RESUMEN
PURPOSE: To experimentally and theoretically investigate the temporal effects of respiratory-gated and intensity-modulated radiotherapy (IMRT) treatment delivery on in vitro survival. METHODS AND MATERIALS: Experiments were designed to isolate the effects of periodic irradiation (gating), partial tumor irradiation (IMRT), and extended treatment time (gating and IMRT). V79 Chinese hamster lung fibroblast cells were irradiated to 2 Gy with four delivery methods and a clonogenic assay performed. Theoretical incomplete repair model calculations were performed using the incomplete repair model. RESULTS: Treatment times ranged from 1.67 min (conformal radiotherapy, CRT) to 15 min (gated IMRT). Survival fraction calculations ranged from 68.2% for CRT to 68.7% for gated IMRT. For the same treatment time (5 min), gated delivery alone and IMRT delivery alone both had a calculated survival fraction of 68.3%. The experimental values ranged from 65.7% +/- 1.0% to 67.3% +/- 1.3%, indicating no significant difference between the experimental observations and theoretical calculations. CONCLUSION: The theoretical results predicted that of the three temporal effects of radiation delivery caused by gating and IMRT, extended treatment time was the dominant effect. Care should be taken clinically to ensure that the use of gated IMRT does not significantly increase treatment times, by evaluating appropriate respiratory gating duty cycles and IMRT delivery complexity.
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Supervivencia Celular/efectos de la radiación , Fibroblastos/efectos de la radiación , Pulmón/efectos de la radiación , Radioterapia Conformacional , Respiración , Animales , Línea Celular , Supervivencia Celular/fisiología , Cricetinae , Reparación del ADN/fisiología , Reparación del ADN/efectos de la radiación , Fibroblastos/fisiología , Pulmón/citología , Modelos Biológicos , Dosis de Radiación , Radioterapia de Intensidad Modulada , Factores de TiempoRESUMEN
Advances in basic neuroscience related to neural stem cells and their malignant counterparts are challenging traditional models of central nervous system tumorigenesis and intrinsic brain repair. Neurogenesis persists into adulthood predominantly in two neurogenic centers: subventricular zone and subgranular zone. Subventricular zone is situated adjacent to lateral ventricles and subgranular zone is confined to the dentate gyrus of the hippocampus. Neural stem cells not only self-renew and differentiate along multiple lineages in these regions, but also contribute to intrinsic brain plasticity and repair. Ionizing radiation can depopulate these exquisitely sensitive regions directly or impair in situ neurogenesis by indirect, dose-dependent and inflammation-mediated mechanisms, even at doses <2 Gy. This review discusses the fundamental neural stem cell concepts within the framework of cumulative clinical experience with the treatment of central nervous system malignancies using conventional radiotherapy.
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Encéfalo/patología , Neoplasias del Sistema Nervioso Central/radioterapia , Glioma/radioterapia , Células Madre Multipotentes/fisiología , Células Madre Neoplásicas/patología , Neuronas/citología , Células Madre/fisiología , Neoplasias del Sistema Nervioso Central/etiología , Neoplasias del Sistema Nervioso Central/patología , Glioma/etiología , Glioma/patología , Humanos , Dosificación Radioterapéutica , Células Madre/patologíaRESUMEN
PURPOSE: Recent discoveries have implicated neural stem cells (NSC) as the source of plasticity and repair in the mature mammalian brain. Treatment-induced NSC dysfunction may lead to observed toxicity. This study evaluates the feasibility of NSC-preserving external beam radiotherapy. METHODS AND MATERIALS: A single computed tomography (CT) dataset depicting a right periventricular lesion was used in this study as this location reflects the most problematic geometric arrangement with respect to NSC preservation. Conventional and NSC preserving radiotherapy (RT) plans were generated for the same lesion using two clinical scenarios: cerebral metastatic disease and primary high-grade glioma. Disease-specific target volumes were used. Metastatic disease was conventionally treated with whole-brain radiotherapy (WBRT) to 3,750 cGy (15 fractions) followed by a single stereotactic radiosurgery (SRS) boost of 1,800 cGy to gross disease only. High-grade glioma was treated with conventional opposed lateral and anterior superior oblique beams to 4,600 cGy (23 fractions) followed by a 1,400 cGy (7 fractions) boost. NSC preservation was achieved in both scenarios with inverse-planned intensity modulated radiotherapy (IMRT). RESULTS: Cumulative dose reductions of 65% (metastatic disease) and 25% (high-grade glioma) to the total volume of the intracranial NSC compartments were achieved with NSC-preserving IMRT plans. The reduction of entry and exit dose to NSC niches located contralateral to the target contributed most to NSC preservation. CONCLUSIONS: Neural stem cells preservation with current external beam radiotherapy techniques is achievable in context of both metastatic brain disease and high-grade glioma, even when the target is located adjacent to a stem cell compartment. Further investigation with clinical trials is warranted to evaluate whether NSC preservation will result in reduced toxicity.
