Abstract 2663: Identification of differential immune landscapes in high-grade serous ovarian tumors of high-risk patients by imaging mass cytometry
Authors: Cun Han , Ferri-Borgogno Sammy , Cheng Rita , Kim Jae-Hoon , Han Gwan Hee , Lu Karen H. , Mok Samuel C. 
Published: September 2020

Abstract
The immune environment of ovarian cancer has previously been linked to clinical outcomes. In particular, BRCA mutated tumors have been associated with the presence of intratumoral T cells, higher mutational load, and improved survival. To better assess the immune environment of such tumors, we used imaging mass cytometry (IMC), a novel imaging platform that allows for high-multiplex biomarker detection and simultaneous spatial evaluation, to generate and compare immune landscapes in advanced stage high-grade serous ovarian tumors (HGSC) with BRCA1 or 2 mutations against wild-type BRCA1 and 2.
A total of 40 advanced stage HGSC tumors were collected under an IRB-approved protocol, 20 patients with known BRCA1 or 2 mutations and 20 wild-type patients. Formalin fixed paraffin embedded (FFPE) tissue sections of 10µm thickness were stained with 34 metal-tagged antibodies to detect various cell specific and immune related markers using the Fluidigm protocol. IMC data was obtained by the Fluidigm Helios CyTOF instrument utilizing the Hyperion Imaging System laser ablation module. Images from a 1mm2 area of each tissue section were processed and converted to tiff files using MCD Viewer (Fluidigm). Tumor tissue was identified by positive pan-Keratin and cytokeratin 8/18 activity. Stroma was noted by positive smooth muscle actin activity. Images were then analyzed for various immune marker activity including CD8, CD4, Granzyme B, CD20, CD68, PDL1, and FoxP3. Statistical analysis was performed using unpaired t-test.
The results demonstrated that HGSCs with BRCA1 mutations had significantly higher number of intratumoral CD8+ cells (32 v. 7 cells/mm2, p=0.02) compared to BRCA2 mutations. A markedly higher number of intratumoral CD20+ cells and CD4+ cells were also observed in HGSCs with BRCA1 than BRCA2 mutations; although, the difference did not reach significance (6.7 v. 1.3 cells/mm2, p=0.09; 93 v. 28 cells/mm2, p=0.08). Compared to wildtype, BRCA1/2 mutated tumors had significantly higher number of CD20+ cells in the tumor aspect and in the whole tissue (4 v. 0.2 cells/mm2, p=0.03; 9 v. 1 cells/mm2, p=0.05). Furthermore, BRCA1/2 mutated tumors also have a higher number of stromal and total CD4+ than wildtype patients (28 v. 14 cells/mm2, p=0.07; 63 v 24 cells/mm2, p=0.7).
In this cohort, we found that HGSC tumors with BRCA1 mutations had increased immune cells compared to BRCA2 mutations, including intratumoral CD8+, CD20+, and CD4+ cells. BRCA mutant tumors also had increased CD20+ and CD4+ cells compared to BRCA wildtype. These immune phenotypes suggest an increased immunogenicity in tumors with BRCA mutations, particularly BRCA1, consistent with previous studies. This may have clinical implications; but, furthermore, may suggest novel approaches to therapy. Overall, this study shows that IMC provides a powerful and comprehensive approach to evaluating the cellular phenotype and spatial interrelationships of the tumor immune environment.
Citation Format: Han Cun, Sammy Ferri-Borgogno, Rita Cheng, Jae-Hoon Kim, Gwan Hee Han, Karen H. Lu, Samuel C. Mok. Identification of differential immune landscapes in high-grade serous ovarian tumors of high-risk patients by imaging mass cytometry [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 2663.