Metabolism and drug design 2018 ADME - Metabolism Organisms posesses a large set of enzymes able to modificate xenobiotics Original issue is to protect internal environment from toxic agents Compounds are generally made more hyrophilic to enhance renal or hepatobiliar excretion ADME - Metabolism Knowledge of metabolic pathways is crucial By metabolization, drug can be - deactivated (most metabolites) - made more potent (prodrugs) - converted to reactive form (toxicity) Enzyme set ready for metabolisation Fraction of drugs metabolized by various systems Oxidation enzymes Cytochrome P450 (CYP, P450) often occures rearrangements in structure after primary oxidation FeO3+ involved in mechanism Oxidation enzymes Cytochrome P450 (CYP, P450) Oxidation enzymes Cytochrome P450 (CYP, P450) human genome has 57 P450 genes Oxidation enzymes Cytochrome P450 (CYP, P450) only few are of high importance Oxidation enzymes Cytochrome P450 (CYP, P450) some uncommon reactions Oxidation enzymes Cytochrome P450 (CYP, P450) Oxidation enzymes Flavin-containing Monooxygenase (FMO) 7 forms of FMO, located in endoplasmatic reticulum distincly from CYP P450, only soft nucleophiles are substrates (N, S, P in phosphines) Oxidation enzymes Flavin-containing Monooxygenase (FMO) Oxidation enzymes Monoamine Oxidase (MAO) two forms – MAO A and MAO B present in mitochondrial membrane of hepatocytes and neurons flavoprotein oxidase protein as FMO, but with different mechanism releasing ammonia and hydrogen peroxide Oxidation enzymes Aldehyde Oxidase Xanthine Dehydrogenase contains molbdenum and iron in active center present in cytosol, mainly in hepatocytes electron can be transferred to an oxidized pyridine nucleoside or to oxygen forming H2O2 substrates are: various aldehydes heterocycles containing N (purines are substrate for both enzymes) Oxidation enzymes Peroxidases similar mechanism with CYP system involving FeO generating of radical radical further undergoes propagation, dimer formation or dealkylation Oxidation enzymes Alcohol Dehydrogenases (ADH) concentrated in liver, specifity for primary and some secondary alcohols reaction is generally reversible at least 7 different genes for ADHs Oxidation enzymes Aldehyde Dehydrogenases (ALDH) concentrated in liver, mainly in mitochondria at least 19 genes for ALDHs are known Reduction enzymes CYP P450, ADH both can catalyze reductions of aldehydes and imines these reverse reactions occures in environments with low O2 tension (e.g. venous section of liver) Reduction enzymes CYP P450, ADH Reduction enzymes NADPH-P450 Reductase main function is to restore P450 system, but has enzymatic activity to some P450 substrates as well Reduction enzymes Aldo-Keto Reductase (AKR) substrates are mainly sugar aldehydes, steroids, prostaglandines Reduction enzymes Quinone Reductase (NQO) present in cytosol, substrates are quinones (both ortho- and para-), iminoquinones, nitro- and azo- compounds Reduction enzymes Glutathione Peroxidase (GPX) reduces hydroperoxides including H2O2 six different enzymes, most of them contains selenocystein in the active site restored by GSH reductase overall reaction can be written as Hydrolysis enzymes Epoxide Hydrolase (GPX) located in microsomes and endoplasmic reticulum catalyses simple addition of water to epoxide: Hydrolysis enzymes Esterases and Amidases heterogenous group of enzymes with similar basic mechanism of action present in all environments in the organism most important: lipases acetylcholin esterase (ACHE) butylcholin esterase Conjugation enzymes UDP-Glucuronosyl Transferases (UGT) conjugation of glucuronic acid to various functional groups: alcohols, phenols, amines, heterocyclic nitrogens, amides thiols, acids conjugates are: - more polar - ionized at physiological pH - have increased Mr - are actively excreted by carriers in liver and kidney Conjugation enzymes UDP-Glucuronosyl Transferases (UGT) Conjugation enzymes UDP-Glucuronosyl Transferases (UGT) endogenous substrates: bilirubin, steroids, lipids, leucotrienes, thyroid hormones, vitamines A, D three major gene families: Conjugation enzymes UDP-Glucuronosyl Transferases (UGT) Conjugation enzymes Cytosolic Sulfotransferases (SULT) works with cosubstrate PAPS (3-phosphoadenosin-5- phosphosulfate) both O-sulfates and N-sulfates can be formed Conjugation enzymes Cytosolic Sulfotransferases (SULT) works with cosubstrate PAPS (3-phosphoadenosin-5- phosphosulfate) Conjugation enzymes Cytosolic Sulfotransferases (SULT) Conjugation enzymes Glutathione-S-Transferases (GTS) - glutathione is endogenous nucleiophile - attacs electrophilic molecules - glutathione conjugates are excreted by transporters into bile - six major classes if GTS Conjugation enzymes Glutathione-S-Transferases (GTS) Conjugation enzymes Glutathione-S-Transferases (GTS) conjugates may be further converted to N-acetylcystein, mercaptouric or thiol derivatives and excreted by urine Metabolism-induced toxicity Part of molecule is altered by oxidation, reduction or conjugation to form a reactive electrophile Electrophiles reacts with internal nucleophiles (proteins, nucleic acids, small peptides) Alkylation of such structures may cause organ toxicity and cancerogenity. Metabolism-induced toxicity Metabolism-induced toxicity two kinds of toxicity – altering protein functions - triggering immunity reaction Toxicophores Epoxides phenytoin is metabolized by P450 to epoxide metabolite causing hepatic necrosis and aplastic anaemia Toxicophores Epoxides carbamazepine epoxide causes teratogenicity and skin rash oxcarbamazepine is much less toxic Toxicophores Quinone Imines phenacetin is oxidated by P450 and causes cellular hepatotoxicity Toxicophores Quinone Imines Antimalaric agent amodiaquine causes hepatotoxicity Toxicophores Quinone Imines One pathway for diclofenac causes hepatotoxicity Toxicophores Quinone Imines ACHE inhibitor tacrine causes hepatotoxicity Toxicophores Quinone Imines Indomethacin caused agranulocytosis Toxicophores Quinone Imines Practolol had to be withdrawn from the market due to skin and eye lesions Atenolol have not toxic acetanilide moiety Toxicophores Quinone Imines Paracetamol has the same toxicologic issue Toxicophores Quinone Imines Carbutamid caused bone marrow toxicity and was withdrawn Tolbutamid have not this problem Toxicophores Quinone Imines Lidocain as antiarrhytmic has short biological half-time. Longer acting analogues procainamide and tocainide are toxic, flecainid do not so. Toxicophores Nitrenium Ions Clozapine forms reactive nitrenium ions causing agranulocytosis Toxicophores Nitrenium Ions Modification of circle brought non-toxic analogues Toxicophores Iminium Ions Antidepressant mianserin causes agranulocytosis Toxicophores Iminium Ions Cardiotonic vesnarinon causes agranulocytosis both due to iminium and quinone imine formation Toxicophores Hydroxylamines hydroxylamines are further metabolized to reactive nitroso derivatives Sulphonamides and dapson are metabolized that way causes agranulocytosis and skin hypersensitivity Toxicophores Thiophene ring thiophene S-oxides and S-chlorides causes covalent protein bonding agranulocytosis by ticlopidine (platelet inhibitor) nephrotoxicity of suprofen (NSA, withdrawn) Toxicophores Thiophene ring ticlopidine metabolism in white blood cells Toxicophores Thioureas thioureas are metabolized to sulfon acids Metiomid was withdrawn due to blood dyscrasias Cimetidine not shows this problem