Androgen receptor functions as transcriptional repressor of cancer-associated fibroblast activation
Andrea Clocchiatti, Soumitra Ghosh, Maria-Giuseppina Procopio, Luigi Mazzeo, Pino Bordignon, Paola Ostano, Sandro Goruppi, Giulia Bottoni, Atul Katarkar, Mitchell Levesque, Peter Kölblinger, Reinhard Dummer, Victor Neel, Berna C. Özdemir, G. Paolo Dotto
Andrea Clocchiatti, Soumitra Ghosh, Maria-Giuseppina Procopio, Luigi Mazzeo, Pino Bordignon, Paola Ostano, Sandro Goruppi, Giulia Bottoni, Atul Katarkar, Mitchell Levesque, Peter Kölblinger, Reinhard Dummer, Victor Neel, Berna C. Özdemir, G. Paolo Dotto
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Research Article Dermatology Oncology

Androgen receptor functions as transcriptional repressor of cancer-associated fibroblast activation

Abstract

The aging-associated increase of cancer risk is linked with stromal fibroblast senescence and concomitant cancer-associated fibroblast (CAF) activation. Surprisingly little is known about the role of androgen receptor (AR) signaling in this context. We have found downmodulated AR expression in dermal fibroblasts underlying premalignant skin cancer lesions (actinic keratoses and dysplastic nevi) as well as in CAFs from the 3 major skin cancer types, squamous cell carcinomas (SCCs), basal cell carcinomas, and melanomas. Functionally, decreased AR expression in primary human dermal fibroblasts (HDFs) from multiple individuals induced early steps of CAF activation, and in an orthotopic skin cancer model, AR loss in HDFs enhanced tumorigenicity of SCC and melanoma cells. Forming a complex, AR converged with CSL/RBP-Jκ in transcriptional repression of key CAF effector genes. AR and CSL were positive determinants of each other’s expression, with BET inhibitors, which counteract the effects of decreased CSL, restoring AR expression and activity in CAFs. Increased AR expression in these cells overcame the consequences of CSL loss and was by itself sufficient to block the growth and tumor-enhancing effects of CAFs on neighboring cancer cells. As such, the findings establish AR as a target for stroma-focused cancer chemoprevention and treatment.

Authors

Andrea Clocchiatti, Soumitra Ghosh, Maria-Giuseppina Procopio, Luigi Mazzeo, Pino Bordignon, Paola Ostano, Sandro Goruppi, Giulia Bottoni, Atul Katarkar, Mitchell Levesque, Peter Kölblinger, Reinhard Dummer, Victor Neel, Berna C. Özdemir, G. Paolo Dotto

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Figure 6

AR represses TP53 gene activation.

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AR represses TP53 gene activation. (A) Immunoblot analysis of p53 expres...
(A) Immunoblot analysis of p53 expression in 2 HDF strains infected with AR-silencing lentiviruses versus control vector. Blots were sequentially probed with antibodies against phospho–Ser15 p53, AR, p53, and β-actin. (B) RT-qPCR analysis of TP53 expression in 3 HDF strains infected with AR-silencing lentiviruses versus control vector. Values for each strain are indicated as dots with mean ± SD. n(HDF strains) = 3; *P < 0.05, 1-way ANOVA with Dunnett’s test. (C) ChIP at TP53 promoter, using anti-AR antibodies in parallel with nonimmune IgGs. Data are expressed as relative fold enrichment over IgGs, and values for each strain are indicated as dots with mean ± SD. Statistical significance was calculated between AR enrichment at the indicated sites relative to the flanking control region. n(HDF strains) = 3; *P < 0.05, ***P < 0.005, 1-way ANOVA with Dunnett’s test. (D) EdU labeling assays of 3 HDF strains stably infected with a TP53-silencing retrovirus (sh TP53) versus control vector (sh CT), with or without subsequent AR gene silencing for 10 days, scoring at least 200 cells per condition. Values for each strain are indicated as dots with mean ± SD. Statistical significance was calculated on differences in changes caused by AR silencing in cells with versus without TP53 silencing or in cells with versus without concomitant AR and TP53 silencing. n(HDF strains) = 3; ***P < 0.005, 1-way ANOVA with Dunnett’s test. (E) RT-qPCR analysis of the indicated genes in 3 HDF strains infected as in D with a TP53-silencing retrovirus (sh TP53) versus control vector (sh CT), with or without subsequent AR gene silencing. Values for each strain are indicated as dots with mean ± SD. n(HDF strains) = 3; *P < 0.05, **P < 0.01, ***P < 0.005, 2-tailed unpaired t test.