R. Tyler Hillman

Multiplexed Imaging Mass Cytometry Reveals Tumor-immune Microenvironment–dependent Hormone Receptor Expression in Adult-Type Ovarian Granulosa Cell Tumors

Abstract Adult-type granulosa cell tumors (AGCT) are rare ovarian tumors with few effective treatments for recurrent disease. To elucidate spatial features and cellular interactions within the AGCT tumor microenvironment, we applied imaging mass cytometry using a 34-marker panel on 130 regions from 24 AGCT samples, profiling more than 900,000 single cells. Analysis confirmed the immune “cold” phenotype of AGCTs and showed higher macrophage abundance in recurrent compared with primary tumors. We observed substantial heterogeneity in tissue architecture across samples, including variable presence of FOXL2+ cells embedded in collagen-rich regions (FOXL2+COL1A1+ cells). Based on tumor microenvironment composition, we defined two AGCT subtypes: AGCT-1 and AGCT-2 with distinct FOXL2+ cell distributions, differences in progesterone receptor expression, and unique transcriptomic profiles. Our findings highlight the role of macrophages, Foxl2+ subpopulations, and the extracellular matrix in AGCT progression and suggest AGCT subtype–specific vulnerabilities that could inform personalized therapies for this rare malignancy. Significance: We discovered two histologically and molecularly distinct forms of AGCTs that differ in cell composition, immune activity, and hormone signals. These findings point to new opportunities for more personalized treatment of this rare ovarian cancer.

Survival outcomes comparing minimally invasive versus open cytoreductive surgery in recurrent adult-type granulosa cell tumors

Adult-type granulosa cell tumors are a rare form of ovarian cancer, 30% of which will recur. Cytoreductive surgery is often performed at the time of a first recurrence, but little is known about the impact of open versus minimally invasive surgical approaches on survival outcomes. To examine associations between surgical approach, clinical variables, and survival outcomes among patients with adult-type granulosa cell tumors who underwent cytoreductive surgery at the time of first recurrence. This is a retrospective cohort study of patients with adult-type granulosa cell tumors enrolled in the MD Anderson Rare Gynecologic Malignancy Registry as of April 2024. Included patients had at least one documented recurrence and underwent secondary cytoreductive surgery as part of their treatment plan. Patients were excluded if surgery was performed prior to January 1, 2000, or if surgery was not intraabdominal. Demographics and clinical variables were compared using descriptive statistics. Surgical complexity was classified as either low, intermediate, or high based on procedures performed. Progression-free and overall survival outcomes were stratified by surgical approach and estimated using Kaplan-Meier curves. A multivariable Cox proportional hazards model was used to adjust progression-free survival at time of first recurrence for age, year of surgery, and extent of disease. Four hundred eighty-five patients with adult granulosa cell tumors were identified, 108 met inclusion criteria. Seventy-eight (72%) had open and 30 (28%) had minimally invasive secondary cytoreductive surgery. Baseline characteristics, including initial stage, self-identified race, or age at diagnosis, did not differ between open and minimally invasive surgery groups. Patients undergoing minimally invasive surgery were significantly younger at the time of surgery than the open group, with a median age of 42 vs 49, respectively (P=.03). For the open group, 33% of surgeries were considered intermediate complexity and 4% high complexity, compared to 7% and 0% in the minimally invasive surgery group, respectively (P=.004). There was no difference in achieving optimal cytoreduction, 85% in the open group and 88% in the minimally invasive surgery group (P=.68). Following secondary cytoreductive surgery, there was no difference in overall survival, median overall survival of 166 months in the open group and 94 months in the minimally invasive group (P=.27), or progression-free survival after first recurrence, 26 months in the open group compared to 21 months in the minimally invasive group (P=.42). The difference in progression-free survival after the first recurrence remained nonsignificant after adjustment for key potential variables, including age, surgical approach, year of surgery, and extent of disease. There was no difference in incisional or port site recurrences at the time of second recurrence among those undergoing open (8.3%) compared to minimally invasive surgery (7.4%) at time of first recurrence (P=.89). In patients with a first recurrence of adult-type granulosa cell tumors, open secondary cytoreductive surgery did not achieve superior outcomes compared to surgery via a minimally invasive approach. Minimally invasive surgery should be considered for carefully selected patients with recurrent adult-type granulosa cell tumors. Future research is needed on patient factors important to the selection of surgical approach in this setting.

