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1.
Crit Care ; 28(1): 72, 2024 03 12.
Artículo en Inglés | MEDLINE | ID: mdl-38475798

RESUMEN

BACKGROUND: New onset refractory status epilepticus (NORSE) is a neurologic emergency without an immediately identifiable cause. The complicated and long ICU stay of the patients can lead to perceiving a prolongation of therapies as futile. However, a recovery is possible even in severe cases. This retrospective study investigates ICU treatments, short- and long-term outcome and ethical decisions of a case series of patients with NORSE. METHODS: Overall, 283 adults were admitted with status epilepticus (SE) to the Neurocritical Care Unit of the University Hospital Zurich, Switzerland, between 01.2010 and 12.2022. Of them, 25 had a NORSE. We collected demographic, clinical, therapeutic and outcome data. Descriptive statistics was performed. RESULTS: Most patients were female (68%), previously healthy (Charlson comorbidity index 1 [0-4]) and relatively young (54 ± 17 years). 96% presented with super-refractory SE. Despite extensive workup, the majority (68%) of cases remained cryptogenic. Most patients had a long and complicated ICU stay. The in-hospital mortality was 36% (n = 9). The mortality at last available follow-up was 56% (n = 14) on average 30 months after ICU admission. The cause of in-hospital death for 89% (n = 8) of the patients was the withholding/withdrawing of therapies. Medical staff except for one patient triggered the decision. The end of life (EOL) decision was taken 29 [12-51] days after the ICU admission. Death occurred on day 6 [1-8.5] after the decision was taken. The functional outcome improved over time for 13/16 (81%) hospital survivors (median mRS at hospital discharge 4 [3.75-5] vs. median mRS at last available follow-up 2 [1.75-3], p < 0.001). CONCLUSIONS: Our data suggest that the long-term outcome can still be favorable in NORSE survivors, despite a prolonged and complicated ICU stay. Clinicians should be careful in taking EOL decisions to avoid the risk of a self-fulfilling prophecy. Our results encourage clinicians to continue treatment even in initially refractory cases.


Asunto(s)
Estado Epiléptico , Humanos , Femenino , Masculino , Estudios Retrospectivos , Mortalidad Hospitalaria , Estado Epiléptico/tratamiento farmacológico , Hospitalización , Enfermedad Aguda
2.
EMBO Rep ; 21(9): e50446, 2020 09 03.
Artículo en Inglés | MEDLINE | ID: mdl-32749065

RESUMEN

Melanoma progression is generally associated with increased transcriptional activity mediated by the Yes-associated protein (YAP). Mechanical signals from the extracellular matrix are sensed by YAP, which then activates the expression of proliferative genes, promoting melanoma progression and drug resistance. Which extracellular signals induce mechanotransduction, and how this is mediated, is not completely understood. Here, using secretome analyses, we reveal the extracellular accumulation of amyloidogenic proteins, i.e. premelanosome protein (PMEL), in metastatic melanoma, together with proteins that assist amyloid maturation into fibrils. We also confirm the accumulation of amyloid-like aggregates, similar to those detected in Alzheimer disease, in metastatic cell lines, as well as in human melanoma biopsies. Mechanistically, beta-secretase 2 (BACE2) regulates the maturation of these aggregates, which in turn induce YAP activity. We also demonstrate that recombinant PMEL fibrils are sufficient to induce mechanotransduction, triggering YAP signaling. Finally, we demonstrate that BACE inhibition affects cell proliferation and increases drug sensitivity, highlighting the importance of amyloids for melanoma survival, and the use of beta-secretase inhibitors as potential therapeutic approach for metastatic melanoma.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales , Melanoma , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Amiloidogénicas , Humanos , Mecanotransducción Celular , Melanoma/tratamiento farmacológico , Melanoma/genética , Factores de Transcripción/genética , Factores de Transcripción/metabolismo
3.
Chemistry ; 25(33): 7948-7952, 2019 Jun 12.
Artículo en Inglés | MEDLINE | ID: mdl-30985041

RESUMEN

Curcumin has chemopreventative properties against a variety of tumours, but has poor bioavailability. Here, two new bis-cyclometallated iridium(III) complexes have been prepared, featuring the natural product curcumin (CUR) or its reduced form, tetrahydrocurcumin (THC), as bidentate, anionic O O-binding ligands. The iridium THC complex is highly luminescent in deoxygenated solution and efficiently generates singlet oxygen under aerated conditions, whereas in the CUR analogue, other non-radiative decay pathways are competitive. The complexes are rapidly taken up by a variety of human tumour cell lines from solutions of micromolar concentration. They show negligible cytotoxicity in the absence of irradiation. When briefly irradiated with visible light, Ir-THC becomes highly phototoxic, inducing rapid apoptosis within 2 h. The results show the high potential of such complexes as sensitizers in photodynamic therapy (PDT).

4.
Biochem Soc Trans ; 43(2): 292-6, 2015 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-25849932

RESUMEN

Microtubules (MTs) are dynamic polymers consisting of α/ß tubulin dimers and playing a plethora of roles in eukaryotic cells. Looking at neurons, they are key determinants of neuronal polarity, axonal transport and synaptic plasticity. The concept that MT dysfunction can participate in, and perhaps lead to, Parkinson's disease (PD) progression has been suggested by studies using toxin-based and genetic experimental models of the disease. Here, we first learn lessons from MPTP and rotenone as well as from the PD related genes, including SNCA and LRRK2, and then look at old and new evidence regarding the interplay between parkin and MTs. Data from experimental models and human cells point out that parkin regulates MT stability and strengthen the link between MTs and PD paving the way to a viable strategy for the management of the disease.


