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Ralph R Weichselbaum

Publications and source records attributed to Ralph R Weichselbaum.

53 records · Page 3Linked to original sources
PubMed2003

Prolonged treatment with angiostatin reduces metastatic burden during radiation therapy.

Ionizing radiation (IR) and concomitant angiostatin (AS) produce greater than additive local antitumor effects. We examined whether prolonged AS treatment added to IR reduces proliferation of lung metastases from LLC primary tumors. Flank tumors were treated with 40 Gy with or without AS (25 mg/kg/day). IR plus a 14-day course of AS improved local tumor control and blocked the increase in lung weights observed in the group receiving IR plus a 2-day course of AS group. Animals treated with prolonged AS exhibited no increase in lung weight and no macrometastases. These findings suggest that long-term treatment with antiangiogenic compounds may be effective in preventing metastases from IR-treated tumors as well as increasing the local antitumor effects of radiotherapy.

Angiostatins↗
PubMed2003

Sustained inhibition of experimental neointimal hyperplasia with a genetically modified herpes simplex virus.

OBJECTIVE: Reported herein is a potential strategy for sustained smooth muscle cell (SMC) inhibition with a virulence-attenuated herpes simplex virus (HSV). Experiments were conducted in vitro to demonstrate selective SMC cytotoxicity and in vivo to demonstrate reduced neointimal hyperplasia (NIH) in a clinically relevant animal model. METHODS: In vitro: Cultured human umbilical artery smooth muscle cells (UASMC) and venous endothelial cells (HUVEC) were exposed to varying multiplicities of infection (MOI) of a gamma(1)34.5-deleted HSV-1 virus (R849). Cell survival was assessed at 48 and 72 hours with a colorimetric MTT viability assay. In vivo: New Zealand White rabbit external jugular veins (n = 21) were exposed to R849 (2.5 x 10(6) pfu/mL) or culture medium at 110 to 120 mm Hg for 10 minutes, then fashioned as vein patches on carotid arteries. Carotid arteries were ligated distally to decrease blood flow and stimulate a hyperplastic response (ultra-low shear stress model). After 2, 4, 12, and 24 weeks, patched segments were perfusion-fixed with glutaraldehyde and morphometrically examined for NIH formation. RESULTS: In vitro: At 48 hours, R849 exhibited preferential cytotoxicity to UASMC compared with HUVEC, with 11% +/- 10% of UASMCs and 49% +/- 8% of HUVECs surviving after infection with MOI = 25 (P <.05). Higher MOI resulted in poor survival of both cell lines. In vivo: Blood flow was similarly reduced in all animals both at surgery (0.9 +/- 0.1 mL/min vs 1.6 +/- 0.3 mL/min) and at harvest (2.7 +/- 0.4 mL/min vs 2.5 +/- 0.5 mL/min). R849-infected patches exhibited markedly less NIH than control patches did at 2 weeks (162 +/- 14 microm vs 49 +/- 6 microm; P <.05), 4 weeks (190 +/- 27 microm vs 67 +/- 8 microm; P <.05), and 12 weeks (233 +/- 18 microm vs 113 +/- 2 microm; P <.05). CONCLUSION: The virulence-attenuated HSV strain R849 demonstrates selective cytotoxicity for SMC and is capable of sustained inhibition of NIH in an experimental model of vein graft failure.

Animals↗
PubMed2003

Receiver operating characteristic analysis: a general tool for DNA array data filtration and performance estimation.

A critical step for DNA array analysis is data filtration, which can reduce thousands of detected signals to limited sets of genes. Commonly accepted rules for such filtration are still absent. We present a rational approach, based on thresholding of intensities with cutoff levels that are estimated by receiver operating characteristic (ROC) analysis. The technique compares test results with known distributions of positive and negative signals. We apply the method to Atlas cDNA arrays, GeneFilters, and Affymetrix GeneChip. ROC analysis demonstrates similarities in the distribution of false and true positive data for these different systems. We illustrate the estimation of an optimal cutoff level for intensity-based filtration, providing the highest ratio of true to false signals. For GeneChip arrays, we derived filtration thresholds consistent with the reported data based on replicate hybridizations. Intensity-based filtration optimized with ROC combined with other types of filtration (for example, based on significances of differences and/or ratios), should improve DNA array analysis. ROC methodology is also demonstrated for comparison of the performance of different types of arrays, imagers, and analysis software.

Data Interpretation, Statistical↗
PubMed2003

Oct-1 is posttranslationally modified and exhibits reduced capacity to bind cognate sites at late times after infection with herpes simplex virus 1.

