Abstract Purpose: To investigate how biologically relevant markers change in response to antiangiogenic therapy in metastatic clear cell renal cancer (mRCC) and correlate these changes with outcome. Experimental Design: The study used sequential tumor tissue and functional imaging (taken at baseline and 12–16 weeks) obtained from three similar phase II studies. All three studies investigated the role of VEGF tyrosine kinase inhibitors (TKI) before planned nephrectomy in untreated mRCC (n = 85). The effect of targeted therapy on ten biomarkers was measured from sequential tissue. Comparative genomic hybridization (CGH) array and DNA methylation profiling (MethylCap-seq) was performed in matched frozen pairs. Biomarker expression was correlated with early progression (progression as best response) and delayed progression (between 12–16 weeks). Results: VEGF TKI treatment caused a significant reduction in vessel density (CD31), phospho-S6K expression, PDL-1 expression, and FOXP3 expression (P < 0.05 for each). It also caused a significant increase in cytoplasmic FGF-2, MET receptor expression in vessels, Fuhrman tumor grade, and Ki-67 (P < 0.05 for each). Higher levels of Ki-67 and CD31 were associated with delayed progression (P < 0.05). Multiple samples (n = 5) from the same tumor showed marked heterogeneity of tumor grade, which increased significantly with treatment. Array CGH showed extensive intrapatient variability, which did not occur in DNA methylation analysis. Conclusion: TKI treatment is associated with dynamic changes in relevant biomarkers, despite significant heterogeneity in chromosomal and protein, but not epigenetic expression. Changes to Ki-67 expression and tumor grade indicate that treatment is associated with an increase in the aggressive phenotype of the tumor. Clin Cancer Res; 19(24); 6924–34. ©2013 AACR.