The Duffy Antigen Receptor for Chemokines (DARC/ACKR1) is an atypical chemokine receptor which promiscuously binds chemokines of both CC and CXC chemokine families. DARC/ACKR1 maintains homeostatic levels of chemokines in circulation through its expression on erythrocytes, and in tissues participates in chemokine transport, where chemokines are transported across the cell layer to help establish chemokine gradients. Establishment of these chemokine gradients is essential for proper immune cell trafficking.
In our recent study, we have shown that DARC/ACKR1 is expressed on the tumor epithelium, and through in silico analysis of data from the Cancer Genome Atlas (TCGA), that higher DARC/ACKR1 expression is positively and significantly correlated with tumor-associated leukocyte abundance when using CIBERSORT deconvolution methods. Specifically, higher DARC/ACKR1 expression was linked with increases in B cell, T cell, monocyte and macrophage populations. Higher DARC/ACKR1 expression is also significantly associated with better survival outcomes, across all breast cancer molecular subtypes.
To further evaluate DARC/ACKR1 expression and influences on the breast TME, we have employed multiple approaches to characterize how DARC/ACKR1 impacts immune cell recruitment and subsequent infiltration into the tumor.
In our patient cohort, we have recently completed Hyperion imaging mass cytometry staining of tumors that have been scored by IHC as DARC/ACKR1 high or low expressing. We used a panel of approximately 30 markers, including structural, immune, and other markers of interest. Following imaging, we have used cell segmentation software to quantify marker expression on a cell-by-cell basis. Using this method, we are able to conduct single-cell analyses of our markers of interest, while also retaining the spatial composition of the cells. We have used the tSNE algorithm to identify cell populations that cluster uniquely within DARC/ACKR1 high or low tumor expressing groups, specifically identifying different clusters of immune cell populations, which show different spatial expression patterns in our comparison groups.
In addition to investigations of immune cell recruitment to the TME, we have also developed a novel DARC/ACKR1 breast cancer transgenic mouse model to study how disease progression may differ with differing DARC/ACKR1 phenotypes. Briefly, DARC -/- female mice were crossed with the male C3(1)Tag +/0 breast cancer transgenic mouse to generate DARC +/-, C3(1)Tag +/0 male mice. These males were subsequently backcrossed to DARC -/- females, to generate the target DARC/ACKR1 +/- or -/-, C3(1)Tag+ mice. These mice show evidence of early disease beginning at 3.5 months of age, and progress to advanced disease by 5 months of age. We have collected tumor tissue and blood specimens from mice ranging from 3.5 to 7 months of age, to characterize disease progression over time between our DARC/ACKR1 expressing and non-expressing groups. We have observed differences in tumors characteristics between our DARC/ACKR1 expressing groups, and upon histological and immunofluorescent evaluation, we have begun to observe differences in immune cell recruitment, following similar patterns to what we observed in our in silico and patient cohort analyses.
Citation Format: Rachel Martini, Brittany Jenkins, Dorrah Deeb, Hiranmayi Ravichandran, Paula Ginter, Esther Cheng, Syed Hoda, Dhananjay Chitale, Haythem Ali, Eleanor Walker, Jessica Bensenhaver, Olivier Elemento, Nancy Manley, Lisa Newman, Melissa Davis. Breast tumor expression of DARC/ACKR1 and impacts on disease progression and immune cell recruitment in the tumor microenvironment [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4975.