The chemokine axis CCR6/CCL20 is involved in cancer progression in a variety of tumors. Here, we show that CCR6 is expressed by melanoma cells. The CCR6 ligand, CCL20, induces migration and proliferation and enhances tumor growth and metastasis Confocal analysis of melanoma tissues showed that CCR6 is expressed by tumor cells, whereas CCL20 is preferentially expressed by nontumoral cells in the stroma of certain tumors. Stromal CCL20, but not tumoral CCR6, predicted poor survival in a cohort of 40 primary melanoma patients. Tumor-associated macrophages (TAM), independently of their M1/M2 polarization profile, were identified as the main source of CCL20 in primary melanomas that developed metastasis. In addition to CCL20, TAMs expressed TNF and VEGF-A protumoral cytokines, suggesting that melanoma progression is supported by macrophages with a differential activation state. Our data highlight the synergistic interaction between melanoma tumor cells and prometastatic macrophages through a CCR6/CCL20 paracrine loop. Stromal levels of CCL20 in primary melanomas may be a clinically useful marker for assessing patient risk, making treatment decisions, and planning or analyzing clinical trials.

The type I interferon (TI-IFN) pathway regulates innate immunity, inflammation, and apoptosis during infection. However, the contribution of the TI-IFN pathway or upstream signaling pathways to tubular injury in kidney disease is poorly understood. Upon observing evidence of activation of upstream regulators of the TI-IFN pathway in a transcriptomics analysis of murine kidney tubulointerstitial injury, we have now addressed the impact of the TI-IFN and upstream signaling pathways on kidney tubulointerstitial injury. In cultured tubular cells and kidney tissue, IFNα/β binding to IFNAR activated the TI-IFN pathway and recruited antiviral interferon-stimulated genes (ISG) and NF-κB-associated proinflammatory responses. TWEAK and lipopolysaccharide (LPS) signaled through TBK1/IKKε and IRF3 to activate both ISGs and NF-κB. In addition, TWEAK recruited TLR4 to stimulate TBK1/IKKε-dependent ISG and inflammatory responses. Dual pharmacological inhibition of TBK1/IKKε with amlexanox decreased TWEAK- or LPS-induced ISG and cytokine responses, as well as cell death induced by a complex inflammatory milieu that included TWEAK. TBK1 or IRF3 siRNA prevented the TWEAK-induced ISG and inflammatory gene expression while IKKε siRNA did not. In vivo, kidney IFNAR and IFNβ were increased in murine LPS and folic acid nephrotoxicity while IFNAR was increased in human kidney biopsies with tubulointerstitial damage. Inhibition of TBK1/IKKε with amlexanox or IFNAR neutralization decreased TI-IFN pathway activation and protected from kidney injury induced by folic acid or LPS. In conclusion, TI-IFNs, TWEAK, and LPS engage interrelated proinflammatory and antiviral responses in tubular cells. Moreover, inhibition of TBK1/IKKε with amlexanox, and IFNAR targeting, may protect from tubulointerstitial kidney injury.

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<p>Supplementary Table S1. List of primary antibodies and primer sequences.</p>

<p>Supplementary Table S1. List of primary antibodies and primer sequences.</p>

<p>Supplementary Figure S1. CCR6 expression, melanoma xenograft model and in vivo effects of exogenous CCL20.</p>

<p>Supplementary Figure S2. TAM CD14+/CD163+ are the major source of CCL20 in human melanoma.</p>

<p>Supplementary Table S2. Melanoma patient characteristics.</p>