Inhibitors of DNA Replication in Eukaryotic cells

DOXORUBICIN

The exact mechanism of action of doxorubicin is complex and still somewhat unclear, though it is thought to interact with DNA by intercalation. Doxorubicin is known to interact with DNA by intercalation and inhibition of macromolecular biosynthesis. This inhibits the progression of the enzyme topoisomerase II, which relaxes supercoils in DNA for transcription. Doxorubicin stabilizes the topoisomerase II complex after it has broken the DNA chain for replication, preventing the DNA double helix from being resealed and thereby stopping the process of replication.

The planar aromatic chromophore portion of the molecule intercalates between two base pairs of the DNA, while the six-membered daunosamine sugar sits in the minor groove and interacts with flanking base pairs immediately adjacent to the intercalation site, as evidenced by several crystal structures.

Intercalation occurs when ligands of an appropriate size and chemical nature fit themselves in between base pairs of DNA.

ETOPOSIDE

Etoposide phosphate (brand names: Eposin, Etopophos, Vepesid, VP-16) is an anti-cancer agent. It inhibits the enzyme topoisomerase II, which unwinds DNA, and by doing so causes DNA strands to break.

Etoposide forms a ternary complex with DNA and the topoisomerase II enzyme, preventing re-ligation of the DNA strands. This causes errors in DNA synthesis and promotes apoptosis of the cancer cell.

CAMPTOTHECIN

Camptothecin (CPT)cytotoxic quinoline alkaloid which inhibits the DNA enzyme topoisomerase I. CPT binds to the topo I and DNA complex (the covalent complex) resulting in a ternary complex, and thereby stabilizing it. This prevents DNA re-ligation and therefore causes DNA damage which results in apoptosis. CPT binds both to the enzyme and DNA with hydrogen bonds.

Toxicity of CPT is primarily a result of conversion of single-strand breaks into double-strand breaks during the S-phase when the replication fork collides with the cleavage complexes formed by DNA and CPT

RIFAMPICIN

Rifampicin inhibits DNA-dependent RNA polymerase in bacterial cells by binding its beta-subunit, thus preventing transcription to RNA and subsequent translation to proteins. Its lipophilic nature makes it a good candidate to treat the meningitis form of tuberculosis, which requires distribution to the central nervous system and penetration through the blood-brain barrier.

Rifampicin acts directly on messenger RNA synthesis. It inhibits only prokaryotic DNA-primed RNA polymerase, especially those that are Gram-stain-positive and Mycobacterium tuberculosis. Much of this acid-fast positive bacteria's membrane is mycolic acid complexed with peptidoglycan, which allows easy movement of the drug into the cell. Evidence shows that in vitro DNA treated with concentrations 5000 times higher than normal dosage remained unaffected; in vivo eukaryotic specimens' RNA and DNA polymerases suffered few problems as well. Rifampicin interacts with the β subunit of RNA polymerase when it is in an α2β trimer. This halts mRNA transcription, therefore preventing translation of polypeptides. It should be made clear, however, that it cannot stop the elongation of mRNA once binding to the template-strand of DNA has been initiated. The Rifampicin-RNA polymerase complex is extremely stable and yet experiments have shown that this is not due to any form of covalent linkage. It is hypothesized that hydrogen bonds and π-π bond interactions between naphthoquinone and the aromatic amino acids are the major stabilizers, though this requires the oxidation of naphthohydroquinone which is found most commonly in Rifampicin. It is this last hypothesis that explains the explosion of multi-drug-resistant bacteria: mutations in therpoB gene that replace phenylalanine, tryptophan, and tyrosine with non-aromatic amino acids result in poor bonding between Rifampicin and the RNA polymerase.

Well due to blocking in RNA transcription, DNA initiation is also not possible since the primer synthesis is also blocked by the drug.

APHIDICOLIN

Aphidicolin is defined as a tetracyclic diterpene antibiotic with antiviral and antimitotical properties. Aphidicolin is a reversible inhibitor of eukaryotic nuclear DNA replication. It blocks the cell cycle at early S phase. It is a specific inhibitor of DNA polymerase A, D in eukaryotic cells and in some viruses and an apoptosis inducer in HeLa cells.

NOVOBIOCIN

The molecular basis of action of novobiocin, and other related drugs clorobiocin and coumermycin A1 has been examined. Aminocoumarins are very potent inhibitors of bacterial DNA gyrase and work by targeting the GyrB subunit of the enzyme involved in energy transduction. Novobiocin as well as the other aminocoumarin antibiotics act as competitive inhibitors of the ATPase reaction catalysed by GyrB. The potency of novobiocin is considerably higher than that of the fluoroquinolones that also target DNA gyrase, but at a different site on the enzyme. The GyrA subunit is involved in the DNA nicking and ligation activity.

CIPROFLOXACIN

Ciprofloxacin is a broad-spectrum antibiotic active against both Gram-positive and Gram-negative bacteria. It functions by inhibiting DNA gyrase, a type II topoisomerase, and topoisomerase IV, enzymes necessary to separate bacterial DNA, thereby inhibiting cell division.

1. Actinomycin -binding between adjacent G-C bases in DNA (intercalation
2. chloramphenicol- inhibits peptidyltransferase of the 70S ribosome
3. erythromycin - binds to the 50S particle and arrests synthesis of the 70S ribosome
4. neomycin- binds to the 30S ribosomal subunits and inhibits binding of a tRNA
5. puromycin- premature chain termination
6. Rifamycin- inhibits RNA synthesis by binding to the β subunit of the RNA polymerase holoenzyme
7. streptomycin as erythromycin
8. tetracyclin -inhibits binding of tRNA to the 30S ribosomal subunit in eukaryotes
9. α-amanitin - inhibits polymerase II
10. chloramphenicol - inhibits peptidyltransferase of the mitochondrial ribosome
11. cycloheximide -inhibits peptidyltransferase
12. diptheria toxin - inhibits factor 2 and translocation