Reduction in IKB Kinase A Expression Promotes the Development of Skin Papillomas and Carcinomas Eunmi Park, Feng Zhu, Bigang Liu, Xiaojun Xia, Jianjun Shen, Tracie Bustos, Susan M. Fischer, and Yinling Hu Department of Carcinogenesis, The University of Texas M. D. Anderson Cancer Center, Smithville, Texas Abstract We reported recently a marked reduction in IKB kinase A (IKKA) expression in a large proportion of human poorly differentiated squamous cell carcinomas (SCC) and the occurrence of Ikka mutations in human SCCs. In addition, overexpression of IKKA in the epidermis inhibited the development of skin carcinomas and metastases in mice. However, whether a reduction in IKKA expression promotes skin tumor development is currently unknown. Here, we assessed the susceptibility of Ikka hemizygotes to chemical carcinogen-induced skin carcinogenesis. Ikka +/ mice devel- oped 2 times more papillomas and 11 times more carcino- mas than did Ikka +/+ mice. The tumors were larger in Ikka +/ than in Ikka +/+ mice, but tumor latency was shorter in Ikka +/ than in Ikka +/+ mice. Some of the Ikka +/ papillomas and most Ikka +/ carcinomas lost the remaining Ikka wild-type allele. Somatic Ikka mutations were detected in carcinomas and papillomas. The chemical carcinogen-induced H-Ras mutations were detected in all the tumors. The phorbol ester tumor promoter induced higher mitogenic and angiogenic activities in Ikka +/ than in Ikka +/+ skin. These elevated activities were intrinsic to keratinocytes, suggesting that a reduction in IKKA expression provided a selective growth advantage, which cooperated with H-Ras mutations to promote papilloma formation. Furthermore, excessive extracellular signal-regulated kinase and IKK kinase activ- ities were observed in carcinomas compared with those in papillomas. Thus, the combined mitogenic, angiogenic, and IKK activities might contribute to malignant conversion. Our findings provide evidence that a reduction in IKKA expression promotes the development of papillomas and carcinomas and that the integrity of the Ikka gene is required for suppressing skin carcinogenesis. [Cancer Res 2007;67(19):9158–68] Introduction We recently reported somatic IjB kinase a (Ikka) mutations in human squamous cell carcinomas (SCC) and a marked reduction in IKKa expression in poorly differentiated human and mouse cutaneous SCCs (1), which highlights the importance of IKKa in human skin cancer. However, the natural role for IKKa in skin tumor development is unclear. The animal model provides an appropriate tool to address these questions. IKKa is one subunit of the IKK complex, which is central for nuclear factor-nB (NF-nB) activation (2). Its involvement in the development of lymphoid organs and innate immunity requires IKK/NF-nB activity (3, 4). IKKa also plays an essential role in the formation of the epidermis during embryonic development in mice (5–7). Ikka / mice develop a striking hyperplastic epidermis that lacks terminal differentiation and these mice die at birth (5–7). Ikka / keratinocytes and skin preserve IKK/NF-nB activity (5, 8). Reintroduction of IKKa or kinase-inactive IKKa induced terminal differentiation in Ikka / keratinocytes, but reintroduction of IKKh, RelA p65, or InBa (an inhibitor for NF-nB) failed to do so (8). Furthermore, Sil et al. (9) reported that IKKa or kinase-inactive IKKa transgene driven by the keratin 14 promoter rescued the skin phenotype of Ikka / mice. These results suggest that the function of IKKa in determining the epidermal formation is IKK/NF-nB independent. We reported recently that Lori.IKKa transgenic mice developed normal skin and that the elevated IKKa in the suprabasal epidermis enhanced terminal differentiation markers (1). These transgenic mice developed significantly fewer malignant carcino- mas and metastases than did wild-type (WT) mice when they were challenged with the chemical carcinogen 7,12-dimethylben- z( a )anthracene (DMBA) and tumor promoter 12-O -tetradecanoyl- phorbol-13-acetate (TPA). The mitogenic and angiogenic activities were reduced in the DMBA/TPA–treated skin of Lori.IKKa mice relative to those in the DMBA/TPA–treated skin of WT mice. These results suggest that elevated IKKa expression antagonizes chemical carcinogen-induced mitogenesis and angiogenesis, thereby repres- sing the development of malignant carcinomas and metastases. However, the effect of reduced IKKa on skin tumor development is unknown. Ras plays a prominent role in the development of human SCCs and H-Ras mutations are frequently found in human SCCs (10, 11). Ras activation is required for chemical carcinogen-induced skin carcinogenesis in mice. The carcinogen DMBA causes activating H-Ras mutations, and the tumor promoter TPA expands the population of Ras-initiated cells (12, 13). Most papillomas eventually regress, and only a few become carcinomas in mice with a C57BL6 or a C56BL/129/Sv background (14, 15). Mice lacking H-Ras developed significantly fewer skin tumors than did WT mice induced by DMBA/TPA (16). In the present study, we tested the susceptibility of Ikka hemizygotes to DMBA/TPA–induced skin carcinogenesis. Ikka +/ mice developed 2 times more benign tumors (papillomas) and 11 times more malignant carcinomas than did WT mice. Furthermore, we found that most Ikka +/ carcinomas and some Ikka +/ papillomas lost the remaining WT Ikka allele. These findings show that reduction in IKKa expression promotes papilloma formation and malignant conversion. Note: Supplementary data for this article are available at Cancer Research Online (http://cancerres.aacrjournals.org/). E. Park and F. Zhu contributed equally to this work. Requests for reprints: Yinling Hu, Department of Carcinogenesis, The University of Texas M. D. Anderson Cancer Center, 1808 Park Road 1-C, Smithville, TX 78957. Phone: 512-237-9338; Fax: 512-237-4375; E-mail: yhu@mdanderson.org. I2007 American Association for Cancer Research. doi:10.1158/0008-5472.CAN-07-0590 Cancer Res 2007; 67: (19). October 1, 2007 9158 www.aacrjournals.org Research Article Research. on March 3, 2016. © 2007 American Association for Cancer cancerres.aacrjournals.org Downloaded from