RESUMEN
The ability of certain enzymatic RNA molecules, or ribozymes, to site-specifically cleave other RNA molecules opens new vistas in gene therapy. Ribozymes can be designed to target specifically a particular mRNA and inhibit protein expression, permitting 'anti-gene' therapy. Here, we describe the progress towards developing ribozymes for use in gene therapy applications. Significant advances have been made in understanding ribozyme transcription unit design and the first clinical tests of ribozyme safety in humans are soon to be initiated.
Asunto(s)
Regulación de la Expresión Génica/efectos de los fármacos , Terapia Genética/métodos , ARN Catalítico/farmacología , Secuencia de Bases , Vectores Genéticos , Humanos , Datos de Secuencia Molecular , Conformación de Ácido Nucleico , Procesamiento Postranscripcional del ARN , ARN Catalítico/química , Transcripción GenéticaRESUMEN
Adenovirus (Ad) vectors for gene therapy are made replication defective by deletion of E1 region genes. For isolation, propagation, and large-scale production of such vectors, E1 functions are supplied in trans from a stable cell line. Virtually all Ad vectors used for clinical studies are produced in the 293 cell, a human embryonic kidney cell line expressing E1 functions from an integrated segment of the left end of the Ad type 5 (Ad5) genome. Replication-competent vector variants that have regained E1 sequences have been observed within populations of Ad vectors grown on 293 cells. These replication-competent variants presumably result from recombination between vector and 293 cell Ad5 sequences. We have developed Ad2-based vectors and have characterized at the molecular level examples of replication-competent variants. All such variants analyzed are Ad2-Ad5 chimeras in which the 293 cell Ad5 E1 sequences have become incorporated into the viral genome by legitimate recombination events. A map of Ad5 sequences within the 293 cell genome developed in parallel is consistent with the proposed recombination events. To provide a convenient vector production system that circumvents the generation of replication-competent variants, we have modified the Ad2 vector backbone by deleting or rearranging the protein IX coding region normally present downstream from the E1 region such that the frequency of recombination between vector and 293 cell Ad5 sequences is greatly reduced. Twelve serial passages of an Ad2 vector lacking the protein IX gene were carried out without generating replication-competent variants. In the course of producing and testing more than 30 large-scale preparations of vectors lacking the protein IX gene or having a rearranged protein IX gene, only three examples of replication-competent variants were observed. Use of these genome modifications allows use of conventional 293 cells for production of large-scale preparations of Ad-based vectors lacking replication-competent variants.
Asunto(s)
Proteínas E1 de Adenovirus/genética , Adenovirus Humanos/genética , Proteínas de la Cápside , Cápside/genética , Vectores Genéticos , Recombinación Genética , Adenovirus Humanos/fisiología , Secuencia de Bases , Línea Celular Transformada , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Replicación del ADN , ADN Viral , Variación Genética , Genoma Viral , Células HeLa , Humanos , Datos de Secuencia Molecular , Mapeo Restrictivo , Homología de Secuencia de Ácido Nucleico , Replicación ViralRESUMEN
The human melanoma tumor Ags, MART1 and gp100, are specifically recognized by HLA-A2-restricted CD8+ CTLs derived from melanoma patients and appear to be involved in tumor regression. In order to develop immunizing vectors for the treatment of patients with metastatic melanoma, replication-defective recombinant adenoviruses, Ad2CMV-MART1 and Ad2CMV-gp100, which encode these tumor Ags, have been generated. Infection of non-Ag expressing HLA-A2+ cell lines A375 and MDA-231 with the vectors resulted in recognition by Ag-specific CTLs as demonstrated by specific target cell lysis and release of cytokines, including IFN-gamma, TNF-alpha, and granulocyte-macrophage-CSF. Sodium butyrate and TNF-alpha can further augment adenovirus-mediated transgene expression and increase recognition by specific CTLs. Although adenovirus-infected cells expressed the E3/19K protein at detectable levels, significant reduction of surface MHC class I expression was observed in only 3 of 10 tumor cell lines infected with either Ad2CMV-MART1 or Ad2CMV-gp100. Because of the suspected homology between the human MART1 and gp100 genes and their murine counterparts, we immunized C57BL/6 mice with these recombinant adenoviruses and demonstrated that immunization with Ad2CMV-gp100 could protect mice from murine melanoma B16 challenge administered intradermally. Depletion of CD8+ but not CD4+ T cells in vivo from Ad2CMV-gp100-vaccinated mice eliminated the protective effect. The anti-gp100 T cells induced by Ad2CMV-gp100 vaccinated appeared to be responsible for the protection. Thus, these recombinant adenoviruses encoding tumor Ags may be useful as vaccines to induce specific T cell immunity for cancer therapy.
