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Background: Patients with high-risk neuroblastoma (NB) have a 5-year event-free survival of less than 50 %, and novel and improved treatment options are needed. Radiolabeled somatostatin analogs (SSTAs) could be a treatment option. The aims of this work were to compare the biodistribution and the therapeutic effects of 177Lu-octreotate and 177Lu-octreotide in mice bearing the human CLB-BAR NB cell line, and to evaluate their regulatory effects on apoptosis-related genes. Methods: The biodistribution of 177Lu-octreotide in mice bearing CLB-BAR tumors was studied at 1, 24, and 168 h after administration, and the absorbed dose was estimated to tumor and normal tissues. Further, animals were administered different amounts of 177Lu-octreotate or 177Lu-octreotide. Tumor volume was measured over time and compared to a control group given saline. RNA was extracted from tumors, and the expression of 84 selected genes involved in apoptosis was quantified with qPCR. Results: The activity concentration was generally lower in most tissues for 177Lu-octreotide compared to 177Lu-octreotate. Mean absorbed dose per administered activity to tumor after injection of 1.5 MBq and 15 MBq was 0.74 and 0.03 Gy/MBq for 177Lu-octreotide and 2.9 and 0.45 Gy/MBq for 177Lu-octreotate, respectively. 177Lu-octreotide treatment resulted in statistically significant differences compared to controls. Fractionated administration led to a higher survival fraction than after a single administration. The pro-apoptotic genes TNSFS8, TNSFS10, and TRADD were regulated after administration with 177Lu-octreotate. Treatment with 177Lu-octreotide yielded regulation of the pro-apoptotic genes CASP5 and TRADD, and of the anti-apoptotic gene IL10 as well as the apoptosis-related gene TNF. Conclusion: 177Lu-octreotide gave somewhat better anti-tumor effects than 177Lu-octreotate. The similar effect observed in the treated groups with 177Lu-octreotate suggests saturation of the somatostatin receptors. Pronounced anti-tumor effects following fractionated administration merited receptor saturation as an explanation. The gene expression analyses suggest apoptosis activation through the extrinsic pathway for both radiopharmaceuticals.
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Background: Co-use of alcohol and other drugs within a certain time frame (i.e., polysubstance use) has become increasingly prevalent, particularly among college-aged individuals, but understanding motives for co-use remains limited. Polysubstance use has been associated with a higher likelihood of negative health consequences as compared to single substance use. Objectives: The current study examined associations between motivations for using alcohol, tobacco, and cannabis among college students who use multiple substances versus students using only one substance or no substances. Additionally, we examined the effect of trauma and daily stress on polysubstance use in self-report data from individuals (N=134) participating in the MAPme Study. Results: First, the observed prevalence of polysubstance use was greater than expected by chance, with most individuals co-using alcohol and cannabis. "Alcohol and Other Drug Users" were more frequently motivated to drink for social (ß=0.27, CI=[0.07, 0.44]), enhancement (ß=0.26, CI=[0.01, 0.42]) and coping (ß=0.21, CI=[0.06, 0.47]) reasons compared to individuals who consumed alcohol alone. Conclusions: Individual differences in motivations for use were partly explained by frequency of alcohol use and alcohol problem severity, but not by history of trauma or stress. Finally, while patterns of correlations among motivations for use across substances suggested a general tendency to be motivated to use substances for similar reasons, this was not supported by confirmatory factor models. Overall, shared motives may inform potential behavioral patterns for co-use of substances during college and might advise future treatment efforts.
Emerging adults tend to use multiple substances, particularly alcohol and cannabisCorrelation patterns suggest shared motives within rather than across substancesAlcohol problem severity and alcohol frequency predict motives for use.
