Abstract
BackgroundMacrophage migration inhibitory factor (MIF) is a widely expressed cytokine involved in a variety of cellular processes including cell cycle regulation and the control of proliferation. Overexpression of MIF has been reported in a number of cancer types and it has previously been shown that MIF is upregulated in melanocytic tumours with the highest expression levels occurring in malignant melanoma. However, the clinical significance of high MIF expression in melanoma has not been reported.MethodsMIF expression was depleted in human melanoma cell lines using siRNA-mediated gene knockdown and effects monitored using in vitro assays of proliferation, cell cycle, apoptosis, clonogenicity and Akt signalling. In silico analyses of expression microarray data were used to correlate MIF expression levels in melanoma tumours with overall patient survival using a univariate Cox regression model.ResultsKnockdown of MIF significantly decreased proliferation, increased apoptosis and decreased anchorage-independent growth. Effects were associated with reduced numbers of cells entering S phase concomitant with decreased cyclin D1 and CDK4 expression, increased p27 expression and decreased Akt phosphorylation. Analysis of clinical outcome data showed that MIF expression levels in primary melanoma were not associated with outcome (HR = 1.091, p = 0.892) whereas higher levels of MIF in metastatic lesions were significantly associated with faster disease progression (HR = 2.946, p = 0.003 and HR = 4.600, p = 0.004, respectively in two independent studies).ConclusionsOur in vitro analyses show that MIF functions upstream of the PI3K/Akt pathway in human melanoma cell lines. Moreover, depletion of MIF inhibited melanoma proliferation, viability and clonogenic capacity. Clinically, high MIF levels in metastatic melanoma were found to be associated with faster disease recurrence. These findings support the clinical significance of MIF signalling in melanoma and provide a strong rationale for both targeting and monitoring MIF expression in clinical melanoma.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2407-14-630) contains supplementary material, which is available to authorized users.
Highlights
Macrophage migration inhibitory factor (MIF) is a widely expressed cytokine involved in a variety of cellular processes including cell cycle regulation and the control of proliferation
Western blotting analysis identified two sequences (MIF-21 and MIF-25) that were effective in reducing MIF levels in melanoma cell lines (Additional file 1: Figure S1A and B)
Comparison of MelCV and Me1007 melanoma cells transfected with MIF-25 siRNAs confirmed a substantial reduction in the total MIF protein measured in cell lysates relative to negative control (NC) siRNA treatment (Figure 1A and B, respectively)
Summary
Macrophage migration inhibitory factor (MIF) is a widely expressed cytokine involved in a variety of cellular processes including cell cycle regulation and the control of proliferation. High levels of MIF have been reported in vivo in several cancer types including breast [5], lung [6] and gastric cancers [7] and the work of several groups points to a correlation between MIF expression and cancer prognosis, e.g. head and neck cancer and glioblastoma [8,9,10]. Findings that MIF is involved in critical pro-survival signalling pathways together with cell cycle control has provided interest in possible associations with the development and progression of cancer. MIF has been shown to play a role in cell proliferation where it has been suggested to be involved in the development and progression of cancer, acting as an extracellular, pro-tumourigenic factor [14,15]
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