Asunto(s)
Microtúbulos/genética , Neuronas/efectos de los fármacos , Enfermedad de Parkinson Secundaria/genética , Ubiquitina-Proteína Ligasas/genética , 1-Metil-4-fenil-1,2,3,6-Tetrahidropiridina/administración & dosificación , Animales , Modelos Animales de Enfermedad , Humanos , Proteína 2 Quinasa Serina-Treonina Rica en Repeticiones de Leucina , Ratones , Microtúbulos/efectos de los fármacos , Microtúbulos/patología , Mutación , Neuronas/metabolismo , Neuronas/patología , Enfermedad de Parkinson Secundaria/inducido químicamente , Enfermedad de Parkinson Secundaria/tratamiento farmacológico , Enfermedad de Parkinson Secundaria/patología , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Rotenona/administración & dosificación , Tubulina (Proteína)/genética , Ubiquitina-Proteína Ligasas/metabolismo , alfa-Sinucleína/genética , alfa-Sinucleína/metabolismo
5.
Cell Death Dis ; 15(1): 28, 2024 01 11.
Artículo en Inglés | MEDLINE | ID: mdl-38199984

RESUMEN

The tumor microenvironment is a complex ecosystem that plays a critical role in cancer progression and treatment response. Recently, extracellular amyloid fibrils have emerged as novel components of the tumor microenvironment; however, their function remains elusive. In this study, we establish a direct connection between the presence of amyloid fibrils in the secretome and the activation of YAP, a transcriptional co-activator involved in cancer proliferation and drug resistance. Furthermore, we uncover a shared mechano-signaling mechanism triggered by amyloid fibrils in both melanoma and pancreatic ductal adenocarcinoma cells. Our findings highlight the crucial role of the glycocalyx protein Agrin which binds to extracellular amyloid fibrils and acts as a necessary factor in driving amyloid-dependent YAP activation. Additionally, we reveal the involvement of the HIPPO pathway core kinase LATS1 in this signaling cascade. Finally, we demonstrate that extracellular amyloid fibrils enhance cancer cell migration and invasion. In conclusion, our research expands our knowledge of the tumor microenvironment by uncovering the role of extracellular amyloid fibrils in driving mechano-signaling and YAP activation. This knowledge opens up new avenues for developing innovative strategies to modulate YAP activation and mitigate its detrimental effects during cancer progression.


Asunto(s)
Melanoma , Neoplasias Pancreáticas , Humanos , Amiloide , Ecosistema , Transducción de Señal , Neoplasias Pancreáticas/genética , Microambiente Tumoral
6.
Front Neurol ; 14: 1098300, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-37234781

RESUMEN

Background: Extracerebral complications in patients with aneurysmal subarachnoid hemorrhage (aSAH) often occur during their stay at the neurocritical care unit (NCCU). Their influence on outcomes is poorly studied. The identification of sex-specific extracerebral complications in patients with aSAH and their impact on outcomes might aid more personalized monitoring and therapy strategies, aiming to improve outcomes. Methods: Consecutive patients with aSAH admitted to the NCCU over a 6-year period were evaluated for the occurrence of extracerebral complications (according to prespecified criteria). Outcomes were assessed with the Glasgow Outcome Scale Extended (GOSE) at 3 months and dichotomized as favorable (GOSE 5-8) and unfavorable (GOSE 1-4). Sex-specific extracerebral complications and their impact on outcomes were investigated. Based on the results of the univariate analysis, a multivariate analysis with unfavorable outcomes or the occurrence of certain complications as dependent variables was performed. Results: Overall, 343 patients were included. Most of them were women (63.6%), and they were older than men. Demographics, presence of comorbidities, radiological findings, severity of bleeding, and aneurysm-securing strategies were compared among the sexes. More women than men suffered from cardiac complications (p = 0.013) and infection (p = 0.048). Patients with unfavorable outcomes were more likely to suffer from cardiac (p < 0.001), respiratory (p < 0.001), hepatic/gastrointestinal (p = 0.023), and hematological (p = 0.021) complications. In the multivariable analysis, known factors including age, female sex, increasing number of comorbidities, increasing World Federation of Neurosurgical Societies (WFNS), and Fisher grading were expectedly associated with unfavorable outcomes. When adding complications to these models, these factors remained significant. However, when considering the complications, only pulmonary and cardiac complications remained independently associated with unfavorable outcomes. Conclusion: Extracerebral complications after aSAH are frequent. Cardiac and pulmonary complications are independent predictors of unfavorable outcomes. Sex-specific extracerebral complications in patients with aSAH exist. Women suffered more frequently from cardiac and infectious complications potentially explaining the worse outcomes.

