SYNTHESIS OF CHIMERIC HETERO-BIVALENT NEAR-IR PROBES FOR MMP POSITIVE CELL IMAGING

Michael   Pun

Matrix metalloproteinase (MMP) is a family of zinc dependent endopeptidase responsible remodeling of the extracellular matrix. Currently there are 23 known types of MMPs in humans whom all exhibit a considerable sequence and structural homology in the pro-peptide, catalytic, and hemopexin domains. The catalytic and hemopexin domains are linked by a hinged peptide region known to have sequences and length that are unique to each MMP. MMP proteolytic activity is crucial in wound healing processes such as reepithelization, scar formation, and inflammation. But several MMPs have also been found to play key roles in metastasis, and angiogenesis in cancer. Because members of the MMP family share a large structural homology, therapeutic drug development targeting MMPs have all been fruitless thus far. We hypothesize that a specific MMP inhibitor can be synthesized by exploiting the distinct distance between the catalytic and hemopexin domains. We test this hypothesis by synthesizing a heterbivalent platform (HBI) consisting of a known catalytic and hemopexin domain inhibitor joined together by a PEG linker tuned to the specific length of an MMP. Cyanine5.5 was applied the molecule allowing for in vitro microscopy and in vivo optical imaging to evaluate the performance of the HBI. Our first attempt at HBI specificity was targeted at membrane-type I matrix metalloproteinase (MT1-MMP or MMP-14), a cell-surface transmembrane protein that is highly correlated with metastasis in breast cancer. The distance between domains of MMP-14 was measured to be ~50 Å, therefore we chose to proceed with a PEG8 spacer to synthesize HBI-8. Our initial confocal microscopy results reveal that the probe specifically targets high MMP-14 expressing MDA-MB-231 cells over lower expressing LNCaP cells. Preliminary in vivo optical imaging showed HBI-8 uptake in MMP-14 (+) tumors after 12 hours. We then synthesized HBI-4, -12 and -24 utilizing PEG-4, -12 and -24 respectively. The HBIs performance was evaluated by confocal microscopy. HBI-24 was found to have high specific uptake to LNCaP cells over MDA-MB-231 and BT-20 cells. This specific uptake correlates to qPCR data for LNCaP specific MMP-16 expression.

SYNTHESIS OF CHIMERIC HETERO-BIVALENT NEAR-IR PROBES FOR MMP POSITIVE CELL IMAGING

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