Peter T. Daley-Yates

Assay of 1-hydroxy-3-aminopropylidene-1,1-bisphosphonate and related bisphosphonates in human urine and plasma by high-performance ion chromatography

A method is described for the analysis of 1-hydroxy-3-aminopropylidene-1, 1-bisphosphonate and related bisphosphonates in human urine and plasma. Samples are spiked with 1-hydroxy-5-aminopentylidene-1,1-bisphosphonate as an internal standard and calcium chloride is added to precipitate the bisphosphonates. Following centrifugation the precipitate is redissolved in acetic acid, and the bisphosphonates are separated by high-performance ion chromatography on a Dionex AS7 column using nitric acid as mobile phase. The bisphosphonates are oxidised to orthophosphate using post-column addition of ammonium persulphate and this is followed by post-column reaction with molybdenum-ascorbate to yield the phosphomolybdate chromophore which is detected at 820 nm. A detection limit of 10 ng/ml is possible.

The clearance and bioavailability of pamidronate in patients with breast cancer and bone metastases

SummaryThe pharmacokinetics and bioavailability of pamidronate were assessed in patients with breast cancer, 6 subjects received the drug intravenously and 7 orally. The initial plasma half-life of pamidronate was short (42±27 min) and the apparent total plasma clearance was high (471±298 ml/min). The renal clearance (74±34 ml/min) was similar to the creatinine clearance (66±19 ml/min). Most of the renal elimination occurred during and immediately post a 4 h infusion of the drug (23.2±7.9% in 24h). The non-renal clearance was ascribed to uptake by bone and deep tissue compartments. Little additional drug appeared in the urine after 24 h. The mean bioavailability was estimated using a parallel study design to be 0.3% for a 300 mg oral dose.

Disposition and nephrotoxicity of 3-amino-1-hydroxypropylidene-1, 1-bisphosphonate (APD), in rats and mice

The purpose of this study was to investigate the disposition and the nephrotoxicity of 3-amino-1-hydroxypropylidene-1, 1-bisphosphonate (APD-pamidronate) in order to elucidate the mechanism of the non-linearity of the renal elimination of this drug. The fate of APD labelled with [14C]APD was studied in mice and rats for a range of doses (0.5-40 mg/kg) and indicators of renal function were monitored. In both species, the percentage of dose excreted during the first 24-h after treatment fell dramatically as a function of the dose. However, the renal burden of APD rose linearly for doses of APD below 10 mg/kg and increased non-linearly over this threshold. In contrast the concentration of APD in both bone and liver, which together account for a large proportion of the dose, appeared to increase proportionally with dose. There was no evidence, therefore, that the non-linear renal elimination of APD was due to an increased uptake of APD by tissues. Conversely, the significant fall in the renal excretion of APD was paralleled by a striking loss in body weight, and for high doses, by a fall in the creatinine clearance. An increased enzymuria suggested the loss of brush border membranes and the release of lysosomal contents by proximal tubular cells. Morphological studies confirmed this and revealed a focal proximal tubular necrosis 6 days post dosing. We conclude that the nephrotoxicity of APD accounts for the non-linear renal elimination of this drug.

The pharmacokinetics and distribution of pamidronate for a range of doses in the mouse

SummaryThe pharmacokinetics of the bisphosphonate drug pamidronate (APD, 3-amino-1-hydroxypropylidene-1,1-bisphosphonate) have been investigated in the mouse by using 14C-APD and following the tissue concentrations for up to 90 days postdose. The accumulation of APD in bone was the highest of all tissues and was linear with increasing dose up to the maximum dose employed (30 mg/kg), which is indicative of the uptake process being a simple chemical phenomenon. Despite the known effects of APD on bone turnover rates and osteoclast activity, the dose appeared to have no significant influence on the biological half-life of APD in bone which was found to be 90–140 days. A high dose of APD (5 mg/kg) appeared to prolong its uptake phase by bone, however, a net movement of APD from the soft tissues is the likely explanation for this finding. The concentrations of APD in the soft tissues investigated (liver, spleen, kidney, lung, and muscle) declined in a biphasic manner, initially in parallel with the fall in the plasma concentration and followed by a gradual fall in APDs concentration in the soft tissues due to renal elimination and a redistribution favoring the calcified tissues. The liver and spleen contained higher concentrations of APD relative to the other soft tissues. The 0–24 hour renal excretion of APD was found to fall with increasing dose above 2.5 mg/kg; this may be due to either nephrotoxicity or increased uptake by soft tissues. For doses over 20 mg/kg, there was some evidence of nephrotoxicity. The data from these studies have been used to formulate a simple physiological model for APD disposition.

A comparison of the pharmacokinetics of 14C-labelled APD and 99mTc-labelled APD in the mouse

The disposition of 14C-labelled and 99mTc-labelled APD has been investigated in the Balb/c mouse following a single ip injection containing APD (total dose 5 mg/kg) labelled partly with carbon-14 and partly with technetium-99m. The plasma clearance of carbon-14 was twice that for technetium-99m and the accumulation of carbon-14 in bone was twice as great as that of technetium-99m. All other tissues assayed (liver, lung, kidney and muscle) contained higher concentrations of technetium-99m than carbon-14. There was a particularly marked accumulation of technetium-99m in the liver. We conclude that pharmacokinetic studies of diphosphonates cannot be conducted reliably with 99mTc-labelled compounds.