7.
F1000Res ; 11: 1149, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-37476819

RESUMEN

Background: In patients with myocardial infarction, atypical symptoms at onset have been demonstrated in women. We aimed to investigate the presence of sex-related differences in symptom presentation in patients with aneurysmal subarachnoid hemorrhage (aSAH) to enable earlier diagnosis and treatment. Methods: We assessed symptoms on admission to hospital in 343 patients with aSAH in this retrospective single-center cohort-study. Univariate statistical analysis was performed by comparing sexes including the whole study population and subgroups (dichotomized using Fisher scale 1-2 vs. 3-4, WFNS grade 1-3 vs. 4-5, and anterior vs. posterior circulation aneurysms, respectively). Results: The majority of patients was female (63.6%, n=218, vs. 36.4%, n=125), the mean age 57.4 years (standard deviation (SD) 13.3) with older women compared to men (59.2, SD 13.8, vs. 54.4, SD 11.6; p=0.003). Anterior communicating artery (AcomA) aneurysms were most common (30.9%, n=106), predominantly in men (43.2%, n=54, vs. 23.9%, n=52; p=0.0002), whereas posterior communicating artery (PcomA) aneurysms were more frequent in women (19.3%, n=42, vs. 8.8%, n=11; p=0.005). Exercise-induced headache was more often reported by men (10.4%, n=13, vs. 5%, n=11; p=0.04) in all patients as well as in the subgroup of WFNS 1-3. Anisocoria was more frequent in women within the subgroup of severely impaired consciousness (WFNS 4-5; 25.3%, n=22, vs. 10.7%, n=6; p=0.032). For all other symptoms, there was no evidence for sex-specific differences in the whole study group as well as in subgroups. Conclusion: Our results show no evidence for relevant sex-related differences in symptom presentation at onset in aSAH patients. Women presenting with an acute onset anisocoria should be screened even more carefully for an underlying ruptured Pcom aneurysm.


Asunto(s)
Aneurisma Roto , Aneurisma Intracraneal , Hemorragia Subaracnoidea , Masculino , Humanos , Femenino , Anciano , Persona de Mediana Edad , Hemorragia Subaracnoidea/complicaciones , Hemorragia Subaracnoidea/diagnóstico , Hemorragia Subaracnoidea/epidemiología , Estudios Retrospectivos , Anisocoria , Aneurisma Intracraneal/complicaciones
8.
J Exp Clin Cancer Res ; 41(1): 34, 2022 Jan 24.
Artículo en Inglés | MEDLINE | ID: mdl-35073946

RESUMEN

BACKGROUND: Acute myeloid leukemia (AML) is characterized by accumulation of aberrantly differentiated hematopoietic myeloid progenitor cells. The karyotyping-silent NUP98-NSD1 fusion is a molecular hallmark of pediatric AML and is associated with the activating FLT3-ITD mutation in > 70% of the cases. NUP98-NSD1 fusion protein promotes myeloid progenitor self-renewal in mice via unknown molecular mechanism requiring both the NUP98 and the NSD1 moieties. METHODS: We used affinity purification coupled to label-free mass spectrometry (AP-MS) to examine the effect of NUP98-NSD1 structural domain deletions on nuclear interactome binding. We determined their functional relevance in NUP98-NSD1 immortalized primary murine hematopoietic stem and progenitor cells (HSPC) by inducible knockdown, pharmacological targeting, methylcellulose assay, RT-qPCR analysis and/or proximity ligation assays (PLA). Fluorescence recovery after photobleaching and b-isoxazole assay were performed to examine the phase transition capacity of NUP98-NSD1 in vitro and in vivo. RESULTS: We show that NUP98-NSD1 core interactome binding is largely dependent on the NUP98 phenylalanine-glycine (FG) repeat domains which mediate formation of liquid-like phase-separated NUP98-NSD1 nuclear condensates. We identified condensate constituents including imitation switch (ISWI) family member SMARCA5 and BPTF (bromodomain PHD finger transcription factor), both members of the nucleosome remodeling factor complex (NURF). We validated the interaction with SMARCA5 in NUP98-NSD1+ patient cells and demonstrated its functional role in NUP98-NSD1/FLT3-ITD immortalized primary murine hematopoietic cells by genetic and pharmacological targeting. Notably, SMARCA5 inhibition did not affect NUP98-NSD1 condensates suggesting that functional activity rather than condensate formation per se is crucial to maintain the transformed phenotype. CONCLUSIONS: NUP98-NSD1 interacts and colocalizes on the genome with SMARCA5 which is an essential mediator of the NUP98-NSD1 transformation in hematopoietic cells. Formation of NUP98-NSD1 phase-separated nuclear condensates is not sufficient for the maintenance of transformed phenotype, which suggests that selective targeting of condensate constituents might represent a new therapeutic strategy for NUP98-NSD1 driven AML.


Asunto(s)
Adenosina Trifosfatasas/metabolismo , Proteínas Cromosómicas no Histona/metabolismo , Hematopoyesis/genética , Proteínas de Complejo Poro Nuclear/metabolismo , Proteómica/métodos , Animales , Humanos , Ratones
9.
Cancer ; 117(24): 5519-28, 2011 Dec 15.
Artículo en Inglés | MEDLINE | ID: mdl-21692061

RESUMEN

BACKGROUND: Emerging evidence has suggested that the capability to sustain tumor formation, growth, and chemotherapy resistance in ovarian as well as other human malignancies exclusively resides in a small proportion of tumor cells termed cancer stem cells. During the characterization of CD44(+) ovarian cancer stem cells, we found a high expression of the genes encoding for claudin-4. Because this tight junction protein is the natural high-affinity receptor for Clostridium perfringens enterotoxin (CPE), we have extensively investigated the sensitivity of ovarian cancer stem cells to CPE treatment in vitro and in vivo. METHODS: Real-time polymerase chain reaction and flow cytometry were used to evaluate claudin-3/-4 expression in ovarian cancer stem cells. Small interfering RNA knockdown experiments and MTS assays were used to evaluate CPE-induced cytotoxicity against ovarian cancer stem cell lines in vitro. C.B-17/SCID mice harboring ovarian cancer stem cell xenografts were used to evaluate CPE therapeutic activity in vivo. RESULTS: CD44(+) ovarian cancer stem cells expressed claudin-4 gene at significantly higher levels than matched autologous CD44(-) ovarian cancer cells, and regardless of their higher resistance to chemotherapeutic agents died within 1 hour after exposure to 1.0 µg/mL of CPE in vitro. Conversely, small-interfering RNA-mediated knockdown of claudin-3/-4 expression in CD44(+) cancer stem cells significantly protected cancer stem cells from CPE-induced cytotoxicity. Importantly, multiple intraperitoneal administrations of sublethal doses of CPE in mice harboring xenografts of chemotherapy-resistant CD44(+) ovarian cancer stem cells had a significant inhibitory effect on tumor progression leading to the cure and/or long-term survival of all treated animals (ie, 100% reduction in tumor burden in 50% of treated mice; P < .0001). CONCLUSIONS: CPE may represent an unconventional, potentially highly effective strategy to eradicate chemotherapy-resistant cancer stem cells.


