Anti-bacterial_Agents_III_2024_LR.pdf

Anti-Bacterial Agents III Dr. Laura Ryan Consultant Microbiologist St. Vincent’s University Hospital 14 02 2024 Agents inhibiting nucleic acid synthesis Objectives of lecture Outline of antibiotics that inhibit nucleic acid synthesis – – – – different mechanisms of action of antibiotics pharmacokine...

Anti-Bacterial Agents III Dr. Laura Ryan Consultant Microbiologist St. Vincent’s University Hospital 14 02 2024 Agents inhibiting nucleic acid synthesis Objectives of lecture Outline of antibiotics that inhibit nucleic acid synthesis – – – – different mechanisms of action of antibiotics pharmacokinetics clinical uses side effects Classification of antimicrobials Characteristics of nucleic acid Nucleic acids are polynucleotides Two main classes are DNA and RNA DNA is the template for synthesis of both DNA and RNA DNA exists as a double helix in the cell RNA exists as a single strand in the cell DNA and RNA consist of strands of nucleotides Structure of RNA and DNA Nucleic Acid Synthesis Enzymes Local unwinding of DNA - DNA gyrase (Topoisomerase II) Synthesis of DNA - DNA polymerase Synthesis of RNA - RNA Polymerase DNA synthesis Occurs when nucleotide units are joined to form DNA DNA replication Polymerase chain reaction Artificial gene sequences DNA replication DNA template DNA double helix unwinds, exposing unpaired bases for new nucleotides to hydrogen bond to Topoisomerases (DNA gyrase) Helicases – uncoil the double-stranded DNA, exposing the nitrogenous bases Complementary base pairing takes place, forming a new doublestranded DNA molecule “Semi-conservative” replication since one strand of the new DNA molecule is from the 'parent' strand Inhibition of nucleic acid synthesis Targets 1. 2. 3. 4. 5. Alteration of the base-pairing properties of the template Inhibition of either DNA or RNA polymerase Direct effects on DNA itself Inhibition of DNA gyrase (Topoisomerase II) Inhibition of nucleotide synthesis (folate anti-metabolites) 1. Alteration of the base-pairing properties of the template Intercalating agents Acridines-proflavine, acriflavine Double the distance between adjacent base pairs, causing frameshift mutations Not in clinical use DNA Intercalation Normal DNA helix Intercalated DNA Phosphodiester backbone A-T T-A G-C Stacked base pairs A-T T-A G-C Intercalation of drug results in overall distortion of helical structure 2. Inhibition of either DNA or RNA polymerase Actinomycin D Binds to Guanine bases in DNA and blocks the movement of RNA Polymerase preventing transcription Not useful as an anti-bacterial agent Used in cancer chemotherapy 2. Inhibition of either DNA or RNA polymerase Rifampicin (a rifamycin) Inhibitors of bacterial RNA polymerase Bind to the enzyme and induce conformational change – which weakens binding of polymerase to initiation sequence of DNA Bactericidal Rifampicin - mechanism of action Inhibits RNA Polymerase Rifampicin Spectrum of activity Gram +ve bacteria Bactericidal against some Gram-ve bacteria All Mycobacteria Administration PO or IV Metabolism Liver Interferes with cytochrome p450 Rifampicin Clinical uses TB Leprosy Legionella Eradication of N. meningitidis, H. influenzae Combination therapy for the treatment of serious S. aureus infections Side effects Hepatic impairment GI upset Colours body fluids an orange colour Increases rate of metabolism of other drugs 3. Direct effects on DNA itself Alkylating agents Form covalent bonds with bases in the DNA and prevent replication e.g. Nitrogen Mustard derivatives 3. Direct effects on DNA itself Metronidazole Mechanism of action – binds to DNA and prevents nucleic acid synthesis (perhaps due to damage to the DNA by toxic oxygen products) – needs to be partially reduced to be effective and this only takes place in anaerobic cells, little activity against aerobic organisms Bactericidal Metronidazole - mechanism of action Inhibits nucleic acid synthesis Causes DNA damage Metronidazole Spectrum of activity – Introduced as anti-protozoal agent – Anaerobic bacteria – either Gram +ve and Gram –ve organisms Administration – PO or IV Pharmacokinetics – well absorbed and distributed – eliminated in the urine, slightly metabolised, mainly un-changed C. difficile infection Normal colon Pseudomembranous colitis Metronidazole – spectrum of activity Metronidazole is active against a wide range of pathogenic microorganisms: Bacteroides fragilis, Fusobacterium necrophorum, F. nucleatum, Clostridioides difficile, Clostridium perfringens, C. septicum, anaerobic cocci Gardnerella vaginalis - anaerobic bacterium, causative in bacterial vaginosis Helicobacter pylori - Gram-negative, microaerophilic, and acidophilic bacterium Trichomonas vaginalis - anaerobic, flagellated protozoan parasite, STI Entamoeba histolytica – anaerobic parasite amoebozoan Giardia lamblia - anaerobic flagellated protozoan parasite Balantidium coli – intestinal protozoan parasite Metronidazole - clinical indications Infections caused by anaerobic bacteria, particularly species of Bacteroides and anaerobic streptococci, e.g., bloodstream infection, peritonitis, brain abscess, necrotising pneumonia, osteomyelitis, puerperal sepsis, pelvic abscess, pelvic inflammatory disease and wound infections Pseudomembranous colitis caused by Clostridioides difficile Urogenital trichomoniasis (Trichomonas vaginalis) Bacterial vaginosis Amoebiasis Giardiasis Acute ulcerative gingivitis Acute dental infections Treatment of Helicobacter pylori infection associated with peptic ulcer as part of triple therapy Metronidazole – side effects – – – – – nausea, vomiting, diarrhoea rash, urticaria, flushing dizziness, headache pancytopenia disulfiram-like effect occurs with alcohol as metronidazole inhibits alcohol and aldehyde dehydrogenases – Increased LFTs, cholestasis or mixed hepatitis 4. Inhibition of DNA gyrase Fluoroquinolones These drugs are selective for the bacterial enzyme because it is structurally different from the mammalian enzyme Rapidly bactericidal Resistance due to reduced uptake and enzyme affinity Fluoroquinolones Ciprofloxacin Levofloxacin Ofloxacin Moxifloxacin Norfloxacin Nalidixic acid Mechanism of action – inhibit topoisomerase II (DNA gyrase) and topoisomerase IV, required for bacterial DNA replication, transcription, repair and recombination Pharmacodynamics – Concentration-dependent killing – Prolonged persistent effects Fluoroquinolones DNA Gyrase (cleaves DNA backbone) RNA Polymerase Target the alpha subunits of DNA gyrase prevents it from supercoiling the bacterial DNA which prevents DNA replication Fluoroquinolone – spectrum of activity Ciprofloxacin most commonly used Broad spectrum ▪ Gram -ve organisms - Enterobacterales (Gram – ve bacilli, e.g. E.coli) - Also effective against H. influenza, N. gonorrhoeae, Shigella sp., Campylobacter, Pseudomonas aeruginosa ▪ Less effective against Gram +ve organisms, some more effective than others (e.g. Levofloxacin, moxifloxacin) ▪ Atypical cover e.g. Chlamydia sp., Legionella sp. ▪Mycobacteria Pharmacokinetics of fluoroquinolones Administration PO or IV, excellent oral bioavailability (Al + Mg antacids interfere with absorption) Distribution - wide esp. kidney, prostate + lung Metabolism - Hepatic – (can inhibit cytochrome P450 enzymes) Excretion - Liver and renal Clinical uses of fluoroquinolones Infections caused by Gram-negative organisms – Pyelonephritis – Abdominal sepsis – Bacterial prostatitis – Pneumonia Hospital acquired Community acquired (Levofloxacin) Pseudomonas aeruginosa infections esp. in CF External otitis media (topically) Osteomyelitis (excellent penetration into bone and joint usually in combination) Gonorrhoea Cervicitis Anthrax Eradication of S. typhi in carriers Eradication of Neisseria meningitidis Fluoroquinolones - side effects Clostridioides difficile infection Tendonitis and tendon rupture Lowers seizure threshold QT interval prolongation - moxifloxacin Constipation Skin rashes Arthropathy CNS – headache, dizziness (CNS pathology) Rare – renal impairment, hypersensitivity reactions, photosensitivity Important interaction between ciprofloxacin + theophylline (P450) leads to theophylline toxicity November 2018 5. Inhibition of Nucleotide Synthesis This can be accomplished by early effects in the metabolic pathway (missing in mammals) Class II reaction - making small molecules, e.g., folate Synthesis (required for nucleotides) Inhibition is produced using anti-metabolites Concept of anti-metabolites Chemotherapy based on synthesis of small molecules that are functional analogues of cell metabolites that inhibit natural metabolic pathways Metabolic map / general plan of nucleotide synthesis Eight distinct types of nucleotide, each containing 3 modular parts (sugar, base and phosphates) Summary of antimetabolites Drug 1. Suphonamides p-aminosalicylic acid 2. Methotrexate Pyrimethamine Trimethoprim 3. Tubercidin Formycin Nebularin Fluoracil Normal cell analogue Reaction or site inhibited PABA Folate synthesis Folate Folate metabolism Purines/ Pyrimidines Nucleic Acid Synthesis Anti-metabolite action of sulphonamides and trimethoprim Para-amino-benzoic acid (PABA) is essential for the synthesis of folic acid in bacteria Folic acid is required for the synthesis of the precursors of