Method and compounds for inhibition of ribonucleases

#1. Field of the Invention

#Embodiments of the present invention relate in general to compounds and methods that are useful in inhibiting enzymes that effectively catalyze the depolymerization of ribonucleic acid, such enzymes being commonly referred to as "ribonucleases". More particularly, embodiments of the present invention relate to low molecular weight inhibitors of ribonucleases.

#2. Description of Related Art

#In the course of the detailed description of certain preferred embodiments to follow, reference will be made to the attached drawings, in which,

#FIG. 1 is a photograph of an electrophoresis gel showing protection of tRNA by an inhibitor of the present invention.

#The principles of the present invention may be applied with particular advantage to obtain compounds and methods for inhibiting enzymes which are effective catalysts of the depolymerization of ribonucleic acid. These enzymes are commonly referred to as "ribonucleases" and include, among others, bovine pancreatic ribonuclease A ("RNase") and like ribonucleases. It is to be understood that the methods and compounds of the present invention may be applied to inhibit a broad range of mammalian ribonucleases in the pancreatic ribonuclease superfamily, including human ribonucleases, as will be more readily apparent based on the disclosure to follow, since every potent ribonuclease in the pancreatic ribonuclease superfamily, including human ribonucleases, is believed to have similar unvaried or conserved substrate binding sites or subsites. The methods and compounds of the present invention surprisingly and advantageously provide a greater inhibition of ribonucleases when compared to existing compounds or methods.

#The methods of the present invention include contacting a ribonuclease with conserved an inhibiting amount of a small molecule compound of the formula I ##STR2## wherein P.sub.1 or P.sub.2 is a phosphate group, P.sub.3 is a phosphate or phosphorothioate group, R.sub.1 is hydrogen or is a substituent having a structure which is capable of occupying the B.sub.1 pyrimidine binding site of a ribonuclease, R.sub.2 is hydrogen or hydroxyl, and R.sub.3 is a substituent having a structure which is capable of occupying the B.sub.2 purine binding site of a ribonuclease. In a certain preferred embodiment, the chirality of the ribose group as shown in formula I (wherein R.sub.3 is attached to the 1' carbon) at the 1' through 4' carbons is R, R, S, R, respectively.

#In certain preferred embodiments, R.sub.1 is a nucleoside or 5'-phospho-nucleotide, preferably a pyrimidine nucleoside or pyrimidine 5'-phospho-nucleotide, more preferably 2'-deoxyuridine, 2'-deoxythymidine, or 2'-deoxycytidine and most preferably 5'-phospho-2'-deoxyuridine, 5'-phospho-2'-deoxythymidine, or 5'-phospho-2'-deoxycytidine or their phospohorothioate derivatives. R.sub.3 is preferably a purine, such as adenine or guanine. It is to be understood that the phosphate groups may be in the acid form or may have a cation attached. It is to be further understood that phosphorothioate derivatives are effective inhibitors and may be used instead of phosphate groups to avoid attack on the compound by phosphatases which may be present during use of the inhibitor.