Asunto(s)
Clostridium perfringens/química , Enterotoxinas/farmacología , Receptores de Hialuranos/biosíntesis , Neoplasias Glandulares y Epiteliales/tratamiento farmacológico , Neoplasias Glandulares y Epiteliales/patología , Células Madre Neoplásicas/efectos de los fármacos , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/patología , Adulto , Anciano , Animales , Carcinoma Epitelial de Ovario , Línea Celular Tumoral , Chlorocebus aethiops , Claudina-3 , Claudinas/genética , Claudinas/metabolismo , Clostridium perfringens/metabolismo , Enterotoxinas/biosíntesis , Enterotoxinas/farmacocinética , Femenino , Citometría de Flujo , Humanos , Inyecciones Intraperitoneales , Ratones , Ratones SCID , Persona de Mediana Edad , Neoplasias Glandulares y Epiteliales/genética , Neoplasias Glandulares y Epiteliales/metabolismo , Células Madre Neoplásicas/metabolismo , Neoplasias Ováricas/genética , Neoplasias Ováricas/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Células Vero , Ensayos Antitumor por Modelo de Xenoinjerto
10.
J Virol ; 83(13): 6779-89, 2009 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-19386711

RESUMEN

Papillomavirus-like particles (VLPs) based on L1 capsid protein represent a promising prophylactic vaccine against human papillomavirus (HPV) infections. However, cell-mediated immune responses against this antigen are believed to be of limited therapeutic value in established HPV-infected cervical lesions and, for this reason, have not been intensively investigated in cervical cancer patients. In this study we analyzed and quantified by real-time PCR (RT-PCR) the RNA expression levels of E6, E7, and L1 genes in flash-frozen HPV-16 cervical carcinomas. In addition, the kinetics of expression of E6, E7, and L1 in HPV-16-infected primary cell lines established as long-term cultures in vitro was also evaluated at RNA and protein levels. Finally, in order to evaluate the therapeutic potential of L1-specific CD4(+) and CD8(+) T lymphocytes responses in cervical cancer patients, L1 VLP-loaded dendritic cells (DCs) were used to stimulate peripheral blood lymphocytes from cervical cancer patients and such responses were compared to those elicited by the E7 oncoprotein. We show that 22 of 22 (100%) flash-frozen cervical biopsy samples collected from HPV-16-positive cervical cancer patients harbor L1, in addition to E6 and E7 RNA, as detected by RT-PCR. E7 RNA copy number (mean, 176.2) was significantly higher in HPV-16-positive cervical cancers compared to the E6 RNA copy number (mean, 47.3) and the L1 copy number (mean, 58.3) (P < 0.0001 and P < 0.001, respectively). However, no significant differences in expression levels between E6 and L1 were found. Kinetic studies of E6, E7, and L1 RNA and protein expression levels in primary tumors showed a sharp reduction in L1 expression after multiple in vitro passages compared to E6 and E7. Autologous DCs pulsed with HPV-16 VLPs or recombinant full-length E7 elicited strong type 1 L1- and E7-specific responses in CD4(+) and CD8(+) T cells from cervical cancer patients. Importantly, L1 VLP-specific CD8(+) T lymphocytes expressed strong cytolytic activity against autologous tumor cells and were as effective as E7-specific cytotoxic T lymphocytes in lysing naturally HPV-16-infected autologous tumor cells. Taken together, these data demonstrate a consistent expression of L1 in primary cervical tumors and the possibility of inducing effective L1/tumor-specific CD4(+) and CD8(+) T-lymphocyte responses in patients harboring HPV-infected cervical cancer. These results may have important implications for the treatment of patients harboring established HPV-infected lesions with L1 VLPs or combined E7/L1 DC-based vaccinations.


Asunto(s)
Papillomavirus Humano 16/inmunología , Infecciones por Papillomavirus/inmunología , Linfocitos T Citotóxicos/inmunología , Neoplasias del Cuello Uterino/inmunología , Adulto , Anciano , Vacunas contra el Cáncer/inmunología , Proteínas de la Cápside/inmunología , Proteínas de la Cápside/metabolismo , Línea Celular Tumoral , Células Dendríticas/inmunología , Femenino , Perfilación de la Expresión Génica , Papillomavirus Humano 16/genética , Humanos , Persona de Mediana Edad , Proteínas Oncogénicas Virales/inmunología , Proteínas Oncogénicas Virales/metabolismo , Proteínas E7 de Papillomavirus , ARN Viral/metabolismo , Proteínas Represoras/metabolismo , Linfocitos T Citotóxicos/virología , Neoplasias del Cuello Uterino/virología , Adulto Joven
11.
BMC Cancer ; 10: 349, 2010 Jul 02.
Artículo en Inglés | MEDLINE | ID: mdl-20598131

