COX-2 Induces IL-11 Production in Human Breast Cancer Cells 1 Balraj Singh, Ph.D., Jacob A. Berry, B.S., Angela Shoher, M.D., 2 and Anthony Lucci, M.D. 3 Department of Surgical Oncology, University of Texas, M.D. Anderson Cancer Center, Houston, Texas Submitted for publication August 25, 2005 Background. Cyclooxygenase-2 (COX-2) is overex- pressed in 40% of human invasive breast cancers. Interleukin-11 (IL-11), a potent mediator of osteoclas- togenesis, is involved in breast cancer metastasis to bone. Since breast cancers that overexpress COX-2 are associated with a higher rate of metastasis to bone, we hypothesized that COX-2 expression in tumor cells would induce IL-11. Materials and methods. We transfected MCF-7 (poorly metastatic) and MDA-231 (highly metastatic) human breast cancer cell lines with COX-2 expression vectors. COX-2 overexpression was confirmed by West- ern blot and PGE 2 immunoassay, and IL-11 production was measured by immunoassay. We also used a nude mouse model to study COX-2 and IL-11 production from breast cancer cells that metastasized to bone. The bone-seeking clones (BSC) were isolated and cul- tured from the long bone metastases. Results. COX-2 transfection caused an approxi- mately 5- to 6-fold increase in IL-11 production in both MCF-7 and MDA-231 cells. MDA-435S-COX2-BSC (cells isolated from bone metastasis) produced elevated lev- els of IL-11 and PGE 2 (an important mediator of COX-2) as compared to the parental MDA-435S-COX2 cells. Furthermore, a treatment with low 1- to 2-M concentration NS-398 or Celecoxib significantly re- duced the production of IL-11 in COX-2-transfected MDA-231 cells, thus confirming the involvement of COX-2 in IL-11 induction. Conclusion. COX-2-mediated production of IL-11 in breast cancer cells may be vital to the development of osteolytic bone metastases in patients with breast can- cer, and a COX-2 inhibitor may be useful in inhibiting this process. © 2006 Elsevier Inc. All rights reserved. Key Words: breast cancer; COX-2; IL-11; bone me- tastasis. INTRODUCTION Cyclooxygenase (COX) enzymes, COX-1 and COX-2, mediate the production of prostaglandins and throm- boxanes from arachidonic acid. Both these enzymes possess two activities, cyclooxygenase and peroxidase, that act sequentially, thus converting arachidonic acid to an intermediate prostaglandin G 2 (PGG 2 ) and then the product PGH 2 . PGH 2 is converted to several eico- sanoids, PGD 2 , PGE 2 , PGF 2 , PGI 2 , and thromboxane A 2 , by specific synthases. COX-1 is expressed constitu- tively, whereas COX-2 is induced in inflammation and cancer. The extracellular stimuli that induce COX-2 include growth factors, cytokines, tumor promoters, hypoxia, ionizing radiation, and carcinogens (reviewed in references [1–3]). There is strong evidence to implicate the role of COX-2 overexpression in breast cancer tumorigenesis. One study showed that 56% of infiltrating mammary carcinomas and intraductal carcinomas expressed sig- nificant levels of COX-2, while benign breast tissue at least 1 cm from a malignant lesion did not express COX-2 [4]. COX-1 expression was ubiquitous and did not differ significantly between benign and malignant tissues. In a murine model of metastatic breast cancer, PGE 2 levels are positively correlated with increased tumorigenic and metastatic potential [5]. Perhaps the most convincing evidence that COX-2 causes breast cancer in animals comes from transgenic mice in which the COX-2 was overexpressed in mammary tissue by using the mouse mammary tumor virus (MMTV) long- terminal repeat promoter. More than 85% of these mice 1 Presented at the 38th Annual Meeting of the Association of Academic Surgery in Houston, TX, 2004. These studies were sup- ported in part by the grants from the Wendy Will Case Cancer Fund, Inc., Methodist Hospital Foundation, and by Grants R21 DK067682 from the National Institutes of Health and DAMD17-03-1-0669 from the United States Army Medical Research and Material Command. 2 Present address: Johns Hopkins University School of Medicine, Baltimore, MD. 3 To whom correspondence and reprint requests should be addressed at the Department of Surgical Oncology, Unit 444, The University of Texas, M.D. Anderson Cancer Center, 1515 Holcombe Boulevard, Hous- ton, TX 77030. E-mail: alucci@mdanderson.org. Journal of Surgical Research 131, 267–275 (2006) doi:10.1016/j.jss.2005.11.582 267 0022-4804/06 $32.00 © 2006 Elsevier Inc. All rights reserved.