Bioequivalence decision for nanoparticular iron complex drugs for parenteral administration based on their disposition.

Although parenteral iron products have been established to medicinal use decades before, their structure and pharmacokinetic properties are not fully characterized yet. With its' second reflection paper on intravenous iron-based nano-colloidal products (EMA/CHMP/SWP/620008/2012) the European Medicine Agency provided an extensive catalogue of methods for quality, non-clinical and pharmacokinetic studies for the comparison of nano-sized iron products to an originator (EMA, 2015). For iron distribution studies, the reflection paper assumed the use of rodents. In our tests, we used a turkey fetus model to investigate time dependent tissue concentrations in pharmacological and toxicological relevant tissues liver, heart and kidney. We found turkey embryos to be a suitable alternative to rodents with high discriminatory sensitivity. Clear differences were found between equimolar doses of iron products with hydroxyethyl amylopectin, sucrose, dextrane and carboxymaltose shell. A linear dose dependency for the tissue accumulation was also demonstrated.

Iron sucrose: assessing the similarity between the originator drug and its intended copies.

Iron sucrose (IS) is a complex nanocolloidal intravenous suspension used in the treatment of iron-deficiency anemia. Follow-on IS products (iron sucrose similars (ISSs)) have obtained marketing authorization by the generic pathway, implying that identical copies of IS may be manufactured. However, recent prospective and retrospective clinical studies showed discrepancies in clinical outcomes, which might be related to differences in physicochemical properties. The aim of this work is to measure and compare the physicochemical properties of IS and three ISSs available in the market using innovative analytical procedures. The comprehensive elucidation of size, size distribution, morphology, and stability of these complex drugs revealed very significant differences between the products. This study serves to provide the basis to define critical quality attributes that may be linked to differences in clinical outcome and thus may contribute to an adequate regulatory approach for IS and its follow-on products.

Complexity of intravenous iron nanoparticle formulations: implications for bioequivalence evaluation.

Intravenous iron formulations are a class of complex drugs that are commonly used to treat a wide variety of disease states associated with iron deficiency and anemia. Venofer® (iron-sucrose) is one of the most frequently used formulations, with more than 90% of dialysis patients in the United States receiving this formulation. Emerging data from global markets outside the United States, where many iron-sucrose similars or copies are available, have shown that these formulations may have safety and efficacy profiles that differ from the reference listed drug. This may be attributable to uncharacterized differences in physicochemical characteristics and/or differences in labile iron release. As bioequivalence evaluation guidance evolves, clinicians should be educated on these potential clinical issues before a switch to the generic formulation is made in the clinical setting.

Investigations of (99m)Tc-labeled glucarate as a SPECT radiotracer for non-small cell lung cancer (NSCLC) and potential tumor uptake mechanism.

This study attempted to evaluate the feasibility of (99m)Tc-labeled glucarate ((99m)Tc-GLA) imaging in non-small cell lung cancer (NSCLC) and the potential tumor uptake mechanism. Cell lysates from two NSCLC cell lines, H292 and H1975, were immunoblotted with anti-glucose transporter 5 (GLUT5) antibody for Western blotting. Thereafter, the two cell lines were used to examine cellular uptake of (99m)Tc-GLA with or without fructose. SPECT/CT imaging studies were performed on small animals bearing H292 and H1975 tumors. Biodistribution studies were also conducted to achieve accurate tissue uptake of this tracer in two tumor models. Hematoxylin & eosin (H&E) staining and GLUT5, Ki67 and cytokeratin-7 (CK-7) immunohistochemistry (IHC) analysis were further investigated on tumor tissues. In Western blotting, H292 cells showed higher levels of GLUT5 compared to the H1975 cells. Meanwhile, the in vitro cell assays indicated GLUT5-dependent uptake of (99m)Tc-GLA in H292 and H1975 cells. The fructose competition assays showed a significant decrease in (99m)Tc-GLA uptake by H292 and H1975 cells when fructose was added. The (99m)Tc-GLA accumulation was as much as two-fold higher in H292 implanted tumors than in H1975 implanted tumors. (99m)Tc-GLA exhibited rapid clearance pharmacokinetics and reasonable uptake in human NSCLC H292 (1.69±0.37 ID%/g) and H1975 (0.89±0.06 ID%/g) implanted tumors at 30min post injection. Finally, the expression of GLUT5, Ki67 and CK-7 on tumor tissues also exhibited positive correlation with the in vitro cell test results and in vivo SPECT/CT imaging results in xenograft tumors. Both in vitro and ex vivo studies demonstrated that the uptake of (99m)Tc-GLA in NSCLC is highly related to GLUT5 expression. Imaging and further IHC results support that (99m)Tc-GLA could be a promising SPECT imaging agent for NSCLC diagnosis and prognosis evaluation.