In herpes simplex virus 1-infected cells, a high level of alpha gene expression requires the transactivation of the genes by a complex containing the viral alpha transinducing factor (alphaTIF) and two cellular proteins. The latter two, HCF-1 and octamer binding protein Oct-1, are transcriptional factors regulated in a cell cycle-dependent manner. alphaTIF is a protein made late in infection but packaged with the virion to transactivate viral genes in newly infected cells. In light of the accumulation of large amounts of alphaTIF, the absence of alpha gene expression late in infection suggested the possibility that one or more transcriptional factors required for alpha gene expression is modified late in infection. Here we report that Oct-1 is posttranscriptionally modified late in infection, that the modification is mediated by the virus but does not involve viral protein kinases or cdc2 kinase activated by the virus late in infection, and that the modified Oct-1 has a reduced affinity for its cognate DNA site. These results are consistent with the hypothesis that modification of Oct-1 transcriptional factor could account at least in part for the shutoff of alpha gene expression late in infection.

Binding Sites↗
PubMed2003

Dual induction of the Epo-Egr-TNF-alpha- plasmid in hypoxic human colon adenocarcinoma produces tumor growth delay.

Gene therapy is a modality for the treatment of solid tumors that involves the introduction of a suicide gene into the tumor cells. Genetic radiotherapy involves the placement of a radiation-sensitive promoter upstream from a suicide gene. Because of their irregular vasculature some solid tumors are chronically hypoxic and hence are resistant to conventional treatment with chemotherapy and ionizing radiation (IR). The purpose of this study was to demonstrate that regional tumor hypoxia could be exploited to improve local tumor control. The cDNA coding the erythropoietin hypoxia-responsive element (EPO) was placed upstream from the Egr-TNF-alpha construct. WIDR human colon adenocarcinoma cells were injected into the right hind limb of nude mice and treated with Epo-Egr-TNF-alpha plasmid with or without IR. Tumor volumes were measured by calipers and tumor necrosis factor (TNF)-alpha content of the tumor was determined by enzyme-linked immunosorbent assay. Treatment with the combined regimen of Epo-Egr-TNF-alpha plasmid + IR resulted in significant tumor growth delay. Tumor TNF-alpha content was increased by 30 per cent in the combined treatment group compared with each treatment alone. Regional tumor hypoxia can be exploited successfully to induce tumor growth delay, enhance local control, and enhance the therapeutic ratio.

Adenocarcinoma↗
PubMed2003

Multicenter randomized Phase II study of paclitaxel (1-hour infusion), fluorouracil, hydroxyurea, and concomitant twice daily radiation with or without erythropoietin for advanced head and neck cancer.

PURPOSE: To expand on our experience with the combination of paclitaxel, fluorouracil, hydroxyurea, and twice daily irradiation (T-FHX) and to assess the impact of weekly administration of erythropoietin (r-HuEpo) on transfusion requirements, we conducted a Phase II multi-institutional trial with a simplified 1-h paclitaxel infusion schedule and randomized patients to receive weekly doses of r-HuEpo. PATIENTS AND METHODS: A total of 90 patients with locally advanced head and neck cancers (stage IV, 96%; N(2)/N(3), 66%) were treated on a regimen of 1-h infusion of paclitaxel (100 mg/m(2)/day, day 1), 120-h infusion of 5-fluorouracil (600 mg/m(2)/day, days 0-5); hydroxyurea 500 mg p.o. every 12 h for 11 doses; and radiation 150cGy bid, days 1-5 of each 14-day cycle repeated for five cycles over 10 weeks (7200-7500 cGy). Before initiating therapy, patients were randomized to receive r-HuEpo 40,000 IU s.c. once weekly. RESULTS: At median follow-up of 40 months, 3-year progression-free survival is 62%, locoregional control is 84%, and systemic control is 79%. Overall survival is 59%. Anemia, leucopenia, dermatitis, and mucositis were the most frequent grade 3 or 4 toxicities. Patients randomized to erythropoietin experienced less grade 2/3 anemia (52 versus 77%; P = 0.02), but transfusion requirements were not significantly different. CONCLUSIONS: T-FHX is an active and tolerable regimen inducing local tumor control and promising survival with organ preservation in high-risk patients. One h infusion of paclitaxel simplified the regimen without compromising efficacy. Addition of erythropoietin does not reduce the need for transfusion with this nonplatinum-containing regimen. T-FHX should be advanced to a randomized trial and compared with a cisplatin-based concomitant regimen.

