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1.
J Cell Mol Med ; 28(9): e18342, 2024 May.
Artículo en Inglés | MEDLINE | ID: mdl-38693852

RESUMEN

Urothelial carcinoma (UC) urgently requires new therapeutic options. Histone deacetylases (HDAC) are frequently dysregulated in UC and constitute interesting targets for the development of alternative therapy options. Thus, we investigated the effect of the second generation HDAC inhibitor (HDACi) quisinostat in five UC cell lines (UCC) and two normal control cell lines in comparison to romidepsin, a well characterized HDACi which was previously shown to induce cell death and cell cycle arrest. In UCC, quisinostat led to cell cycle alterations, cell death induction and DNA damage, but was well tolerated by normal cells. Combinations of quisinostat with cisplatin or the PARP inhibitor talazoparib led to decrease in cell viability and significant synergistic effect in five UCCs and platinum-resistant sublines allowing dose reduction. Further analyses in UM-UC-3 and J82 at low dose ratio revealed that the mechanisms included cell cycle disturbance, apoptosis induction and DNA damage. These combinations appeared to be well tolerated in normal cells. In conclusion, our results suggest new promising combination regimes for treatment of UC, also in the cisplatin-resistant setting.


Asunto(s)
Apoptosis , Inhibidores de Histona Desacetilasas , Inhibidores de Poli(ADP-Ribosa) Polimerasas , Neoplasias de la Vejiga Urinaria , Humanos , Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Apoptosis/efectos de los fármacos , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Cisplatino/farmacología , Daño del ADN/efectos de los fármacos , Sinergismo Farmacológico , Inhibidores de Histona Desacetilasas/farmacología , Inhibidores de Histona Desacetilasas/uso terapéutico , Ácidos Hidroxámicos/farmacología , Ácidos Hidroxámicos/uso terapéutico , Ftalazinas/farmacología , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Neoplasias de la Vejiga Urinaria/tratamiento farmacológico , Neoplasias de la Vejiga Urinaria/patología , Neoplasias de la Vejiga Urinaria/metabolismo , Neoplasias Urológicas/tratamiento farmacológico , Neoplasias Urológicas/patología
2.
Int J Mol Sci ; 24(6)2023 Mar 16.
Artículo en Inglés | MEDLINE | ID: mdl-36982764

RESUMEN

Chimeric antigen receptor (CAR) T-cell therapy has led to profound and durable tumor responses in a relevant subset of patients with relapsed/refractory (r/r) B-cell lymphomas. Still, some patients show insufficient benefit or relapse after CAR T-cell therapy. We performed a retrospective study to investigate the correlation between CAR T-cell persistence in the peripheral blood (PB) at 6 months, assessed by droplet digital PCR (ddPCR), with CAR T-cell treatment outcome. 92 patients with r/r B-cell lymphomas were treated with CD19-targeting CAR T-cell therapies at our institution between 01/2019-08/2022. Six months post-treatment, 15 (16%) patients had no detectable circulating CAR-T constructs by ddPCR. Patients with CAR T-cell persistence had a significantly higher CAR T-cell peak (5432 vs. 620 copies/ug cfDNA, p = 0.0096), as well as higher incidence of immune effector cell-associated neurotoxicity syndrome (37% vs. 7%, p = 0.0182). After a median follow-up of 8.5 months, 31 (34%) patients relapsed. Lymphoma relapses were less frequent among patients with CAR T-cell persistence (29% vs. 60%, p = 0.0336), and CAR T-cell persistence in the PB at 6 months was associated with longer progression-free survival (PFS) (HR 2.79, 95% CI: 1.09-7.11, p = 0.0319). Moreover, we observed a trend towards improved overall survival (OS) (HR 1.99, 95% CI: 0.68-5.82, p = 0.2092) for these patients. In our cohort of 92 B-cell lymphomas, CAR T-cell persistence at 6 months was associated with lower relapse rates and longer PFS. Moreover, our data confirm that 4-1BB-CAR T-cells have a longer persistence as compared to CD-28-based CAR T-cells.


Asunto(s)
Linfoma de Células B , Linfocitos T , Humanos , Estudios Retrospectivos , Recurrencia Local de Neoplasia/etiología , Linfoma de Células B/terapia , Linfoma de Células B/etiología , Inmunoterapia Adoptiva/efectos adversos
3.
Int J Cancer ; 150(5): 856-867, 2022 03 01.
Artículo en Inglés | MEDLINE | ID: mdl-34536301

RESUMEN

Transcriptome-based molecular subtypes of muscle-invasive bladder cancer (MIBC) have been shown to be both prognostic and predictive, but are not used in routine clinical practice. We aimed to develop a feasible, reverse transcription quantitative polymerase chain reaction (RT-qPCR)-based method for molecular subtyping. First, we defined a 68-gene set covering tumor intrinsic (luminal, basal, squamous, neuronal, epithelial-to-mesenchymal, in situ carcinoma) and stromal (immune, extracellular matrix, p53-like) signatures. Then, classifier methods with this 68-gene panel were developed in silico and validated on public data sets with available subtype class information (MD Anderson [MDA], The Cancer Genome Atlas [TCGA], Lund, Consensus). Finally, expression of the selected 68 genes was determined in 104 frozen tissue samples of our MIBC cohort by RT-qPCR using the TaqMan Array Card platform and samples were classified by our newly developed classifiers. The prognostic value of each subtype classification system and molecular signature scores were assessed. We found that the reduced marker set combined with the developed classifiers were able to reproduce the TCGA II, MDA, Lund and Consensus subtype classification systems with an overlap of 79%, 76%, 69% and 64%, respectively. Importantly, we could successfully classify 96% (100/104) of our MIBC samples by using RT-qPCR. Neuronal and luminal subtypes and low stromal gene expressions were associated with poor survival. In conclusion, we developed a robust and feasible method for the molecular subtyping according to the TCGA II, MDA, Lund and Consensus classifications. Our results suggest that stromal signatures have a superior prognostic value compared to tumor intrinsic signatures and therefore underline the importance of tumor-stroma interaction during the progression of MIBC.