TERT promoter mutations and survival outcomes in adult-type granulosa cell tumors

To evaluate survival outcomes among patients with adult-type granulosa cell tumors who have telomerase reverse transcriptase (TERT) promoter mutations. This is a retrospective cohort study using the MD Anderson Rare Gynecologic Malignancy Registry. Patients with adult granulosa cell tumors who underwent molecular testing for TERT promoter and FOXL2 c.C402G mutations were included. We used descriptive statistics to compare demographic and clinical variables and estimated progression-free and overall survival with Kaplan-Meier curves. Cox proportional hazards regression and log-rank tests were employed for comparisons, with multivariable analyses adjusting for various factors. Among 70 patients, 28 (40%) had TERT+ tumors. The median age at diagnosis was 40 years (range 12-71) for TERT- patients and 46 years (range 25-76) for TERT+ patients. At diagnosis, 22 (63%) of 35 TERT- patients were stage I, 10 (29%) stage II, and 3 (9%) stage III, while in the TERT+ group, 17/23 (74%) were stage I, 3 (13%) stage II, and 3 (13%) stage II. Univariable analysis showed no difference in time from diagnosis to first recurrence (p=0.19) and from first recurrence to second recurrence (p=0.24) based on tumor TERT status. The median time from first to second recurrence in the TERT- group was 27.3 months (95% CI 14.1 to 40.0) and in the TERT+ group was 14.8 months (95% CI 8.1 to 21.0). There was no observed difference in overall survival between the groups (HR=0.53; 95% CI 0.19 to 1.45; p=0.21).Multivariable analysis adjusting for age at diagnosis, TERT promoter mutation status, systemic chemotherapy, and stage demonstrated a significant difference in progression-free survival based on TERT mutation status (HR=2.89; 95% CI 1.32 to 6.36). After adjustment for covariates, patients with adult granulosa cell tumors and TERT+ tumors had shorter progression-free survival after first recurrence. TERT promoter mutations may identify a subset of patients with recurrent adult granulosa cell tumors and less favorable outcomes.

Neuroendocrine neoplasms of the ovary: a review of 63 cases

To describe the clinicopathological characteristics and survival outcomes of ovarian neuroendocrine neoplasms from a curated registry. This is a retrospective cross-sectional study of patients in our registry with confirmed ovarian neuroendocrine neoplasms. We excluded patients with small cell carcinoma not otherwise specified, small cell hypercalcemic type, and those with neuroendocrine 'features' or 'differentiation.' Clinicopathological characteristics were described in two separate groups: patients with carcinoid tumors and patients with neuroendocrine carcinomas. Progression-free and overall survival were estimated with the Kaplan-Meier product-limit estimator in these two groups, and multivariable analysis was done to identify predictors of survival for neuroendocrine carcinomas only. A total of 63 patients met inclusion criteria, 13 (21%) with carcinoid tumors and 50 (79%) with neuroendocrine carcinomas. In the carcinoid tumor group, one patient (8%) was misdiagnosed. Two patients (15%) had a recurrence and the 5-year overall survival rate was 80% (95% CI 45% to 100%), with a lower bound of the median survival of 4.8 years (95% CI). In the neuroendocrine carcinoma group, 23 patients (46%) were misdiagnosed, 16 of whom (69%) received therapy with the presumption of a non-neuroendocrine carcinoma diagnosis. Thirty patients (60%) had a recurrence, and the 5-year overall survival rate was 24% (10%, 38%), with a median survival of 1.6 years (1.3, 3.3). Patients with carcinomas stage III or IV had an increased risk of progression/recurrence (HR=5.6; 95% CI 1.9 to 17.0) and death (HR=8.1; 95% CI 2.2 to 29.7) compared with those with stage I or II. Pure histology was associated with an increased risk of progression/recurrence (HR=2.3; 95% CI 1.0 to 5.2) compared with admixed histology. Most patients had neuroendocrine carcinomas, which were associated with a higher recurrence rate and worse survival than carcinoid tumors. A high proportion of patients in both groups were initially misdiagnosed, and a new association with endometrial hyperplasia was observed. Neuroendocrine admixed histology is associated with a higher risk of progression.

