Pharmacology of gastric acid secretion

Omeprazole as a pro-drug

(has to be converted to an active substance): weak base, accumulates in canaliculi (acidic areas, protonated), modifies pump and inhibits it. Omeprazole is weak base so accumulates in acidic environment where it's converted to sulfenic acid then sulphonamide then disulfide with ATPase to inhibit it

Prostaglandins:

A group of bioactive, hormone-like chemicals derived from fatty acids that have a wide variety of biological effects including roles in inflammation, platelet aggregation, vascular smooth muscle dilation and constriction, cell growth, protection of from acid in the stomach, and many more. Arachidonic acid metabolites, membrane permeant, rapidly metabolised, act on GPCRs, coupled to e.g. adenylyl cyclase or PLC. Synthesised from phospholipids. Steroid inhibit phospholipase A2 for anti-inflammatory effect. Aspirin inhibits COX via covalent inhibition (transacetylation) and competitive inhibition (salicylic acid) thus inhibiting protective action of prostaglandins so can cause ulcers

Mucosa

A mucus-secreting membrane

Intrinsic factor

A substance produced by the mucosa of the stomach and intestines that is essential for the absorption of vitamin B12. Also known as gastric intrinsic factor (GIF), is a glycoprotein produced by the parietal cells of the stomach.

Physiology of gastric acid secretion

Acid promotes pepsinogen (chief cells) conversion to active pepsin and reduces bacterial colonisation. Parietal cell acid lumen via unique (good drug target as won't affect other cells) H+/K+ ATPase (proton pump). Acid secretion stimulated by acetylcholine (vagus), gastrin (G cells antral mucosa), parietal cells have M1 and G(astrin) receptors but most acid secretion stimulated indirectly via H2 receptor from paracrine cell Nerve endings in the stomach secrete two stimulatory neurotransmitters: acetylcholine and gastrin-releasing peptide. Their action is both direct on parietal cells and mediated through the secretion of gastrin from G cells and histamine from enterochromaffine-like cells. Gastrin acts on parietal cells directly and indirectly too, by stimulating the release of histamine. The release of histamine is the most important positive regulation mechanism of the secretion of gastric acid in the stomach. Its release is stimulated by gastrin and acetylcholine and inhibited by somatostatin. Parietal cells secrete acid in response to three types of stimuli: 1. Histamine, stimulates H2 histamine receptors (most significant contribution). 2. Acetylcholine, from parasympathetic activity via the vagus nerve and enteric nervous system, stimulating M3 receptors. 3. Gastrin, stimulating CCK2 receptors (least significant contribution, but also causes histamine secretion by local ECL cells) Activation of histamine through H2 receptor causes increases intracellular cAMP level while Ach through M3 receptor and gastrin through CCK2 receptor increases intracellular calcium level. These receptors are present on basolateral side of membrane. Increased cAMP level results in increased protein kinase A. Protein kinase A phosphorylates proteins involved in the transport of H+/K+ ATPase from the cytoplasm to the cell membrane. This causes resorption of K+ ions and secretion of H+ ions. Gastrin primarily induces acid-secretion indirectly, increasing histamine synthesis in ECL cells, which in turn signal parietal cells via histamine release/H2 stimulation. Gastrin itself has no effect on the maximum histamine-stimulated gastric acid secretion. Ach stimulates acid secretion by secretion by vagus nerve and acting on M1/M3 Gq receptors. Also directly affects ECL cells and indirectly affects G cells (via interneuron which releases Gastrin releasing peptide). Parietal cells also have gastrin receptor (CCK2, Gq)

4 subtypes of histamine receptor:

All G protein-coupled H1 = IP3/Ca DAG/PKC e.g. smooth muscle contraction H2 = Gs/cAMP e.g. parietal cell HCL secretion H3/H4 = Gi/o inhibit VGCC e.g. neurones, presynaptic inhibition of transmitter release. Drugs have different equilibrium constants for different receptors (lower = higher affinity)

Aspirin

Also called acetylsalicylic acid

How does aspirin aid the metabolism of arachidonic acid?