Asunto(s)
Genes Relacionados con las Neoplasias , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Neoplasias de la Vejiga Urinaria/genética , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Perfilación de la Expresión Génica , Humanos , Masculino , Persona de Mediana Edad , Pronóstico , Neoplasias de la Vejiga Urinaria/clasificación , Neoplasias de la Vejiga Urinaria/mortalidad , Neoplasias de la Vejiga Urinaria/patología , Adulto Joven
4.
BMC Emerg Med ; 22(1): 109, 2022 06 15.
Artículo en Inglés | MEDLINE | ID: mdl-35705901

RESUMEN

BACKGROUND: Misdiagnosis is a major public health problem, causing increased morbidity and mortality. In the busy setting of an emergency department (ED) patients are diagnosed under difficult circumstances. As a consequence, the ED diagnosis at hospital admittance may often be a descriptive diagnosis, such as "decreased general condition". Our objective was to determine in how far patients with such an unspecific ED diagnosis differ from patients with a specific ED diagnosis and whether they experience a worse outcome. METHODS: We conducted a prospective observational study in Bern university hospital in Switzerland for all adult non-trauma patients admitted to any internal medicine ward from August 15th 2015 to December 7th 2015. Unspecific ED diagnoses were defined through the clinical classification software for ICD-10 by two outcome assessors. As outcome parameters, we assessed in-hospital mortality and length of hospital stay. RESULTS: Six hundred eighty six consecutive patients were included. Unspecific diagnoses were identified in 100 (14.6%) of all consultations. Patients receiving an unspecific diagnosis at ED discharge were significantly more often women (56.0% vs. 43.9%, p = 0.024), presented more often with a non-specific complaint (34% vs. 21%, p = 0.004), were less often demonstrating an abnormal heart rate (5.0% vs. 12.5%, p = 0.03), and less often on antibiotics (32.0% vs. 49.0%, p = 0.002). Apart from these, no studied drug intake, laboratory or clinical data including change in diagnosis was associated significantly with an unspecific diagnosis. Unspecific diagnoses were neither associated with in-hospital mortality in multivariable analysis (OR = 1.74, 95% CI: 0.60-5.04; p = 0.305) adjusted for relevant confounders nor with length of hospital stay (GMR = 0.87, 95% CI: 0.23-3.32; p = 0.840). CONCLUSIONS: Women and patients with non-specific presenting complaints and no abnormal heart rate are at risk of receiving unspecific ED diagnoses that do not allow for targeted treatment, discharge and prognosis. This study did not find an effect of such diagnoses on length of hospital stay nor in-hospital mortality.


Asunto(s)
Servicio de Urgencia en Hospital , Alta del Paciente , Adulto , Femenino , Mortalidad Hospitalaria , Humanos , Tiempo de Internación , Estudios Prospectivos
5.
Molecules ; 26(5)2021 Feb 27.
Artículo en Inglés | MEDLINE | ID: mdl-33673611

RESUMEN

Cisplatin-based treatment is the standard of care therapy for urothelial carcinomas. However, complex cisplatin resistance mechanisms limit the success of this approach. Both apoptosis and autophagy have been shown to contribute to this resistance. Prodigiosin, a secondary metabolite from various bacteria, exerts different biological activities including the modulation of these two cellular stress response pathways. We analyzed the effect of prodigiosin on protein levels of different autophagy- and apoptosis-related proteins in cisplatin-sensitive and -resistant urothelial carcinoma cells (UCCs). Furthermore, we investigated the effect on cell viability of prodigiosin alone or in combination with cisplatin. We made use of four different pairs of cisplatin-sensitive and -resistant UCCs. We found that prodigiosin blocked autophagy in UCCs and re-sensitized cisplatin-resistant cells to apoptotic cell death. Furthermore, we found that prodigiosin is a potent anticancer agent with nanomolar IC50 values in all tested UCCs. In combination studies, we observed that prodigiosin sensitized both cisplatin-sensitive and -resistant urothelial carcinoma cell lines to cisplatin treatment with synergistic effects in most tested cell lines. These effects of prodigiosin are at least partially mediated by altering lysosomal function, since we detected reduced activities of cathepsin B and L. We propose that prodigiosin is a promising candidate for the therapy of cisplatin-resistant urothelial carcinomas, either as a single agent or in combinatory therapeutic approaches.


Asunto(s)
Antineoplásicos/química , Productos Biológicos/química , Prodigiosina/química , Neoplasias de la Vejiga Urinaria/tratamiento farmacológico , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Autofagia/efectos de los fármacos , Productos Biológicos/farmacología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Cisplatino/farmacología , Resistencia a Antineoplásicos , Sinergismo Farmacológico , Quimioterapia Combinada , Humanos , Prodigiosina/farmacología
6.
Mol Carcinog ; 59(7): 862-870, 2020 07.
Artículo en Inglés | MEDLINE | ID: mdl-32386086

RESUMEN

The strength of the interaction between T-cell receptors (TCRs) and their ligands, peptide/major histocompatibility complex complexes (pMHCs), is one of the most frequently discussed and investigated features of T cells in immuno-oncology today. Although there are many molecules on the surface of T cells that interact with ligands on other cells, the TCR/pMHC is the only receptor-ligand pair that offers antigen specificity and dictates the functional response of the T cell. The strength of the TCR/pMHC interaction, along with the environment in which this interaction takes place, is key to how the T cell will respond. The TCR repertoire of T cells that interact with tumor-associated antigens is vast, although typically of low affinity. Here, we focus on the low-affinity interactions between TCRs from CD8+ T cells and different models used in immuno-oncology.


