5570 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 23
Letters
Table 3. IC50 Values of Diaryl Isoindolin-2-yl Phosphonates (2a-h)a
IC50 (µM)
Supporting Information Available: General synthetic proce-
dures, spectral and analytical data for new compounds, and detailed
descriptions of all biochemical experiments used for their charac-
terization. This material is available free of charge via the Internet
compound
Xaa
Ar
DPP8
DPP IV
DPP II
n.a.d
2a
2b
2c
2d
2e
2f
Lys
Lys(Z)
Ile
allo-Ile
Lys
Lys(Z)
Ile
Ph-
Ph-
Ph
2.11 ( 0.23c
7.8 ( 1.5b
36 ( 2
>500
>50
2.2 ( 0.12
>250
>1000
>500
>250
>1000
>250
>250
References
Ph
16.0 ( 1.0
0.53 ( 0.11c
12.5 ( 2.5b
48 ( 3
>250
Pct
Pct
Pct
Pct
>1000
>250
(1) (a) Augustyns, K.; Van der Veken, P.; Senten, K.; Haemers, A. The
therapeutic potential of inhibitors of dipeptidyl peptidase IV (DPP
IV) and related proline specific dipeptidyl peptidases. Curr. Med.
Chem. 2005, 12, 971-998. (b) Van der Veken, P.; Haemers, A.;
Augustyns, K. Prolyl peptidases related to dipeptidyl peptidase IV:
potential for drug discovery. Curr. Topics Med. Chem. 2007, 7, 621-
635.
(2) Lambeir, A. M.; Durinx, C.; Scharpe´, S.; De Meester, I. Dipeptidyl-
peptidase IV from bench to bedside: an update on structural
properties, functions and clinical aspects of the enzyme DPP IV. Crit.
ReV. Clin. Lab. Sci. 2003, 40, 209-294.
(3) Amori, R. E.; Lau, J.; Pittas, A. G. Efficacy and safety of incretin
therapy in type II diabetes. Systematic review and meta-analysis.
JAMA, J. Am. Med. Assoc. 2007, 298, 194-206.
(4) (a) Lankas, G. R.; Leiting, B.; Roy, R. S.; Eiermann, G. J.; Beconi,
M. G.; Biftu, T.; Chan, C. C.; Edmondson, S.; Feeney, W. P. et al.
Dipeptidyl-peptidase IV inhibition for the treatment of type II
diabetessPotential importance of selectivity over dipeptidyl pepti-
dases 8 and 9. Diabetes 2005, 54, 2988-2994. (b) Lankas, G.;
Leiting, B.; Roy, R. S.; Eiermann, G. J.; Biftu, T.; Kim, D.; Ok, H.;
Weber, A.; Thornberry, N. A. Inhibition of DPP8/9 results in toxicity
of preclinical species: potential importance of selective dipeptidyl
peptidase IV inhibition for the treatment of type II diabetes. Diabetes
2004, 53, A2-A2, suppl. 2.
2g
2h
>1000
>100
allo-Ile
17.5 ( 0.2
a Ph ) phenyl; Pct ) 4-acetamidophenyl. b Mean of two independent
measurements. c Mean of three to four independent measurements. d n.a.)
not analyzed.
Table 4. Kinetic Analysis of DPP8 Inhibition by Selected Compounds
compound
Kd(µM)
kinact (s-1
)
kapp (M-1 s-1
)
7a
2e
2d
2h
7.3 ( 1.1a
0.77 ( 0.26
39 ( 6
0.005 ( 0.0006a
0.0029 ( 0.0004
0.0017 ( 0.0001
0.0013 ( 0.000046
680b
3800b
40b
9.1 ( 1.3
140b
a Mean of two measurements. b Calculated value (kinact/ Kd).
(5) (a) Powers, J. C.; Asgian, J. L.; Ekici, O. D.; James, K. E. Irreversible
inhibition of serine, cysteine and threonine proteases. Chem. ReV.
2002, 102, 4639-4750. (b) Evans, M. J.; Cravatt, B. F. Mechanism-
based profiling of enzyme families. Chem. ReV. 2006, 106, 8, 3279-
3301.
(6) (a) Jiaang, W. T.; Chen, Y. S.; Hsu, T.; Wu, S. H.; Chien, C. H.;
Chang, C. N.; Chang, S. P.; Lee, S. P.; Chen, X. Novel isoindoline
compounds for potent and selective inhibition of prolyl dipeptidase
8. Bioorg. Med. Chem. Lett. 2005, 15, 687-791. (b) Lu, I. L.; Lee,
S. J.; Tsu, H.; Kao, K. H.; Chien, C. H.; Chang, Y. Y.; Chen, Y. S.;
Cheng, J. H.; Chang, C. N.; Chen, T. W.; Chang, S. P.; Chen, X.;
Jiaang, W. T. Glutamic acid analogues as potent dipeptidyl peptidase
IV and 8 inhibitors. Bioorg. Med. Chem. Lett. 2005, 15, 3271-3275.