Assuntos
Motivação , Estresse Psicológico , Estudantes , Transtornos Relacionados ao Uso de Substâncias , Humanos , Masculino , Feminino , Estudantes/psicologia , Adulto Jovem , Universidades , Estresse Psicológico/psicologia , Estresse Psicológico/epidemiologia , Transtornos Relacionados ao Uso de Substâncias/epidemiologia , Transtornos Relacionados ao Uso de Substâncias/psicologia , Individualidade , Adolescente , Adulto , Consumo de Álcool na Faculdade/psicologia , Adaptação Psicológica , Consumo de Bebidas Alcoólicas/epidemiologia , Consumo de Bebidas Alcoólicas/psicologiaRESUMO
INTRODUCTION: Echinoderm microtubule-associated protein-like 4 (EML4)-Anaplastic Lymphoma Kinase (ALK) rearrangements occur in 3% to 7% of lung adenocarcinomas and are targets for treatment with tyrosine kinase inhibitors (TKIs). Here we have developed three novel EML4-ALK-positive patient-derived Non-Small-Cell-Lung-Cancer (NSCLC) cancer cell lines, CUTO8 (variant 1), CUTO9 (variant 1) and CUTO29 (variant 3) and included a fourth ALK-positive cell line YU1077 (variant 3) to study ALK-positive signaling and responses. Variants 1 and 3 are the most common EML4-ALK variants expressed in ALK-positive NSCLC, and currently cell lines representing these EML4-ALK variants are limited. MATERIALS AND METHODS: Resazurin assay was performed to evaluate cell viability. Protein levels were determined using western blotting. RNA sequencing was performed in all four cell lines to identify differentially expressed genes. Whole-genome sequencing was performed to determine the presence of EML4-ALK fusion and ALK tyrosine kinase inhibitor resistance mutations. RESULTS: In this study, we have confirmed expression of the corresponding ALK fusion protein and assessed their sensitivity to a range of ALK tyrosine kinase inhibitors. These patient derived cell lines exhibit differential sensitivity to lorlatinib, brigatinib and alectinib, with EML4-ALK variant 3 containing cell lines exhibiting increased sensitivity to lorlatinib and brigatinib as compared to alectinib. These cell lines were further characterized by whole genome sequencing and RNA-seq analysis that identified the ribonucleotide reductase regulatory subunit 2 (RRM2) as a downstream and potential therapeutic target in ALK-positive NSCLC. CONCLUSION: We provide a characterization of four novel EML4-ALK-positive NSCLC cell lines, highlighting genomic heterogeneity and differential responses to ALK TKI treatment. The RNA-Seq characterization of ALK-positive NSCLC CUTO8, CUTO9, CUTO29 and YU1077 cell lines reported here, has been compiled in an interactive ShinyApp resource for public data exploration (https://ccgg.ugent.be/shiny/nsclc_rrm2_2022/).
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Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Ribonucleosídeo Difosfato Redutase , Quinase do Linfoma Anaplásico/genética , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Proteínas de Fusão Oncogênica/genética , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Ribonucleosídeo Difosfato Redutase/metabolismoRESUMO
The first-in-class inhibitor of ALK, c-MET and ROS1, crizotinib (Xalkori), has shown remarkable clinical efficacy in treatment of ALK-positive non-small cell lung cancer. However, in neuroblastoma, activating mutations in the ALK kinase domain are typically refractory to crizotinib treatment, highlighting the need for more potent inhibitors. The next-generation ALK inhibitor PF-06463922 is predicted to exhibit increased affinity for ALK mutants prevalent in neuroblastoma. We examined PF-06463922 activity in ALK-driven neuroblastoma models in vitro and in vivo In vitro kinase assays and cell-based experiments examining ALK mutations of increasing potency show that PF-06463922 is an effective inhibitor of ALK with greater activity towards ALK neuroblastoma mutants. In contrast to crizotinib, single agent administration of PF-06463922 caused dramatic tumor inhibition in both subcutaneous and orthotopic xenografts as well as a mouse model of high-risk neuroblastoma driven by Th-ALK(F1174L)/MYCN Taken together, our results suggest PF-06463922 is a potent inhibitor of crizotinib-resistant ALK mutations, and highlights an important new treatment option for neuroblastoma patients.
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Lactamas Macrocíclicas/uso terapêutico , Proteína Proto-Oncogênica N-Myc/antagonistas & inibidores , Neuroblastoma/tratamento farmacológico , Inibidores de Proteínas Quinases/uso terapêutico , Receptores Proteína Tirosina Quinases/antagonistas & inibidores , Aminopiridinas , Quinase do Linfoma Anaplásico , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Ensaios Clínicos como Assunto , Crizotinibe , Lactamas , Lactamas Macrocíclicas/farmacologia , Camundongos Endogâmicos BALB C , Camundongos Nus , Mutação/genética , Proteína Proto-Oncogênica N-Myc/metabolismo , Neuroblastoma/patologia , Células PC12 , Inibidores de Proteínas Quinases/farmacologia , Pirazóis/farmacologia , Pirazóis/uso terapêutico , Piridinas/farmacologia , Piridinas/uso terapêutico , Ratos , Receptores Proteína Tirosina Quinases/genética , Receptores Proteína Tirosina Quinases/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
A vast array of oncogenic variants has been identified for anaplastic lymphoma kinase (ALK). Therefore, there is a need to better understand the role of ALK in cancer biology in order to optimise treatment strategies. This review summarises the latest research on the receptor tyrosine kinase ALK, and how this information can guide the management of patients with cancer that is ALK-positive. A variety of ALK gene alterations have been described across a range of tumour types, including point mutations, deletions and rearrangements. A wide variety of ALK fusions, in which the kinase domain of ALK and the amino-terminal portion of various protein partners are fused, occur in cancer, with echinoderm microtubule-associated protein-like 4 (EML4)-ALK being the most prevalent in non-small-cell lung cancer (NSCLC). Different ALK fusion proteins can mediate different signalling outputs, depending on properties such as subcellular localisation and protein stability. The ALK fusions found in tumours lack spatial and temporal regulation, which can also affect dimerisation and substrate specificity. Two ALK tyrosine kinase inhibitors (TKIs), crizotinib and ceritinib, are currently approved in Europe for use in ALK-positive NSCLC and several others are in development. These ALK TKIs bind slightly differently within the ATP-binding pocket of the ALK kinase domain and are associated with the emergence of different resistance mutation patterns during therapy. This emphasises the need to tailor the sequence of ALK TKIs according to the ALK signature of each patient. Research into the oncogenic functions of ALK, and fast paced development of ALK inhibitors, has substantially improved outcomes for patients with ALK-positive NSCLC. Limited data are available surrounding the physiological ligand-stimulated activation of ALK signalling and further research is needed. Understanding the role of ALK in tumour biology is key to further optimising therapeutic strategies for ALK-positive disease.