Asunto(s)
Adenoviridae/genética , Virus Defectuosos/genética , Vectores Genéticos/genética , Inmunoterapia Activa , Melanoma Experimental/prevención & control , Melanoma/terapia , Glicoproteínas de Membrana/inmunología , Proteínas de Neoplasias/inmunología , Proteínas Recombinantes de Fusión/inmunología , Linfocitos T Citotóxicos/inmunología , Vacunas Sintéticas , Proteínas E3 de Adenovirus/fisiología , Animales , Antígenos de Neoplasias , Butiratos/farmacología , Ácido Butírico , Línea Celular , ADN Recombinante , Regulación Viral de la Expresión Génica/efectos de los fármacos , Antígeno HLA-A2/inmunología , Humanos , Epítopos Inmunodominantes/genética , Epítopos Inmunodominantes/inmunología , Linfocitos Infiltrantes de Tumor/inmunología , Antígeno MART-1 , Melanoma/inmunología , Melanoma Experimental/inmunología , Glicoproteínas de Membrana/genética , Ratones , Ratones Endogámicos C57BL , Proteínas de Neoplasias/genética , Trasplante de Neoplasias , Proteínas Recombinantes de Fusión/genética , Factor de Necrosis Tumoral alfa/farmacología , Vacunación/métodos , Vacunas Sintéticas/uso terapéutico , Antígeno gp100 del MelanomaRESUMEN
Adenovirus-mediated gene transfer to retinal cells was evaluated using the replication-defective recombinant adenovirus vector Ad2/CMVlacZ-1 (coding for beta-galactosidase) both in an in vitro murine culture model and in vivo in adult mice. In vitro, no difference in infectability of neuronal and glial cells was observed, and 50% of neurons expressed the exogenous gene at low viral concentration (10 pfu/cell). In vivo, intraocular injection of 3 x 10(6) pfu Ad2/CMVlacZ-1 resulted in expression of the transferred beta-galactosidase gene in retinal pigment epithelium and ganglion cells. These results demonstrate that Ad2/CMVlacZ-1 is an effective vector for gene transfer into retinal cells.