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Neoplasias Encefálicas/radioterapia , Ventrículos Cerebrales , Glioma/radioterapia , Plasticidad Neuronal , Traumatismos por Radiación/prevención & control , Células Madre/efectos de la radiación , Neoplasias Encefálicas/diagnóstico por imagen , Neoplasias Encefálicas/secundario , Neoplasias Encefálicas/cirugía , Irradiación Craneana/métodos , Estudios de Factibilidad , Glioma/cirugía , Humanos , Radiografía , Radiocirugia/métodos , Dosificación Radioterapéutica , Radioterapia de Intensidad Modulada/métodos , Células Madre/fisiologíaRESUMEN
The sequence of incremental dose delivery during a radiotherapy fraction can potentially influence the radiobiological effect. This would be most noticeable during the long fractions characteristic of hypo-fractionated stereotactic radiotherapy and radiosurgery. We demonstrate here the spatio-temporal variation of dose delivery by the CyberKnife to a lung tumor and propose strategies to reduce and/or correct for any resultant dose-time cytotoxic effects.
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Fraccionamiento de la Dosis de Radiación , Neoplasias Pulmonares/cirugía , Modelos Biológicos , Radiometría/métodos , Radiocirugia/métodos , Planificación de la Radioterapia Asistida por Computador/métodos , Algoritmos , Simulación por Computador , Relación Dosis-Respuesta en la Radiación , Humanos , Radiobiología/métodos , Dosificación Radioterapéutica , Efectividad Biológica Relativa , Factores de TiempoRESUMEN
PURPOSE: The expression of epidermal growth factor receptor (EGFR)-CD533, a truncation mutant of the wild-type EGFR, radiosensitizes carcinoma and malignant glioma cell lines. This deletion mutant disrupts EGFR activation and downstream signaling through the formation of inhibitory dimerizations. In this study, the effects of EGFR-CD533 on other ErbB receptor tyrosine kinase (RTK) family members were quantified to better understand the mechanism of EGFR-CD533-mediated radiosensitization. METHODS AND MATERIALS: Breast carcinoma cell lines with different ErbB RTK expression profiles were transduced with EGFR or ErbB2 deletion mutants (EGFR-CD533 and ErbB2-CD572) using an adenoviral vector. ErbB RTK activation, mitogen activated protein kinase (MAPK) and phosphatidylinositol-3-kinase (PI3K)/p70S6K signaling, and clonogenic survival were determined for expression of each deletion mutant. RESULTS: EGFR-CD533 radiosensitizes carcinoma cells with either high EGFR expression (MDA-MB231) or low EGFR expression (T47D) through significant blockade of the ErbB RTK network. Analysis of clonogenic survival demonstrate significant enhancement of the alpha/beta ratios, as determined by the linear-quadratic model. Split-dose survival experiments confirm that EGFR-CD533 reduces the repair of cellular damage after ionizing radiation. CONCLUSION: Expression of EGFR-CD533 inhibits the ErbB RTK network and radiosensitizes carcinoma cells irrespective of the ErbB RTK expression patterns, and ErbB2-CD572 does not radiosensitize cells with low EGFR expression. These studies demonstrate that the mechanism of action for EGFR-CD533-mediated radiosensitization is inhibition of the ErbB RTK network, and is an advantage for radiosensitizing multiple malignant cell types.
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Neoplasias de la Mama , Receptores ErbB/genética , Mutación/genética , Tolerancia a Radiación/genética , Receptor ErbB-2/genética , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/radioterapia , Línea Celular Tumoral , Daño del ADN/genética , Receptores ErbB/metabolismo , Femenino , Humanos , Receptor ErbB-2/metabolismo , Receptor ErbB-3/genética , Receptor ErbB-3/metabolismo , TransfecciónRESUMEN
Ionizing radiation induces in autocrine growth-regulated carcinoma and malignant glioma cells powerful cytoprotective responses that confer relative resistance to consecutive radiation exposures. Understanding the mechanisms of these responses should provide new molecular targets for tumor radiosensitization. ERBB and other receptor Tyr kinases have been identified as immediate early response gene products that are activated by radiation within minutes, as by their physiological growth factor ligands, and induce secondary stimulation of cytoplasmic protein kinase cascades. The simultaneous activation of all receptor Tyr kinases and nonreceptor Tyr kinases leads to complex cytoprotective responses including increased cell proliferation, reduced apoptosis and enhanced DNA repair. Since these responses contribute to cellular radioresistance, ERBB1, the most extensively studied ERBB receptor, is examined as a target for tumor cell radiosensitization. The three methods of ERBB1 inhibition include blockade of growth factor binding by monoclonal antibody against the ligand-binding domain, inhibition of the receptor Tyr kinase-mediating receptor activation, and overexpression of a dominant-negative epidermal growth factor receptor-CD533 that lacks the COOH-terminal 533 amino acids and forms nonfunctional heterodimeric complexes with wild-type receptors. All the three approaches enhance radiation toxicity in vitro and in vivo. The different mechanisms of inhibition have contributed to the understanding of cellular responses to radiation, vary in relative effectiveness and pose different challenges for translation.
