Phase I clinical and pharmacokinetic trial of Brequinar sodium (DuP 785; NSC 368390).

Brequinar sodium is a 4-quinolinecarboxylic acid analogue that inhibits dihydroorotate dehydrogenase and subsequent de novo pyrimidine biosynthesis. It has shown dose-dependent antineoplastic activity against several mouse and human tumor models. This trial evaluated Brequinar given as a single daily i.v. bolus over a 5-day period repeated every 28 days. One hundred seven courses of treatment at dosages ranging from 36 to 300 mg/m2/day x 5 were administered to 45 patients (31 male and 14 female) with refractory solid tumors; median age was 58 years (range 30-74); median Southwest Oncology Group performance status was 1 (range, 0-3). Thirty patients had prior cytotoxic chemotherapy. Dose-limiting toxicities were thrombocytopenia and a severe desquamative maculopapular dermatitis. Two of 5 good risk patients at 300 mg/m2 and 3 of 6 poor risk patients at 170 mg/m2 developed a platelet count less than 25 x 10(3)/microliters. Two of 5 good risk patients at 300 mg/m2 and 1 of 6 poor risk patients at 170 mg/m2 developed a severe desquamative dermatitis. Moderate to severe mucositis was usually associated with the thrombocytopenia and/or the dermatitis. Nonhematological drug-related toxicities included nausea and vomiting, malaise, anorexia, diarrhea, phlebitis, reversible transaminase elevation, and mucositis. Other hematological toxicities were anemia, granulocytopenia, and leukopenia. There were no drug-related deaths. There were no objective tumor responses. Plasma and urine levels of Brequinar were quantified by high pressure liquid chromatography in 28 patients. Plasma levels and areas under the curve increased proportionally with increased dose. Brequinar had a harmonic mean terminal t1/2 of 8.1 +/- 3.6 h with a model-independent determined apparent volume of distribution at steady state of 9.0 +/- 2.9 liters/m2 and a total body clearance of 19.2 +/- 7.7 ml/min/m2. Renal excretion was a minor route of elimination for Brequinar. The maximally tolerated dose of Brequinar on a daily x 5 i.v. schedule was 250 mg/m2 for good risk patients. For the daily x 5 i.v. schedule, the recommended dose of Brequinar for phase II evaluation is 250 mg/m2 for good risk patients and 135 mg/m2 for poor risk patients.

Effect of the fatty acid oxidation inhibitor 2-tetradecylglycidic acid (TDGA) on glucose and fatty acid oxidation in isolated rat soleus muscle.

1. The effect of 2-tetradecylglycidic acid (TDGA), a potent, specific inhibitor of long-chain fatty acid oxidation, on fatty acid and glucose oxidation by isolated rat soleus muscle was studied. 2. TDGA inhibited [1-14C]palmitate oxidation by soleus muscle in a concentration-dependent manner. 3. TDGA inhibited the activity of soleus muscle mitochondrial carnitine palmitoyltransferase A (CPT-A). 4. Added palmitate (0.5 mM) significantly inhibited D-[U-14C]glucose oxidation and, under conditions where TDGA inhibited palmitate oxidation, the oxidation of D-[U-14C]glucose by isolated soleus muscle was significantly stimulated. 5. TDGA stimulation of glucose oxidation was reversed by octanoate, a medium-chain fatty acid whose oxidation is not inhibited by TDGA. 6. When nondiabetic rats were treated with TDGA (10 mg/kg p.o./day x 3 days), fasting plasma glucose was significantly lowered and the ability of isolated contralateral soleus muscles to oxidize palmitate was inhibited while glucose oxidation was significantly stimulated.

Inhibition of mitochondrial carnitine palmitoyl transferase A in vivo with methyl 2-tetradecylglycidate (methyl palmoxirate) and its relationship to ketonemia and glycemia.

The oral hypoglycemic agent, methyl 2-tetradecylglycidate (Me-TDGA), which inhibits in vitro mitochondrial carnitine palmitoyl transferase A (CPT-A) was used to study the relationship of CPT inhibition to changes in ketonemia and glycemia in normal and diabetic rats. After oral administration of Me-TDGA, the CPT activity of isolated rat liver mitochondria was substantially reduced with only the presumed outer enzyme fraction CPT-A released by digitonin treatment showing reduced activity. Mitochondrial fatty acyl-CoA synthetase was not inhibited. Oral doses of 0.1-2.5 mg/kg Me-TDGA produced both a dose-dependent lowering of plasma ketones and an inhibition of liver CPT. With single doses in excess of 2.5 mg/kg, po, heart and skeletal muscle CPT were also consistently inhibited. The effect on the liver enzyme persisted for at least 48 hr following 1 mg/kg, po, while the effect on ketones disappeared by 36 hr. The degree of inhibition of liver CPT produced by Me-TDGA was not altered by diabetes or the dietary state. At low doses (0.05-0.25 mg/kg, po), the most sensitive parameter was inhibition of hepatic CPT. Both plasma ketones and CPT were lowered with doses 10-fold less (0.1 mg/kg) than were required for blood glucose lowering, thus making Me-TDGA the most potent hypoketonemic compound known. In conclusion, inhibition of liver beta-oxidation at the stage of CPT-A by Me-TDGA can explain the potent hypoketonemic effects of this compound in fasted normal and diabetic rats. Higher acute doses are needed for both inhibition of muscle CPT and lowering of blood glucose.

