期刊
MOLECULES
卷 24, 期 2, 页码 -出版社
MDPI
DOI: 10.3390/molecules24020255
关键词
MT1-MMP; TIMP; renal carcinoma; cancer therapy; prion; GPI anchor; protein engineering
资金
- National Natural Science Foundation of China (NSFC) [31270850]
- Suzhou Industrial Park Supplement Fund
- Key Program Special Fund, XJTLU [KSF-E-11]
Renal carcinoma cells express Membrane Type 1-Matrix Metalloproteinase (MT1-MMP, MMP-14) to degrade extracellular matrix components and a range of bioactive molecules to allow metastasis and cell proliferation. The activity of MT1-MMP is modulated by the endogenous inhibitors, Tissue Inhibitor of Metalloproteinases (TIMPs). In this study, we describe a novel strategy that would enable a designer TIMP-1 tailored specifically for MT1-MMP inhibition (V4A/P6V/T98L; K-i(app) 1.66 nM) to be targeted to the plasma membrane for more effective MT1-MMP inhibition. To achieve this, we fuse the designer TIMP-1 to the glycosyl-phosphatidyl inositol (GPI) anchor of the prion protein to create a membrane-tethered, high-affinity TIMP variant named T1(Pr MT1) that is predominantly located on the cell surface and co-localised with MT1-MMP. Confocal microscopy shows that T1(Pr MT1) is found throughout the cell surface in particular the membrane ruffles where MT1-MMP is most abundant. Expression of T1(Pr MT1) brings about a complete abrogation of the gelatinolytic activity of cellular MT1-MMP in HT1080 fibrosarcoma cells whilst in renal carcinoma cells CaKi-1, the GPI-TIMP causes a disruption in MMP-mediated proteolysis of ECM components such as fibronectin, collagen I and laminin that consequently triggers a downstream senescence response. Moreover, the transduced cells also suffer from an impairment in proliferation and survival in vitro as well as in NOD/SCID mouse xenograft. Taken together, our findings demonstrate that the GPI anchor of prion could be exploited as a targeting device in TIMP engineering for MT1-MMP inhibition with a potential in renal carcinoma therapy.
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