Supplementary Figure 1-6, Supplementary Tables 1-3. Supplementary Table 1 Clinic pathologic characteristics of 54 cases of lung cancer patients; Supplementary Table 2 The number of mouse for TNFa expression quantitative analysis; Supplementary Table 3 TNFa is crucial for B[a]P-induced lung carcinogenic effect in vivo; Figure S1 miR-494 was not involved in B[a]P/B[a]PDE induced PHLPP2 downregulation; Figure S2 miR-205 regulated AP-1-mediated TNFa transcription; Figure S3 B[a]P induced lung inflammatory responses and carcinoma in wild-type and TNFa-/-mice; Figure S4 lung carcinoma formation in C57BL/6J wild type mice following B[a]P exposure; Figure S5 PHLPP2 downregulation in TNFa knockdown Beas-2B cells and C57BL/6J TNFa-/- mice following B[a]P/B[a]PDE exposure; Figure S6 Illustration of molecular mechanisms leading to Lung carcinogenesis following B[a]P exposure.
ARTICLE ABSTRACT
Purpose: The carcinogenic capacity of B[a]P/B[a]PDE is supported by epidemiologic studies. However, the molecular mechanisms responsible for B[a]P/B[a]PDE-caused lung cancer have not been well investigated. We evaluated here the role of novel target PHLPP2 in lung inflammation and carcinogenesis upon B[a]P/B[a]PDE exposure.Experimental Design: We used the Western blotting, RT-PCR, [35S]methionine pulse and immunohistochemistry staining to determine PHLPP2 downregulation following B[a]P/B[a]PDE exposure. Both B[a]PDE-induced Beas-2B cell transformation model and B[a]P-caused mouse lung cancer model were used to elucidate the mechanisms leading to PHLPP2 downregulation and lung carcinogenesis. The important findings were also extended to in vivo human studies.Results: We found that B[a]P/B[a]PDE exposure downregulated PHLPP2 expression in human lung epithelial cells in vitro and in mouse lung tissues in vivo. The ectopic expression of PHLPP2 dramatically inhibited cell transformation upon B[a]PDE exposure. Mechanistic studies showed that miR-205 induction was crucial for inhibition of PHLPP2 protein translation by targeting PHLPP2-3′-UTR. Interestingly, PHLPP2 expression was inversely associated with tumor necrosis factor alpha (TNFα) expression, with low PHLPP2 and high TNFα expression in lung cancer tissues compared with the paired adjacent normal lung tissues. Additional studies revealed that PHLPP2 exhibited its antitumorigenic effect of B[a]P/B[a]PDE through the repression of inflammatory TNFα transcription.Conclusions: Our studies not only first time identify PHLPP2 downregulation by lung carcinogen B[a]P/B[a]PDE, but also elucidate a novel molecular mechanisms underlying lung inflammation and carcinogenesis upon B[a]P/B[a]PDE exposure. Clin Cancer Res; 21(16); 3783–93. ©2015 AACR.
