RESUMO
Small cell lung cancer (SCLC) accounts for 25% of all lung cancers, and is almost uniformly fatal. Unlike other lung cancers, ras mutations have not been reported in SCLC, suggesting that activation of ras-associated signal transduction pathways such as the raf-MEK mitogen-activated protein kinases (MAPK) are associated with biological consequences that are unique from other cancers. The biological effects of raf activation in small cell lung cancer cells was determined by transfecting NCI-H209 or NCI-H510 SCLC cells with a gene encoding a fusion protein consisting of an oncogenic form of human Raf-1 and the hormone binding domain of the estrogen receptor (DeltaRaf-1:ER), which can be activated with estradiol. DeltaRaf-1:ER activation resulted in phosphorylation of MAPK. Activation of this pathway caused a dramatic loss of soft agar cloning ability, suppression of growth capacity, associated with cell accumulation in G1 and G2, and S phase depletion. Raf activation in these SCLC cells was accompanied by a marked induction of the cyclin-dependent kinase (cdk) inhibitor p27(kip1), and a decrease in cdk2 protein kinase activities. Each of these events can be inhibited by pretreatment with the MEK inhibitor PD098059. These data demonstrate that MAPK activation by DeltaRaf-1:ER can activate growth inhibitory pathways leading to cell cycle arrest. These data suggest that raf/MEK/ MAPK pathway activation, rather than inhibition, may be a therapeutic target in SCLC and other neuroendocrine tumors.
Assuntos
Quinases relacionadas a CDC2 e CDC28 , Carcinoma de Células Pequenas/patologia , Proteínas de Ciclo Celular , Neoplasias Pulmonares/patologia , Proteínas Proto-Oncogênicas c-raf/metabolismo , Proteínas Supressoras de Tumor , Proteínas E1A de Adenovirus/farmacologia , Proteína Quinase CDC2/metabolismo , Proteínas Quinases Dependentes de Cálcio-Calmodulina/metabolismo , Ciclo Celular , Divisão Celular , Quinase 2 Dependente de Ciclina , Inibidor de Quinase Dependente de Ciclina p27 , Quinases Ciclina-Dependentes/antagonistas & inibidores , Quinases Ciclina-Dependentes/metabolismo , Ciclinas/metabolismo , Inibidores Enzimáticos/metabolismo , Estradiol/farmacologia , Flavonoides/farmacologia , Humanos , Proteínas Associadas aos Microtúbulos/metabolismo , Quinases de Proteína Quinase Ativadas por Mitógeno , Fosforilação , Proteínas Quinases/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas c-raf/biossíntese , Proteínas Proto-Oncogênicas c-raf/genética , Receptores de Estrogênio/genética , Receptores de Estrogênio/metabolismo , Proteínas Recombinantes de Fusão/biossíntese , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Tamoxifeno/análogos & derivados , Tamoxifeno/farmacologia , Células Tumorais CultivadasRESUMO
Patients with malignant gliomas have a very poor prognosis. To explore a novel and more effective approach for the treatment of malignant gliomas, a strategy that combined tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) gene therapy and radiation treatment (RT) was designed in this study. Plasmid pE4-GFP was constructed by including the radioinducible early growth response gene 1 (Egr-1) promoter, and it yielded the best response with fractionated RT. Plasmid pE4-TRAIL was constructed by including the Egr-1 promoter and evaluated using U251 and U87 glioma cells. In the assay of apoptosis and killing activities, pE4-TRAIL exhibited radioresponse. pE4-TRAIL combined with RT is capable of inducing cell death synergistically. The expression of TRAIL death receptors was evaluated; which may be influenced by RT. Glioma cells with wild-type p53 showed upregulated expression of death receptors, and more synergistic effects on killing activities are expected. pE4-TRAIL was transfected into the subcutaneous U251 glioma cells in nude mice by the in vivo electroporation method. In the mice treated with pE4-TRAIL and RT, apoptotic cells were detected in pathological sections, and a significant difference of tumor volumes was observed when compared with the other groups (P<0.001). Our results indicate that radioresponsive gene therapy may have great potential as a novel therapy because this therapeutic system can be spatially or temporally controlled by exogenous RT and provides specificity and safety.
Assuntos
Neoplasias Encefálicas/radioterapia , Terapia Genética , Ligante Indutor de Apoptose Relacionado a TNF/genética , Animais , Neoplasias Encefálicas/patologia , Neoplasias Encefálicas/terapia , Linhagem Celular Tumoral , Eletroquimioterapia , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos NusRESUMO
The cell cycle dependence of sister chromatid exchange (SCE) induced by topoisomerase II inhibitors was studied in Chinese hamster V79 cells. 4'-(9-Acridinylamino)methansulfon-m-anisidide, which increases the concentration of covalently linked DNA-topoisomerase II complexes (cleavable complexes), induces SCE strongly in only a short period of the cell cycle. The sensitive period was identified an occurring in early to mid-S phase through the use of labeled thymidine incorporation and flow cytometry. Novobiocin, an inhibitor which prevents formation of the cleavable complex, did not induce SCEs in any part of the cell cycle. However, novobiocin did decrease the level of 4'-(9-acridinylamino)methansulfon-m-anisidide-induced SCEs. These results indicate that the cleavable complex may be important in 4'-(9-acridinylamino)methansulfon-m-anisidide-induced SCE.