- AA is metabolized by COX to prostanoids e.g. prostaglandins - Aspirin inhibits COX o Covalent inhibition via transacetylation o Competitive inhibition by salicylic acid - Prostanoids control mucous and HCO3- secretion in stomach Aspirin can exacerbate ulcers

Briefly describe the role of antihistamines, proton pump blockers and antibiotics in the treatment of gastric ulcer. Name a specific drug for each process.

- Antihistamines o Block H2 histamine receptor o This normally stimulates parietal cells to secrete acid o E.g. ranitidine and cimetidine which are selective antagonists o Treats symptoms and not cause - Proton pump blocker o Blocks H/K ATPase which secrete protons from parietal cells o E.g. esomeprazole (S enantiomer, prodrug, covalently inhibits pump) o Symptoms not cause - Antibiotics o Kill helicobacter pylori o Responsible for >95% of all ulcers o Cause not just symptoms E.g. amoxicillin

Mucosal strengtheners:

DENOL (bismuth chelate) - may kill helicobacter pylori, stimulates mucosal prostaglandins and/or bicarbonate secretion, side effects = darkens tongue, motions black, nausea and vomiting. SULCRALFATE - complex of Al(OH)3/sulphated sucrose, little antacid effect, protects mucosa from acid - pepsin attack, side effects = lower gastro-intestinal symptoms

Proton pump inhibitors:

Decrease the amount of acid produced by the stomach Omeprazole and lansoprazole = treatment of choice erosive oesophagitis, heals gastric and duodenal ulcers, blocks H+/K+ ATPase proton pump, duration of treatment: 4 weeks for duodenal ulcer, 8 weeks for gastric ulcer. Maintenance treatment now possible. Side effects (usually well tolerated) - possible drug interactions as a result of metabolism by same liver enzyme: Warfarin and Phenytoin effects are increased

Describe how and why the pro-drug omeprazole is converted into the active compound and how the active form then inhibits its target.

Forms a disulfide bond with the H+/K+ ATPase (proton pump to inhibit it) Omeprazole (prodrug) --> Activated omeprazole (sulfenic acid) --> sulfenamide - Lone pair on S results in tetrahedral geometry and thus chirality - Omeprazole is a weak base so accumulates in acidic environment - In acid environment (pH 1) compound is converted o > sulfuric > sulfenamide > disulfide with H/K ATPase to inhibit it Chemical mechanism: (SEE PDF) (S-O- and N-H+) 1. Nitrogen lone pair in ring attacks N= and electrons go to N+ 2. +/- proton move from N-H to S-O- 3. N lone pair attacks S and OH2+ leaves (arguably OH2) 4. SH on ATPase attacks S to form disulfide bond

Helicobacter pylori and pathology:

H. pylori secretes virulence factors (CagA) and ureas and makes NH4+ (epithelial cell toxin) Discovered in 1982 by Warren and Marshall. Associated with >95% ulcers. Secretes virulence factors and urease making epithelial cell toxin NH4+. Marshall infected himself and gave himself an ulcer. Carried by 50% of 40yr old adults but other factors also required for ulceration e.g. stress and alcohol Treatments to eradicate H pylori: standard triple therapy: broad spectrum antibiotic (amoxycillin), metronidazole (anti-protozoan/bacterial - used for abscess in dentist), bismuth chelate (mucosal strengthener) - all for 2 weeks

Ranitidine

H2 receptor antagonist Prevents insertion of pump into membrane

Paracetamol

Has good antipyretic and analgesic effects and inhibits prostaglandin synthesis in CNS but weak anti-inflammatory effects. However, is selective for COX-2 which is barely found in most cells (inducible) and thus has weak gastric/platelet side effects while aspirin is non-selective