Asunto(s)
Neoplasias/inmunología , Neoplasias/terapia , Receptores de Antígenos de Linfocitos T/inmunología , Animales , Linfocitos T CD8-positivos/inmunología , Antígenos de Histocompatibilidad/inmunología , Humanos , Inmunoterapia/métodos
7.
Int J Mol Sci ; 21(24)2020 Dec 11.
Artículo en Inglés | MEDLINE | ID: mdl-33322422

RESUMEN

Human genomes contain about 100,000 LINE-1 (L1) retroelements, of which more than 100 are intact. L1s are normally tightly controlled by epigenetic mechanisms, which often fail in cancer. In bladder urothelial carcinoma (UC), particularly, L1s become DNA-hypomethylated, expressed and contribute to genomic instability and tumor growth. It is, however, unknown which individual L1s are activated. Following RNA-immunoprecipitation with a L1-specific antibody, third generation nanopore sequencing detected transcripts of 90 individual elements in the VM-Cub-1 UC line with high overall L1 expression. In total, 10 L1s accounted for >60% of the reads. Analysis of five specific L1s by RT-qPCR revealed generally increased expression in UC tissues and cell lines over normal controls, but variable expression among tumor cell lines from bladder, prostate and testicular cancer. Chromatin immunoprecipitation demonstrated active histone marks at L1 sequences with increased expression in VM-Cub-1, but not in a different UC cell line with low L1 expression. We conclude that many L1 elements are epigenetically activated in bladder cancer in a varied pattern. Our findings indicate that expression of individual L1s is highly heterogeneous between and among cancer types.


Asunto(s)
Elementos de Nucleótido Esparcido Largo/genética , Retroelementos/genética , Neoplasias Testiculares/genética , Anciano , Anciano de 80 o más Años , Inmunoprecipitación de Cromatina , Metilación de ADN/genética , Metilación de ADN/fisiología , Femenino , Histonas/metabolismo , Humanos , Inmunoprecipitación , Masculino , Persona de Mediana Edad , Secuenciación de Nanoporos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa
8.
BMC Cancer ; 19(1): 806, 2019 Aug 14.
Artículo en Inglés | MEDLINE | ID: mdl-31412811

RESUMEN

BACKGROUND: Few diagnostic and prognostic biomarkers are available for head-and-neck squamous cell carcinoma (HNSCC). Long non-coding RNAs (lncRNAs) have shown promise as biomarkers in other cancer types and in some cases functionally contribute to tumor development and progression. Here, we searched for lncRNAs useful as biomarkers in HNSCC. METHODS: Public datasets were mined for lncRNA candidates. Two independent HNSCC tissue sets and a bladder cancer tissue set were analyzed by RT-qPCR. Effects of lncRNA overexpression or downregulation on cell proliferation, clonogenicity, migration and chemosensitivity were studied in HNSCC cell lines. RESULTS: Data mining revealed prominently CASC9, a lncRNA significantly overexpressed in HNSCC tumor tissues according to the TCGA RNAseq data. Overexpression was confirmed by RT-qPCR analyses of patient tissues from two independent cohorts. CASC9 expression discriminated tumors from normal tissues with even higher specificity than HOTAIR, a lncRNA previously suggested as an HNSCC biomarker. Specificity of HNSCC detection by CASC9 was further improved by combination with HOTAIR. Analysis of TCGA pan-cancer data revealed significant overexpression of CASC9 across different other entities including bladder, liver, lung and stomach cancers and especially in squamous cell carcinoma (SCC) of the lung. By RT-qPCR analysis we furthermore detected stronger CASC9 overexpression in pure SCC of the urinary bladder and mixed urothelial carcinoma with squamous differentiation than in pure urothelial carcinomas. Thus, CASC9 might represent a general diagnostic biomarker and particularly for SCCs. Unexpectedly, up- or downregulation of CASC9 expression in HNSCC cell lines with low or high CASC9 expression, respectively, did not result in significant changes of cell viability, clonogenicity, migration or chemosensitivity. CONCLUSIONS: CASC9 is a promising biomarker for HNSCC detection. While regularly overexpressed, however, this lncRNA does not seem to act as a major driver of development or progression in this tumor.


Asunto(s)
Biomarcadores de Tumor/genética , Neoplasias de Cabeza y Cuello/diagnóstico , Neoplasias de Cabeza y Cuello/genética , ARN Largo no Codificante/genética , Carcinoma de Células Escamosas de Cabeza y Cuello/diagnóstico , Carcinoma de Células Escamosas de Cabeza y Cuello/genética , Regulación hacia Arriba , Anciano , Carcinoma de Células Escamosas/diagnóstico , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patología , Línea Celular Tumoral , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Neoplasias de Cabeza y Cuello/patología , Humanos , Persona de Mediana Edad , Pronóstico , Sensibilidad y Especificidad , Carcinoma de Células Escamosas de Cabeza y Cuello/patología , Neoplasias de la Vejiga Urinaria/diagnóstico , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/patología
9.
Int J Mol Sci ; 20(9)2019 Apr 30.
Artículo en Inglés | MEDLINE | ID: mdl-31052182

