RESUMEN
The C(17,20)-lyase is a key enzyme in the biosynthesis of androgens by both the testes and adrenals. A complete inhibition of this enzyme would provide an alternative means of androgen suppression for the treatment of prostatic cancers. In the present study, the inhibitory effects of new non-steroidal compounds were tested in vitro on rat C(17,20)-lyase versus abiraterone, a reference steroidal inhibitor. Their activities were also evaluated in vivo on plasma testosterone (T) and luteinizing hormone (LH) levels and on testes, adrenals, seminal vesicles (SV) and ventral prostate (VP) weights after 3 days of oral treatment to adult male rats (50mg/kg per day p.o.). Inhibition in the nanomolar range was obtained with TX 977, the lead racemate product in this series, and optimization is ongoing based on a slight dissociation observed between its two diastereoisomers, TX 1196-11 (S) and TX 1197-11 (R). These non-steroidal compounds (including YM 55208, a reference competitor) proved to be more active in vivo than abiraterone acetate in this model, but the observed impact on adrenal weight suggests that the specificity of lyase inhibition versus corticosteroid biosynthesis deserves further investigations with this new class of potentially useful agents for the treatment of androgen-dependent prostate cancer.
Asunto(s)
Inhibidores Enzimáticos/farmacología , Esteroide 17-alfa-Hidroxilasa/antagonistas & inhibidores , Administración Oral , Androstenos , Androstenoles/farmacología , Animales , Inhibidores Enzimáticos/química , Concentración 50 Inhibidora , Hormona Luteinizante/sangre , Masculino , Microsomas/enzimología , Tamaño de los Órganos , Ratas , Ratas Wistar , Estereoisomerismo , Testículo/enzimología , Testosterona/sangreRESUMEN
A prenatal tumor located in the lumbar paravertebral area was discovered during a routine ultrasound examination at 32 weeks of pregnancy and surgically removed at 4 months of life. The histopathological diagnosis was first suggested to be an infantile desmoid fibromatosis. The tumor karyotype showed a three-way translocation involving both chromosomes 2 and a chromosome 11, t(2;11;2)(p23;p15;q31). Fluorescence in situ hybridization with a probe flanking the ALK gene at 2p23 demonstrated a rearrangement, as previously described in inflammatory myofibroblastic tumors (IMTs). In light of the genetic analysis, the histopathological diagnosis was revised to IMT, although inflammatory cells were scarce. IMTs are pseudosarcomatous inflammatory lesions that primarily occur in the soft tissue and viscera of children and young adults. Our report describes for the first time the occurrence of IMT during prenatal life. The ALK rearrangement may represent the molecular definition of a subgroup of mesenchymal tumors, not always with complete morphological features of IMT, similar to the model of EWS rearrangement in the Ewing sarcoma family of tumors.
Asunto(s)
Cromosomas Humanos Par 11/genética , Cromosomas Humanos Par 2/genética , Sondas de ADN , Enfermedades Fetales/genética , Granuloma de Células Plasmáticas/clasificación , Granuloma de Células Plasmáticas/genética , Proteínas Tirosina Quinasas/genética , Translocación Genética/genética , Adulto , Quinasa de Linfoma Anaplásico , Pintura Cromosómica , Femenino , Enfermedades Fetales/clasificación , Enfermedades Fetales/diagnóstico por imagen , Enfermedades Fetales/patología , Granuloma de Células Plasmáticas/diagnóstico por imagen , Granuloma de Células Plasmáticas/patología , Humanos , Hibridación Fluorescente in Situ , Cariotipificación , Embarazo , Proteínas Tirosina Quinasas Receptoras , Ultrasonografía PrenatalRESUMEN
During Drosophila oogenesis, asymmetrically localized Gurken activates the EGF receptor (Egfr) and determines dorsal follicle cell fates. Using a mosaic follicle cell system we have identified a mutation in the D-cbl gene which causes hyperactivation of the Egfr pathway. Cbl proteins are known to downregulate activated receptors. We find that the abnormal Egfr activation is ligand dependent. Our results show that the precise regulation of Egfr activity necessary to establish different follicle cell fates requires two levels of control. The localized ligand Gurken activates Egfr to different levels in different follicle cells. In addition, Egfr activity has to be repressed through the activity of D-cbl to ensure the absence of signaling in the ventral most follicle cells.