H. pylori

Helicobacter pylori o >95% ulcers o Secretes virulence factors (CagA) and ureas and makes NH4+ (epithelial cell toxin) o Also inflammatory response can be harmful o Other factors required (e.g. stress) o Triple therapy for 2 weeks - Broad spectrum antibiotic (amoxicillin) - Metronidazole (anti-protozoan/bacterial) - Bismuth chelate (mucosal strengthener)

arachidonic acid

Metabolism aid by aspirin

Prostaglandin analogues:

Misoprostol. Actions = stable PGE1 analogue, inhibits acid secretion, but augments mucus and HCO3- secretion. Mucosal protective (+small antisecretory effect). Prevents NSAID - associated ulcers. Promotes healing of gastric and duodenal ulcers. Side effects = diarrhoea due to opening of Cl channel. They tend to be unstable and act on prostaglandin receptors on parietal cells that inhibit acid secretion. Could use alongside COX inhibitor to avoid side effects

gastric mucosa

Mucosa that lines the stomach The gastric mucosa is the mucous membrane layer of the stomach, which contains the glands and the gastric pits.

Proton pump blocker in the treatment of gastric ulcer?

Omeprazole

parietal cells of stomach

Secrete HCl and intrinsic factor Also known as oxyntic or delomorphous cells) are the epithelial cells that secrete hydrochloric acid (HCl) and intrinsic factor. These cells are located in the gastric glands found in the lining of the fundus and in the cardia of the stomach. They contain an extensive secretory network (called canaliculi) from which the HCl is secreted by active transport into the stomach. The enzyme hydrogen potassium ATPase (H+/K+ ATPase) is unique to the parietal cells and transports the H+ against a concentration gradient Parietal cells are primarily regulated via histamine, acetylcholine and gastrin signaling from both central and local modulators

Protective action of mucus/HCO3-:

Secreted from mucus cells/surface or pit cells. Stimulated by prostaglandins (lipid mediators)

Endocrine

The body's "slow" chemical communication system; a set of glands that secrete hormones into the bloodstream Secrete internally

gastric glands of stomach

The glands and gastric pits are located in the stomach lining. The glands themselves are in the lamina propria of the mucous membrane and they open into the bases of the gastric pits formed by the epithelium.[

NSAID (Aspirin)

non-steroid anti-inflammatory drug Inhibits COX2 but not COX1 Inhibition of cylcogenoxygenase 2 -decrease in prostoglandins -increase in GA secretion -decrease in blood flow

Acid secretion stimulated by

o ACh (vagus) - Parietal cells have M1 and/or M3 receptors (Gq) - NB. ACh also indirectly affects parietal cells via: · Direct effects on ECL cells · Indirect effects on G cells (via an interneuron which releases gastrin-releasing peptide) o Gastrin (G cells) - Parietal cells have G (CCK2) receptors (Gq) o Histamine - Released by paracrine cells e.g. ECL (enterochromafin-like) cells - Binds H2 receptor (Gs) Most important

Proton pump inhibitors

o Drug interactions often occur as a result of metabolism by same liver enzyme - This increases effective concentration - E.g. warfarin and phenytoin effects are increased o E.g. omeprazole and lansoprazole o Treatment of choice for erosive oesophagitis o Heal gastric/duodenal ulcers o Maintenance treatment possible o Omeprazole - Weak base so selectively accumulates in acidic environments - Irreversibly changes the structure of the proton pump so is an irreversible inhibitor

Chelates and complexes (mucosal strengtheners, heal ulcers)

o E.g. DENOL (bismuth chelate) and Sucralfate o Stimulate prostaglandins and/or bicarbonate o Denol may also kill h.pylori - Darkens tongue, motions, nausea and vomiting o Sulcralfate may also be antacid Lower GI side effects

H2 antagonists

o E.g. cimetidine and ranitidine o Heal gastric/duodenal ulcers o Treatment indefinite, does not modify natural history of disease o Cimetidine can cause gynaecomastia o Invented by Sir James Black (along with beta blockers) o Imidazole ring (in histamine and cimetidine) - Dissolves in water, basic o Thiazole ring (famotidine, nizatidine) o Relationship between affinity and intrinsic activity - Histamine has low affinity but very high efficacy - Cimetidine has high affinity but low efficacy · Antagonists on the whole tend to have higher affinity and by definition must have low efficacy Partial agonists e.g. n-guanylhistamine