Asunto(s)
Adenoviridae/genética , Técnicas de Transferencia de Gen , Retina/metabolismo , beta-Galactosidasa/genética , Animales , Células Cultivadas , Proteína Ácida Fibrilar de la Glía/análisis , Inmunohistoquímica , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C3H , Proteínas de Neurofilamentos/análisis , Neuroglía/metabolismo , Neuronas/metabolismo , Retina/citologíaRESUMEN
Retroviral vectors were constructed in which the U3 promoter/enhancer of Moloney murine leukemia (Mo-MLV) was replaced by the corresponding region from five related murine retroviruses--AKR murine leukemia virus (AKV), Harvey murine sarcoma virus (HaMSV), myeloproliferative sarcoma virus (MPSV), SL3-3, and the NZB-xenotropic virus (Xeno). In these vectors the chimeric long terminal repeat (chLTR) drives the expression of the chloramphenicol acetyl transferase (CAT) reporter gene that is followed by an internal SV40 virus early region promoter linked to the neomycin phosphotransferase II (NEO) gene. As an initial measure of the relative promoter/enhancer strength of the chLTR vectors, the murine NIH-3T3 cell line and the human JURKAT cell lines were transfected and assayed for CAT reporter activity. Relative to the MoMLV vector, the HaMSV construct was the most active in NIH-3T3 cells whereas the SL3-3 vector displayed the greatest activity in JURKAT cells. Retroviral vector producer cell populations and cell clones were established for each chLTR vector, and all were capable of yielding high vector titers (> 10(5) G418R cfu/ml on NIH-3T3). Supernatant from these cells was used to transduce both mouse and human cell lines and primary cells. In NIH-3T3 cells and two murine fibrosarcoma cell lines, the HaMSV chLTR vector was slightly more active than the MoMLV chLTR vector. In the human HepG2 and HeLa cell lines, the MPSV chLTR vector was the most active. Data from the human JURKAT T-cell line and a T cell line derived from an ADA-deficient severe combined immunodeficiency (SCID) patient demonstrate that the SL3-3 chLTR is the most active in these lymphoid cell lines. The greatest difference in the comparison of the different chLTR vectors was observed in primary human umbilical vein endothelial cells, where the MoMLV vector produced up to 100 times more CAT activity than the SL3-3 vector. These data suggest that the use of specific promoter/enhancer elements may lead to higher levels of gene expression following retroviral-mediated gene transfer into specific cell types and these observations may be useful in the design of human gene therapy experiments.
Asunto(s)
Quimera/genética , Elementos de Facilitación Genéticos , Regulación Viral de la Expresión Génica/fisiología , Vectores Genéticos , Regiones Promotoras Genéticas , Retroviridae/genética , Animales , Secuencia de Bases , Línea Celular , Cloranfenicol O-Acetiltransferasa/metabolismo , Células HeLa , Humanos , Datos de Secuencia Molecular , Secuencias Repetitivas de Ácidos Nucleicos , Transducción GenéticaRESUMEN
Adenovirus vectors are a promising vehicle to deliver cystic fibrosis transmembrane conductance regulator (CFTR) cDNA to airway epithelia. However, the value of adenovirus vectors will depend on the efficiency with which the vector can correct the defective fluid transport that is though to underlie the pathogenesis of the disease. To address the efficiency of gene transfer, we applied adenovirus vectors expressing CFTR (Ad2/CFTR-1) or beta-galactosidase to the mucosal surface of primary cultures of airway epithelial cells grown as polarized epithelial monolayers on permeable filter supports. These conditions provide a model that reproduces the physiology of the airways in vivo. We found that after adding 1 moi Ad2/CFTR-1 to the mucosal surface, cAMP agonists stimulated fluid secretion that was within the range observed in epithelia from normal subjects. When we measured electrolyte transport, we found that as little as 0.1 moi partially restored cAMP-stimulated Cl- secretion, and at 10 moi Cl- secretion was in the normal range. A related vector encoding beta-galactosidase generated activity in approximately 20% of cells at an moi of 1 and 90% of cells at an moi of 10. These data suggest that Ad2/CFTR-1 is very efficient at restoring normal fluid and electrolyte transport to CF airway epithelia. Thus, they suggest that relatively low input doses could be used for gene transfer to CF airway epithelia.
