Uridine Diphosphate Glucuronosyltransferases UGTs EC 24117

Subcellular location: Luminal side of the ER. Organ distribution: Liver, kidney, and intestine, Cofactor: Uridine diphosphoglucuronic acid (UDPGA), Overall reaction: See Fig. 2.6.

Dipeptidase

Mercapturic Acid

Figure 2.6. (a) Mechanism of formation of glucuronide conjugate. X is a nucleophilic site on the molecule such as an oxygen atom (in the case of alcohols, phenols and carboxylic acids), a nitrogen atom (in the case of amines, azaheterocyclic molecules), or a carbon atom (not common, but seen with activated carbons such as phenylbutazone). (b) Mechanism of breakdown of GSH conjugate to mercapturic acid (or N-acetyl cysteine conjugate). UDP uridine diphosphate; NAT N-acetyltransferase.

Isoforms, substrates, and inhibitors: See Table 2.12. Typically, the glucuronides are considered to be the final metabolic products. However, other metabolic modifications have been reported, including diglucuronide conjugates. Gemfibrozil glucuronide is a potent inhibitor of CYP2C8.

A number of transporters (mostly hepatic such as MRP3 and MRP2) are known to transport glucuronide conjugates.

Sometimes, albeit rarely, glucuronides have significant pharmacological activity (i.e., morphine glucuronide).

Quantification of glucuronide metabolites:

• In vitro use of 14C-UDPGA as a cofactor followed by LC separation and radioactive detection.

• Quantification of the parent drug is conducted in the presence and absence of b-glucuronidase to regenerate the aglycan parent. The difference in quantification will be the amount of glucuronide metabolite formed.

Considerations for in vitro incubations:

Detergents (Brij 58, lubrol, and Triton X-100) or alamethicin provide access to the luminal side of the ER.

• Alamethicin is a pore-forming peptide; it should be dissolved in methanol and mixed with microsomes (20-50 mg/mg LM) on ice for 15-20 min prior to incubation.

• Detergents inhibit P450 activity, and since a number of metabolites are formed by P450 and UGT enzymes, alamethicin is used more often than detergents in in vitro incubations.

1,4-Saccharolactone inhibits b-glucuronidase activity (see Sect. 2.3.9.2).

General Glucuronide Stability:

• Glucuronides of primary and secondary amines are acid labile.

• Acyl glucuronides are base labile.

Acyl glucuronides: A number of glucuronide conjugates of car-boxylic acids (acyl glucuronides) are considered reactive electro-philic metabolites. They are capable of undergoing hydrolysis, intramolecular rearrangement, and intermolecular reactions with proteins leading to covalent drug/protein adducts. A number of acyl glucuronides lead to idiosyncratic hepatotoxicity that is considered to be immune-mediated.

Table 2.12. Major sites of UGT expression and the enzymes' substrates and inhibitors (Uchaipichat et al. 2006)

Major sites of

UGTa

expression

Substrates

Inhibitors

1A1 (15%)

Liver, intestine

Bilirubin, ß-estradiol, ethynylestradiol, morphine, SN-38

Atazanavir

1A3

Liver

Bile acids, cyproheptadine, alizarin, hyodeoxycholic acid

1A4 (20%)

Liver

Imipramine, trifluoroperazine

Hecogenin

1A5

Liver, brain

1A6

Liver, brain

Seratonin, naphthol

Amitriptyline, phenylbutazone

1A7

Stomach

Phenylbutazone, sulfinpyrizone

1A8

Intestine

1A9

Liver, kidney

Propofol

Androsterone, phenylbutazone, sulfinpyrizone

1A10

Stomach, intestine

Amitriptyline

2B4

Liver

Xenobiotics, bile acids, hyodeoxycholic acid

2B7 (35%)

Liver, kidney, intestine

AZT, morphine

Amitriptyline, androsterone

2B10

Liver, prostate, mammary

2B11

Liver, kidney, prostate, adrenal

2B15

Liver, prostate

(S)-Oxazepam

Amitriptyline, quinidine, quinine

2B17

Prostate

Amitriptyline, quinidine, quinine

AZT 30-Azido-30-deoxythimidine aPercent of marketed drugs that are metabolized by the major UGT enzymes (Williams et al. 2004)

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