Quantitative Assessment and Prognostic Associations of the Immune Landscape in Ovarian Clear Cell Carcinoma

Abstract

Ovarian clear cell carcinoma (OCCC) is a rare subtype of epithelial ovarian cancer characterised by a high frequency of loss-of-function ARID1A mutations and a poor response to chemotherapy. Despite their generally low mutational burden, an intratumoural T cell response has been reported in a subset of OCCC, with ARID1A purported to be a biomarker for the response to the immune checkpoint blockade independent of micro-satellite instability (MSI). However, assessment of the different immune cell types and spatial distribution specifically within OCCC patients has not been described to date. Here, we characterised the immune landscape of OCCC by profiling a cohort of 33 microsatellite stable OCCCs at the genomic, gene expression and histological level using targeted sequencing, gene expression profiling using the NanoString targeted immune panel, and multiplex immunofluorescence to assess the spatial distribution and abundance of immune cell populations at the protein level. Analysis of these tumours and subsequent independent validation identified an immune-related gene expression signature associated with risk of recurrence of OCCC. Whilst histological quantification of tumour-infiltrating lymphocytes (TIL, Salgado scoring) showed no association with the risk of recurrence or ARID1A mutational status, the characterisation of TILs via multiplexed immunofluorescence identified spatial differences in immunosuppressive cell populations in OCCC. Tumour-associated macrophages (TAM) and regulatory T cells were excluded from the vicinity of tumour cells in low-risk patients, suggesting that high-risk patients have a more immunosuppressive microenvironment. We also found that TAMs and cytotoxic T cells were also excluded from the vicinity of tumour cells in ARID1A-mutated OCCCs compared to ARID1A wild-type tumours, suggesting that the exclusion of these immune effectors could determine the host response of ARID1A-mutant OCCCs to therapy. Overall, our study has provided new insights into the immune landscape and prognostic associations in OCCC and suggest that tailored immunotherapeutic approaches may be warranted for different subgroups of OCCC patients.

The Evolution of Ovarian Carcinoma Subclassification

“…While there are obscure cases with diffuse intratumoral stromal inflammation that are MMRd and might be classified as CCC [ 95 ], MMRd does not occur in prototypical CCCs; hence, MMRd is better considered in the context of the endometrioid histotypes (see above) [ 45 ]. Recent studies evaluating the immune microenvironment of CCC suggest that tumor-associated macrophages may be a marker for immunosuppressive microenvironments [ 96 , 97 ]. Perhaps an overlay of the immune microenvironment with tumor intrinsic oncogenic alterations will better explain which patients respond to immune therapy.…”

Comparative Analysis of the Antitumor Immune Profiles of Paired Radiotherapy-naive and Radiotherapy-treated Cervical Cancer Tissues

“…This panel consists of 730 immune response-related genes, covering 24 different immune cell types, and both the adaptive and innate immune responses (14). Analysis was performed as reported previously (9,(14)(15)(16)(17). Briefly, total RNA was extracted from frozen tumor tissues using a NucleoSpin RNA kit (Macherey-Nagel, Düren, Germany).…”

Definitive study shows no association between ARID1A mutation status and clinical outcome in endometriosis‐related ovarian cancers‡

“…The CD8 + TILs scoring methodology was a semiquantitative approach that only considered CD8 + TILs within the epithelial component of the tumour, disregarding stromal CD8 + cells [2]. Recently, Khalique et al [4] quantified and assessed the spatial locations of various immune subpopulations in 31 microsatellite stable CCOCs using multiplexed immunofluorescence. ARID1A mutant cases showed significantly higher CD8 + cells and CD68 + cells (tumour‐associated macrophages, TAMs) in the stroma relative to tumour.…”

“…This interaction would suggest that targeting TAMs could be relevant in synergising immune checkpoint‐based immunotherapy in various tumour types, and in the context of endometriosis‐related ovarian cancers, specifically ARID1A‐ mutated tumours. Khalique et al [4] also found significantly higher numbers of immunosuppressive subpopulations (TAMs and FOXP3 + /CD4 + regulatory T‐cells) in the stroma relative to tumours of patients with improved outcomes, suggesting that the ‘tumour‐exclusion’ of these cells is important in maintaining an effective anti‐tumour immune response. It would be useful to validate these spatial findings in larger cohorts, such as that of Heinze, Nazeran et al [2].…”