Adult↗
PubMed2002

Angiostatin potentiates cyclophosphamide treatment of metastatic disease.

PURPOSE: We examined the interaction between cyclophosphamide (CPA) and angiostatin (AS) on the growth of primary Lewis lung carcinoma (LLC) tumors and on the development of LLC pulmonary metastases. We studied the effects of AS and CPA on the stages of angiogenesis employing in vitro assays. METHODS: Primary tumor growth and pulmonary metastases were measured to evaluate the effects of treatment with AS alone, CPA alone or the combination of CPA and AS. We examined the effects of CPA plus AS on endothelial cell (HUVEC) survival, migration and tube formation. RESULTS: Combined treatment with CPA and AS did not significantly affect primary tumor growth when compared with CPA treatment alone. However, a significant decrease in the number of pulmonary metastases was observed following CPA plus AS treatment when compared with CPA treatment alone ( P<0.001). AS did not enhance CPA-mediated HUVEC cytotoxicity, and CPA failed to enhance AS-mediated inhibition of migration. However, tube formation was inhibited following combined treatment with CPA and AS when compared with either treatment alone. CONCLUSIONS: AS enhanced the antimetastatic effects of CPA without significantly influencing the effects of CPA on primary tumor growth. CPA plus AS inhibited tube formation, suggesting that interrupting specific steps in the angiogenesis process might be an effective approach to the treatment of subclinical distant metastases.

Angiogenesis Inhibitors↗
PubMed2002

Activation of the nuclear transcription factor kappaB (NFkappaB) and differential gene expression in U87 glioma cells after exposure to the cytoprotector amifostine.

PURPOSE: Amifostine has been approved as a therapy to decrease the incidence of moderate-to-severe xerostomia in patients undergoing postoperative radiation treatment for head-and-neck cancer. As a reducing agent capable of participating in intracellular reductive/oxidative processes, it has the potential to affect redox-sensitive transcription factors and gene expression. Amifostine's active free thiol WR-1065 was investigated to determine its effect on nuclear transcription factor kappaB (NFkappaB) activation and subsequent gene expression in U87 glioma cells. METHODS AND MATERIALS: The human glioma cell line U87 was grown to confluency and then exposed to WR-1065 at a concentration of 40 microM for times ranging from 30 min to 24 h. Changes in cell cycle were monitored by flow cytometry. The effect of WR-1065 on NFkappaB activation was determined by a gel shift assay. Changes in gene expression as a function of time of exposure to WR-1065 were determined by Northern blot and the Atlas Human cDNA Expression Array (Clontech, Palo Alto, CA). Changes in gene expression using the Atlas Array were verified by reverse transcriptase-polymerase chain reaction (RT-PCR) with gene-specific primers. RESULTS: Exposure of U87 cells to 40 microM WR-1065 resulted in a marked activation of NFkappaB between 30 min and 1 h after treatment. Expression of MnSOD, an NFkappaB-responsive gene, was enhanced by over 2-fold after 16 h of treatment and remained elevated at 24 h. During this period of time, no changes in cell cycle distribution were observed. To assess changes in the expression levels of NFkappaB-responsive genes as a function of WR-1065 exposure, cDNA arrays containing 49 genes identified as having DNA-binding motifs for NFkappaB were used. Only five genes were found to be significantly affected at 1, 4, and/or 16 h of treatment. GST-3 and c-myc were repressed up to 2- and 4-fold, respectively. The expression levels of IL-2Ra, RANTES, and c-myb, in contrast, were enhanced up to 14-, 3-, and 2-fold, respectively. The remaining genes having NFkappaB-responsive elements in their promoter regions were either not expressed (20 genes) or were not affected (24 genes) by exposure to WR-1065. CONCLUSIONS: The redox-sensitive transcription factor NFkappaB can be activated in U87 glioma cells by the active thiol form of the cytoprotector amifostine. Activation of NFkappaB by the antioxidant WR-1065 is accompanied by a reduced expression of the oncogene c-myc and an enhanced expression of the antioxidant gene MnSOD, a gene whose expression in tumor cells is relatively low, but when overexpressed has been correlated with a suppression of the malignant phenotype. Activation of NFkappaB by WR-1065, however, results in selective rather than global changes in the expression of genes containing NFkappaB-responsive elements.

Amifostine↗
PubMed2002

Tumor production of angiostatin is enhanced after exposure to TNF-alpha.

