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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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 13

ZEB1/TGF-β/SMAD signaling transactivates Vegfa and Ang2 genes in LLC tumor ECs.

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ZEB1/TGF-β/SMAD signaling transactivates Vegfa and Ang2 genes in LLC tum...
(A) Immunoblot analysis of control and ZEB1-deleted LLC-ECs that were treated with vehicle or 2 ng/mL r.TGF-β1 protein for 3 hours. (B) Immunofluorescence analysis of VEGFA (top) and ANG2 (bottom) expression in the indicated cells as described in A. Nuclei, DAPI (blue). (C) Immunoblot (left) and qRT-PCR (right) analyses of vehicle- or r.TGF-β1–treated LLC-ECs for p-SMAD2/3 and SMAD2/3 protein expression and Vegfa and Ang2 mRNA expression (n = 3 independent experiments). (D) ChIP-PCR for assessing SMAD2/3 level on the promoters of Vegfa, Ang2, and Id1 in vehicle- and r.TGF-β1–treated LLC-ECs (n = 3 independent experiments). Around 5 × 106 LLC-ECs were used for each ChIP-PCR experiment. (E) ChIP-PCR for assessing SMAD2/3 level on the promoters of Vegfa, Ang2, and Id1 in control and ZEB1-deleted LLC-ECs (n = 3 independent experiments). Left panel: immunoblot analysis for confirming reduced ZEB1 and p-SMAD2/3 expression in ZEB1-deleted LLC-ECs. (F) ChIP-PCR showing no binding of ZEB1 on the proximal promoters of Vegfa and Ang2 in LLC-ECs (n = 3 independent experiments). (G and H) Immunoblotting (G) and qRT-PCR (H) for assessing ZEB1 protein and mRNA expression in LLC-ECs treated with vehicle, r.TGF-β1 (2 ng/mL, 3 hours), or r.VEGFA (20 ng/mL, 18 hours) (n = 3 independent experiments). (I) Schematic diagram showing the effect of vascular ZEB1 loss on tumor growth and metastasis, tumor angiogenesis, vascular functionality and integrity, and tumor immune microenvironment. All data are represented as mean ± SD. *P < 0.05; **P < 0.01. Differences were tested using unpaired 2-sided Student’s t test (C) and 1-way ANOVA with Tukey’s post hoc test (H).