CCR6+ B lymphocytes responding to tumor cell-derived CCL20 support hepatocellular carcinoma progression via enhancing angiogenesis.

BACKGROUND & AIMS: Different immune cells in tumor microenvironment shape tumor progression. CCL20 over-expression was reported as one of the "stemness" trait in TP53 mutated hepatocellular carcinoma (HCC). We aimed to understand the effect of CCL20 on HCC progression. METHODS: In two HCC cohort patients (n=95, n=85 respectively), serum CCL20 concentration was quantified by using ELISA. Expressions of CCL20 and CCR6 in 41 paired HCC tumor and adjacent non-tumor tissues were determined by quantitative Real-Time PCR, confirmed by immunohistochemistry (CCL20) or by flow cytometry analysis (CCR6). Chemotaxis of splenocytes or purified CD19+ B cells to tumor cell-derived CCL20, and angiogenesis of different CD19+ B subtypes responding to tumor cell-derived CCL20 were measured in vitro. H22 murine hepatoma cells were inoculated into immunocompetent or immunodeficient SCID mice, tumor growth and metastasis were monitored after the mice were treated with anti-CCL20 neutralizing antibody or depleted B cells by anti-CD20. RESULTS: Elevation of pretherapy serum CCL20 in HCC patients and increase of CCR6 expression in HCC tissues were closely associated with tumor metastasis and disease poor prognosis. In HCC tissues, CCL20 expression was positively correlated with CCR6 (R2 =0.3134, P=0.0002), and CCR6 was exclusively identified in tumor infiltrated immune cells. CD19+CD5+ B lymphocytes expressed higher CCR6, responded to tumor cell-derived CCL20 and enhanced angiogenesis in vitro. Neutralizing CCL20 activity in immunocompetent mice, not in SCID mice, attenuated tumor incidence, restrained tumor growth and distal metastasis. Tumor angiogenesis was significantly inhibited after CCL20 activity was blockade. In addition, inhibiting B lymphocyte infiltration into tumor mileum also attenuated tumor growth. CONCLUSIONS: Tumor cell-derived CCL20 interacts with CCR6 highly expressed CD19+CD5+ B cells, to promote HCC progression, which might be via enhancing angiogenesis.