RESUMEN

BACKGROUND: Development of innovative, effective therapies against recurrent/chemotherapy-resistant ovarian cancer remains a high priority. Using high-throughput technologies to analyze genetic fingerprints of ovarian cancer, we have discovered extremely high expression of the genes encoding the proteins claudin-3 and claudin-4. METHODS: Because claudin-3 and -4 are the epithelial receptors for Clostridium perfringens enterotoxin (CPE), and are sufficient to mediate CPE binding, in this study we evaluated the in vitro and in vivo bioactivity of the carboxy-terminal fragment of CPE (i.e., CPE290-319 binding peptide) as a carrier for tumor imaging agents and intracellular delivery of therapeutic drugs. Claudin-3 and -4 expression was examined with rt-PCR and flow cytometry in multiple primary ovarian carcinoma cell lines. Cell binding assays were used to assess the accuracy and specificity of the CPE peptide in vitro against primary chemotherapy-resistant ovarian carcinoma cell lines. Confocal microscopy and biodistribution assays were performed to evaluate the localization and uptake of the FITC-conjugated CPE peptide in established tumor tissue. RESULTS: Using a FITC-conjugated CPE peptide we show specific in vitro and in vivo binding to multiple primary chemotherapy resistant ovarian cancer cell lines. Bio-distribution studies in SCID mice harboring clinically relevant animal models of chemotherapy resistant ovarian carcinoma showed higher uptake of the peptide in tumor cells than in normal organs. Imunofluorescence was detectable within discrete accumulations (i.e., tumor spheroids) or even single chemotherapy resistant ovarian cancer cells floating in the ascites of xenografted animals while a time-dependent internalization of the FITC-conjugated CPE peptide was consistently noted in chemotherapy-resistant ovarian tumor cells by confocal microscopy. CONCLUSIONS: Based on the high levels of claudin-3 and -4 expression in chemotherapy-resistant ovarian cancer and other highly aggressive human epithelial tumors including breast, prostate and pancreatic cancers, CPE peptide holds promise as a lead peptide for the development of new diagnostic tracers or alternative anticancer agents.


Asunto(s)
Adenocarcinoma de Células Claras/tratamiento farmacológico , Adenocarcinoma/tratamiento farmacológico , Carcinoma Papilar/tratamiento farmacológico , Carcinoma de Células Escamosas/tratamiento farmacológico , Cistadenocarcinoma Seroso/tratamiento farmacológico , Enterotoxinas/farmacología , Neoplasias Ováricas/tratamiento farmacológico , Fragmentos de Péptidos/farmacología , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Adenocarcinoma de Células Claras/genética , Adenocarcinoma de Células Claras/metabolismo , Animales , Carcinoma Papilar/genética , Carcinoma Papilar/metabolismo , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Chlorocebus aethiops , Claudina-3 , Claudina-4 , Clostridium perfringens , Cistadenocarcinoma Seroso/genética , Cistadenocarcinoma Seroso/metabolismo , Resistencia a Antineoplásicos , Femenino , Fibroblastos , Citometría de Flujo , Humanos , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones , Ratones SCID , Neoplasias Ováricas/genética , Neoplasias Ováricas/metabolismo , Fragmentos de Péptidos/farmacocinética , ARN Mensajero/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Esferoides Celulares/efectos de los fármacos , Distribución Tisular , Trasplante Heterólogo , Células Tumorales Cultivadas , Neoplasias del Cuello Uterino/tratamiento farmacológico , Neoplasias del Cuello Uterino/genética , Neoplasias del Cuello Uterino/metabolismo , Células Vero
12.
Gynecol Oncol ; 119(1): 140-5, 2010 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-20673976

RESUMEN

OBJECTIVE: To compare the in vitro sensitivity/resistance to patupilone versus paclitaxel in uterine serous papillary carcinoma (USPC) with high versus low HER-2/neu expression. METHODS: Six primary USPC cell lines, half of which overexpress HER-2/neu at a 3+ level, were evaluated for growth rate and tested for their in vitro sensitivity/resistance to patupilone versus paclitaxel by MTS assays. Quantitative RT-PCR was used to identify potential mechanisms underlying the differential sensitivity/resistance to patupilone versus paclitaxel in primary USPC cell lines. RESULTS: Cell lines overexpressing HER-2/neu showed higher proliferation when compared to low HER-2/neu-expressing cell lines. Compared to low-expressing cell lines, high HER-2/neu expressors were significantly more sensitive to patupilone than to paclitaxel (P<0.0002). In contrast, there was no appreciable difference in sensitivity to patupilone versus paclitaxel in primary USPC cell lines with low HER-2/neu expression. Higher levels of ß-tubulin III (TUBB3) and P-glycoprotein (ABCB1) were detected in USPC cell lines with high versus low HER-2/neu expression (P<0.05). CONCLUSIONS: USPC overexpressing HER-2/neu display greater in vitro sensitivity to patupilone and higher levels of the patupilone molecular target TUBB3 when compared to low HER-2/neu expressors. Due to the adverse prognosis associated with HER-2/neu overexpression in USPC patients, patupilone may represent a promising novel drug to combine to platinum compounds in this subset of aggressive endometrial tumors.


