Objective: Frequent loss of function mutations in the chromatin remodeling protein and tumor suppressor, AT-rich interactive domain-containing protein 1A (ARID1A), have been noted in ovarian cancers, particularly clear-cell and mucinous histotypes. We initially investigated the relationship between differential ARID1A protein levels and patient survival in a well-annotated ovarian cancer tissue microarray. Our analyses revealed distinct cellular localization patterns for ARID1A using moderately stringent assay conditions and we observed a relationship between cytoplasmic staining of ARID1A and poor disease outcome. We further investigated the relevance of cytoplasmic ARID1A in vitro in a cell line model of ARID1A-mutated, clear-cell ovarian cancer and found that ectopic expression of a cytoplasm-localized variant of ARID1A enhances cellular proliferation relative to wild type ARID1A restored cells.
Methods: Tissue microarrays representing a natural distribution of ovarian cancer histotypes (N=259) were stained for ARID1A and scored. Four patient subgroups were further defined based on the absence or presence of distinct ARID1A staining in nuclear and/or cytoplasmic tumor cell compartments. Relationships between ARID1A staining pattern and patient outcome were investigated using univariate and multivariate Cox regression modeling with Wald testing and Kaplan-Meier method with log-rank testing. Nuclear localization sequence variants of ARID1A were generated and phenotypic assessments of cellular proliferation were performed following ectopic expression of cytoplasm-localized versus wild type, i.e. nuclear-localized, ARID1A in a cell line model of ARID1A-mutated, clear cell ovarian cancer cells (TOV21G).
Results: Using a moderately stringent IHC assay for ARID1A, loss of nuclear ARID1A was observed in 7% of ovarian cancer patients, and women with versus without loss of nuclear ARID1A had similar survival. Prevalent cytoplasmic staining was observed in 14% of cases overall and was more common in mucinous (50%), clear cell (42%) and serous (11%) adenocarcinomas than other histotypes (p=0.002). Prevalent cytoplasmic ARID1A was not only associated with an increased risk of death in the entire cohort (HR=1.67, 95% CI=1.137-2.454, p=0.009), but was also an independent predictor of worse survival (adjusted HR=1.530, 95% CI=1.017-2.301, p=0.041) after adjusting for patient age at diagnosis, site of disease, stage, cell type and grade. Comparative in vitro analyses of cytoplasmic and wild type ARID1A variants, ectopically-expressed in clear-cell ovarian cancer cells revealed that cytoplasmic ARID1A confers an increased proliferative phenotype relative to wild type ARID1A restored cells.
Conclusions: ARID1A cellular localization and abundance varied across ovarian cancer histotypes and cytoplasmic localization was indicative of worse outcome. Our in vitro studies suggest cytoplasmic ARID1A increases the proliferative potential of clear-cell ovarian tumor cells and provides phenotypic evidence supporting the poor disease outcomes we observe in patients exhibiting cytoplasmic ARID1A. These findings support further investigations combining assessments of ARID1A cellular localization and abundance patterns towards the identification of ovarian cancer patients with an elevated risk to experience poor disease outcome.
Citation Format: Nicholas W. Bateman, Kevin A. Byrd, Kelly A. Conrads, John I. Risinger, Carl Morrison, Kunle Odunsi, Chad A. Hamilton, G. Larry Maxwell, Kathleen M. Darcy, Thomas P. Conrads. Cytoplasmic ARID1A and poor outcome in ovarian cancer patients. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research: Exploiting Vulnerabilities; Oct 17-20, 2015; Orlando, FL. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(2 Suppl):Abstract nr A19.