DRUG-NUCLEIC ACID INTERACTIONS

Abstract

As our knowledge of the action of antibiotics has increased, it has become clear that many of them attack on specific molecular target and there has been concentration on tracking down these molecules and investigating the nature of their interaction with drugs. A large number of antibiotics exert their effect primarily by interacting directly with the genetic material of cells i.e. with DNA. In doing so, these compounds impair the ability of DNA to act as a template for the processes of nucleic acid expression and synthesis. Daunomycin is an anticancer antibiotic, isolated from Streptomyces peuc&tius; active mainly against acute lymphocytic leukemia. It inhibits in vitro growth of both normal and cancer cell lines and as a consequence nuclear damage is characteristically observed. Chromosomal and genetic abberation of several types are also produced by daunomycin. Actinomycin D is an antibiotic metabolite containing two identical cyclic pentapeptide chains attached to a phenoxazone chromophore. It is very potent antitumor agent that inhibits DNA directed RNA synthesis and has found clinical applications in the treatment of chloriocarcinoma, but the extreme toxicity of the drug has percluded it from general use. The present study has been undertaken to find a new insight of

the conformational features of these two drugs in solution by using One dimensional and Two dimensional NMR techniques. The structure of d-GpG.daunomycin complex in stoichiometric ratio 1:2 CD/N) has been deduced by means of 2D NMR techniques and variable temperature one dimensional NMR experiments. One dimensional NMR has been used to find chemical shifts due to stacking, change in Tn, etc. while two dimensional NMR techniques; Correlation Spectroscopy <COSY> and nuclear Overhauser enhancement Spectroscopy CNOESY) serve as tools to assign all proton NMR signals unambiguously and determine conformational features such as sugar puckering, helix sense, glycosidic bond rotation and interproton distances in case of oligonucleotides. Rigorous theoretical potential energy calculations on the complexes of these two drugs with various nucleic acid bases, base pairs and dinucleotide model systems have been carried out to investigate various forces and their magnitude involved in complex formation and with a view to find base/base sequence specificity at the site of interaction. Classical potential function has been used to estimate conformational energy. Total interaction energy calculated is a sum of «l«ot.ro«t«tic, di«P~«ion, polarisation and repulsuon Ur^. Minimum energy conformation of the complex has been worked out in each case. The charge distribution on each molecule under study, has been calculated by using orbital method ofComplete Neglect Of Differential Overlap CCNDOX iii Our NMR results are indicative of presence of specific conformations of daunomycin and actinomycin D at NMR concentrations. Presence of more than one conformers of daunomycin molecule has been observed in solution. Complex of d-CpG and daunomycin exhibits interesting results as change in helical sense of d-CpG after complex formation. A B-Z transition is expected in d-CpG from native to complexed state and base to sugar orientation has been found to be SYN and not ANTI/hi^h ANTI as observed in d-CpG alone. In case of actinomycin D it has been observed that peptide chains do not experience any change on increase in temperature. Resonances from the protons of two separate chains have been successfully assigned Presence of an inverted dimer of actinomycin D in solution has been confirmed on the basis of nOe's seen in NOESY spectrum of the drug. Energy calculations have shown that both drugs,daunomycin as well as actinomycin D, prefer C,G containing binding sites. However, the specific sequence of preferential sites is being found different for two drugs. Theoretical calculations for probes into helix unwinding capacity for both drugs suggest value of Aa- unwinding angle- as around 10° and 28 for daunomycin and actinomycin D, respectively. These obsereved values of unwnding angles are well in accordance with that calculated and reported by other workers using X-ray and other theoretical methods.