Omeprazole: The First Proton Pump Inhibitor

Behnaz Shafii

Michigan State University

January 23rd, 2008

Organic Seminar

Outline

- Gastric acid secretion

- Helicobacter pylori, common diseases and treatments

- Proton Pump Inhibitors (PPIs)

- Development of omeprazole

- Mechanism of action

- Pkas and their importance

- Anti-cancer activity and future

- Conclusion

Stomach Cells

Gastric glands

Blood vessels

Stomach

Parietal cell K+

H+

Cytoplasm

Lumen

Mucous cells

� ATP is consumed for ion exchange

http://www.britannica.com/eb/art-68634

Mechanism of Proton Transport

Lys

Asp

Glu

H3 O +

� ATP is consumed for ion exchange

NH3+

COO-

COO-

COO-

Asp

Glu

Glu

Lys

H3O+

NH3+

COO-COO-

COO-

H3O+

K+ K+

H3O+

Glu

Glu

Lys

Ala

ValK+

COO-

COO-

COO-NH3+

Asp

Shin et al, Dig Dis Sci., 51, 2006, 823-833

Lower esophageal

sphincter closed

Lower esophageal

sphincter open

allowing reflux

Esophagus

� Gastroesophageal Reflux Disease (GERD)

Lower esophageal

sphincter closed

Lower esophageal

sphincter open

allowing reflux

� Gastroesophageal Reflux Disease (GERD)

Common Disease Related to Inappropriate Levels of Gastric Acid

Gastric ulcer

� Peptic Ulcer Diseases

Duodenal ulcer

Discovery of Helicobacter pylori (Hp)

Nobel Prize in Medicine 2005- Robin Warren

- Barry Marshall

http://nobelprize.org/nobel_prizes/medicine/laureates/2005/press.html

H. pylori

(NH2)2CO+H2O CO2+2NH3

Urease

Chronic Peptic Ulcer and Cancer

Chronic Ulcer Cancerhttp://nobelprize.org/nobel_prizes/medicine/laureates/2005/press.html

� Once the bacteria have entered stomach, they cause inflammation and

may lead to different diseases.

http://www.worldgastroenterology.org

Helicobacter pylori Infection Globally in 2006

� Helicobacter pylori is common

and infects half of the world’s population.

0

20

40

60

80

100

Per

cen

tag

e

Mexico, Central

and South

America

Africa Asia United States

and Canada

Australia

� Triple therapy:A proton pump inhibitor + two antibiotics

Most Common Helicobacter pylori Treatments

� Quadruple therapyA proton pump inhibitor + two antibiotics + bismuth

Clarithromycin Amoxicillin Bismuth subsalicylate

Proton Pump Inhibition

Histamine

Histamine H2

receptor antagonists

Acetylcholine

Muscarinic antagonists

Gastrin

Parietal Cell Proton pump inhibitors

Olbe, L. et al. Nature reviews, drug discovery, 2003, 2, 132-139

K+

H+

++

Lumen

Cytoplasm

Development of Omeprazole

H 81/75

No effect in humans

H 116/18

Olbe. L, Proton Pump Inhibitors, 1999, 3-20

CMN 131

Severe liver toxicity

H 77/67

Starting point for the X-Y-Z general structures

Development of Omeprazole

H 124/26

Leading compound with

good anti-secretary effect

Olbe. L, Proton Pump Inhibitors, 1999, 3-20

NS

HN

N

H 83/69 (Timoprazole)

Potent anti-secretary but blocking

the uptake of iodine

H 149/94 (Picoprazole)Potent anti-secretary

without thyroid effect

Development of Omeprazole

+2.295-OCH33-CH3, 4-OCH3, 5-CH3

+1.825-OCH34-OCH3, 5-CH3

+1.235-OCH34-CH3, 5-CH3

+0.945-OCH33-CH3, 5-CH3

+0.765-OCH34-CH3

05-OCH3H

∆pKa (Pyridine)R2R1

NH

N

S

O

N

R2 4

5

6

7

3 4

5

6

R1

Olbe. L, Proton Pump Inhibitors, 1999, 3-20

Omeprazole

Development of Omeprazole

H 168/68 (Omeprazole)

