Journal Article

Mutant K-<i>ras</i><sup>V12</sup> increases COX-2, peroxides and DNA damage in lung cells

Anna Maciag, Gunamani Sithanandam and Lucy M. Anderson

in Carcinogenesis

Volume 25, issue 11, pages 2231-2237
Published in print November 2004 | ISSN: 0143-3334
Published online November 2004 | e-ISSN: 1460-2180 | DOI: http://dx.doi.org/10.1093/carcin/bgh245
Mutant K-rasV12 increases COX-2, peroxides and DNA damage in lung cells

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K-ras is frequently mutated in lung adenocarcinomas. Recent discovery that wild-type K-ras is tumor suppressive in the lung raises a question: how is mutant K-ras aggressively oncogenic? We hypothesized that mutant K-ras might lead to generation of reactive oxygen species (ROS) and DNA damage, contributing to malignant transformation. We stably transfected human mutant K-rasV12 into non-transformed peripheral mouse lung epithelial cells (E10 line). Constitutively active mutant K-rasV12 in E10 cells led to a highly significant (P < 0.001) increased level of peroxides, and a corresponding increase in the amount of DNA strand-break damage, compared with the parental line E10 and the vector control. Levels of superoxide were not increased, suggesting a direct source of peroxides, such as cyclooxygenase-2 (COX-2). COX-2 protein and activity measured as prostaglandin E2 level were up-regulated in cells expressing mutant K-rasV12; COX-2 activity correlated with K-ras activity (K-ras p21-GTP). Both peroxide generation and DNA single strand breaks were significantly reduced by pre-treatment with COX-2-specific inhibitor SC 58125, confirming COX-2 as the source of the ROS. COX-2 has been repeatedly implicated in lung cancer, and is known to be regulated by ras and to release ROS. Our data suggest that up-regulation of COX-2, with a consequent increase in peroxides and DNA damage, contributes to the dominant oncogenicity of mutant K-ras.

Keywords: CM-H2DCF-DA, 5,6-chloromethyl-2′,7′-dichlorodihydrofluorescein diacetate; COX, cyclooxygenase; DCF, 2′,7′-dichlorofluorescein; HBSS, Hanks' balanced salt solution; NBT, nitroblue tetrazolium; PGE2, prostaglandin E2; ROS, reactive oxygen species

Journal Article.  4373 words.  Illustrated.

Subjects: Clinical Cytogenetics and Molecular Genetics

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