RESUMEN

Class I histone deacetylases (HDACs) generally promote cell proliferation and tumorigenesis, whereas class IIA HDACs like HDAC4 and HDAC5 may promote or impede cancer development in a tissue-dependent manner. In urothelial carcinoma (UC), HDAC5 is often downregulated. Accordingly, HDAC5 was weakly expressed in UC cell lines suggesting a possible tumor-suppressive function. We therefore characterized the effects of stable HDAC5 expression in four UC cell lines (RT112, VM-Cub-1, SW1710 and UM-UC-3) with different phenotypes reflecting the heterogeneity of UC, by assessing proliferation, clonogenicity and migration ability. Further, we detailed changes in the proteome and transcriptome by immunoblotting, mass spectrometry and RNA sequencing analysis. We observed that HDAC5 overexpression in general decreased cell proliferation, but in one cell line (VM-Cub-1) induced a dramatic change from an epitheloid to a mesenchymal phenotype, i.e., epithelial-mesenchymal transition (EMT). These phenotypical changes were confirmed by comprehensive proteomics and transcriptomics analyses. In contrast to HDAC5, overexpression of HDAC4 exerted only weak effects on cell proliferation and phenotypes. We conclude that overexpression of HDAC5 may generally decrease proliferation in UC, but, intriguingly, may induce EMT on its own in certain circumstances.


Asunto(s)
Carcinoma/metabolismo , Proliferación Celular , Transición Epitelial-Mesenquimal , Histona Desacetilasas/genética , Neoplasias de la Vejiga Urinaria/metabolismo , Urotelio/patología , Carcinoma/genética , Línea Celular Tumoral , Células HEK293 , Histona Desacetilasas/metabolismo , Humanos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Neoplasias de la Vejiga Urinaria/genética , Urotelio/metabolismo
10.
BJU Int ; 122(4): 695-704, 2018 10.
Artículo en Inglés | MEDLINE | ID: mdl-29802777

RESUMEN

OBJECTIVE: To assess the predictive value of pre-chemotherapy matrix metalloproteinase 7 (MMP-7), soluble Fas (sFas) and Fas ligand (FasL) serum levels, as well as their changes during therapy. PATIENTS AND METHODS: Serum levels of MMP-7, Fas and FasL were determined by ELISA in 96 patients with castration-resistant prostate cancer (CRPC): 21 docetaxel-resistant patients who received one single series and 75 docetaxel-sensitive patients who received repeated series of docetaxel. In addition to the 96 pretreatment serum samples, 987 sera collected during chemotherapy were also analysed. RESULTS: Higher pretreatment serum MMP-7, sFas and prostate-specific antigen (PSA) levels were significantly associated with both docetaxel resistance (P = 0.007, P = 0.001, P < 0.001, respectively) and shorter cancer-specific survival (P < 0.001, P = 0.041, P < 0.001, respectively). High MMP-7 level remained an independent predictor of both docetaxel resistance (hazard ratio [HR] 2.298, 95% confidence interval [CI]: 1.354-3.899; P = 0.002) and poor cancer-specific survival (HR 2.11, 95% CI: 1.36-3.30; P = 0.001) in multivariable analyses. Greater increase in MMP-7 levels in the second treatment holiday and greater increase in PSA levels in the first and second treatment holidays were predictive of survival. CONCLUSIONS: Pretreatment serum MMP-7 levels may help to select patients with CRPC who are likely to benefit from docetaxel chemotherapy. Furthermore, MMP-7 levels alone or in combination with PSA levels could be used for therapy monitoring. Correlative studies embedded in clinical trials are necessary to validate these biomarkers for clinical decision-making.


Asunto(s)
Antineoplásicos/uso terapéutico , Docetaxel/uso terapéutico , Resistencia a Antineoplásicos/efectos de los fármacos , Proteína Ligando Fas/sangre , Metaloproteinasa 7 de la Matriz/sangre , Neoplasias de la Próstata Resistentes a la Castración/sangre , Adulto , Anciano , Anciano de 80 o más Años , Protocolos de Quimioterapia Combinada Antineoplásica , Estudios de Cohortes , Humanos , Masculino , Persona de Mediana Edad , Valor Predictivo de las Pruebas , Antígeno Prostático Específico/sangre , Neoplasias de la Próstata Resistentes a la Castración/tratamiento farmacológico , Neoplasias de la Próstata Resistentes a la Castración/mortalidad , Neoplasias de la Próstata Resistentes a la Castración/patología , Análisis de Supervivencia , Resultado del Tratamiento
11.
Int J Mol Sci ; 19(2)2018 Feb 16.
Artículo en Inglés | MEDLINE | ID: mdl-29462944

RESUMEN

Therapeutic efficacy of cisplatin-based treatment of late stage urothelial carcinoma (UC) is limited by chemoresistance. To elucidate underlying mechanisms and to develop new approaches for overcoming resistance, we generated long-term cisplatin treated (LTT) UC cell lines, characterised their cisplatin response, and determined the expression of molecules involved in cisplatin transport and detoxification, DNA repair, and apoptosis. Inhibitors of metallothioneins and Survivin were applied to investigate their ability to sensitise towards cisplatin. Cell growth, proliferation, and clonogenicity were examined after cisplatin treatment by MTT 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, EdU (5-ethynyl-2'-deoxyuridine) incorporation assay, and Giemsa staining, respectively. Cell cycle distribution and apoptosis were quantified by flow cytometry. mRNA and protein expressions were measured by real-time quantitative (qRT)-PCR, western blot, or immunofluorescence staining. LTTs recovered rapidly from cisplatin stress compared to parental cells. In LTTs, to various extents, cisplatin exporters and metallothioneins were induced, cisplatin adduct levels and DNA damage were decreased, whereas expression of DNA repair factors and specific anti-apoptotic factors was elevated. Pharmacological inhibition of Survivin, but not of metallothioneins, sensitised LTTs to cisplatin, in an additive manner. LTTs minimise cisplatin-induced DNA damage and evade apoptosis by increased expression of anti-apoptotic factors. The observed diversity among the four LTTs highlights the complexity of cisplatin resistance mechanisms even within one tumour entity, explaining heterogeneity in patient responses to chemotherapy.


