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Supplementary Figures 1 - 3 from HOXB7 Promotes Malignant Progression by Activating the TGFβ Signaling Pathway

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posted on 2023-03-30, 23:27 authored by Shou Liu, Kideok Jin, Yvonne Hui, Jie Fu, Chunfa Jie, Sheng Feng, David Reisman, Qian Wang, Daping Fan, Saraswati Sukumar, Hexin Chen

Fig. S1. TGF-β/SMAD3 signaling mediates the promotion of migration and invasion by HOXB7. Fig. S2 Effects of HOXB7 overexpression and TGF-β2 knockdown on cellular morphology and proliferation rate. Fig. S3 Effects of HOXB7 overexpression on cellular morphology and proliferation rate of Her2 mouse mammary tumor cell line, H605

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ARTICLE ABSTRACT

Overexpression of HOXB7 in breast cancer cells induces an epithelial–mesenchymal transition and promotes tumor progression and lung metastasis. However, the underlying mechanisms for HOXB7-induced aggressive phenotypes in breast cancer remain largely unknown. Here, we report that phosphorylation of SMAD3 was detected in a higher percentage in primary mammary tumor tissues from double-transgenic MMTV-Hoxb7/Her2 mice than tumors from single-transgenic Her2/neu mice, suggesting activation of TGFβ/SMAD3 signaling by HOXB7 in breast tumor tissues. As predicted, TGFβ2 was high in four MMTV-Hoxb7/Her2 transgenic mouse tumor cell lines and two breast cancer cell lines transfected with HOXB7, whereas TGFβ2 was low in HOXB7-depleted cells. HOXB7 directly bound to and activated the TGFβ2 promoter in luciferase and chromatin immunoprecipitation assays. Increased migration and invasion as a result of HOXB7 overexpression in breast cancer cells were reversed by knockdown of TGFβ2 or pharmacologic inhibition of TGFβ signaling. Furthermore, knockdown of TGFβ2 in HOXB7-overexpressing MDA-MB-231 breast cancer cells dramatically inhibited metastasis to the lung. Interestingly, HOXB7 overexpression also induced tumor-associated macrophage (TAM) recruitment and acquisition of an M2 tumor-promoting phenotype. TGFβ2 mediated HOXB7-induced activation of macrophages, suggesting that TAMs may contribute to HOXB7-promoted tumor metastasis. Providing clinical relevance to these findings, by real-time PCR analysis, there was a strong correlation between HOXB7 and TGFβ2 expression in primary breast carcinomas. Taken together, our results suggest that HOXB7 promotes tumor progression in a cell-autonomous and non–cell-autonomous manner through activation of the TGFβ signaling pathway. Cancer Res; 75(4); 709–19. ©2014 AACR.