Clinical and Genomic Landscape of RAS Mutations in Gynecologic Cancers

Abstract Purpose: We aimed to describe RAS mutations in gynecologic cancers as they relate to clinicopathologic and genomic features, survival, and therapeutic implications. Experimental Design: Gynecologic cancers with available somatic molecular profiling data at our institution between February 2010 and August 2022 were included and grouped by RAS mutation status. Overall survival was estimated by the Kaplan–Meier method, and multivariable analysis was performed using the Cox proportional hazard model. Results: Of 3,328 gynecologic cancers, 523 (15.7%) showed any RAS mutation. Patients with RAS-mutated tumors were younger (57 vs. 60 years nonmutated), had a higher prevalence of endometriosis (27.3% vs. 16.9%), and lower grades (grade 1/2, 43.2% vs. 8.1%, all P < 0.0001). The highest prevalence of KRAS mutation was in mesonephric-like endometrial (100%, n = 9/9), mesonephric-like ovarian (83.3%, n = 5/6), mucinous ovarian (60.4%), and low-grade serous ovarian (44.4%) cancers. After adjustment for age, cancer type, and grade, RAS mutation was associated with worse overall survival [hazard ratio (HR) = 1.3; P = 0.001]. Specific mutations were in KRAS (13.5%), NRAS (2.0%), and HRAS (0.51%), most commonly KRAS G12D (28.4%) and G12V (26.1%). Common co-mutations were PIK3CA (30.9%), PTEN (28.8%), ARID1A (28.0%), and TP53 (27.9%), of which 64.7% were actionable. RAS + MAPK pathway-targeted therapies were administered to 62 patients with RAS-mutated cancers. While overall survival was significantly higher with therapy [8.4 years [(95% confidence interval (CI), 5.5–12.0) vs. 5.5 years (95% CI, 4.6–6.6); HR = 0.67; P = 0.031], this effect did not persist in multivariable analysis. Conclusions: RAS mutations in gynecologic cancers have a distinct histopathologic distribution and may impact overall survival. PIK3CA, PTEN, and ARID1A are potentially actionable co-alterations. RAS pathway-targeted therapy should be considered.

Serial cytoreductive surgery and survival outcomes in recurrent adult-type ovarian granulosa cell tumors

Few studies have evaluated the role of cytoreductive surgery in patients with recurrent adult granulosa cell tumors of the ovary. Despite a multitude of treatment modalities in the recurrent setting, the optimal management strategy is not known. Cytoreductive surgery offers an attractive option for disease confined to the abdomen/pelvis. However, few studies have evaluated the role of surgery compared with systemic therapy alone following the first recurrence and subsequent disease progressions. This study aimed to determine the impact of secondary, tertiary, and quaternary cytoreductive surgery on survival outcomes in recurrent adult granulosa cell tumors of the ovary. This is a multicenter, retrospective cohort study evaluating patients with recurrent adult granulosa cell tumors of the ovary enrolled in the MD Anderson Rare Gynecologic Malignancy Registry from 1970 to 2022. Study inclusion criteria consisted of histology-proven recurrent disease, at least 1 documented recurrence, and treatment/treatment planning at the MD Anderson Cancer Center or Lyndon B. Johnson General Hospital. The primary exposure was cytoreductive surgery, and the outcomes of interest were progression-free survival and overall survival. Survival analyses were restricted to eligible patients with resectable disease without medical barriers to surgery at each progression episode. Demographic and clinicopathologic characteristics were summarized using descriptive statistics. Progression-free survival (after first, second, and third progression) and overall survival were estimated with methods of Kaplan and Meier, and were modeled via Cox proportional hazards regression. Multivariable analyses were performed for progression-free survival after first progression and overall survival. Among the 369 patients with adult granulosa cell tumors of the ovary in the registry, 149 patients met the study inclusion criteria. Secondary cytoreductive surgery was associated with a significant improvement in progression-free survival on univariable (hazard ratio, 0.37; 95% confidence interval, 0.17-0.81, P=.01) and multivariable analyses (hazard ratio, 0.42; 95% confidence interval, 0.19-0.92; P=.03). Those who underwent secondary cytoreductive surgery had a significantly improved median overall survival compared with those who did not undergo cytoreductive surgery (181.92 vs 61.56 months, respectively; P=.002). Overall survival benefit remained statistically significant on multivariable analysis (hazard ratio, 0.28; 95% confidence interval, 0.11-0.67; P=.004). Tertiary cytoreductive surgery was similarly associated with a significant improvement in progression-free survival (hazard ratio, 0.43; 95% confidence interval, 0.26-0.70; P=.001). Despite a similar trend, quaternary cytoreductive surgery was not associated with a significant improvement in progression-free survival (hazard ratio, 0.74; 95% confidence interval, 0.42-1.26; P=.27). Among those with resectable disease and no medical contraindications to surgery, cytoreductive surgery may have a beneficial impact on progression-free survival and overall survival in patients with recurrent adult granulosa cell tumors of the ovary.