Assuntos
Amsacrina/farmacologia , Troca de Cromátide Irmã/efeitos dos fármacos , Inibidores da Topoisomerase II , Ciclo Celular/efeitos dos fármacos , Linhagem Celular , DNA/biossíntese , Citometria de Fluxo , Novobiocina/farmacologiaRESUMO
Therapeutic trials were performed on the HepG2 human hepatoblastoma implanted s.c. in the athymic nude mouse. Animals were treated with polyclonal and monoclonal antiferritin and control antibodies labeled with either iodine-131 (131I) or yttrium-90 (90Y). Administration of 400 muCi of 131I-labeled polyclonal antiferritin or 300 muCi of 90Y-labeled polyclonal antiferritin significantly increased survival (P less than 0.001). There were no tumor cures with radiolabeled polyclonal antibody therapy. Animals treated with 200 or 300 muCi of 131I-labeled monoclonal antiferritin (QCI054) did not show increased survival compared to controls. Although 400 muCi of 131I-labeled QCI significantly prolonged survival, treatment resulted in no long-term survivors. Monoclonal antiferritin labeled with 90Y significantly prolonged survival of animals (P less than 0.001) at doses of 100, 200, or 300 muCi compared with untreated controls. Fifty % of the animals treated with 200 muCi and 75% of the animals treated with 300 muCi showed no evidence of disease at 140 days following treatment. Four hundred muCi of 90Y-labeled QCI proved toxic to the animals. Increased survival was accompanied by a decrease in tumor mitotic rate and increase in cellular polymorphism as determined by pathological examination. The radiation dose absorbed in the tumor correlated directly with tumor response following treatment. The absorbed dose in tumors for complete decay of the isotope ranged from 165 and 330 cGy at the periphery and center of small tumors for an administered activity of 200 muCi of 131I-labeled polyclonal antiferritin, to 7,573 and 12,400 cGy for 300 muCi of 90Y-labeled monoclonal antiferritin QCI.
Assuntos
Anticorpos Monoclonais/uso terapêutico , Carcinoma Hepatocelular/radioterapia , Radioisótopos do Iodo/uso terapêutico , Neoplasias Hepáticas/radioterapia , Radioisótopos de Ítrio/uso terapêutico , Animais , Carcinoma Hepatocelular/patologia , Linhagem Celular , Ferritinas/imunologia , Humanos , Imunoterapia , Neoplasias Hepáticas/patologia , Camundongos , Camundongos Nus , Transplante de Neoplasias , Transplante HeterólogoRESUMO
Recent studies documenting the response of several human tumor cell lines to protracted, continuous irradiation and to acutely delivered radiation, suggest that tumor control may be enhanced with a tumor therapy combining external beam fractionated therapy and protracted irradiation from radiolabeled antibodies. We have evaluated the cytotoxic effect of continuous, protracted irradiation (0.005 Gy/hr to 0.50 Gy/hr) and acutely delivered high-dose-rate irradiation (1.0 Gy/min) on monolayer cultures of human tumor cell lines. Colony formation in these studies was analyzed by the seven-parameter simulation model of Dillehay. Additionally, for some cell lines, cultures were challenged during irradiation with an acute dose of 2.5, 5.0, 7.5 or 10.0 Gy at high-dose-rate. Results from these various studies indicate an altered cellular radiosensitivity occurs for some cell lines during protracted irradiation. At least two mechanisms have been identified that underlie this altered radiosensitivity. One mechanism, G2B, associated with redistribution within the cell cycle, has been previously described by Mitchell and others, and is associated with the phenomenon termed "the inverse dose-rate effect". The other mechanism is only observable following a challenge by acute high-dose-rate irradiation. We have termed this phenomenon "protracted-exposure-sensitization." The characteristics of these two mechanisms are discussed from the perspective of clinical therapeutic exploitation of combined external beam fractionated therapy and radiolabeled immunoglobulin therapy.
