Biomarkers Of Immune Infiltration For Multiplex Immunofluorescence In Breast Cancer

Introduction: Only a subset of patients have a clinical response to cancer immunotherapy. The molecular mechanisms that mediate the immunological response or tolerance to treatment are just beginning to be understood. Several tumor signaling pathways are involved in those mechanisms: Wnt/β-catenin, STAT3, NFkB, PI3K/PTEN/AKT, and TP53. To characterize the tumor, its immune infiltrate, and its microenvironment, we examined expression of several biomarkers and determined the relationships among them in various breast cancer subtypes. Methods: 148 breast cancer (BCa) cases from the University of Chicago Breast Cancer SPORE tissue bank under IRB approved protocols were classified into four biological subtypes based on immunohistochemical (IHC) staining: luminal A, luminal B, Her2 and basal-like. IHC staining for PD-L1, CD8, FoxP3, CD68, CD163, and β-catenin was performed on tissue microarrays. CD8+ and FoxP3+ cells were counted manually and recalculated for 1 mm 2 . Macrophage phenotype was determined using single and double staining with CD68 for total population and CD68/CD163 for M2. CD68+ and CD163+ cells were counted manually, confirmed by Image Analysis, and recalculated for 1 mm 2 . PD-L1 and β-catenin (membrane-associated, cytosolic and nuclear) were scored as negative, weak positive, moderate, and strong. Results: The study cohort subtypes were 20.9% basal-like, 10.4% Her2, 58.3% luminal A, and 10.4% luminal B. The ratio of M2 to M1 cells increased with disease progression (p + cell infiltrate and low or absent CD8+ T cells. The ratio of FoxP3 Tregs to CD8+ cells in basal-like tumors were 1.7, 2.5, and 4 -fold higher than in Her2, luminal A, and luminal B tumors, respectively (p Conclusions and Future Directions: Our data suggests that IHC using a panel of antibodies may be a robust and suitable method for evaluating level of immune infiltration. Future studies will evaluate multiplex immunofluorescence of multiple biomarkers in breast tissue. Citation Format: April Swoboda, Galina Khramtsova, Lise Sveen, Andrey Khramtsov, Rita Nanda, Olufunmilayo Olopade. Biomarkers of immune infiltration for multiplex immunofluorescence in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2950. doi:10.1158/1538-7445.AM2017-2950