Nucleic acid therapeutics

The discovery of ribozymes by Cech and Altman has fundamentally changed the view of the function of RNA in chemistry, biology, and medicine (1–3). Prior to this discovery, RNA had been viewed as a passive molecule that only carried information or provided structure to RNA-protein complexes. It is now clear that RNA can act as an enzyme and is capable of catalyzing RNA splicing and cleavage, as well as several other chemical reactions. These novel activities of RNA now permit the development of enzymatic RNA molecules as therapeutic agents that can suppress the expression of deleterious proteins by catalyzing the trans-cleavage of the corresponding mRNAs (4). RNA targets for ribozyme-based therapeutics may encode oncoproteins, growth factors, proinflammatory cytokines and their corresponding cell-surface receptors, and signal transduction molecules; viral and microbial mRNAs or genomic RNAs are also readily cleaved by this approach. RNA-cleaving ribozymes gain their target specificity from Watson-Crick base-pairing between the ribozyme’s binding-arm sequences and sequences that flank the cleavage site of the target RNA. Once bound, their mechanism of cleavage involves attack of the 2′-OH that is 5′ to the scissile bond in the target, thus destabilizing the target RNA’s phosphate backbone. Upon cleavage, the resultant products dissociate from the ribozyme complex and the ribozyme is released and may bind and cleave other targets again. The cleavage event renders the mRNA untranslatable and leads to further degradation of the target by cellular ribonucleases. With the recent completion of the sequencing of the human genome and several viral and bacterial pathogens, the number of potential ribozyme drug targets is enormous. This number will increase as the functions of all the genes in the human genome become clearer and the number of pathogens being sequenced increases. However, if they are to reach the clinic, ribozymes directed at these targets will need to be designed so that they can survive in nuclease-rich biological tissues and fluids, have a favorable pharmacokinetic (PK) profile, and prove effective in preclinical cell culture and animal models. Several groups have demonstrated the efficacy of ribozymes in cell culture and animal models. To date, only three ribozymes — one vector-expressed and two synthetic — have been reported in clinical trials. The first, a retrovirally expressed ribozyme that targets the HIV tat and rev exons, entered clinical testing in late 1996 and is currently in phase II testing for patients with AIDS-related lymphoma. The second, ANGIOZYME, is directed against the mRNA for Flt-1 (VEGF-R1), the high-affinity receptor for VEGF. This antiangiogenic ribozyme entered phase I trials in late 1998, phase I/II trials in late 1999, and is currently in phase II trials for several tumor types. The third, HEPTAZYME, a ribozyme targeting the 5′–untranslated region (5′-UTR) of the hepatitis C virus (HCV) RNA genome, has recently completed a phase I/II clinical trial in patients with chronic hepatitis C. This perspective will focus on nuclease-resistant, chemically synthesized ribozymes, as exemplified by ANGIOZYME and HEPTAZYME. These ribozymes can be readily synthesized, can be administered subcutaneously (s.c.) or intravenously (i.v.), have excellent specificity, are well tolerated, and function well in several in vitro and in vivo model systems. The clinical efficacy of these two ribozymes is being assessed currently. If they, or others in preclinical development, show a clinical effect, then the potential utility of nuclease-resistant ribozymes in medicine will be realized, with many more applications to follow.