Anti-inflammatories

Question 1

What are NSAIDs?

Answer 1

they inhibit the production of inflammatory mediators such as prostaglandins and thromboxanes
- cyclooxygenases (COX) are enzymes which synthesis these paracrine mediators
- NSAIDs inhibit COX enzymes so inhibit the production of PGs and thromboxanes
- therefore they prevent paracrine signalling between cells at sites of inflammation

Question 2

how are paracrine mediators produced?

Answer 2

arachidonic acid is produced from membrane phospholipids
- it acts as a second messenger and as a substrate for lipoxygenases and cyclooxygenases which leads to generation of paracrine mediators
- lipoxygenase pathway produces leukotrines
- cyclooxygenase pathway forms PGs and thromboxanes

Question 3

what are prostaglandins?

Answer 3

• PGs are created by cells and act only in the surrounding area before they are broken down
• PGs control neighbourhood processes such as vasodilation (PGE2), aggregation of platelets during clotting (PGD), constriction of uterus during labour (PGF) and a hyperalgesic (makes you more sensitive to pain) (PGE2)

Question 4

what is the role of PGs?

Answer 4

• deliver and strengthen pain signals to induce inflammation
• chemoattractants - recruit immune cells

Question 5

what do NSAIDs result in?

Answer 5

1. anti-inflammatory - modify the inflammatory reaction
   - decrease vasodilation and in turn oedema
   - effective against headaches as they reduce the vasodilator effect of PGs on cerebral tissue
2. analgesic - reduce pains
   - decrease production of PGs in damaged and inflamed tissue which sensitises nociceptors to inflammatory mediators e.g. bradykinin, 5-HT
3. antipyretic - lowers raised temperature
   - thermostat in hypothalamus is activated by IL-1 induced COX2 production of PGe

Question 6

what is the structure of the COX enzyme?

Answer 6

• made up of 2 identical subunits, each with 2 catalytic sites
• 2 active sites are collectively termed as prostaglandin synthase:
  
  • one side has a cyclooxygenase active site
  • the other side has a peroxidase site which activates haem groups in
    the COX reaction
• enzyme complex is a dimer of 2 subunits, so there are 2 cyclooxygenase sites and 2 peroxidase sites in close proximity
• each subunit has a carbon-rich knob which anchors the complex to the ER membrane
• cyclooxygenase active site is buried deep inside the protein which is accessible through a tunnel in the knob which acts as a funnel to guide arachidonic acid into the enzyme

Question 7

what are the types of COX enzymes?

Answer 7

1. COX1: constitutive expression
   - important for platelets, stomach, kidney and colon
2. COX2: inducible expression
   - most cells, especially inflammtory cells after stimulation with cytokines, growth factors or tumour promoters
   - immediate-early response gene

3 COX3: in CNS, target for paracetamol?

Question 8

what is the action of NSAIDs?

Answer 8

• COX1 and COX2 inhibitors prevent the cyclooxygenation reaction of arachidonic acid to prevent production of PGs
• aspirin inhibits expression of transcription factor NF-kappaB which has a key role in triggering gene transcription of inflammatory mediators

Question 9

how are NSAIDs selective?

Answer 9

• COX1 contains isoleucine residue while COX2 contains valine residues, allowing the creation of NSAIDs which are selective to COX1 or COX2
• COX2 is inducible, so is only produced when there is tissue damage
• COX1 is constitutive so maintains homeostatic mechanisms e.g. PGs in the GI tract maintain the mucus layer to protect GI cells from acid

Question 10

how may NSAIDs have unwanted side effects?

Answer 10

if PG production is inhibited by COX1-inhibitors, the mucus layer in the GI tract is reduced, so acid from the stomach may harm GI cells, leading to ulcers

Question 11

what is the action of aspirin?

Answer 11

suicide inhibitor:
- aspirin is composed of an acetyl group which attaches to salicylic acid
- when aspirin attacks cyclooxygenase, its acetyl group covalently binds to the serine residue in COX to permanently deactivate the enzyme
- this prevents arachidonic acid from reaching the cyclooxygenase site through the tunnel
- aspirin therefore blocks the active site

Question 12

what are the 2 isoforms of COX?

Answer 12

COX1: constitutive, expressed in most cells including platelets, has a housekeeping role in homeostasis

COX2: induced in inflammatory cells when activated and produces prostanoid mediators of inflammation

Question 13

do NSAIDS show a lot of selectivity? how do types of NSAIDs differ?

Answer 13

no, they show little selectively.

main differences are in toxicity, duration of action and pain tolerence

Question 14

what are the side effects of NSAIDs?

