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The antiviral activities of antileukemic drugs 1-beta-D-arabinofuranosylcytosine (Cytarabine; Ara-C), 2,2'-anhydro-1-beta-D-arabinofuranosylcytosine (Ancitabine; Cyclo-C), and N4-behenoyl-1-beta-D-arabinofuranosylcytosine (Enocitabine; BH-AC) were evaluated in vitro against human cytomegalovirus (HCMV) in comparison with those of five other antiviral drugs. Both Ara-C and Cyclo-C showed the strongest inhibitory effect to HCMV. BH-AC inhibited the replication of HCMV and depicted almost as the same dose-response curve as Ganciclovir (DHPG). In the presence of Ara-C, Cyclo-C, or BH-AC, triphosphate forms of the nucleoside analogs were detected in the HCMV-infected cells, and synthesis of HCMV DNA was strongly suppressed. Thus, Ara-C, Cyclo-C, and BH-AC were not only antileukemic, but also antiviral in vitro. However, Ara-C and Cyclo-C may not be suitable as anti-HCMV agents, because they are cytotoxic or excreted rapidly in the urine in vivo [Van Voris, 1984; Hirayama et al., 1974]. Because of lower toxicity and longer retention in vivo, BH-AC may be expected as an anti-HCMV agent in patients with leukemia, in addition to serving as an antileukemic drug.

The basic biochemical characteristics of cyclocytidine hydrochloride (cC.HCl) and arabinosylcytosine (araC) were compared. It was demonstrated that despite different lipophilicity and different pK (4.15 for araC and 6.60 for cC.HCl), the mechanism of inhibition of DNA synthesis by both compounds is the same (ID50 for araC was 0.048 mumol/l and for cC. HCl 0.23 mumol/l). The compounds had a different mechanism of inhibition of RNA synthesis (ID50 for araC was 2.69 mmol/l and for cC.HCl 1.08 mmol/l) and showed a marginal effect on protein synthesis. Hydrolysis of the 0(2),2'-anhydro bond in cC.HCl and formation of araC in vivo was characterized by a Km = 280 mumol/l using HPLC. Deamination of araC in vivo was studied in healthy mice (Km = 247 mumol/l), 8.6% of arabinosyluracil 15 minutes after araC administration) and in mice with sensitive and araC resistant leukemia L1210 (15.5% and 8.5% of arabinosyluracil 15 minutes after araC administration, respectively). On the basis of different physico-chemical properties of cC.HCl and different mechanisms of inhibition of RNA synthesis it can be assumed that cC.HCl, when therapeutically used, may have its own mechanism of biological effect(s) and that its application may be therapeutically advantageous in some aspects as compared to araC.

Conversion rates of the prodrug ancitabine to the antileukemic cytarabine have been measured in vivo (rabbits) and in vitro (in the presence of rabbit blood and human red blood cells, blood, and plasma) using HPLC analyses for the prodrug, drug, and its inactive metabolite, 1-beta-D-arabinosyluracil. These observed pH-dependent in vitro rate constants were consistent with those for chemical hydrolysis determined from controls using Tris buffers. Hydrolysis of ancitabine to cytarabine is chemically, not enzymatically, mediated. The blood concentration-time course for administered compound was described by a two-compartment open model following a rapid intravenous injection of prodrug, drug, or metabolite in each of three rabbits. The in vivo conversion rate constant (kc) following a rapid intravenous prodrug injection was estimated by simultaneous nonlinear regression of ancitabine and cytarabine blood concentration-time courses using equations for two-compartment prodrug and drug with all possible models describing potential conversion sites. The best fit was obtained for the case allowing simultaneous conversion of the prodrug in both central and peripheral compartments to the drug in the central compartment with a common value for kc. The resulting kc value (0.09 h-1, three rabbits) is similar to that for chemical hydrolysis (0.07 h-1) at 38.8 degrees C. Reasons why this agreement is regarded as fortuitous are discussed.

The rationale and history of the development of physiologically based pharmacokinetic models are briefly reviewed in this paper. The methods of model construction and the previous application of this type of model to anticancer drugs are discussed. Future research should be focused on the following areas: (1) interspecies scaling, (2) the effects of disease states on the pharmacokinetics of anticancer drugs, and (3) the applications of pharmocokinetics to the studies of growth behavior of cancer cells. The ultimate goal will be to utilize this basic information to design an optimal dosage regimen and treatment schedule for the safe and effective cancer chemotherapy of each individual patient.

2,2'-Anhydro-1-beta-D-arabinofuranosyl-5-fluorocytosine (anhydro-ara-FC) was compared with cytosine arabinoside (ara-C) in the treatment of ic implanted murine Glioma 261. Both drugs given in ip doses of 500 mg/kg immediately inhibited the uptake and incorporation of tritiated thymidine into the DNA of brain tumor, small intestine, and spleen. Inhibition of DNA synthesis in the tumor recovered within 12 hours of anhydro-ara-FC administration, yet it remained depressed greater than 50% of control 12 hours after ara-C administration. Inhibition in the small intestine recovered within 24 hours of drug administration with either agent while inhibition in the spleen remained depressed greater than 24 hours. Anhydro-ara-FC administered ip in single doses less than or equal to 1500 mg/kg or in multiple doses less than or equal to 200 mg/kg three times a week for ten doses failed to prolong the survival of tumor-bearing mice, and minimal increased survival followed drug administration of 200 mg/kg every 24 hours for five doses. In contrast, ara-C in doses of 50, 100, or 200 mg/kg three times weekly for ten doses significantly increased the survival of tumor-bearing animals between 17% and 36%.