Asunto(s)
Antineoplásicos/farmacología , Carcinoma Papilar/tratamiento farmacológico , Cistadenocarcinoma Seroso/tratamiento farmacológico , Epotilonas/farmacología , Paclitaxel/farmacología , Receptor ErbB-2/biosíntesis , Neoplasias Uterinas/tratamiento farmacológico , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/biosíntesis , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/metabolismo , Antineoplásicos Fitogénicos/farmacología , Carcinoma Papilar/enzimología , Carcinoma Papilar/metabolismo , Línea Celular Tumoral , Cistadenocarcinoma Seroso/enzimología , Cistadenocarcinoma Seroso/metabolismo , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Femenino , Humanos , Tubulina (Proteína)/biosíntesis , Tubulina (Proteína)/metabolismo , Neoplasias Uterinas/enzimología , Neoplasias Uterinas/metabolismo
13.
Int J Gynecol Cancer ; 20(9): 1440-7, 2010 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-21370592

RESUMEN

INTRODUCTION: Epithelial cell adhesion molecule (EpCAM) is a surface glycoprotein highly differentially expressed in many epithelial malignancies. The goal of this study was to evaluate the expression of EpCAM and the potential of MT201 (adecatumumab), a human monoclonal antibody targeting EpCAM, against multiple primary cervical carcinoma cell lines. METHODS: Epithelial cell adhesion molecule expression was evaluated by real-time polymerase chain reaction and flow cytometry in a total of 8 primary cervical cancer cell lines. Sensitivity to MT201-mediated cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity was tested in standard 4-hour 51Cr release assays. To investigate the effect of interleukin-2 (IL-2) on MT201-mediated ADCC, 4-hour 51Cr release assays were also conducted in the presence of low doses of IL-2. RESULTS: High messenger RNA expression by real-time polymerase chain reaction and high EpCAM surface expression by flow cytometry were detected in 4 (50%) of 8 primary cervical carcinoma cell lines. With no exception, the primary cell lines derived from clinically aggressive tumors showed EpCAM overexpression. Whereas these cell lines were highly resistant to complement-dependent cytotoxicity and natural killer (NK)-dependent cytotoxicity in vitro (range of killing, 4%-19%), EpCAM-positive cell lines showed high sensitivity to MT201-mediated ADCC (range of killing, 23%-59%). Incubation with IL-2 in addition to MT201 significantly increased the cytotoxic activity against EpCAM-positive cervical cancer cell lines (P = 0.007). Addition of human serum also further increased the MT201-mediated killing of EpCAM-positive cell lines (P = 0.03). CONCLUSIONS: Epithelial cell adhesion molecule is highly expressed in primary cervical carcinoma cell lines, and these biologically aggressive tumors are highly sensitive to MT201-mediated cytotoxicity in vitro. MT201 may represent a novel, potentially highly effective treatment option for patients with cervical carcinoma, especially for those with advanced, recurrent, or metastatic disease refractory to standard salvage therapy.


Asunto(s)
Anticuerpos Monoclonales/uso terapéutico , Antígenos de Neoplasias/genética , Carcinoma/genética , Carcinoma/terapia , Moléculas de Adhesión Celular/genética , Moléculas de Adhesión Celular/uso terapéutico , Inmunoterapia/métodos , Neoplasias del Cuello Uterino/genética , Neoplasias del Cuello Uterino/terapia , Adulto , Anciano , Anticuerpos Monoclonales/farmacología , Anticuerpos Monoclonales Humanizados , Antígenos de Neoplasias/inmunología , Antígenos de Neoplasias/metabolismo , Carcinoma/metabolismo , Carcinoma/patología , Moléculas de Adhesión Celular/antagonistas & inhibidores , Moléculas de Adhesión Celular/inmunología , Moléculas de Adhesión Celular/metabolismo , Moléculas de Adhesión Celular/farmacología , Técnicas de Cultivo de Célula , Línea Celular Tumoral , Molécula de Adhesión Celular Epitelial , Femenino , Citometría de Flujo , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Humanos , Persona de Mediana Edad , Resultado del Tratamiento , Neoplasias del Cuello Uterino/metabolismo , Neoplasias del Cuello Uterino/patología , Adulto Joven
14.
Nat Commun ; 11(1): 1345, 2020 03 12.
Artículo en Inglés | MEDLINE | ID: mdl-32165637

RESUMEN

In several metazoans, the number of active replication origins in embryonic nuclei is higher than in somatic ones, ensuring rapid genome duplication during synchronous embryonic cell divisions. High replication origin density can be restored by somatic nuclear reprogramming. However, mechanisms underlying high replication origin density formation coupled to rapid cell cycles are poorly understood. Here, using Xenopus laevis, we show that SSRP1 stimulates replication origin assembly on somatic chromatin by promoting eviction of histone H1 through its N-terminal domain. Histone H1 removal derepresses ORC and MCM chromatin binding, allowing efficient replication origin assembly. SSRP1 protein decays at mid-blastula transition (MBT) when asynchronous somatic cell cycles start. Increasing levels of SSRP1 delay MBT and, surprisingly, accelerate post-MBT cell cycle speed and embryo development. These findings identify a major epigenetic mechanism regulating DNA replication and directly linking replication origin assembly, cell cycle duration and embryo development in vertebrates.


Asunto(s)
Proteínas de Unión al ADN/metabolismo , Histonas/metabolismo , Proteínas de Xenopus/metabolismo , Xenopus laevis/genética , Xenopus laevis/metabolismo , Animales , Blástula/embriología , Blástula/metabolismo , Cromatina/genética , Cromatina/metabolismo , Replicación del ADN , Proteínas de Unión al ADN/genética , Proteínas del Grupo de Alta Movilidad , Histonas/química , Histonas/genética , Dominios Proteicos , Origen de Réplica , Proteínas de Xenopus/genética , Xenopus laevis/embriología
15.
Biochim Biophys Acta Mol Basis Dis ; 1866(1): 165581, 2020 01 01.
Artículo en Inglés | MEDLINE | ID: mdl-31672549