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Neoplasias/enzimologia , Receptores Proteína Tirosina Quinases/fisiologia , Quinase do Linfoma Anaplásico , Animais , Desenvolvimento Embrionário , Humanos , Linfoma Anaplásico de Células Grandes/enzimologia , Neoplasias/genética , Neoplasias/terapia , Neuroblastoma/enzimologia , Proteínas de Fusão Oncogênica/genética , Mutação Puntual , Receptores Proteína Tirosina Quinases/antagonistas & inibidores , Receptores Proteína Tirosina Quinases/química , Receptores Proteína Tirosina Quinases/genética , Transdução de SinaisRESUMO
BACKGROUND: Heritability estimates from twin studies of the multi-faceted phenotype of nicotine dependence (ND) range from moderate to high (31-60%), but vary substantially based on the specific ND-related construct examined. The current study estimated the aggregate role of common genetic variants on key ND constructs. METHOD: Genomic-relationship-matrix restricted maximum likelihood (GREML) was used to decompose phenotypic variance across multiple ND indices using 796 125 polymorphisms from 2346 unrelated 'lifetime ever smokers' of European ancestry. Measures included DSM-IV ND and Fagerström Test for Nicotine Dependence (FTND) summary measures and constituent constructs (e.g. withdrawal severity, tolerance, heaviness of smoking and time spent smoking). Exploratory and confirmatory factor models were used to describe the covariance structure across ND measures; resulting factor(s) were the subject(s) of GREML analyses. RESULTS: Factor models indicated highly correlated DSM-IV and FTND factors for ND (0.545, 95% confidence interval 0.50-0.60) that could be represented as a higher-order factor (NIC DEP). Additive genetic influence on NIC DEP was 33% (s.e. = 0.14, p = 0.009). Post-hoc analyses indicated moderate genetic effects on the DSM-IV (34%, s.e. = 0.14, p = 0.008) and FTND (26%, s.e. = 0.14, p = 0.032) factors, both of which were influenced by the same genetic effects (r G-SNP = 1.00, s.e. = 0.09, p < 0.00001). CONCLUSIONS: Overall, common single nucleotide polymorphisms accounted for a large proportion of the genetic influences on ND-related phenotypes that have been observed in twin studies. Genetic contributions across distinct ND scales were largely influenced by shared genetic factors.
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Genômica/métodos , Tabagismo/genética , Tabagismo/fisiopatologia , Adulto , Feminino , Predisposição Genética para Doença , Humanos , Masculino , Pessoa de Meia-Idade , Fenótipo , Polimorfismo de Nucleotídeo ÚnicoRESUMO
OBJECTIVES: To evaluate the influence of shock wave therapy (SWT) on radiographic evidence of bone healing after tibial plateau leveling osteotomy (TPLO). METHODS: Healthy dogs between two to nine years of age that underwent TPLO were randomly assigned to receive either electro-hydraulic SWT (1,000 shocks) or sham treatment (SHAM). Treatment or SHAM was administered to the osteotomy site immediately postoperatively and two weeks postoperatively. Three blinded radiologists evaluated orthogonal radiographs performed eight weeks postoperatively with both a 5-point and a 10-point bone healing scale. Linear regression analysis was used to compare median healing scores between groups. RESULTS: Forty-two dogs (50 stifles) were included in the statistical analysis. No major complications were observed and all osteotomies healed uneventfully. The median healing scores were significantly higher at eight weeks postoperatively for the SWT group compared to the SHAM group for the 10-point (p <0.0002) and 5-point scoring systems (p <0.0001). CLINICAL SIGNIFICANCE: Shock wave therapy applied immediately and two weeks postoperatively led to more advanced bone healing at the eight week time point in this study population. The results of this study support the use of electro-hydraulic SWT as a means of accelerating acute bone healing of canine osteotomies. Additional studies are needed to evaluate its use for acceleration of bone healing following fracture, or with delayed union.