Hydroxyurea could be a good clinically relevant iron chelator.

Our previous study showed a reduction in serum ferritin of ß-thalassemia patients on hydroxyurea therapy. Here we aimed to evaluate the efficacy of hydroxyurea alone and in combination with most widely used iron chelators like deferiprone and deferasirox for reducing iron from experimentally iron overloaded mice. 70 BALB/c mice received intraperitonial injections of iron-sucrose. The mice were then divided into 8 groups and were orally given hydroxyurea, deferiprone or deferasirox alone and their combinations for 4 months. CBC, serum-ferritin, TBARS, sTfr and hepcidin were evaluated before and after iron overload and subsequently after 4 months of drug therapy. All animals were then killed. Iron staining of the heart and liver tissue was done using Perl's Prussian Blue stain. Dry weight of iron in the heart and liver was determined by atomic absorption spectrometry. Increased serum-ferritin, TBARS, hepcidin and dry weight of iron in the liver and heart showed a significant reduction in groups treated with iron chelators with maximum reduction in the group treated with a combination of deferiprone, deferasirox and hydroxyurea. Thus hydroxyurea proves its role in reducing iron from iron overloaded mice. The iron chelating effect of these drugs can also be increased if given in combination.

Biodistribution and predictive hepatic gene expression of intravenous iron sucrose.

INTRODUCTION: We have examined iron biodistribution and hepatic gene expression in rats following administration of the generic Iron Sucrose Azad (ISA) or the reference iron sucrose drug Venofer®. METHODS: ISA and Venofer® were administered intravenously to normal, non-anemic, male rats at 15 mg/kg (a supra-therapeutic dose-level). To evaluate biodistribution, tissue iron levels were determined over 28 days for plasma, liver, spleen, bone marrow, heart, kidney, lung and stomach using a validated ICP-MS method. Hepatic gene expression was evaluated by microarray analysis of mRNA from samples taken 24 h after drug administration. RESULTS: Iron concentration/time profiles for plasma and tissues were quantitatively similar for ISA and Venofer. Following administration, circulating iron levels briefly exceeded transferrin binding capacity and there was a transient increase in hepatic iron. Bone marrow iron levels remained elevated throughout the study. No increases in tissue iron levels were observed in the heart, stomach or lungs. Spleen iron levels increased over the course of the study in treated and control rats. Small, transient increases were recorded in the kidneys of treated rats. The effects of ISA and Venofer® on hepatic gene transcription were similar. Principal components analysis showed that there was no systematic effect of either treatment on transcriptional profiles. Only a small number of genes showed significant modulation of expression. No transcriptional pattern matches with toxicity pathways were found in the ToxFX database for either treatment. No modulation of key genes in apoptosis, inflammation or oxidative stress pathways was detected. DISCUSSION: These findings demonstrated that the biodistribution of administered iron is essentially similar for Iron Sucrose Azad and Venofer®, that iron sucrose partitions predominantly into the liver, spleen and bone marrow, and that hepatic gene expression studies did not provide any evidence of toxicity in animals treated at a supra-therapeutic dose-level.

Nanoparticle iron medicinal products - Requirements for approval of intended copies of non-biological complex drugs (NBCD) and the importance of clinical comparative studies.

Currently, most countries apply the standard generic approach for the approval of intended copies of originator nanoparticle iron medicinal products, requiring only demonstration of bioequivalence to a reference medicinal product by bioavailability studies. However, growing evidence suggests that this regulatory approach is not appropriate. Clinical and non-clinical studies have shown that intended copy preparations of nanoparticle iron medicinal products can differ substantially from the originator product in their efficacy and potentially in their safety profile. An adapted regulatory pathway (separate from the standard generic approach) with defined data requirements is needed for approval of intended copies of iron medicinal products. Here, we discuss the difficulties involved in assessing therapeutic equivalence of nanoparticle iron medicinal products and suggest key concepts of a regulatory approach. Standardized non-clinical comparative studies are necessary but, as demonstrated in the reported clinical data, they may not be sufficient to demonstrate a comparable efficacy and safety profile. Validated, prospective, comparative clinical studies might be needed, in addition to non-clinical studies, in order to enable appropriate assessment of therapeutic equivalence. Furthermore, including brand names in addition to the International Non-proprietary Names (INNs) in safety reports could enable effective safety monitoring of intended copies and originator products.