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Proteínas Tirosina Quinasas Receptoras/efectos de la radiación , Animales , Humanos , Neoplasias/radioterapia , Proteínas Tirosina Quinasas Receptoras/antagonistas & inhibidoresRESUMEN
PURPOSE: To test whether or not the commonly prescribed daily dose of 2 Gy (whole fraction), when delivered as various partial fraction (PF) dose sequences simulating clinical treatment fields, produces equal biologic effects. METHODS AND MATERIALS: Eleven actively proliferating cell lines derived from human and animal tissues were used in this study. 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide (MTT) and clonogenic assays were used to determine the radiation effects on cell proliferation and survival, respectively. The 2 Gy dose was divided into 2 or more PFs for delivery to simulate the delivery of clinical treatment fields. Most irradiation sequences contained two parts consisting of at least 1 small PF, denoted by S which was 0.5 Gy or less, and a large PF, denoted by L which was 1 Gy or more. Irradiation schemes were designed to include the following conditions: (a) the 2 Gy dose divided into combinations of an L-dose and one or more S-doses; (b) the L-dose given either before or after the S-doses; and (c) delivery of all partial fractions within a fixed total time. RESULTS: Significant differences in biologic effect were observed between sequences in which the L-dose was given before or after the S-doses in both the MTT and clonogenic assays. Nearly all the latter schemes, that is S-L, produced greater cytotoxic effects than the L-S schemes. CONCLUSIONS: These data demonstrate that the biologic effects of 2 Gy may differ in different clinical settings depending on the size and sequence of the partial fractions. The variation between cytotoxic effects is likely a result of the combination of low-dose hyper-radiosensitivity (HRS) and higher-dose increased radioresistance (IRR) effects established recently. We suggest that to ensure the optimal biologic effect of a prescribed dose of 2 Gy clinically, it is critical to consider the sequence in which the treatment fields are delivered when partial fractions of different sizes are used.
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Línea Celular Tumoral/efectos de la radiación , Fraccionamiento de la Dosis de Radiación , Relación Dosis-Respuesta en la Radiación , Animales , Línea Celular Tumoral/fisiología , Supervivencia Celular , Colorantes , Humanos , Tolerancia a Radiación , Efectividad Biológica Relativa , Sales de Tetrazolio , Tiazoles , Ensayo de Tumor de Célula MadreRESUMEN
PURPOSE: The commonly expressed variant epidermal growth factor receptor (EGFR), the type III EGFR variant (EGFRvIII), functions as an oncoprotein promoting neoplastic transformation and tumorigenicity. The role of EGFRvIII in cellular responses to genotoxic stress, such as ionizing radiation, is only minimally defined. Thus, we have investigated EGFRvIII as a potential modulator of cellular radiation responses and explored the feasibility of adenovirus (Ad)-mediated expression of dominant-negative EGFR-CD533 as a gene therapeutic approach for inhibiting EGFRvIII function in vitro and in vivo. EXPERIMENTAL DESIGN AND RESULTS: EGFR-CD533 and EGFRvIII were expressed in vitro and in vivo in malignant U-373 MG glioma cells through transduction with an Ad vector, Ad-EGFR-CD533 and Ad-EGFRvIII, respectively. In vivo studies defined the importance of EGFRvIII as a modulator of radiation responses, demonstrating a 2.6-fold activation of EGFRvIII in U-373 malignant glioma tumors. Concomitant expression of EGFR-CD533 inhibited the radiation-induced activation of EGFRvIII in vitro and completely abolished the enhanced clonogenic survival conferred by EGFRvIII. The ability of EGFR-CD533 to inhibit EGFRvIII function was further confirmed in vivo through complete inhibition of EGFRvIII-mediated increased tumorigenicity and radiation-induced activation of EGFRvIII. Growth delay assays with U-373 xenograft tumors demonstrated that the expression of EGFR-CD533 significantly enhanced radiosensitivity of tumor cells under conditions of intrinsic and Ad-mediated EGFRvIII expression. CONCLUSIONS: We conclude that EGFRvIII confers significant radioresistance to tumor cells through enhanced cytoprotective responses, and we have demonstrated that dominant-negative EGFR-CD533 effectively inhibits EGFRvIII function. These data affirm the broad potential of EGFR-CD533 to radiosensitize human malignant glioma cells.
