Abstract Background: VEGFR-2 plays a crucial role in mediating angiogenic endothelial cell responses via the VEGF pathway and VEGFR-2 inhibitors are in clinical use. Angiogenesis is a host-driven process and heritable variation in KDR, the gene encoding VEGFR-2, may impact on angiogenesis. We have previously characterized KDR germline genetic variation by resequencing (Ye et al., AACR, 2007;737). However, very limited data are available on the molecular function of KDR germline variants and their resulting effect on tumoral expression and vascularization. The purpose of this study was to identify functional variants affecting KDR expression and microvessel density (MVD) in lung cancer. Methods: The molecular function of the non-synonymous variants R106W, V297I, Q472H and C482R was assessed by testing their effect on VEGFR-2 phosphorylation (measured via immunoblotting) after VEGFA165 exposure in HEK293 cells. Four 5’ upstream region SNPs were examined by luciferase reporter assay in endothelial SVEC4-10 cells. To validate these results and examine the effects of other KDR SNPs, KDR mRNA and VEGFR-2 protein expression were measured by quantitative PCR and immunohistochemistry in non-small cell lung cancer (NSCLC) samples from Caucasian patients and MVD was assessed by immunohistochemistry (anti-CD31). Patients were genotyped for KDR SNPs and their association with mRNA, protein expression, or MVD was evaluated by linear regression. Results: Among the non-synonymous SNPs, the only significant observation was that Q472H had a 46% increase in VEGFR-2 phosphorylation relative to the wild-type variant after VEGFA165 stimulation (p=0.035); in agreement with this finding, Q472H was associated with 22% greater MVD in 168 NSCLC patients (p=0.05). Of the putatively regulatory SNPs, -2008G and -1942G had no effect on luciferase expression. -2854C and -2455A showed 10-20% higher luciferase expression than the wild-type variant (p<0.05) and they were associated with 23% greater tumor KDR mRNA expression in 66 NSCLC patients (p=0.014), but had no effect on tumor VEFGR-2 expression; -271A reduced luciferase expression by approximately 50% (Ye et al., AACR 2007;737) and correlated with 10% lower tumor VEGFR-2 expression in 162 NSCLC patients (p=0.01); -906C and 23408G were associated with 20-25% higher tumor KDR mRNA levels (p=0.009 and 0.025, respectively), although these variants had no effect on tumor VEGFR-2 expression. Conclusions: This study has identified novel functional variants which correlate with in vitro gene expression or VEGFR-2 phosphorylation and KDR expression or MVD in NSCLC. This information may be critical for understanding the role of heritable KDR variations as contributing factors to prognosis and lung cancer outcome. Acknowledgment: This research was supported by the American Cancer Society. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5136. doi:10.1158/1538-7445.AM2011-5136