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Doenças do Cão/terapia , Ondas de Choque de Alta Energia/uso terapêutico , Osteotomia/veterinária , Animais , Cães , Feminino , MasculinoRESUMO
Nicotine dependence (ND) is a heterogeneous phenotype with complex genetic influences that may vary across ethnicities. The use of intermediate phenotypes may clarify genetic influences and reveal specific etiological pathways. Prior work in European Americans has found that the four Primary Dependence Motives (PDM) subscales (Automaticity, Craving, Loss of Control, and Tolerance) of the Wisconsin Inventory of Smoking Motives represent core features of nicotine dependence and are promising intermediate phenotypes for understanding genetic pathways to ND. However, no studies have examined PDM as an intermediate phenotype in African American smokers, an ethnic population that displays unique patterns of smoking and genetic variation. In the current study, 268 African American daily smokers completed a phenotypic assessment and provided a sample of DNA. Associations among haplotypes in the NCAM1-TTC12-ANKK1-DRD2 gene cluster, a dopamine-related gene region associated with ND, PDM intermediate phenotypes, and ND were examined. Dopamine-related genetic variation in the DBH and COMT genes was also considered on an exploratory basis. Mediational analysis was used to test the indirect pathway from genetic variation to smoking motives to nicotine dependence. NCAM1-TTC12-ANKK1-DRD2 region variation was significantly associated with the Automaticity subscale and, further, Automaticity significantly mediated associations among NCAM1-TTC12-ANKK1-DRD2 cluster variants and ND. DBH was also significantly associated with Automaticity, Craving, and Tolerance; Automaticity and Tolerance also served as mediators of the DBH-ND relationship. These results suggest that PDM, Automaticity in particular, may be a viable intermediate phenotype for understanding dopamine-related genetic influences on ND in African American smokers. Findings support a model in which putatively dopaminergic variants exert influence on ND through an effect on patterns of automatic routinized smoking.
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Negro ou Afro-Americano/genética , Negro ou Afro-Americano/psicologia , Dopamina/genética , Motivação/genética , Fumar/genética , Fumar/psicologia , Adulto , Fissura , Tolerância a Medicamentos , Feminino , Variação Genética , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Testes Neuropsicológicos , Polimorfismo de Nucleotídeo Único , Estados Unidos , Adulto JovemRESUMO
RATIONALE: Nicotine dependence (ND) is a heterogeneous phenotype with complex genetic influences. The use of intermediate ND phenotypes may clarify genetic influences and reveal specific etiological pathways. Prior work has found that the four Primary Dependence Motives (PDM) subscales (Automaticity, Craving, Loss of Control, and Tolerance) of the Wisconsin Inventory of Smoking Motives (WISDM) represent heavy, pervasive smoking, which is a core feature of nicotine dependence, making these motives strong candidates as intermediate phenotypes. OBJECTIVE: This study examines the WISDM PDM as a novel intermediate phenotype of nicotine dependence. METHODS: The study used data from 734 European Americans who smoked at least 5 cigs/day [M = 16.2 (SD = 9.5) cigs/day], completed a phenotypic assessment, and provided a sample of DNA. Based on prior evidence of the role of genetic variation in the NCAM1-TTC12-ANKK1-DRD2 region on chromosome 11q23 in smoking behavior, associations among 12 region loci with nicotine dependence and PDM phenotypes were examined using haplotype and individual loci approaches. In addition, mediational analysis tested the indirect pathway from genetic variation to smoking motives to nicotine dependence. RESULTS: NCAM1-TTC12-ANKK1-DRD2 region loci and haplotypes were significantly associated with the motive of Automaticity and, further, Automaticity significantly mediated associations among NCAM1-TTC12-ANKK1-DRD2 cluster variants and nicotine dependence. CONCLUSIONS: These results suggest that motives related to automaticity are a viable intermediate phenotype for understanding genetic contributions to nicotine dependence. Further, NCAM1-TTC12-ANKK1-DRD2 variants may increase the likelihood that a person will become dependent via a highly automatic smoking ritual that can be elicited with little awareness.