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Receptores ErbB/genética , Glioma/radioterapia , Adenoviridae/genética , Animales , Western Blotting , Células CHO , Línea Celular Tumoral , Cricetinae , Cricetulus , Receptores ErbB/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica , Genes Dominantes/genética , Vectores Genéticos/genética , Glioma/genética , Glioma/patología , Humanos , Ratones , Ratones Desnudos , Mutación , Fosforilación/efectos de la radiación , Radiación Ionizante , Factores de Tiempo , Transfección , Tirosina/metabolismo , Ensayos Antitumor por Modelo de Xenoinjerto/métodosRESUMEN
PURPOSE: To develop a theoretical model for oxygen delivery to tumors, and to use the model to simulate the effects of changing the affinity of hemoglobin for oxygen on tumor oxygenation. METHODS AND MATERIALS: Hemoglobin affinity is expressed in terms of P(50), the partial pressure of oxygen (Po(2)) at half saturation. Effects of changing P(50) on arterial Po(2) are predicted using an effective vessel approach to describe diffusive oxygen transport in the lungs, assuming fixed systemic oxygen demand and fixed blood flow rate. The decline in oxygen content of blood as it flows through normal tissue before entering the tumor region is assumed fixed. The hypoxic fraction of the tumor region is predicted using a three-dimensional simulation of diffusion from a network of vessels whose geometry is derived from observations of tumor microvasculature in the rat. RESULTS: In air-breathing rats, predicted hypoxic fraction decreases with moderate increases in P(50), but increases with further increases of P(50), in agreement with previous experimental results. In rats breathing hyperoxic gases, and in humans breathing either normoxic or hyperoxic gases, increased P(50) is predicted to improve tumor oxygenation. CONCLUSIONS: The results support the administration of synthetic agents to increase P(50) during radiation treatment of tumors.
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Hemoglobina A/metabolismo , Modelos Biológicos , Neoplasias/sangre , Oxígeno/sangre , Animales , Hipoxia de la Célula/fisiología , Humanos , Neoplasias/irrigación sanguínea , Presión Parcial , Intercambio Gaseoso Pulmonar/fisiología , Radiobiología , RatasRESUMEN
PURPOSE: In linac-based stereotactic radiosurgery, treatment is delivered intermittently via multiple individual small radiotherapy arcs. The time lapses between the individual arcs permit greater damage repair and increased tumor cell survival in comparison with continuous irradiation. Because pentoxifylline (PTX) has been reported to prevent radiation-induced cell cycle arrest at the G2/M checkpoint, where damage repair is critically linked to cell survival, we hypothesized that PTX would exert a favorable radiosensitization effect by reducing the recovery observed during intermittent stereotactic radiosurgery. METHODS AND MATERIALS: The human glioma cell line T98G was used to study the effects of continuous vs. intermittent irradiation with or without PTX. Cell cycle patterns were studied using flow cytometry. Clonogenic assays of single cells and spheroid outgrowth assays provided a quantitative measure of PTX-mediated radiosensitization. The PTX effect upon cells in low oxygen conditions was also studied in vitro after enzymatic oxygen scavenging. RESULTS: Flow Cytometry: T98G cells exposed to both continuous and intermittent irradiation exhibit similar arrest at the G2/M checkpoint. The addition of 2 mM PTX significantly reduced the radiation-induced G2/M block in both irradiation schemes. Clonogenic Assays: The same PTX concentration applied before a continuous dose of 12 Gy, two intermittent doses of 6 Gy, or three intermittent doses of 4 Gy, all given within a 1-h interval, consistently caused radiosensitization. The drug enhancement ratios for PTX were 1.5, 2.7, and 6.0 for the continuous and two different intermittent dose schedules, respectively. Adding PTX after irradiation yielded lower enhancement ratios than pre-irradiation application. A similar pattern was observed after total doses of 4, 6, 9, or 12 Gy, as well. In low oxygen conditions, PTX was seen to have the same effects as in normoxic conditions. Spheroid Outgrowth Assays: The in vitro PTX effects were replicated in the spheroid outgrowth assays. CONCLUSION: In human glioma cells, PTX abrogates the radiation-induced G2/M block observed after either continuous radiation exposure or intermittent exposures modeling clinical linac-based radiosurgery. The PTX-mediated reduction of the G2/M block translates into radiosensitization, most notably during intermittent exposures, and is presumably a consequence of diminished DNA damage repair at the G2/M checkpoint, though other contributing effects cannot be ruled out. The radiosensitization effect of PTX is sustained under low oxygen conditions. These results support consideration of the clinical evaluation of PTX to enhance the efficacy of linac-based radiosurgery involving intermittent irradiation through multiple arcs.
