LRRN4 and UPK3B Are Markers of Primary Mesothelial Cells Mutsumi Kanamori-Katayama 1 , Ai Kaiho1, Yuri Ishizu1, Yuko Okamura-Oho 2 , Okio Hino 3 , Masaaki Abe3, Takumi Kishimoto4, Hisahiko Sekihara5, Yukio Nakamura6, Harukazu Suzuki1, Alistair R. R. Forrest1*, Yoshihide Hayashizaki1* 1 OMICs Science Center, RIKEN Yokohama Institute, Yokohama, Kanagawa, Japan, 2 Advanced Computational Sciences Department, RIKEN Advanced Science Institute, Wako-shi, Saitama, Japan, 3 Department of Pathology and Oncology, Juntendo University School of Medicine, Tokyo, Japan, 4 Department of Respiratory Medicine, Japan Labor Health and Welfare Organization Okayama Rosai Hospital, Okayama, Japan, 5 Japan Labour Health and Welfare organization, Kawasaki, Japan, 6 Cell Engineering Division, RIKEN BioResource Center, Tsukuba, Ibaraki, Japan Abstract Background: Mesothelioma is a highly malignant tumor that is primarily caused by occupational or environmental exposure to asbestos fibers. Despite worldwide restrictions on asbestos usage, further cases are expected as diagnosis is typically 20– 40 years after exposure. Once diagnosed there is a very poor prognosis with a median survival rate of 9 months. Considering this the development of early pre clinical diagnostic markers may help improve clinical outcomes. Methodology: Microarray expression arrays on mesothelium and other tissues dissected from mice were used to identify candidate mesothelial lineage markers. Candidates were further tested by qRTPCR and in-situ hybridization across a mouse tissue panel. Two candidate biomarkers with the potential for secretion, uroplakin 3B (UPK3B), and leucine rich repeat neuronal 4 (LRRN4) and one commercialized mesothelioma marker, mesothelin (MSLN) were then chosen for validation across a panel of normal human primary cells, 16 established mesothelioma cell lines, 10 lung cancer lines, and a further set of 8 unrelated cancer cell lines. Conclusions: Within the primary cell panel, LRRN4 was only detected in primary mesothelial cells, but MSLN and UPK3B were also detected in other cell types. MSLN was detected in bronchial epithelial cells and alveolar epithelial cells and UPK3B was detected in retinal pigment epithelial cells and urothelial cells. Testing the cell line panel, MSLN was detected in 15 of the 16 mesothelioma cells lines, whereas LRRN4 was only detected in 8 and UPK3B in 6. Interestingly MSLN levels appear to be upregulated in the mesothelioma lines compared to the primary mesothelial cells, while LRRN4 and UPK3B, are either lost or down-regulated. Despite the higher fraction of mesothelioma lines positive for MSLN, it was also detected at high levels in 2 lung cancer lines and 3 other unrelated cancer lines derived from papillotubular adenocarcinoma, signet ring carcinoma and transitional cell carcinoma. Citation: Kanamori-Katayama M, Kaiho A, Ishizu Y, Okamura-Oho Y, Hino O, et al. (2011) LRRN4 and UPK3B Are Markers of Primary Mesothelial Cells. PLoS ONE 6(10): e25391. doi:10.1371/journal.pone.0025391 Editor: Sumitra Deb, Virginia Commonwealth University, United States of America Received July 19, 2011; Accepted September 2, 2011; Published October 3, 2011 Copyright: 2011 Kanamori-Katayama et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was funded by a Research Grant from the Japanese Ministry of Education, Culture, Sports, Science and Technology to the RIKEN Omics Science Center to Yoshihide Hayashizaki. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: alistair.forrest@gmail.com (ARRF); rgscerg@gsc.riken.jp (YH) Introduction Mesothelioma is one of the most fatal and difficult to treat cancers in the world and in Japan the median survival rate from diagnosis is 9 months with 94% of patients not surviving at 3 years [1]. In recent years the incidence of diagnosed cases of mesothelioma in Japan has doubled from 500 cases in 1995 to 1068 in 2007. The majority of mesothelioma cases can be directly linked to asbestos exposure and is estimated to be responsible for at least 90% of all cases [2]. Asbestos has long been used for its insulating properties and was used for insulation around pipes, inside ceiling cavities, brake linings and compounded into asbestos composite sheeting for an inexpensive heat and fire resistant building material. In the 1960s it was recognized that occupational asbestos exposure was a strong risk factor for the development of mesothelioma [3,4,5]. At particular risk were asbestos mine workers at locations such as Wittenoom Western Australia [6] and Western Cape South Africa [5]. The realization that Asbestos exposure was responsible led to legislation in multiple countries which progressively reduced the use of asbestos and potential exposure and finally led to complete bans in 52 countries [7]. Despite this, the past large scale use of asbestos means that there is still a large amount of it in the general community that needs to be safely removed, with demolition workers likely to be at risk. The latent period between exposure and diagnosis is in the order of 20– 40 years meaning that workers who were exposed a long time ago may still develop the disease [8]. Considering these two factors, the number of mesothelioma cases is likely to continue to rise at least for the next 20 years. PLoS ONE | www.plosone.org 1 October 2011 | Volume 6 | Issue 10 | e25391