Asunto(s)
Tipificación del Cuerpo/genética , Proteínas de Drosophila , Drosophila/embriología , Receptores ErbB/genética , Oogénesis/genética , Proteínas Tirosina Fosfatasas , Proteínas Proto-Oncogénicas/genética , Transducción de Señal/genética , Factor de Crecimiento Transformador alfa , Alelos , Animales , Células Clonales/metabolismo , Análisis Mutacional de ADN , Drosophila/genética , Desarrollo Embrionario y Fetal/genética , Receptores ErbB/metabolismo , Femenino , Regulación Enzimológica de la Expresión Génica/fisiología , Proteínas de Insectos/genética , Proteínas de la Membrana/genética , Mutación/genética , Proteínas del Tejido Nervioso/genética , Folículo Ovárico/citología , Folículo Ovárico/crecimiento & desarrollo , Folículo Ovárico/metabolismo , Óvulo/crecimiento & desarrollo , Embarazo , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas c-cbl , Sulfotransferasas/genética , Factores de Crecimiento Transformadores/genéticaRESUMEN
During Drosophila oogenesis, signaling between the germline and the soma leads to the establishment of polarity in the egg and embryo. This process involves the interaction of gurken (grk), a TGFalpha-like protein, with torpedo (top), the Drosophila EGF receptor (Egfr). In early stage egg chambers, grk RNA is present predominantly along the posterior cortex of the oocyte, and in mid stage egg chambers, the grk transcript becomes tightly localized to the future dorsal anterior corner of the oocyte. This localization of grk RNA restricts the distribution of Gurken protein and is critical in defining both the anterior-posterior and dorsal-ventral axes of the egg. We have determined the genomic sequence of the grk gene. By testing the requirement of various fragments of grk RNA in the localization process, we find localization signals present in both the 5' and 3' regions of the gene. Sequences in the 5' noncoding region allow for accumulation of the transcript within the oocyte in early stage egg chambers, while signals in the coding region and the 3'UTR are necessary for localization in mid to late stage egg chambers. Active translation is not required for localization of the grk RNA. The mechanism of gurken RNA localization, therefore, differs from that of other localized RNAs studied to date.
Asunto(s)
Proteínas de Drosophila , Drosophila/genética , Proteínas de Insectos/genética , Oocitos/fisiología , Oogénesis/genética , Transcripción Genética , Factor de Crecimiento Transformador alfa , Factores de Crecimiento Transformadores/genética , Regiones no Traducidas 3'/genética , Animales , Polaridad Celular , Femenino , Hibridación in Situ , Oocitos/citología , Proteínas Recombinantes de Fusión/biosíntesisRESUMEN
During Drosophila oogenesis, localization of the transforming growth factor alpha (TGFalpha)-like signaling molecule Gurken to the oocyte membrane is required for polarity establishment of the egg and embryo. To test Gurken domain functions, full-length and truncated forms of Gurken were expressed ectopically using the UAS/Gal4 expression system, or in the germline using the endogenous promoter. GrkDeltaC, a deletion of the cytoplasmic domain, localizes to the oocyte membrane and can signal. GrkDeltaTC, which lacks the transmembrane and cytoplasmic domains, retains signaling ability when ectopically expressed in somatic cells. However, in the germline, the GrkDeltaTC protein accumulates throughout the oocyte cytoplasm and cannot signal. In addition, we found that several strong gurken alleles contain point mutations in the transmembrane region. We conclude that secretion of Gurken requires its transmembrane region, and propose a model in which the gene cornichon mediates this process.
Asunto(s)
Membrana Celular/metabolismo , Proteínas de Drosophila , Drosophila/genética , Proteínas de Insectos/genética , Proteínas de Insectos/metabolismo , Oocitos/metabolismo , Factor de Crecimiento Transformador alfa , Factores de Crecimiento Transformadores/genética , Factores de Crecimiento Transformadores/metabolismo , Secuencia de Aminoácidos , Animales , Animales Modificados Genéticamente , Transporte Biológico , Receptores ErbB/metabolismo , Femenino , Datos de Secuencia Molecular , MutaciónRESUMEN
A translocation, t(17;22)(q22;q13), was identified in two cases of dermatofibrosarcoma protuberans (DP). They bring to four the number of DP cases characterized by an identical t(17;22)(q22;q13), which can be considered as a new tumor-associated chromosome rearrangement. To date, this translocation has been found only in DP and its juvenile form, giant-cell fibroblastoma. This finding has two major consequences. First, it casts light on the development and significance in DP of ring chromosomes which consistently harbor sequences derived from chromosomes 17 and 22. Second, the identification of this new chromosome marker, and eventually of the underlying molecular rearrangement, should help to classify DP, a soft-tissue tumor of still uncertain cell origin. In addition, it could be used to differentiate DP from truly benign or malignant entities, in order that this tumor of intermediate malignancy could be adequately managed.
Asunto(s)
Cromosomas Humanos Par 17/genética , Cromosomas Humanos Par 22/genética , Dermatofibrosarcoma/genética , Neoplasias Cutáneas/genética , Translocación Genética , Adolescente , Femenino , Humanos , Hibridación Fluorescente in Situ , Cariotipificación , MasculinoRESUMEN
In Drosophila, the dorsal-ventral polarity of the egg chamber depends on the localization of the oocyte nucleus and the gurken RNA to the dorsal-anterior corner of the oocyte. Gurken protein presumably acts as a ligand for the Drosophila EGF receptor (torpedo/DER) expressed in the somatic follicle cells surrounding the oocyte. cornichon is a gene required in the germline for dorsal-ventral signaling. cornichon, gurken, and torpedo also function in an earlier signaling event that establishes posterior follicle cell fates and specifies the anterior-posterior polarity of the egg chamber. Mutations in all three genes prevent the formation of a correctly polarized microtubule cytoskeleton required for proper localization of the anterior and posterior determinants bicoid and oskar and for the asymmetric positioning of the oocyte nucleus.