Answer 14

1. Gut - PGs normally inhibit acid secretion to protect mucosa, so NSAIDs prevent this
   - causes diarrhoea, dyspepsia, nausea, vomiting, gastric bleeding and ulceration
   - co-administration of misoprostal (PG analogue) helps
   - can be relieved by use of COX2-selective drugs
2. Renal function: PGs maintain renal blood flow, so NSAIDs may cause issues with renal function, leading to renal failure
3. liver damage: the metabolite of paracetamol in the phase 1 reaction is toxic to liver
   - patients must be careful if cytochrome p450 is induced as an overdose may cause liver failure
4. bronchospasm/asthma attacks
5. skin rashes

Question 15

what is the risk of GI complications with NSAIDs?

Answer 15

35-45% users sustain some form of GI damage

Question 16

what are the advantages of COX1/COX2-selective NSAIDs?

Answer 16

COX2-selective NSAIDS such as rofecoxib (Vioxx), valdecoxib, parecoxib and celecoxib can reduce burden of GI toxicity:
- analgesic and anti-inflammatory]

COX1-selective NSAIDs may be antithrombotic and can help prevent stroke (COX1 is involved in clot formation)

Question 17

what are the disadvantages of COX1/COX2-selective NSAIDs?

Answer 17

COX2-selective drugs may cause increased arterial blood pressure, increased atherogenesis, or increased thrombotic tendency
- there is evidence for these NSAIDs to have cardiovascular toxicity if COX2 is consistently inhibited
- increased blood pressure and increased salt retention -> kidney failure

COX1-selective drugs are not suitable with someone with a bleeding disorder, as these drugs prevent clot formation

Question 18

aspirin vs paracetamol:

Answer 18

aspirin:
- Anti-platelet action
- Reduced risk of colonic and rectal cancer
- Reduced risk of Alzheimer’s
- Weak acid, rapid and efficient absorption in the ileum
- Suicide inhibitor (irreversible)

paracetamol
- Analgesic-antipyretic due to CNS effects
- Weak anti-inflammatory
- Cox3/1 selective
- Well absorbed, metabolized in liver
- Less side-effects than aspirin with long term use, but large doses may increase kidney damage
- N-acetyl-p-benzoquinone imine is hepatotoxic in uncongugated form
- Competitive Inhibitor

Question 19

what is ibuprofen?

Answer 19

same as aspirin except it is a competitive inhibitor

Question 20

what are the clinical uses of NSAIDs?

Answer 20

• antithrombotic = aspirin (for patients with high risk of arterial thrombosis)
• analgesia:
  short term: aspirin, paracetamol, ibuprofen,
  long-term = naproxen (codeine)
• anti-inflammatory = ibuprofen, aspirin and naproxen
• antipyretic (reduces fever) = paracetamol

Question 21

what is rheumatoid arthritis?

Answer 21

• chronic inflammatory condition and autoimmune disease
• possibly genetic
• 3x more prevalent in women than in men
• affects joints, inflammation of synovium and erosion of cartilage and bone
• treated with DMARDs (reverse/halt the disease) and NSAIDs (alleviate symptoms)
• no cure
• smoking is a risk factor

Question 22

what are the mediators involved in the pathogenesis of rheumatoid arthritis?

Answer 22

• T cells activate macrophages which release cytokines involved in driving the inflammatory response:
  e.g. IL-1 and TNF-alpha

Question 23

how can rheumatoid arthritis be treated?

Answer 23

• methotrexate: low doses have cytotoxic and immunosuppresant activity (high doses for chemotherapy)
  
  • folic acid is the antagonist
• DMARDs: mixed group of drugs with different mechanisms of action
  
  • clinical effects are slow, so NSAIDs are provided as cover during
    induction phase

Question 24

give examples of some DMARDs:

Answer 24

1. sulfasalazine: sulfa drug used for chronic inflammatory bowel disease
   - bacteria in colon produce 5-aminosalicylic acid which acts as a free radical to decrease damage by neurtrophils
2. penicillamine (d-isoform): metabolite of penicillin, used as a heavy metal chelator in poisoning
3. gold compounds e.g. auranofin: inhibits induction of IL-1 and TNF-alpha

Question 25

what is the role of immunosuppressant drugs?

Answer 25

they inhibit the induction phase of the inflammatory response: - cyclosporin and glucocorticoids inhibit transcription of pro-inflammatory cytokines such as IL-2

Question 26

what is the mechanism of action of cyclosporin?

Answer 26

inhibits IL-2 synthesis to cause decrease in T cell proliferation: 1. cyclosporin binds to cyclophilin, a cytosolic immunophilin protein 2. the drug-immunophilin complex bind to and inhibit calcineurin to prevent activation/signalling via transcription factor NF-kappaB, which is regulated by calcium signalling in T cells - calcineurin is a phosphatase regulated by calcium and targets NF-kappaB transcription factor to produce cytokines 3. inhibition of calcineurin means NF-kappaB can no longer drive the production and secretion of cytokines IL-2, IL-1 and TNF-alpha

Question 27

what is the mechanism of action of glucocorticoids?