Assuntos
Tolerância a Radiação , Células Tumorais Cultivadas/efeitos da radiação , Ciclo Celular/efeitos da radiação , Sobrevivência Celular/efeitos da radiação , Humanos , Método de Monte CarloRESUMO
Using 90Yttrium radiolabeled antibodies, radioimmunotherapy was compared to fractionated external beam radiotherapy in the treatment of human glioma xenografts. Antibody treatments required administration of an approximately threefold greater total dose compared to external beam treatments to achieve the same tumor regrowth delay. Following multi-fraction external beam radiation treatments, tumor regrowth delay demonstrated a large fractionation effect (alpha/beta = 2.3 Gy, 95% confidence limits 0.4-4.2 Gy), suggesting that much of the ineffectiveness of the antibody treatments could be caused by a large dose-rate effect in this system. Despite the large fractionation effect, the regrowth delay was small for a large single-fraction external beam irradiation, possibly because of tumor hypoxia. When compared to external beam radiation, radiolabeled antibody treatments resulted in a comparatively diminished tumor bed effect, suggesting radioimmunotherapy spares normal tissue surrounding the tumor.
Assuntos
Glioma/radioterapia , Radioimunoterapia , Radioisótopos de Ítrio/uso terapêutico , Animais , Glioma/patologia , Humanos , Camundongos , Camundongos Nus , Transplante de Neoplasias , Doses de Radiação , Transplante HeterólogoRESUMO
Glioblastoma U251 and U87 cells irradiated with single fraction high dose rate radiation (1.1 Gy/min) were relatively insensitive to inactivation of colony forming ability, similar to other glioblastoma cell lines. Initial rates of cell kill with continuous low dose rate irradiation (0.075 Gy/hr to 0.49 Gy/hr) were low, but at times greater than 20 hours and with dose rates of 0.25 Gy/hr or higher, the rate of cell kill increased. Population doubling times for these cell lines were about 24 hours, suggesting that cell cycle redistribution may be responsible for the increased sensitivity. DNA histograms obtained by flow cytometry support this hypothesis, with cells accumulating in the G2 and M phases of the cell cycle. These results suggest that low dose rate irradiation may be effective in treating glioblastomas. Optimization of time intervals between radiation treatments as well as dose rates used for glioblastoma patients may be influenced by these findings, resulting in better integration of continuous low-dose-rate irradiation (radioactive antibodies and implants) and high-dose-rate irradiation (fractionated external beam) into therapeutic programs.
Assuntos
Glioma/radioterapia , Ciclo Celular/efeitos da radiação , Linhagem Celular , Sobrevivência Celular/efeitos da radiação , Relação Dose-Resposta à Radiação , Glioma/patologia , Humanos , Técnicas In Vitro , Dosagem Radioterapêutica , Fatores de TempoRESUMO
PURPOSE: Recently, polymeric controlled delivery of chemotherapy has been shown to improve survival of patients with malignant glioma. We evaluated whether we could similarly deliver halogenated pyrimidines to experimental intracranial human malignant glioma. To address this issue we studied the in vitro release from polymers and the in vivo drug delivery of IUdR to experimental human U251 glioblastoma xenografts. METHODS AND MATERIALS: In vitro: To measure release, increasing (10%, 30%, 50%) proportions of IUdR in synthetic [(poly(bis(p-carboxyphenoxy)-propane) (PCPP):sebacic acid (SA) polymer discs were serially incubated in buffered saline and the supernatant fractions were assayed. In vivo: To compare local versus systemic delivery, mice bearing flank xenografts had intratumoral or contralateral flank IUdR polymer (50% loading) treatments. Mice bearing intracranial (i.c.) xenografts had i.c. versus flank IUdR polymer treatments. Four or 8 days after implantation of polymers, mice were sacrificed and the percentage tumor cells that were labeled with IUdR was measured using quantitative microscopic immunohistochemistry. RESULTS: In vitro: Increasing percentage loadings of IUdR resulted in higher percentages of release: 43.7 + 0.1, 70.0 + 0.2, and 90.2 + 0.2 (p < 0.001 ANOVA) for the 10%, 30%, and 50% loadings, respectively. In vivo: For the flank tumors, both the ipsilateral and contralateral IUdR polymers resulted in similarly high percentages labeling of the tumors versus time. For the ipsilateral IUdR polymers, the percentage of tumor cellular labeling after 4 days versus 8 days was 45.8 +/- 7.0 versus 40.6 +/- 3.9 (p = NS). For the contralateral polymer implants, the percentage of tumor cellular labeling were 43.9 +/- 10.1 versus 35.9 +/- 5.2 (p = NS) measured 4 days versus 8 days after implantation. For the i.c. tumors treated with extracranial IUdR polymers, the percentage of tumor cellular labeling was low: 13.9 +/- 8.8 and 11.2 +/- 5.7 measured 4 and 8 days after implantation. For the i.c. tumors having the i.c. IUdR polymers, however, the percentage labeling was comparatively much higher: 34.3 +/- 4.9 and 35.3 +/- 4.0 on days 4 and 8, respectively. For the i.c. tumors, examination of the percentage cellular labeling versus distance from the implanted IUdR polymer showed that labeling was highest closest to the polymer disc. CONCLUSION: Synthetic, implantable biodegradable polymers provide the local, controlled release of IUdR and result in the high, local delivery of IUdR to experimental intracranial human malignant glioma. This technique holds promise for the local delivery of IUdR for radiosensitization of human brain tumors.