Infection of tumors with an adenoviral vector expressing a chimeric gene composed of the CArG elements of the Egr-1 promoter and a cDNA encoding TNF-alpha (Ad.Egr-TNF) has previously been shown to result in the production of high intratumoral levels of TNF-alpha and thereby tumor regression. The antitumor effects of TNF-alpha were ascribed to vascular thrombosis. We and others, have reported that inhibition of tumor vessel thrombosis using anticoagulation therapy does not abrogate the antitumor effects after TNF-alpha treatment. To investigate the potential antiangiogenic effects of TNF-alpha, we studied the generation of angiostatin after intratumoral injection of Ad.Egr-TNF. We report an increase in plasma angiostatin levels both during and after treatment with Ad.Egr-TNF that parallel tumor regression. We also report that TNF-alpha enhances angiostatin production by inducing the activity of plasminogen activator and the release of MMP-9 by tumor cells. These studies support a model in which the antiangiogenic effects of TNF-alpha on the tumor microvasculature are mediated by generation of angiostatin.

Adenocarcinoma↗
PubMed2002

Inhibition of spontaneous metastases formation by amifostine.

Amifostine was investigated for its ability to inhibit spontaneous metastases formation using the well-characterized murine sarcoma, Sa-NH. Amifostine was administered intraperitoneally at a dose of 50 mg/kg every other day for 6 days to C3Hf/Kam mice until tumors reached an average size of 8-8.5 mm in diameter. Amifostine was again administered immediately after surgical removal of the tumor-bearing limbs by amputation, and then once more 2 days later. Twenty-one days later, animals were evaluated for the presence of spontaneously developed pulmonary metastases. Nontumor-bearing control animals were sham treated using the same dosing and surgery schedules. Treatment with amifostine appeared to slightly delay tumor growth, that is, 13 vs. 12 days for tumors to reach an average diameter of 8 mm. Amifostine reduced both the incidence of pulmonary metastases formed in experimental animals from 77% to 57% (p < 0.05), and their average number per animal from 12.8 +/- 5.4 (SEM) to 2.9 +/- 1.1 (SEM). The effect of amifostine exposure on serum levels of the angiogenesis inhibitor angiostatin was also determined using Western blot analysis. Consistent with the antimetastatic effect, exposure of animals to 50 mg/kg of amifostine resulted in a 4-fold enhanced serum level of angiostatin above control levels. This phenomenon occurred in tumor-bearing and nontumor-bearing animals. The effects of amifostine on matrix metalloproteinase (MMP) enzymatic activity was also determined using gelatin zymography. Conditioned growth medium collected from Sa-NH cells grown to confluency was exposed to various concentrations of SH, i.e., 2-[(aminopropyl)amino]ethane-thiol (WR-1065), the active thiol form of amifostine, for either 30 min or 18 hr. WR-1065, as a function of increasing dose and time, inhibited the enzymatic activities of MMP-2 and MMP-9. At a concentration and time of exposure likely to be achieved in vivo, that is, 40 microM and 30 min, MMP-2 and MMP-9 activities were reduced to between 30% and 40% of control values. Consistent with these affects, WR-1065 was also found to be effective in inhibiting the ability of Sa-NH cells to migrate through Matrigel membranes. After an 18-hr exposure under in vitro conditions, WR-1065 at concentrations of 4, 40 and 400 microM, and 4 mM, inhibited Sa-NH migration to 11%, 44%, 81% and 97% of control values, respectively. The abilities of amifostine and its active thiol WR-1065 to stimulate angiostatin production in mice, and to inhibit the MMP enzymatic activities and invasion ability of Sa-NH cells under in vitro conditions, are consistent with the observed antimetastatic effects exhibited against Sa-NH tumors growing in vivo.

Amifostine↗
PubMed2002

Combined gene therapy and ionizing radiation is a novel approach to treat human esophageal adenocarcinoma.