Relation of oxidation of long-chain fatty acids to gluconeogenesis in the perfused liver of the guinea pig: effect of 2-tetradecylglycidic acid (McN-3802).

1. The potent, specific inhibitor of long-chain fatty acid oxidation, 2-tetradecylglycidic acid (McN-3802), at 10 microM totally abolished ketogenesis from endogenous substrates in the isolated perfused guinea pig liver. This effect was accompanied by a marked inhibition of gluconeogenesis from lactate plus pyruvate and by a shift toward a more oxidized state of the mitochondrial (3-hydroxybutyrate/acetoacetate ratio) and cytoplasmic (lactate/pyruvate ratio) compartments. 2. The addition of octanoate (88-500 microM) almost completely reversed the inhibitory effect of 2-tetradecylglycidic acid on gluconeogenesis. Octanoate oxidation, measured by the rate of ketogenesis, was not inhibited. This protective effect of octanoate against inhibition of gluconeogenesis by 2-tetradecylglycidic acid was seen even though in some experiments the mitochondrial redox state remained two to three times the magnitude observed prior to octanoate addition. 3. Gluconeogenesis from 4 mM glycerol was not inhibited and gluconeogenesis from 4 mM propionate was only slightly inhibited by 2-tetradecylglycidic acid. 4. Thus, it would appear that fatty acid oxidation in guinea pig liver is essential for maintaining maximal rates of gluconeogenesis, especially from substrates dependent on pyruvate carboxylation for conversion to glucose. 5. A single dose of 2-tetradecylglycidic acid (orally, 10-25 mg/kg) given to guinea pigs previously fasted 72 h produced a highly significant decrease of total ketones, of the 3-hydroxybutyrate/acetoacetate ratio and of plasma glucose. A smaller hypoglycemic effect was seen when the drug was administered to animals fasted for only 24 h or 48 h. 6. It appears from evidence in vivo and in vitro that the guinea pig and rat respond similarly to inhibition of fatty acid oxidation. This may be important since it has been suggested that the role of fatty acid oxidation in glucose synthesis is markedly different in these two species.

Common mechanism for the adaptive increase in hepatic ethanol and acetaldehyde metabolism due to chronic pretreatment with ethanol.

Perfused livers from ethanol pretreated rats utilized ethanol and acetaldehyde at higher rates than appropriate controls. This adaptive increase in hepatic ethanol and acetaldehyde uptake was associated with a marked (greater than 60%) increase in hepatic oxygen uptake. Ethanol uptake in both ethanol-treated and control livers was similarly sensitive to inhibition by 4-methylpyrazole, rotenone, and antimycin A. The adaptive increase in ethanol uptake was apparently specifically abolished by ouabain, an inhibitor of the sodium-plus potassium-activated ATPase. The data are consistent with the hypothesis that chronic treatment with ethanol increases ATPase activity. The ADP produced from these initiating events enters the mitochondrial space and stimulates electron transport and oxygen uptake. As a consequence of these events, a greater rate of NADH reoxidation occurs, resulting in a greater rate of production of NAD+ which stimulates ethanol oxidation via alcohol dehydrogenase and acetaldehyde oxidation via aldehyde dehydrogenase(s).

Significant pathways of hepatic ethanol metabolism.

Rat liver microsomes oxidized ethanol two to three times faster than propanol when incubated with either an NADPH- or an H2O2-generating system. In addition, solubilized, purified microsomal subfractions were found to contain protein with an electrophoretic mobility identical to rat liver catalase on SDS polyacrylamide gels, suggesting that the separation of catalase from cytochrome P-450 and other microsomal components may not be feasible. These data support the postulate that catalase is responsible for NADPH-dependent microsomal ethanol oxidation. Direct read-out techniques for pyridine nucleotides, the catalase-H2O2 complex, and cytochrome P-450 were utilized to evaluate the specificity of inhibitors of alcohol dehydrogenase (4-methylpyrazole; 4 mM) and catalase (aminotriazole; 1.0 g/kg) qualitatively in perfused rat livers. 4-Methylpyrazole and aminotriazole are specific inhibitors for alcohol dehydrogenase and catalase, respectively, under these conditions. Neither inhibitor nor a combination of them altered the mixed function oxygen of p-nitroanisole to p-nitrophenol as observed by oxygen uptake and product formation. When ethanol utilization was measured over the concentration range 20-80 mM in perfused liver, a concentration dependence was observed. At low concentrations of ethanol, ethanol oxidation was almost totally abolished by 4-methylpyrazole; however, the contribution of 4-methylpyrazole-insensitive ethanol uptake increased as a function of ethanol concentration. At 80 mM ethanol, ethanol utilization was nearly 50% methylpyrazole-insensitive. This portion of ethanol oxidation, however, was abolished by aminotriazole. The data indicate that alcohol dehydrogenase and catalase-H2O2 are responsible for hepatic ethanol oxidation. At low ethanol concentrations (less than 20 mM), alcohol dehydrogenase is predominant; however, at higher ethanol concentrations (up to 80 mM), the contribution of catalase-H2O2 to overall ethanol utilization is significant. No evidence that the endoplasmic reticulum is involved in ethanol metabolism in the perfused liver emerged from these studies.