Abstract
DARC (aka Duffy), a chemokine receptor, is expressed on erythrocytes and endothelial cells in blood vessels. It has been linked to inhibition of tumor metastasis, through blockade of tumor cell invasion into vessels, and to regulation of plasma levels of some cytokines which influence leukocyte egress from blood into tissues. The Duffy null (Fy-) phenotype, in which DARC is not expressed on red blood cells, is especially prevalent among people of West African descent. These populations are also highly susceptible to aggressive subtypes of breast and prostate cancers with higher rates of metastases. There is limited evidence, from a Chinese study, that the Fy- phenotype may be associated with increased breast cancer (BrCa) invasiveness and lower rates of survival. However, little is known about how the presence or absence of DARC expression may affect breast cancer metastases and overall survival among African-American and European- American women. Our lab is investigating whether Fy- phenotypes could be associated with increased BrCa metastasis by genotyping the Fy- (null and weak) alleles and assessing DARC gene expression in breast and lymphoblast cells using qPCR and Immunofluorescence (IF). Because there are two isoforms of DARC and the Fy- SNP is located in the promoter of one and the 5′UTR of the other, it is plausible that the alternate isoform is specific to these cell types and may still be expressed. Here we present data from the preliminary stages of this project; including characterizing Duffy alleles in our cell line models. We have genotyped the 3 known DARC polymorphisms that result in Fy- (and weak) erythrocyte phenotypes in breast cancer and lymphoblast cell lines. Specifically, we validated that HAPMAP Yoruba lines are homozygous for the -541T>C point mutation Fy- allele (rs2814778) and half of the African-American lines are heterozygous while the remaining are homozygous for the Fy- allele and the CEPH-European lines do not have the Fy- allele. Our breast cancer (BrCa) lines are derived from tumors of varying molecular subtypes, isolated from African-American women. We show that these BrCa lines are homozygous for the Fy- allele and yet still express the DARC protein. The specific isoform expressed in these cells (DARC-2) may not be the same as used to phenotype the Fy- status in erythrocytes (DARC-1). This suggests that a Fy- genotype may still have some tissue specific expression of DARC-2 in epithelial cells. Therefore, we will also show whether these SNPs correspond with expression of the DARC gene in HAPMAP lymphoblasts and delineate which isoform is being expressed in both cell types. In conclusion, we hypothesize that Fy- status as determined in endothelial and erythrocytes, in people of African descent may remove the metastatic protection of DARC and alternate expression of DARC-2 on epithelial cells may increase the rate of metastases in these populations. Currently, we are analyzing clinical samples (blood and breast tissues) from local breast cancer patients for Fy- status and will determine if this correlates with metastatic disease. In addition, biochemical experiments will be conducted to determine whether DARC-related chemokines are in higher prevalence in the Fy- patients, how the isoforms differ in their affinity to the chemokine ligands and whether this is associated with specific breast cancer subtypes.
Citation Format: Melissa Davis, Rupa Hire, Jonathan Swanson, Kauthar Mumin, Michele Monteil, Michele Lou. Investigating correlations of DARC underexpression (Duffy Null phenotypes) with increased breast cancer lymph node metastasis. [abstract]. In: Proceedings of the Sixth AACR Conference: The Science of Cancer Health Disparities; Dec 6–9, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2014;23(11 Suppl):Abstract nr C69. doi:10.1158/1538-7755.DISP13-C69
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