Figure 6. CCR4 antagonism ameliorates the malignancy phenotype of melanoma.

(A–B) The CCR4 antagonist reduces melanoma cell migration. Local and HBMMC were incubated in serum-free medium (SFM), SFM supplemented with migration-stimulating astrocyte-derived soluble factors (HA CM), SFM + 10 μM CCR4 antagonist, SFM supplemented with HA CM and 10 μM CCR4 antagonist. (A) Representative images of migrated local and HBMMC cells following 24 h incubation (B) Quantification of A. Data shown are number of migrated cells counted cells in four independent fields/well; **p < 0.005, ***p < 0.0005. (n = 3). (C–D) CCR4 over-expressing melanoma cells (CCR4^hi) were s.c inoculated into nude mice. Two wks post cell inoculation mice were treated, twice a week, with CCR4 antagonist (1.5 μg) or with DMSO (vehicle Control). 12 wks following inoculation local tumors were excised and brains were removed and analyzed for metastasis formation. (C) Tumors of antagonist-treated and control mice. (D) Brain micrometastasis in antagonist-treated and control mice (n = 6). (E) CCR4-sorted melanoma cells were s.c inoculated into nude mice. One wk post cell inoculation mice were treated, every 2 days, with CCR4 antagonist (1.5 μg) or with DMSO (Control). 12 wks following inoculation local tumors were excised and brains were removed and analyzed for metastasis formation. (F) Antagonist-treated and control mice. (G) Tumors of antagonist-treated and control mice. (H) Brain micrometastasis in antagonist-treated and control mice (n = 5).