BACKGROUND: The ability to infect tumor cells limits the antitumor effects of gene therapy. The addition of radiotherapy to treatment with Ad.Egr.TNF.11D, a replication-deficient adenovirus containing a radiation-inducible promoter, early growth response-1, and the tumor necrosis factor-alpha (TNFalpha) complementary DNA may enhance the therapeutic ratio. METHODS: Seg-1 human esophageal adenocarcinoma cells were treated with Ad.Egr.TNF.11D with or without radiation. TNFalpha levels were quantified with enzyme-linked immunosorbent assay. Athymic nude mice bearing Seg-1 tumors were randomized to buffer, ionizing radiation, Ad.Egr.TNF.11D, and combination therapy. Tumor growth delay was used to compare treatment regimens. TNFalpha levels were measured in tumor homogenates and plasma. RESULTS: Seg-1 cells treated with Ad.Egr.TNF.11D and ionizing radiation demonstrated increased TNFalpha levels at 72 hours compared with cells exposed to vector alone (124 +/- 0 pg/mL vs. 31.11 +/- 22 pg/mL; P =.008). In vivo, Ad.Egr.TNF.11D-treated tumors expressed low TNFalpha levels (151.5 +/- 107.11 pg/mg protein) compared with tumors receiving combined treatment (793.92 +/- 489.13 pg/mg protein; P =.067). Increased TNFalpha levels were associated with increased tumor growth delay after combined treatment (P <.05). CONCLUSIONS: Radiotherapy enables focal stimulation of TNFalpha expression in Ad.Egr.TNF.11D-infected cells and thus improves local tumor control.

Adenocarcinoma↗
PubMed2002

Radiation-induced tumour necrosis factor-alpha expression: clinical application of transcriptional and physical targeting of gene therapy.

Promising data are emerging on a new anticancer agent, Ad.EGR-TNF, an adenoviral vector, which contains radio-inducible DNA sequences from the early growth response (EGR1) gene promoter and cDNA for the gene encoding human tumour necrosis factor-alpha. Ad.EGR-TNF combines the well-documented broad-spectrum anticancer activity of TNFalpha with the proven clinical usefulness of radiotherapy. Systemic delivery of the TNFalpha protein has had limited success clinically because of severe dose-limiting toxic effects. This limitation has been overcome by the use of a gene delivery approach, combined with a radiation-inducible promoter to express the TNFalpha protein in the irradiated tumour tissue. Preclinical and early phase I clinical testing indicates that effective concentrations of TNFalpha can be delivered to the tumour site without significant systemic exposure or toxic effects. The combination of radiation and TNFalpha gene delivery has produced striking antitumour effects in model systems in animals. In the clinical setting, potent anticancer activity has been observed with a high rate of complete and partial objective tumour responses. A novel mechanism of destruction of the tumour vasculature seems to be central to this distinct antitumour activity. This review summarises the rationale, mechanistic basis, preclinical data, and preliminary clinical findings for this new treatment model.

Clinical Trials as Topic↗
PubMed2002

Antimetastatic effectiveness of amifostine therapy following surgical removal of Sa-NH tumors in mice.

The effects of dose per fraction on the ability of amifostine exposure to elevate angiostatin levels in the serum of mice and to inhibit spontaneous metastases formation using the well-characterized murine Sa-NH sarcoma were investigated. Amifostine was administered intraperitoneally at doses of 50, 100, or 200 mg/kg every other day for 6 days to C3Hf/Kam mice until tumors reached an average size of 8 mm in diameter. Amifostine was again administered immediately following surgical removal of the tumor-bearing limbs by amputation, and then once more 2 days later. Nontumor-bearing control animals were treated using the same dosing and surgery schedules. The average number of pulmonary metastases per animal was determined for each experimental group. A significant reduction (P <.05) in the average number of pulmonary metastases was observed only in the group of animals exposed to a dose per fraction of 50 mg/kg. A dose of 100 mg/kg was less effective while 200 mg/kg had no effect on metastases formation in this study. The effects of amifostine exposure on serum levels of the angiogenesis inhibitor angiostatin were also determined using Western analysis. Correlating with the antimetastatic effect measured, exposure of animals to 50 mg/kg of amifostine resulted in a four-fold enhanced serum level of angiostatin above control levels. This phenomenon occurred in both tumor-bearing as well as nontumor-bearing animals. In contrast, a dose of 200-mg/kg amifostine administered intraperitoneally under these conditions had no measurable effect on angiostatin serum levels in this animal system. The enhanced ability of relatively low doses of amifostine to inhibit spontaneous metastases formation suggests that effective antimetastatic therapies with amifostine can be designed with minimal toxic side effects. While the dose responses for angiostatin production and metastases inhibition by amifostine are well correlated, the precise mechanism of action underlying these phenomena is unclear but is suggestive of a redox driven process(es).

Amifostine↗
PubMed2002

Transcriptional control of viral gene therapy by cisplatin.

