Abstract
BackgroundLife of patients with uveal melanoma (UM) is largely threatened by liver metastasis. Little is known about the drivers of liver organotropic metastasis in UM. The elevated activity of transcription of oncogenes is presumably to drive aspects of tumors. We hypothesized that inhibition of transcription by cyclin-dependent kinase 7/9 (CDK7/9) inhibitor SNS-032 diminished liver metastasis by abrogating the putative oncogenes in charge of colonization, stemness, cell motility of UM cells in host liver microenvironment.MethodsThe effects of SNS-032 on the expression of the relevant oncogenes were examined by qRT-PCR and Western blotting analysis. Proliferative activity, frequency of CSCs and liver metastasis were evaluated by using NOD-SCID mouse xenograft model and NOG mouse model, respectively.ResultsThe results showed that CDK7/9 were highly expressed in UM cells, and SNS-032 significantly suppressed the cellular proliferation, induced apoptosis, and inhibited the outgrowth of xenografted UM cells and PDX tumors in NOD-SCID mice, repressed the cancer stem-like cell (CSC) properties through transcriptional inhibition of stemness-related protein Krüppel-like factor 4 (KLF4), inhibited the invasive phonotypes of UM cells through matrix metalloproteinase 9 (MMP9). Mechanistically, SNS-032 repressed the c-Myc-dependent transcription of RhoA gene, and thereby lowered the RhoA GTPase activity and actin polymerization, and subsequently inhibited cell motility and liver metastasis.ConclusionsIn conclusion, we validate a set of transcription factors which confer metastatic traits (e.g., KLF4 for CSCs, c-Myc for cell motility) in UM cells. Our results identify SNS-032 as a promising therapeutic agent, and warrant a clinical trial in patients with metastatic UM.
Highlights
Uveal melanoma (UM) is the most common intraocular malignancy in adults which arises from melanocytes of the choroid, ciliary and iris
Reagents and DNA constructs, Western blot analysis, cell viability assay, colony-formation assay, apoptosis assay by flow cytometry, measurement of mitochondrial transmembrane potential, real-time quantitative RT-PCR, dual luciferase reporter assay, melanosphereformation assay, aldehyde dehydrogenase positive cells assay, limiting dilution assay in NOD-SCID mice, woundhealing scratch assay, migration and invasion assay, lentivirus transduction, fluorescence signals of actin polymerization (F-actin) staining assay, small GTPases activities assay, chromatin immunoprecipitation assay All above methods were performed as described previously reported [23,24,25,26,27,28], with details provided in the Additional file 2
SNS-032 inhibits proliferation of uveal melanoma (UM) cells through blocking Yes-associated protein (YAP) signaling To assess the effects of SNS-032 on UM cells, we first detected the expression of cyclin-dependent kinase 7/9 (CDK7/9) in UM cells
Summary
The effects of SNS-032 on the expression of the relevant oncogenes were examined by qRT-PCR and Western blotting analysis. Proliferative activity, frequency of CSCs and liver metastasis were evaluated by using NODSCID mouse xenograft model and NOG mouse model, respectively
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