Anticancer agents Topoisomerase inhibitors Topoisomerases are separated into two types - topoisomerases I and II. They are the enzymes of the cell nucleus responsible for the control, behavior and modification or topology of DNA during the replication and translation of genetic material. Both these classes of enzyme utilize a conserved tyrosine, however these enzymes are structurally and mechanistically different. 2 Their function consists in causing breaks in one or both strands of the double helix of DNA. These breaks make possible the passage of the helix strand through the cleft and next the re-connection of the DNA helix, which means a renewed ligation. During this process a covalent bond between topoisomerase and DNA is formed, called the cleavage complex. Topoisomerase inhibitors stabilize the cleavage complex, which maintains the break of the helix and inhibits the physiological function of DNA. 3 Topoisomerases I and II differ in the following properties: . topoisomerase I is not a specific enzyme for the cell cycle, while the activity of topoisomerase II is greatest in the phase of logarithmic growth and in rapidly growing tumors . topoisomerase I induces a break in only one strand of the DNA helix; topoisomerase II is responsible for a break in one or both strands of the helix . the activity of topoisomerase I does not depend on ATP; the activity of topoisomerase II depends on ATP. 4 Type I topoisomerases are subdivided into two subclasses: . type IA topoisomerases which share many structural and mechanistic features with the type II topoisomerases, and . type IB topoisomerases, which utilize a controlled rotary mechanism. Examples of type IA topoisomerases include topo I and topo III. 5 Historically, type IB topoisomerases were referred to as eukaryotic topo I, but IB topoisomerases are present in all three domains of life. Interestingly, type IA topoisomerases form a covalent intermediate with the 5' end of DNA, while the IB topoisomerases form a covalent intermediate with the 3' end of DNA. Recently, a type IC topoisomerase has been identified, called topo V. While it is structurally unique from type IA and IB topoisomerases, it shares a similar mechanism with type IB topoisomerase. 6 Type II topoisomerase is also split into two subclasses: . type IIA and . type IIB which share similar structure and mechanisms. Examples of type IIA topoisomerases include eukaryotic topo II, E. coli gyrase, and E. coli topo IV. Examples of type IIB topoisomerase include topo VI. Both type I and type II topoisomerases change the linking number of DNA. Type IA topoisomerases change the linking number by one, type IB and type IC topoisomerases change the linking number by any integer, while type IIA and type IIB topoisomerases change the linking number by two. 7 8 Large amounts of topoisomerase I are present in all phases of the cell cycle. Topoisomerase is the target of action of anticancer agents. Anticancer activity is demonstrated by the alkaloid camptothecin, which is found in the tree Camptotheca acuminata, growing in China. Because of the significant toxicity of this alkaloid, semi-synthetic derivatives (topotecan and irinotecan) showing better therapeutic properties are used in therapy. Camptothecin: R1 = R2 = R3 = H R2 R1 R3 O Topotecan: R = H, R = -CH N(CH ) , R = OH N 1 2 2 3 2 3 HYCAMTIN N O H O Irinotecan, R = -C H , R = H R = N N O H5C2 1 2 5 2 3 O CAMPTO, CAMTOSAR OH 9 The modification of camptothecin by addition of the phenolic hydroxyl and dimethylaminomethyl groups improved water solubility and reduced the occurrence of unpredictable side- effects without compromising its antitumor activity. The lactone form of topotecan is pharmacologically active, although the equilibrium between the lactone and hydrolysis product favors the ring-opened form at physiological pH. Both camptothecin and topotecan are inhibitors of topoisomerase I and cause single-strand breaks in DNA. Either or both of these effects may be cytotoxic. R2 R1 R 3 O Camptothecin: R = R = R = H N 1 2 3 N Topotecan: R1 = H, R2 = -CH2N(CH3)2, R3 = OH O HYCAMTIN H5C2 O OH 10 Irinotecan is a pro-drug. Its metabolite (7-ethyl-10-hydroxycamptothecin) demonstrates approx. 100 times greater activity than irinotecan. Irinotecan, apart from anticancer action, is also an acetylcholinesterase inhibitor. R2 R1 R 3 O N N O Irinotecan, R1= -C2H5, R2 = H H O H C R3 = N N O CAMPTO, CAMTOSAR5 2 O OH 11 Topotecan and irinotecan are effective in the treatment of . microcellular and non microcellular carcinoma of the lung, . ovaries (also cancer resistant to therapy with cisplatin and carboplatin), . colon and rectum, . pancreas, kidneys and esophagus. Additionally, . topotecan is used in the treatment of acute leukemia and . irinotecan in the treatment of carcinoma of the cervix, breast, stomach, liver and in malignant tumors of the head and neck. 1212 During therapy with topotecan and irinotecan the following side- effects can appear: . suppression of bone marrow, . disturbance of the function of the gastric tract, . thrombocytopenia, . eosinophilia, . influenza-like symptoms, . increased activity of hepatic enzymes and . others. 