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Antígeno CD56/genética , Motivação/genética , Fenótipo , Proteínas Serina-Treonina Quinases/genética , Proteínas/genética , Receptores de Dopamina D2/genética , Tabagismo/genética , Adolescente , Adulto , Feminino , Variação Genética/genética , Humanos , Masculino , Polimorfismo de Nucleotídeo Único/genética , Fumar/epidemiologia , Fumar/genética , Tabagismo/diagnóstico , Tabagismo/epidemiologia , População Branca/genética , Adulto JovemRESUMO
OBJECTIVE: To describe veterinarians' treatment recommendations and decision-making factors for dogs with cranial cruciate ligament disease (CCLD). METHODS: An online survey of American College of Veterinary Surgeons (ACVS)-Diplomates (surgeon group) and primary care veterinarians (practitioner group) was performed. The survey included questions on treatment recommendations for common case scenarios (small or large breed dog with complete or partial CCLD), treatment decision factors, non-surgical treatment options, and actual treatment, if any, provided for a client-owned dog as well as one owned by their family or close friend. RESULTS: The response rate was 42% for the surgeon group (n = 305/723) and four percent for the practitioner group (n = 1145/ 27,771). Extracapsular stabilization (ES) was the most common treatment recommendation for CCLD in small (9.1 kg) breed dogs amongst surgeons and practitioners. Tibial plateau levelling osteotomy (TPLO) was the most common treatment recommendation for CCLD in large (27.2 kg) breed dogs amongst both groups. The two most important treatment decision factors were dog size (78% of practitioners, 69% of surgeons) and activity level (63% of practitioners, 52% of surgeons). The most common treatment provided for a dog of their own or close relation in the surgeon group was TPLO (64%) followed by ES (15%), whereas in the practitioner group it was ES (38%) followed by TPLO (30%). CLINICAL SIGNIFICANCE: Extracapsular stabilization and TPLO are the most commonly employed surgical procedures in the surveyed population; dog size and activity level (but not age) are the major factors influencing treatment decisions.
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Ligamento Cruzado Anterior , Doenças do Cão/terapia , Cirurgia Veterinária/estatística & dados numéricos , Medicina Veterinária/estatística & dados numéricos , Animais , Ligamento Cruzado Anterior/cirurgia , Coleta de Dados , Doenças do Cão/cirurgia , Cães , Cirurgia Veterinária/métodos , Estados Unidos , Medicina Veterinária/métodosRESUMO
This study investigated the stability of genetic and environmental effects on the common liability to alcohol, tobacco, and cannabis dependence across adolescence and young adulthood. DSM-IV symptom counts from 2,361 adolescents were obtained using a structured diagnostic interview. Several sex-limited longitudinal common pathway models were used to examine gender differences in the magnitude of additive genetic (A), shared environment, and non-shared environmental effects over time. Model fitting indicated limited gender differences. Among older adolescents (i.e., age > 14), the heritability of the latent trait was estimated at 0.43 (0.05, 0.94) during the first wave and 0.63 (0.21, 0.83) during the second wave of assessment. A common genetic factor could account for genetic influences at both assessments, as well as the majority of the stability of SAV over time [rA = 1.00 (0.55, 1.00)]. These results suggest that early genetic factors continue to play a key role at later developmental stages.
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Alcoolismo/genética , Abuso de Maconha/genética , Fumar/genética , Meio Social , Adolescente , Criança , Manual Diagnóstico e Estatístico de Transtornos Mentais , Feminino , Humanos , Masculino , Fenótipo , Adulto JovemRESUMO
BACKGROUND: Personalized treatment for psychopathologies, in particular alcoholism, is highly dependent upon our ability to identify patterns of genetic and environmental effects that influence a person's risk. Unfortunately, array-based whole genome investigations into heritable factors that explain why one person becomes dependent upon alcohol and another does not, have indicated that alcohol's genetic architecture is highly complex. That said, uncovering and interpreting the missing heritability in alcohol genetics research has become all the more important, especially since the problem may extend to our inability to model the cumulative and combinatorial relationships between common and rare genetic variants. As numerous studies begin to illustrate the dependency of alcohol pharmacotherapies on an individual's genotype, the field is further challenged to identify new ways to transcend agnostic genomewide association approaches. We discuss insights from genetic studies of alcohol related diseases, as well as issues surrounding alcohol's genetic complexity and etiological heterogeneity. Finally, we describe the need for innovative systems-based approaches (systems genetics) that can provide additional statistical power that can enhance future gene-finding strategies and help to identify heretofore-unrealized mechanisms that may provide new targets for prevention/treatments efforts. Emerging evidence from early studies suggest that systems genetics has the potential to organize our neurological, pharmacological, and genetic understanding of alcohol dependence into a biologically plausible framework that represents how perturbations across evolutionarily robust biological systems determine susceptibility to alcohol dependence.
