In vivo inhibition of adenosine deaminase by 2'-deoxycoformycin in mouse blood and leukemia L1210 cells

Abstract

Adenosine deaminase (ADA) activities in mouse whole blood, washed erythrocytes and L1210 cells were 0.48, 0.93 and 4.76 units/ml respectively. Methods were developed to determine the second-order association rate constant ( k 1) of a tight-binding ADA inhibitor, deoxycoformycin (DCF), and ADA in mouse blood and L1210 cells in vivo. After i.v. injection of DCF, the inhibition of the enzyme was of a monophasic pseudo-first-order nature in blood and biphasic (with an initial lag of 3–5 min) in L1210 cells. In contrast, i.p. injection of DCF produced the opposite pattern, monophasic in L1210 cells and biphasic in blood. The apparent k 1 values determined from the linear portions of these curves were compared with the k 1 values obtained in vitro. The mean k 1 values in vivo were: 4.2 × 10 4 and 1.4 × 10 4M −1 sec −1 in blood after i.v. and i.p. injections, respectively, and 2.6 × 10 3 and 2.2 × 10 4 M −1 sec −1 in L1210 cells after i.v. and i.p. injections respectively. The k 1 values with either whole blood or L1210 in vitro (3.1 × 10 4 and 5.5 × 10 3 M −1 sec −1, respectively) were of the same order of magnitude as those obtained with these tissues in vivo. In contrast, the k 1 values were about 150 to 1400-fold higher when either blood hemolysates (4.8 x 10 −6M −1 sec −1) or homogenized L1210 cells (7.5 x 10 6 −1 sec −1) were used. The 150 to 1400-fold higher k 1 values for blood hemolysates and homogenized L1210 cells than for intact cell samples (whole blood or whole L1210 ascitic fluid) suggest that the cell membrane plays a role in the interaction of DCF and ADA in these cell lines. The similarity of the rates of association of DCF and ADA in vivo and in vitro for mouse blood and ascites L1210 cells suggests that data obtained in vitro may be used to estimate the k 1 values in in vivo conditions.