence. YM155 has been indicated to significantly reduce stemness-derived tumorsphere formation. However, the pharmaceutical mechanism of YM155 against cancer stemness is unclear. This study investigated the potential mechanism of YM155 against cancer stemness in lung cancer. Tumorspheres derived from epidermal growth factor receptor (EGFR)-mutant HCC827 and EGFR wild-type A549 cells expressing higher cancer stemness markers (CD133, Oct4, and Nanog) were used as cancer stemness models. We observed that EGFR autophosphorylation (Y1068) was higher in HCC827- and A549-derived tumorspheres than in parental cells; this autophosphorylation induced tumorsphere formation by activating G9a-mediated stemness. Notably, YM155 inhibited tumorsphere formation by blocking the autophosphorylation of EGFR and the EGFR-G9a-mediated stemness pathway. The chemical and genetic inhibition of EGFR and G9a revealed the significant role of the EGFR-G9a pathway in maintaining the cancer stemness property. In conclusion, this study not only revealed that EGFR could trigger tumorsphere formation by elevating G9a-mediated stemness but also demonstrated that YM155 could inhibit this formation by simultaneously blocking EGFR autophosphorylation and G9a activity, thus acting as a potent agent against lung cancer stemness.
Asunto(s)
Antineoplásicos/farmacología , Receptores ErbB/metabolismo , Antígenos de Histocompatibilidad/metabolismo , N-Metiltransferasa de Histona-Lisina/metabolismo , Imidazoles/farmacología , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/metabolismo , Naftoquinonas/farmacología , Afatinib , Línea Celular Tumoral , Evaluación Preclínica de Medicamentos , Receptores ErbB/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/fisiología , N-Metiltransferasa de Histona-Lisina/antagonistas & inhibidores , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Metilación/efectos de los fármacos , Factor 3 de Transcripción de Unión a Octámeros/metabolismo , Fosforilación/efectos de los fármacos , Fosforilación/fisiología , Quinazolinas/farmacología , ARN Mensajero/metabolismoRESUMEN
Mutations in JAK2, MPL and CALR genes have been identified in the majority of myeloproliferative neoplasm (MPN) patients, and patients negative for these three mutations are the so-called triple-negative (TN) MPN. In this study, we examined the mutational profiles of 16 triple-negative MPN patients including 7 essential thrombocythemia (ET), 1 primary myelofibrosis and 8 polycythemia vera (PV). Targeted next-generation sequencing was performed using the ACTOnco Comprehensive Cancer Panel (Ion AmpliSeq Comprehensive Cancer Panel, Life Technologies) to target all coding exons of 409 cancer-related genes. Overall, 30 nonsynonymous somatic mutations were detected in 12 (75%) patients with a range of 1-5 mutations per sample. Notably, one ET patient was found to have JAK2V617F and KITP551L mutations at very low allele frequency. One MPLP70L and 1 MPLM602T mutations were identified each in 1 ET and 1 PV, respectively. Other recurrent mutations were also identified including KMT2C, KMT2D, IRS2, SYNE1, PDE4DIP, SETD2, ATM, TNFAIP3 and CCND2. In addition, germline mutations were also found in some cancer-related genes. Copy number changes were rare in this cohort of TN MPNs. In conclusion, both somatic and germline mutations can be detected in TN MPN patients.
Asunto(s)
ADN de Neoplasias/genética , Mutación , Trastornos Mieloproliferativos/genética , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Calreticulina/genética , Análisis Mutacional de ADN/métodos , ADN de Neoplasias/sangre , Femenino , Mutación de Línea Germinal , Humanos , Janus Quinasa 2/genética , Masculino , Persona de Mediana Edad , Trastornos Mieloproliferativos/sangre , Receptores de Trombopoyetina/genética , Análisis de Secuencia de ADN/métodosRESUMEN
Essential thrombocythemia (ET) is a BCL-ABL1-negative myeloproliferative neoplasm. We have reported that increased activated B cells can facilitate platelet production mediated by cytokines regardless JAK2 mutational status in ET. Recently, calreticulin (CALR) mutations were discovered in ~30% JAK2/MPL-unmutated ET and primary myelofibrosis. Here we sought to screen for CALR mutations and to evaluate B cell immune profiles in a cohort of adult Taiwanese ET patients. B cell populations, granulocytes/monocytes membrane-bound B cell-activating factor (mBAFF) levels, B cells toll-like receptor 4 (TLR4) expression and intracellular levels of interleukin (IL)-1ß/IL-6 and the expression of CD69, CD80, and CD86 were quantified by flow cytometry. Serum BAFF concentration was measured by ELISA. 48 healthy adults were used for comparison. 19 (35.2%) of 54 ET patients harbored 8 types of CALR exon 9 mutations including 4 (7.4%) patients with concomitant JAK2V617F mutations. Compared to JAK2V617F mutation, CALR mutations correlated with younger age at diagnosis (p=0.04), higher platelet count (p=0.004), lower hemoglobin level (p=0.013) and lower leukocyte count (p=0.013). Multivariate analysis adjusted for age, sex, follow-up period and hematological parameters confirmed that increased activated B cells were universally present in JAK2-mutated, CALR-mutated and triple-negative ET patients when compared to healthy adults. JAK2- and CALR-mutated ET have significantly higher fraction of B cells with TLR4 expression when compared to triple-negative ET (p=0.019 and 0.02, respectively). CALR-mutated ET had significantly higher number of CD69-positive activated B cells when compared to triple-negative ET (p=0.035). In conclusion, increased B cell activation is present in ET patients across different mutational subgroups.