Angiopoietin-2 blockade suppresses growth of liver metastases from pancreatic neuroendocrine tumors by promoting T cell recruitment
Eunhyeong Lee, … , Gavin Thurston, Minah Kim
Eunhyeong Lee, … , Gavin Thurston, Minah Kim
Published October 16, 2023
Citation Information: J Clin Invest. 2023;133(20):e167994. https://doi.org/10.1172/JCI167994.
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Research Article Angiogenesis

Angiopoietin-2 blockade suppresses growth of liver metastases from pancreatic neuroendocrine tumors by promoting T cell recruitment

Abstract

Improving the management of metastasis in pancreatic neuroendocrine tumors (PanNETs) is critical, as nearly half of patients with PanNETs present with liver metastases, and this accounts for the majority of patient mortality. We identified angiopoietin-2 (ANGPT2) as one of the most upregulated angiogenic factors in RNA-Seq data from human PanNET liver metastases and found that higher ANGPT2 expression correlated with poor survival rates. Immunohistochemical staining revealed that ANGPT2 was localized to the endothelial cells of blood vessels in PanNET liver metastases. We observed an association between the upregulation of endothelial ANGPT2 and liver metastatic progression in both patients and transgenic mouse models of PanNETs. In human and mouse PanNET liver metastases, ANGPT2 upregulation coincided with poor T cell infiltration, indicative of an immunosuppressive tumor microenvironment. Notably, both pharmacologic inhibition and genetic deletion of ANGPT2 in PanNET mouse models slowed the growth of PanNET liver metastases. Furthermore, pharmacologic inhibition of ANGPT2 promoted T cell infiltration and activation in liver metastases, improving the survival of mice with metastatic PanNETs. These changes were accompanied by reduced plasma leakage and improved vascular integrity in metastases. Together, these findings suggest that ANGPT2 blockade may be an effective strategy for promoting T cell infiltration and immunostimulatory reprogramming to reduce the growth of liver metastases in PanNETs.

Authors

Eunhyeong Lee, Sophie O’Keefe, Alessandra Leong, Ha-Ram Park, Janani Varadarajan, Subrata Chowdhury, Shannon Hiner, Sungsoo Kim, Anahita Shiva, Richard A. Friedman, Helen Remotti, Tito Fojo, Hee Won Yang, Gavin Thurston, Minah Kim

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Figure 1

Increased ANGPT2 expression in PanNET liver metastases.

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Increased ANGPT2 expression in PanNET liver metastases. (A and B) Volcan...
(A and B) Volcano plot (A) and heatmap (B) from RNA-Seq transcriptome analyses of differentially expressed angiogenesis-related genes in liver metastases of patients with PanNETs (n = 30) and normal human liver tissues (n = 119), with ANGPT2 noted in red. (A) The horizontal dotted line in the volcano plot represents a FDR of 1%; the vertical lines represent the threshold (±2-fold) of the log2 fold change. (B) The heatmap shows normalized RNA-Seq data for angiogenesis-related genes (rows) from 149 samples (columns). (C) Kaplan-Meier survival curves of patients with PanNETs with low (n = 9) or high (n = 11) concentrations of plasma ANGPT2 (cutoff, 0.66 ng/mL). (D and E) Increased ANGPT2 immunoreactivity in liver metastases of patients with PanNETs (n = 11) compared with that in normal livers (n = 5) (scale bar: 50 μm) (D) and quantification of ANGPT2+ vessels (unpaired t test) (E). (F–H) Whole-liver-lobe cross-sections showing the metastatic tumor progression of RT2;AB6F1 mice (scale bar: 1 mm) (F). Mice at 15, 18, and 20 weeks of age were stratified by percentage area of metastasis (low, <1%; medium, 1%–10%; high, >10%). Quantification of ANGPT2+ vessels (1-way ANOVA with Tukey’s multiple comparisons test) (G) and representative images showing increased ANGPT2 during metastatic growth (scale bar: 50 μm) (H). (I) Analysis of plasma ANGPT2 concentrations by ELISA (1-way ANOVA with Tukey’s multiple comparisons test). (J and K) Greater vascular leakage marked by extravasated fibrin in metastatic colonies of the liver with high ANGPT2 compared with colonies with little or no ANGPT2 staining in RT2;AB6F1 mice at 20 weeks of age (scale bar: 50 μm) (J) and its quantification (unpaired t test) (K). The cutoff value for high and low ANGPT2 was 13.8, the average ANGPT2 expression (ANGPT2/CD31, %). For E, G, I, and K, each data point represents an individual human or mouse. Data are displayed as the mean ± SEM.