Purine nucleoside phosphorylase as enzymatic activator of nucleoside prodrugs

Abstract

A process for inhibiting a mammalian cancerous cell or virally infected cell includes providing a Trichomonas vaginalis purine nucleoside phosphorylase enzyme or a tail mutant purine nucleoside phosphorylase enzyme in proximity to the mammalian cancerous cell or the virally infected cell and exposing the enzyme to a purine nucleoside phosphorylase enzyme cleavable substrate to yield a cytotoxic purine analog. The process includes introducing to the cell a vector containing the phosphorylase enzyme, or a DNA sequence coding for the same and delivering to the cell an effective amount of the substrate such as 9-(β-D-arabinofuranosyl)-2-fluoroadenine (F-araA).

1 . A process for inhibiting a mammalian cancerous cell or virally infected cell comprising:
 providing a  Trichomonas vaginalis  purine nucleoside phosphorylase enzyme in proximity to the cancerous mammalian cell or the virally infected cell; and   exposing the enzyme to a cleavable substrate of 9-(β-D-arabinofuranosyl)-2-fluoroadenine (fludarabine) to yield a cytotoxic purine analog.   
     
     
         2 . The process of  claim 1  wherein providing the enzyme is by administering a viral vector coding a nucleotide sequence for said enzyme expressible in said cell. 
     
     
         3 . The process of  claim 1  wherein providing said enzyme is by direct injection, infection, lipofection, or biolistic administration of a nucleotide sequence for the enzyme expressible in the cell. 
     
     
         4 . The process of  claim 1  wherein providing said enzyme is by direct injection of the enzyme proximal to said cell. 
     
     
         5 . The process of  claim 1  wherein providing said enzyme is by administration to a subject or a subject cell modified to express  Trichomonas vaginalis  purine nucleoside phosphorylase. 
     
     
         6 . The process of  claim 1  wherein providing is by intracellular delivery of an expressible nucleotide sequence encoding said enzyme. 
     
     
         7 . A composition produced by the process of  claim 1  comprising:
 mammalian cancerous or virally infected cell lysate; 
   Trichomonas vaginalis  purine nucleoside phosphorylase enzyme; and 
 9-(β-D-arabinofuranosyl)-2-fluoroadenine. 
 
     
     
         8 . The composition of  claim 7  further comprising a viral protein. 
     
     
         9 . A commercial kit for inhibiting a mammalian cancerous cell or virally infected cell according to  claim 1  comprising:
 a vector containing an expressible nucleotide sequence coding for a  Trichomonas vaginalis  purine nucleoside phosphorylase enzyme; and 
 instructions for the introduction of said vector to the cell to express said  Trichomoas vaginalis  purine nucleoside phosphorylase enzyme followed by administration of a purine nucleoside phosphorylase enzyme cleavable substrate of 9-(β-D-arabinofuranosyl)-2-fluoroadenine (fludarabine) to yield a cytotoxic purine analog. 
 
     
     
         10 . The kit of  claim 9  wherein said vector is a retrovirus, adenovirus, herpes virus, measles virus, adeno-associated virus, or a vaculovirus.