INHIBITION OF PLASMODIUM-FALCIPARUM PROLIFERATION IN-VITRO BY RIBOZYMES

Catalytic RNA (ribozymes) suppressed the growth of the human malarial parasite Plasmodium falciparum in vitro. The phosphorothioated hammerh ead ribozymes targeted unique regions of the P. falciparum carbamoylph osphate synthetase II gene. The P. falciparum carbamoyl-phosphate synt hetase II gene encodes the first and limiting enzyme in the pathway, a nd its mRNA transcript contains two large insert regions absent in oth er carbamoyl-phosphate synthetases, including that from humans, These inserts are ideal targets for nucleic acid therapy, Exogenous delivery of ribozymes to cultures reduced malarial viability up to 55% at 0.5 mu M ribozyme concentrations, which is significantly greater than cont rol levels (5-15% reduction), suggesting a sequence-specific inhibitio n, This inhibition was shown to be stage-specific, with optimal inhibi tions being detected after 24 h, coincident with maximal production of the carbamoyl-phosphate synthetase enzyme in the course of the life c ycle of the parasite, A decrease in total carbamoyl-phosphate syntheta se activity was observed only in cultures treated with the ribozymes. The task of developing alternative therapeutic agents against malaria is urgent due to the evolution of drug-resistant strains of P. falcipa rum, the most virulent of all human malarial parasites, Another critic al issue to be addressed is the possibility of eliminating or reducing any systemic toxicity to the host, which can potentially be provided by nucleic acid therapy, This work is the first reported assessment of the ability of ribozymes as antimalarials. Ribozyme inhibition assays can also aid in identifying important antimalarial loci for chemother apy The malarial parasite can, in turn, be a useful in vivo host to st udy the catalysis and function of new ribozyme designs.