Advanced Pharmacology-I(PHR5001)

Lecture 12: Anti-ulcer Agents

(H2 receptor antagonists)

Dr. M G AzamAsstt. Professor

Dept. of Pharmacy, NSU1

IntroductionDefinition: Anti-ulcer (Antisecretory) agents are the drugs which decreases the secretion of gastric acid in the stomach. Histamine - Powerful stimulant of Hydrochloric acid

secretion from parietal (oxyntic)cells of gastric mucosa. Histamine (In larger dose )- increase secretion of pepsin.

These actions are mediated by H2-receptor. The secretion of acid by these parietal cells are regulated by

various mediators or receptors.a) Histamine H2 receptorsb) G receptors-release gastrin & release gastric acidc) Acetylcholine (Ach) M3 receptor

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What is Peptic Ulcer ?• A peptic ulcer disease or PUD is an ulcer (defined as mucosal

erosions ≥ 0.5 cm) of an area of the gastrointestinal tract exposed to the acid and pepsin secretion

• Gastritis is the precursor to PUD and it is clinically difficult to differentiate the two– Stomach (called gastric ulcer): Relieved by food but pain may

persist even after eating. Infrequent or absent remissions (>55 yrs)

– Duodenum (called duodenal ulcer): burning upper abdomen pain relieved by food but reappears 1-3 hrs after meals. Worse pain when stomach empty

– Esophagus (called Esophageal ulcer)

Causes of ulcers: Hypersecretion of acid and pepsin and GI infection of gm-ve bacteria H.

Pylori. Stress, alcohol, ulcerogenic drugs (e.g. NSAIDs), male genders, age, & diet

Why Ulceration Occurs? • High [H+] in the gastric lumen• Require defense mechanisms to protect oesophagus and stomach

– Mucus secretion: slows ion diffusion– Prostaglandins: I2 and E2 (alcohol, aspirin )– Bicabonate ions– High Blood Flow (nitric oxide)

• Imbalance primarily between Aggressive factors & Defensive factors:

Aggressive

factors, e,g,

acid, pepsin,

bile etc.

Defensive

factors, e.g.

mucus, HCO3,

PG

Gastroesophageal Reflux Disease (GERD)• Common and GI motility disorder• Acidity of Gastric contents – most common factor• Acid contents reflux back into esophagus• Intense burning, sometimes belching• Can lead to esophagitis, and esophageal ulcers

– Barrett’s esophagus (A complication of severe chronic GERD involving changes in the cells of the tissue that line the bottom of the esophagus. These esophageal cells become irritated when the contents of the stomach back up (refluxes) and there is a small but definite increased risk of adenocarcinoma of the esophagus.)

• Commonly associated with obesity• Improves with lifestyle management

Physiology of Acid Secretion

Schematic diagram the physiologic control of hydrogen ion secretion by the gastric parietal cell. ECL (enterochromaffin-like) cell; G(CCK-B), gastrin-cholecystokinin-B receptor; H, histamin; M1, M3, muscarinic receptors; ST2, somatostatin-2 receptor; ATPase, H+/K+ ATPase proton pump.

• Histamine uses the CAMP pathway whereas gastrin and Ach uses Ca2+ dependent pathway. Both pathway activates the H+/K+ ATPase. Ach release from postganglionic vagal fibres can stimulate directly gastric acid secretion though M3 (a specific muscarnic cholinergic receptor subtype).

Ach. also indirectly affects the parietal cells though the stimulation of histamine release from the entrochromaffin-like cells in the fundus and the stimulation of gastric release from G cells in the gastric antrum.

• Histamine is released from ECL cells through the multifactorial pathway and is critical regulator of acid production through the H2 sub type of receptor.

• The release of gastric is regulated through the multifactorial pathway and it stimulates acid secretion predominantly in an indirect manner by causing the release of histamine from ECL cells; a direct effect of gastric or parietal cell is also seen but is less important.