Asunto(s)
Antineoplásicos/farmacología , Carcinoma/metabolismo , Cisplatino/farmacología , Resistencia a Antineoplásicos , Neoplasias de la Vejiga Urinaria/metabolismo , Urotelio/efectos de los fármacos , Apoptosis , Ciclo Celular , Línea Celular Tumoral , Daño del ADN , Humanos , Metalotioneína/metabolismo , Urotelio/metabolismo
12.
J Biol Chem ; 291(16): 8399-413, 2016 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-26884329

RESUMEN

Hepatic stellate cells (HSCs) were recently identified as liver-resident mesenchymal stem cells. HSCs are activated after liver injury and involved in pivotal processes, such as liver development, immunoregulation, regeneration, and also fibrogenesis. To date, several studies have reported candidate pathways that regulate the plasticity of HSCs during physiological and pathophysiological processes. Here we analyzed the expression changes and activity of the RAS family GTPases and thereby investigated the signaling networks of quiescent HSCs versus activated HSCs. For the first time, we report that embryonic stem cell-expressed RAS (ERAS) is specifically expressed in quiescent HSCs and down-regulated during HSC activation via promoter DNA methylation. Notably, in quiescent HSCs, the high level of ERAS protein correlates with the activation of AKT, STAT3, mTORC2, and HIPPO signaling pathways and inactivation of FOXO1 and YAP. Our data strongly indicate that in quiescent HSCs, ERAS targets AKT via two distinct pathways driven by PI3Kα/δ and mTORC2, whereas in activated HSCs, RAS signaling shifts to RAF-MEK-ERK. Thus, in contrast to the reported role of ERAS in tumor cells associated with cell proliferation, our findings indicate that ERAS is important to maintain quiescence in HSCs.


Asunto(s)
Metilación de ADN/fisiología , Células Estrelladas Hepáticas/enzimología , Sistema de Señalización de MAP Quinasas/fisiología , Proteína Oncogénica p21(ras)/biosíntesis , Regiones Promotoras Genéticas/fisiología , Animales , Proteínas Reguladoras de la Apoptosis/genética , Proteínas Reguladoras de la Apoptosis/metabolismo , Factores de Transcripción Forkhead/genética , Factores de Transcripción Forkhead/metabolismo , Células Estrelladas Hepáticas/citología , Masculino , Diana Mecanicista del Complejo 2 de la Rapamicina , Complejos Multiproteicos/genética , Complejos Multiproteicos/metabolismo , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Proteína Oncogénica p21(ras)/genética , Proteínas Proto-Oncogénicas c-akt/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , Ratas , Ratas Wistar , Serina-Treonina Quinasas TOR/genética , Serina-Treonina Quinasas TOR/metabolismo , Proteínas Señalizadoras YAP
13.
Int J Mol Sci ; 18(7)2017 Jul 05.
Artículo en Inglés | MEDLINE | ID: mdl-28678170

RESUMEN

Disturbances in histone acetyltransferases (HATs) are common in cancers. In urothelial carcinoma (UC), p300 and CBP are often mutated, whereas the GNAT family HATs GCN5 and PCAF (General Control Nonderepressible 5, p300/CBP-Associated Factor) are often upregulated. Here, we explored the effects of specific siRNA-mediated knockdown of GCN5, PCAF or both in four UC cell lines (UCCs). Expression of various HATs and marker proteins was measured by qRT-PCR and western blot. Cellular effects of knockdowns were analyzed by flow cytometry and ATP-, caspase-, and colony forming-assays. GCN5 was regularly upregulated in UCCs, whereas PCAF was variable. Knockdown of GCN5 or both GNATs, but not of PCAF alone, diminished viability and inhibited clonogenic growth in 2/4 UCCs, inducing cell cycle changes and caspase-3/7 activity. PCAF knockdown elicited GCN5 mRNA upregulation. Double knockdown increased c-MYC and MDM2 (Mouse Double Minute 2) in most cell lines. In conclusion, GCN5 upregulation is especially common in UCCs. GCN5 knockdown impeded growth of specific UCCs, whereas PCAF knockdown elicited minor effects. The limited sensitivity towards GNAT knockdown and its variation between the cell lines might be due to compensatory effects including HAT, c-MYC and MDM2 upregulation. Our results predict that developing drugs targeting individual HATs for UC treatment may be challenging.


Asunto(s)
Carcinoma/genética , Neoplasias Uretrales/genética , Neoplasias Uretrales/patología , Factores de Transcripción p300-CBP/genética , Apoptosis/genética , Biomarcadores de Tumor , Carcinoma/metabolismo , Carcinoma/patología , Línea Celular Tumoral , Supervivencia Celular/genética , Senescencia Celular/genética , Expresión Génica , Técnicas de Silenciamiento del Gen , Humanos , Unión Proteica , Interferencia de ARN , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN Interferente Pequeño/genética , Neoplasias Uretrales/metabolismo , Factores de Transcripción p300-CBP/metabolismo
14.
Int J Mol Sci ; 18(8)2017 Aug 02.
Artículo en Inglés | MEDLINE | ID: mdl-28767070

