The in vitro destruction of tumor cells by specifically sensitized mouse lymphocytes was inhibited by adenosine; this inhibition was markedly potentiated by the presence of an inhibitor of adenosine deaminase. The inhibition of cytolysis by adenosine was accompanied by a rapid elevation in lymphocytic adenosine 3',5'-monophosphate (cyclic AMP) concentrations. Both the inhibition of cytolysis and the elevation of cyclic AMP were reversed by prolonged incubation of the lymphocytes in the presence of adenosine or, more rapidly, by removal of the adenosine. Low concentrations of adenosine also caused an elevation of cyclic AMP in human lymphocytes, and this effect of adenosine may contribute to the lack of immune response associated with adenosine deaminase deficiency.
A variety of imidazo[4,5-c]pyridines (3-deazapurines) were synthesized. With use of these aglycons as pentosyl acceptors, the corresponding ribonucleosides and 2'-deoxyribonucleosides were prepared by an enzymatic method involving transfer of the pentosyl moiety from appropriate pyrimidine nucleosides. With most of the imidazo[4,5-c]pyridines, the products obtained from the enzyme-catalyzed reactions were pentosylated exclusively in the 1-position. However, some 3-pentosylation occurred with aglycons that had H or N3 in the 4-position. In addition to the 2'-deoxy congener of the ribonucleoside of 4-amino-1H-imidazo[4,5-c]pyridine, the 5'-deoxy and 2',5'-dideoxy congeners were synthesized. All of the aglycons and their nucleosides were tested for toxicity to mammalian cells in culture. None were markedly cytotoxic. These compounds were also evaluated for their ability to inhibit lymphocyte-mediated cytolysis in vitro. 3-Deazaadenosine (23) and its 2'-deoxy congener (38) were the most potent inhibitors (ED50 = 20 microM). In addition to these two in vitro tests, in vivo inhibition of the inflammatory response in the rat carregeenan pleurisy model was determined. 3-Deazaadenosine (23) was the most potent compound (ED50 = 3 mg/kg) in this in vivo test.
ABSTRACr 3-Deazaadenosine (deazaAdo) inhibits lymphocyte-mediated cytolysis in vitro at micromolar concentrations and is potentiated markedly in this activity by Lhomocysteine thiolactone. DeazaAdo alone causes a rapid, dose-dependent-buildup of S{[3HJadenosylhomocysteine (AdoHcy) and S{[3Hladenosylmethionine in cytolytic lymphocytes labeled with LJ2-3Hlmethionine; smaller amounts of S-3 I3H]deazaadenosylomocysteine (deazaAdoHcy) are also formed in these cells. Te simultaneous addition of deazaAdo and L-homocysteine thiolactone to the lymphocytes results in a massive intracellular accumulation of deazaAdo~cy. Both the inhibition of lymphocyte-mediated cytolysis and the cellular accumulation of 13HJdo cy caused by deazaAdo alone are reversed rapidly by removal of drug from the medium. However, the inhibition of cytolysis and the large cellular buildup of deazaAdoHcy resulting from treatmento the lymphocytes with deazaAdo plus L-homocysteine-thiolactone are dissipated more slowly under these same conditions. Unlike adenosine, deazaAdo is not potentiated in its inhibition of lymphocyte-mediated cytolysis by Ro 20-1724 4(3-butoxy-4-methoxybenzyl).2-imidazolidinonel an inhibitor of cyclic AMP phosphodiesterase, and has little or no effect upon the level of fymphocytic cyclic AMP. DeazaAdo is not metabolized detectably to 5'-nucleotides in the Iymphocytes and does not cause a decrease in the pool sizes of CTP, UTP, ATP, or GTP. Both AdoHcy and deazaAdoHcy have been reported to be powerful inhibitors of a variety of S-adenosylmethionine-utilizing methytansferases. The present results, therefore, indicate that the effect of deazaAdo upon lymphocyte-mediated cytolysis is due ultimately to the inhibition of an unidentified but crucial methyltransferase within the cytolytic lymphocytes and provide an insight into the biochemical processes involved in lymphocyte-mediated cytolysis. Among their many important functions in cellular immunity, sensitized thymus-derived lymphocytes can effect the specific lysis of antigen-bearing target cells; this phenomenon, lymphocyte-mediated cytolysis (LMC), may play a substantial role in tumor and graft rejection (for reviews see refs.
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