SF083

Drug Absorption: Diffusion depends on...

1. The non-ionized molecules
     pH of the medium is important

2. Depends on the amount of free, unbound drug:
     Extent of protein binding (plasma or cell)
     Extent of binding to formulation/excipients

SF083

Transporter that influence drug passage across membrane

1. Organic anion transport proteins (OATs or OATPs)
   
   
2. Organic cation transport proteins (OCTs)
         (New nomenclature “SLC---”)
   
   
3. P-glycoprotein (P-gp or MDR1)
   
   
4. Multidrug resistance-assoc. protein (MRPs)

SF083

Limitations for Drug Absorption (3)

1. Tissue perfusion (blood flow)
2. Diffusion-limited absorption
1. Partition coefficient
2. Surface area
3. First pass effect
4. Distribution and metabolism

SF083

Biopharmaceutics Classification System for Assessing Oral Absorption of Drugs (Table)

SF083

General Properties of Absorption Barriers (Table)

SF083

8 Factors influencing GI drug absorption

Drug Driven:

1. Formulation
2. Water solubility
3. Lipid solubility
4. pKa

Physiology Driven:

5. GI motility
6. gastric pH
7. Posture
8. Other drugs (anti-cholinergics)

SF083

Bioavailability

Fraction of drug reaching systemic circulation

Intravenous dosing has a bioavailability of 1.0 (all drug enters circulation)
Oral drugs have bioavailability less than 1.0 because either because drugs are not absorbed in the gut or because they are metabolized before they reach the bloodstream

SF083

Factors influencing drug Distribution in Body

1. pKa of cpd and pH of tissue compartment

Acidic drugs more likely to be concentrated in blood compartment

Low pKa drugs are uncharged (protonated) at low pH. Drugs are absorbed from the stomach (acidic) into the bloodstream (neutral) where they become charged (deprotonated). Drug accumulates in the blood because it can no longer diffuse across the membrane.

Basic drugs more likely to be concentrated in tissue

High pKa are mostly uncharged (protonated) at neutral/high pH. Drugs can freely move from the blood into tissues because they are uncharged. Once in the tissues (slightly acidic) the drug becomes charged (deprotonated), causing accumulation in the tissue.

2. Drug binding to pr./tissues. Tissue binding in non-target sites (depots) increases the time it takes for levels of the drug to reach equilibrium between the drug, target and depot.

3. Specialized distribution barriers (BBB, placenta)

SF083

Apparent Volume of Distribution calc.

Determined by following formula: IV Dose/[blood]

VD = (mg/Kg)/(mg/L) = L/Kg

Note: not a real volume or space, but rather a calculated
 value used to determine the tissue distribution of a drug

SF083

Basic Principles of Drug Metab.

1. Metabolism often removes biological activity, but can also result in active drug
   
   
2. Most metabolism follows first-order rate kinetics rate of loss is proportional to concentration in plasma half-lives of a drug in plasma
   
   
3. First order enzyme kinetics takes 5 half-lifes to essentially remove drug from body (~4%)

SF083

3 Phases of Drug Metabolism

SF083

CYP 450 Major Player in Phase I Rx

1. CYP 450s (biggest player)

2. MOA

3. Proteases

4. Esterases

5. Amidases

SF083

Clearance in Relationship to Drug Elimination

Clearance (Cl) – the volume of plasma that would contain the amount of drug excreted per unit time (minute)

Volume of plasma that would have to lose all of the drug that it contains within a unit of time (usually 1 min) to account for an observed rate of drug elimination

Clearance expresses the rate or efficiency of drug removal from the plasma vol/time (ml/min)

Cl = ke (Vd) or Cl = (0.693/t_1/2)*Vd

ke = Elimination Constant – Measure of the rate at which a drug is eliminated from circulation

SF083

Drug Filtration Rate

Dose Adjustment for patients with Renal Impairment

Drug Filtration Rate  =  GFR x fu x [Drug]

     (fu = free fraction)

 C_ss = (Dose/τ)/CL_E

Maintain usual dosing interval but reduce dose in proportion to  ↓CL_E

or
  

Maintain usual dose but increase dosing interval in proportion to  ↓CL_E

SF083

Elimination by the kidneys is dependent on...

• rate of filtration (glomerular)
• Affects all drugs and metabolites of appropriate molecular size.
• Influenced by protein binding
• the extent of tubular secretion
• the extent of renal tubular reabsorption of the drug
•   Not influenced by protein binding
•   May be affected by other drugs, etc.
• Mechanism: Reabs by non-ionic diffusion
• Affects weak acids and weak bases
• Only important if excretion of free drug is major elimination pathway.
  