Optimal compound

�Prilosec or Losec (Omeprzole) was the world best selling drug by year 2000

( Worldwide sales of US$6.1 b/year)

�Marketed by AstraZeneca

http://www.astrazeneca.com/

Marketed PPIs

Omeprazole Lansoprazole

Pantoprazole Rabeprazole

Tenatoprazole

Mechanism of Action

Shin et al, Cell. Mol. Life Sci., 64, 2007, 1-18

H3CO

NH

N

S

O

N

H3C OCH3

CH3

H

H3CO

NH

N

S

O

N

H3C OCH3

CH3

H

H-

-

-

Omeprazole

Mechanism of Action of Omeprazole

Shin et al, Cell. Mol. Life Sci., 64, 2007, 1-18

S

N

NH

CH3

OCH3

CH3

N

SH K -ATPase

H3CO

H3CO

NH

N

S

O

H

N

CH3

OCH3H3C

N

CH3

OCH3

CH3

N

NH

SOH

H3CO

S

N

N

CH3

OCH3

CH3

NH3CO-H2O

+H2O

Methylation on 6th position

Wallmark, B. et al, J. Med. Chem, 1986, 1327, 1329

� Prevention of formation of the spiro-intermediate by a 6-methyl group

� Strong steric interaction with the imidazole ring

(revealed by molecular modeling)

N

CH3

OCH3

H3C

H3C

SO

N

NH

H3CO

Half-lives of the inhibitory effect

on acid secretion in humans

28 Hours for omeprazole

46 Hours for pantoprazole

Half-lives of PPIs

� De novo synthesis of proton pump protein?

Protein pump half-life 54 Hours

glutathione

�Reduction of disulfide by an endogenous cellular reducing agents

such as glutathione

Shin, J. et al, Gastroenterology, 2002, 123, 1588-1597

Labeling and Inhibition of ATPase by Omeprazole

CPM 0 150 300 450 600 0 150 300 450 600

6

810

Sachs, G. et al, JBC, 1997, 272, 22438-22446

[3H] Omeprazole

10 min 45 min

47% inhibition 83% inhibition KDa

Cys 813

Cys 321

Cys 892

Why PPIs Have Different Half-lives?

Omeprazole

Pantoprazole

Shin, J. et al, Gastroenterology, 2002, 123, 1588-1597

7-8 loop3-4 loop

Cys 892 Cys 321

5-6 loop

Cys 813

Cys 822

28 Hour half-life

46 Hour half-life

34568

Membrane

Protonation:

� Pyridine

� Benzimidazole or imidazopyridine

Rate Limiting Step

Omeprazole

Tenatoprazole

pKa1

pKa2

-

Sachs, G. et al, JACS, 2004, 126, 7800-7811

� Biphasic rate

Co

nv

ers

ion

ra

te c

on

sta

nts

(m

in -1

)

0

0.1

0.2

0.3

0.4

0.5

0.6

-1 0 1 2 3 4 5 6 7 8

pH

Lanzoprazole

Tenatoprazole

Pantoprazole

Omeprazole

Conversion Rates of PPIs at Different pHs

Sachs, G. et al, JACS, 2004, 126, 7800-7811

Protonation of Benzimidazole and Pyridine

[Bz-Py.H+] = [Bz-Py] × 10 (pKa1-pH)

Bz-PyBz-PyH+

BzH+-Py

BzH+-PyH+

-

-

-

[Bz.H+-Py] = [Bz-Py] × 10 (pKa2-pH)

