Many cells comprise the tumor microenvironment, but endothelial cells are particularly crucial. They line blood vessels, regulating blood flow and inflammation, and their cell junctions are the gateway for life-giving nutrients or tumor-killing immune cells. As such, they represent the first contact of immune cells with the tumor microenvironment. It is not surprising, then, that many tumors recruit endothelial cells to promote angiogenesis—blood vessel formation—without which tumors cannot grow much bigger than a few millimeters in diameter.
A highly vascularized cancer that is especially reliant on tumor endothelial cells is clear cell renal cell carcinoma (ccRCC). Almost 80% of malignant tumors in the kidney are ccRCC, and the 5-year survival rate remains disappointing. Current standard of care is a combination of drugs that target vascular growth factor receptors in order to kill tumor endothelial cells (TECs) and deprive the tumor of its blood supply in combination with antibodies that augment anti-tumor immune responses.
There are still unanswered questions of what factors make TECs support tumor growth. Previous studies have investigated the overall tumor microenvironment via single-cell RNA profiling, but none have analyzed TECs specifically to determine if they differ from normal endothelial cells. To close this gap, Dr. Yuexin Xu, a staff scientist in Dr. Edus “Hootie” Warren’s lab, performed a deep dive into TEC identity in a study recently published in BJC Reports. Their lab used flow cytometry to specifically enrich for either TECs or normal endothelial cells (NECs), then used single cell sequencing to analyze each cell type, similar to how they previously profiled infiltrating immune cells in ccRCC tumors.
Dr. Xu and colleagues already knew that NECs need to have a variety of gene expression patterns to specialize as venous, arterial, capillary, or lymphatic vasculatures. By contrast, they found that TECs have a more homogenous phenotype. TECs also express more angiogenic factors compared to NECs, and they often down-regulate MHC markers, which could help the tumors evade detection by the immune system.
There was one gene that stood out as a reliable and specific marker for TECs: insulin-like growth factor-binding protein 3 (IGFBP-3). This caught the authors’ attention, as IGFBP3 has been used as a biomarker for other cancers. “Increased expression of IGFBP3 in RCC (and also in lung and colorectal cancer) was associated with inferior overall survival”, the study points out. “However, direct targeting of IGFBP3 as a common TEC-directed therapy may not be straightforward since it appears to have tumor-promoting and anti-tumor effects in different cancers.”
Xu also compared endothelial cells from ccRCC to those from a study on liver cancer (hepatocellular carcinoma, or HCC). She found that normal endothelial cells differed in gene expression patterns by over 50%, suggesting that these cells have organ-specific roles. In contrast, not only are TECs within each tumor type homogenous, more of their gene expression profiles were shared between the two separate cancers than for their NEC counterparts. This means that TECs from different tumors share similar tumor-promoting phenotypes. It also raises the possibility of using a common TEC targeting strategy to treat different types of tumor.
One of the challenges of studying TECs is the ability to recapitulate their unique cell phenotypes in tissue culture. To meet this challenge, Dr. Xu and her colleagues turned to their longtime collaboration with Dr. Scott Tykodi and Dr. Shreeram Akilesh, with whom they recently developed a 3D model of renal carcinoma. They used similar techniques to successfully culture both TECs and NECs isolated from ccRCC nephrectomy surgeries that could stably retain their phenotypes for 2-3 passages.
With a stable model in hand, they could conduct experiments to understand the phenotypic differences between TECs and NECs. Interestingly, they found that TECs have enhanced binding to leukocytes such as T cells or monocytes but not natural killer cells. They observed that “CD4+CD25+ T regulatory cells were much more likely to adhere to TECs versus NECs,” suggesting that immunosuppressive T cells are allowed to bypass the TEC barrier and enter the tumor, while other subsets of immune cells are prevented from doing so. “In this way, due to their altered binding affinity for different immune cell subsets, TECs may play a critical role in conditioning the tumor microenvironment to dampen immune responses and promote tumor growth,” says Dr. Akilesh.
These findings show how much more there is to learn about TECs, and how gaining this knowledge may “provide opportunities to exploit or modify their phenotypes as a therapeutic strategy,” the authors conclude. Dr. Akilesh adds: “Understanding the phenotype of TECs will be important in improving the effectiveness of T cell-based therapies targeting the tumor. The shared phenotypes of TECs across tumor types also raises the hope that this knowledge may be applied to treatment of diverse cancer types.” Knowledge opens doors, and this knowledge is the key to understanding the gateway into the tumor.
Fred Hutch/University of Washington/Seattle Children’s Cancer Consortium members Drs. Ying Zheng, Edus Warren, Scott Tykodi, and Shreeram Akilesh contributed to this research.
The spotlighted research was funded by the Department of Defense, the National Cancer Institute, and the Fred Hutchinson Cancer Center/UW/Seattle Children’s Cancer Consortium and Cancer Therapeutics Endowment.
Xu, Y., Miller, C.P., Xue, J., Ying, Z., Warren, E.H., Tykodi, S.S., Akilesh, S. (2024). Single cell atlas of kidney cancer endothelial cells reveals distinct expression profiles and phenotypes. BJC Rep. 2024 Mar 14;2(1):23. doi: 10.1038/s44276-024-00047-9
Hannah Lewis is a postdoctoral research fellow with Jim Boonyaratanakornkit’s group in the Vaccine and Infectious Disease Division (VIDD). She is developing screens to find rare B cells that produce protective antibodies against human herpesviruses. She obtained her PhD in molecular and cellular biology from the University of Washington.
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