Methionine tumor starvation by erythrocyte-encapsulated methionine gamma-lyase activity controlled with per os vitamin B6.

Erymet is a new therapy resulting from the encapsulation of a methionine gamma-lyase (MGL; EC number 4.4.1.11) in red blood cells (RBC). The aim of this study was to evaluate erymet potential efficacy in methionine (Met)-dependent cancers. We produced a highly purified MGL using a cGMP process, determined the pharmacokinetics/pharmacodynamics (PK/PD) properties of erymet in mice, and assessed its efficacy on tumor growth prevention. Cytotoxicity of purified MGL was tested in six cancer cell lines. CD1 mice were injected with single erymet product supplemented or not with vitamin B6 vitamer pyridoxine (PN; a precursor of PLP cofactor). NMRI nude mice were xenografted in the flank with U-87 MG-luc2 glioblastoma cells for tumor growth study following five intravenous (IV) injections of erymet with daily PN oral administration. Endpoints included efficacy and event-free survival (EFS). Finally, a repeated dose toxicity study of erymet combined with PN cofactor was conducted in CD1 mice. Recombinant MGL was cytotoxic on 4/6 cell lines tested. MGL half-life was increased from <24 h to 9-12 days when encapsulated in RBC. Conversion of PN into PLP by RBC was demonstrated. Combined erymet + PN treatment led to a sustained Met depletion in plasma for several days with a 85% reduction of tumor volume after 45 days following cells implantation, and a significant EFS prolongation for treated mice. Repeated injections in mice exhibited a very good tolerability with only minor impact on clinical state (piloerection, lean aspect) and a slight decrease in hemoglobin and triglyceride concentrations. This study demonstrated that encapsulation of methioninase inside erythrocyte greatly enhanced pharmacokinetics properties of the enzyme and is efficacy against tumor growth. The perspective on these results is the clinical evaluation of the erymet product in patients with Met starvation-sensitive tumors.

Bioavailability of Cu, Zn and Mn from Mineral Chelates or Blends of Inorganic Salts in Growing Turkeys Fed with Supplemental Riboflavin and/or Pyridoxine.

An 84-day feeding trial was conducted in growing turkeys to measure the bioavailability of Cu, Zn and Mn from a commercial mineral chelate and corresponding inorganic salts in composite feeds containing supplemental riboflavin (B2) and/or pyridoxine (B6). A total of 320, 28-day-old British United Turkeys (BUT) were assigned to eight dietary treatments in a 2 × 4 factorial arrangement comprising two trace mineral sources: chelated trace mineral blend (CTMB) and its corresponding inorganic trace minerals blend (ITMB) fed solely or with supplements of vitamin B2 (8 ppm) or B6 (7 ppm) or 8 ppm B2 + 7 ppm B6. Each treatment was replicated four times with 10 turkeys each. It was observed that turkeys fed with diets supplemented solely with ITMB elicited higher (P < 0.05) Zn excretion than their counterparts fed with diets containing ITMB with supplements of vitamins B2 and/or B6. Manganese retention was lower (P < 0.05) in turkeys fed with diets supplemented solely with ITMB than those fed with diets containing vitamins B2 and/or B6 additives. Combination of CTMB or ITMB with B6 improved (P < 0.05) the concentration of Mn in the liver and Cu in the bone. It was concluded that the minerals in CTMB were more available to the animals than ITMB. Furthermore, vitamins B2 and/or B6 supplementation improved the bioavailability of the inorganic Cu, Zn and Mn in growing turkeys and tended to reduce the concentration of these trace elements in birds' excreta.

CSF glutamate/GABA concentrations in pyridoxine-dependent seizures: etiology of pyridoxine-dependent seizures and the mechanisms of pyridoxine action in seizure control.

Several lines of evidence suggest that the binding affinity of glutamate decarboxylase (GAD) to the active form of pyridoxine is low in cases of pyridoxine-dependent seizures (PDS) and that a quantitative imbalance between excitatory (i.e. glutamate) and inhibitory (i.e. gamma-aminobutyric acid, GABA) neurotransmitters could cause refractory seizures. However, inconsistent findings with GAD insufficiency have been reported in PDS. We report a case of PDS that is not accompanied by an elevated cerebrospinal fluid (CSF) glutamate concentration. Intravenous pyridoxine phosphate terminated generalized seizures which were otherwise refractory to conventional anti-epileptic medicines. No seizure occurred once oral pyridoxine (13.5 mg/kg per day) was started in combination with phenobarbital sodium (PB, 3.7 mg/kg per day). The electroencephalogram (EEG) normalized approximately 8 months after pyridoxine was started. The patient is gradually acquiring developmental milestones during the 15 months follow-up period. The CSF glutamate and GABA concentrations were determined on three separate occasions: (1) during status epilepticus; (2) during a seizure-free period with administration of pyridoxine and PB; and (3) 6 days after suspension of pyridoxine and PB and immediately before a convulsion. The CSF glutamate level was below the sensitivity of detection (<1.0 microM) on each of the three occasions; the CSF GABA level was within the normal range or moderately elevated. The CSF and serum concentrations of vitamin B6-related substances, before pyridoxine supplementation, were within the normal range. We suggest that (1) PDS is not a discrete disease of single etiology in that insufficient activation of GAD may not account for seizure susceptibility in all cases and (2) mechanism(s) of anti-convulsive effect of pyridoxine, at least in some cases, may be independent of GAD activation.