Sachs, G. et al, JACS, 2004, 126, 7800-7811

XH3CO

NH

N

S

O

N

H3C OCH3

CH3

H

XH3CO

NH

N

S

O

N

H3C OCH3

CH3

H

H

H

H

H

H

XH3CO

NH

N

S

O

H

N

CH3

OCH3H3C

XH3CO

NH

N

S

O

N

H3C OCH3

CH3

H

H

1 2

3 4

pKa Measurements by UV Spectroscopy

Sachs, G. et al, JACS, 2004, 126, 7800-7811

H3CO

NH

N

S

CH3

O

H3CO

N

N

S

CH3

O

CH3

F2HCO

NH

N

S

CH3

O

F2HCO

N

N

S

CH3

O

CH3

pKa Measurements by UV Spectroscopy

5 6

Sachs, G. et al, JACS, 2004, 126, 7800-7811

OmeprazolePantoprazole

Tenatoprazole

Rabeprazole

Lansoprazole

Conversion Rates of Methyl-PPIs at Different Medium pH

N1-methyl lanzoprazole

N1-methyl tenatoprazole

N1-methyl pantoprazole

N1-methyl omeprazole

Sachs, G. et al, JACS, 2004, 126, 7800-7811

0

0.02

0.04

0.06

0.08

-0.5 0 0.5 1 1.5 2

Co

nv

ers

ion

ra

te c

on

sta

nts

(m

in -1

)

pH pH

0

0.1

0.2

0.3

0.4

0.5

-1 0 1 2 3 4 5 6 7 8

Lanzoprazole

Tenatoprazole

Pantoprazole

Omeprazole

N1-methyl lanzoprazole

0

0.2

0.4

0.6

0.8

1

230 250 270 290 310 330 350

Ab

sorb

an

ce

Wavelength (nm)

UV Spectra of N1-methyl Lansoprazole in Different pHs

pH 5.60pH 5.02

pH 4.53

pH 1.00

pH 4.03

pH 3.83

pH 1.60

pH 3.24

pH 1.30

pH 2.53

pH 2.30pH 2.00

pH 0.60

pH 0.30

pH 0.00

pH-0.30

Sachs, G. et al, JACS, 2004, 126, 7800-7811

pKa1

pKa2

246 nm

291 nm

pKa Measurements

� N-methylation and protonated pyridinylmethyl sulfinyl effect

pKa2= 0.7

pKa= 1.35

pKa= 1.43

0.7-1.43 = -0.73pyridinylmethylsulfinyl effect

1.43-1.35 = +0.08methylation effect

Sachs, G. et al, JACS, 2004, 126, 7800-7811

Pkas of Proton Pump Inhibitors

pKa1 pKa2

4.06 0.25+_

3.83 0.15+_

3.83 0.24+_

4.53

4.04

0.79

0.62

0.11

0.62

-0.12Actual measurements

Calculated

Omeprazole

Lansoprazole

Pantoprazole

Rabeprazole

Tenatoprazole

Sachs, G. et al, JACS, 2004, 126, 7800-7811

Conversion Rates and Pkas of Proton Pump Inhibitors

pKa1 = 3.83

pKa2 = 0.62

pKa1 = 3.83

pKa2 = 0.11

Lansoprazole Pantoprazole

[Bz-Py.H+] = [Bz-Py] × 10

[Bz.H+-Py] = [Bz-Py] × 10

Sachs, G. et al, JACS, 2004, 126, 7800-7811

(pKa1-pH)

(pKa2-pH)

Bz Py Bz Py

Conversion Rates of Proton Pump Inhibitors

� Substituent effects on the nucleophilicity of pyridine at pKa1 (when pH > 4.0)

� Conversion rates of PPIs are strongly affected by the second protonation

at pKa2 (when pH < 4.0)

First protonation

Second protonation

Sachs, G. et al, JACS, 2004, 126, 7800-7811

Why PPIs Have Different Half-lives?

Omeprazole

Pantoprazole

Shin, J. et al, Gastroenterology, 2002, 123, 1588-1597

7-8 loop3-4 loop

Cys 892 Cys 321

5-6 loop

Cys 813

Cys 822

28 Hour half-life

46 Hour half-life

34568

Membrane

� Drug-drug interaction with other anti-acid drugs

� Since PPIs require stimulation of gastric acid secretion, they are

more effective when administered one hour before breakfast.

