Structure-Activity Relationships of Sulfonimide and Ester-Based Inhibitors of

Plasminogen Activator Inhibitor-1

Karen L. Sanders,1 Nadine C. El-Ayache,1 Gregory A. Abernathy,1 Jacinda M. Lisi,1 Melinda S. Myers,1 Paul R. North,1 Shih-Hon Li,2 Mark Warnock,2 Daniel A. Lawrence,2 Cory D. Emal1*; (1) Chemistry Department, Eastern Michigan University, Ypsilanti, 48197; (2) Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI, 48109

Introduction/BackgroundDesign and Synthesis

The inhibition of plasminogen activator-inhibitor-1 (PAI-1) isanticipated to increase our understanding of various human

ailments including diabetes, stroke, and atherosclerosis, withwhich high levels of PAI-1 have been associated.(1) PAI-1prevents certain serine proteases from cleaving peptide bondsand thus is able to regulate various cellular processes such ascontrolling the levels of other intracellular proteins, such as tissue-

type plasminogen activator (tPA) and urokinase-type plasminogenactivator (uPA).(2)

Scheme 2: Synthesis of ester-based PAI-1 inhibitors (4)

Effect of modifying the gallate to a protocatechuate on PAI-1 inhibition:

IC50 = 4.69 µM

Effect on PAI-1 inhibition of manipulating the carbamate side chain:

IC50 = 4.75 µM IC50 = 0.159 µM

IC50:

X = OH: 0.104 µMX = H: (ND)

IC50:

X = OH: 0.062 µMX = H: 0.048 µM

IC50:

X = OH: 0.105 µMX = H: 0.029 µM

Scheme 1: Synthesis of sulfonimide-based PAI-1 inhibitors (5)

O

O

OH

O

OO

O

OHO BnO BnO

OBn

HN

OO

O

OBnO

OBn

H3N

OO

1) BnCl, KI, K2CO3

(CH3)2CO, reflux

2) NaOH,C2H5OH

reflux, 18 hr

EDCįHCl, DMAP

CH2Cl2

TFA

CH2Cl2

triphosgene

NaHCO3 (aq.)

CH2Cl2 pyridine

R-OH

BocHN

OH

OH

CF3CO2

RO Cl

O

O O

SNHR

O O

MeO

MeO SN

O O

HO

HO

R

S

O O

OH

OH

SN

O O

MeO

MeO

R

S

O O

OMe

OMe

ArSO2Cl BBr3

NaH

HOO

O

O

OH

OH

HOO

O

O

OH

OH

OO

O

HO

OH

OH

• Route of developing our PAI-1 inhibitor scaffolds• high-throughput screen of MicroSource SPECTRUM library.(4)

• hypothesized effective inhibitors based on tannic acid as a lead compound (IC50 = 6.6 nM)• gallate/digallate-containing species

• synthesized a number of novel small-molecule inhibitors (4,5)

Future Directions• Test the inhibitory effect of electronically larger species attachments.• Synthesize inhibitors with nonsymmetrical gallate attachments.

• Incorporate different functionality into linking unit.

References

AcknowledgementsFunding from: - National Institutes of Health (HL089407); Camille and Henry Dreyfus Foundation; Eastern

Michigan University

1) Ren, Y.; Himmeldirk, K.; Chen, X. J. Med. Chem. 2006, 49, 2829-2837.

2) Wang, Z.; Mottonen, J.; Goldsmith, E. J. Biochemistry. 1996, 35 (51), 16443-16448. 3) Miyazaki, H.; Ogiku, T.; Hiroshi, S.; Moritani, Y.; Ohtanl, A. Chem. Pharm. Bull. 2009, 57 (9), 979-985.

4) Cale, J. M.; Li, S.; Warnock, M.; Su, E. J.; North, P. R.; Sanders, K. L.; Puscau, M. M.; Emal, C. D.; Lawrence, D. A. J. Biol. Chem. 2010, in press.5) El-Ayache, N. C.; Li, S. H.; Warnock, M.; Lawrence, D. A.; Emal, C. D. Bioorg. Med. Chem. Lett. 2010, 966-970.

Fibrinolysis:

Green arrows = stimulatory effectRed arrows = inhibitory effect

The goal of this research is to develop small-molecule inhibitors of PAI-1, a major natural inhibitor of fibrinolysis, the process which leads

to the breakdown of blood clots. The structural complexity of the protein allows for multiple potential binding sites. Several classes of PAI-1 inhibitors have been reported, recently including a furan-2-one and pyrrolin-2-one series. (3)

Conclusions

PAI-1 Inhibition Assay Results

IC50 = 0.02 µM IC50 = 0.022 µMIC50 = 0.027 µM

• Sulfonamide inhibitors have an optimum methylene unit length of 6.5 which correlates to a chain length of approximately 780-1000 pm.

• The protocatechuate modification tends to moderately improve PAI-1 inhibition compared to gallate moieties.

• Potency of ester-based inhibitors is enhanced by the side-chain:• Inhibition improves as size of carbamate handle increases.• Ring ending carbamate handles or linker handles composed of

approximately 7 methylene units give most potent inhibitors in series.• Drawbacks of these inhibitors:• low binding affinity for PAI-1

• inability to inhibit PAI-1 in the presence of vitronectin

Effect of side-chain length on PAI-1 inhibition:

R IC50 Values (µM)

H N.D.

(CH2)2CH3 2.670

(CH2)4CH3 0.240

(CH2)5CH3 0.284

(CH2)7CH3 0.086

(CH2)9CH3 0.250

(CH2)11CH3 2.600

OO

O

OBnO

OBn

HN

OO

O

OBnO

OBn

N

C

O

OO

O

OHO

OH

R-OHtriethylamine

CH2Cl2

10% Pd¼/C

H2

CH2Cl2

O OR

HN

O OR

HOO

HN

O O

HOO

HN

O O

Cl

OO

O

HO

HO HN

O O

7