The transcription factor LSF: a novel oncogene for hepatocellular carcinoma

Abstract

The transcription factor LSF (Late SV40 Factor), also known as TFCP2, belongs to the LSF/CP2 family related to Grainyhead family of proteins and is involved in many biological events, including regulation of cellular and viral promoters, cell cycle, DNA synthesis, cell survival and Alzheimer's disease. Our recent studies establish an oncogenic role of LSF in Hepatocellular carcinoma (HCC). LSF overexpression is detected in human HCC cell lines and in more than 90% cases of human HCC patients, compared to normal hepatocytes and liver, and its expression level showed significant correlation with the stages and grades of the disease. Forced overexpression of LSF in less aggressive HCC cells resulted in highly aggressive, angiogenic and multi-organ metastatic tumors in nude mice. Conversely, inhibition of LSF significantly abrogated growth and metastasis of highly aggressive HCC cells in nude mice. Microarray studies revealed that as a transcription factor LSF modulated specific genes regulating invasion, angiogenesis, chemoresistance and senescence. LSF transcriptionally regulates thymidylate synthase (TS) gene, thus contributing to cell cycle regulation and chemoresistance. Our studies identify a network of proteins, including osteopontin (OPN), Matrix metalloproteinase-9 (MMP-9), c-Met and complement factor H (CFH), that are directly regulated by LSF and play important role in LSF-induced hepatocarcinogenesis. A high throughput screening identified small molecule inhibitors of LSF DNA binding and the prototype of these molecules, Factor Quinolinone inhibitor 1 (FQI1), profoundly inhibited cell viability and induced apoptosis in human HCC cells without exerting harmful effects to normal immortal human hepatocytes and primary mouse hepatocytes. In nude mice xenograft studies, FQI1 markedly inhibited growth of human HCC xenografts as well as angiogenesis without exerting any toxicity. These studies establish a key role of LSF in hepatocarcinogenesis and usher in a novel therapeutic avenue for HCC, an invariably fatal disease.

Keywords: FQI1; Late SV40 Factor (LSF); angiogenesis; c-Met; cell cycle regulation; hepatocellular carcinoma (HCC); matrix metalloproteinase-9 (MMP-9); metastasis; osteopontin (OPN); small molecule inhibitors; thymidylate synthase (TS).

Figure 1

Schematic structure of human LSF protein. The numbers indicate amino acid residues. NLS: nuclear localizationsignal.

Figure 2

LSF inhibition causes slower cell cycle progression through S-phase. Control-1 and LSFdn-15 clones of QGY-7703 cells were synchronized by double thymidine block and then allowed to enter the cell cycle by thymidine removal. Samples were collected every two hours and cell cycle was analyzed by flow cytometry of propidium iodide-stained DNA. A. Histograms of the cell cycle profile. B. Graphical representation of the percentage of cells in eachphase of the cell cycle.

Figure 3

Dual color 2D DIGE detecting protein expression in Control-1 and LSFdn-15 clones of QGY-7703 cells. Leftpanel shows single color and overlay images. Right panel shows peak intensities of Spot 20 and Spot 43. Spot 20and Spot 43 represent Dihydrofolate reductase (DHFR) and thymidylate synthase (TS), respectively. LSF and hepatocellular carcinoma (HCC).

Figure 4

Model for molecular mechanism by which LSF promotes hepatocarcinogenesis. LSF transcriptionally upregulates osteopontin (OPN), matrix metalloproteinase-9 (MMP-9), complement factor H (CFH) and fibronectin 1 (FN1). Secreted OPN binds to CD44 receptor resulting in activation of c-Met and its downstream signaling Akt and ERK. Collectively these events contribute to proliferation, invasion, angiogenesis and metastasis. LSF upregulates thymidylate synthase (TS) thus contributing to cell cycle progression and chemoresistance.