RESUMEN

Therapeutic efficacy of cisplatin-based chemotherapy for advanced-stage urothelial carcinoma (UC) is limited by drug resistance. The nuclear factor (erythroid-derived 2)-like 2 (NRF2) pathway is a major regulator of cytoprotective responses. We investigated its involvement in cisplatin resistance in long-term cisplatin treated UC cell lines (LTTs). Expression of NRF2 pathway components and targets was evaluated by qRT-PCR and western blotting in LTT sublines from four different parental cells. NRF2 transcriptional activity was determined by reporter assays and total glutathione (GSH) was quantified enzymatically. Effects of siRNA-mediated NRF2 knockdown on chemosensitivity were analysed by viability assays, γH2AX immunofluorescence, and flow cytometry. Increased expression of NRF2, its positive regulator p62/SQSTM1, and elevated NRF2 activity was observed in 3/4 LTTs, which correlated with KEAP1 expression. Expression of cytoprotective enzymes and GSH concentration were upregulated in some LTTs. NRF2 knockdown resulted in downregulation of cytoprotective enzymes and resensitised 3/4 LTTs towards cisplatin as demonstrated by reduced IC50 values, increased γH2AX foci formation, and elevated number of apoptotic cells. In conclusion, while LTT lines displayed diversity in NRF2 activation, NRF2 signalling contributed to cisplatin resistance in LTT lines, albeit in diverse ways. Accordingly, inhibition of NRF2 can be used to resensitise UC cells to cisplatin, but responses in patients may likewise be variable.


Asunto(s)
Cisplatino/farmacología , Citoprotección/efectos de los fármacos , Resistencia a Antineoplásicos/efectos de los fármacos , Factor 2 Relacionado con NF-E2/metabolismo , Proteínas de Neoplasias/metabolismo , Neoplasias de la Vejiga Urinaria/metabolismo , Urotelio/metabolismo , Línea Celular Tumoral , Humanos , Neoplasias de la Vejiga Urinaria/patología , Urotelio/patología
15.
Tumour Biol ; 37(2): 1763-9, 2016 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-26314857

RESUMEN

According to GLOBOCAN 2012, the worldwide burden of cancer increased and is expected to worsen within the next decades. Therefore, universal combat against cancer will not succeed with treatment solely; effective prevention and early detection are urgently needed to tackle the cancer crisis. Emerging data demonstrate that long non-coding RNAs are involved in numerous biological and pathological processes like development and differentiation and in a variety of human diseases including cancer. Located at 18q21, LINC-ROR (regulator of reprogramming) is a modulator of ESCs maintenance and hypoxia-signaling pathways in hepatocellular cancer cells. The aim of this study was to examine the expression of LINC-ROR in various cell lines and representative samples of human cancers by quantitative real-time RT-PCR to provide a snapshot on how LINC-ROR expression may be deregulated in cancer. More than 30 cell lines and 112 patient specimens from various tissues were assessed for relative expression of LINC-ROR. Our results revealed that the expression of LINC-ROR was lower in all somatic cancer cell lines compared to stem cells or cells with stem cell-like capabilities, like the embryonic carcinoma cell line, NTERA-2. In tissues, expression patterns vary, but some cancerous tissues displayed increased LINC-ROR expression compared to corresponding normal tissues. Thus, we hypothesize that LINC-ROR may have a key function in a subpopulation of cells from the tumor bulk, i.e., the cancer stem cells associated with specific properties including resistance to adverse environmental conditions.


Asunto(s)
Neoplasias/genética , ARN Largo no Codificante/genética , Diferenciación Celular/genética , Línea Celular , Línea Celular Tumoral , Reprogramación Celular/genética , Células HEK293 , Células Hep G2 , Células Endoteliales de la Vena Umbilical Humana , Humanos , Hipoxia/genética , Células MCF-7 , Células Madre Neoplásicas/metabolismo , Transducción de Señal/genética
16.
Mol Cancer ; 14: 108, 2015 May 21.
Artículo en Inglés | MEDLINE | ID: mdl-25994132

RESUMEN

BACKGROUND: Urothelial carcinoma (UC) is the fifth most common cancer in the developed world. Delineation of differentiation subtypes in UC highlighted the importance of aberrant differentiation. Understanding underlying mechanisms may facilitate diagnosis and development of efficient therapy strategies. It is well accepted that epigenetic mechanisms are involved. Long noncoding RNAs (lncRNAs), a new class of epigenetic factors, are thought to mediate molecular differences between cell types to control cellular identity. The present study focuses on the lncRNA HOTAIR, originating from the HOXC locus. Its overexpression induces an aggressive phenotype in many cancers and aberrant expression of homeotic HOX transcription factors, especially HOXD10, that regulate differentiation and tissue homeostasis. The aim of the present study was to determine the functional role of HOTAIR in UC with regard to aggressive phenotype, regulation of aberrant differentiation and altered HOX gene expression. METHODS: We determined RNA expression levels of HOTAIR and HOX genes in UC tissues and cell lines. Knockdown of HOTAIR and ectopic overexpression was performed to determine the effect on reported target genes in UC. Cell lines were stably transfected with HOTAIR to investigate changes in phenotype and HOX gene expression. RESULTS: HOTAIR was overexpressed in approximately half of UC tissues and cell lines. Effects of HOTAIR overexpression differed between cell lines. Whereas VM-CUB1 cells acquired the expected phenotype with increased proliferation, clonogenicity, anchorage independent growth, migratory activity and epithelial-to-mesenchymal transition, 5637 cells grew more slowly displaying induction of senescence and related immune response genes. Other UC lines showed intermediate effects. Expression profiling revealed divergent effects on HOX genes, cell cycle regulators and differentiation according with the phenotypic differences between HOTAIR-overexpressing VM-CUB1 and 5637 cells. CONCLUSIONS: Our data indicate that HOTAIR overexpression may affect differentiation state and aggressiveness of UC cells, but in a cell-type dependent manner. Our functional studies and the comparison of our expression data sets with those from other cancer cell types, which revealed minimal overlaps, indicate that effects of HOTAIR are strongly tissue-dependent and can even differ within one cancer type. Thus, HOTAIR functions and target genes cannot simply be transferred from one cancer type to the other.


