Digital Spatial Profiling In Her2 Positive Breast Cancer: The Road To Precision Medicine.

Abstract Background: Breast cancer (BC) has been considered immunologically quiescent, this could be explained by tumor and tumor immune microenvironment (TIME) characteristics. We aimed to characterize the TIME of human epidermal growth factor 2 positive (HER2+) BC using GeoMx Digital spatial profiling (DSP) in primary and metastatic samples. Methods: DSP was performed on 15 primary and metastatic FFPE samples from 8 patients; including 2 brain metastases. Visualization markers for Pan-Cytokeratin (PanCK, tumor), CD45 (immune), CD3 (T cells) were used to guide selection of immune cold, warm and hot regions of interest (ROI). Each ROI segmented tumor (PanCK+) or stroma (PanCK-) based on PanCK tumor + or - areas of illumination (AOI). For each AOI 71 protein targets were tested. Gene expression profiling for PAM50 subtypes and Tumor Inflammation Score (TIS) was done using NanoString's nCounter PanCancer IO360 assay with PAM50 spike. TIS was determined by threshold of 6.5. Results: Primary HER2+ samples had the highest amount of tumor infiltrating lymphocytes (CD3) and overall immune cells (CD45) compared to all metastatic disease in tumor and stroma (Table 1). Brain metastases had lowest TIS. Stroma of primary tumors had higher protein expression of immune activation and checkpoint markers CD27, ICOS, 41BB and TIM3 compared to brain and soft tissue metastases. HER2 protein expression most correlated with metastases. Metastatic tumor had higher proliferation by Ki67. Patient 2 had archival primary tumor (n=2) and soft tissue metastasis (ST) (n=1). TIS was obtained from immune hot, warm and cold ROI of each sample. Primary tumor stroma had highest expression of STING and Granzyme B compared to ST, suggesting that the tumor underwent immune escape. Conclusions: Primary tumors had a higher number of immune cells than metastatic sites. These findings suggest that immunotherapy in early stage BC could be more effective than in the advanced BC. This project will be part of the GeoMx Breast Cancer Consortium. Table 1.Summary of patient, tissue, diagnosis, ER/PR/HER2 status, PAM50 subtype and TIS statusPatientTissueDiagnosisER(%)/PR(%)/HER2(+)PAM50 SubtypeTIS Status1BreastIDC, DCIS2/0/3Luminal AHigh1BreastTumor emboliN/ALuminal AHigh1LungMetastatic0/0/3HER2 enrichedHigh2BreastICD, DCIS0/0/3HER2 enrichedLow2BreastResidual IDC0/0/N/AHER2 enrichedHigh2Soft tissueLocal recurrence0/0/3HER2 enrichedLow3BrainMetastatic15/0/3HER2 enrichedLow4BreastIDC0/0/3Basal likeHigh4BreastLocal recurrenceN/ALuminal AHigh5BrainMetastatic0/0/3Basal likeLow6BreastIDC, DCIS80/70/2*Luminal AHigh7BreastIDC, DCIS0/0/3HER2 enrichedHigh7BreastLocal recurrenceN/ABasal likeHigh8BreastIDC, DCIS10/15/1Luminal ALow8Soft tissueLocal recurrence0/0/3Basal likeLowAbbreviations: ER: estrogen receptor; PR: progesterone receptor; HER2: human epidermal growth factor 2; TIS: tumor inflammation score; IDC: invasive ductal carcinoma; DCIS: ductal carcinoma in situ*HER2/CEP17 ratio: 2.13 Citation Format: Ilana Schlam, Sarah E. Church, Krysta Chaldekas, Brent T. Harris, Briana M. Hudson, Andrew M. White, Sandra M. Swain. Digital spatial profiling in HER2 positive breast cancer: The road to precision medicine [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2718.