Mechanism

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ComparisonH1 receptor antagonist H2 receptor antagonist

Two aryl, heteroaryl rings in place of imidazole (bulky groups)

Connecting chain of aryl and terminal nitrogen is of 2 to 3 atoms

Ionic flexible chain at end. Ionized at pH 7.4

Hydrophobic (due to aryl rings). High partition coefficient value

Low dipole moment

Imidazole or other 5 membered heterocyclic ring

(Imidazole is not necessary) Connecting chain of ring and

terminal nitrogen is of 4 atoms Polar π e containing systems,

amphoteric, unionized at pH 7.4 Hydrophobic (due to polar grp).

Low partition coefficient High dipole moment

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Treatment of ulcerH2 receptor antagonists are the most popular

drug for the treatment of peptic ulcer. H2 receptor antagonists Must bind but not activate H2 receptor site

They bring about sympathomimetic relief and promote ulcer healing.

Gastric acid secretion involve H2 receptor which is competitively blocked by H2 blockers.

In 1972, Black and co-workers first described selective H2 receptor blokade for acid secretion. With successful introduction of cimetidine, in 1977 other analogs like ranitidine, famotidine & roxatidine has been synthesized.

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Historical developmentNHN

C C NH2

Histamine

H H

H H

NHN

H2C

H2C NH C

NH

NH2

Guanidine

Guanylhistamine

weak antagonist &lead molecule

NHN

H2C

H2C

H2C

HN C

NH

NH2

Extension of side chain

SK & F 91581

NHN

H2C

H2C

H2C CH2

Extension of side chain

NH C NH

SThiourea(Neutral)

Burimamide(No agonist but Orally not absorbed)

CH3

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NHN

H2C S

H2C CH2 NH C NH

N Cimetidine

Insertion of e- withdrawing grp.

to decrease basicity & increase potency

CN

CH3

Imidazole ring replaced by various Nitrogen containing heterocyclic ring to optimize the activity. Furan (Ranitidine) & thiazole (Nizatidine) showed good activity.Substitution in side chain with different basic as well as neutral groups were carried out like nitroamine ketene (Ranitidine) & sulfonylamidine (Famotidine) for better activity.Substitution of dimethyl amino methyl (Ranitidine & Nizatidine) and amidine (Famotidine) groups on ring showed better activity.

NHN

H2C S

H2C CH2

Insertion of sulfurin side chain

NH C NH

S

Metiamide(kidney damage& granulocytopeniadue to thiourea)

4-methyl grpsubst. in ring

Increase bindingreduce pKa of ring 'N'Increase permeability

CH3

NHN

H2C S

H2C CH2 NH C NH

NH less potent compound

thiourea replacedby guanidine

CH3

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Ranitidine

NO2

ON S N

HNH

Furan ring

Nitro amine ketene

NH2

S

SN

NH2

H2N

NN

SO

NH2O

famotidine

Amidine grp

Sulfonylamidine grp

Thiazole

NO2

S

NNS

HN

HN

Thiazole

Nizatidine

O

SNH

OOO

N

N

Lafutidine

Roxatidine

-specific & competitive H2-antagonist-metabolite of roxatidine acetate-Dose: 150mg

O NH

O

OHN

O

O

NH

NO

NH

S

NHN

Oxmetidine

benzo-1,3-dioxole

H2 receptor antagonists

Classification of Anti-ulcer DrugsI. Gastric Antisecretory Drugs

HCl secretion1. Antagonize Rs. on Parietal CellParietal Cell--- H2,M3, G2. Inhibitor of HH++-Pump-Pump

II. Antacids--- Neutralizations of secreted acid and pepsin activity

III. Agents kill HP : HP : Eradication of Helicobacter pylori by triple therapy: Almost all duodenal and 2/3 gastric ulcer pt’s infected with HP

Omeprazole / Lansoprazole - 20 / 30 mg bd

Clarithromycin - 500 mg bd

Amoxycillin / Metronidazole - 1gm / 500 mg bd

Given for 14 days

ProglumideAChPGE2 Histamine Gastrin

Adenyl cyclase

_+

ATP cAMP

Protein Kinase (Activated)