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Alcoolismo/genética , Alcoolismo/epidemiologia , Alcoolismo/psicologia , Epistasia Genética , Predisposição Genética para Doença , Variação Genética , Estudo de Associação Genômica Ampla , Humanos , FenótipoRESUMO
BACKGROUND: Acne vulgaris is a disorder of the sebaceous follicles. Propionibacterium acnes can be involved in inflammatory acne. OBJECTIVES: This case-control study aimed at investigating the occurrence and localization of P. acnes in facial biopsies in acne and to characterize the P. acnes phylotype in skin compartments. METHODS: Specific monoclonal and polyclonal antibodies were applied to skin biopsies of 38 patients with acne and matching controls to localize and characterize P. acnes and to determine expression of co-haemolysin CAMP factor, a putative virulence determinant. RESULTS: Follicular P. acnes was demonstrated in 18 (47%) samples from patients with acne and eight (21%) control samples [odds ratio (OR) 3·37, 95% confidence interval (CI) 1·23-9·23; P = 0·017]. In 14 (37%) samples from patients with acne, P. acnes was visualized in large macrocolonies/biofilms in sebaceous follicles compared with only five (13%) control samples (OR 3·85, 95% CI 1·22-12·14; P = 0·021). Macrocolonies/biofilms consisting of mixed P. acnes phylotypes expressing CAMP1 were detected in both case and control samples. Only four samples tested positive for the presence of Staphylococcus spp. and fungi were not observed. CONCLUSIONS: We have for the first time visualized different P. acnes phylotypes in macrocolonies/biofilms in sebaceous follicles of skin biopsies. Our results support the hypothesis that P. acnes can play a role in the pathogenesis of acne as acne samples showed a higher prevalence of follicular P. acnes colonization, both in terms of follicles containing P. acnes and the greater numbers of bacteria in macrocolonies/biofilms than in control samples.
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Acne Vulgar/microbiologia , Biofilmes/crescimento & desenvolvimento , Propionibacterium acnes/fisiologia , Pele/microbiologia , Adolescente , Adulto , Biópsia/métodos , Estudos de Casos e Controles , Feminino , Humanos , Masculino , Microscopia Confocal , Pessoa de Meia-Idade , Fenótipo , Adulto JovemRESUMO
Neuroblastoma is a neural crest-derived embryonal tumour of the postganglionic sympathetic nervous system and a disease with several different chromosomal gains and losses, which include MYCN-amplified neuroblastoma on chromosome 2, deletions of parts of the chromosomes 1p and 11q, gain of parts of 17q and triploidy. Recently, activating mutations of the ALK (Anaplastic Lymphoma Kinase) RTK (Receptor Tyrosine Kinase) gene have been described in neuroblastoma. A meta-analysis of neuroblastoma cases revealed that ALK mutations (49 of 709 cases) in relation to genomic subtype were most frequently observed in MYCN amplified tumours (8.9%), correlating with a poor clinical outcome. MYCN proteins target proliferation and apoptotic pathways, and have an important role in the progression of neuroblastoma. Here, we show that both wild-type and gain-of-function mutants in ALK are able to stimulate transcription at the MYCN promoter and initiate mRNA transcription of the MYCN gene in both neuronal and neuroblastoma cell lines. Further, this stimulation of MYCN gene transcription and de novo MYCN protein expression is abrogated by specific ALK inhibitors, such as crizotinib (PF-2341066), NVP-TAE684, and by small interfering RNA to ALK resulting in a decrease in proliferation rate. Finally, co-transfection of ALK gain-of-function mutations together with MYCN leads to an increase in transformation potential. Taken together, our results indicate that ALK signalling regulates initiation of transcription of the MYCN gene providing a possible explanation for the poor clinical outcome observed when MYCN is amplified together with activated ALK.
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Neuroblastoma/genética , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteínas Oncogênicas/genética , Proteínas Oncogênicas/metabolismo , Receptores Proteína Tirosina Quinases/genética , Receptores Proteína Tirosina Quinases/metabolismo , Quinase do Linfoma Anaplásico , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Transformação Celular Neoplásica/efeitos dos fármacos , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Cicloeximida/farmacologia , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Humanos , Camundongos , Mutação/genética , Proteína Proto-Oncogênica N-Myc , Células NIH 3T3 , Neuroblastoma/tratamento farmacológico , Neuroblastoma/metabolismo , Neuroblastoma/patologia , Células PC12 , Regiões Promotoras Genéticas/efeitos dos fármacos , Biossíntese de Proteínas/efeitos dos fármacos , Biossíntese de Proteínas/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Transcrição Gênica/efeitos dos fármacosRESUMO
Anaplastic lymphoma kinase (ALK) is a receptor tyrosine kinase (RTK), which is transiently expressed during development of the central and peripheral nervous system. ALK has been recently identified as a major neuroblastoma predisposition gene and activating mutations have also been identified in a subset of sporadic neuroblastoma tumors. Two hot spots of ALK mutations have been observed at positions F1174 and R1275. Here, we studied stably transfected cell lines expressing wild-type or F1174L- or R1275Q-mutated ALK in parallel with a neuroblastoma cell line (CLB-GE) in which the allele mutated at position F1174 is amplified. We observed that the mutated ALK variants were essentially intracellular and were largely retained in the reticulum/Golgi compartments. This localization was corroborated by a defect of N-linked glycosylation. Although the mutated receptors exhibited a constitutive activation, the minor pool of receptor addressed to the plasma membrane was much more tyrosine phosphorylated than the intracellular pool. The use of antagonist monoclonal antibodies suggested that the constitutive activity of the mutated receptors did not require the dimerization of the receptor, whereas adequate dimerization triggered by agonist monoclonal antibodies increased this activity. Finally, kinase inactivation of the mutated receptors restored maturation and cell-surface localization. Our results show that constitutive activation of ALK results in its impaired maturation and intracellular retention. Furthermore, they provide a rationale for the potential use of kinase inhibitors and antibodies in ALK-dependent tumors.
