Sunitinib has been used as the main therapy to treat the metastatic clear cell renal cell carcinoma (ccRCC) as it could function via suppressing the tumor growth and angiogenesis. Yet most ccRCC tumors may still regrow due to the development of sunitinib-resistance, and detailed mechanisms remain to be further investigated. The angiopoietin family includes angiopoietin-1 and angiopoietin-2 (ANGPT-1 and -2). It was reported that estradiol regulates expression of ANGPT-1, but not ANGPT-2, through estrogen receptor α (ERα) in an experimental stroke model. To date, there is no finding to link the E2/ER signal on regulating ANGPT-2. Our study is the first to explore (i) how estrogen receptor β (ERβ) can up-regulate ANGPT-2 in RCC cells, and (ii) how ERβ-increased ANGPT-2 can promote the HUVEC tube formation and reduce sunitinib sensitivity. Mechanistic studies revealed that ERβ could function via transcriptional regulation of the cytokine ANGPT-2 in the ccRCC cells. We found the up-regulated ANGPT-2 of RCC cells could then increase the Tie-2 phosphorylation to promote the angiogenesis and increase sunitinib treatment resistance of endothelial cells. In addition to the endothelial cell tube formation and aortic ring assay, preclinical studies with a mouse RCC model also confirmed the finding. Targeting this newly identified ERβ/ANGPT-2/Tie-2 signaling pathway with the FDA-approved anti-estrogen, Faslodex, may help in the development of a novel combined therapy with sunitinib to better suppress the ccRCC progression.