(7) (a) Belyaev, A.; Zhang, X. M.; Augustyns, K.; Lambeir, A. M.; De
Meester, I.; Vedernikova, I.; Scharpe´, S.; Haemers, A. Structure-
activity relationship of diaryl phosphonate esters as potent irreversible
dipeptidyl peptidase IV inhibitors. J. Med. Chem. 1999, 42, 1041-
1052. (b) Senten, K.; Daniels, L.; Van der Veken, P.; De Meester,
I.; Lambeir, A. M.; Scharpe´, S.; Haemers, A.; Augustyns, K. Rapid
parallel synthesis of dipeptide diphenyl phosphonate esters as
inhibitors of dipeptidyl peptidases. J. Comb. Chem. 2003, 5, 336-
344.
(8) (a) De Meester, I.; Belyaev, A.; Lambeir, A. M.; De Meyer, G. R.
Y.; Van Osselaer, N.; Haemers, A.; Scharpe´, S. In vivo inhibition of
dipeptidyl-peptidase IV by Pro-Pro diphenyl phosphonate (prodipine).
Biochem. Pharmacol. 1997, 54, 173-179. (b) Jung, F. J.; Yang, L.;
De Meester, I.; Augustyns, K.; Cardell, M.; Hillinger, S.; Vogt, P.;
Lardinois, D.; Scharpe´, S.; Weder, W.; Korom, S. CD 26/DPP IV-
targeted therapy of acute lung rejection in rats. J. Heart Lung Transpl.
2006, 25, 1109-1116. (c) Zhai, W.; Cardell, M.; De Meester, I.;
Augustyns, K.; Hillinger, S.; Arni, S.; Jungraithmayr, W.; Scharpe´,
S.; Weder, W.; Korom, S. Intragraft DPP IV inhibition attenuates
post-transplant ischemia/reperfusion injury after prolonged ischemia.
J. Heart Lung Transpl. 2007, 26, 174-180.
Figure 4. Kinetic analysis of compound 2e inhibition of DPP8. The
observed pseudo-first-order rate constants (kobs (s-1)) exhibit a hyper-
bolic relationship with the inhibitor concentration. The dissociation
constant Kd was determined to be 0.77 µM, and the kinact was 0.0029
( 0.0004 s-1
.
enzyme-inhibitor complex as well as the first-order rate
constant for the irreversible inactivation of the enzyme.
Figure 4 shows a typical hyperbolic relation between the
observed first-order rate constant and the inhibitor concentration
(as exemplified with data for compound 2e).
In conclusion, we developed a series of irreversible DPP8
inhibitors. Potent inhibition of DPP8 could be obtained with
bis(4-acetamidophenyl) pyrrolidin-2-yl and isoindolin-1-yl phos-
phonate derivatives 7a and 2e, both containing a dibasic P2
lysine residue. Compound 2e proved to combine a good affinity
(Kd) and pronounced selectivity for DPP8 with regard to DPP
V and DPP II. This molecule promises to be a useful tool in
the study of the non-DPP II/non-DPP IV members of the
proline-selective dipeptidyl peptidases (DPP8 and DPP9). The
compounds’ inhibitory potential with respect to the latter enzyme
will be determined.10
(9) Van der Veken, P.; El Sayed, I.; Joossens, J.; Stevens, C. V.;
Augustyns, K.; Haemers, A. Lewis acid catalysed synthesis of
N-protected diphenyl 1-aminoalkylphosphonates. Synthesis 2005,
634-638.
(10) Efforts to establish the identity of an enzyme we recently purified
from bovine testes with DPP9 are currently underway; Dubois, V.;
Lambeir, A. M.; Van der Veken, P.; Augustyns, K.; Creemers, J.;
Chen, X.; Scharpe´, S.; De Meester, I. Purification and characterisation
of DPP IV-like enzymes from bovine testes: evidence for identifica-
tion as natural DPP9-like enzyme. Front. Biosci. Accepted.
Acknowledgment. Pieter Van der Veken, Inger Brandt, and
Marie-Berthe Maes are indebted to the Flemish Fund for
Scientific Research (FWO-Vlaanderen) for financial support.
The Flemish Fund for Scientific Research is also acknowledged
for support to this project (Grant Number 0.410.05). The authors
thank Nicole Lamoen and Willy Bollaert for their excellent
technical assistance.
JM701005A