RESUMEN

Exposure to environmental toxins, including hydrocarbon solvents, increases the risk of developing Parkinson's disease. An emergent hypothesis considers microtubule dysfunction as one of the crucial events in triggering neuronal degeneration in Parkinson's disease. Here, we used 2,5-hexanedione (2,5-HD), the toxic metabolite of n-hexane, to analyse the early effects of toxin-induced neurodegeneration on the cytoskeleton in multiple model systems. In PC12 cells differentiated with nerve growth factor for 5 days, we found that 2,5-HD treatment affected all the cytoskeletal components. Moreover, we observed alterations in microtubule distribution and stability, in addition to the imbalance of post-translational modifications of α-tubulin. Similar defects were also found in vivo in 2,5-HD-intoxicated mice. Interestingly, we also found that 2,5-HD exposure induced significant changes in microtubule stability in human skin fibroblasts obtained from Parkinson's disease patients harbouring mutations in PRKN gene, whereas it was ineffective in healthy donor fibroblasts, suggesting that the genetic background may really make the difference in microtubule susceptibility to this environmental Parkinson's disease-related toxin. In conclusion, by showing the imbalance between dynamic and stable microtubules in hydrocarbon-induced parkinsonism, our data support the crucial role of microtubule defects in triggering neurodegeneration.


Asunto(s)
Hexanonas/farmacología , Microtúbulos/efectos de los fármacos , Enfermedad de Parkinson/etiología , Enfermedad de Parkinson/metabolismo , Animales , Línea Celular , Citoesqueleto/efectos de los fármacos , Citoesqueleto/metabolismo , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Humanos , Ratones , Microtúbulos/metabolismo , Factores de Crecimiento Nervioso/metabolismo , Células PC12 , Trastornos Parkinsonianos/inducido químicamente , Trastornos Parkinsonianos/metabolismo , Ratas , Tubulina (Proteína)/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo
16.
Neurobiol Aging ; 61: 66-74, 2018 01.
Artículo en Inglés | MEDLINE | ID: mdl-29040870

RESUMEN

Loss-of-function caused by mutations in the parkin gene (PARK2) lead to early-onset familial Parkinson's disease. Recently, mechanistic studies proved the ability of parkin in regulating mitochondria homeostasis and microtubule (MT) stability. Looking at these systems during aging of PARK2 knockout mice, we found that loss of parkin induced an accelerated (over)acetylation of MT system both in dopaminergic neuron cell bodies and fibers, localized in the substantia nigra and corpus striatum, respectively. Interestingly, in PARK2 knockout mice, changes of MT stability preceded the alteration of mitochondria transport. Moreover, in-cell experiments confirmed that loss of parkin affects mitochondria mobility and showed that this defect depends on MT system as it is rescued by paclitaxel, a well-known MT-targeted agent. Furthermore, both in PC12 neuronal cells and in patients' induced pluripotent stem cell-derived midbrain neurons, we observed that parkin deficiencies cause the fragmentation of stable MTs. Therefore, we suggest that parkin acts as a regulator of MT system during neuronal aging, and we endorse the hypothesis that MT dysfunction may be crucial in the pathogenesis of Parkinson's disease.


Asunto(s)
Envejecimiento/patología , Neuronas Dopaminérgicas/patología , Mutación con Pérdida de Función , Microtúbulos/patología , Enfermedad de Parkinson/etiología , Ubiquitina-Proteína Ligasas/deficiencia , Ubiquitina-Proteína Ligasas/genética , Acetilación , Animales , Humanos , Ratones Noqueados , Microtúbulos/metabolismo , Mitocondrias/metabolismo , Células PC12 , Paclitaxel/farmacología , Enfermedad de Parkinson/genética , Ratas , Ubiquitina-Proteína Ligasas/fisiología
17.
Sci Rep ; 6: 33289, 2016 09 15.
Artículo en Inglés | MEDLINE | ID: mdl-27628239

RESUMEN

α-Synuclein is a presynaptic protein associated to Parkinson's disease, which is unstructured when free in the cytoplasm and adopts α helical conformation when bound to vesicles. After decades of intense studies, α-Synuclein physiology is still difficult to clear up due to its interaction with multiple partners and its involvement in a pletora of neuronal functions. Here, we looked at the remarkably neglected interplay between α-Synuclein and microtubules, which potentially impacts on synaptic functionality. In order to identify the mechanisms underlying these actions, we investigated the interaction between purified α-Synuclein and tubulin. We demonstrated that α-Synuclein binds to microtubules and tubulin α2ß2 tetramer; the latter interaction inducing the formation of helical segment(s) in the α-Synuclein polypeptide. This structural change seems to enable α-Synuclein to promote microtubule nucleation and to enhance microtubule growth rate and catastrophe frequency, both in vitro and in cell. We also showed that Parkinson's disease-linked α-Synuclein variants do not undergo tubulin-induced folding and cause tubulin aggregation rather than polymerization. Our data enable us to propose α-Synuclein as a novel, foldable, microtubule-dynamase, which influences microtubule organisation through its binding to tubulin and its regulating effects on microtubule nucleation and dynamics.


Asunto(s)
Enfermedad de Parkinson/genética , Agregación Patológica de Proteínas/genética , Tubulina (Proteína)/metabolismo , alfa-Sinucleína/metabolismo , Humanos , Microtúbulos/química , Microtúbulos/metabolismo , Enfermedad de Parkinson/metabolismo , Enfermedad de Parkinson/patología , Unión Proteica , Pliegue de Proteína , Multimerización de Proteína/genética , Tubulina (Proteína)/química , Tubulina (Proteína)/genética , alfa-Sinucleína/química , alfa-Sinucleína/genética
18.
Funct Neurol ; 28(3): 205-21, 2013.
Artículo en Inglés | MEDLINE | ID: mdl-24139657

RESUMEN

Awake surgery requires coordinated teamwork and communication between the surgeon and the anesthesiologist, as he monitors the patient, the neuroradiologist as he interprets the images for intraoperative confirmation, and the neuropsychologist and neurophysiologist as they evaluate in real-time the patient's responses to commands and questions. To improve comparison across published studies on clinical assessment and operative settings in awake surgery, we reviewed the literature, focusing on methodological differences and aims. In complex, interdisciplinary medical care, such differences can affect the outcome and the cost-benefit ratio of the treatment. Standardization of intraoperative mapping and related controversies will be discussed in Part II.