Assuntos
Arginina , Mutação , Fenilalanina , Receptores Proteína Tirosina Quinases/química , Receptores Proteína Tirosina Quinases/metabolismo , Quinase do Linfoma Anaplásico , Animais , Linhagem Celular Tumoral , Membrana Celular/enzimologia , Retículo Endoplasmático/enzimologia , Ativação Enzimática , Glicosilação , Complexo de Golgi/enzimologia , Humanos , Camundongos , Peso Molecular , Células NIH 3T3 , Dobramento de Proteína , Transporte Proteico/genética , Receptores Proteína Tirosina Quinases/genéticaRESUMO
Many different types of cancer originate from aberrant signaling from the anaplastic lymphoma kinase (ALK) receptor tyrosine kinase (RTK), arising through different translocation events and overexpression. Further, activating point mutations in the ALK domain have been recently reported in neuroblastoma. To characterize signaling in the context of the full-length receptor, we have examined whether ALK is able to activate Rap1 and contribute to differentiation/proliferation processes. We show that ALK activates Rap1 via the Rap1-specific guanine-nucleotide exchange factor C3G, which binds in a constitutive complex with CrkL to activated ALK. The activation of the C3G/Rap1 pathway results in neurite outgrowth of PC12 cells, which is inhibited by either overexpression of Rap1GAP or siRNA-mediated knockdown of Rap1 itself or the guanine nucleotide exchange factor C3G. Significantly, this pathway also appears to function in the regulation of proliferation of neuroblastoma cells such as SK-N-SH and SH-SY5Y, because abrogation of Rap1 activity by Rap1-specific siRNA or overexpression of Rap1GAP reduces cellular growth. These results suggest that ALK activation of Rap1 may contribute to cell proliferation and oncogenesis of neuroblastoma driven by gain-of-function mutant ALK receptors.
Assuntos
Fator 2 de Liberação do Nucleotídeo Guanina/metabolismo , Neuroblastoma/patologia , Proteínas Tirosina Quinases/metabolismo , Proteínas rap1 de Ligação ao GTP/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Quinase do Linfoma Anaplásico , Animais , Proliferação de Células , Transformação Celular Neoplásica , Ativação Enzimática/efeitos dos fármacos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Proteínas Ativadoras de GTPase/genética , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Fator 2 de Liberação do Nucleotídeo Guanina/deficiência , Fator 2 de Liberação do Nucleotídeo Guanina/genética , Humanos , Camundongos , Neuritos/metabolismo , Neuroblastoma/genética , Proteínas Nucleares/metabolismo , Células PC12 , Inibidores de Proteínas Quinases/farmacologia , Proteínas Tirosina Quinases/antagonistas & inibidores , Pirimidinas/farmacologia , Ratos , Receptores Proteína Tirosina Quinases , Proteínas rap1 de Ligação ao GTP/deficiência , Proteínas rap1 de Ligação ao GTP/genéticaRESUMO
Past studies highlight a narrowing gender gap and the existence of a shared etiology across substances of abuse; however, few have tested developmental models using longitudinal data. We present data on developmental trends of alcohol, tobacco, and marijuana use, abuse and dependence assessed during adolescence and young adulthood in a community-based Colorado twin sample of 1733 respondents through self-report questionnaires and structured psychiatric interviews. Additionally, we report on the rates of multiple substance use and disorders at each developmental stage, and the likelihood of a substance use disorder (SUD; i.e., abuse or dependence) diagnosis in young adulthood based on adolescent drug involvement. Most notably, we evaluate whether the pattern of multiple substance use and disorders and likelihood ratios across substances support a model of generalized risk. Lastly, we evaluate whether the ranked magnitudes of substance-specific risk match the addiction liability ranking. Substance use and SUDs are developmental phenomena, which increase from adolescence to young adulthood with few and inconsistent gender differences. Adolescents and young adults are not specialized users, but rather tend to use or abuse multiple substances increasingly with age. Risk analyses indicated that progression toward a SUD for any substance was increased with prior involvement with any of the three substances during adolescence. Despite the high prevalence of alcohol use, tobacco posed the greatest substance-specific risk for developing subsequent problems. Our data also confirm either a generalized risk or correlated risk factors for early onset substance use and subsequent development of SUDs.
