J. Roth et al. / Bioorg. Med. Chem. Lett. 21 (2011) 7180–7184
7183
Figure 3. Lineweaver–Burk plot of MMP-13 inhibition of fTHP-15 hydrolysis by compound 20 (left) or 24 (right).
References and notes
Table 4
IC50 or Ki values (
20, and 24
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are selective exosite-binding inhibitors of MMP-13, and thus
represent leads for the development of the next generation in
metalloproteinase therapeutics.
We initially assumed that compound 4 would bind within the
same S10 specificity pocket as compound 1, and designed multiple
analogs to achieve interactions present in the crystal structure of
compound 1 bound to MMP-13.4e However, because none of these
changes led to substantially improved activity in the compound 4
analogs, we suspected that compound 4 and its analogs did not
bind in the S10 pocket as did compound 1. Preliminary hydro-
gen–deuterium exchange mass spectrometry experiments6 found
protection of MMP-13 from exchange by compound 1 at the previ-
ously observed site of compound 1 binding,4e but no protection
from exchange by compound 20 at this site (M. Chalmers, P. Griffin,
P. Hodder, G. Fields, and W. R. Roush, unpublished results). Dual
inhibition kinetic assays4f demonstrated non-mutually exclusive
binding between compound 4 and either compound 1 or 3 (D. Min-
ond, T. P. Spicer, W. R. Roush, G. B. Fields, and P. S. Hodder, manu-
script in preparation). We are pursuing detailed biochemical
characterization and competition experiments to further evaluate
the unique binding of compound 4 and its analogs, and will report
those results in due course.
Acknowledgments
This research was supported by the NIH Molecular Library
Screening Center Network Grant U54MH074404 (Dr. Hugh Rosen,
Principal Investigator) and NIH R01CA098799 (G.B.F.). We thank
Dr. Patrick Griffin and Dr. Michael Chalmers, of the Scripps Florida
Department of Molecular Therapeutics, for the preliminary hydro-
gen–deuterium exchange mass spectrometry experiments de-
signed to identify the binding site for the MMP-13 exosite
inhibitors 4, 20, and 24.
5. Lauer-Fields, J. L.; Minond, D.; Chase, P. S.; Baillargeon, P. E.; Saldanha, S. A.;
Stawikowska, R.; Hodder, P.; Fields, G. B. Bioorg. Med. Chem. 2009, 17,
990.
6. Lauer-Fields, J. L.; Chalmers, M. J.; Busby, S. A.; Minond, D.; Griffin, P. R.; Fields,
G. B. J. Biol. Chem. 2009, 284, 24017.
Supplementary data
7. Ki values were determined by non-linear regression (hyperbolic equation)
analysis using the mixed inhibition model which allows for simultaneous
determination of the mechanism of inhibition.10 The mechanism of inhibition
was determined using the ‘‘alpha’’ parameter derived from the mixed-model
Supplementary data associated with this article can be found, in