The kinetics of conversion of the prodrug ancitabine to the anticancer drug cytarabine have been studied in aqueous solutions in the pH range of 1.5-10.7, temperature range of 19.5-80.0 degrees C, ionic strength range of 10(-4) to 1.5, and in the presence of several general-base catalysts. Under all conditions ancitabine was quantitatively converted to cytarabine. The pH-rate profiles were linear with slope = 1 in alkaline pH, becoming pH independent in the region of maximum stability at pH less than or equal to 4, where buffer catalysis was found to be insignificant and kobs approximately equal to (1.12 X 10(11) h-1)-exp [-10121 deg/T]. At 30 degrees C, pH less than or equal to 4, it is calculated that an aqueous ancitabine solution will maintain 90% of its initial concentration for 12 d. A novel method for measuring general-base catalysis in competition with predominating specific-base catalysis and in the presence of secondary salt effects at constant ionic strength was developed. Three mechanisms of hydrolytic prodrug conversion are proposed: nucleophilic hydroxide addition, general base-assisted nucleophilic water attack, and spontaneous water attack.

We prepared the 5'- and 3'-O-phosphorothioate esters of the antitumor agent O2 : 2'-anhydro-1-beta-D-arabinosylcytosine. We also included in this study esters of 2'-thio-2'-deoxycytidine, namely, 2'-S-dCyd-2' : 3'-P, 2'-S-dCyd-2'-P, and 2'-S-dCyd-3'-P, along with natural nucleotides. These compounds were subjected to the action of Escherichia coli alkaline phosphatase, potato acid phosphatase, and bovine pancreatic ribonuclease A. The data were analyzed by Lineweaver-Burk plots to obtain Km and KI values. Only 2'-S-dCyd-2'-P was a substrate for alkaline phosphatase; the anhydro-araCyt phosphorothioates were good competitive inhibitors, while 2'-S-dCyd-3'-P did not associate with the enzyme. Acid phosphatase hydrolyzed all four monoesters investigated, including the S-phosphorothioate. The cyclic phosphorothioate, 2'-S-dCyd-2' : 3'-P was neither hydrolyzed by, nor associated with, ribonuclease A. ORD spectroscopy was also used in an attempt to relate the structural features of analogs to the peculiarity of their hydrolysis.

Ara-C should be converted to ara-CTP to inhibit DNA polymerase in the malignant cells but is rapidly inactivated to uracil arabinoside (ara-U) by cytidine deaminase in human tissue. Therefore, production as well as maintenance of ara-CTP in the cells is a function of both phosphorylation and deamination of ara-C, but is more dependent on the latter, because the deamination is several times superior to the former in terms of enzymatic activities. In chemotherapy with ara-C, the rate of the inactivation should be estimated for evaluating antitumor effect of the agent. Determination of serum or plasma deaminase activity can be a useful parameter of the inactivation. Attempts have been made to enhance the antitumor activity of ara-C by preventing deamination and a number of ara-C derivatives resistant to the deamination such as cyclocytidine, ara-C-5'-ester and acyl ara-C have been introduced. Cyclo-C gradually receives non-enzymatic hydrolysis to produce ara-C in neutral medium, which is useful for maintaining plasma ara-C level. Acyl ara-C such as behenoyl-ara-C (BHAC) is well incorporated into the cells and is highly distributed to lipophilic components such as membrane, microsome and mitochondria in the cells. The extremely gradual conversion of BHAC to ara-C in the cells is considered to be useful for maintaining effective intracellular concentration. A part of BHAC could be phosphorylated before deacylation. After intravenous administration of BHAC, the plasma drug concentrations are maintained significantly longer than those after the administration of the equivalent dose of ara-C. Therefore, BHAC is more resistant to the deamination than cyclo-C and the antitumor effect of the former is suspected to be milder but prolonged than that of ara-C or cyclo-C.

The pharmacokinetics of the drug cyclocytidine in humans were modeled by using a physiological and anatomical approach. Each pertinent tissue is represented by a single compartment, and these compartments are linked together by the circulatory system. Each compartment is then represented by an ordinary differential equation that represents the rate of change in drug concentration as function of convecting transport, metabolism, and urinary clearance. The models for cyclocytidine and cytarabine are linked together by a hydrolysis term in each equation set. The resulting equation sets are then solved numerically to predict the concentration of both drug species in situ. The models use physiological blood flows, tissue volumes, and clearance parameters. The results of the model show that cyclocytidine can act as a reservoir for cytarabine in vivo over the time studied. This effect is confined to relatively long times and relatively low plasma concentrations.

Since cyclocytidine (2,2'-anhydro-1-beta-arabinofuranosylcytosine hydrochloride) was introduced as an antineoplastic agent for the treatment of lymphatic leukemia, sinus acceleration and an increase in systemic blood pressure has been reported as its systemic effects in the clinical cases. These cardiovascular effects of cyclocytidine were observed also in anesthetized dogs, but not in reserpine-pretreated animals. Increases in heart rate and in systemic blood pressure were prevented by propranolol and phentolamine, respectively. The mechanism of these sympathomimetic effects was further analysed in the excised, blood-perfused canine sinoatrial node and papillary muscle preparations with a support dog. Positive chronotropic and inotropic responses to cyclocytidine were abolished by desipramine, propanolol, and pretreatment with reserpine but not by tetrodotoxin and hexamethonium. The tyramine-like actions of cyclocytidine at adrenergic neuronal terminals were discussed in conjunction with the uptake mechanism of the drug into the tumor cells.