Vitamin B6 and hemodialysis: the impact of high-flux/high-efficiency dialysis and review of the literature.

High-flux/high-efficiency (HF/HE) dialysis is associated with improved clearance for larger molecules, which include a wide variety of middle molecules and water-soluble vitamins. Our study attempted to measure in vivo clearances of serum pyridoxal-5'-phosphate (PLP), the active metabolite of vitamin B6, on standard cuprophan versus cellulose triacetate HF/HE dialysis for patients maintained on 10 mg daily pyridoxine supplements. A longitudinal evaluation of PLP after 3 months on HF/HE dialysis was performed simultaneously. The average in vivo PLP clearance for six patients on standard hemodialysis increased by more than 50%, from 86 +/- 61.7 mL/min using a cuprophan membrane to 173 +/- 90.2 mL/min using a cellulose triacetate dialyzer, at average blood flows of 375 mL/min (P < 0.05). Levels of PLP decreased from a baseline of 50 +/- 13.8 ng/mL to 24 +/- 9.7 ng/mL (P < 0.05) after 3 months of HF/HE treatments; the levels returned to 45 +/- 6.4 ng/mL on resumption of standard dialysis treatments. Although not achieving statistical significance, the average hematocrit increased from 31.2% +/- 1.66% to 32.7% +/- 1.24% while on HF/HE dialysis without an increase in erythropoietin requirements. We conclude that HF/HE dialysis treatments can have a dramatic impact on vitamin B6 homeostasis. Further investigation to evaluate the effects of different membranes and reprocessing should be performed on more heterogeneous patient populations in whom compliance problems with diet and vitamin supplementation may exist. The increased clearance of vitamin B6 may have significantly more detrimental effects in these settings.

Effect of feeding various dietary vitamin B6 concentrations during gestation and lactation on vitamin B6 level in liver and carcass of rat dams.

The present investigation was designed to determine the vitamin B6 concentration in liver and carcass of rat dams fed various dietary vitamin B6 levels during gestation and lactation. Therefore, gravid female rats received 0.6, 3, 6, 18 or 180 mg vitamin B6 per kg diet. After parturition each group was divided into 2 groups of 8 dams each, which received then a diet with 3, respectively 6 mg/kg vitamin B6 during lactation. At the end of the experiment at day 14 of lactation weight gain and food consumption as well as liver and carcass weights did not differ within the groups. The present data clearly show that both in liver and carcass of lactating rats, there exists only a slight dose-response-relationship between the dietary vitamin B6 intake and the vitamin B6 concentration in body tissue. Moreover, liver reflects the various vitamin B6 supplies during gestation and lactation somewhat better than carcass. The distribution of the vitamins did not differ among the groups and was therefore independent of the vitamin B6 supply. The present findings, especially regarding the liver, elucidate, that an adequate vitamin B6 supply during lactation can not compensate for a lack of vitamin B6 during gestation and vice versa a high dose of vitamin B6 during gestation did not completely protect against a suboptimal vitamin B6 during lactation.

Pharmacokinetics of vitamin B6 supplements in humans.

The use of vitamin B6 supplements is widespread today. Doses used are often elevated far above the physiological range and reach levels up to 600-fold higher than recommended dietary allowances for healthy people. While the toxic effects caused by chronic high doses of vitamin B6 have been described earlier, pharmacokinetic data on vitamin B6 supplements are rare. This article reviews the pharmacokinetic data of vitamin B6 from human subjects.

Vitamin B6 metabolism by human liver.