Asunto(s)
Adenoviridae/genética , Cloruros/metabolismo , AMP Cíclico/fisiología , Fibrosis Quística/metabolismo , Técnicas de Transferencia de Gen , Vectores Genéticos , Pulmón/metabolismo , Proteínas de la Membrana/metabolismo , Fibrosis Quística/genética , Fibrosis Quística/terapia , Regulador de Conductancia de Transmembrana de Fibrosis Quística , Epitelio/metabolismo , Humanos , Proteínas de la Membrana/genética , beta-Galactosidasa/metabolismoRESUMEN
Gene therapy for cystic fibrosis (CF) will require the safe transfer of CFTR cDNA to airway epithelia in vivo. We showed previously that a recombinant adenovirus, Ad2/CFTR-1, expresses CFTR in vitro. As adenovirus rarely integrates, treatment will require repeated vector administration. We applied Ad2/CFTR-1 to intrapulmonary airway epithelia of cotton rats and nasal epithelia of Rhesus monkeys. In both species we detected CFTR mRNA and protein after repeated administration and in monkeys, protein was detected six weeks after repeat administration. The vector did not replicate and was rapidly cleared. Despite an antibody response, there was no evidence of a local or systemic inflammatory response after repeat administration. These data indicate that repetitive administration of Ad2/CFTR-1 is both safe and efficacious.
Asunto(s)
ADN Complementario/administración & dosificación , ADN Complementario/genética , Terapia Genética/métodos , Proteínas de la Membrana/genética , Adenoviridae/genética , Animales , Secuencia de Bases , Fibrosis Quística/genética , Fibrosis Quística/terapia , Regulador de Conductancia de Transmembrana de Fibrosis Quística , Cartilla de ADN/genética , Epitelio/metabolismo , Femenino , Expresión Génica , Terapia Genética/efectos adversos , Vectores Genéticos , Humanos , Macaca mulatta , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Sistema Respiratorio/metabolismo , Sistema Respiratorio/patología , Seguridad , SigmodontinaeRESUMEN
To evaluate the potential of direct transfer of cystic fibrosis transmembrane conductance regulator (CFTR) cDNA for the treatment of cystic fibrosis (CF), we administered an E1-deficient adenovirus, encoding CFTR, to a defined area of nasal airway epithelium of three individuals with CF. This treatment corrected the Cl- transport defect that is characteristic of CF-affected epithelia. After treatment, there was a decrease in the elevated basal transepithelial voltage, and the normal response to a cAMP agonist was restored. We found no evidence of viral replication or virus-associated adverse effects, even at the highest dose tested (25 MOI). These data represent a small step in achieving long-term improvement of CF lung function by gene therapy.
Asunto(s)
Cloruros/metabolismo , Fibrosis Quística/terapia , Terapia Genética/métodos , Proteínas de la Membrana/genética , Mucosa Nasal/metabolismo , Adenoviridae/genética , Adulto , Amilorida/farmacología , Transporte Biológico , Fibrosis Quística/genética , Regulador de Conductancia de Transmembrana de Fibrosis Quística , Epitelio/metabolismo , Femenino , Técnicas de Transferencia de Gen , Vectores Genéticos/genética , Humanos , Masculino , Potenciales de la Membrana/efectos de los fármacos , Persona de Mediana Edad , Mucosa Nasal/patología , Seguridad , Terbutalina/farmacologíaRESUMEN
A new adenovirus-based vector (Ad2/CFTR-1) has been constructed in which the cDNA encoding the cystic fibrosis transmembrane conductance regulator (CFTR), the cystic fibrosis (CF) gene product, replaces the early region 1 coding sequences, E1a and E1b. The virus retains the E3 region. Ad2/CFTR-1 and a related construct encoding beta-galactosidase replicate in human 293 cells which provide E1 gene functions in trans. Replication of these recombinant viruses was not detected in a variety of other cells, although very limited viral DNA synthesis and transcription from the E4 and L5 regions could be measured. These E1-deletion vectors were also deficient in cellular transformation, shut-off of host cell protein synthesis, and production of cytopathic effects, even at high multiplicities of infection. Ad2/CFTR-1 produced CFTR protein in a variety of cells including airway epithelia from CF patients. Expression of functional CFTR protein in a CF airway epithelial monolayer was detected by correction of the Cl- transport defect characteristic of CF. Surprisingly low multiplicities of infection (0.1 moi) were sufficient to generate CFTR Cl- current across a CF epithelial monolayer in vitro. These data, together with the lack of obvious toxicity, suggest that Ad2/CFTR-1 should be suitable for CF gene therapy.