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Fase G2/efectos de los fármacos , Glioma/cirugía , Pentoxifilina/farmacología , Fármacos Sensibilizantes a Radiaciones/farmacología , Radiocirugia/métodos , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/efectos de la radiación , Reparación del ADN , Citometría de Flujo/métodos , Fase G2/efectos de la radiación , Glioma/patología , Humanos , Dosis de Radiación , Protectores contra Radiación/farmacología , Radiobiología , Esferoides Celulares/efectos de los fármacos , Esferoides Celulares/efectos de la radiación , Células Tumorales Cultivadas/efectos de los fármacos , Células Tumorales Cultivadas/efectos de la radiaciónRESUMEN
PURPOSE: Genetic radiotherapy, the combination of gene therapy and radiation therapy, for cancer treatment is evolving from laboratory studies to clinical trials. Genetic radiotherapy involves the viral infection of cells that change the sensitivity of transduced cells to radiation. Because there is no patient outcome data for genetic radiotherapy, prospective models are needed to determine the expected benefit of this new modality. Such a prospective model has been developed in this work. METHODS AND MATERIALS: An existing tumor control probability (TCP) calculation model developed for external beam radiotherapy was modified for genetic radiotherapy. Specifically, the (1) transduced fraction and (2) enhancement factor of the transduced cells were included in the model. Parametric studies of the effects of these two variables on TCP for head-and-neck cancer were performed. RESULTS: Using reasonable transduction fraction and enhancement factor values (0.8 and 1.4, respectively), the model predicts an increase in the TCP for genetic radiotherapy over radiotherapy alone by up to 15% for the same radiotherapy dose. The theoretical limit of TCP increase was calculated to be near 70%, which may occur with improved techniques that increase the transduced fraction or because of a strong bystander effect. To maintain existing TCP, dose reductions from 5 Gy (reasonable values) to >30 Gy (ideal case) are predicted for genetic radiotherapy over radiotherapy alone. CONCLUSIONS: Our results indicate that genetic radiotherapy has the potential to significantly improve tumor control over radiotherapy alone.
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Terapia Genética/métodos , Neoplasias de Cabeza y Cuello/terapia , Modelos Biológicos , Modelos Estadísticos , Transducción Genética/métodos , Resultado del Tratamiento , Terapia Combinada/métodos , Simulación por Computador , Relación Dosis-Respuesta en la Radiación , Neoplasias de Cabeza y Cuello/radioterapia , Humanos , Tolerancia a Radiación/genética , Radioterapia/métodos , Dosificación Radioterapéutica , Sensibilidad y EspecificidadRESUMEN
PURPOSE: We recently reported that an adenovirus-expressing mutant HSV-TK75 (AdCMV-TK75) radiosensitized rat syngeneic gliomas in combination with low concentrations of acyclovir (ACV) much more effectively than a virus expressing wild-type herpes simplex virus thymidine kinase (HSV-TK). In this report we have examined whether similar radiosensitizing effects are also seen with human glioma cells in vitro and in vivo. METHODS AND MATERIALS: Human U87 MG glioma cells were transduced with AdCMV-TK75 and exposed to ACV followed by single-dose irradiation and colony-forming survival assays. Similarly, U87 MG xenografts were infused with AdCMV-TK75 or Adbetagal control virus, followed by ACV administration and fractionated irradiation. Therapeutic efficacy was monitored by tumor growth. RESULTS: U87 MG cells transduced with AdCMV-TK75 were significantly more sensitive to ACV (3 microM) than cells transduced with either wild-type HSV-TK or control virus. To determine whether human cells also demonstrate improved radiosensitization similar to that seen with rat glioma cells and tumors, we transduced U87 MG cells with either AdCMV-TK75, AdCMV-TK, expressing wild-type HSV-TK, or Adbetagal and then treated the cells with 3 microM of ACV. Cells transduced with AdCMV-TK75 were significantly more radiosensitive (dose enhancement ratio [D(37)]: 2.6) by colony-forming survival assay than cells transduced with either AdCMV-TK or Adbetagal. Furthermore, we found that U87 MG xenografts infused with AdCMV-TK75 by slow positive pressure infusion were more radiosensitive after administration of ACV than tumors infused with Adbetagal. A more dramatic result was achieved when fractionated irradiation was carried out concurrently with ACV administration, in which case AdCMV-TK75-treated tumors did not grow at all. CONCLUSIONS: These results demonstrate that transduction of human glioma cells in vitro and infusion of xenografts in vivo with AdCMV-TK75 and treatment with concentrations of ACV that can be achieved in vivo produce similar radiosensitization, as previously reported with rat glioma cells and intracerebral syngeneic tumors. In addition, concurrent treatment with ACV and radiation therapy is more efficient than when ACV is administered before radiation therapy.
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Aciclovir/uso terapéutico , Adenoviridae/genética , Antivirales/uso terapéutico , Glioma/radioterapia , Herpesviridae/enzimología , Mutación , Tolerancia a Radiación , Timidina Quinasa/genética , Adenoviridae/enzimología , Animales , Terapia Combinada , Glioma/tratamiento farmacológico , Glioma/virología , Humanos , Ratones , Ratones Desnudos , Radiobiología , Timidina Quinasa/metabolismo , Transducción Genética , Trasplante Heterólogo , Células Tumorales Cultivadas/efectos de los fármacos , Células Tumorales Cultivadas/efectos de la radiación , Ensayo de Tumor de Célula MadreRESUMEN
PURPOSE: During a high-dose-rate (HDR) brachytherapy treatment, as the source steps through different dwell positions, the dose rate at any fixed point within the implant varies, because the distance between the point and the source continually changes. The instantaneous dose rate may vary by a factor of 100 or more, in a complex dwell position sequence. Two different points which receive the same total dose may have received that dose with a very different sequence of dose rates. Any effects due to the complex changes in dose rate, including the sequence of dose delivery, are ignored. We investigated the possible effects of the sequence in which dose is delivered at two different dose rates, representative of dose rates that occur during an HDR treatment. METHODS AND MATERIALS: The target consisted of a tube containing a 1.0 cm(3) suspension of V-79 Chinese hamster cells. Two fixed source dwell positions near and far from the target, representing high and intermediate dose rates, were considered. The experiments compared the survival of V-79 cells exposed to an irradiation sequence consisting of either an HDR component followed by an intermediate-dose-rate component (H-I arm), or the reverse (I-H arm). In either case, the total dose and the dose ratio were the same, only the order in which the high- or intermediate-dose-rate components of the dose were delivered was changed. RESULTS: When the intermediate-dose-rate component was given before the HDR component, there was increased survival. All data pairs from three experiments showed greater survival for the I-H arm than the H-I arm by amounts ranging from 4% to 24%. Simple linear-quadratic models such as the Lea-Catchside model, which is invariant to time reversal of irradiation sequence, do not predict these results. CONCLUSIONS: These results suggest that targets receiving the same total dose of radiation during an HDR implant may not experience the same biological effect. This may be related to induced radioresistance or sublethal damage repair.