Prediction of the renal clearance of cimetidine using endogenous N -1-methylnicotinamide

We investigated the influence of the type rather than the degree of renal insufficiency on the renal clearance of drugs. Different models of site specific experimental renal failure (ERF) have been developed in the rat; proximal tubular necrosis, induced by cisplatin; papillary necrosis, induced by 2-bromoethylamine, and glomerulonephritis, induced by sodium aurothiomalate or by antiglomerular basement membrane antibody. Several parameters of kidney function were assessed: the clearance of inulin, PAH, and endogenous N-1-methylnicotinamide (NMN). Plasma BUN and creatinine concentrations, and the presence of proteinuria and glucosuria were also measured. Our results showed a nonparallel decrease in glomerular filtration rate (GFR)and tubular secretion as measured by the secretory clearance of endogenous NMN or by the secretory clearance of p-aminohippuric acid (PAH), that is incompatible with the “intact nephron hypothesis.” As a result, the renal clearance of cimetidine, a drug eliminated mainly by renal secretion, correlated better with the renal clearance of endogenous NMN than with the GFR.We conclude that (i) our models of ERF demonstrated the existence of glomerulo-tubular imbalance that is contrary to expectations based on the intact nephron hypothesis; (iii) the type of the renal disease has a direct influence on the renal clearance of cimetidine; (in) the clearance of endogenous NMN may be a valuable noninvasive test for assessing renal tubular secretion which could be useful in predicting the clearance of drugs eliminated predominantly by tubular secretion.

Variability in the renal clearance of cephalexin in experimental renal failure

This study forms a part of an investigation into the extent to which the type of renal damage influences the renal clearance of drugs. We have already demonstrated an effect of different types of experimental renal failure (ERF) on the renal clearance of two cations: cimetidine, a drug that is filtered and secreted by the nephron, and lithium, which is filtered and reabsorbed by more than one segment of the nephron. In this report the renal clearance of cephalexin (CLCEX)is investigated, a drug that has a different mode of renal elimination, since it is filtered, secreted, and reabsorbed by the proximal tubules. The aim was to extend our earlier studies to an organic onion, and to provide an opportunity to evaluate the feasibility of using the renal clearance of N-1-methylnicotinamide (NMN) to predict the renal clearance of anionic drugs in renal failure. Different models of sitespecific ERF have been developed in the rat; proximal tubular necrosis (induced by cisplatin), papillary necrosis (induced by 2-bromoethylamine), andglomerulonephritis (induced by sodium aurothiomalate or by antiglomerular basement membrane antibodies). Glomerular function (GFR)was assessed by the clearance of inulin (CLNULIN),and tubular function was assessed by the clearance of endogenous NMN (CLNLM.Our results show that even if the models of ERF used were not absolutely site-specific, glomerular function and tubular function did not decrease to the same extent in the different ERF. Therefore, glomerulo-tubular imbalance existed, which is incompatible with the “intact nephron hypothesis, ” i.e., the site of the damage along the nephron and not only the degree of renal dysfunction, is a potential source of variability in the clearance of certain drugs. The renal clearance ofcephalexin was estimated more accurately by CLNMNthan GFR (r= 0.90). We conclude that the clearance of the endogenous cation NMN can be used to predict the renal clearance of drugs that are not only filtered by the glomeruli but also secreted and/or reabsorbed by the proximal tubules, and in the limited examples investigated appears to apply to both anionic and cationic compounds. In this respect the GFRalone was not an adequate parameter for the prediction of the renal clearance of such drugs.

Plasma protein binding of APD: Role of calcium and transferrin

The bisphosphonate drug APD (pamidronate, 3-amino-1-hydroxypropylidene-1,1-bisphosphonate) has been shown to bind to human plasma proteins. This was an unexpected observation since this hydrophilic, anionic drug is not typical of molecules that exhibit this characteristic. At a concentration of 5 micrograms/ml the extent of binding of APD to fresh human plasma in vitro was variable between subjects 30.2% +/- 8.5% (mean +/- S.D., n = 10). Binding was not influenced by the time or concentration of APD over the range 0.05-10.0 micrograms/ml. At 20 and 50 micrograms/ml some precipitation of APD occurred. Both calcium and iron play a role in the binding of APD to plasma proteins, addition of calcium to plasma increased the degree of binding of APD, whereas the calcium chelators EDTA and EGTA reduced the binding of APD. Similarly, addition of iron to plasma increased the binding and the inclusion of the iron chelator desferrioxamine diminished the binding of the drug. The effects of iron and desferrioxamine were less pronounced than those of calcium and EDTA, indicating that the majority of the binding involves calcium ions and a smaller contribution is made by ferric ions. The equilibrium dissociation constants (Kd) for APD binding to calcium and iron binding sites on plasma proteins were estimated to be 852 microM and 29 microM, respectively. Calcium binding sites were of high capacity but low affinity and the iron binding sites were of lower capacity and higher affinity. Electrophoresis of plasma proteins following incubation with [14C]APD revealed binding to the transferrin and globulin fractions. However, there was some dissociation of protein bound APD during the electrophoresis. The consequences of hypercalcaemia on the pharmacokinetics of APD are discussed.

Drug Pharmacokinetics in Renal Failure — Influence of Disease Type

For many drugs the major route of elimination from the body is renal excretion. Therefore, a loss of renal function will lead to excessive accumulation of the drug or its metabolites in the body, unless the drug dosage is reduced. The problem is particularly acute for drugs that have a narrow therapeutic index intended for patients with severe renal insufficiency. A failure to appropriately adjust dosage can lead to toxicity, which for some drugs may be life-threatening.