Survival outcomes in patients with recurrent mixed sex cord-stromal tumors of the ovary

Mixed sex cord-stromal tumors of the ovary contain combinations of granulosa cell tumor components-either adult or juvenile subtypes-and/or Sertoli-Leydig cell tumor elements. The objective of this study is to evaluate survival outcomes in recurrent mixed sex cord-stromal tumors. This is a retrospective cohort study of recurrent mixed ovarian sex cord-stromal tumors identified through the MD Anderson Rare Gynecologic Malignancy Registry between 2000 and 2025. Comparative cohorts with recurrent, histologically uniform adult granulosa cell tumors, juvenile granulosa cell tumors, and Sertoli-Leydig cell tumors were included. Demographic and clinical characteristics were compared using descriptive statistics. Progression-free survival after first recurrence and overall survival from first recurrence were assessed using Kaplan-Meier methods and compared using log-rank tests. Sixteen patients with recurrent mixed ovarian sex cord-stromal tumors were identified: 6 (37.5%) with adult granulosa cell plus Sertoli-Leydig cell tumors, 4 (25%) with juvenile granulosa cell plus Sertoli-Leydig cell tumors, and 6 (37.5%) with adult plus juvenile granulosa cell tumors. When comparing adult granulosa cell tumors to adult plus juvenile granulosa cell tumors, significant differences in median progression-free survival-2 (21.2 vs 8.7 months, p = .03) and overall survival (181.9 vs 83.8 months, p = .001) were observed. No significant differences in progression-free survival-2 (p = .7) or overall survival (p = .8) were noted between juvenile granulosa cell tumors and adult plus juvenile granulosa cell tumors. Among tumors with molecular testing results, 25% (1 of 4) of adult plus juvenile granulosa cell tumors, 25% (1 of 4) of adult granulosa cell plus Sertoli-Leydig cell tumors, and 33% (1 of 3) of juvenile granulosa cell plus Sertoli-Leydig cell tumors were positive for the c.C402G FOXL2 mutation. Recurrent adult plus juvenile granulosa cell tumors may exhibit more aggressive clinical behavior than uniform adult granulosa cell tumors, aligning more closely with juvenile granulosa cell tumors in recurrence outcomes.

Comparative Tumor Microenvironment Analysis of Primary and Recurrent Ovarian Granulosa Cell Tumors

Abstract Adult-type granulosa cell tumors (aGCT) are rare ovarian sex cord tumors with few effective treatments for recurrent disease. The objective of this study was to characterize the tumor microenvironment (TME) of primary and recurrent aGCTs and to identify correlates of disease recurrence. Total RNA sequencing (RNA-seq) was performed on 24 pathologically confirmed, cryopreserved aGCT samples, including 8 primary and 16 recurrent tumors. After read alignment and quality-control filtering, DESeq2 was used to identify differentially expressed genes (DEG) between primary and recurrent tumors. Functional enrichment pathway analysis and gene set enrichment analysis was performed using “clusterProfiler” and “GSVA” R packages. TME composition was investigated through the analysis and integration of multiple published RNA-seq deconvolution algorithms. TME analysis results were externally validated using data from independent previously published RNA-seq datasets. A total of 31 DEGs were identified between primary and recurrent aGCTs. These included genes with known function in hormone signaling such as LHCGR and INSL3 (more abundant in primary tumors) and CYP19A1 (more abundant in recurrent tumors). Gene set enrichment analysis revealed that primarily immune-related and hormone-regulated gene sets expression was increased in recurrent tumors. Integrative TME analysis demonstrated statistically significant depletion of cancer-associated fibroblasts in recurrent tumors. This finding was confirmed in multiple independent datasets. Implications: 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.