Assuntos
Antimetabólitos/administração & dosagem , Neoplasias Encefálicas/tratamento farmacológico , Glioblastoma/tratamento farmacológico , Idoxuridina/administração & dosagem , Polímeros , Animais , Antimetabólitos/metabolismo , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Divisão Celular , Preparações de Ação Retardada , Portadores de Fármacos , Glioblastoma/metabolismo , Glioblastoma/patologia , Humanos , Idoxuridina/metabolismo , Masculino , Camundongos , Camundongos Nus , Fatores de Tempo , Transplante Heterólogo , Células Tumorais CultivadasRESUMO
PURPOSE: Low-dose-rate radiation therapy has been widely used in the treatment of urogenital malignancies. When continuously exposed to low-dose-rate ionizing radiation, target cancer cells typically exhibit abnormalities in replicative cell-cycle progression. Cancer cells that arrest in the G2 phase of the cell cycle when irradiated may become exquisitely sensitive to killing by further low-dose-rate radiation treatment. Oncogenic human papillomaviruses (HPVs), which play a major role in the pathogenesis of uterine cervix cancers and other urogenital cancers, encode E6 and E7 transforming proteins known to abrogate a p53-dependent G1 cell-cycle checkpoint activated by conventional acute-dose radiation exposure. This study examined whether expression of HPV E6 and E7 oncoproteins by cancer cells alters the cell-cycle redistribution patterns accompanying low-dose-rate radiation treatment, and whether such alterations in cell-cycle redistribution affect cancer cell killing. METHODS AND MATERIALS: RKO carcinoma cells, which contain wild-type P53 alleles, and RKO cell sublines genetically engineered to express HPV E6 and E7 oncoproteins, were treated with low-dose-rate (0.25-Gy/h) radiation and then assessed for p53 and p21WAF1/CIP1 polypeptide induction by immunoblot analysis, for cell-cycle redistribution by flow cytometry, and for cytotoxicity by clonogenic survival assay. RESULTS: Low-dose-rate radiation of RKO carcinoma cells triggered p53 polypeptide elevations, p21WAF1/CIP1 induction, and arrest in the G1 and G2 phases of the cell cycle. In contrast, RKO cells expressing E6 and E7 transforming proteins from high-risk oncogenic HPVs (HPV 16) arrested in G2, but failed to arrest in G1, when treated with low-dose-rate ionizing radiation. Abrogation of the G1 cell-cycle checkpoint activated by low-dose-rate radiation exposure appeared to be a characteristic feature of transforming proteins from high-risk oncogenic HPVs: RKO cells expressing E6 from a low-risk nononcogenic HPV (HPV 11) exposed to low-dose-rate radiation arrested in both G1 and G2. Surprisingly, despite differences in cell-cycle redistribution accompanying low-dose-rate radiation treatment associated with high-risk HPV transforming protein expression, no consistent differences in clonogenic survival following low-dose-rate radiation treatment were found for RKO cell sublines expressing high-risk HPV oncoproteins and arresting only in G2 during low-dose-rate radiation exposure vs. RKO cell sublines exhibiting both G1 and G2 cell-cycle arrest when irradiated. CONCLUSION: The results of this study demonstrate that neither HPV oncoprotein expression nor loss of the radiation-activated G1 cell-cycle checkpoint alter the sensitivity of RKO carcinoma cell lines to low-dose-rate radiation exposure in vitro. Perhaps for urogenital malignancies associated with oncogenic HPVs in vivo, HPV oncoprotein-mediated abrogation of the G1 cell-cycle checkpoint may not limit the potential efficacy of low-dose-rate radiation therapy.
