The CMF-regimen. Toxicity patterns following stepwise combinations of cyclophosphamide, methotrexate and fluorouracil.

The contribution of the agents used in the CMF regimen, i.e., cyclophosphamide (CY), methotrexate (MTX) and fluorouracil (FUra), to the development of toxicity was determined in tumor-bearing WAG/Rij rats. Data from untreated (U) rats were compared with data from rats treated with single-agent therapy (C-, M- and F-treatment groups), with data from double-agent therapy (CM-, MF- and CF-treatment groups) and with data from the triple combination: the CMF-treatment group. Doses of agents of interest were the same in all treatment groups. The sequence of administration was (1) CY; (2) MTX and (3) FUra which is similar to clinical treatment with CMF. Systemic levels of CY, MTX and FUra were comparable to those found in patients treated according to the CMF regimen. Toxicity was evaluated by body-weight changes, water and food consumption, white blood cell (WBC) and platelet cell (Pts) counts. With the exception of WBC and Pts nadirs, estimated toxicity parameters reflected toxicity over the whole treatment period of 14 days. The toxicity was generally mild and well tolerated, with one fatality in the M-treatment group. CY was the main contributor to toxicity; it caused both myelotoxicity and gastro-intestinal toxicity. The contribution of FUra was judged to be negligible. MTX + FUra did not increase host toxicity in a synergistic or even an additional fashion. The absence of addition or synergism of toxic side-effects can be explained both by site-specific interactions at the pharmacodynamic level and by interactions at the pharmacokinetic level.

The CMF-regimen. Modulation of cyclophosphamide uptake and clearance by methotrexate and fluorouracil.

Influence of the 2 antimetabolites used in the CMF-regimen, methotrexate (MTX, M) and fluorouracil (FUra, F) on in vivo pharmacokinetics of orally administered cyclophosphamide (CY, C), were studied in WAG/Rij rats. Blood plasma concentrations of CY following oral administration were monitored in single-agent CY, in CY + MTX (CM), in CY + FUra (CF) and in CY + MTX + FUra (CMF) treatments. Each treatment group consisted of at least 10 rats. CY was determined in 50 microliters of plasma by capillary gas chromatography on the first day of chemotherapy. Statistical analysis of blood plasma concentration data revealed a significant influence of both MTX and FUra on CY input/output function (p:0.01). MTX and FUra significantly increased the area under the plasma concentration time-curve, whereas tmax was significantly prolonged in CF and CMF treatment groups (p:0.01). It is suggested that MTX and FUra interact at the site of CY pre-systemic metabolism, including first-pass metabolism, subsequently resulting in prolonged absorption.

In vivo distribution of hydrocortisone over whole blood: a novel method for the extraction of erythrocytes.

Ten mg hydrocortisone (HC) was administered intravenously to a healthy volunteer after a dexamethasone suppression test and HC concentrations were determined from 1-270 min in plasma, plasma water and on erythrocytes. HC was extracted from erythrocyte concentrates with high efficiency by HC-poor plasma or by human or bovine albumin solutions. Determination of HC in the plasma of the volunteer mainly gave insight about the concentration time course of HC bound to plasma proteins. One minute after HC injection the amount associated with erythrocytes was about half the amount bound to plasma proteins. Decrease of HC in plasma, and hence on plasma proteins, was monophasic from 30-270 min with a half-life of 116 min. Decrease of HC associated with erythrocytes and HC free in plasma water was biphasic from 30-270 min and initially HC diminished about five times faster from these compartments than from plasma proteins. At the end of the observation period half-lives on plasma proteins, erythrocytes and in plasma water were similar, i.e., at 120 min. It is concluded from these as well as from previous in vitro experiments that erythrocytes gain importance as HC carriers at increasing HC blood concentrations. Once charged, erythrocytes yield HC much more readily than do plasma proteins. This "last come first go" phenomenon of association of HC with erythrocytes is known also to exist for certain drugs. It indicates erythrocytes as important transporters of non-freely in water soluble compounds.

Distribution of drugs over whole blood: I. The transport function of whole blood for valproate.

A method is presented for the determination of valproate (VPA) in approximately 40 microliters erythrocyte concentrations. Valproate partition between plasma proteins, ultrafiltrate, and erythrocytes at various concentrations was studied in vitro and in vivo. Partition ratios depended on VPA concentration in whole blood and the ratio of erythrocytes/ultrafiltrate, which increased with rising concentrations in the ultrafiltrate fraction. Shifts in ratios were studied by expanding the ultrafiltrate fraction of VPA-spiked blood. It appeared that VPA delivery to the expanded ultrafiltrate compartment originated in a disproportionately large part from erythrocytes. In vivo the half-life of VPA in the erythrocyte fraction was 0.7 h, whereas in the ultrafiltrate fraction and protein-bound fraction half-lives of 2 and 5 h were observed. It is concluded that for VPA in relation to erythrocytes a "last-come-first-go" system is plausible.

