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@ARTICLE{Sananes:136966,
      author       = {A. Sananes and I. Cohen and A. Shahar and A. Hockla and E.
                      De Vita$^*$ and A. Miller$^*$ and E. S. Radisky and N. Papo},
      title        = {{A} potent, proteolysis-resistant inhibitor of
                      kallikrein-related peptidase 6 ({KLK}6) for cancer therapy,
                      developed by combinatorial engineering.},
      journal      = {The journal of biological chemistry},
      volume       = {293},
      number       = {33},
      issn         = {1083-351X},
      address      = {Bethesda, Md.},
      publisher    = {Soc.},
      reportid     = {DKFZ-2018-01395},
      pages        = {12663 - 12680},
      year         = {2018},
      abstract     = {Human tissue kallikrein (KLK) proteases are hormone-like
                      signaling molecules with important functions in cancer
                      pathophysiology. KLK-related peptidase 6 (KLK6),
                      specifically, is highly up-regulated in several types of
                      cancer, where its increased activity promotes cancer
                      invasion and metastasis. This characteristic suggests KLK6
                      as an attractive target for therapeutic interventions.
                      However, inhibitors that specifically target KLK6 have not
                      yet been reported, possibly because KLK6 shares a high
                      sequence homology and structural similarity with other
                      serine proteases and resists inhibition by many polypeptide
                      inhibitors. Here, we present an innovative combinatorial
                      approach to engineering KLK6 inhibitors via flow
                      cytometry-based screening of a yeast-displayed mutant
                      library of the human amyloid precursor protein Kunitz
                      protease inhibitor domain (APPI), an inhibitor of other
                      serine proteases, such as anionic and cationic trypsins. On
                      the basis of this screening, we generated
                      APPIM17L,I18F,S19F,F34V (APPI-4M), an APPI variant with a
                      KLK6 inhibition constant (Ki ) of 160 pm and a turnover time
                      of 10 days. To the best of our knowledge, APPI-4M is the
                      most potent KLK6 inhibitor reported to date, displaying
                      146-fold improved affinity and 13-fold improved proteolytic
                      stability compared with WT APPI (APPIWT). We further
                      demonstrate that APPI-4M acts as a functional inhibitor in a
                      cell-based model of KLK6-dependent breast cancer invasion.
                      Finally, the crystal structures of the APPIWT/KLK6 and
                      APPI-4M/KLK6 complexes revealed the structural and
                      mechanistic bases for the improved KLK6 binding and
                      proteolytic resistance of APPI-4M. We anticipate that
                      APPI-4M will have substantial translational potential as
                      both imaging agent and therapeutic.},
      cin          = {G404},
      ddc          = {570},
      cid          = {I:(DE-He78)G404-20160331},
      pnm          = {317 - Translational cancer research (POF3-317)},
      pid          = {G:(DE-HGF)POF3-317},
      typ          = {PUB:(DE-HGF)16},
      pubmed       = {pmid:29934309},
      pmc          = {pmc:PMC6102146},
      doi          = {10.1074/jbc.RA117.000871},
      url          = {https://inrepo02.dkfz.de/record/136966},
}