13 Epipodophyllotoxins Natural glycoside – podophyllotoxin – present in the roots of the plant May Apple (Podophyllum notatum/peltatum) is not used in medicine because of its high toxicity. Teniposide and etoposide – semisynthetic derivatives – are less toxic. 14 OH H CO OCH3 3 4' O H Teniposide, VEHEM, VUMON O O 9 O Etoposide, R = -CH3; ETOPOL, VEPESID H O 6 O R O O 4 OH OH Teniposide (Vumon, VM-26) is a chemotherapeutic medication mainly used in the treatment of childhood acute lymphocytic leukemia. It is in a class of drugs known as podophyllotoxin derivatives and slows the growth of cancer cells in the body. 15 Mechanism of action Teniposide causes dose-dependent single- and double-stranded breaks in DNA and DNA-protein cross-links. The mechanism of action appears to be related to the inhibition of type II topoisomerase activity since teniposide does not intercalate into DNA or bind strongly to DNA. The cytotoxic effects of teniposide are related to the relative number of double-stranded DNA breaks produced in cells, which are a reflection of the stabilization of a topoisomerase II-DNA intermediate. Administration The medication is injected though a vein and burns if it leaks under the skin. It is sometimes used in combination with other anticancer drugs. Side-effects Teniposide, when used with other chemotherapeutic agents for the treatment of ALL, results in severe myelosuppression. Other common side effects include, hypersensitivity reactions, and alopecia. 16 Etoposide phosphate (brand names: Eposin, Etopophos, Vepesid, VP-16) is an inhibitor of the enzyme topoisomerase II. It is used as a form of chemotherapy for malignancies such as Ewing‘s sarcoma, lung cancer, testicular cancer, lymphoma, non- lymphocytic leukemia, and glioblastoma multiforme. It is often given in combination with other drugs. It is also sometimes used in a conditioning regimen prior to a bone marrow or blood stem cell transplant. Its chemical make-up derives from podophyllotoxin, a toxin found in the American Mayapple. 17 Administration It is given intravenously or orally in capsule form. If the drug is given by IV it must be done slowly over a 30 to 60 minute period because it can lower blood pressure as it is being administered. Blood pressure is checked often during infusing, with the speed of administration adjusted accordingly. In general, patients are advised to call their doctor in case of fever symptoms of infection or painful injection sites, as these may progress severely without adequate medical attention. Patients are advised to drink large amounts of fluids after treatment to prevent damage to the bladder and kidneys, typically 1.5 to 3.5 litres of water on the day of treatment and for several days after. 18 Side-effects Common are: . low blood pressure . hair loss . pain and or burning at the IV site . constipation or diarrhea . metallic food taste . bone marrow suppression, leading to: - decreased white blood cell counts (leading to increased susceptibility to infections) - low red blood cell counts (anemia) - low platelet counts (leading to easy bruising and bleeding) Less common are: . nausea and vomiting . allergic type reactions . rash . fever, often occurring shortly after IV administration and not due to infection . mouth sores . acute myeloid leukemia 19 Anticancer agents • Cytostatic antibiotics The following are considered clinically important antitumor agents: . actinomycins . anthracyclines . mitomycins . bleomycins. 20 Actinomycins All actinomycins are derivatives of 3-phenoxazone-1,9-dicarboxylic acid (actinocin). Sar L-Pro L-Pro Sar An example of an antineoplastic L-MeVal D-Val D-Val L-MeVal actinomycin - dactinomycin O L-Thr L-Thr O C O O C (actinomycin D), the main antibiotic N NH constituent of Streptomyces parvullus. 2 O O Dactinomycin consists of a planar, CH3 CH3 tricyclic phenoxazone ring in the quinone oxidation state and two Dactinomycin, ACTINOMYCIN D, identical pentapeptide lactone COSMEGEN appendages. N2,1,N2’,1’'-(2-Amino-4,6-dimethyl-3-oxo- The pentapeptides are made up of 3H-phenoxazine-1,9-diylodicarbonyl)- L-proline, L-threonine and three bis[threonyl-D-valilprolil(N-methylglicyl- nonessential amino acids – D-valine, (N-methyl-valine)1,5-3,1-lactone sarcosine and methylvaline. 21 The mechanism of action Sequention of bases O O CH2 P _O O O O Actinomycins can intercalate or guanine cytosine P _ CH O O 2 N O HNH insert into DNA between base- O N NH N N N pair steps. H NH O O CH2 The preferred base-pair - O O O P ACTINOMYCIN O _O P _ guanine-cytosine and cytosine- O O O CH guanine. 2 HNH O N O N N N N N O O HNH CH2 cytosine guanine O O O O In the intercalation process, the P P _ _O O O O CH3 helix must unwind some of the CH2 O HNH N O base pairs in order for there to N NH N N N O O CH2 be space for the actinocin O O thymine adenine O O P P _ moiety.