Asunto(s)
Enfermedades del Sistema Nervioso/cirugía , Procedimientos Neuroquirúrgicos , Vigilia , Anestesia , Estimulación Eléctrica , Humanos , Procesamiento de Imagen Asistido por Computador , Inmunohistoquímica , Imagen por Resonancia Magnética , Enfermedades del Sistema Nervioso/diagnóstico , Enfermedades del Sistema Nervioso/psicología , Pruebas Neuropsicológicas , Procedimientos Neuroquirúrgicos/métodos , Selección de Paciente , Cuidados Preoperatorios , Resultado del Tratamiento
19.
Sci Rep ; 3: 1837, 2013.
Artículo en Inglés | MEDLINE | ID: mdl-23670541

RESUMEN

The role of microtubule (MT) dysfunction in Parkinson's disease is emerging. It is still unknown whether it is a cause or a consequence of neurodegeneration. Our objective was to assess whether alterations of MT stability precede or follow axonal transport impairment and neurite degeneration in experimental parkinsonism induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) in C57Bl mice. MPTP induced a time- and dose-dependent increase in fibres with altered mitochondria distribution, and early changes in cytoskeletal proteins and MT stability. Indeed, we observed significant increases in neuron-specific ßIII tubulin and enrichment of deTyr tubulin in dopaminergic neurons. Finally, we showed that repeated daily administrations of the MT stabilizer Epothilone D rescued MT defects and attenuated nigrostriatal degeneration induced by MPTP. These data suggest that alteration of ΜΤs is an early event specifically associated with dopaminergic neuron degeneration. Pharmacological stabilization of MTs may be a viable strategy for the management of parkinsonism.


Asunto(s)
1-Metil-4-fenil-1,2,3,6-Tetrahidropiridina/farmacología , Neuronas Dopaminérgicas/efectos de los fármacos , Epotilonas/uso terapéutico , Intoxicación por MPTP/prevención & control , Microtúbulos/efectos de los fármacos , Fármacos Neuroprotectores/uso terapéutico , Trastornos Parkinsonianos/prevención & control , Animales , Transporte Axonal/efectos de los fármacos , Western Blotting , Dopaminérgicos/toxicidad , Neuronas Dopaminérgicas/metabolismo , Neuronas Dopaminérgicas/patología , Técnicas para Inmunoenzimas , Intoxicación por MPTP/inducido químicamente , Intoxicación por MPTP/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Microtúbulos/metabolismo , Microtúbulos/patología , Degeneración Nerviosa , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Neuronas/patología , Trastornos Parkinsonianos/inducido químicamente , Trastornos Parkinsonianos/patología , Procesamiento Proteico-Postraduccional/efectos de los fármacos , Sustancia Negra/efectos de los fármacos , Sustancia Negra/metabolismo , Sustancia Negra/patología , Tubulina (Proteína)/metabolismo , Moduladores de Tubulina/uso terapéutico , Tirosina 3-Monooxigenasa/metabolismo
20.
PLoS One ; 7(5): e37467, 2012.
Artículo en Inglés | MEDLINE | ID: mdl-22666358

RESUMEN

Data from both toxin-based and gene-based models suggest that dysfunction of the microtubule system contributes to the pathogenesis of Parkinson's disease, even if, at present, no evidence of alterations of microtubules in vivo or in patients is available. Here we analyze cytoskeleton organization in primary fibroblasts deriving from patients with idiopathic or genetic Parkinson's disease, focusing on mutations in parkin and leucine-rich repeat kinase 2. Our analyses reveal that genetic and likely idiopathic pathology affects cytoskeletal organization and stability, without any activation of autophagy or apoptosis. All parkinsonian fibroblasts have a reduced microtubule mass, represented by a higher fraction of unpolymerized tubulin in respect to control cells, and display significant changes in microtubule stability-related signaling pathways. Furthermore, we show that the reduction of microtubule mass is so closely related to the alteration of cell morphology and behavior that both pharmacological treatment with microtubule-targeted drugs, and genetic approaches, by transfecting the wild type parkin or leucine-rich repeat kinase 2, restore the proper microtubule stability and are able to rescue cell architecture. Taken together, our results suggest that microtubule destabilization is a point of convergence of genetic and idiopathic forms of parkinsonism and highlight, for the first time, that microtubule dysfunction occurs in patients and not only in experimental models of Parkinson's disease. Therefore, these data contribute to the knowledge on molecular and cellular events underlying Parkinson's disease and, revealing that correction of microtubule defects restores control phenotype, may offer a new therapeutic target for the management of the disease.


Asunto(s)
Fibroblastos/patología , Microtúbulos/metabolismo , Enfermedad de Parkinson/genética , Enfermedad de Parkinson/patología , Adulto , Anciano , Femenino , Fibroblastos/efectos de los fármacos , Glucógeno Sintasa Quinasa 3/metabolismo , Glucógeno Sintasa Quinasa 3 beta , Humanos , Masculino , Microtúbulos/efectos de los fármacos , Persona de Mediana Edad , Paclitaxel/farmacología , Enfermedad de Parkinson/metabolismo , Fenotipo , Fosforilación/efectos de los fármacos
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