Assuntos
Ciclinas/metabolismo , Fase G1/efeitos da radiação , Fase G2/efeitos da radiação , Proteínas Oncogênicas Virais/metabolismo , Proteínas Repressoras , Proteína Supressora de Tumor p53/metabolismo , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Inibidor de Quinase Dependente de Ciclina p21 , Ciclinas/efeitos da radiação , Dano ao DNA , Humanos , Proteínas E7 de Papillomavirus , Doses de Radiação , Tolerância a Radiação , Células Tumorais Cultivadas/metabolismo , Células Tumorais Cultivadas/efeitos da radiação , Proteína Supressora de Tumor p53/efeitos da radiaçãoRESUMO
PURPOSE: To describe and evaluate a new, simple, inexpensive method for directly measuring the radiation dose and its spatial distribution generated from explanted tissues of animals previously injected with radiolabeled immunoconjugates or other agents. METHODS AND MATERIALS: This technique uses the newly developed radiochromic dye medium (Gafchromic) which responds reproducibly for therapeutic dose exposures, has high spatial resolution, does not require film processing, and is relatively insensitive to ambient light. We have evaluated the dose distribution from LS174T tumors and selected normal tissues in nude mice previously injected with 90Y labeled anti-carcinoembrionic antigen antibodies. Individual tissues from sacrificed animals are halved and the flat section of the tissue is placed onto the dosimetry media and then frozen. The dosimetry medium is exposed to beta and Bremsstrahlung radiation originating from the frozen tissues. The relative darkening of the dosimetry medium depends on the dose deposited in the film. The dosimetry medium is scanned with a commercial flatbed scanner and the image intensity is digitally stored and quantitatively analyzed. Isodose curves are generated and compared to the actual tissue outline. RESULTS: The absorbed dose distribution due to 90Y exposure show only slight gradients in the interior of the tissue, with a markedly decreasing dose near the edges of the tissue. In addition, the isodose curves follow the tissue outline except in regions having radii of curvature smaller than the range of the beta-particle (R90 = 5 mm). These results suggest that the shape of the tumor, and its curvature, are important in determining the minimum dose delivered to the tumor by radiation from 90Y monoclonal antibodies, and hence in evaluating the tumor response to the radiation. The dose and spatial dose distribution were calculated assuming that the total 90Y activity is distributed uniformly throughout a half ellipsoid. The calculated spatial dose distributions for the half ellipsoids were similar to those observed from the dosimetry media that had been exposed to radioactivity contained in the tumors. CONCLUSION: This method provides direct dose evaluation without elaborate summary calculations based on activity measurements from serial slices. The measured radiation dose actually indicates the dose rate at the time of animal sacrifice. Quantitative analysis of radiation emitted from the tissues is relatively fast, making it feasible to examine a number of tissues under a variety of conditions.
Assuntos
Radioimunoterapia , Dosagem Radioterapêutica , Animais , Humanos , Camundongos , Camundongos Nus , Transplante de Neoplasias , Neoplasias Experimentais/radioterapia , Transplante HeterólogoRESUMO
PURPOSE: To measure, quantify, and evaluate the planar dose-rate distribution for human tumor xenografts implanted into mice that are treated with 90Y-labeled monoclonal antibodies or bispecific antibodies and 90Y-labeled haptens. METHODS AND MATERIALS: Twenty-five LS174T human colon carcinoma tumors grown subcutaneously in nude mice were treated with 90Y by either directly labeled ZCE025 or bispecific ECA001-DBX antibody systems. A simple, quick technique using GAF radiochromic medium determined the dose-rate distribution in a plane passing through the tumor center. The dose-rate distribution is generated from exposure to activity situated in one-half of the tumor (0.045 to 0.83 g). RESULTS: Planar dose-rate distributions were obtained from the tumor xenografts. Planar dose-rate histograms were computed along with the coefficients of variance and skewness of the distributions. The observed dose-rate distributions were quantitatively compared to those calculated for a uniformly distributed activity in a half-ellipsoid of the same volume and approximate shape as the tumor half. The observed dose-rate distributions were usually broader with a more positive coefficient of skewness than the dose-rate distributions calculated from the uniformly active half-ellipsoids. For 90Y, tumor shape plays an important role in determining the minimum tumor dose. For these tumors, the tumor minimum dose-rate is always observed along the edge, usually where the edge curvature is most convex. Larger tumors tended to have broader dose-rate distributions and more positive coefficients of skewness. Exceptions to this trend were associated with dose-rate maxima displaced from the central regions due to activity heterogeneity or tumor size greatly exceeding the range of emission. Calculations for dose rate from the conventional Medical Internal Radiation Dose (MIRD) formulation exceeded the average and minimum dose rate derived from radiochromic media. The coefficient of skewness became more positive for increasing time between injection and tumor excision, consistent with the activity evolving into a more uniform activity distribution. CONCLUSION: Using radiochromic media to measure the spatial dose-rate distribution is a valuable method for comparing the dose-rate heterogeneity among experimental tumor xenografts in animals treated with radiolabeled antibodies. Tumor size (relative to the particle range) and changes in activity distribution radiolabeled antibodies. Tumor size (relative to the particle range) and changes in activity distribution affect the dose-rate distribution that are reflected by changes in the coefficients of skewness and variation of the dose-rate area histogram. The increase in coefficients of variation and skewness with tumor size and time results from the size of the 90Y beta particle penetration range that either exceeds or is comparable to the tumor dimensions. The minimum dose rate is more dependent, relative to the average and the maximum dose rates, on the curvature of the tumor surface.