The B6 vitamers (pyridoxine, pyridoxamine, and pyridoxal) are primarily metabolized in liver to pyridoxal 5'-phosphate (PLP) and the deadend catabolite 4-pyridoxic acid. We have built on the elegant early work of Snell and others to describe the activities of the human liver enzymes responsible for vitamin B6 metabolism and to develop a model of the relative rates of these interconversions in vivo. This model is consistent with changes in plasma B6 after a load, the clearance of different vitamers (e.g., pyridoxine versus pyridoxal), and with the low plasma PLP in patients with cirrhosis. Because cirrhotics were found to be capable of PLP synthesis, we have used oral supplementation with pyridoxine to restore plasma PLP to the normal range, and have evaluated the effects of this intervention on amino acid metabolism. No significant differences were observed in plasma or urinary clearance of methionine (or cystathionine) after an oral load, nor in amino acid clearance from circulation after a protein load for cirrhotic patients before and after restoration of normal plasma PLP. Hence, the abnormal metabolism of vitamin B6 does not appear to be an important factor in the deranged amino acid metabolism in this disease. Nonetheless, this approach may be generally useful in assessing the importance of PLP in other abnormalities.

Plasma pyridoxal 5'-phosphate concentrations in obese and nonobese black women residing near Petersburg, VA.

The vitamin B-6 status of 15 obese and 15 nonobese black women aged 21-51 y who were not taking vitamin supplements was assessed by using plasma pyridoxal 5'-phosphate (PLP) measurements. Ages, heights, and ideal body weights of the two groups were similar as were reported energy, protein, and vitamin B-6 intakes obtained by using 24-h intake data collected on two nonconsecutive days separated by at least 1 wk. The reported vitamin B-6 intakes were 1.18 +/- 0.44 mg/d (means +/- SD). Plasma PLP levels in the obese and nonobese black women were similar; these levels were also similar to those observed previously for white obese and nonobese women having similar physical characteristics. All subjects had plasma PLP levels indicative of adequate status with one possible exception. Obesity did not affect the plasma PLP levels in these black women.

A simple liquid-chromatographic method for measuring vitamin B6 compounds in plasma.

This relatively simple high-performance liquid chromatographic (HPLC) method for measuring all seven known forms of vitamin B6 in plasma from individuals supplemented with pyridoxine hydrochloride shows good analytical recovery (85-98%) and precision. Within-run and between-run CVs for plasmas supplemented with standards were 4% and 7%, respectively. The major forms of B6 found in unsupplemented plasma from normal subjects were pyridoxal phosphate and 4-pyridoxic acid, with pyridoxal just detectable. The HPLC procedure correlated well (r = 0.94) with a modification of an enzymatic method involving apotryptophanase (Anal Biochem 1972;45:567-76) for measuring plasma pyridoxal phosphate, and also (r = 0.94) with a routine method for determining 4-pyridoxic acid in urine (Clin Chem 1964;10:479-89). Elimination of pyridoxine from the plasma of both normal and hyperoxaluric individuals was shown to be very rapid, with half-lives (t1/2) of 45 and 40 min, respectively. Finally, we present evidence for the existence of two other forms of B6 and discuss the possibility of a new metabolic pathway in vitamin B6 metabolism.

[Anticarcinogenic action of vitamins PP and B6 in the natulan initiation of malignant growth in mice]. / Antikantserogennoe deistvie vitaminov PP i B6 pri initsiatsii natulanom zlokachestvennogo rosta u myshei.

Parenteral administration of vitamins PP and B6 at the initiation stage of natulan-induced carcinogenesis was shown to significantly inhibit formation of lung adenomas. The preventive effect was found to depend on treatment schedule. Biochemical aspects of anticarcinogenic action of the vitamins require special investigation.

Pharmacokinetics of doxylamine, a component of Bendectin, in the rhesus monkey.

The elimination of doxylamine and metabolites was determined after iv administration of [14C]doxylamine succinate at 0.7 and 13.3 mg/kg to the adult female rhesus monkey. Although the total recovery of radioactivity was the same for the low- and high-dose studies (90.2%), the rate of plasma elimination of doxylamine and its demethylated metabolite (desmethyldoxylamine) was slower for the high dose group. The 24 hr urinary excretion of doxylamine metabolites, desmethyl- and didesmethyldoxylamine, was significantly increased and the polar doxylamine metabolites were significantly decreased as the iv doxylamine succinate dose was increased. The plasma elimination of gas chromatograph (GC)-detected doxylamine was determined after oral administration of Bendectin (doxylamine succinate and pyridoxine hydrochloride) at 7, 13.3, and 27 mg/kg to adult female rhesus monkeys. As the dose increased, the clearance of doxylamine decreased. A statistically evaluated fit of the oral data to a single-compartment, parallel first-order elimination model and a single-compartment, parallel first- and second-order (Michaelis-Menten) elimination model indicated that the more complex model containing the second-order process was most consistent with the observed elimination data.