  

SF083

Excretion methods for class 1-4 drugs

SF083

Enterohepatic Cycling

Drugs excreted into the bile may be subject to enterohepatic cycling, causing the drug to persist in the body longer than expected. Usually due to the conversion of the drug metabolite back into the active form of the drug by intestinal enzymes or intestinal microflora, which is then reabsorbed into circulation. Drugs subject to enterohepatic cycling are typically:

• large (MW>300)
• polar drugs
• Actively secreted into bile (often glucuronide conjugation)
  
  

PH083

Describe how disease of specific organ systems (eg liver, kidney) may alter drugs in body

Liver:

Alteration of liver function will primarily affect the rate of drug metabolism.

Retention would be caused be caused by a decrease Phase I/II metabolism, resulting in less polar forms of the drug, which are more difficult to excrete. Accumulation  may cause toxicity if too much accumulates. Decreased production of active drug would occur if the liver was responsible for converting a prodrug to an active drug (eg codeine to morphine). This may result in decreased therapeutic effect.

Kidney:

Alteration of kidney function would primarily affect the excretion of  the drug. Kidney damage would decrease filtration rate, causing retention of drug and drug metabolites, which may result in toxicity.

SF085

What can you use Vd for?

Calc Vd and Loading dose

• Needed for determining clearance of a drug from the body
• Needed for determining loading dose of a drug

Vd = Dose / [plasma time 0]

loading dose = Vd x desired plasma conc

SF085

What is the importance of Cl?

Calc Dosing Rate and Maitenance Dose

• Provides an index of the efficiency by which a drug is removed from the body
• Is subject to changes due to disease state, genetic and environmental factors
• Needed for determining Dosing Rate and Maintenance Dose.
  
  Cl = (0.693/t_1/2)*Vd
  
  Dosing Rate = Cl x target concentration 
  Maintenance Dose = (Dosing Rate/F) x Dosing Interval
  
  F stands for bioavailability or how well the drug is absorbed.
  
  F = (AUC oral/ AUC IV) x 100. AUC stands for area under curve. The idea is that when a drug is given intravenously, there is 100% bioavailability; yet when it is given in other ways (such as orally), there is less bioavailability.

SF085

Reading a drug absorption graph

SF085

Factors influencing PK/PD propetries of drugs

• Environmental factors – (diet, lifestyle, other drugs)
• drug interactions (DIs)
• inhibitors of enzyme or transport processes
• inducers of enzyme or transport processes   
• Genetic considerations
• Mutations in enzyme and/or transporters
• Bioavailablity, metabolism, elimination
• Physiology/pathophysiology based considerations
• alterations in renal function
• alterations in hepatic function

SF085

Enzyme Inhibitors vs Enzyme Inducers

Enzyme inhibitor                        

Drug A blocks Metabolism of drug B 

PD-increase response

PK-increase plasma t1/2

Effects are immediate

Ex Cimetidine-H2 receptor antagonist, used to treat
peptic ulcer. Potent inhibitor of several different cyp450 enzymes.
Drug interactions include warfarin, benzodiazepines,
phenytoin, morphine
PK effects- decreased clearance; increased half-life
PD effects- increased response; increased duration

Enzyme Inducer
Drug A increases amt of enzyme for metab
- Decreased response
- Decrease plasma t1/2
- Effects are delayed

- Mechanism involves binding of the drug to site on
gene encoding the enzyme which turns on
synthesis of more enzyme

- Effects not only from drugs (also env (smoking) and herbal supplements)

Ex 1 - Phenytoin- anticonvulsant medication. Over time phenytoin causes an increased expression level for various cyp450 enzymes (cyp3A4)
Ex 2 - Rifampicin - antimicrobial agent; induce cyp450

PK - increased clearance; decreased half-life
PD - decreased response

SF085

Polymorphic distribution for drug metabolism

Genetic Variations leading to these polymorphisms (3)

Mostly EM (extensive metaboliser) but small amounts of PM (poor metabolisers) and UM (Ultrametabolisers)

1. CYP gene family
   Ex CYP2D6   -> PM/EM/UM
   Caucasians highest frequency of PM phenotype = inc toxicity
   Asians highest frequency of IM phenotype
   Africans highest frequency of the UM phenotype = Reduced Respiration
   Clinical Ex: Codeine to Morphine
2. UDP glucoronosyl transferase
3. N-acetyl transferase
   Ex Slow Acetylator NAT-2 phenotype most common in caucasians and africans. Leads to inc chance of Lupus

SF085

In general, what are the 6 sources of variability in Drug Metabolism

SF104 Tutorial

1. Due to induction – other drugs, environment
2. Due to diet – natural inducers and inhibitors
3. Due to inhibition – disease processes and drugs
4. Due to genetics – defect in metabolic pathways
5. Due to development – special populations
6. Due to disease – impairment of organ function

