Conjugation Reactions Flashcards
What phase of metabolism is oxidation (cytochrome P450)?
PHASE 1 metabolic reaction: metabolite formed with certain functional groups
What phase is glucuronidation?
(Conjugation reaction): PHASE 2 metabolic reaction
What phase follows phases 1 and 2?
Excretion (biliary or renally)
What do phase 2 reactions require?
They require a functional group as a site for conjugation (often phase 1 metabolite)
Describe what a drug undergoes in the hepatocyte.
- Drug present in blood
- Transporters facilitate uptake into the cell for metabolism
- Follows either phase 1 or phase 2
- Transporters for elimination
What type of molecules are conjugating molecules?
Endogenous - glucose or amino acid derivatives
What physicochemical properties do resulting conjugates have?
They are extremely polar (pka <3), terminal metabolites (not going to be metabolised further)
What is biliary and renal excretion of conjugates facilitated by?
Facilitated via efflux transporters (e.g. MRP2, MRP4)
Where are glucuronides formed?
In the liver
o Some Glucuronides are present in blood in systemic circulation – how do they get there if they’re formed in the liver?
MRP3 and MRP4 on side of hepatocyte efflux glucuronide into systemic circulation. Once it is in systemic circulation, it can be excreted via the kidneys
What is the general principle metabolism reactions follow?
Drug with certain functional group (e.g. COOH)
+
Endogenous compound (e.g. glucuronic acid)
–>
Conjugate (e.g. glucuronide)
What are the types of conjugation reactions?
- direct conjugation
- conjugation of the phase 1 metabolite
What is direct conjugation?
- often parallel reaction to CYP metabolism: diclofenac, propofol
- already HAVE particular groups in their structure needed for conjugation
What phase 2 reactions occur with OH functional group?
Glucuronidation or Sulphation
What phase 2 reactions occur with COOH functional group?
Glucuronidation or Glycine conjugation
What phase 2 reactions occur with NH2 functional group?
Glucuronidation or Acetylation
What is a common mechanism for the activation step of conjugating agents?
- Conjugating agent gets activated as a coenzyme (for glucuronides, sulphates and acetyl conjugates) OR
- Drug (for amino acid conjugates) - complex, involves intermediary metabolism. Reactive intermediates - co-enzymes or co-substrates
What is a common mechanism for the synthetic step?
(Simple, transferase enzymes)
1. Enzymes transfer conjugate to molecule and a metabolite is formed
What enzymes facilitate glucuronidation?
UGT enzymes (UDP-glucuronosyltransferases)
Where are UGT enzymes located?
• UGT enzymes located on luminal side of the endoplasmic reticulum – membrane bound.
Present mainly in the liver, but also in the intestine and kidney
What catalyses the addition of glucuronic acid to a substrate to form glucuronide metabolite?
Glycoproteins
What is glucuronic acid a derivative of?
Glucose - readily available in the body
Are conjugate metabolites generally active or inactive?
As they are terminal metabolites that are going to be eliminated, they are generally inactive*
*exceptions: morphine 6-glucuronide, ezetimibe glucuronide
What drugs are conjugation metabolic pathways most important for?
Lamotrigine, mycophenolic acid, valproic acid (epileptic drugs glucuronidated)
What glucuronides cause renal toxicity?
acyl glucuronides of NSAIDs
What glucuronides contribute towards DDIs?
gemfibrozil glucuronide (inhibitor of OATP1B1 and CYP2C8)
What compounds do UGTs conjugate?
Multiple endogenous compounds - bilirubin, steroid hormones, thyroxine and bile acids
What is the difference in levels of UGT between adults and babies?
o New-born baby does not have some levels of enzymes
o UGT1A1 activity and protein expression reach adult levels at 3-6 months of age
o Morphine glucuronidation (UGT2B7) deficient in young infants
What is congenital jaundice?
Increase in bilirubin levels that have not been conjugated (UGT1A1)
What is Gilbert’s syndrome?
Unconjugated hyperbilirubinemia, deficiency in UGT1A1 in adults (UGT1A1*28)
o Occurs in 2-13% of Caucasian
What are patients with UGT1A1*28 variant associated with?
Associated with higher risk of neutropenia (low neutrophils) as a side effect when receiving irinotecan
What is Irinotecan disposition?
- Does undergo some P450 metabolism which form the inactive metabolites
- Forms SN-38: ACTIVE METABOLITE – glucuronidated via UGT1A1 etc
- Build up of UGT1A1*2B leads to higher risk of neutropenia – therefore start at lower dose
What UGT enzymes are the most abundant glucuronidation enzymes in both healthy kidneys and tumour tissue?
UGT1A9, UGT2B7 and UGT1A6
reduced renal glucuronidation in tumours
What glucuronidation is seen in obese patients?
Increased
What enzymes facilitate sulphation reactions?
SULT - sulphur transferase
Where are SULT enzymes located?
Mainly in the cytosol - in contrast to membrane-bound UGTs.
Present in the liver and small intestine (SULT1A1)
*o SULT1A3 expressed in the human foetal liver
Is an activation step required for SULT enzymes?
YES - via ATP, cofactor PAPS (3’phosphoadenosine 5’-phosphate). More readily saturable than UGTs: – concentration of PAPS is app. 10-fold < UDPGA
What drugs do SULT enzymes metabolise?
Paracetamol, salbutamol and troglitazone
What are SULT linked to the hepatoxicity of ?
some drugs e.g. troglitazone
What is Glycine conjugation limited to?
Limited to acids: drug becomes activated as acyl coenzyme A intermediate
What is Acetylation limited to?
Limited to amines: formation of N-acetyl conjugates.
- acetyl CoA is the coenzyme
- Genetic polymorphisms in N-acetyl transferase (NAT)
- Rapid and slow acetylators (sulfasalazine)
What is a phase 3 reaction: conjugate hydrolysis and enterohepatic recycling?
Form glucuronide but then if it gets excreted into the bile and goes into the intestine, it can get hydrolysed and converted back to the parent via enzymes.
- this causes recycling of the parent molecule and prolonging the potential effect of the drug
- occurs in the small intestine
What enzymes are used in phase 3 reactions?
o Beta-gluronidase (for glucuronide metabolites) o Sulphatase (for sulphate metabolites)