� Protecting PPIs from gastric acid prior to absorption (formulated

with an acid-resistant coating)

Acid Related Activity of PPIs

Gastric glands

Blood vessels

Stomach

Enzyme-Inhibitor Complex in Acidic Compartment of the Stomach Cell

S

N

NH

CH3

OCH3

CH3

N

S

H3CO

H+

K+

Parietal cell

Cytoplasm

Mucous cells

NH

NS N

CH3

H3COO

OCH3H3C

H

K+

H+

Lumen

Esophagus

Stomach

Duodenum

Small intestine(jejunum and ileum)

Acid resistance coat

pH < 2

pH 2-5

pH 6.5-7.5

Release of PPIs in the Body

N

H3C

OCH3CH3

S

O

NH

N

OCH3

What other things can PPIs do?

Inappropriate gastric acid levels could be inhibited by PPIs

but:

Other Properties of PPIs

Alternative Uses

Minoxidil tablet

(Hypertension treatment)

(Topical foam)

Inappropriate gastric acid levels could be inhibited by PPIs

but:

Other Properties of PPIs

What is more about PPIs?

Glucose Metabolism in Mammalian Cells

Gateby, R. et al. Nature reviews, Cancer, 2004, 4, 891-899

Effect of Medium Acidity on the Viability of Gastric Cancer Cells

0

20

40

60

80

100

pH 7.5 pH 6.5 pH 5.5

Non-cancer cells

Gastric cancer cells

AGS

SNU-1

SNU-16

SNU-601

MKN-28

MKN-45

KATO IIIRGM

Ce

ll v

iab

ility

(%

)

Yeo, M. et al, Cancer Sci. 2008, 99, 185-192

Reduction of Cell Viability by Treatment with Pantoprazole

0 20 40 60 80 100

AGS

KatoIII

MKN28

MKN45

SNU1

SNU601

Cell viability (%) after PPI treatment

Yeo, M. et al, Cancer Sci. 2008, 99, 185-192

� 0.8 mM PPI for 16 h

Pretreatment of Cancer Cells with Omeprazole

Fais, S. et al, JNCI. 2004, 96, 1702-1713

� Melanoma cell line (MelM6)

0

20

40

60

80

100

0 0.5 5 50%

of

de

ad

ce

lls

[cisplatin] µM

0

20

40

60

80

100

0 0.5 5 50

% o

f d

ea

d c

ell

s

[cisplatin] µM

cisplatinsalineomeprazole (pretreatment)

cisplatinsalineomeprazole + cisplatin

Pretreatment of Cancer Cells with Omeprazole

Fais, S. et al, JNCI. 2004, 96, 1702-1713

� Melanoma cell line (MelM6)

0

20

40

60

80

100

0 0.1 1 10 100 1000

% o

f d

ea

d c

ell

s

0

20

40

60

80

100

0 0.1 1 10 100

% o

f d

ea

d c

ell

s

[5FU] µg/ml [vinblastin] ng/ml

5FU saline

omeprazole (pretreatment)

vinblastinsaline

omeprazole (pretreatment)

Conclusion

� Different conversion rates of proton pump inhibitors in the body

is mostly due to the protonation of benzimidazole ring.

� Inappropriate levels of Gastric acid can be inhibited by proton

pump inhibitors.

�There might be chances of using current FDA approved drugs for

other possible treatments rather than those usages they currently

have.

� PPIs are potentially potent anti-cancer agents (treatment or

pretreatment of cancer cells) which their anti-cancer activity

definitely needs further work.

Clinical Gastroenterology and Hepatology, 2007, 5,5

Acknowledgment

- Dr. Kevin Walker

- Dr. Babak Borhan

- Dr. Daniel Jones

- Dr. Jetze Tepe

- Dr. Ahmad Shafii

- Mark, Washington, Danielle, Yemane, Irosha

Group members:

- Chrysoula, Ramin, Roozbeh,

- Atefeh, Afra, Maryam, Rafida, Behrouz

My friends: