KAP1 (TRIM28) is a transcriptional regulator in embryonic development that controls stem cell self-renewal, chromatin organization and the DNA damage response, acting as an essential co-repressor for KRAB family zinc finger proteins (KRAB-ZNF). To gain insight into the function of this large gene family, we developed an antibody that recognizes the conserved zinc fingers linker region (ZnFL) in multiple KRAB-ZNF. Here we report that the expression of many KRAB-ZNF along with active SUMOlyated KAP1 is elevated widely in human breast cancers. KAP1 silencing in breast cancer cells reduced proliferation and inhibited the growth and metastasis of tumor xenografts. Conversely, KAP1 overexpression stimulated cell proliferation and tumor growth. In cells where KAP1 was silenced, we identified multiple downregulated genes linked to tumor progression and metastasis, including EREG/epiregulin, PTGS2/COX2, MMP1, MMP2 and CD44, along with downregulation of multiple KRAB-ZNF proteins. KAP1-dependent stabilization of KRAB-ZNF required direct interactions with KAP1. Together, our results show that KAP1-mediated stimulation of multiple KRAB-ZNF contributes to the growth and metastasis of breast cancer.

The detachment of normal epithelial cells from matrix triggers an apoptotic response known as anoikis, during homeostatic turnover. Metastatic tumor cells evade anoikis, by mechanisms that are only partly characterized. In particular, the epithelial–mesenchymal transition (EMT) in a subset of invasive tumor cells confers anoikis-resistance. In some cases, EMT up-regulates the cancer stem cell marker CD44S and the enzyme hyaluronic acid synthase-2 (HAS2). CD44S is the major receptor for hyaluronan in the extracellular matrix. Herein, we demonstrate that CD44S, unlike the CD44E isoform expressed in normal epithelial cells, contributes to the protection against anoikis. This protection requires the interaction of CD44S with hyaluronan (HA). CD44S–HA interaction is proposed to play an important role in tumor metastasis through enhanced cell survival under detached conditions.

The transcriptional regulator TRIM28 plays an important role in development, stem cells self-renewal, chromatin organization and DNA damage response. KAP1 is an essential co-repressor for KRAB zinc finger proteins (KRAB-ZNFs). Though KRAB-ZNFs represent the largest family of human transcription factors, their biological functions are largely unknown. We showed that KAP1 and certain KRAB-ZNFs were overexpressed in breast tumors and breast cancer cell lines at both mRNA and protein levels. More significantly, an active SUMOylated form of KAP1 was markedly increased in breast cancer cells. KAP1 depletion in several breast cancer cell lines slowed cell proliferation and inhibited primary tumor growth and metastasis of MDA-MB-231LN xenografts, while KAP1 overexpression conversely stimulated cell proliferation and tumor growth. KAP1 knockdown led to down-regulation of PTGS2/COX2, EREG, CD44, MMP1, MMP2 and ID transcription factors, previously implicated in cancer progression and metastasis. These results indicate that KAP1 and KRAB-ZNFs are up-regulated in breast cancer cells and contribute to increased cell proliferation, tumor growth and metastasis. Citation Format: Joseph Addison, Colton Koontz, Sarah McLaughlin, Ryan Livengood, Elena Pugacheva, Dongquan Chen, Chad Creighton, Alexey V. Ivanov. TRIM28 is overexpressed in breast cancer and regulates the expression of multiple cancer-associated genes. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-2. doi:10.1158/1538-7445.AM2014-LB-2