The
vascular endothelial growth factor (
VEGF)-
VEGF receptor (VEGFR) signaling pathway is involved in
cancer-related
biological functions and is a therapeutic target in
cancer. However, the influence of epigenetic regulation of
VEGF-VEGFR signaling-related genes remains unclear. Here, we evaluated the effects of FLT1 and KDR promoter hypermethylation combined with drugs targeting
VEGF-VEGFR signaling on
cancer-related phenotypes in
renal cancer cells (RCCs) and examined changes in FLT1 and KDR promoter hypermethylation in tissues from patients with
renal cancer.
RESULTS: In vitro experiments were performed to evaluate the effects of beavacizumab (an anti-
VEGF antibody), an anti-FLT1
peptide, an anti-KDR antibody, and the VEGFR
tyrosine kinase inhibitors (TKIs)
sunitinib and
axitinib in 13 RCC lines with different levels of FLT1 and/or KDR promoter methylation and in 2 FLT1 or KDR in vitro knockdown models. The synergistic effects of
sunitinib and
axitinib treatment were also evaluated in four RCC lines having different levels of FLT1 and/or KDR methylation. In our in vitro experiments,
bevacizumab and an anti-KDR antibody did not affect the proliferation of RCCs having FLT1 and/or KDR hypermethylation. In contrast, in RCCs with FLT1 hypermethylation, proliferation inhibition was counteracted by treatment with an anti-FLT1
peptide and both
VEGF-TKIs (
sunitinib and
axitinib). Demethylation with
sunitinib or
axitinib synergistically increased proliferation inhibition in the RCCs exhibiting FLT1 hypermethylation. Using in vitro FLT1 or KDR knockdown models, decreased proliferation inhibition following anti-FLT1
peptide,
sunitinib, and
axitinib treatment was observed only in FLT1-knockdown cells. In patients with
renal cancer who received
sunitinib, FLT1 promoter methylation was higher in
renal cancer tissues from eight nonresponders (stable or progressive disease assessed by the Response Evaluation Criteria in Solid Tumors) than in
cancer tissues from five responders (complete response or partial response).
CONCLUSIONS: The present data showed that hypermethylated FLT1 was important for the efficacy of anti-
VEGF/VEGFR drugs targeting FLT1 or intracellular VEGFR signaling. FLT1 hypermethylation causing alterations of FLT1 function could serve as a useful
biomarker for predicting changes in FLT1 status in RCCs.