Plasma spermidine and putrescine as markers of cancer management.

The hypothesis was tested that dying tumor cells release spermidine and putrescine into the bloodstream. Therefore, a rhabdomyosarcoma was implanted in rats, irradiated and its growth and subsequent reduction compared with spermidine and putrescine plasma concentrations of the venous effluent from the tumor side and contralateral side. In the experimental set-up used the hypothesis could not be verified. It appeared that irradiation only, implantation and growth of a tumor only and an irradiated tumor, all caused elevated spermidine and putrescine concentrations in the effluent of both sides in approximately the same order of magnitude, compared to untreated controls.

Distribution of drugs over whole blood: II. The transport function of whole blood for phenytoin.

Phenytoin (DPH) partition between the three main blood compartments, i.e., plasma proteins, erythrocytes, and plasma water, was studied at various concentrations in vitro and in vivo. In vitro, the partition ratio of DPH in a system of erythrocytes in plasma water was 4.5 at concentrations between 0.8 and 100.8 micrograms DPH/ml. In vitro in whole blood (hence, in the presence of plasma proteins), this ratio was approximately 3.9. At 38 degrees C, blank erythrocytes were already in equilibrium with DPH-spiked plasma 3 min after contact, whereas at 20 degrees C, equilibration took 10 minutes or more. By adding blank ultrafiltrate to blood containing DPH, DPH concentrations of blood compartments shifted. It appeared that with the added blank ultrafiltrate, DPH was delivered overproportionally from erythrocytes and less from the protein fraction. In vivo, the elimination half-life of DPH in erythrocytes was 21.4 h and in plasma proteins 67.9 h. These results are similar to those obtained with valproate. It is concluded that erythrocytes have a low affinity for DPH. Their high-capacity transport system, having a "last-come-first-go" mechanism, plays a quantitatively important role in the transport of DPH.

Distribution of drugs over whole blood: III. The transport function of whole blood for hydrocortisone.

Hydrocortisone (HC) distribution over the three main components of blood, i.e., plasma water, plasma proteins, and erythrocytes, was studied in vitro at various HC concentrations in plasma, in a suspension of washed erythrocytes in plasma water, and in whole blood. The distribution ratio of HC in the system of erythrocytes in plasma water was 2.1 when HC, 0.18-10.8 micrograms/ml, was added. In whole blood, however, this ratio was 2.4 for the same concentration range. In the HC range of 0.18-0.68 micrograms/ml of whole blood, the uptake percentage of HC by erythrocytes increased from about 16 to 28% of the amount of HC added. By adding blank ultrafiltrate to HC-enriched blood, it appeared that the erythrocyte fraction released HC in overproportional quantities compared with the release by plasma proteins. Similar findings have previously been reported regarding the drugs valproic acid and phenytoin. Migration of HC from HC-spiked plasma to blank erythrocytes reached an equilibrium within 5 min.

Pharmacokinetics of antimicrobial drugs in cystic fibrosis. Aminoglycoside antibiotics.

Patients with cystic fibrosis (CF) show abnormal aminoglycoside pharmacokinetics. After a conventional dose, the serum concentrations in CF patients are lower than those in nonCF patients. The lower serum concentrations in CF might be explained by increased total body clearance and/or a larger volume of distribution. The therapeutic range of aminoglycosides is narrow due to oto- and nephrotoxicity. The changed pharmacokinetics and the narrow therapeutic range make it difficult to ensure that patients with CF are adequately and safely treated with aminoglycosides. The mode of administration of aminoglycosides influences the antibacterial effect of these agents on Pseudomonas aeruginosa and the development of possible side effects. The therapeutic implications of these facts are discussed.

The elimination of spermidine from plasma in rats.

Infused spermidine in rats with bypassed livers was eliminated very slowly, whereas endogenous spermidine in saline-infused controls with bypassed livers increased. In the same animals endogenous putrescine plasma levels did not differ significantly between spermidine- or saline-infused animals, whereas endogenous cadaverine levels increased during spermidine infusion and decreased afterwards. It is concluded that the liver is the most important site for plasma spermidine elimination.

Interactions of methotrexate and cyclophosphamide with the pharmacokinetics of 5-fluorouracil in an animal model.

The interaction of methotrexate and/or cyclophosphamide with the pharmacokinetics of 5-fluorouracil (5-FU) was studied in tumor-bearing WAG/Rij rats. Four groups were formed including treatment with single-agent 5-FU (eight rats); 5-FU plus methotrexate (11 rats); 5-FU plus cyclophosphamide (12 rats); and 5-FU, cyclophosphamide, and methotrexate (13 rats). The area-under-the-plasma-concentration/time curve, total-body clearance, elimination half-life, mean residence time, and steady-state volume of distribution were computed and compared. The mean residence time and elimination half-life of 5-FU increased when methotrexate was included in the combination. The increase was significant (P less than 0.05) for 5-FU, cyclophosphamide, and methotrexate versus 5-FU and cyclophosphamide.