Ca++

+

Ca++

Proton pump

KK+ H+

Gastric acid

Parietal cellLumen of stomach

AntacidOmeprazole

Ranitidine

H2M3

Misoprostol

_

__

_

+

PGE receptor

+

+

Gastrin receptor+

+

+Compete Gastrin-R

HCl & pepsin ProbanthinePirenzepine

Classification of Anti-ulcer Drugs

1. Reduction in Gastric acid secretion:• H2 antihistamines: Cimetidine, Ranitidine, Famotidine, Nizatidine

and Roxatidine• Proton pump inhibitors: Omeprazole, Lansoprazole

Pantoprazole, Rabeprazole and Esomeprazole• Anticholinergics: Pirenzepine, Propantheline and Oxyphenonium• Prostaglandin analogue: Misoprostol

3. Ulcer protectives: Sucralfate, Colloidal Bismuth sudcitrate4. Anti-H. pylori Drugs: Amoxicillin, Clarithromycin,

metronidazole, tinidazole and tetracycline

2. Acid Neutralizing agents: (ANTACIDS)• Systemic: Sodium Bicarbonate and Sod. Citrate• Nonsystemic: Magnesium hydroxide, Mag. Treisilicate, Aluminium

hydroxide gel, Magaldrate and calcium carbonate

H2 Antagonists• Cimetidine, Ranitidine, Famotidine, Roxatidine, Nizatidine• MOA:

– Reversible competitive inhibitors of H2 receptor– Highly selective, no action on H1 or H3 receptors– All phases of gastric acid secretion– Very effective in inhibiting nocturnal acid secretion (as it

depends largely on Histamine )– Modest impact on meal stimulated acid secretion (as it depends

on gastrin, acetylcholine and histamine)– Volume of pepsin content and IF are also reduced– Volume reduced by 60 – 70% - anti ulcerogenic effect

Adverse Effect:Gynecomastia, prolactin , CYP450 , headache

Proton Pump Inhibitors• Most effective drugs in antiulcer therapy• Prodrugs requiring activation in acidic pH.• Block enzymes responsible for secreting HCl - binds

irreversibly to H+K+ATPase • Prototype: Omeprazole

Omeprazole

• Substituted Benzimidazole derivative • Diffuses into G. canaliculi = accumulationpH < 5 (proton catalyzed )= tetracyclic sulfenamide + sulphenic acid• Covalent binding with sulfhydryl cysteines of H⁺K⁺ ATPase• Irreversible inactivation of the pump molecule• Acid suppressants regardless of stimulating factors• Also inhibits gastric mucosal carbonic anhydrase

CLINICAL USES OF AGENTS AFFECTING GASTRIC ACIDITY:

1. H2 –histamine receptor antagonist eg. Ranitidine,Cimetidine,Famotidine, Nizatidine

- Peptic ulcer (Gastric ulcer and duodenal ulcer) - Reflex oesophagitis 2. Proton Pump Inhibitor : Omeprazole Lansoprazole, Esomeprazole,Rabeprazole,Pantoprazole - Peptic ulcer - Reflex oesophagitis - As one component Of therapy for H-pylori infection - Zollinger-Ellison syndrome

3. Antacid eg. Mg-trisillicate, Al-hydroxide alginates - Dyspepsia - Symptomatic relief of peptic ulcer - Esophageal reflex4. Bismuth chelate -One component of therapy for H-pylori infection.

Screening methods:-1. Pyloric ligation induced gastric ulceration

2. Ethanol induced mucosal damage in rats

3. Stress ulcer through immobilization stress Lower & upper extremities fixed, wrapped in a wire gaze, kept at dark for 24 hr.

4. Indomethacin induced ulcers in rats After 10 min of admistering 20 mg/kg indomethacin, test drug is

given orally. 6hr later , sacrifice in co2 , stomach is removed.

Pyloric ligation induced gastric ulceration• After 1h of treatment with test or std. (Ranitidine 50 mg/kg) ,

animals are anaesthetized with the help of anesthetic ether; the abdomen is opened by a small midline incision. Pyloric portion of the stomach is slightly lifted out and ligated.

