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Thiol‐Reactive Inhibitors of Cdc48/p97 AAA ATPase as Potential Anti‐Cancer Therapeutics
Author(s) -
Chou TsuiFen,
Jones Amanda C,
Stoltz Brian M.,
Deshaies Raymond J.
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.1_supplement.791.9
Subject(s) - hela , aaa proteins , cancer cell , biochemistry , atpase , chemistry , microbiology and biotechnology , in vitro , biology , cancer , enzyme , genetics
Cdc48/p97 is an important AAA ATPase not only due to its diverse cellular functions but also because it has been implicated in mediating turnover of proteins involved in tumorgenesis, such as cyclin E and IkBa, the inhibitor of nuclear factor‐kB. We have set out to develop small‐molecule inhibitors of Cdc48/p97 to use as tools to study Cdc48/p97 function and to evaluate whether it is a viable therapeutic target. Using p97 crystal structures as a guide, we first derivatizated a PP1 like compound to incorporate electrophiles that are likely to be targeted by an active‐site cysteine of wild type p97. One of the inhibitors (ACJI‐99C) identified from our initial efforts exhibited an IC 50 ~0.6 μM for wild type murine p97 and ~300 μM for wild type yeast Cdc48 and ~100 μM for hamster NSF in an in vitro ATPase activity assay. ACJI‐99C‐induced inactivation of p97 was protected by cysteine and DTT. Cys522 of p97 was shown to form a covalent bond with ACJI‐99C by tandem mass spectroscopy. Furthermore, ACJI‐99C exhibited anti‐proliferation activity toward cancer cell lines with sub‐micromolar GI 50 . After 1 hr incubation with ACJI‐99C, accumulation of poly‐ubiquitinated proteins was observed in HeLa and RPMI‐8226 cancer cells, which recapitulated the effect of p97 depletion by siRNA‐mediated knockdown. In conclusion, we have identified a cell‐permeable covalent inhibitor of p97 and demonstrated its potential to be an anti‐cancer agent.