Asunto(s)
Adenovirus Humanos/genética , Fibrosis Quística/terapia , Terapia Genética , Vectores Genéticos , Adenovirus Humanos/crecimiento & desarrollo , Adulto , Animales , Northern Blotting , Línea Celular , Canales de Cloruro/metabolismo , Clonación Molecular , Regulador de Conductancia de Transmembrana de Fibrosis Quística , Células HeLa , Humanos , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratas , beta-Galactosidasa/genética , beta-Galactosidasa/metabolismoRESUMEN
Wild type polyomavirus expression is suppressed in embryonal carcinoma (EC) cell lines. This suppression is alleviated when the EC cells are induced to differentiate. Several characterized host range mutants of polyoma overcome suppression and are able to express and replicate in the undifferentiated EC cells. These previously described isolates were obtained by serial passage of a wild type strain through PCC4 or F9 EC lines. We present a new pyPCC4 isolate (LPT) derived without selection in EC cells. Isolates with host range specificity for a given EC line have been reported to share several common rearrangements and features. These features are also observed in LPT. We report a novel feature shared by these mutants, including LPT, capable of expression in the EC cell line PCC4. In 8 of 10 isolates a novel sequence is created within the enhancer region by rearrangement junctions with near perfect homology to the AP-1 core consensus sequence, 'TGACT(C/A)A'. That the precise location of these junctions varies among these isolates suggest a functional role for this conserved sequence. Our goal is to understand the function of various mutations in host range mutants of polyoma. In order to understand the rearrangements necessary for expression and replication of polyoma in PCC4 cells, we have further characterized the limits of the B enhancer in these cells as compared to those described in permissive cell systems. We have been able to locate the origin proximal limit of the B enhancer for replication close to nt 5189 and distinguish it from the origin proximal limit of the B enhancer for transcription near nt 5215. The two B enhancer cores overlap but do not coincide and are conserved in both cell lines.
Asunto(s)
Regulación Neoplásica de la Expresión Génica , Células Madre Neoplásicas , Poliomavirus/genética , Teratoma/genética , Animales , Secuencia de Bases , Secuencia de Consenso , Células Madre de Carcinoma Embrionario , Elementos de Facilitación Genéticos , Ratones , Datos de Secuencia Molecular , Mutagénesis Sitio-Dirigida , Teratoma/microbiología , TransactivadoresRESUMEN
A specific teratogenic response is elicited in the mouse as a result of exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; dioxin). The characteristic spectrum of structural malformations induced in mice following exposure to TCDD and structurally related congeners is highly reproducible and includes both hydronephrosis and cleft palate. In addition, prenatal exposure to TCDD has been shown to induce thymic hypoplasia. These three abnormalities occur at doses well below those producing maternal or embryo/fetal toxicity and are thus among the most sensitive indicators of dioxin toxicity. In all other laboratory species tested, TCDD causes maternal and embryo/fetal toxicity but does not induce a significant increase in the incidence of structural abnormalities even at toxic dose levels. Developmental toxicity occurs in a similar dose range across species; however, mice are particularly susceptible to development of TCDD-induced terata. Recent experiments using an organ culture were an attempt to address the issue of species and organ differences in sensitivity to TCDD. Human palatal shelves examined in this in vitro system were found to approximate the rat in terms of sensitivity for induction of cleft palate. Investigators have suggested that altered regulation of growth factors and their receptors may involve inappropriate proliferation and differentiation of target cells, ultimately producing TCDD-induced terata. Why the teratogenic effects of TCDD are so highly species and tissue specific, and which animal species most accurately predicts the response of the human embryo/fetus, at the levels of exposure experienced by humans, still remains to be clarified.