Evaluation of 99mTc-glucarate as a breast cancer imaging agent in a xenograft animal model.

INTRODUCTION: The use of [(99m)Tc]glucarate has been reported as an infarct-avid agent with the potential for very early detection of myocardial infarction. [(99m)Tc]Glucarate has also been postulated as an agent for non-invasive detection of tumors. The aim of our study was to develop a Glucarate kit and evaluate [(99m)Tc]glucarate as a potential cancer imaging agent in female SCID mice bearing human MDA-MB-435 breast tumors. METHODS: Glucarate in a kit formulation was labeled with (99m)Tc and evaluated for radiolabelling efficiency and radiochemical purity. The Glucarate kit stability was assessed by monthly quality controls. The pharmacokinetics of [(99m)Tc]glucarate were determined in female SCID mice bearing MDA-MB-435 human breast carcinoma tumors at 0.5, 1, 2, 4 and 24 h. Nuclear imaging studies were performed with a micro-single photon emission tomography (SPECT)/computed tomography (CT) system at 2 h post injection, while magnetic resonance imaging (MRI) was employed for tumor morphology analysis and metastatic deposit localization. RESULTS: The Glucarate kits exhibited a stable shelf life of 6 months. [(99m)Tc]Glucarate was obtained with radiochemical purity greater than 95%. Biodistribution studies demonstrated moderate tumor uptake coupled with high renal clearance. Tumor-to-muscle ratios were 4.85 and 5.14 at 1 and 4 h post injection. MRI analysis showed tumors with dense cellular growth and moderate central necrosis. [(99m)Tc]Glucarate uptake in the primary MDA-MB-435 shoulder tumors and metastatic lesions were clearly visualized with micro-SPECT/CT imaging. CONCLUSIONS: Selective tumor uptake and rapid clearance from nontarget organs makes [(99m)Tc]glucarate a potential agent for breast cancer imaging that awaits validation in a clinical trial.

MRP-1 protein expression and glutathione content of in vitro tumor cell lines derived from human glioma carcinoma U-87-MG do not interact with 99mTc-glucarate uptake.

The main cause of the multidrug resistance (MDR) of glioma cells is the overexpression of MRP-1, often associated with high levels of glutathione (GSH). We investigated whether MRP-1-related GSH content can influence (99m)Tc-glucarate entry by comparing its uptake with that of (99m)Tc-sestamibi (MIBI), an MRP- 1 probe, in an in vitro model of a sensitive cell line (U-87-MG) and a resistant derived cell line expressing MRP-1 (U-87-MG-R). Drug resistance was assessed by immunoblotting, GSH measurement, and Alamar Blue assay. To correlate MDR phenotype with tracer accumulation, uptakes were performed with and without modulators and after GSH depletion. Similar accumulation of (99m)Tc-glucarate was observed in the two cell lines, and the use of MDR reversals did not enhance its uptake. Our results clearly demonstrate that (99m)Tc-glucarate uptake is not related to MRP-1 expression or GSH levels. In contrast, (99m)Tc- MIBI accumulation is inversely proportional to the cell MDR phenotype. The combination of (99m)Tc-glucarate and (99m)Tc-MIBI may be a useful tool for the noninvasive detection of malignant sites and their chemoresistance status.

99mTc glucarate high-resolution imaging of drug sensitive and drug resistant human breast cancer xenografts in SCID mice.

BACKGROUND AND AIM: Previous studies have showed that 99mTc labelled glucarate (GLA) might be an agent for non-invasive detection of breast tumours. In xenografted BT20 breast tumours, GLA was found to have higher uptake than 99mTc sestamibi (MIBI). It is unclear whether GLA can localize in all cell line breast cancer xenografts, as well as breast tumours with multidrug resistance (MDR). The present study aimed to investigate the properties of GLA in detecting drug sensitive and drug resistant MCF7 breast cancer xenografts in mice by using dynamic single photon emission computed tomography (SPECT) imaging. METHODS: MCF7/S cells are drug sensitive breast carcinoma cells. MCF7/D40 cells are 40-fold more resistant to doxorubicin compared to MCF7/S. Subcutaneous tumours were grown in SCID mice for 10-14 days after injection of 1 x 10(6) cells into the right thigh. Anaesthetized mice with MCF7/S (MIBI, n=9; GLA, n=8) and MCF7/D40 (MIBI, n=6; GLA, n=5) tumours were imaged using a high-resolution SPECT system called FASTSPECT. Dynamic images were acquired for 2 h after intravenous injection of GLA or MIBI. Expression of MDR P-glycoprotein (Pgp) in the tumours was demonstrated in the MCF7/D40 tumours by western blotting, not in the MCF7/S tumours. RESULTS: The xenografted tumours were visualized unequivocally within 10-30 min in GLA images and remained detectable for at least 2 h after injection. Drug resistant tumours, from which MIBI was rapidly expelled, retained GLA as readily as did drug sensitive tumours. The biodistribution data of GLA demonstrated significantly higher accumulation (%ID/g) compared to MIBI. CONCLUSION: MCF7 tumour xenografts can be detected by 99mTc glucarate imaging. More importantly, 99mTc glucarate can potentially localize drug resistant breast tumours.