Ultimately, parietal cell to get acid release

o Gs (histamine) - cAMP, PKA, phosphorylation of vesicle/cytoskeletal proteins causing drag of protons to membrane o Gq (probably gastrin and ACh) Calcium

Causes of ulcers

o Helicobacter pylori o Stress o Alcohol o NSAIDs o Zollinger-ellison syndrome (a rare tumour) - Ectopic tumours of G cells causing hypersecretion of gastrin and therefore acid Treated with omeprazole

Histamine summary

o Histadine > histamine o Histidine decarboxylase o B6 cofactor o Stored in vesicles o 4 subtypes of receptor, all GPCR - H1: smooth muscle contraction (Gq) - H2: almost uniquely on parietal cell (Gs) (e.g. cimetidine) - H3/4: neurons, presynaptic inhibition (Gi) - Make it a good selective target (we have agonists/antagonists for each class) · NB. Drugs are not entirely specific, they just have different equilibrium constants (KB) for different receptors o Lower = higher affinity

Selective anti-muscarinic agent

o More side effects o M1/3 receptors o E.g. pirenzepine (M1) o Heal gastric/duodenal ulcers o Do not cross BBB (no CNS effects) Dry mouth, blurred vision

Stomach cells

o Parietal (or oxyntic) cells secrete acid - Canaliculi on surface due to proton pump vesicles fusing with plasma membrane on stimulation - Need acid to activate pepsinogen (to pepsin) and reduce bacterial colonization · Stomach more acidic than lysosome - The proton pump is a H+/K+ ATPase (unique) · An enzyme with 2 subunits · Potassium retrieved from lumen in the process · It therefore must be actively transported out by cell - Cl- also pumped out - Ultimate source of H+ is CO2 in the blood · Often get alkaline tide in blood after meal - Also secret intrinsic factor required for vitamin B12 uptake (Pernicious anaemia) o Mucous neck cells produce mucous Glycosylated in Golgi

Mucous and HCO3- accumulates near cell surface

o Secreted from mucous cells o Stimulated by prostaglandins - Arachidonic acid metabolites (AA often in 2 position of phospholipid) · COX performs first step in the series of reactions o COX-1 and COX-2 (the latter is inducible, former is constitutive) o Inhibited by aspirin - NB. Failures of Vioxx (selective COX-2 inhibition) - Membrane permeant - Rapidly metabolized - Act on GPCRs (Gs or Gq) - Act autocrine (on self) or paracrine - Prostaglandin analogues as drugs · E.g. misoprostol (stable PGE1 analogue) o Inhibits acid secretion, augments mucous / bicarbonate secretion o Mucousal protective · Could use alongside COX inhibition to avoid side effects · Also promote ulcer healing · Open Cl- channel so cause diarrhea

Antacids

o Symptom relief and can assist healing o Aluminum (constipation) and magnesium (diarrhea) compounds o Sodium bicarbonate (belching) - NB. Blood pressure o Algnates (increases viscosity, oesophagitis)

Selective anti-muscarinic agents

Vagus nerve releases Ach acting on muscarinic M1 Ach receptor on parietal cells. Pirenzepine heals gastric and duodenal ulcers, selective M1 antagonist (doesn't cross blood brain barrier so no effects on CNS), duration treatment 4-6 weeks. Side effects = dry mouth, blurred vision