Assuntos
Anticorpos Biespecíficos/uso terapêutico , Anticorpos Monoclonais/uso terapêutico , Neoplasias do Colo/radioterapia , Radioimunoterapia/métodos , Dosagem Radioterapêutica , Radioisótopos de Ítrio/uso terapêutico , Animais , Humanos , Camundongos , Camundongos Nus , Transplante Heterólogo , Células Tumorais CultivadasRESUMO
PURPOSE: To identify those factors that predict variability in tumor response to 90Y-radioimmunotherapy based on measurement of incorporated activity and physical dimensions of individual tumors and to apply the concept of effective dose to radioimmunotherapy. METHODS AND MATERIALS: Human colon carcinoma xenografts growing in nude mice were treated with anti-CEA antibodies labeled with 90Y directly or through a bispecific antibody/labeled hapten system. Tumor response was measured as the delay in growth to eight times the treatment volume. Noninvasive activity (based on bremsstrahlung radiation) and dimension measurements were made in these animals at several times after label injection. The following parameters were compared for their ability to predict individual tumor response: (a) injected activity, (b) injected activity times a factor based on average uptake as a function of volume, (c) in vivo activity per volume measured in each animal at a single time, (d) the integral over time of in vivo activity per volume in each animal, and (e) the minimum dose for each animal in a uniformly active ellipsoid whose total activity and dimensions varied over time the same as the tumor. RESULTS AND CONCLUSION: After correcting for differences in injected activity, two parameters account for much of the variability in tumor response. One of these is the general trend of larger tumors to take up less activity per volume. Additional variability can be accounted for by the in vivo activity per volume measurements. The minimum dose as introduced here is likely to be useful in estimating the biologically effective dose delivered by each treatment.
Assuntos
Anticorpos Biespecíficos/uso terapêutico , Anticorpos Monoclonais/uso terapêutico , Antígeno Carcinoembrionário/imunologia , Neoplasias do Colo/radioterapia , Radioimunoterapia/métodos , Radioisótopos de Ítrio/uso terapêutico , Animais , Anticorpos Biespecíficos/metabolismo , Anticorpos Monoclonais/metabolismo , Divisão Celular/efeitos da radiação , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Humanos , Camundongos , Camundongos Nus , Eficiência Biológica Relativa , Transplante Heterólogo , Células Tumorais Cultivadas , Radioisótopos de Ítrio/farmacocinéticaRESUMO
Radiolabeled antibodies provide a potential basis for selective radiotherapy of human gliomas. Monoclonal P96.5, a mouse IgG2a immunoglobulin, defines an epitope of a human melanoma cell surface protein and specifically binds the U-251 human glioma as measured by immunoperoxidase histochemistry. 111In-radiolabeled P96.5 specifically targets the U-251 human glioma xenograft and yields 87.0 microCuries (uCi) of tumor activity per gram per 100 uCi injected activity compared to 4.5 uCi following administration of radiolabeled irrelevant monoclonal antibody. Calculations of targeting ratios demonstrate deposited dose to be 11.6 times greater with radiolabeled P96.5 administration compared to irrelevant monoclonal antibody. Tumor dose found in normal organs is less than 20% of the tumor dose, further supporting specific targeting of the human glioma xenograft by this antibody. Monoclonal antibodies QCI054 and ZME018, which define a tumor-associated and a second melanoma-associated antigen, respectively, demonstrate positive immunoperoxidase staining of the tumor, but comparatively decreased targeting. To test the therapeutic potential of 90Y-radiolabeled P96.5, QCI054, and ZME018, tumors and normal sites were implanted with miniature thermoluminescent dosimeters (TLD). Average absorbed doses of 3770 +/- 445 (mean +/- SEM), 2043 +/- 134, and 645 +/- 48 cGy in tumor, 353 +/- 41, 243 +/- 22, and 222 +/- 13 cGy in a contralateral control intramuscular site, 980 +/- 127, 815 +/- 41, and 651 +/- 63 cGy in liver, and 275 +/- 14, 263 +/- 11, and 256 +/- 18 cGy in total body were observed 7 days following administration of 100 uCi 90Y-radiolabeled P96.5, QCI054, and ZME018, respectively. To test the therapeutic potential, tumor-bearing nude mice were given intracardiac injections of either buffer or 90Y-radiolabeled P96.5, QCI054, or ZME018. Striking tumor regression and prolonged survival were measured following administration of 90Y-labeled P96.5. Average maximal decreases in tumor volume were 42.7 +/- 11.9 and 94.2 +/- 3.3 percent 28 and 58 days following 100 and 200 uCi 90Y-radiolabeled P96.5 administration, respectively. The time required to achieve four times the initial tumor volume was 6.1 +/- 0.9 days for buffer; 43 +/- 12 and 63 +/- 10 days for 50 and 100 uCi 90Y-radiolabeled P96.5; 7 +/- 2, 20 +/- 1, and 53 +/- 4 for 50, 100, and 200 uCi 90Y-radiolabeled QCI054; and 9 +/- 1, 13 +/- 1, and 29 +/- 3 days for 50, 100, and 200 uCi 90Y-radiolabeled ZME018, respectively. Average tumor regrowth failed to occur 180 days following administration of 200 uCi 90Y-labeled P96.5.(ABSTRACT TRUNCATED AT 400 WORDS)
Assuntos