Asunto(s)
Anomalías Inducidas por Medicamentos/etiología , Dibenzodioxinas Policloradas/toxicidad , Efectos Tardíos de la Exposición Prenatal , Anomalías Inducidas por Medicamentos/embriología , Animales , Embrión de Pollo , Fisura del Paladar/inducido químicamente , Cricetinae , Femenino , Muerte Fetal/inducido químicamente , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Hidronefrosis/inducido químicamente , Macaca mulatta , Ratones , Técnicas de Cultivo de Órganos , Dibenzodioxinas Policloradas/farmacología , Embarazo , Conejos , Ratas , Especificidad de la Especie , Timo/anomalíasRESUMEN
2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is an extremely potent teratogen in mice. Hydronephrosis and cleft palate are the most sensitive measures of teratogenicity in mice following exposure to TCDD and other structurally related polyhalogenated aromatic hydrocarbons. Despite a relatively long half-life, investigators have identified a critical window for the induction of cleft palate in C57BL/6N mice. To characterize the critical period for renal teratogenesis, pregnant C57BL/6N mice were treated once by gavage with 0-24 micrograms TCDD/kg body wt on Gestation Day (GD) 6, 8, 10, 12, or 14. All dams were killed on GD 18, and the fetuses were examined for the presence of hydronephrosis and cleft palate. Maternal liver-to-body weight ratios were significantly elevated above controls on all days, while maternal weight gain was unaffected. Fetal mortality was increased relative to controls only at 24 micrograms TCDD/kg on GD 6. There was no significant difference in fetal body weights between control and TCDD-treated fetuses. The incidence of cleft palate increased in a dose-related fashion from GD 6 to GD 12, and identification of GD 12 as the critical window for induction of clefting of the hard palate was confirmed. Hydronephrosis was observed at all dose levels, regardless of exposure day, and the incidence was close to 100% at 3 micrograms TCDD/kg and higher doses on GD 12 and earlier. At all doses on GD 14, both the incidence and severity of hydronephrosis were decreased relative to all other days. There was a dose-related increase in the severity of the renal lesion on each day, but between GD 6 and 12 severity was constant. Thus, while palatal sensitivity to TCDD increased with gestational age between GD 6 and 12, there was no difference among these days in development of hydronephrosis. The data suggest, however, that on GD 14 the urinary tract may be less sensitive to TCDD.
Asunto(s)
Dioxinas/toxicidad , Hidronefrosis/inducido químicamente , Dibenzodioxinas Policloradas/toxicidad , Animales , Peso Corporal/efectos de los fármacos , Fisura del Paladar/inducido químicamente , Femenino , Feto/efectos de los fármacos , Edad Gestacional , Semivida , Hidronefrosis/congénito , Hidronefrosis/fisiopatología , Cinética , Hígado/efectos de los fármacos , Ratones , Ratones Endogámicos C57BL , Tamaño de los Órganos/efectos de los fármacos , EmbarazoRESUMEN
Fischer 344 rats were exposed acutely to 2,3,4,7,8-pentachlorodibenzofuran (4-PeCDF) during the organogenic period to evaluate its potential as an inducer of teratogenic and embryolethal effects. All dams were treated by gavage with a single dose of 0, 30, 100, or 300 micrograms 4-PeCDF/kg body wt on gestation Day (gd) 8, 10, or 12. An additional treatment group was included on gd 12 and administered 10 micrograms 4-PeCDF/kg body wt po. All animals were killed on gd 20 and maternal and fetal toxicities were assessed. Determination of embryotoxicity involved both soft tissue and skeletal examinations. 4-PeCDF induced a dose-related decrease in corrected maternal weight gain following treatment on gd 8 and 10, as well as resulted in a concomitant increase in the liver/body weight ratios, first evident at 30 micrograms/kg for all 3 days of exposure. The maternal thymus weight decreased relative to body weight compared with those of controls. Embryo-fetal toxicity was evident from the high mortality (greater than 80%) observed at 300 micrograms/kg for all 3 days of exposure. Mean fetal weight, a sensitive indicator of fetal toxicity, decreased compared to that of controls at 30, 100, and 300 micrograms/kg following treatment on either gd 8, 10, or 12.4-PeCDF induced cleft palate in survivors at a dose of 300 micrograms/kg for all 3 days of exposure. In conclusion, 4-PeCDF is maternally and fetally toxic regardless of the gestation day of exposure, but induced terata only at doses where overt maternal and fetal toxicity were observed, in contrast to previously reported studies in the mice where teratogenic effects were observed at nonfetotoxic dose levels. Thus, the mouse may be a more sensitive model for evaluating specific toxic responses induced prenatally following exposure to the structurally related polyhalogenated aromatic hydrocarbons which include the dioxins, furans, biphenyls, and naphthalenes.
