Angiogenesis and growth kinetics of tumors treated with GW2580. (A-D) Tumors were harvested on day 14 after tumor implantation and GW2580 treatment, and mRNA expression was quantified using RT-PCR for the stated genes. Relative mRNA expression was normalized to β-actin (n = 6/group). (E) MMP-9 protein levels from tumors were analyzed by Western blot and normalized to α-tubulin. (F) Quantification of MMP-9 protein levels by ImageJ software (n = 3/group). (G) Representative CD31⁺ vascular staining of 3LL tumors from control and GW2580-treated mice is shown. (H) Quantification of CD31⁺ area was performed using ImageJ software (n ≥ 5/group). Tumor volume was monitored by caliper measurements of 3LL tumors (I-J; n = 6/group) and B16F1 tumors (K; n = 3/group) in mice treated with control diluent, 20 mg/kg, or 80 mg/kg GW2580 twice a day, or 160 mg/kg GW2580 once daily. Scale bars represent 100 μm.