Accumulation of technetium-99m glucarate: in vitro cell cultures and in vivo tumour models.

99mTc-glucarate is an investigational radiopharmaceutical which has been shown to accumulate in acute cerebral and myocardial injuries and in some tumours. In the present work, a survey of possible factors affecting the cellular accumulation of 99mTc-glucarate was carried out in cell lines and strains in vitro and in murine tumours in vivo. Accumulation was enhanced under hypoxic conditions in 12 of the 16 human and murine cell lines and strains studied, and inhibited in the presence of nitroimidazoles. At temperatures lower than 37 degrees C, accumulation was reduced, but a hypoxic/aerobic differential was maintained. Aerobic accumulation of 99mTc-glucarate was enhanced by cyanide. In transplanted tumours in mice, 99mTc-glucarate showed high tumour/muscle and tumour/blood ratios at early times after injection. Pharmacological enhancement of the extent of hypoxia by the administration of hydralazine or nitro-L-arginine resulted in significantly increased accumulation of 99mTc-glucarate in the tumour. The in vitro and in vivo properties of 99mTc-glucarate suggest that it may be useful for tumour imaging in the clinic, although the exact mechanism(s) by which it localizes in tumours remains unknown.

Inhibition of liver metastases and tumor cell invasion in spontaneous liver metastasis model (LMFS) by sodium D-glucaro-delta-lactam (ND2001).

We examined the inhibitory effect of sodium D-glucaro-delta-lactam (sodium 5-amino-5-deoxy-D-glucosaccharic acid-delta-lactam: ND2001) upon liver metastases of the LMFS tumor. A permanent cell line, LMFS, was established from a spontaneously occurring murine retroperitoneal tumor of BALB/c mouse origin, and after a subcutaneous injection, the LMFS cells proliferated at the inoculation site (100% take) with liver metastases. ND2001 had little effect on the cell growth, cell cycle and phagokinesis of the LMFS cells in vitro. However, when the invasive activity was measured by the Boydem chamber method, the number of LMFS cells was reduced, with inhibition rates of 98.0%. After the LMFS cells treated with ND2001 in vitro, the numbers of hepatic metastases of subcutaneous inoculation of treated cells were reduced dose-dependently, and those of intravenous inoculation were not found by microscopical study. When the LMFS tumor-bearing mice were treated with ND2001 (0, 30, 100 mg/kg/d) from day 1, ND2001 (30 mg/kg) inhibited the liver metastases with a rate of 56.4%, and when given from day 15, ND2001 (100 mg/kg) inhibited with a rate of 47.5%. But ND2001 showed neither cytocidal nor anti-tumor activity. Combination therapy of primary tumor resection and ND2001 administration revealed that preoperative use of ND2001 was more effective in preventing liver metastases. These results suggested that ND2001 might have a potential use as an anti-metastatic agent for operative patients without metastasis.

Very early noninvasive detection of acute experimental nonreperfused myocardial infarction with 99mTc-labeled glucarate.