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Braquiterapia/métodos , Supervivencia Celular/efectos de la radiación , Animales , Supervivencia Celular/fisiología , Células Cultivadas , Cricetinae , Cricetulus/fisiología , Tolerancia a Radiación , Dosificación RadioterapéuticaRESUMEN
BACKGROUND AND PURPOSE: The type-III EGFR variant (EGFRvIII) is known to promote enhanced tumorigenicity. We have previously defined the importance of EGFRvIII in cellular radiation responses using Chinese hamster ovary cells (CHO). In the current study, we have extended our investigations of EGFRvIII to human tumor cells in vitro and in vivo and further verified the important role of EGFRvIII in modulating radiosensitivity. MATERIAL AND METHODS: The cell lines MDA-MB-231, U-87 MG, A-431 and U-373 MG were used. Adenoviral (Ad) vectors were produced to overexpress EGFRvIII in vitro or in xenograft tumors in vivo. The EGFR, EGFRvIII expression and tyrosine phosphorylation (Tyr-P) levels were quantified by Western blotting. The relative radiosensitivities were assessed in vitro by standard colony formation and in vivo by tumor growth delay assays. RESULTS: The presence of EGFRvIII was verified in all xenograft tumors tested with no detectable expression in the corresponding cells under in vitro culture conditions. MDA-MB-231 xenograft tumors demonstrated EGFRvIII expression levels, which were 1.9-fold higher relative to EGFRwt compared to a 14.5-fold higher Tyr-P. Ionizing radiation of these tumors at 4 Gy induced an average 3.2-fold increase in EGFRvIII Tyr-P. EGFRvIII expression in U-373 MG cells significantly enhanced survival after 4Gy, which was completely abolished by dominant-negative EGFR-CD533. Finally, the ability of EGFRvIII to accelerate tumor growth during irradiation was confirmed in vivo. CONCLUSION: EGFRvIII is frequently expressed in a variety of different tumor types and can confer significant radioresistance, thus further providing evidence for EGFRvIII as an additional important target in our approaches to radiosensitize malignant solid tumors.
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Receptores ErbB/fisiología , Tolerancia a Radiación/efectos de los fármacos , Animales , Receptores ErbB/análisis , Receptores ErbB/metabolismo , Humanos , Ratones , Trasplante de Neoplasias , Fosforilación , Trasplante Heterólogo , Células Tumorales Cultivadas , Tirosina/metabolismoRESUMEN
A clinically relevant dose (2.0 Gy) of ionizing radiation (IR) was employed to determine if subsequent exposure to the protein kinase C (PKC) and Chk 1 inhibitor UCN-01 for 24 h could abrogate IR-induced G2/M arrest and promote apoptosis in U937 leukemic cells ectopically expressing Bcl-2 (U937/Bcl-2). To this end, empty-vector control (U937/pCEP4) and U937/Bcl-2 cells were exposed to two UCN-01 concentrations following IR: i) a 50 nM concentration, which by itself was minimally toxic to both cell lines, and ii) a 150 nM concentration, which modestly induced apoptosis (e.g., ~19%) in control cells after 24 h. The effects of UCN-01 on IR responses were examined in relation to apoptosis induction, suspension culture growth inhibition, loss of clonogenic survival, and cell cycle perturbations. IR (2 Gy) alone minimally induced apoptosis in both U937 transfectant cell lines (e.g., <5% at 24 h in each case). Although UCN-01 failed to potentiate IR-mediated apoptosis at either early (e.g., 24 h) or late (e.g., 72 h) intervals, exposure to 50 or 150 nM UCN-01 resulted in a significant, albeit modest, reduction in proliferation and colony formation in irradiated U937/pCEP4 and U937/Bcl-2 cells. Despite failing to enhance apoptosis, UCN-01 treatment abrogated IR-induced G2/M arrest in both cell lines, an event associated with enhanced activation of cyclin-dependent kinase 1 (cdk1), promotion of G0/G1 arrest, and dephosphorylation of the retinoblastoma protein (pRb). Together, these findings indicate that exposure of U937 cells ectopically-expressing Bcl-2 to the combination of UCN-01 + IR leads to a further reduction in cell proliferation, and that this phenomenon appears to involve a non-apoptotic mechanism.