 Take a quick look at if you have time

SF139

Allergic Drug Reactions

Immune system-mediated drug reactions (type I -IV)

• 5-10% of adverse drug reactions
• Immune responses to drugs occur because drugs or their metabolites bind to self-proteins and these are recognized as foreign by the immune system
• Such responses may be Type I-IV
• Drugs can bind directly to MHC -> makes MHC foreign and illicits immune response

Type I: IgE & mast cell/basophil mediated occur immedately (anaphylaxis) or accelerated (1-72 hours)
Type II: IgG or IgM mediated against cell or tissue antigens
Type III: IgG mediated against soluble proteins
Type IV: Delayed cell-mediated reactions

SF139

Allergic or Hypersensitivity reactions mechanism

• Uncertain but anaphylactoid or pseudo-allergic may involve direct:
  1. histamine release
  2. prostaglandins/leukotrienes or
  3. kinins
  
• with
  1. hives
  2. angioedema
  3. asthma
  4. hypotension
  
  
• Ex. Stevens-Johnson (like burns on skin), Toxic epidermal necrolysis
  
  
• Drugs are haptens that stimulate immune responses by modifying self-proteins
• Allergic drug reactions require sensitization and this takes time – very rare that sensitivity will occur to a drug that a person has never seen before
• Sensitization may occur through cross-reactivity of structurally related drugs
• Clinical manifestations reflect immune mechanisms

SF139

Case 1

56 yrs old woman, vague hx penicillin allergy, had received amoxicillin 6 mths ago.
Is prescribed cloxacillin -> 2nd dose develops pruritis, hives, swelling

What type of reaction? 

Type I – b/c it occurs quickly and with hives (typical of IgE)

What is the mechanism?

Mast cell degranulation

What are possible & probable causes?

Cloxicillin possibly

When did sensitization occur?

She had amox 6 months before, so she is showing cross-reactivity with cloxicillin (strcut similar)

How could it be proved that cloxacillin was responsible?

Skin test

SF139

Case 2

26 yr old drug addict admitted with endocarditis
Treated with i.v.benzylpenicillin for 8 weeks.
After 6 weeks noted to be pale, Hb 8g/L and reticulocytosis on blood smear

What is the mechanism?
Type II
What class of antibodies are involved?
IgM, IgG

Pen binds to cell memb, Ab binf to pen, activation of complement, NK cells attack cell and we get RBC lysis -> low Hb

SF139

Case 3

34 yr old woman with Crohn’s disease
Treatment with high dose steroids fails and started on Infliximab, chimeric murine/human anti-TNF antibody
First dose tolerated but 9 days after 2nd dose she develops generalized hives, leukocytosis, swollen painful joints, fever, hematuria and vasculitic rash

What is the likely diagnosis?

Type III b/c it is targeted against a soluble molecule. Also, Vasculitic rash is typical of type III

What is the mechanism?

Ag-Ab complex builds up – immune complex related vasculitis. Binding of IgM or IgG Abs to soluble antigen causes an insoulble complex to form that is deposited on surface of the tissue. Complement is activated.

This brings upon serum sickness - fever, vasculitis, nephritis and arthritis

Why is there a delay?

Takes 9 days for immune response – Ag-Ab complexing – thus faster if we gave the drug again

SF139

Case 4

54 yr old paraplegic man has recurring skin breakdown on buttocks
Controls local infection with bacitracin ointment
After several months, nurse notices blistering red rash on his buttocks, which he had not felt

What type of rxn?

Type IV reaction

What immune mechanism would explain clinical symptoms and signs?
Involves activated T cells. The macrophage eats up bacitracin, grinds it up, expresses it and T-cells then target it as foreign

How could you prove the bacitracin responsible?

Skin test

What is this common in?

Contact dermititis - nickel on skin for example

SF147

Classification of HS Reactions (model #2)

Type: 1 - Immediate

Immu mech: Pre-formed IgE activates mast cells.
3 key players:

Mast cell
IgE
Antigen

Ex: Hay fever, asthma

Type: 2 - Cytotoxic

Immu mech: IgG, IgM binds to immobilized antigens (on a cell, RBC examples later), usually followed by complement activation

Ex: Goodpasture Disease; autoimmune hemolytic anemia

Type: 3 - Immune Complex-Mediated

Immu mech: IgG, IgM, IgA form immune complexes with antigens (solube antigen = not on a cell); complement and leukocytes become activated

Ex: Glomerulonephritis

Type: 4 - Cell-Mediated

Immu mech: Mononuclear cells become activated to secrete cytokines and/or become cytolytic; antibody not involved

Ex: Granulomatous disease; rejection in transplantation

SF147

How is IgE involved in Type 1 HS reactions?