Asunto(s)
Regulación Neoplásica de la Expresión Génica , Proteínas de Homeodominio/genética , ARN Largo no Codificante/metabolismo , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/patología , Urotelio/patología , Anciano , Anciano de 80 o más Años , Línea Celular Tumoral , Proliferación Celular , Células Clonales , Femenino , Perfilación de la Expresión Génica , Proteínas de Homeodominio/metabolismo , Humanos , Estimación de Kaplan-Meier , Masculino , Persona de Mediana Edad , Análisis Multivariante , Invasividad Neoplásica , Análisis de Secuencia por Matrices de Oligonucleótidos , Especificidad de Órganos , Fenotipo , ARN Largo no Codificante/genética , Urotelio/metabolismo
17.
BMC Cancer ; 14: 628, 2014 Aug 29.
Artículo en Inglés | MEDLINE | ID: mdl-25167871

RESUMEN

BACKGROUND: Notch signalling regulates cell fate in most tissues, promoting precursor cell proliferation in some, but differentiation in others. Accordingly, downregulation or overactivity variously contributes to cancer development. So far, little is known about Notch pathway activity and function in the normal urothelium and in urothelial carcinoma (UC). We have therefore investigated expression of Notch pathway components in UC tissues and cell lines and studied the function of one receptor, NOTCH1, in detail. METHODS: Expression of canonical Notch pathway components were studied in UC and normal bladder tissues by immunohistochemistry and quantitative RT-PCR and in UC cell lines and normal cultured urothelial cells by qRT-PCR, immunocytochemistry and Western blotting. Pathway activity was measured by reporter gene assays. Its influence on cell proliferation was investigated by γ-secretase inhibition. Effects of NOTCH1 restoration were followed by measuring cell cycle distribution, proliferation, clonogenicity and nuclear morphology. RESULTS: NOTCH1 and its ligand, DLL1, were expressed at plasma membranes and in the cytoplasm of cells in the upper normal urothelium layer, but became downregulated in UC tissues, especially in high-stage tumours. In addition, the proteins were often delocalized intracellularly. According differences were observed in UC cell lines compared to normal urothelial cells. Canonical Notch pathway activity in reporter assays was repressed in UC cell lines compared to normal cells and a mammary carcinoma cell line, but was induced by transfected NOTCH1. Inhibitors of Notch signalling acting at the γ-secretase step did not affect UC cell proliferation at concentrations efficacious against a cell line with known Notch activity. Surprisingly, overexpression of NOTCH1 into UC cell lines did not significantly affect short-term cell proliferation, but induced nuclear abnormalities and diminished clonogenicity. CONCLUSION: Our data indicate that canonical Notch signalling is suppressed in urothelial carcinoma mainly through downregulation of NOTCH1. These findings can be explained by proposing that canonical Notch signalling may promote differentiation in the urothelium, like in many squamous epithelia, and its suppression may therefore be advantageous for tumour progression. As an important corollary, inhibition of canonical Notch signalling is unlikely to be efficacious and might be counter-productive in the treatment of urothelial carcinoma.


Asunto(s)
Carcinoma/metabolismo , Receptores Notch/metabolismo , Transducción de Señal , Neoplasias de la Vejiga Urinaria/metabolismo , Anciano , Anciano de 80 o más Años , Proteínas de Unión al Calcio/genética , Proteínas de Unión al Calcio/metabolismo , Carcinoma/genética , Carcinoma/patología , Línea Celular Tumoral , Proliferación Celular , Femenino , Expresión Génica , Humanos , Péptidos y Proteínas de Señalización Intercelular/genética , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Ligandos , Masculino , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Persona de Mediana Edad , Transporte de Proteínas , Receptor Notch1/genética , Receptor Notch1/metabolismo , Receptores Notch/genética , Proteínas Serrate-Jagged , Ensayo de Tumor de Célula Madre , Vejiga Urinaria/metabolismo , Neoplasias de la Vejiga Urinaria/genética , Neoplasias de la Vejiga Urinaria/patología
18.
Curr Oncol ; 31(4): 2067-2075, 2024 04 05.
Artículo en Inglés | MEDLINE | ID: mdl-38668056

RESUMEN

Donor cell leukemia (DCL) is a rare complication after allogeneic hematopoietic stem cell transplantation (HSCT) accounting for 0.1% of relapses and presenting as secondary leukemia of donor origin. Distinct in phenotype and cytogenetics from the original leukemia, DCL's clinical challenge lies in its late onset. Its origin is affected by donor cell anomalies, transplant environment, and additional mutations. A 43-year-old woman, treated for early stage triple-negative breast cancer, developed mixed-phenotype acute leukemia (MPAL), 12 years later. Following induction chemotherapy, myeloablative conditioning, and allo-HSCT from her fully HLA-matched brother, she exhibited multiple cutaneous relapses of the original leukemia, subsequently evolving into DCL of the bone marrow. Cytogenetic analysis revealed a complex male karyotype in 20 out of 21 metaphases, however, still showing the MPAL phenotype. DCL diagnosis was confirmed by 90.5% XY in FISH analysis and the male karyotype. Declining further intensive chemotherapy including a second allo-HSCT, she was subsequently treated with repeated radiotherapy, palliative systemic therapies, and finally venetoclax and navitoclax but died seven months post-DCL diagnosis. This case underlines DCL's complexity, characterized by unique genetics, further complicating diagnosis. It highlights the need for advanced diagnostic techniques for DCL identification and underscores the urgency for early detection and better prevention and treatment strategies.


