New adenosine kinase inhibitors with oral antiinflammatory activity: synthesis and biological evaluation

Abstract

Several 5-iodotubercidin analogues in the pyrazolo[3,4-d]pyrimidine ring system were synthesized as potential inhibitors of adenosine kinase by a direct Lewis acid-catalyzed glycosylation procedure using both the preformed carbohydrate and the heterocyclic base as starting materials. The 5'-hydroxyl, -chloro, -azido, -deoxy, -amino, and -fluoro derivatives were prepared and evaluated in three systems for biological activity relative to adenosine, the true substrate, and 5-iodotubercidin, a known inhibitor. First, each compound was studied kinetically for inhibition of purified human placental adenosine kinase activity. The order of potency was: iodotubercidin > hydroxyl > amino > or = deoxy > fluoro > chloro >> azido. The Ki values for the 5'-hydroxyl and 5'-amino compounds, the two most potent inhibitors, were 80 and 150 nM, respectively. The inhibition appeared to be essentially competitive in nature, although a noncompetitive component of significance for the more potent inhibitors cannot be ruled out. Second, a bioassay was conducted in which the toxicity of 6-methylmercaptopurine riboside toward human CEM lymphoblasts was reversed by varying concentrations of the compounds. The order of effectiveness of the compounds in this system, representing a functional inhibition of adenosine kinase in cultured cells, was about the same as that with the purified enzyme, except that the 5'-chloro and 5'-fluoro compounds were ineffective. Third, the 5'-hydroxyl derivative was evaluated in vivo in a rat pleurisy inflammation model and displayed biological activity at a dose of 30 mg/kg given orally. Finally, the in vitro toxicity of each compound was assessed in CEM lymphoblasts. Results indicated that the two most potent inhibitors in the pyrazolo[3,4-d]pyrimidine ring system, the 5'-hydroxyl (7) and the 5'-amino (20), were 15-fold and 75-fold, respectively, less growth inhibitory than 5-iodotubercidin.

Adenosine Kinase: Exploitation for Therapeutic Gain

“…ADK inhibitor development was ini- Fig. 3) and 59-amino-59-deoxyadenosine as lead compounds (Cottam et al, 1993;Kowaluk et al, 1998;Wiesner et al, 1999), which were studied kinetically for inhibition of purified ADK activity (Cottam et al, 1993). A valuable strategy to modify and optimize existing lead molecules to improve their potency, bioavailability, or toxicity profile is based on fragmentation of existing leads and NMR-based screening of those fragments with the goal to identify suitable replacement of the fragments and incorporation of the newly identified fragments into the original scaffold (Hajduk et al, 2000).…”

“…3), which is a derivative of adenosine in which the 5-aza group of the purine ring has been replaced by a carbon that is linked to an iodine moiety; those compounds compete with adenosine for binding to the enzyme (Kowaluk et al, 1998;Kowaluk and Jarvis, 2000;McGaraughty et al, 2001bMcGaraughty et al, , 2005. Development of ADK inhibitors was initially based on the generation of 5-iodotubercidin analogs with modifications of the 59-group of the ribose moiety to include hydroxyl-, chloro-, azido-, deoxy-, amino-, or fluoro-groups in the 59-position; however, none of those compounds exceeded the potency of 5-iodotubercidin (Cottam et al, 1993). The ADK inhibitors 5-iodotubercidin, 59-amino-59-deoxyadenosine, 59-deoxy-5-iodotubercidin, as well as novel classes of ADK inhibitors such as 4-(N-phenylamino)-5-phenyl-7-(59-deoxyribofuranosyl)pyrrolo[2,3-day] pyrimidine (GP683), were shown to inhibit seizures in the maximal electroshock (MES) model in rats (Wiesner et al, 1999).…”

Structure–activity relationship for adenosine kinase from Mycobacterium tuberculosis

“…While it is known not to be a substrate for mammalian Ado kinases [40][41][42], there was strong evidence that 5′-amino-5′-deoxy-Ado (32) was a substrate for M. tuberculosis Ado kinase. With the M. tuberculosis enzyme, a peak appeared in the region consistent with the formation of a monophosphate, but the peak area did not increase linearly with time.…”

“…Therefore, we believe that the product is unstable under our assay conditions. The activity of 5′-amino-5′-deoxy-Ado (32) is noteworthy because this compound is known not to be a substrate for human Ado kinase; indeed, it is a potent inhibitor of Ado kinases [40][41][42][43]. The fact that the 5′-hydroxyl group is the site of catalysis for this enzyme makes this compound more intriguing than changes to other sites.…”

Identification and Characterization of a Unique Adenosine Kinase from Mycobacterium tuberculosis

“…Iodotubercidin is an Ado analog that is a potent competitive inhibitor of human AK, with a K i between 3 and 30 nM (8,22). Double-reciprocal plots of AK activity in the presence of iodotubercidin were constructed with five data points for each concentration of iodotubercidin (the regression coefficients were at least 0.94).…”