• The stomach is replaced carefully and the abdominal wall is closed by interrupted sutures. Rats are sacrificed by an over dose of anaesthetic ether after four hours of pyloric ligation.

• Abdomen is opened and ligature is placed around esophagus. Stomach is removed and contents are drained to centrifuge tube.

• The volume of the gastric juice is measured and centrifuged at 2000 rpm for 10 min. From the supernatant, aliquots (1 ml of each) are taken for the determination of pH, total and free acidity. Each stomach is examined (by a 10Χ magnifier lens) for lesions in the fore stomach portion & indexed according to severity.

% Inhibition of Ulceration =

0 - no ulcer, 1 - spot ulcer, 2 - deep ulcer, 3 - perforation

Ethanol induced ulcer modelThe ulcer is induced by administering ethanol to Albino rats

(fasted for 36 h). One group represented the control group, which received

ethanol Second & Third• Groups received methanolic extract of A. Indicum 250 and• 500 mg/kg and, Ranitidine, in the dose of 50 mg/kg were• administered orally for Four group as reference standard• drug. The gastric ulcers were induced in rats by• administrating absolute ethanol (90%) (0.5 ml/100g) orally,• after 45 min of methanolic extract and ranitidine treatment.• They were kept in specially constructed cages to prevent• coprophagia during and after the experiment. The animals• were anaesthetized 1h latter with anaesthetic ether and• stomach was incised along the greater curvature and• ulceration will be scored

Dr. M G AzamAsstt. Professor

Dept. of Pharmacy, NSU23

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Antibody is a class of immunological proteins (globular glycoproteins/ “Immunoglobulin” produced by plasma cells (B lymphocytes) in response to a specific antigen.

Antibody is a class of immunological proteins (globular glycoproteins/ “Immunoglobulin” produced by plasma cells (B lymphocytes) in response to a specific antigen.

• Antibodies function as markers, binding to the antigen so that the antigen molecules can be recognized and destroyed by phagocytes.

• The part of the antigen that the antibody binds to is called the epitope. The epitope is thus a short amino acid sequence that the antibody is able to recognize.

• Antibodies are useful tools for the immune system to combat pathogens.

Polyclonal Abs• Mixture of antibodies similar to sera Antibodies that are obtained from different B cell resources.

They are a combination of immunoglobulin molecules secreted against a specific antigen, each identifying a different epitope.

Biomedical research Diagnosis of diseases Treatment of diseases such as infection, cancer, etc.

* Used in

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Structure of antibody

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Consist of four polypeptide chains, somewhat resembling to a Y shape. i) 2 identical Light chains ( L chains) ii) 2 identical Heavy chains ( H chains)

Disulphide bonds hold 4 polypeptide chains

Ig domain: 110 amino acids; globular domain used in many proteins.Variable domains,Constant domains, Hinge.Fab: fragment antigen bindingFc: fragment crystallizable (effector functions)

CDR: Complementarity-Determining Regions

Immunoglobulin (Ig) Class

Heavy Chain

Diagram Distribution

IgA αor

External Secretions

IgD δB Cell surface

receptor

IgE εCells that secrete

histamines

IgGγ

Main antibody in serum

Most Stable

IgM μ

First antibody secreted in

development

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• A mouse is immunized with the specific antigen“X”. This will allow the mouse to produce antibodies.

• The spleen cells producing specific antibodies are removed.• Fusion of spleen cells with tumor cells (myeloma cells) in a culture medium is done.

The resulting cell is called a hybridoma.• Hybridoma cells are continuously growing cell line. Each hybridoma will produce

relatively large quantities of monoclonal antibody molecules. These hybridomas have antibody producing capability from lymphocytes & have ability to grow continuously (immortal) from malignant cells.

• Separate the hybridomas from unfused cells.• Culture selected hybridoma cells for the production of monoclonal antibodies in large

quantity. • Remain indefinitely and maintained by in vitro; that is, in culture vessels or in vivo-in

the body of an animal so that antibodies will be produced in body and can be recovered later from body fluid.