DNA and RNA in both bacteria and mammals Mammals obtain their folic acid in their diet so they do not need to synthesise it The Action of sulphonamides and trimethoprim on bacterial folate synthesis Sulphonamides Mechanism of action Structural analogue of PABA Sulphonamides compete with PABA for the enzyme dihydropteroate synthetase. Thus some local anaesthetics (PABA esters) can antagonise the antibacterial effect of sulphonamides. Mechanism results in growth inhibition is bacterostatic Action negated by presence of pus/tissue breakdown which contains thymidine and purines which bacteria use to bypass their need for folic acid. Resistance (common) due to synthesis of an enzyme which is insensitive to the drug (plasmid mediated) Pharmacokinetics of sulphonamides Administration – PO/IV/IM – not usually topical due to allergic reactions (except for silver sulfadiazine-infected burns) Absorption good oral absorption with the exception of sulfasalazine (used for U.C.) Distribution wide, crosses placenta + BBB Metabolism liver Excretion kidney Clinical uses of sulphonamides Anti-microbial agent – Decreasing therapeutic importance with the exception of cotrimoxazole - due to resistance – Broad spectrum - Gram +ve / Gram –ve Cotrimoxazole – Pneumocystis jiroveci (fungus) - cotrimoxazole – Nocardia infections - cotrimoxazole – Toxoplasmosis (Protozoa) - cotrimoxazole Other uses – Inflammatory bowel disease – sulfasalazine – Ulcerative colitis – sulfasalazine – Burns - prevents bacterial colonisation Sulphonamides – side effects GI - nausea, vomiting and diarrhoea Headache and mental depression Cyanosis due to methaemoglobinaemia Hepatitis Hypersensitivity reactions - rash, anaphylaxis, fever – Stevens-Johnson Syndrome – Erythema multiforme - skin + mucus membrane lesions +/- systemic illness. Photosensitivity reactions Bone marrow suppression, anaemia, agranulocytosis, thrombocytosis Crystallisation in the renal tract leading to crystalluria - precipitation of acetylated metabolites in the urine Kernicterus (now called chronic bilirubin encephalopathy [CBE]) risk esp. in premature infants. Sulphomanides may unbind bilirubin from albumin, increasing blood level. Unbound bilirubin can cross into the brain Increased activity by methotrexate/warfarin (displaced from albumin) Trimethoprim Mechanism of action Structural analogue of pteridine moiety in folate - folate antagonist Trimethoprim competes with folate for the enzyme dihydrofolate reductase Bacterial enzyme is much more sensitive than the mammalian enzyme Mechanism inhibits growth is bacteriostatic Trimethoprim is active against most bacterial pathogens except enterococci and P. aeruginosa Used in combination with sulphonamides, e.g., cotrimoxazole Pharmacokinetics of trimethoprim Administration PO Absorption fully absorbed in the GIT Distribution Wide distribution -high conc. in kidney + lungs, relatively high concentration in CSF Excretion Kidney – up to 60% of orally administered dose can be recovered in the urine 24 hours later (active form) Clinical uses of trimethoprim Active against most bacterial pathogens Main use - urinary tract infections Prophylaxis of UTI Respiratory tract infections Cotrimoxazole – Pneumocystis jiroveci (fungus) - cotrimoxazole – Nocardia infections - cotrimoxazole – Toxoplasmosis (Protozoa) - cotrimoxazole Trimethoprim – side effects Fewer than with cotrimoxazole Nausea, vomiting + rashes Folate deficiency causes megaloblastic anaemia - give folic acid supplements Hypersensitivity reactions of cotrimoxazole are caused by sulphonamide and are not dose related Cotrimoxazole Sulfamethoxazole (400mg) + Trimethoprim (80mg) Both drugs inhibit different enzymes in the same pathway effective at 1/10 doses Combination has slowed resistance Main use is in the treatment of Pneumocystis jiroveci (PJP) Increasingly being used to treat resistant Gram –ve organisms No significant benefit in UTIs over trimethoprim Summary Rifampicin inhibits RNA polymerase Metronidazole binds to DNA and prevents nucleic acid synthesis perhaps due to damage to the DNA by toxic oxygen products Fluoroquinolones are selective for the bacterial DNA gyrase enzyme because it is structurally different from the mammalian enzyme Sulphonamides mimic the natural metabolite PABA and disrupt folate production in bacteria. Selective toxicity is achieved because folate synthesis does not occur in eukaryotes Trimethoprim disrupts folate synthesis by inhibiting the enzyme dihydrofolate reductase Combination therapy of sulphonamides and trimethoprim (cotrimxazole) have synergistic effect by blocking two steps in the same pathway Questions?