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Alcaloides/farmacología , Antineoplásicos/farmacología , Ciclo Celular/efectos de los fármacos , Ciclo Celular/efectos de la radiación , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Células U937/efectos de los fármacos , Células U937/efectos de la radiación , Apoptosis/efectos de los fármacos , Apoptosis/efectos de la radiación , División Celular/efectos de los fármacos , División Celular/efectos de la radiación , Terapia Combinada , Humanos , Potenciales de la Membrana/efectos de los fármacos , Potenciales de la Membrana/efectos de la radiación , Mitocondrias/efectos de los fármacos , Mitocondrias/efectos de la radiación , Proteína Quinasa C/antagonistas & inhibidores , Radiación Ionizante , Estaurosporina/análogos & derivados , Transfección , Ensayo de Tumor de Célula Madre , Células U937/metabolismoRESUMEN
It has been suggested that continued tobacco use during radiation therapy contributes to maintenance of neoplastic growth despite treatment with radiation. Nicotine is a cigarette component that is an established risk factor for many diseases, neoplastic and otherwise. The hypothesis of this work is that nicotine promotes the proliferation, migration, and radioresistance of human malignant glioma cells. The effect of nicotine on cellular proliferation, migration, signaling, and radiation sensitivity were evaluated for malignant glioma U87 and GBM12 cells by use of the AlamarBlue, scratch healing, and clonogenic survival assays. Signal transduction was assessed by immunoblotting for activated EGFR, ERK, and AKT. At concentrations comparable with those found in chronic smokers, nicotine induced malignant glioma cell migration, growth, colony formation, and radioresistance. Nicotine increased phosphorylation of EGFR(tyr992), AKT(ser473), and ERK. These molecular effects were reduced by pharmacological inhibitors of EGFR, PI3K, and MEK. It was therefore concluded that nicotine stimulates the malignant behavior of glioma cells in vitro by activation of the EGFR and downstream AKT and ERK pathways.
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Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Receptores ErbB/metabolismo , Glioma/patología , Nicotina/efectos adversos , Nicotina/farmacología , Tolerancia a Radiación/efectos de los fármacos , Receptores ErbB/antagonistas & inhibidores , Receptores ErbB/fisiología , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Quinasas MAP Reguladas por Señal Extracelular/fisiología , Humanos , Fosforilación , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Proto-Oncogénicas c-akt/fisiología , Transducción de Señal/efectos de los fármacos , Estimulación Química , Células Tumorales CultivadasRESUMEN
Bioluminescence imaging (BLI) is a relatively new noninvasive technology used for quantitative assessment of tumor growth and therapeutic effect in living animal models. BLI involves the generation of light by luciferase-expressing cells following administration of the substrate luciferin in the presence of oxygen and ATP. In the present study, the effects of hypoxia, hypoperfusion, and pH on BLI signal (BLS) intensity were evaluated in vitro using cultured cells and in vivo using a xenograft model in nude mice. The intensity of the BLS was significantly reduced in the presence of acute and chronic hypoxia. Changes in cell density, viability, and pH also affected BLS. Although BLI is a convenient non-invasive tool for tumor assessment, these factors should be considered when interpreting BLS intensity, especially in solid tumors that could be hypoxic due to rapid growth, inadequate blood supply, and/or treatment.
RESUMEN
The purpose of this study was to investigate the increase in cell kill that can be achieved by tumor irradiation with heterogeneous dose distributions targeting hypoxic regions that can be visualized with non-invasive imaging. Starting with a heterogeneous distribution of microvessels, a microscopic two-dimensional model of tumor oxygenation was developed using planar simulation of oxygen diffusion. Non-invasive imaging of hypoxia was simulated taking partial volume effect into account. A dose-modulation scheme was implemented with the goal of delivering higher doses to the hypoxic pixels, as seen in simulated hypoxia images. To determine the relative cell kill in response to hypoxia-targeting irradiation, tumor cell survival fractions were compared to those resulting from treatments delivering the same average dose to the lesion in a spatially uniform fashion. It was shown that hypoxia-targeting dose modulation may be better suited for tumors with low α/ß, low hypoxic fraction and spatially aggregated hypoxic features. Most importantly, it was determined that at low fraction doses there is no cell kill increase from targeting hypoxic regions alone versus escalating the total tumor dose. However, for higher doses per fraction (≥8 Gy/fraction), the effectiveness of hypoxia-targeting irradiation increases, resulting in the tumoricidal effect of up to 30% higher than that of uniform tumor irradiation delivering the same average tumor dose.