H+/K+ ATPase or "proton pump" in acid secretion

- Located in the cannalicular membrane - Hydrogen ions are generated within the parietal cell from dissociation of water. The hydroxyl ions formed in this process rapidly combine with carbon dioxide to form bicarbonate ion, a reaction cataylzed by carbonic anhydrase. - Bicarbonate is transported out of the basolateral membrane in exchange for chloride. The outflow of bicarbonate into blood results in a slight elevation of blood pH known as the "alkaline tide". This process serves to maintain intracellular pH in the parietal cell. - Chloride and potassium ions are transported into the lumen of the cannaliculus by conductance channels, and such is necessary for secretion of acid. - Hydrogen ion is pumped out of the cell, into the lumen, in exchange for potassium through the action of the proton pump; potassium is thus effectively recycled. - Accumulation of osmotically-active hydrogen ion in the cannaliculus generates an osmotic gradient across the membrane that results in outward diffusion of water - the resulting gastric juice is 155 mM HCl and 15 mM KCl with a small amount of NaCl. Parietal cells secrete acid. On simulation, proton pump vesicles fuse with PM to produce canaliculi. Need to activate pepsinogen to pepsin and reduce bacterial colonisation. Stomach more acidic then lysosome. K retrieved from lumen in process thus must be actively transported out of cell. Ultimate source of H is CO2 in blood. Often get alkaline tide in blood after meal. Also secrete intrinsic factor for B12 synthesis (pernicious anaemia). Mucous neck cells produce mucous (glycosylated by Golgi)

Briefly describe why acetylsalicylic acid (aspirin) but not paracetamol exasperates stomach ulcers.

- Paracetamol has good analgesic and antipyretic activity due to CNS prostaglandin synthesis inhibition but weak anti-inflammatory activity o No gastric or platelet side effects o This is because it is a selective (although weak) COX-2 inhibitor - COX-1 is a constitutive enzyme, found in most cells - COX-2 is inducible (barely found in normal cells) - Aspirin is non-selective Prostaglandins induce mucous and HCO3- secretion protecting the stomach epithelium from the acid secreted by parietal cells

Omeprazole is a proton pump inhibitor used for the treatment of gastric ulcers. a) How does it act? b) AstraZeneca recently made the pure (S)-enantiomer esomeprazole. What is the advantage of this compound OR pure enantiomers in general terms? / b) Why did AstraZeneca make the s-enantiomer esomeprazole?

- The S isomer is the active isomer - One of the cytochrome P450 enzymes converts the S to the R isomer in humans (effectively doubling concentration of active form) - However, the particular isozyme (CYP2C19) that does this conversion is not equal in all human populations (efficacy, expression level etc) o E.g. poor metabolizers - Leads to different levels of active drug if the racemic mixture is given - Although the S-isomer is more potent in humans, the R-isomer is more potent in rats - NB. Part of the reason for production of the esomeprazole drug was because omeprazole was running out of patent time

Why are antihistamines such as ranitidine now used mostly to treat heartburn rather than ulcer, for which they were designed? b) Given the structure of ranitidine, what chemical is it based on, and what are the salient chemical modifications?

- Ulcer now primarily treated with antibioitics (as well as proton pump inhibitors) to kill H. pylori - Draw histamine o Extend chain o Separate 2 pharmacophores - Ranitidine prevents insertion of the pump to the membrane - Furan ring mimics imidazole ring of histamine

H2

= histamine G coupled receptor

H2 receptor antagonists

Cimetidine Ranitidine Famotidine Nizatidine Mepyramine Thioperamine Can heal gastric and duodenal ulcers, reduces acid by blocking H2 receptors on surface of parietal cells. Duration of treatment 4-8 weeks at full dose. Maintenance treatment indefinite -half dose nocte. Doesn't modify natural history of disease when stopped. Also used in peptic oesophagitis, NSAID ulcer, undiagnosed dyspepsia (young). Side effect (usually well tolerated): cimetidine - gynaecomastia - enzyme inhibitor therefore interactions. Cimetidine has higher affinity than histamine but 0 efficacy (histamine has positive efficacy). 2 pharmacophores separated by chain

Antacids:

Counteract or neutralize acidity, usually in the stomach For symptom relief, can assist ulcer healing. 2 main types: aluminium compounds, S/E = constipation. Magnesium compounds, S/E = diarrhoea. Sodium bicarbonate, S/E = belching - beware patients with sodium restricted diets! Alginates - increases viscosity, may be useful in oesophagitis

gastric pits of stomach

Exocrine ducts from gastric glands that lead to the stomach lumen Cells: -Stomach lumen -> epithelial cells -Mucous neck cells -Oxynitic cells -Endocrine cells -Chief cells

What 3 agonists principally regulate acid secretion?