• Patient is exposed to antigen and immune response to antigen develops
• CD4+ T helper produce Th2 cytokines (e.g. IL-4, 5, 13)
• CD4+ T helper cells help B cells to produce antibody to the antigen
• B cells undergo “class switching” and start making IgE antibody (induced by the Th2 cytokines)
• IgE binds to Fc receptors on the surface of mast cells and basophils
• Once patient has been sensitized, IgE is “ready to go” on surface of mast cells; can bind the antigen upon subsequent exposure

SF147

What happens when IgE Binds Antigen?

What are the effects?

Cross-linking of antibody and antigen required on mast cell surface

Note: can also be accomplished when C3a and C5a bind to their receptors on the mast cell surface

This results in the release of:

• Histamine
• Platelet Activating Factor
• Prostglandin (PGD2)
• Leukotrines C, D and E
• Thromboxane
• Chemotactic factors for neutrophils & eosinophils
• Enzymes (proteases)

1. Smooth muscle contraction
2. Increased vascular permeability
3. Chemotaxis of eosinophils
4. Platelet activation
5. Protease effects, activation of kinin pathway
   
   Anaphylactic shock may develop due to bronchoconstriction, airway obstruction and circulatory collapse

SF147

Type 2: Cytotoxic Response Involving
Antibodies and Immobilized Antigen

4 clinical examples

Some type 2 HS reactions involve complement:

Typically mediated by IgG or IgM, which are antibody isotypes that fix complement in humans (recall complement cascade from Inflammation lectures)

Example 1: Autoimmune
Hemolytic Anemia - antigen is on the cell and antibodies recognize it, activate complement and C5-9 MEMBRANE
ATTACK COMPLEX (MAC), forms holes, destroys permeability barrier

Example 2: Goodpasture’s Disease

The antigen is extracellular and the immobilized antigen is type IV collagen on the glomerular basement membrane (BM) and in the lung. In the kidney, antibody binds to BM and activates complement. This is followed by inflammation and tissue injury. 1 in a million.

Example 3: Opsonization (coats and makes things easier to phagocytose) by C3b or Antibodies leads to phagocytosis and cell destruction

Some Type 2 HS Reactions do not require complemen:

Example 4: Antibody-Dependant Cytotoxicity: antibody-antigen-Fc receptor interaction forms a “bridge” between a cytotoxic cell (e.g. natural killer cell) and a target cell.

SF147

In type 3: How do immune complexes cause tissue
injury?

Complement becomes activated, Inflammatory response develops and leukocytes are recruited, become activated and release mediators such as proteases and reactive oxygen intermediates, which can injury the tissue.

Problems they cause:

Immune complexes may be deposited in blood vessels causing vasculitis or in the glomeruli in the kidney causing glomerulonephritis (GN).

Immune complexes are a feature of some autoimmune
disease such as systemic lupus erythematosus (SLE).

SF147

Type 4: Cell Mediated Cytotoxicity
2 Major mechanisms and examples

Antibodies are not involved (contrast other HS responses).

Two major mechanisms:

1. Delayed-type HS reactions – macrophages (or other antigen-presenting cell), CD4+ T helper cells

Example 1: Contact hypersensitivity (poison ivy, nickel)

• chemical ligands are not proteins, referred to as “haptens”, which bind covalently to endogenous “carrier proteins”
• hapten-carrier protein complex interacts with Langerhans’ cell in the skin (antigen-presenting cell) and are phagocytosed
• inside the Langerhans cell, antigen is cleaved into peptides, which are presented in class II MHC (HLA) molecule to CD4+ T helper cells
• CD4+ T cell becomes activated and secrete cytokines such as interferon-γ (a Th1 cytokine) and chemokines, which attract other mononuclear cells
• Antigen presenting cell becomes activated to release mediators of inflammation

Example 2: Tuberculosis

• Response of Th cells and macrophages to persistent antigen results in the development of granulomas.

2. Cytotoxic killer lymphocytes - CD8+ T lymphocytes and natural killer (NK) cells

• CD8+ T cell recognizes antigenic peptide presented in self, class I MHC molecule on the surface of a nucleated cell
• CD4+ T helper cell recognize antigenic peptide presented in self, class II MHC molecule on the surface of an antigen-presenting cell
• CD4+ T helper cell helps to activate CD8+ T cell by providing cytokines such as IL-2
• Activated CD8+ T cell binds to the target cell by engaging class I MHC +antigenic peptide with its T cell receptor
• Activated CD8+ T cell kills the target cell through a process involving perforin (like MAC), which creates a pore in the target cell membrane
• NK cells use a similar mechanism for target cell lysis, but the mechanism for recognizing target cells is somewhat different