IL-36γ drives skin toxicity induced by EGFR/MEK inhibition and commensal Cutibacterium acnes
Takashi K. Satoh, … , Emmanuel Contassot, Lars E. French
Takashi K. Satoh, … , Emmanuel Contassot, Lars E. French
Published December 5, 2019
Citation Information: J Clin Invest. 2020;130(3):1417-1430. https://doi.org/10.1172/JCI128678.
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Research Article Dermatology Inflammation

IL-36γ drives skin toxicity induced by EGFR/MEK inhibition and commensal Cutibacterium acnes

Abstract

Epidermal growth factor receptor (EGFR) and MEK inhibitors (EGFRi/MEKi) are beneficial for the treatment of solid cancers but are frequently associated with severe therapy-limiting acneiform skin toxicities. The underlying molecular mechanisms are poorly understood. Using gene expression profiling we identified IL-36γ and IL-8 as candidate drivers of EGFRi/MEKi skin toxicity. We provide molecular and translational evidence that EGFRi/MEKi in concert with the skin commensal bacterium Cutibacterium acnes act synergistically to induce IL-36γ in keratinocytes and subsequently IL-8, leading to cutaneous neutrophilia. IL-36γ expression was the combined result of C. acnes–induced NF-κB activation and EGFRi/MEKi–mediated expression of the transcription factor Krüppel-like factor 4 (KLF4), due to the presence of both NF-κB and KLF4 binding sites in the human IL-36γ gene promoter. EGFRi/MEKi increased KLF4 expression by blockade of the EGFR/MEK/ERK pathway. These results provide an insight into understanding the pathological mechanism of the acneiform skin toxicities induced by EGFRi/MEKi and identify IL-36γ and the transcription factor KLF4 as potential therapeutic targets.

Authors

Takashi K. Satoh, Mark Mellett, Barbara Meier-Schiesser, Gabriele Fenini, Atsushi Otsuka, Hans-Dietmar Beer, Tamara Rordorf, Julia-Tatjana Maul, Jürg Hafner, Alexander A. Navarini, Emmanuel Contassot, Lars E. French

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Figure 5

KLF4 is critical for IL-36γ transcriptional activity upon EGFR/MEK inhibition.

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KLF4 is critical for IL-36γ transcriptional activity upon EGFR/MEK inhib...
(A) KLF4-overexpressing primary keratinocytes were exposed to C. acnes for 24 hours. (B) Flag-tagged wild-type (WT) and dominant-negative (DN) KLF4 were overexpressed in response to doxycycline using a Tet-on system for 24 hours, followed by exposure to Pam3CSK4 for another 24 hours. The cell lysates were collected for Western blotting and quantitative PCR (qPCR). Data represent mean ± SEM (n = 3). (C) KLF4 siRNA–treated PHKs were exposed to erlotinib and C. acnes for 6 hours and IL36G levels were analyzed by qPCR. Data represent mean ± SEM (n = 3). (D) Keratinocyte cell lines in which KLF4 was knocked out by CRISPR/Cas9 were exposed to trametinib (2 μg/mL) for 24 hours and total cell lysates were collected for Western blotting with antibodies against KLF4 and β-actin. The cells were exposed to trametinib for 24 hours and isolated RNA was analyzed by qPCR. Data represent mean ± SEM (n = 3). All blots were run contemporaneously with the same protein samples. Data are representative of 3 independent experiments. (E) Mutations generated by CRISPR/Cas9 in the KLF4 binding site. Red nucleotides are the PAM sequence and blue nucleotides hybridize to the sgRNA. KLF4 binding site–mutant cells were exposed to trametinib and Pam3CSK4 for 24 hours. Total RNA was analyzed by qPCR. Data represent mean ± SEM (n = 3). Data were analyzed with 1-way ANOVA followed by Dunnett’s (B and E) or Tukey’s multiple-comparisons test (C) or with 2-tailed unpaired t test (D). **P < 0.01; ***P < 0.001.