Inactivation of endothelial ZEB1 impedes tumor progression and sensitizes tumors to conventional therapies
Rong Fu, … , Tao Lu, Zhao-Qiu Wu
Rong Fu, … , Tao Lu, Zhao-Qiu Wu
Published February 10, 2020
Citation Information: J Clin Invest. 2020;130(3):1252-1270. https://doi.org/10.1172/JCI131507.
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Research Article Angiogenesis

Inactivation of endothelial ZEB1 impedes tumor progression and sensitizes tumors to conventional therapies

Abstract

Current antiangiogenic therapy is limited by its cytostatic property, scarce drug delivery to the tumor, and side toxicity. To address these limitations, we unveiled the role of ZEB1, a tumor endothelium–enriched zinc-finger transcription factor, during tumor progression. We discovered that the patients who had lung adenocarcinomas with high ZEB1 expression in tumor endothelium had increased prevalence of metastases and markedly reduced overall survival after the diagnosis of lung cancer. Endothelial ZEB1 deletion in tumor-bearing mice diminished tumor angiogenesis while eliciting persistent tumor vascular normalization by epigenetically repressing TGF-β signaling. This consequently led to improved blood and oxygen perfusion, enhanced chemotherapy delivery and immune effector cell infiltration, and reduced tumor growth and metastasis. Moreover, targeting vascular ZEB1 remarkably potentiated the anticancer activity of nontoxic low-dose cisplatin. Treatment with low-dose anti–programmed cell death protein 1 (anti–PD-1) antibody elicited tumor regression and markedly extended survival in ZEB1-deleted mice, conferring long-term protective anticancer immunity. Collectively, we demonstrated that inactivation of endothelial ZEB1 may offer alternative opportunities for cancer therapy with minimal side effects. Targeting endothelium-derived ZEB1 in combination with conventional chemotherapy or immune checkpoint blockade therapy may yield a potent and superior anticancer effect.

Authors

Rong Fu, Yi Li, Nan Jiang, Bo-Xue Ren, Chen-Zi Zang, Li-Juan Liu, Wen-Cong Lv, Hong-Mei Li, Stephen Weiss, Zheng-Yu Li, Tao Lu, Zhao-Qiu Wu

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Figure 10

ZEB1 deletion epigenetically represses TGF-β signaling in LLC tumor ECs.

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ZEB1 deletion epigenetically represses TGF-β signaling in LLC tumor ECs....
(A) Comparison of indicated gene expression in the in vitro–cultured control (i.e., Ad–β-gal–infected) versus ZEB1-deleted (i.e., Ad-Cre–infected) LLC tumor ECs (n = 3 independent experiments). (B) Immunoblot analysis of control and ZEB1-deleted tumor ECs as described in A. (C) Luciferase reporter analysis of control and ZEB1-deleted LLC-ECs that were transfected with Tgfb1, Tgfb2, and Tgfb3 promoter reporter constructs (n = 3 independent experiments). Right panel: immunoblot analysis of ZEB1 expression in control and ZEB1-deleted LLC-ECs. (D) ChIP-PCR for confirming the loading of ZEB1 on the proximal but not distal promoters of Tgfb1, Tgfb2, and Tgfb3 in LLC-ECs (n = 3 independent experiments). Around 5 × 106 LLC-ECs were used for each ChIP-PCR experiment. (E) ChIP-PCR for analyzing the enrichments of H3K4Ac, H3K14Ac, H3K18Ac, and H3K27me3 on Tgfb1 promoter in control and ZEB1-deleted LLC-ECs (n = 3 independent experiments). (F) IP analysis confirming association of endogenous ZEB1 with CBP or p300 in LLC-ECs (n = 3 independent experiments). (G) Sequential ChIP-PCR for analyzing the co-occupancy of ZEB1 and CBP on the promoters of Tgfb1, Tgfb2, and Tgfb3 (n = 3 independent experiments). (H) Luciferase reporter assays for analyzing Tgfb1 promoter activity in HEK293T cells that were cotransfected with the indicated constructs (n = 3 independent experiments). Right panel: immunoblot analysis confirming ectopic expression of HA-tagged ZEB1, p300, and CBP in HEK293T cells. Asterisks and arrows mark nonspecific bands and specific bands, respectively, with the expected molecular weights. All data are represented as mean ± SD. *P < 0.05; **P < 0.01. Differences were tested using unpaired 2-sided Student’s t test (A and C) and 1-way ANOVA with Tukey’s post hoc test (H).