Sex cord-stromal tumors of the ovary: road map for progress

Comparative genomics of high grade neuroendocrine carcinoma of the cervix

In order to improve treatment selection for high grade neuroendocrine carcinomas of the cervix (NECC), we performed a comparative genomic analysis between this rare tumor type and other cervical cancer types, as well as extra-cervical neuroendocrine small cell carcinomas of the lung and bladder. We performed whole exome sequencing on fresh-frozen tissue from 15 NECCs and matched normal tissue. We then identified mutations and copy number variants using standard analysis pipelines. Published mutation tables from cervical cancers and extra-cervical small cell carcinomas were used for comparative analysis. Descriptive statistical methods were used and a two-sided threshold of P < .05 was used for significance. In the NECC cohort, we detected a median of 1.7 somatic mutations per megabase (range 1.0-20.9). PIK3CA p.E545K mutations were the most frequency observed oncogenic mutation (4/15 tumors, 27%). Activating MAPK pathway mutations in KRAS (p.G12D) and GNAS (p.R201C) co-occurred in two tumors (13%). In total we identified PI3-kinase or MAPK pathway activating mutations in 67% of NECC. When compared to NECC, lung and bladder small cell carcinomas exhibited a statistically significant higher rate of coding mutations (P < .001 for lung; P = .001 for bladder). Mutation of TP53 was uncommon in NECC (13%) and was more frequent in both lung (103 of 110 tumors [94%], P < .001) and bladder (18 of 19 tumors [95%], P < .001) small cell carcinoma. These comparative genomics data suggest that NECC may be genetically more similar to common cervical cancer subtypes than to extra-cervical small cell neuroendocrine carcinomas of the lung and bladder. These results may have implications for the selection of cytotoxic and targeted therapy regimens for this rare disease.

PPP2R1A mutations portend improved survival after cancer immunotherapy

Immune checkpoint blockade (ICB) therapy is effective against many cancers, although resistance remains a major issue and new strategies are needed to improve clinical outcomes

Gain-of-Function Chromatin Remodeling Activity of Oncogenic FOXL2C134W Reprograms Glucocorticoid Receptor Occupancy to Drive Granulosa Cell Tumors

Abstract Adult type ovarian granulosa cell tumors (AGCT) are rare malignancies with the near universal c.C402G (p.Cys134Trp) somatic mutation in FOXL2, a forkhead box family transcription factor important for ovarian function. Relapsed AGCT is incurable, but the mechanism of the unique FOXL2 mutation could confer therapeutic vulnerabilities. To identify FOXL2C134W-dependent pharmacologic synergies, we created and characterized endogenous FOXL2 isogenic AGCT cells and an AGCT tumoroid biobank. A drug screen identified that glucocorticoids promote FOXL2C134W-dependent AGCT growth. Epigenetic investigation revealed that the Cys134Trp mutation exposes latent DNA sequence–specific chromatin remodeling activity in FOXL2. FOXL2C134W-dependent chromatin remodeling activity redirected glucocorticoid receptor chromatin occupancy to drive hyaluronan synthase 2 gene expression and increase extracellular hyaluronan secretion. Treatment of AGCT models with hyaluronidase reduced viability, and dexamethasone rescued this effect. Combinatorial drug–drug interaction experiments demonstrated that dexamethasone antagonizes the potency of paclitaxel, a chemotherapy agent frequently used in the treatment of AGCT. Thus, gain-of-function pioneering activity contributes to the oncogenic mechanism of FOXL2C134W and creates a potentially targetable synergy with glucocorticoid signaling. Significance: Glucocorticoids promote granulosa cell tumor growth via epigenetic coregulation with the disease driver FOXL2C134W, providing mechanistic insight into disease oncogenesis and uncovering a potential treatment strategy.