Asunto(s)
Trasplante de Células Madre Hematopoyéticas , Trasplante Homólogo , Trasplante de Células Madre Hematopoyéticas/efectos adversos , Humanos , Adulto , Femenino , Leucemia/terapia , Donantes de Tejidos , Masculino
19.
Front Immunol ; 15: 1268652, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-38558822

RESUMEN

Introduction: A multitude of findings from cell cultures and animal studies are available to support the anti-cancer properties of cannabidiol (CBD). Since CBD acts on multiple molecular targets, its clinical adaptation, especially in combination with cancer immunotherapy regimen remains a serious concern. Methods: Considering this, we extensively studied the effect of CBD on the cytokine-induced killer (CIK) cell immunotherapy approach using multiple non-small cell lung cancer (NSCLC) cells harboring diverse genotypes. Results: Our analysis showed that, a) The Transient Receptor Potential Cation Channel Subfamily V Member 2 (TRPV2) channel was intracellularly expressed both in NSCLC cells and CIK cells. b) A synergistic effect of CIK combined with CBD, resulted in a significant increase in tumor lysis and Interferon gamma (IFN-g) production. c) CBD had a preference to elevate the CD25+CD69+ population and the CD62L_CD45RA+terminal effector memory (EMRA) population in NKT-CIK cells, suggesting early-stage activation and effector memory differentiation in CD3+CD56+ CIK cells. Of interest, we observed that CBD enhanced the calcium influx, which was mediated by the TRPV2 channel and elevated phosphor-Extracellular signal-Regulated Kinase (p-ERK) expression directly in CIK cells, whereas ERK selective inhibitor FR180204 inhibited the increasing cytotoxic CIK ability induced by CBD. Further examinations revealed that CBD induced DNA double-strand breaks via upregulation of histone H2AX phosphorylation in NSCLC cells and the migration and invasion ability of NSCLC cells suppressed by CBD were rescued using the TRPV2 antagonist (Tranilast) in the absence of CIK cells. We further investigated the epigenetic effects of this synergy and found that adding CBD to CIK cells decreased the Long Interspersed Nuclear Element-1 (LINE-1) mRNA expression and the global DNA methylation level in NSCLC cells carrying KRAS mutation. We further investigated the epigenetic effects of this synergy and found that adding CBD to CIK cells decreased the Long Interspersed Nuclear Element-1 (LINE-1) mRNA expression and the global DNA methylation level in NSCLC cells carrying KRAS mutation. Conclusions: Taken together, CBD holds a great potential for treating NSCLC with CIK cell immunotherapy. In addition, we utilized NSCLC with different driver mutations to investigate the efficacy of CBD. Our findings might provide evidence for CBD-personized treatment with NSCLC patients.


Asunto(s)
Cannabidiol , Carcinoma de Pulmón de Células no Pequeñas , Células Asesinas Inducidas por Citocinas , Neoplasias Pulmonares , Animales , Humanos , Carcinoma de Pulmón de Células no Pequeñas/terapia , Cannabidiol/farmacología , Neoplasias Pulmonares/terapia , Proteínas Proto-Oncogénicas p21(ras) , ARN Mensajero
20.
Cancers (Basel) ; 16(10)2024 May 15.
Artículo en Inglés | MEDLINE | ID: mdl-38791965

RESUMEN

(1) Background: Treosulfan and melphalan (TreoMel)-based high-dose chemotherapy (HDCT) has shown promising safety and efficacy as a conditioning regimen for acute myeloid leukemia (AML) patients undergoing autologous stem cell transplantation (ASCT). However, despite intensive first-line induction treatment and upfront consolidation with HDCT and ASCT, AML relapse rates are still high, and further efforts are needed to improve patient outcomes. The aim of this study was to compare two melphalan dose schedules in regard to the safety of TreoMel HDCT and patient outcomes. (2) Methods: We retrospectively analyzed the safety and efficacy of two melphalan dose schedules combined with standard-dose treosulfan in AML patients undergoing HDCT and ASCT at the University Hospital of Bern, Switzerland, between August 2019 and August 2023. Patients received treosulfan 42 g/m2 combined with either melphalan 140 mg/m2 (TreoMel 140) or melphalan 200 mg/m2 (TreoMel 200). Co-primary endpoints were progression-free survival (PFS), overall survival (OS), as well as safety profile. (3) Results: We included a total of 51 AML patients: 31 (60.8%) received TreoMel 140 and 20 (39.2%) TreoMel 200. The patients' basal characteristics were comparable between both cohorts. No significant differences in the duration of hospitalization or the adverse event profile were identified. There were no statistically significant differences in relapse (0.45 vs. 0.30, p = 0.381) and mortality rates (0.42 vs. 0.15, p = 0.064) between the melphalan 140 mg/m2 and 200 mg/m2 cohorts, nor for PFS (HR: 0.81, 95% CI: 0.29-2.28, p = 0.70) or OS (HR: 0.70, 95% CI: 0.19-2.57, p = 0.59) for the TreoMel 140 vs. TreoMel 200 cohort. (4) Conclusions: A higher dose of melphalan (TreoMel 200) was well tolerated overall. No statistically significant differences for patient outcomes could be observed, possibly due to the relatively small patient cohort and the short follow-up. A longer follow-up and prospective randomized studies would be required to confirm the safety profile and clinical benefit.

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