Utilizing Single Cell RNA Sequencing to Target Ovarian Cancer

Despite ovarian cancer’s rarity, affecting about 1.3% of women, it is the fifth leading cause of cancer-related deaths among women, often diagnosed late with a survival rate of less than 50% after five years. Recent FDA drug approvals have increased treatment options, specifically highlighting targeted therapies like mirvetuximab soravtansine-gynx for specific patient subsets. Single cell RNA sequencing has emerged as a powerful tool in understanding the cellular diversity within tumors, contributing to the variation in treatment efficacy and toxicity.

Recent advancements have utilized the Chromium Single Cell Immune Profiling assay to develop an in-depth tumor microenvironment atlas, focusing on primary tumors, metastases, and particularly ascites – the fluid accumulation common in advanced cases. This study identified immune cells in ascites that could potentially enhance the immune response against primary and metastatic tumors.

A pivotal finding was the identification of a specific T-cell population (CD8+ effector memory T cells expressing granzyme K) within ascites, which seems to share receptors with exhausted T cells found in tumors. These findings suggest that these T cells initially active in ascites could become exhausted upon infiltrating tumors, linking to immune response mechanisms that might be leveraged to improve treatment outcomes.

Given the immunogenic nature of ovarian cancer, despite typically poor responses to current immunotherapies, further research into the function and therapeutic potential of these T-cell populations could be crucial. This could lead to more effective strategies in managing ovarian cancer, potentially improving the prognosis for affected patients.

Unraveling the T-cell response to ovarian cancer tumors

To study the relationship between tumors, metastases, and ascites, the researchers examined samples from patients with high-grade serous ovarian cancer (HGSOC)—the most common type of ovarian cancer, accounting for 70% of all cases. 75% of people already have widespread metastases and ascites once they are diagnosed. And effective treatment options are limited.

Immunotherapies are effective for some patients—nearly 10% of 125 patients previously treated with chemotherapy responded to avelumab in a phase 1b clinical trial (5). Researchers attribute varied immunotherapeutic responses to differences in how many TILs effectively invade the primary tumor and its metastases.

For example, last year, researchers from the Moffitt Cancer Center study used our Chromium Single Cell Immune Profiling and our Single Cell Multiome ATAC + Gene Expression assays to analyze 83,454 tissue-resident memory T cells from 122 HGSOC tumors (6). They validated that ovarian cancer is an immunogenic disease—3% of total T-cell receptors (TCRs) in CD8⁺ T cells from tumors recognized ovarian cancer antigens—and, therefore, should be responsive to immunotherapies despite low response rates observed in clinical trials.

But why these tumors rarely respond to immunotherapy isn’t clear.

Rethinking the role of ascites in shaping the ovarian cancer TME

Researchers think the ascites-enriched T cells may play a critical role in the immune response to ovarian cancer tumors, and provide a path to improving immunotherapy responses in these tumors.

In the Nature Cancer study published in July, the scientists used the Chromium Single Cell Immune Profiling assay to compare immune cellular compositions of five

separate tumor-related sites—including primary ovarian tumors, omentum metastases, ascites, pelvic lymph nodes, and peripheral blood—from 14 patients with advanced ovarian cancer with varying responses to chemotherapy. They categorized cells, based on canonical gene expression markers, into five major cell lineages: lymphoid cells, myeloid cells, stromal cells, endothelial cells, and cancer cells.

As expected, primary tumors and their metastases had the same, basic cellular makeup; pelvic lymph nodes were enriched with B cells and CD4⁺ T cells; peripheral blood contained lymphocytes and monocytes.

Ascites had large numbers of stromal and immune cells, particularly CD8⁺ T cells, macrophages, and dendritic cells, which represent the classic makeup of an inflammatory environment. All ascites samples also contained cancer cells.

The researchers took a closer look at the T-cell populations in samples from patients with HGSOC. Using unsupervised clustering, they identified five CD4⁺ clusters, five CD8⁺ clusters, and two unconventional clusters. A cluster of CD8⁺ effector memory T cells highly expressing the gene-expressing protease granzyme K (GZMK) was highly enriched in astrocytes. Interestingly, previous studies found that lung cancer tumors enriched with these GZMK⁺ T cells responded better to immunotherapy.

Further analysis using the TCR-tracking algorithm STARTRAC revealed that ascites-derived GZMK⁺ T cells shared TCR sequences with exhausted T cells in primary tumors and metastases. This suggests that these cells likely infiltrate tumors where they then differentiate into exhausted T cells.

The authors suggest that, since high levels of GZMK⁺ T cells are found in lung cancer tumors associated with better lung cancer prognosis, manipulating these cells in patients could improve outcomes in ovarian cancer patients.

“As reported, pre-exhausted GZMK⁺ T subpopulations were regarded as pre-activated T cells which would accumulate in responsive lung cancer and melanoma tumors following immune-checkpoint-based treatment. We suspected that accelerating the migration of ascites-derived [GZMK⁺ CD8⁺ effector memory T cells] into tumor sites could be a potential therapeutic strategy for [ovarian cancer],” wrote the authors.

Further studies clarifying the function of this unique cell population in ovarian cancer samples and beyond could help researchers better understand varied immunotherapy responses in a variety of tumors.

References:

1. https://www.aacr.org/patients-caregivers/awareness-months/ovarian-cancer-awareness-month/
2. https://ocrahope.org/news/2022-breakthroughs-in-ovarian-cancer-research-and-treatment/#:~:text=Ovarian%20cancer%20immunotherapy&text=Some%20monoclonal%20antibodies%20are%20considered,cells%20to%20kill%20cancer%20cells.
3. https://www.fda.gov/drugs/resources-information-approved-drugs/fda-grants-accelerated-approval-mirvetuximab-soravtansine-gynx-fra-positive-platinum-resistant
4. Zheng X, et al. Single-cell analyses implicate ascites in remodeling the ecosystems of primary and metastatic tumors in ovarian cancer. Nat Cancer 4: 1138–1156 (2023). doi: 10.1038/s43018-023-00599-8
5. Disis ML, et al. Efficacy and Safety of Avelumab for Patients With Recurrent or Refractory Ovarian Cancer Phase 1b Results From the JAVELIN Solid Tumor Trial. JAMA Oncol 5: 393-401 (2019). doi:10.1001/jamaoncol.2018.6258
6. Anadon CM, et al. Ovarian cancer immunogenicity is governed by a narrow subset of progenitor tissue-resident memory T cells. Cancer Cell 40 (5): P545-557.E13 (2022). doi: 10.1016/j.ccell.2022.03.008