6-Thioguanosine Diphosphate and Triphosphate Levels in Red Blood Cells and Response to Azathioprine Therapy in Crohn’s Disease  Markus F. Neurath, Ralf Kiesslich, Ute Teichgräber, Christine Fischer, Ute Hofmann, Michel Eichelbaum, Peter R. Galle, Matthias Schwab  Clinical Gastroenterology and Hepatology  Volume 3, Issue 10, Pages 1007-1014 (October 2005) DOI: 10.1016/S1542-3565(05)00697-X Copyright © 2005 American Gastroenterological Association Terms and Conditions

Figure 1 Azathioprine and 6-MP metabolism.19,21,38 Azathioprine given orally is converted rapidly to 6-MP by nonenzymatic processes. 6-MP biotransformations occur along competing catabolic (XO, xanthin oxidase; TPMT) and anabolic (HPRT, hypoxanthine phosphoribosyltransferase) enzymatic pathways. 6-thioinosine 5′-monophosphate (6-TIMP) generated by the HPRT pathway may be transformed either into S-methyl-mercaptopurine ribonucleotide by methylation or into active 6-TGN via rate-limiting multiple enzymatic reactions involving inosine monophosphate dehydrogenase (IMPDH) and guanosine monophosphate synthetase (GMPS). Furthermore, 6-TG can be converted into 6-TGN via the HPRT pathway. Importantly, 6-TGN comprise a heterogeneous mixture of metabolites (blue) including TGMP, TGDP, and TGTP phosphates. 6-TGN also can be converted into the corresponding 2′-deoxy-analogues of TGDP and TGTP. Although the latter metabolites may be incorporated into DNA (2′-deoxy-6-TGTP) or have no described immunomodulatory function, 6-TGTP targets the Rac1 GTPase pathway that regulates T-cell apoptosis. 6-MTGN, methyl-thioguanine nucleotides; NDPK, nucleoside diphosphokinase. Clinical Gastroenterology and Hepatology 2005 3, 1007-1014DOI: (10.1016/S1542-3565(05)00697-X) Copyright © 2005 American Gastroenterological Association Terms and Conditions

Figure 2 Correlation between 6-TGN and (A) TGMP, (B) TGDP, and (C) TGTP levels in RBCs from 47 azathioprine-treated patients with Crohn’s disease. Statistically significant correlations using the Spearman rank correlation test are indicated. 6-TGN levels showed a significant positive correlation with TGDP and TGTP levels. (A) r = .17, P = .25; (B) r = .82, P < .0001; and (C) r = .88, P < .0001. Clinical Gastroenterology and Hepatology 2005 3, 1007-1014DOI: (10.1016/S1542-3565(05)00697-X) Copyright © 2005 American Gastroenterological Association Terms and Conditions

Figure 3 Correlation between 6-TGN and the sum of (A) TGDP and TGTP levels and (B) TGDP and TGTP levels in RBCs from 47 azathioprine-treated patients with Crohn’s disease. Statistically significant correlations using the Spearman-rank correlation test are indicated. 6-TGN levels showed a significant positive correlation with TGDP plus TGTP levels. A weaker correlation was noted between TGDP and TGTP levels. (A) r = .9, P < .0001; (B) r = .83, P < .0001. Clinical Gastroenterology and Hepatology 2005 3, 1007-1014DOI: (10.1016/S1542-3565(05)00697-X) Copyright © 2005 American Gastroenterological Association Terms and Conditions

Figure 4 Correlation between (A) 6-TGN or (B) TGDP levels in RBCs from 47 Crohn’s disease patients with azathioprine dosage. Statistically significant correlations using the Spearman-rank correlation test are indicated. TGDP levels showed a significant positive correlation with the azathioprine dosage used. No significant correlation was noted between 6-TGN levels and azathioprine dosage. (A) r = .27, P = .07; (B) r = .38, P = .009. Clinical Gastroenterology and Hepatology 2005 3, 1007-1014DOI: (10.1016/S1542-3565(05)00697-X) Copyright © 2005 American Gastroenterological Association Terms and Conditions

Figure 5 Correlation between (A) TGTP or (B) the sum of TGTP and TGDP levels in RBCs from 47 Crohn’s disease patients with azathioprine dosage. Statistically significant correlations using the Spearman-rank correlation test are indicated. No significant correlations were noted between TGTP levels and the azathioprine dosage and TGDP plus TGTP levels and the azathioprine dosage. (A) r = .21, P = .16; (B) r = .24, P = .11. Clinical Gastroenterology and Hepatology 2005 3, 1007-1014DOI: (10.1016/S1542-3565(05)00697-X) Copyright © 2005 American Gastroenterological Association Terms and Conditions

Figure 6 Hypothetical model of predicting azathioprine responsiveness in Crohn’s disease. In the presence of high 6-TGN levels, patients with high levels of active, Rac1-binding TGTP and low levels of TGDP are more likely to respond than patients with low levels of TGTP and high levels of inactive TGDP. Clinical Gastroenterology and Hepatology 2005 3, 1007-1014DOI: (10.1016/S1542-3565(05)00697-X) Copyright © 2005 American Gastroenterological Association Terms and Conditions