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Fraccionamiento de la Dosis de Radiación , Modelos Biológicos , Hipoxia de la Célula/efectos de la radiación , Línea Celular Tumoral , Supervivencia Celular/efectos de la radiación , Humanos , Neoplasias/diagnóstico por imagen , Neoplasias/patología , Tomografía de Emisión de PositronesRESUMEN
PURPOSE: To compare the ability of single- and dual-isotope prostate seed implants to escalate biologically effective dose (BED) to foci of disease while reducing prescription dose to the prostate. METHODS AND MATERIALS: Nine plans, using 125I, 103Pd, and 131Cs alone and in combination were created retrospectively for 2 patients. Ultrasound and MRI/MRS datasets were used for treatment planning. Voxel-by-voxel BED was calculated for single- and dual-isotope plans. Equivalent uniform BED (EUBED) was used to compare plans. The MRS-positive planning target volumes (PTVi) were delineated along with PTV (prostate+5 mm), rectum, and urethra. Single-isotope implants, prescribed to conventional doses, were generated to achieve good PTV coverage. The PTVi were prospectively used to generate implants using mixtures of isotopes. For mixed-radioisotope implants, we also explored the impact on EUBED of lowering prescription doses by 15%. RESULTS: The EUBED of PTVi in the setting of primary 125I implant increased 20-66% when 103Pd and 131Cs were used compared with 125I boost. Decreasing prescription dose by 15% in mixed-isotope implants results in a potential 10% reduction in urethral EUBED with preservation of PTV coverage while still boosting PTVi (up to 80%). When radiobiologic parameters corresponding to more-aggressive disease are assigned to foci, faster-decaying isotopes used in mixed implants have the potential to preserve the equivalent biological effect of mono-isotope implants considering less-aggressive disease distributed in the entire prostate. CONCLUSIONS: This is a hypothesis-generating study proposing a treatment paradigm that could be the middle ground between whole-gland irradiation and focal-only treatment. The use of two isotopes concurrent with decreasing the minimal peripheral dose is shown to increase EUBED of selected subvolumes while preserving the therapeutic effect at the level of the gland.
Asunto(s)
Braquiterapia/métodos , Modelos Biológicos , Próstata/efectos de la radiación , Neoplasias de la Próstata/radioterapia , Traumatismos por Radiación/prevención & control , Braquiterapia/efectos adversos , Radioisótopos de Cesio/uso terapéutico , Estudios de Factibilidad , Semivida , Humanos , Radioisótopos de Yodo/uso terapéutico , Imagen por Resonancia Magnética , Espectroscopía de Resonancia Magnética , Masculino , Tamaño de los Órganos , Órganos en Riesgo/efectos de la radiación , Paladio/uso terapéutico , Próstata/diagnóstico por imagen , Próstata/patología , Neoplasias de la Próstata/diagnóstico , Neoplasias de la Próstata/diagnóstico por imagen , Radioisótopos/uso terapéutico , Dosificación Radioterapéutica , Planificación de la Radioterapia Asistida por Computador/métodos , Recto/efectos de la radiación , Efectividad Biológica Relativa , Estudios Retrospectivos , Carga Tumoral , Ultrasonografía , Uretra/efectos de la radiaciónRESUMEN
Although tyrosyl-DNA phosphodiesterase (TDP1) is capable of removing blocked 3' termini from DNA double-strand break ends, it is uncertain whether this activity plays a role in double-strand break repair. To address this question, affinity-tagged TDP1 was overexpressed in human cells and purified, and its interactions with end joining proteins were assessed. Ku and DNA-PKcs inhibited TDP1-mediated processing of 3'-phosphoglycolate double-strand break termini, and in the absence of ATP, ends sequestered by Ku plus DNA-PKcs were completely refractory to TDP1. Addition of ATP restored TDP1-mediated end processing, presumably due to DNA-PK-catalyzed phosphorylation. Mutations in the 2609-2647 Ser/Thr phosphorylation cluster of DNA-PKcs only modestly affected such processing, suggesting that phosphorylation at other sites was important for rendering DNA ends accessible to TDP1. In human nuclear extracts, about 30% of PG termini were removed within a few hours despite very high concentrations of Ku and DNA-PKcs. Most such removal was blocked by the DNA-PK inhibitor KU-57788, but approximately 5% of PG termini were removed in the first few minutes of incubation even in extracts preincubated with inhibitor. The results suggest that despite an apparent lack of specific recruitment of TDP1 by DNA-PK, TDP1 can gain access to and can process blocked 3' termini of double-strand breaks before ends are fully sequestered by DNA-PK, as well as at a later stage after DNA-PK autophosphorylation. Following cell treatment with calicheamicin, which specifically induces double-strand breaks with protruding 3'-PG termini, TDP1-mutant SCAN1 (spinocerebellar ataxia with axonal neuropathy) cells exhibited a much higher incidence of dicentric chromosomes, as well as higher incidence of chromosome breaks and micronuclei, than normal cells. This chromosomal hypersensitivity, as well as a small but reproducible enhancement of calicheamicin cytotoxicity following siRNA-mediated TDP1 knockdown, suggests a role for TDP1 in repair of 3'-PG double-strand breaks in vivo.