Asunto(s)
Proteínas de Drosophila , Drosophila/embriología , Factor de Crecimiento Transformador alfa , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Polaridad Celular/genética , ADN/genética , Drosophila/genética , Drosophila/fisiología , Receptores ErbB/genética , Receptores ErbB/fisiología , Femenino , Genes de Insecto , Hormonas de Insectos/genética , Datos de Secuencia Molecular , Mutación , Oocitos/metabolismo , Oocitos/ultraestructura , Ovario/citología , Ovario/embriología , ARN/genética , ARN/metabolismo , Transducción de Señal , Factores de Crecimiento Transformadores/genéticaRESUMEN
The consistent involvement of the region 12q13-q15 in numerous human tumors speaks in favor of the presence of genes that may contribute to oncogenesis. Mapping genes within this region of chromosome 12 is a necessary step toward the identification of those that play a role in this process. We have undertaken a multiplex analysis using translocation breakpoint mapping to order from the centromere to the telomere a series of 24 loci from the region 12q12-q22. Thirteen adipose tissue tumors with seven different chromosome changes involving the long arm of chromosome 12 (12q) were used. Since most of these loci are genes or anonymous DNA segments largely available to the scientific community, this map should be useful for investigation of genetic disorders associated with chromosome 12q. While these breakpoints were used as natural landmarks to order groups of loci, this work has positioned them more accurately, leading to a better chromosomal definition of the translocations than the one derived from standard cytogenetic studies.
Asunto(s)
Mapeo Cromosómico , Cromosomas Humanos Par 12/ultraestructura , Neoplasias/genética , Translocación Genética , Reordenamiento Génico , Marcadores Genéticos , Humanos , Lipoma/genética , Liposarcoma Mixoide/genética , Mapeo RestrictivoRESUMEN
We have isolated mutants of the yeast Saccharomyces cerevisiae that are defective in localization of nuclear proteins. Chimeric proteins containing the nuclear localization sequence from SV40 large T-antigen fused to the N-terminus of the mitochondrial F1 beta-ATPase are localized to the nucleus. Npl (nuclear protein localization) mutants were isolated by their ability to grow on glycerol as a consequence of no longer exclusively targeting SV40-F1 beta-ATPase to the nucleus. All mutants with defects in localization of nucleolar proteins and histones are temperature sensitive for growth at 36 degrees C. Seven alleles of NPL3 and single alleles of several additional genes were isolated. NPL3 mutants were studied in detail. NPL3 encodes a nuclear protein with an RNA recognition motif and similarities to a family of proteins involved in RNA metabolism. Our genetic analysis indicates that NPL3 is essential for normal cell growth; cells lacking NPL3 are temperature sensitive for growth but do not exhibit a defect in localization of nuclear proteins. Taken together, these results indicate that the mutant forms of Npl3 protein isolated by this procedure are interfering with nuclear protein uptake in a general manner.
Asunto(s)
Proteínas Fúngicas/metabolismo , Mutación , Proteínas Nucleares/metabolismo , Proteínas de Unión al ARN/metabolismo , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae/metabolismo , Secuencia de Aminoácidos , Secuencia de Bases , Transporte Biológico/genética , Western Blotting , División Celular/genética , Clonación Molecular , ADN Recombinante , Proteínas Fúngicas/genética , Genes Fúngicos , Datos de Secuencia Molecular , Proteínas Nucleares/genética , ARN de Hongos/metabolismo , Proteínas de Unión al ARN/genética , Mapeo Restrictivo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/aislamiento & purificación , Homología de Secuencia de Aminoácido , TemperaturaRESUMEN
We have used a new combination of previously-described methods to obtain a 29-fold purification of plasma membranes from Dictyostelium discoideum. In this procedure, the pellet from a cell lysate is centrifuged through a high-pH sucrose gradient and then through a Renografin gradient. Electron microscopy shows that the resultant "Renografin membranes" are essentially homogeneous. As measured by enzymatic marker assays, contamination with mitochondria, lysosomes, and endoplasmic reticulum is minimal. As assayed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), the protein composition of Renografin membranes is similar to that of highly purified membranes isolated using concanavalin A stabilization and detergent extraction. Using Renografin membranes, we have examined developmental changes in the membrane protein composition. In agreement with previous investigations, we observe major changes in lectin-binding glycoproteins and cell-surface-labeled proteins during the first 18 h of D. discoideum development. In contrast to most previous work, which may have employed plasma membranes of lesser purity, we also observe major changes in silver-stained membrane proteins. We conclude that many developmentally regulated proteins, previously thought to be minor membrane constituents, are a larger proportion of the plasma membrane than originally believed. The observed changes in membrane protein composition may correlate with changes in plasma membrane functions during development. For instance, ponticulin, the major salt-sensitive F-actin-binding protein in plasma membranes from vegetative cells, increases at least twofold in plasma membranes during early development when the cells are chemotaxing into large aggregates. The amount of plasma membrane ponticulin then decreases during the pseudoplasmodial stage.