Histamine binds H2 --> cAMP --> H+ (acid) pumped by K/H+ ATPase Acetylcholine binds M3 (muscarinic) --> Ca2+ --> H+ (acid) pumped by K/H+ ATPase Gastrin binds CCK2 --> Ca2+ --> H+ (acid) pumped by K/H+ ATPase

How does the histamine cAMP pathway cause gastric acid secretion?

Histamine binds to H2 ATP --> cAMP by adenylcyclase activated by Gas cAMP activates PKA Vesicle/cytoskeletal protein phosphorylated = H+/K+ ATPase insertion into membrane

Histamine vs cimetidine

Histamine vs Cimetidine (H2 antagonist) - Histamine has lower affinity - Cimetidine has zero efficacy, histamine has positive efficacy - Cimetidine is a selective, competitive antagonist - Extending the chain and separating the 2 pharmacophores

Hydrochloric acid is formed in the following manner:

Hydrogen ions are formed from the dissociation of carbonic acid. Water is a very minor source of hydrogen ions in comparison to carbonic acid. Carbonic acid is formed from carbon dioxide and water by carbonic anhydrase. The bicarbonate ion (HCO3−) is exchanged for a chloride ion (Cl−) on the basal side of the cell and the bicarbonate diffuses into the venous blood, leading to an alkaline tide phenomenon. Potassium (K+) and chloride (Cl−) ions diffuse into the canaliculi. Hydrogen ions are pumped out of the cell into the canaliculi in exchange for potassium ions, via the H+/K+ ATPase. These receptors are increased in number on lumenal side by fusion of tubulovesicles during activation of parietal cells and removed during deactivation. This receptor maintains a million-fold difference in proton concentration. ATP is provided by the numerous mitochondria. As a result of the cellular export of hydrogen ions, the gastric lumen is maintained as a highly-acidic environment. The acidity aids in digestion of food by promoting the unfolding (or denaturing) of ingested proteins. As proteins unfold, the peptide bonds linking component amino acids are exposed. Gastric HCl simultaneously cleaves pepsinogen, a zymogen, into active pepsin, an endopeptidase that advances the digestive process by breaking the now-exposed peptide bonds, a process known as proteolysis.

Zollinger-Ellison syndrome

Hypersecretion of gastric acid that produces peptic ulcers as a result of a non-beta-cell tumor of the pancreatic islets Ectopic tumours of G cells (gastrinomas), hyper-secretion of gastrin, hypersecretion of acid. Treated with e.g. omeprazole (H+ pump inhibitor)

The mechanism of proton pumping by H+/K+ ATPase

Involves a cycle of conformational changes which leads to changes in affinity for H+/K+

Gastric ulcers

Peptic ulcers that occur in the stomach Erosion of the gastric mucosa Disintegration of the stomach lining which interferes with the production of mucous that coats the lining allowing stomach acid to come in to contact with the stomach. Can be caused by helicobacter pylori, stress, alcohol, excessive NSAIDs and Zollinger-Ellison syndrome

Omeprazole

Proton pump inhibitor Chiral atom is sulfur

Paracrine

Relating to or denoting a hormone that has effect only in the vicinity of the gland secreting it.

Histamine biosynthesis and storage:

Released by ECL (enterochromaffin-like) cells Histidine + B6 --> Histamine by Histidine decarboxylase

Active form of omeprazole

S enantiomer but CYP2C19 (cyt P450 family) converts S to R, effectively doubling concentration of active form. Expression and efficacy of this enzyme different in different people (some are poor metabolisers) so if racemic mix given, different people will have different levels of active S. R-isomer more potent in rats

Chelates and complexes:

Tri potassium Dicitratobismuthate (DENOL) and sucralfacte - heal duodenal and gastric ulcers.