Production of Monoclonal Antibodies

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Purification of monoclonal antibody :

Affinity chromatography is used for purification of mAbs. This method separates molecules on their differing affinities for a ligand.

Antigen can be bound to the support

matrix (small, chemically reactive beads) in order to purify antigen-specific

antibody from a polyclonal antiserum.Specific antibody is eluted by altering the pH, which disrupt antibody-antigen bonds.

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Application of monoclonal antibody :

1. Research tools:

2. Clinical diagnosis:

3. Treatment of diseases:

Identification of cell surface marker Purification of proteins and enzymes Detection assays Genetic engineering Structure of cell membrane

Cancers Leukemia Viral disease Allergic reaction AIDS

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Sufix of Antibody

• Diagnostic testsMonoclonal antibodies are widely used as

diagnostic and research reagents. • Once monoclonal antibodies for a given substance

have been produced, they can be used to detect the presence of this substance. The Western blot test detect the protein on a membrane.

• Monoclonal antibodies can also be used to purify a substance with techniques called affinity chromatography.

• Other monoclonal antibodies allow rapid diagnosis of hepatitis, influenza, herpes and streptococcal infections.

• Antibodies are used in several diagnostic tests to detect small amounts of drugs, toxins or hormones.

• Monoclonal Antibodies also used in ELISA (enzyme linked immunosorbant Assay) for detection of viruses.

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Monoclonal antibody in cancer therapy:

A. Naked mAbs: Without any drugs, radioactive materials or toxins.B. Conjugated mAbs: They are combined with chemotherapy drugs (Chemolabeled Ab) or radioactive material or toxins

(Immunotoxin)

• By means of genetic engineering, so-called chimeric antibodies are prepared that contain both human and mouse sections.

• The mouse section is located in the variable region of the antibody that contains the antigen- binding site, while the human section is in the constant region.

• After the variable region binds to the cancer antigen, the human section activates complement and cytotoxic T cells to destroy the cancer cell.

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When a monoclonal antibody attaches to a cancer cell, it can:

• Make the cancer cell more visible to the immune system

• Block growth signals

• Deliver radiation to cancer cells.

• Slip powerful drugs into cancer cells

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mAbs as antitumor agentsTrastuzumabTrastuzumab

HER2 (Human Epidermal Growth Factor Receptor 2) also known as CD340 (cluster of differentiation340) /p185 is a protein that in humans is encoded by the ERBB2 gene. Amplification or over-expression of this gene has been shown to play an important role in the pathogenesis and progression of certain aggressive types of breast cancer

Trastuzumab is effective only in cancers where HER2 is over-expressed. After binding to HER2, the Mab causes an increase in p27, a protein that halts cell proliferation. Another MAb, Pertuzumab, which inhibits dimerization of HER2 and HER3 receptors, was approved by the FDA for use in combination with trastuzumab in June 2012

AntibodyAntibody Target Target Treatment(s)Treatment(s) Approved Approved inin

TrastuzumabTrastuzumab HER2HER2 Breast cancer & lymphomasBreast cancer & lymphomas 19981998

Rituximab Rituximab CD20CD20 B cell lymphomasB cell lymphomas 19971997

Tositumomab Tositumomab CD20CD20 B cell lymphomasB cell lymphomas 20032003

AlemtuzumabAlemtuzumab CD52CD52 Chronic lymphocytic Chronic lymphocytic leukemialeukemia

20012001

Bevacizumab Bevacizumab Vascular Vascular endothelial endothelial growth factorgrowth factor

Colorectal cancerColorectal cancer 20042004

Cetuximab Cetuximab Epidermal Epidermal growth factor growth factor receptorreceptor

Colorectal cancerColorectal cancer 20042004

Panitumumab Panitumumab EGF receptorEGF receptor Colorectal cancerColorectal cancer

Gemtuzumab Gemtuzumab CD33CD33 Acute myelogenous Acute myelogenous leukemialeukemia

20002000

Ibritumomab Ibritumomab CD20CD20 Non Hodgkin lymphomaNon Hodgkin lymphoma 20022002

Antitumor therapyAntitumor therapy

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RituximabPharmacology and M/A• Rituxan is the first MAb approved in 1997 by FDA. Chimeric MAb that binds to

the antigen CD20 found on B-lymphocytes (normal B cell and most malignant B cells).

