482 R. E. HANDSCHUMACHER AND A. D. WELCH 



The nutritional and enzymic studies described above, coupled with in- 

 formation concerning the effect of mercaptopurine on biosynthetic reactions 

 in vivo and in vitro, indicate that this antimetabolite has several sites of 

 action. What cannot be said with certainty is that these actions are neces- 

 sarily those which lead to inhibition of growth, or that the most critical 

 site of action has been disclosed. Formation of ribonucleotides of the natural 

 purines, particularly those of guanine and hypoxanthine, is competitively 

 inhibited by mercaptopurine and, thus, at sufficiently high levels of the 

 analog, utilization of preformed purines for the biosynthesis of nucleic acids 

 and coenzymes is restricted. 180, 185, 187, m Generally, incorporation of ade- 

 nine into nucleic acid or acid-soluble adenine derivatives is not affected ap- 

 preciably by mercaptopurine. 



Of much interest have been repeated observations indicating that mer- 

 captopurine blocks the synthesis de novo of purines; this was evidenced by 

 some of the earlier microbial growth experiments in defined media in the 

 absence of exogenous purines. 187 ' 168 The incorporation of formate 4, 191193 

 and of glycine 3 137 ' 143 into both purines of the acid-soluble fraction and 

 the nucleic acids of mammalian as well as microbial systems is depressed 

 by mercaptopurine. These results, however, do not conform with the ob- 

 servation that neither mercaptopurine nor its ribonucleotide inhibits the 

 synthesis of inosinic acid from glycine in soluble enzyme preparations from 

 pigeon liver. 186 The resolution of this apparent impasse may be found in a 

 preliminary report which shows that mercaptopurine ribonucleotide in- 

 hibits the conversion of inosinic acid to adenylosuccinate (the intermediate 

 in the formation of adenylic acid) in a cell-free extract of S. faecalis. 19 * Less 

 marked inhibition was also seen in the oxidation by an unidentified system 

 of inosinic acid to xanthylic acid, the presumed precursor of guanine ribo- 

 nucleotides. The same report confirmed earlier studies 172, 179 showing that a 

 resistant strain of this organism exhibited an impaired capacity for the for- 

 mation of the ribonucleotides of mercaptopurine, guanine, and hypoxan- 

 thine; however, it was suggested that a sufficient amount of the analog 

 ribonucleotide was formed to block the conversion of all the natural purines, 

 except xanthine, into ribonucleotides and their subsequent incorporation 

 into nucleic acids. 181 



These findings would imply that the apparent inhibition of the synthesis 

 de novo of purines by mercaptopurine may be related to interference with 

 the conversion of inosinic acid to the appropriate adenine and guanine 



191 R. W. Brockman, L. L. Bennett, Jr., and H. E. Skipper, Proc. Am. Assoc. Cancer 

 Research 2, 191 (1957). 



192 H. E. Skipper, Ann. N. Y. Acad. Set. 60, 315 (1954). 



193 H. G. Mandel, J. K. Inscoe, H. M. Maling, and P. K. Smith, ./. Pharmacol. Expil. 

 Therap. 120, 195 (1957). 



194 J. S. Salser and M. E. Balis, Federation Proc. 18, 314 (1959). 