Answer 27

they act at the level of gene transcription by binding to DNA itself: - they act as repressors of transcription of genes that make pro-inflammatory cytokines - they affect IL-1, TNF-alpha and INF-gamma synthesis - effective in induction and effector phases of the immune response - they also inhibit NF-kappaB

Question 28

how are biopharmaceuticals used as anti-inflammatory drugs?

Answer 28

Humanised monoclonal antibodies: - can bind to cytokines and prevent their inflammatory response - high affinity and selectivity for target - they neutralise the action of soluble or membrane-bound pro-inflammatory cytokines - long half-life - the antibodies are engineered with human sequences to prevent rejection Soluble receptors: - proteins engineered that mimic the binding site of a known cytokine receptor - protein is injected into body and cytokine binds to it but has no downstream inflammatory activity

Question 29

what are the issues with biopharmaceuticals' use in anti-inflammatory drugs?

Answer 29

very expensive

Question 30

what is COPD? How is it treated?

Answer 30

chronic obstructive pulmonary disease: - disease driven by macrophages - symptoms: breathlessness and eventually respiratory failure - emphysema - treatment: B2 agonists, muscarinic antagonists and corticosteroids - cannot be cured

Question 31

what is asthma?

Answer 31

- shortness of breath especially when exhaling - symptoms: cough, wheezing, inflammation of airways, bronchial hyperactivity, reversible airway obstruction

Question 32

how can asthma be treated?

Answer 32

- bronchodilators: salbutamol is a B2-adrenoreceptor agonist (GPCR via Gs and adenylyl cyclase) on airway smooth muscle to relax the airways - polymorphisms in B2-adrenoreceptors cause reduced efficacy of bronchodilators - anti-inflammatory agents: prednisolone and omalizumab reduce cytokine production

Question 33

what are the types of respiratory allergies?

Answer 33

1. allergic rhinitis (hayfever) - issue in upper airways - allergen activates mast cells in the nasal mucosa - nasal congestion, sneezing and allergic conjunctivitis 2. allergic asthma (atopic) - issue in lower airways - allergen activates mast cells in lower respiratory tract - early/immediate phase reactions are reversible airway obstructions - bronchial hyperactivity - abnormal sensitivity to many stimuli, causing bronchoconstriction - acute attacks are reversible but can progress to chronic state - patients have an increased no. in mast cells in the bronchi

Question 34

what is involved in the late phase reactions of asthma?

Answer 34

- cytokines and leukocyte infiltration (especially eosinophils), leading to inflammation - some patients develop chronic asthma: chronic inflammation, tissue damage and airway remodelling - 50% of asthma patients suffer with this

Question 35

what causes asthma?

Answer 35

- associated with overactivity of T-helper cells as they drive the immune response and production of antibodes to the allergen - antibody is bound to mast cells and eosinophils - when receptors are activated from allergen binding to antibody, histamine is released and binds to smooth muscle receptors to cause constriction - histamine drives production of PGs which also cause bronchoconstriction

Question 36

what are mast cells and their role in asthma?

Answer 36

mediators of type 1 hypersensitivity reactions - cause: smooth muscle contraction, increased vascular permeability, mucous secretion, platelet activation, stimulation of nerve endings and recruitment of eosinophils - they secrete cytokines and chemokines during the immediate phase to set the scene for the late phase of an asthma attack - IgE has natural role in allergic reaction

Question 37

what is the pathology associated with chronic inflammation of the airways?

Answer 37

- clogged airways are full of mucus which restricts oxygen exchange - thickening and growth of smooth muscle cells affects the stretchiness of airways, so future allergen responses are worse - increase in mucus-producing cells

Question 38

what are the unwanted side effects of prednisolone steroid use?

Answer 38

Cushing's syndrome: - hypertension - moon face - poor wound healing - muscle wasting - increased abdominal fat

Question 39

what are the new therapies for treating asthma?

Answer 39

- humanised antibodies and soluble receptors to IgE (omalizumab), cytokines and chemokines (biopharmaceuticals) - prostaglandin D2-R antagonists: PG D2 is synthesised in mast cells by PGD synthase - PGD2 acts on DP1 receptors on vessels to mediate vasodilation, and also on DP2 receptors to attract T-helper2 cells and eosinophils - therefore PGD2-R antagonists prevent the vasodilation and recruitment - antagonists of DP1 and DP2 receptors, and inhibitors of PGDs are in clinical development for asthma therapy