Assuntos
Transtornos Relacionados ao Uso de Substâncias/epidemiologia , Adolescente , Fatores Etários , Consumo de Bebidas Alcoólicas/epidemiologia , Alcoolismo/epidemiologia , Interpretação Estatística de Dados , Etnicidade , Feminino , Humanos , Estudos Longitudinais , Masculino , Abuso de Maconha/epidemiologia , Fumar Maconha/epidemiologia , Modelos Estatísticos , Razão de Chances , Medição de Risco , Fatores de Risco , Fatores Sexuais , Fumar/epidemiologia , Adulto JovemRESUMO
BACKGROUND: Failure of ulcer healing may be critically important to the development of serious gastrointestinal complications in patients on long-term NSAIDs. AIM: To determine the effect of indometacin, celecoxib, a cyclooxygenase-2-specific inhibitor, and nabumetone, a pro-drug, on ulcer healing rates in the rat. METHODS: Gastric ulcers were induced using a cryoprobe. An NSAID or a vehicle control was administered to groups of eight rats for 3 or 6 days (2 mg/kg indometacin, 9 mg/kg celecoxib or 40 mg/kg nabumetone). The ulcer area was measured and epithelial proliferation at the ulcer margins was measured histochemically. The effect of the drugs on intestinal prostaglandin levels was also assessed. RESULTS: The mean ulcer sizes in the four groups on day 3 were comparable. On day 6, control animals and those receiving nabumetone showed significant ulcer healing (P < 0.02), while the mean ulcer sizes in the indometacin (P < 0.01) and celecoxib (P < 0.02) groups were significantly larger than those in the control group. Higher doses of nabumetone (160 mg/kg), however, impaired healing. Intestinal prostaglandins were reduced (P < 0.01) only in indometacin-treated animals. The epithelial proliferation index was significantly lower among indometacin- (P=0.02) and celecoxib-treated (P=0.03) animals compared to controls at day 3. CONCLUSIONS: Celecoxib and indometacin both decreased the epithelial proliferative response and delayed healing of cryoprobe-induced gastric ulcers. In contrast, nabumetone impaired ulcer healing only at very high doses.
Assuntos
Anti-Inflamatórios não Esteroides/farmacologia , Úlcera Gástrica/tratamento farmacológico , Cicatrização/efeitos dos fármacos , Animais , Butanonas/farmacologia , Celecoxib , Divisão Celular/efeitos dos fármacos , Inibidores de Ciclo-Oxigenase/farmacologia , Células Epiteliais/efeitos dos fármacos , Mucosa Gástrica/efeitos dos fármacos , Mucosa Gástrica/patologia , Indometacina/farmacologia , Masculino , Nabumetona , Prostaglandinas/metabolismo , Pirazóis , Ratos , Ratos Sprague-Dawley , Úlcera Gástrica/induzido quimicamente , Úlcera Gástrica/fisiopatologia , Sulfonamidas/farmacologiaAssuntos
Inibidores de Ciclo-Oxigenase/farmacologia , Dinoprostona/biossíntese , Mucosa Gástrica/efeitos dos fármacos , Isoenzimas/efeitos dos fármacos , Lactonas/farmacologia , Prostaglandina-Endoperóxido Sintases/efeitos dos fármacos , Ciclo-Oxigenase 2 , Inibidores de Ciclo-Oxigenase 2 , Mucosa Gástrica/metabolismo , Humanos , Proteínas de Membrana , SulfonasRESUMO
OBJECTIVE: Although Peer Review Organizations (PROs) and researchers rely on physicians to assess quality of care, little is known about what physicians think about when they judge quality. We sought to identify features of individual cases that are associated with physicians' judgments. DESIGN: Using 1994 Medicare data, we selected hospitalizations for 1134 beneficiaries in 42 acute care hospitals in California and Connecticut. The sample was enriched with 17 surgical and six medical complications identified using diagnosis and procedure codes. PRO physicians confirmed quality problems using a structured implicit chart review instrument and provided written open-ended comments about each case. We coded physicians' comments for factors presumed to influence judgments about quality. RESULTS: In crude and adjusted comparisons, reviewers questioned quality more frequently in cases with serious or fatal outcomes, technical mishaps and inadequate documentation. Among surgical (but not medical) patients, they were less likely to record poor quality among patients presenting with an acute illness. CONCLUSION: Factors other than the adequacy of key processes of care are associated with physician-reviewers' judgments about quality.