The stability of 5,6-dihydrofluorouracil in plasma and the consequences for its analysis.

This study shows that 5,6-dihydrofluorouracil kept in plasma at room temperature deteriorates in time. Preservation is gained by low temperature and low pH of plasma samples. Therefore, plasma samples should be buffered and frozen immediately after being obtained, and thawing should proceed to 5 degrees C only.

Tobramycin in patients with cystic fibrosis. Adjustment in dosing interval for effective treatment.

The efficacy of the dosing regimen of tobramycin was investigated in 28 patients with cystic fibrosis who had an acute exacerbation of chronic pulmonary infection with Pseudomonas aeruginosa. The initial dose of tobramycin was 3.3 mg/kg of body weight three times daily (ie, 10 mg/kg/day). A highly significant relationship was found between the serum concentration of tobramycin before the dose and the change in the forced expiratory volume in one second (FEV1), both measured on the tenth day of treatment (rs = 0.75; p less than 0.001). In nine of the 16 patients who had a six-hour serum concentration of 1 mg/L or less on the tenth day of treatment, the eight-hour dosing interval of tobramycin was shortened to achieve a serum concentration of tobramycin of about 1 mg/L before the dose. In the other seven patients, the dosage of tobramycin was not changed. On the 20th day, seven of the nine patients in whom the dosing interval was shortened exhibited an increase in FEV1 of 20 percent or more. Such an increase was observed only in one of the seven patients in whom the dosing interval was not reduced (p less than 0.05). We conclude that individualizing the dosage of tobramycin in patients with cystic fibrosis results in a better clinical outcome.

pH modulates cisplatin toxicity.

Cisplatin in normal saline of pH 2.5 caused haemolysis of rat erythrocytes, whereas cisplatin in normal saline of pH 3.5 did not. Even a difference of 0.2 pH units appeared to be of significant importance: haemolysis of rat erythrocytes was observed with cisplatin in saline of pH 3.0 but not with cisplatin in saline of pH 3.2. The LD in mice was 15.4 mg/kg for cisplatin in saline of pH 2.5 versus 24.0 mg/kg for cisplatin in saline of pH 3.5. Experiments with cisplatin should include careful control of pH.

Pharmacokinetic interactions of cyclophosphamide and 5-fluorouracil with methotrexate in an animal model.

In tumor-bearing WAG/Rij rats the interaction of cyclophosphamide and/or 5-fluorouracil (5-FU) with methotrexate as manifested at the pharmacokinetic level was studied. Four groups were formed of at least ten animals. The control group, which received single-agent methotrexate, was compared with groups that received methotrexate plus cyclophosphamide, methotrexate plus 5-FU, and methotrexate plus cyclophosphamide plus 5-FU. There appeared to be an increase of 40% in the clearance of methotrexate by the triple combination. Cyclophosphamide especially diminished the terminal part of the concentration-time curve of methotrexate.

The influence of anaesthesia on plasma spermidine concentration and renal excretion in rats.

Anaesthesia with ether or a mixture of fentanyl and fluanisone diminished the clearance of infused spermidine from plasma in WAG/Rij rats considerably. The renal excretion of infused spermidine in unanaesthetized animals was only very small. The diminished plasma clearance of infused spermidine in anaesthetized animals therefore cannot be explained by loss of renal excretion.

Does determination of polyamines in body fluids require heating with hydrochloric acid?

In this study it is shown that hydrolysis of plasma and urine by heating with concentrated hydrochloric acid does not increase the recovery of spermidine from these body fluids.

Pharmacokinetics of tobramycin in patients with cystic fibrosis. Implications for the dosing interval.

The pharmacokinetics of tobramycin were evaluated in 15 patients (8 to 22 years of age) with cystic fibrosis (CF). A dose of 3.0 to 3.3 mg/kg of body weight was given intravenously over 20 minutes, and concentrations in serum were followed up to eight hours after initiation of the infusion. In the calculation of pharmacokinetic parameters, a two-compartment open model was used. The elimination half-life of the drug was highly inversely correlated with age (p less than 0.0004), and body weight (p less than 0.00002). Total body clearance (TBC), and volume of distribution at steady state (VDSS) were directly correlated with age and body weight. However, when TBC and VDSS were corrected for BSA, no correlation could be demonstrated. The mean one-hour and eight-hour serum concentrations of tobramycin were 5.40 and 0.45 microgram X ml-1, respectively. Between patients, considerable differences were found in the time after administration at which the serum concentration decreased below 1 microgram X ml-1. This interpatient variation has clinical implications for tobramycin therapy in CF, in particular for the dosing interval.