• CD20 is involved in cell cycle initiation, regulation and differentiation by activation of B-cells from the G0 to G1 phase. It also operates as a calcium channel.

• Used in low, intermediate and high grade lymphomas/Non-Hodgkin Lymphoma (NHL) (in combination with cyclophosphamide, doxorubicin, vincristine and prednisone).

• It can also be given with radioimmunotherapy given directly to the tumor site

• M/A: Rituximab causes CD20-positive cell death by antibody dependent cell-mediated cytotoxicity, direct effects via CD20 ligation, and complement-mediated lysis.

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AntibodyAntibody Target Target Treatment(s)Treatment(s)Supportive CareSupportive Care

OTK3 (muromonab-CD3)OTK3 (muromonab-CD3) T-cell CD3 receptorT-cell CD3 receptor Transplant RejectionTransplant Rejection

DaclizumabDaclizumab IL2 ReceptorIL2 Receptor Transplant RejectionTransplant Rejection

Infliximab Infliximab TNF TNF αα Rheumatoid arthritisRheumatoid arthritis

Omalizumab Omalizumab Ig E Ig E Allergy related AsthmaAllergy related Asthma

Natalizumab Natalizumab T cell VLA4 receptorT cell VLA4 receptor Multiple sclerosis therapyMultiple sclerosis therapy

Adalimumab Adalimumab TNF TNF αα Inflammatory diseasesInflammatory diseases

Basiliximab Basiliximab IL2 ReceptorIL2 Receptor Transplant RejectionTransplant Rejection

Therapeutic agents:

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Tissue transplantation

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CD3 (cluster of differentiation 3) T-cell co-receptor is a protein complex and is composed of four distinct chains. These chains associate with a molecule known as the T-cell receptor (TCR)

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Rheumatoid Arthritis is a chronic, autoimmune disease characterized by Severe joint inflammation, Increased synovial fluid , and thickened synovial membrane. It occurs due to destruction of bone and cartilage in several joints & Elevated levels of pro-inflammatory cytokines TNF(Tumor necrosis factor)-α, IL-1, IL-6

Efalizumab

• Psoriasis is a disease of the immune system that involves T lymphocytes.

• It results from complex communications that cause activation of T lymphocytes and trafficking to the skin.

• Further reactivation causes inflammation and overproduction of skin, resulting in lesions and plaques 42

Efalizumab is a humanized IgG1 antibody produced by recombinant DNA technology. It exhibits immunosuppressive function. It binds to CD11a, which is the α-subunit of leukocyte function antigen (LFA)-1. Efalizumab decreases the cell surface expression of CD11a, which is expressed on all leukocytes.

Efalizumab is a humanized IgG1 antibody produced by recombinant DNA technology. It exhibits immunosuppressive function. It binds to CD11a, which is the α-subunit of leukocyte function antigen (LFA)-1. Efalizumab decreases the cell surface expression of CD11a, which is expressed on all leukocytes.

In the treatment of diabetes, novel and improved therapeutic modalities for those individuals with impaired insulin secretory function would be helpful. Currently, long-acting basal insulins are given daily, or more often, and are associated with both hypoglycemia and weight gain.

Therefore, a highly specific, ultra-long-acting activator of INSR, such as a monoclonal antibody, would represent a new paradigm in diabetes therapy.

XMetA, an allosteric activator of INSR both in vitro and in vivo, has the potential to normalize glycemic control in a model of insulinopenic diabetes without causing hypoglycemia or promoting weight gain.

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• Of the more than 20 monoclonal antibodies generated to combat infectious diseases that are in clinical development in 2011, most are in phase 1 or 2 and are directed against either viruses or bacterial toxins.

Thank

ou

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