Profiling of suppressive immune subsets in metastasis negative and positive sentinel lymph nodes from patients with HER2- breast cancer

Methods Suppressive immune subsets were profiled by multi-color flow cytometry in two different BrCa SLN cohorts. In the first cohort, collected at the Providence Cancer Center (08/11-07/13), we looked at frequencies of HLA-DRCD14+ myeloid cells and their expression of co-stimulatory and inhibitory receptors. Four metastasis+ SLN and 11 metastasisSLN were assessed in this cohort. SLN in the second cohort were collected at the VUmc in Amsterdam and Kennemer Gasthuis in Haarlem (10/13-06/14) and contained 2 metastasis+ and 7 metastasisSLN. In this cohort the frequencies of HLA-DRCD14+ cells as well as CD25_hi FoxP3+ T regulatory cells (Treg) were analyzed. Since all tumors corresponding to the metastasis + SLN turned out to be HER2 negative, only metastasisSLN from HER2tumors were included. Apart from 2 tumors in the Providence cohort, all tumors did express progesterone and/or estrogen receptors. Results Elevated frequencies of HLA-DRCD14+ immature myeloid cells could be detected in the metastasis+ SLN in both cohorts. This difference was statistically significant for the Providence cohort (p < 0.0001) (3.8 fold increase). No differences were observed for expression levels of the co-stimulatory molecules CD80 and CD86 or the inhibitory molecules PD-L1 and B7H4 on HLADRCD14+ cells between positive or negative SLN and expression was low for all these markers. Due to the small number of metastasis+ SLN in the Dutch cohort statistics could not yet be performed, but a 2.3 fold increase in HLA-DRCD14+ cells was seen. In this cohort, frequencies of Treg were found to be 4-fold higher in the metastasis+ SLN compared to metastasisSLN (0.31 ± 0.22 vs. 1.22 ± 0.24 Treg of CD4 T cells). Moreover, a significant correlation was observed between the frequencies of HLA-DRCD14+ immature myeloid cells and the frequencies of Treg in these BrCa SLN (r = 0.718, p < 0.01).