Ionizing radiation (IR) and radical oxygen intermediates (ROIs) activate the early growth response-1 (Egr1) promoter through specific cis-acting sequences termed CArG elements. Ad.Egr.TNF.11D, a replication-deficient adenoviral vector containing CArG elements cloned upstream of the cDNA for human recombinant TNF-alpha was used to treat human esophageal adenocarcinoma and rat colon adenocarcinoma cells in culture and as xenografts in athymic nude mice. Cisplatin, a commonly used chemotherapeutic agent, causes tumor cell death by producing DNA damage and generating ROIs. The present studies demonstrate induction of TNF-alpha production in tumor cells and xenografts treated with the combination of Ad.Egr.TNF.11D and cisplatin. The results show that the Egr1 promoter is induced by cisplatin and that this induction is mediated in part through the CArG elements. These studies also demonstrate an enhanced antitumor response without an increase in toxicity following treatment with Ad.Egr.TNF.11D and cisplatin, compared with either agent alone. Chemo-inducible cancer gene therapy thus provides a means to control transgene expression while enhancing the effectiveness of commonly used chemotherapeutic agents.

Adenocarcinoma↗
PubMedDate not supplied

Relevance and irrelevance of DNA damage response to radiotherapy.

Ionizing radiation (IR) has been used to treat human malignancies since the early part of the 20th century. To date, most of the advances in radiotherapy have focused on optimization of treatment delivery schedules and technologic improvements in the physical targeting of dose. By comparison, many of the discoveries regarding the molecular basis of DNA damage and repair have not yet been translated to clinical practice. This article offers some perspectives regarding modulators of radiation effects and the challenges faced as we approach newer molecular targets. Our goal is to frame the issues that contribute to the apparent disconnect between laboratory discoveries and improvements in clinically relevant therapeutics.

DNA↗
PubMedDate not supplied

Vascular endothelial growth factor enhances endothelial cell survival and tumor radioresistance.

PURPOSE: Vascular endothelial growth factor (VEGF) is an important mediator of endothelial cell proliferation and survival. The purpose of the present studies was to investigate the role of VEGF in the tumor response to ionizing radiation. METHODS: Two ras-transformed murine fibrosarcoma cell lines, VEGF+/+ and VEGF-/- were exposed to ionizing radiation (0, 1, 3, 5, 7 or 9 Gy) in vitro, and clonogenic survival was determined. VEGF+/+ and VEGF-/- xenografts were generated in athymic nude mice and then treated with ionizing radiation (ten 5-Gy fractions = 50 Gy). Mean fractional tumor volume was used to evaluate treatment efficacy. To determine whether VEGF enhances tumor radioresistance by targeting endothelial cells, we performed clonogenic survival assays with human umbilical vein endothelial cells. Surviving fractions were calculated after treatment with ionizing radiation (5 Gy) and recombinant hVEGF165 (0, 1, 10, and 100 ng/mL). To determine whether VEGF neutralization enhances tumor radiosensitivity, we employed anti-VEGF165 monoclonal antibody to treat human tumor xenografts. Tumors were exposed to ionizing radiation (four 5-Gy fractions = 20 Gy) and treated with anti-VEGF antibody (0, 5, and 25 microg/kg in four intraperitoneal doses). Mean fractional tumor volume was used to evaluate treatment efficacy. To elucidate the molecular mechanism contributing to the observed anti-VEGF/ionizing radiation interaction, we exposed human umbilical vein endothelial cells to ionizing radiation (5 Gy) in the presence of anti-VEGF antibody (1 microg/mL). Sodium dodecyl sulfate polyacrylamide gel electrophoresis of cell lysates was probed for mitogen-activated protein kinase (MAPK) and MAPK kinase (MEK1/MEK2). RESULTS: The in vitro radiosensitivities of the VEGF+/+ and VEGF-/- clones were equivalent (D0 = 146 vs 149). However, the VEGF+/+ xenografts were more resistant to the cytotoxic effects of ionizing radiation than the VEGF-/- xenografts. VEGF+/+ xenografts demonstrated a faster doubling time (4.5 vs 6.0 days) and a shorter growth delay (15 vs 23 days) than VEGF-/- xenografts. The surviving fraction of human umbilical vein endothelial cells after exposure to ionizing radiation was significantly enhanced in the presence of VEGF (6.4% vs 12.5%). Western blot analysis demonstrated that stimulation of MAPK and MEK1/MEK2 was abrogated after exposure to anti-VEGF antibody. DISCUSSION: These findings represent the first genetic evidence that factors other than inherent tumor cell radiosensitivity are important determinants of radiocurability. Antitumor strategies targeting VEGF and other endothelial cell survival mechanisms may be used to enhance the cytotoxic effects of radiotherapy.

Animals↗