Synthesis and biological activity of tubercidin analogs of ppp5'A2'p(5'A2'p)n5'A

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Abstract
A series of tubercidin (7-deazaadenosine) analogs of 2-5A of the general formula p5''(c7A)2''p[5''(c7A)2''p]n5''(c7A) (n = 0-5) were prepared by Pb ion catalyzed polymerization of the 5''-phosphoroimidazolidate of tubercidin. Through the corresponding imidazolidates, these oligonucleotide 5''-monophosphates were converted to the 5''-triphosphates. All reported structures were corroborated by enzyme digestion and 1H or 31P NMR. When evaluated for its ability to bind to the 2-5A-dependent endonuclease of mouse L cells, the tubercidin analog of trimeric 2-5A, i.e., ppp5''(c7A)2''p5''(c7A) 2''p5''(c7A), and the corresponding tetramer were bound as effectively as 2-5A itself. Nonetheless, it and the corresponding tetramer, ppp5''-(c7A)2''p5''(c7A)2''p5''(c7A)2''p5''(c7A), failed to stimulate the 2-5A-dependent endonuclease as judged by its inability to inhibit translation in extracts of mouse L cells programmed with encephalomyocarditis virus RNA and to give rise to rRNA cleavage in the same cell system under conditions where 2-5A showed activity at 10-9 M. The trimer, ppp5''(c7A)2''p5''(c7A)2''p5''(c7A), was an antagonist of 2-5A action in the L cell extract. In the lysed rabbit reticulocyte system, both the trimeric and tetrameric tubercidin 2-5A analogs were bound to the 2-5A-dependent endonuclease as well as 2-5A, but in this case, the tetramer triphosphate, ppp5''(c7A)2''p5''(c7A)2''p5''(c7A)2''p5''(c7A), was just as potent an inhibitor of translation as 2-5A tetramer triphosphate. This inhibition was prevented by the established 2-5A antagonist p5''A2''p5''A2''p5''A. The tubercidin analogs of 2-5A also were bound to the endonuclease of Daudi lymphoblastoid cells albeit slightly less effectively than 2-5A. The purine N(7) moieties of 2-5A are not involved in binding to the 2-5A-dependent endonuclease but one or more are required for activation of the 2-5A-dependent endonuclease of mouse L cells. These results also serve to underscore the inherent differences between the mouse L cell enzyme and the rabbit reticulocyte RNase L.