2003 Effect of the Administration of Lovastatin on the Development of Pulmonary Fibrosis Following WholeLung Irradiation in a Mouse Model

J.P. Williams1, E. Hernady1, C. Johnston2, C. Reed2, P. Okunieff1, J.N. Finkelstein2

1Department of Radiation Oncology, University of Rochester Medical Center, Rochester, NY, 2Departments of Pediatrics &Neonatology, University of Rochester Medical Center, Rochester, NY

Purpose/Objective: Pulmonary pneumonitis and fibrosis have long been known to represent the spectrum of expression of lunginjury in response to ionizing radiation. Our approach to the investigation of the late radiation response in the lung is governedby the overall hypothesis that repair and regeneration are the consequences of multicellular interactions and a network ofinteracting mediators. The specific hypothesis addressed in this study is that radiation-induced pulmonary injury leads topersistent changes in parenchymal cell cytokine and chemokine gene expression that leads to recruitment and activation ofmononuclear cells through the activation of the CCR2 chemokine receptor and expression of C-C chemokines, notablymonocyte chemoattractant protein-1 (MCP-1). Statins have been shown to reduce MCP-1 expression in a number of animalmodels. Because of this, statin could be useful in reducing monocyte recruitment.

Materials/Methods: Using our standard mouse (C57Bl/6) model for radiation-induced pulmonary effects, all treatment groupsreceived 15 Gy whole thorax irradiation. In one group, we administered lovastatin (10 �g/kg 3 � per week by gavage) to micestarting at the time of irradiation. In a second group of mice, statin treatment was delayed until 8 weeks post-radiation. A thirdgroup received radiation alone. Animals were sacrificed at 12, 16 and 24 weeks post-irradiation. At each selected interval, 5-6animals were sacrificed, subjected to exhaustive bronchopulmonary lavage and the cells processed for flow cytometricevaluation after immunostaining. In the remaining 5-6 animals, the lung tissue was assessed by RNase protection, immuno-histochemistry, ELISA and in situ hybridization, for mRNA and protein analysis. In addition, breathing rates were measuredas a means of assessing ongoing lung injury.

Results: Measurements of MCP-1 protein in lung homogenates showed that radiation-induced increases in tissue abundance ofMCP-1 protein were reduced by statin treatment (Table 1). It also appeared that both early and late treatment were effectivein reducing measured MCP-1 levels. This reduction correlated with an improved rate of survival and a decrease in breathingrates as compared to radiation alone at all time points.

Conclusions: Additional experiments with more animals over a longer period of time are needed to assess whether thistreatment would reduce cell recruitment and subsequent fibrosis. Nonetheless, statins may offer a potential treatment for theamelioration of late effects in breast and lung radiation therapy patients.

This work was supported by CA 11051-29.

Table. MCP-1 protein in lung homogenate at 16 weeks post-radiation

Treatment MCP-1

Sham-treated 9.5 � 415 Gy alone 53.7 � 1115 Gy � statin 22.7 � 415 Gy � statin (8 weeks) 32.5 � 3

2004 Potential Radiation Sensitizing Effect of SU5416 by Down-Regulating the COX-2 Expression in HumanLung Cancer Cells

J. Kim, D. Saha, H. Wu, Q. Cao, H. Choy

Department of Radiation Oncology, Vanderbilt University Medical Center, Nashville, TN

Purpose/Objective: Targeting tumor angiogenesis is a promising strategy of cancer treatment because angiogenesis is anessential part of tumor growth. We evaluated the role of the novel anti-angiogenic drug SU5416, inhibitor of VEGF receptor-2(Flk-1/KDR), as a potential radiosensitizer and its potential mechanism.

Materials/Methods: For immunoblot analysis, human large cell lung cancer cells (NCI-H460) were grown in RPMI media,serum-starved for 24 h and treated with increasing doses of SU5416. Cells were lysed 24 h after the treatment with SU 5416and analyzed for COX-1 and COX-2 expressions. In some experiments cells were treated with SU5416 for 1 h prior to theaddition of phorbol 12-myristate 13-acetate (PMA) to inhibit the induction of COX-2 expression. For northern blot analysis,total RNA was isolated from H460 cell monolayers using Tri reagent and phase lock gel tubes under similar conditions. Wealso conducted a endothelial cell migration assay using the serum collected from the animals after treatment with SU5416 plusradiation. For in vivo studies, H460 cell xenografts were used in four groups of athymic nude mice; 1) DMSO (vehicle) alone,2) radiation alone (2 Gy/day, days 0-4), 3) SU5416 alone (25 mg/kg/day, days 0-4), 4) SU5416 plus radiation. Tumor growthdelay was then measured for each treatment group at a regular interval for two months and an enhancement factor was thendetermined. Tumor tissues from each group were also subjected to immunoblot analysis for COX-2 expression.

Results: SU5416 caused a dose-dependent decrease of endogenous COX-2 expression in H460 cells as well as in other humanlung carcinoma cell lines. However, COX-1 expression remained unchanged under similar conditions. SU5416 also preventedthe PMA in induced COX-2 expression in early hours (2-4 h). PMA-induced COX-2 transcription was also suppressed bySU5416. We have also demonstrated the similar results using another VEGF receptor-2 inhibitor SU6668. In our in vivo studies,SU5416 in combination with radiation demonstrated an enhancement of tumor growth delay when compared with either drugor radiation alone with an enhancement factor of 1.41. We also observed that SU5416 plus radiation significantly down-regulated COX-2 expression in the tumors compared to control. A higher degree of prevention of migration was observed withthe serum obtained from the animals treated with the combination of SU5416 and radiation.

215Proceedings of the 44th Annual ASTRO Meeting