Abstract 2503: Tumor microenvironment composition correlates with relapse in ovarian granulosa cell tumors
Authors: Khlebus Eleonora , Vuttaradhi Veena K. , Welte Thomas , Khurana Namrata , Celestino Joseph , Beird Hannah C. , Gumbs Curtis , Little Latasha , Legarreta Alejandra Flores , Nguyen Tri , Lawson Barrett , Broaddus Russell R. , Gershenson David M. , Futreal P. Andrew , Hillman R. Tyler 
Published: April 2023

Abstract
Background: Adult-type granulosa cell tumors (AGCT) are rare ovarian sex cord tumors that exhibit near-universal FOXL2 c.C402G (p.Cys134Trp) hotspot mutations. AGCT recurrence is difficult to predict and is almost always incurable after relapse. Little is known about the relationship between intra-tumor immune and stromal composition and AGCT relapse.
Objective: To compare global gene expression profiles between primary and recurrent AGCTs, characterize the tumor microenvironment (TME), and identify correlates of disease recurrence.
Methods: Total RNA sequencing was performed on 24 pathologically confirmed, cryopreserved AGCT samples, including 8 primary and 16 recurrent tumors. Standard methods were applied for read alignment, quality control, and quantification of gene-specific read counts. DESeq2 was used to identify statistically significant (adjusted P-value &lt; 0.05) differentially expressed genes between primary and recurrent tumors with fold change &gt; 2. Gene set enrichment analysis was performed using clusterProfiler. Integrative TME composition de-convolution was performed using multiple published algorithms including CIBERSORTx, quanTIseq, xCell, MCP-counter, and EPIC. TME analysis results were externally validated using data from smaller, previously published RNA sequencing datasets.
Results: Thirty-one genes were identified as differentially expressed between primary and recurrent AGCTs, including NELL2, GDF6, TUBB2B, AQP3. These included genes with known function in hormone signaling such as LHCGR (adjusted P-value = 0.002) and INSL3 (adjusted P-value = 0.017) which were highly expressed in primary tumors and CYP19A1 (adjusted P-value = 0.009) which was highly expressed in recurrent tumors. Gene set enrichment analysis revealed increased expression of hormone-regulated and immune-related gene sets in recurrent tumors. Integrative, multi-platform TME analysis showed recurrent AGCT to exhibit reduced fractions of cancer-associated fibroblasts and enrichment of myeloid lineages such as neutrophils and macrophages.
Conclusions: Recurrent AGCTs exhibit alterations in hormone pathway gene expression as well as decreased infiltration of cancer-associated fibroblasts, suggesting dual roles for hormonal signaling and TME remodeling underpinning disease relapse.
Citation Format: Eleonora Khlebus, Veena K. Vuttaradhi, Thomas Welte, Namrata Khurana, Joseph Celestino, Hannah C. Beird, Curtis Gumbs, Latasha Little, Alejandra Flores Legarreta, Tri Nguyen, Barrett Lawson, Russell R. Broaddus, David M. Gershenson, P. Andrew Futreal, R. Tyler Hillman. Tumor microenvironment composition correlates with relapse in ovarian granulosa cell tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2503.