Abstract P2043: Role of GRK4, a Risk Factor for Hypertension, in Breast Cancer

Abstract

Clinical studies have shown that breast cancer risk is increased in hypertensive women. The molecular mechanism linking hypertension and breast cancer remains undetermined. Several lines of evidence suggest that G-protein coupled receptor kinase 4 (GRK4) is a candidate protein. The link between high salt and hypertension has been well documented. Renal sodium excretion is regulated by dopaminergic D1-like receptor (D 1 R). Sustained activation of GRK4 impairs D 1 R function and causes hypertension. On the other hand, GRK4 expression is higher in breast cancer than in normal breast tissues. GRK4 variants A142V and R65L have been identified as both hypertension and breast cancer risk loci. To determine the role of GRK4 in breast cancer, we first examined GRK4 expression and SNPs in 7 breast cancer cell lines and benign breast epithelial cell line, MCF-10A. We found that MCF-10A cells do not express detectable level of GRK4 protein whereas all 7 breast cancer cell lines express different levels of GRK4. Except for BT20, all other breast cancer lines have 1-3 SNPs on GRK4 gene of which A142V occurs in all 6 lines. MDA-MB-468 cells contain both A142V and R65L at both alleles. These 2 SNPs have been demonstrated to be associated with enhanced GRK4 activity and reduced sodium excretion in the kidney. To further explore the function of GRK4 in breast cancer cells, we generated three stable GRK4 knock-down MDA-MB-468 lines using inducible lentiviral shRNA vectors. GRK4 knock-down induced by doxycycline (Dox) significantly reduced growth rates of these cells with greatest reduction in line 167 (68%). Growth rate changes correlated with that in GRK4 mRNA levels. 5-bromo-2-deoxyuridine (BrdU) incorporation test demonstrated that reduced growth rate was predominantly a result of reduced cell proliferation. Percentage of BrdU labeled cells were decreased from 45±3% in the cells without Dox to 32±5% in the cells treated with 0.1 μg/ml Dox (p<0.005). There was no change in growth rate in the cells expressing non-targeting shRNA. Our results demonstrate that GRK4 plays a role in promoting breast cancer proliferation and suggest that GRK4 could be a new therapeutic target for the treatment of hypertension and breast cancer.