BACKGROUND: 99mTc glucarate has recently been reported to be an infarct-avid agent. The feasibility of imaging with 99mTc glucarate was evaluated for the early diagnosis of nonreperfused and reperfused myocardial infarction and compared with localization of simultaneously administered 111In anti-myosin. METHODS AND RESULTS: Four groups of six rabbits each were studied. The left anterior descending coronary artery (LAD) was kept persistently occluded (n = 6) or reperfused after 40 minutes (n = 6) in rabbits. After confirmation of LAD occlusion by 201Tl scintigraphy, a mixture of 99mTc glucarate (15.7 +/- 1.6 mCi) and 111In anti-myosin (0.53 +/- 0.03 mCi) was administered intravenously. Another group of rabbits (n = 6) with 5 or 15 minutes of LAD occlusion were used to assess the affinity of 99mTc glucarate for the ischemic myocardium. The remaining 6 rabbits with reperfused myocardial infarction were used for the assessment of subcellular localization of 99mTc glucarate. 99mTc glucarate cleared rapidly from circulation (elimination t1/2, 36 minutes). Infarcts were visualized within 10 minutes in reperfused and within 30 minutes in nonreperfused coronary territories after intravenous administration. 111In anti-myosin delineated reperfused infarcts within 1 to 3 hours, but no uptake was seen in persistently occluded rabbits. 99mTc glucarate uptake in reperfused and nonreperfused infarct centers was 28 and 12 times greater, respectively, than that in normal myocardium (P = .0001). A direct correlation between glucarate and anti-myosin localization (r = .60 for nonreperfused; 0.76 for reperfused; P < .0001) was observed. Ischemic hearts showed no glucarate uptake. Subcellularly, 99mTc glucarate localized predominantly in the nuclear fraction of the infarct, with lesser extents in the mitochondrial and cytoplasmic fractions. CONCLUSIONS: Noninvasive imaging of myocardial infarcts with 99mTc glucarate is possible within minutes in persistently occluded or reperfused myocardial infarcts. Early detectability results from the rapid blood clearance and high avidity of glucarate for the acutely necrotic myocardial tissue.

Targeting human breast tumour in xeno-grafted SCID mice with 99Tcm-glucarate.

99Tcm-glucarate accumulation in human mammary BT-20 tumours hosted in severe combined immune deficiency (SCID) mice was compared to 111In-monoclonal antibody 103D2-F(ab')2, 99Tcm-methoxyisobutyl isonitrile (99Tcm-MIBI) and 99Tcm-diethylenetriamine pentaacetate (99Tcm-DTPA). The intracellular tumour distribution of 99Tcm-glucarate was also determined. SCID mice injected with a mixture of 99Tcm-glucarate and 111In-103D2-F(ab')2 were imaged serially up until 24 h. Computer planimetered tumour-to-blood activity (in the heart) ratios (T/BH) to 8 h were significantly greater for 99Tcm-glucarate than 111In-103D2. The mean (+/-S.D.) tumour-to-blood ratio (T/B) from biodistribution was 1.21 +/- 0.31 and 0.35 +/- 0.06 (P < 0.0001) at 5 h, and 1.526 +/- 0.29 and 0.75 +/- 0.2 (P < 0.0001) at 8 h, for 99Tcm-glucarate and 111In-103D2 respectively. At 24 h, T/B for 111In-103D2 (1.76 +/- 0.22) exceeded that of 99Tcm-glucarate (1.44 +/- 0.2, P = 0.01). 99Tcm-glucarate uptake in the tumours at 5 h (1.133 +/- 0.25 %ID g-1) and 8 h (1.213 +/- 0.23 %ID g-1) was significantly greater than that of 99Tcm-MIBI (0.340 +/- 0.09, P = 0.0002; 0.220 +/- 0.04, P = 0.0001) and 99Tcm-DTPA (0.091 +/- 0.03, P < 0.0002; 0.016 +/- 0.01, P < 0.0001) respectively. Intracellular tumour distribution of 99Tcm-glucarate was 50.91 +/- 6.55% in the nuclear fraction, 34.34 +/- 2.88% in the cytoplasmic fraction and 14.75 +/- 7.66% in the mitochondrial fraction. Thus glucarate may provide a 99Tcm-based mammary tumour imaging modality for visualization of tumours very quickly after tracer administration with maximal targeting in the nuclei of the tumour cells.

Metabolism, uptake, and excretion of a D-glucaric acid salt and its potential use in cancer prevention.

D-Glucaric acid (GA) is a nontoxic, natural compound. One of its derivatives is the potent beta-glucuronidase inhibitor D-glucaro-1,4-lactone (1,4-GL). The goal of this study was to demonstrate the in vivo formation of 1,4-GL from a D-glucarate salt and determine its metabolism, uptake by selected organs, and excretion following oral administration of potassium hydrogen D-[14C]glucarate to male and female Sprague-Dawley rats. 1,4-GL increases detoxification of carcinogens and tumor promoters/progressors by inhibiting beta-glucuronidase and preventing hydrolysis of their glucuronides. 1,4-GL and its precursors, such as potassium hydrogen D-glucarate and calcium D-glucarate, may exert their anticancer action, in part, through alterations in steroidogenesis accompanied by changes in the hormonal environment and the proliferative status of the target organ. Thus, GA derivatives may be useful as new or adjuvant cancer preventive and therapeutic agents. In our study, 1,4-GL was found to be formed from the D-glucarate salt in the stomach of rats. It was apparently absorbed from the gastrointestinal tract, transported with the blood to different internal organs, and excreted in the urine and to a lesser extent in bile. There were no significant differences in the metabolism of PHG between male and female rats. Thus, formation of 1,4-GL from D-glucaric acid derivatives may be prerequisite for their inhibition of chemical carcinogenesis in rodents and prevention of breast, prostate, and colon cancer in humans.