Anticorpos Monoclonais/uso terapêutico , Glioblastoma/radioterapia , Radioisótopos de Índio/uso terapêutico , Radioisótopos de Ítrio/uso terapêutico , Animais , Glioblastoma/química , Glioblastoma/mortalidade , Humanos , Imuno-Histoquímica , Radioisótopos de Índio/administração & dosagem , Masculino , Camundongos , Camundongos Nus , Transplante de Neoplasias , Doses de Radiação , Taxa de Sobrevida , Transplante Heterólogo , Radioisótopos de Ítrio/administração & dosagemRESUMO
A computer model that simulates the killing of exponentially growing cells by low-dose-rate radiation is described. The model incorporates cell killing by single-hit damage and double-hit (sublethal) damage, as well as repair of sublethal damage, delay of cell cycle progression, blockage and increased sensitivity of cells in the G2 phase of the cell cycle, and cell division. Seven cellular parameters determine the rate of cell killing. Initial estimates of most of these parameters can be made from independent experiments. Parameters were obtained that gave the best fit to the data for four cell lines, using constant or variable dose rates, and using as end points either the fraction of single cells forming colonies or the total number of clonogenic cells in a mass culture. Some of the parameters were determined to be insignificant or similar for the four cell lines. The main differences between the cell lines in patterns of cell killing over a range of dose rates appeared to be determined by differences in the values of four of the parameters.
Assuntos
Divisão Celular/efeitos da radiação , Sobrevivência Celular/efeitos da radiação , Simulação por Computador , Reparo do DNA , Interfase/efeitos da radiação , Modelos Biológicos , Doses de RadiaçãoRESUMO
The ability of mutagens to transform benign papillomas to malignancy in the mouse skin model of multistage carcinogenesis [Hennings et al. Nature 303, 67-68 (1983)] suggests that multiple events may underlie carcinogenic progression, and that mutagenic exposures separated by time can act synergistically. Such synergism may result from initial mutagenic exposure which induces heritable sensitivity to subsequent mutagenic exposures. For example, progeny of X-irradiated V79 cells are hypersensitive to subsequent mutation induced by psoralen plus long-wave ultraviolet light, PUVA [Frank and Williams, Science 216, 307-308 (1982)]. In the present studies 100 to 200 surviving clones of short-wave ultraviolet light (UVC) irradiated V79 cells were assayed for mutation at two loci. Cultures derived from these cells were found to be hypermutable at the hypoxanthine guanine phosphoribosyl transferase (HGPRT) locus following exposure to PUVA, but showed mutant frequencies similar to control cells following UVC challenge at the HGPRT and ATPase loci.
Assuntos
Mutação , Radiogenética , Raios Ultravioleta , Adenosina Trifosfatases/genética , Animais , Linhagem Celular , Hipoxantina Fosforribosiltransferase/genéticaRESUMO
High interstitial pressure limits the uptake of systemically-administered macromolecular agents. Fast infusion, which has been found to spread by convection macromolecules in the brain, was examined in a xenograft tumor system. Subcutaneous human U251 glioblastoma tumors (0.3-1.3 cm3) were infused for up to 40 minutes starting at 20 microliters/minutes while line pressure was recorded. The spread of blue dextran (molecular weight 2 x 10(6) was examined in excised tumors. Resistance to infusion decreased with time so that the infusion rate could be increased without an increase in line pressure. Blue dextran was spread up to the length of the tumor (maximum of 1.5 cm), but the spreading appeared to be asymmetric. The results suggest the pressures produced by the infusion dilated the tumor tissue, thus producing increased hydraulic conductivity. Although this produces rapid convective spread, the spread is asymmetric. Possible methods for obtaining a more uniform or controlled convective spread of macromolecular agents are discussed.
Assuntos
Neoplasias Cutâneas/metabolismo , Animais , Glioblastoma/metabolismo , Humanos , Camundongos , Camundongos Nus , Transplante de Neoplasias , Pressão , Transplante HeterólogoRESUMO
The alkaline elution of bromodeoxyuridine-containing (BrdUrd) DNA and chlorodeoxyuridine-containing ( CldUrd ) DNA was studied in two CHO lines, the parental AA8 and a mutant line, EM9 , which has a defect in repairing strand breaks and a 12-fold elevated baseline frequency of SCE. BrdUrd-DNA was found to have alkali-labile sites as well as direct breaks, neither of which were increased significantly by prior treatment of AA8 cells with an inhibitor (benzamide) or poly(adenosine diphosphoribose) polymerase. CldUrd -DNA, which gives higher frequencies of SCEs than BrdUrd-DNA, had more strand breaks than BrdUrd-DNA in AA8 cells after treatment with benzamide, while without benzamide there was no difference. The accumulation of breaks in CldUrd -DNA by benzamide was shown to occur rapidly, to reach a maximum by 90 min, and to be readily reversible after benzamide removal. Under all conditions, EM9 cells had more strand breaks than AA8 . These observed differences in strand breaks were not due to differences in incorporation efficiencies. For the different halogenated pyrimidines and cell types, there was a good correlation between the number of strand breaks and reduction in plating efficiencies.