Asunto(s)
Benzofuranos/toxicidad , Teratógenos , Animales , Peso Corporal/efectos de los fármacos , Femenino , Feto/efectos de los fármacos , Edad Gestacional , Masculino , Tamaño de los Órganos/efectos de los fármacos , Embarazo , Ratas , Ratas Endogámicas F344RESUMEN
Octachlorodibenzo-p-dioxin (OCDD) is a widespread environmental contaminant which has been reported to be nontoxic after acute administration. The disposition of [14C]OCDD was studied in male Fischer 344 rats in order to better assess the significance of chronic environmental exposure to OCDD. Rats were treated with 50 micrograms OCDD/kg iv, and 50, 500, or 5000 micrograms/kg po and held in individual metabolism cages for 3 days. Additional rats treated iv were held up to 56 days to follow elimination of OCDD-derived radioactivity and to determine terminal tissue distribution. Feces was the major route of elimination after both routes of exposure with little radioactivity ever appearing in the urine. Gastrointestinal absorption was nonlinear between 500 and 5000 micrograms/kg, never exceeding 10% of the administered dose. Liver was the major depot, followed by adipose tissue and skin. No metabolites of OCDD were detected in tissues, bile, or excreta. The whole-body half-life for the elimination of OCDD was between 3 and 5 months. Repeated oral exposure resulted in linear accumulation of OCDD in the tissues. Thus, OCDD, while poorly absorbed, can accumulate upon low-dose, repeated exposure and concentrate in the liver and adipose tissue.
Asunto(s)
Dioxinas/farmacocinética , Dibenzodioxinas Policloradas/farmacocinética , Absorción , Animales , Semivida , Masculino , Dibenzodioxinas Policloradas/análogos & derivados , Ratas , Ratas Endogámicas F344 , Distribución TisularRESUMEN
Octachlorodibenzo-p-dioxin (OCDD) is a ubiquitous environmental pollutant which has been reported to be nontoxic following acute exposure but has recently been shown to accumulate upon repeated exposure. To determine if this accumulation results in toxic effects, male Fischer 344 rats were treated with 50 micrograms/kg [14C]OCDD by gavage for 10, 20, 40, or 65 times (once a day, 5 days/week) and terminated 3 days postexposure. OCDD accumulated linearly with increasing number of doses and the liver was the major depot, while the adipose served as a secondary sink. Hepatic accumulation resulted in alteration of several biochemical parameters. In animals given 65 doses of OCDD, 7-ethoxyresorufin-O-deethylase activity was elevated 40-fold over controls. Total cytochrome P-450 content doubled and exhibited a 2-nm blue-shift in the Soret maximum for the CO-reduced complex. Treatment-related cytoplasmic fatty vacuolization in the liver was observed concomitant with the biochemical alterations. Thus, subchronic exposure to OCDD appears to cause effects similar to those observed following exposure to low levels of TCDD, but is only 1/100-1/1000 as potent. Such a potency, given the persistent environmental levels to which man may be exposed during a lifetime, suggests that OCDD may pose a potential risk to human health.