Periodic Reporting for period 1 - EC-InterCom (Pyrimidine de novo synthesis in tumor endothelium: an overlooked target?)
Reporting period: 2023-01-01 to 2024-12-31
Endothelial cells form the wall of blood vessels and are at the blood-tissue interface. All systemically applied anticancer treatments must cross the layer of endothelial cells to reach their intended targets within the cancer cells. For this reason, endothelial cells are a major cell type exposed to systemically applied cancer therapy. Therapy directed at nucleotide metabolism is the oldest widely used cancer treatment, yet how targeting nucleotide metabolism impacts endothelial cells and tumor endothelium is not known. Here I aimed to investigate how specific endothelial disruption of de novo pyrimidine synthesis, a pathway that provides cells with pyrimidine nucleotides for proliferation, affects tumor growth, angiogenesis, and metabolic interactions within tumors. The key objectives were (i) to characterize the metabolic communication between endothelial cells and other tumor-associated cell types, (ii) to assess whether endothelial de novo pyrimidine synthesis contributes to angiogenesis (new vessel formation) and tumor vascularization, and (iii) to identify novel metabolic targets in endothelial cells that could enhance the efficacy of pyrimidine de novo synthesis inhibitors in vivo.
I focused on lung tumors, as lungs are highly vascularized organ, and I thus expected that the impact of endothelial pyrimidine de novo synthesis disruption in the lung would be significant. I used in vitro and in vivo models to study how disruption of endothelial de novo pyrimidine synthesis affects angiogenesis and tumor growth. In vitro I disabled an essential biosynthetic enzyme of de novo synthesis pathway by Crispr/Cas9 editing in primary human umbilical cord endothelial cells, and in vivo I used an inducible endothelium-specific mouse model. I assessed multiple angiogenic parameters as well as tumor growth kinetics and analyzed tumors and their environment by multi-omics approaches. I integrated single-cell RNA sequencing to map transcriptional changes in all cells of tumor microenvironment, spectral flow cytometry to characterize immune populations, spatial transcriptomics (Visium) to assess transcriptional alterations in the tissue architecture, and metabolic spatial MALDI imaging and LC-MS metabolomics to profile metabolic changes in the tumor.
I found that de novo pyrimidine synthesis in the endothelium contributes to angiogenesis and its ablation leads to a complex, although overall negative, effect on the angiogenic capacity. Surprisingly, pyrimidine de novo synthesis deletion in the endothelium resulted in accelerated lung tumor growth, suggesting that the endothelial pyrimidine synthesis pathway has an anti-tumorigenic effect (unlike in cancer cells, where it is pro-tumorigenic). The increased tumor growth could not be ascribed to elevated tumor angiogenesis and blood supply. Instead, my data suggest this is due to an altered tumor immune environment and/or direct metabolite exchange between endothelial cells and tumor cells. Overall, my research predicts that suppression of de novo synthesis pathway in the endothelium will reduce the efficacy of cancer therapy.