Assuntos
Bromodesoxiuridina/toxicidade , Replicação do DNA/efeitos dos fármacos , Desoxiuridina/análogos & derivados , Animais , Benzamidas/toxicidade , Bromodesoxiuridina/metabolismo , Linhagem Celular , Cricetinae , Cricetulus , Reparo do DNA , Desoxiuridina/metabolismo , Desoxiuridina/toxicidade , Feminino , Cinética , Mutação , Ovário , Troca de Cromátide Irmã/efeitos dos fármacosRESUMO
Pommier et al. (1985) suggested that sister-chromatid exchange (SCE) results from exchange of topoisomerase II subunits. "Homologous displacement", an alternative mechanism, is proposed in which strand switching occurs during removal of parental helical turns by topoisomerases. The steps in the SCE model proposed by Ishii and Bender (1980) for SCE occurring at a blocked replication fork could occur by this mechanism and would require the action of both topoisomerases I and II. Homologous displacement involving topoisomerase II alone provides a mechanism for the strand switching required in the models of Kato (1977) and Cleaver (1981) in which SCE occur between replicated double strands. These mechanisms and models are discussed in relation to current knowledge of the locations and functions of topoisomerases during DNA replication.
Assuntos
Replicação do DNA , DNA Topoisomerases Tipo II/metabolismo , DNA Topoisomerases Tipo I/metabolismo , Modelos Genéticos , Troca de Cromátide Irmã , RepliconRESUMO
The mutant Chinese hamster cell line, EM9, is characterized by a high baseline sister-chromatid exchange (SCE) frequency, increased sensitivity to cell killing, and a defect in DNA strand-break repair. The molecular basis for this pleotrophic phenotype is not known. We examined, at the chromosomal level, the increased sensitivity of this mutant to incorporated BrdUrd. By varying the amount of BrdUrd in template DNA and measuring the frequency of SCEs and chromosomal aberrations, we demonstrated the enhanced sensitivity of EM9 to BrdUrd present in the template strand of DNA. Our results show that a 6-fold increase in SCEs occurs due to DNA replication over a BrdUrd-substituted template relative to a dThd-substituted template. With regard to aberration production in EM9, there is a significant enhancement of aberrations and a specific bias toward damage for the chromatid with Brdurd in the template strand. While these cells share some phenotypic properties with cells from patients with Bloom's syndrome, the genotypic similarities have not yet been established.
Assuntos
Bromodesoxiuridina/toxicidade , Aberrações Cromossômicas/efeitos dos fármacos , Metanossulfonato de Etila/toxicidade , Troca de Cromátide Irmã/efeitos dos fármacos , Animais , Linhagem Celular , Deleção Cromossômica/efeitos dos fármacos , Cricetinae , Replicação do DNA , Sinergismo FarmacológicoRESUMO
The majority of the high (12-fold elevated) baseline sister-chromatid exchanges (SCEs) that occur in the CHO mutant line EM9 appear to be a consequence of incorporated BrdUrd, and they arise during replication of DNA containing BrdUrd in a template strand. In normal CHO cells the alkaline elution patterns of DNA newly replicated on a BrdUrd-containing template are significantly altered compared with those seen during the replication on an unsubstituted template. The nascent DNA synthesized on such an altered template is delayed in reaching mature size, possibly because replication forks are temporarily blocked at sites occurring randomly along the template. Transient blockage of replication forks may be a prerequisite for SCE. The delay in replication on BrdUrd-substituted templates was greater in EM9 cells than in parental AA8 cells and was also greater in AA8 cells treated with benzamide, an inhibitor of poly(ADPR) polymerase, than in untreated AA8 cells. Under these conditions, treatment with benzamide also produced a 7-fold increase in SCEs in AA8. An EM9-derived revertant line that has a low baseline SCE frequency showed less delay in replication on BrdUrd-substituted templates than did EM9. However, under conditions where the template strand contained CldUrd, which was shown to produce 4-fold more SCEs than BrdUrd in AA8 cells, the replication delay in AA8 was not any greater in the CldUrd-substituted cells. Thus, other factors besides the delay appear to be involved in the production of SCEs by the template lesions resulting from incorporation of the halogen-substituted pyrimidine molecules.