Pharmacokinetics relevant to the anti-carcinogenic and anti-tumor activities of glucarate and the synergistic combination of glucarate:retinoid in the rat.

Alone and in synergistic combination with retinoids, dietary glucarate inhibits both the chemical induction and growth of rat mammary tumors. To investigate the pharmacokinetics of glucarate, [14C]glucarate was synthesized, converted to the calcium salt, and administered to rats bearing primary mammary tumors. When given by gavage, [14C]glucarate, as the calcium salt, showed a biphasic response in the blood. After peaking within 1 hr of administration at a level of 0.4 mumol/mL (normal endogenous level is approximately 0.04 mumol/mL), its plasma concentration dropped to 0.1 mumol/mL at 3 hr. In the second phase, there was a semilog increase to 0.6 mumol/mL at 15 hr, followed by a slow rise to 0.75 mumol/mL at 24 hr. Of the 38% of the administered glucarate that was recovered, 38% was excreted in the urine, and 30% remained in the gastrointestinal tract at 24 hr. Glucarate was concentrated 3- to 4-fold in the liver and intestinal mucosa, compared to the level in serum. With minor exception, the pharmacokinetics of [14C]13-cis-retinoic acid administered by gavage to rats was similar or not the semipurified diets were supplemented with 64 mmol/kg of calcium glucarate. During the interval between 5 and 10 hr post-administration of [14C]13-cis-retinoid, there was a transient 35-50% rise in the plasma level in rats on the glucarate-supplemented diet. This rise had no observable effect on the level of retinoid in major organs or in the tumor. A glucarate-binding protein was detected in the tumor cytosol. This potential receptor had a Ka of 1.49 x 10(7) M-1.

Determinants of urinary thioethers, D-glucaric acid and mutagenicity after exposure to polycyclic aromatic hydrocarbons assessed by air monitoring and measurement of 1-hydroxypyrene in urine: a cross-sectional study in workers of coke and graphite-electrode-producing plants.

A cross-sectional epidemiological study was performed on 286 workers from two coke oven and one graphite electrode plants. The aim was to evaluate the usefulness of monitoring 1-hydroxypyrene (1-HOP) in urine for assessing exposure to polycyclic aromatic hydrocarbons (PAHs), and that of the urinary excretion of thioethers and D-glucaric acid, and the mutagenic activity of urine as indicators or biological effects of PAHs. The results confirm that 1-HOP determination in urine probably reflects exposure to PAHs by all routes and is not significantly influenced by the smoking habit. In comparison with the total PAHs in the air and 1-hydroxypyrene in urine, taken as reference exposure parameters, the results indicate that urinary D-glucaric acid excretion is not positively influenced by PAHs exposure; thioethers determination in urine is of poor value, since the smoking habit is a strong confounding factor. The determination of urinary mutagenicity might contribute to the detection of groups of workers exposed to potentially genotoxic PAHs.

Effect of hypoxia on the accumulation of technetium-99m-glucarate and technetium-99m-gluconate by Chinese hamster ovary cells in vitro.

The accumulation of 99mTc-glucarate, an agent recently reported to localize in acutely infarcted myocardium, zones of acute cerebral injury and tumors, has been compared with 99mTc-gluconate in an in vitro system of cultured Chinese hamster ovary fibroblasts. The effects on accumulation of hypoxia and competition with fructose have been studied. Both labeled glucose analogs showed a two- to threefold enhanced accumulation in hypoxic cells relative to aerobic cells. No such enhanced accumulation under hypoxia was observed for the nonsugar tracers pertechnetate and 99mTc-DTPA. The presence of 20 mM fructose reduced the accumulation of 99mTc-glucarate by 30% (p = 0.067) and 99mTc-gluconate by 40% (p < 0.05) in hypoxic cells, but had no significant effect in aerobic cells. These results suggest that both compounds at least partially share a common mechanism of uptake and/or accumulation with fructose.