Abstract
BackgroundActive fibroblasts, the predominant and the most active cells of breast cancer stroma, are responsible for tumor growth and spread. However, the molecular mediators and pathways responsible for stromal fibroblast activation, and their paracrine pro-carcinogenic effects are still not well defined. The CHEK2 tumor suppressor gene codes for a protein kinase, which plays important roles in the cellular response to various genotoxic stresses.MethodsImmunoblotting, quantitative RT-PCR and Immunofluorescence were used to assess the expression of CHEK2 in different primary breast fibroblasts and in tissues. The effect of CHEK2 on the expression and secretion of SDF-1 and IL-6 was evaluated by immunoblotting and ELISA. The WST-1 colorimetric assay was used to assess cell proliferation, while the BD BioCoat Matrigel invasion chambers were utilized to determine the effects of CHEK2 on the migratory and the invasiveness capacities of breast stromal fibroblasts as well as breast cancer cells.ResultsWe have shown that CHEK2 is down-regulated in most cancer-associated fibroblasts (CAFs) as compared to their corresponding tumor counterpart fibroblasts (TCFs) at both the mRNA and protein levels. Interestingly, CHEK2 down-regulation using specific siRNA increased the expression/secretion of both cancer-promoting cytokines SDF-1 and IL-6, and transdifferentiated stromal fibroblasts to myofibroblasts. These cells were able to enhance the proliferation of non-cancerous epithelial cells, and also boosted the migration/invasion abilities of breast cancer cells in a paracrine manner. The later effect was SDF-1/IL-6-dependent. Importantly, ectopic expression of CHEK2 in active CAFs converted these cells to a normal state, with lower migration/invasion capacities and reduced paracrine pro-carcinogenic effects.ConclusionThese results indicate that CHEK2 possesses non-cell-autonomous tumor suppressor functions, and present the Chk2 protein as an important mediator in the functional interplay between breast carcinomas and their stromal fibroblasts.
Highlights
Active fibroblasts, the predominant and the most active cells of breast cancer stroma, are responsible for tumor growth and spread
We have shown that CHEK2 inhibits the procarcinogenic effects of breast stromal fibroblasts and has a noncell-autonomous tumor suppressive function through repressing the expression/secretion of Stromal-derived factor-1 (SDF-1) and IL-6
CHEK2 is down-regulated in cancer-associated fibroblasts We started the present study by assessing the level of the Chk2 protein in Cancer-associated fibroblasts (CAFs) and their corresponding tumor counterpart fibroblasts (TCFs) isolated from the same patients
Summary
The predominant and the most active cells of breast cancer stroma, are responsible for tumor growth and spread. Cancer-associated fibroblasts (CAFs)-related procarcinogenic effects are mediated through paracrine factors, CHEK2 is another tumor suppressor gene, which is implicated in the pathogenesis of various types of sporadic tumors and is a low penetrance-predisposing gene to sarcoma, brain tumors and familial breast cancer [5]. Al-Rakan et al BMC Cancer (2016) 16:575 exon 3 These 2 mutations are associated with approximately 2- fold increased risk of breast cancer [5,6,7]. A novel recurrent CHEK2 Y390C mutation has been recently identified in high-risk Chinese breast cancer patients. This mutation impairs CHEK2 activity and is associated with increased breast cancer risk [8]
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