Evidence of different pharmacokinetics between cyclosporine and tacrolimus in renal transplant recipients: why cyclosporine is monitored by C2 level and tacrolimus by trough level.

The clinical efficacy of calcineurin inhibitors administered to renal transplant recipients is considered to be a strong function of the area under the concentration time curve (AUC). Monitoring of blood concentrations for two similar calcineurin inhibitors, cyclosporine (CyA) and tacrolimus (TAC) are different. Namely, CyA blood concentration is usually monitored at two hours after administration (C2), a surrogate for peak concentration (Cp), and TAC at trough concentration (Ct). We examined the behavior of blood concentration curves simultaneously for both CyA and TAC in renal transplant recipients with similar clinical backgrounds. Furthermore, we analyzed the correlation of Cp and Ct vs AUC implementing an area under the trough level, or area above the trough level as new pharmacokinetic parameters, so that C2 for CyA and Ct for TAC has validated using controlled clinical data. We observed differences in the pharmacokinetics between.

Enhanced delivery of doxorubicin to tumor by long-circulating thermosensitive liposomes and local hyperthermia.

Doxorubicin (DXR) was encapsulated in long-circulating, thermosensitive liposomes (180-200 nm), prepared from dipalmitoylphosphatidylcholine (DPPC)/distearoylphosphatidylcholine (DSPC) (9:1 (m/m)) and 6 mol% of ganglioside GM1 (GM1), with 95-98% entrapping efficiency by the pH-gradient method. 45% of the entrapped DXR was released from these GM1/DPPC/DSPC liposomes by incubation at 42 degrees C for 5 min in 20% serum or saline (this degree of release was lower than that of hydrophilic drugs such as cisplatin, due to the basic and amphiphilic nature of DXR). Inclusion of GM1 (6 mol%) endowed DPPC/DSPC liposomes with prolonged circulation ability, resulting in increased blood levels of liposomes and decreased reticuloendothelial system uptake over 6 h after injection. Concomitantly, DXR levels in blood remained high for long time. Accumulation of DXR into tumor tissue of tumor-bearing mice (mouse colon carcinoma 26) by local hyperthermia after injection of DXR loaded, long-circulating, thermosensitive (DXR-GM1/DPPC/DSPC) liposomes was 2.5-times or 6-times higher than that after treatment with DXR-DPPC/DSPC liposomes or free DXR in combination with hyperthermia, respectively. Furthermore, the treatment with DXR-GM1/DPPC/DSPC liposomes and hyperthermia resulted in effective tumor-growth retardation and increased survival time. Our results indicate that the combination of drug-loaded, long-circulating, thermosensitive liposomes with local hyperthermia at the tumor site could be clinically useful for delivering a wide range of chemotherapeutic agents in the treatment of solid tumors.

Formation of platelet-activating factor-like phospholipids by Fe2+/ascorbate/EDTA-induced lipid peroxidation.

We have identified novel phospholipids together with platelet-activating factor and its 1-acyl analogues in purified fractions from a bovine brain lipid extract. These novel compounds were phospholipids with an sn-2-short-chain monocarboxylyl, dicarboxylyl or omega-hydroxymonocarboxylyl group. The profiles of these three types of phospholipids suggest that they were formed by lipid peroxidation. To examine this possibility, we peroxidized synthetic phosphatidylcholines (PC) with an sn-2-polyunsaturated fatty acyl group and PC from bovine brain, with Fe2+/ascorbate/EDTA, and analyzed the secondary degradation products retaining a glycerol backbone by fast atom bombardment-mass spectrometry and GC-MS. Results showed the formation of four kinds of PC with a short-chain monocarboxylate, dicarboxylate, dicarboxylate semialdehyde or omega-hydroxymonocarboxylate moiety. The chain lengths of these PC were related to the position of the double bond vicinal to the esterified carbonyl group in the sn-2-long-chain acyl moiety of the parent PC. The molecular heterogeneity of secondary products formed by the oxidative degradation of bovine brain PC resembled those of the unique phospholipids that we previously detected in the fractions with platelet-activating factor-like activity purified from a bovine brain lipid extract, although the former lacked the species with an acetyl group. These results suggest that all the novel phospholipids with a short-chain acyl group in the brain lipid extract except that with an acetyl group were produced by lipid peroxidation.

Asparagine-linked carbohydrate of Penicillium notatum phospholipase B.

The asparagine-linked sugar chains of phospholipase B from Penicillium notatum were released by hydrazinolysis. The carbohydrate components were re-N-acetylated and then reductively aminated with 2-aminopyridine. The pyridyl-aminated derivatives were analyzed by size-fractionation and reverse-phase HPLC. The results indicated that the enzyme contains only one kind of oligosaccharide. The carbohydrate was purified by HPLC, and then subjected to NMR and MS analyses for determination of its structure. Based on the results, the structure was determined to be as follows. [sequence: see text]

Involvement of prostaglandin E2 in rabbit corneal injury by anterior segment ischaemia.

The involvement of prostaglandins (PGs) in the development of anterior segment ischaemia after occlusion of the bilateral long posterior ciliary arteries was investigated in rabbit eyes. In this experimental ischaemia, the tissue weight and protein content in the peripheral cornea and the protein content in the aqueous humour increased on the first postoperative day. Topically applied cyclooxygenase inhibitor diclofenac (0.1%) reduced corneal inflammation and further suppressed the elevation in the tissue weight and protein content in the peripheral cornea on day 1 after ischaemia, but did not affect the changes in the aqueous humour. Subconjunctivally administered PGE1 and PGE2 induced corneal oedema and increased corneal protein content in diclofenac-treated and ischaemia-induced eyes, but PGD2, PGF2alpha, and the stable PGI2 analogue cicaprost did not evoke any change. In fact, PGE2 content was markedly increased in the aqueous humour on day 1 after ischaemia, and diclofenac suppressed the increase. In addition, CPT-cAMP increased the corneal tissue weight and protein content in organ culture. These observations suggest that PGE2 may play an important role in developing corneal oedema at the initial stage of ischaemic damage, possibly through the cAMP-mediated pathway.

Enhanced tumor targeting of doxorubicin by ganglioside GM1-bearing long-circulating liposomes.

Doxorubicin (DXR) was encapsulated in long-circulating liposomes, composed of ganglioside GM1 (GM1)/distearoylphosphatidylcholine (DSPC)/cholesterol (CH) (0.13:1:1 in molar ratio) and sized to approximately 100 nm in mean diameter, with 98% entrapping efficiency by the transmembrane pH gradient method. Free DXR, DXR-DSPC/CH and DXR-GM1/DSPC/CH liposomes were injected intravenously into Colon 26 tumor-bearing Balb/c mice via the tail vein at a dose of 5.0 mg DXR/kg. DXR-GM1/DSPC/CH liposomes gave a higher blood level of the drug than did DXR-DSPC/CH liposomes or free DXR up to 24 hours after injection, and the area under the blood concentration-time curve (AUC) for DXR-GM1/DSPC/CH liposomes was 1.5 or 526 times higher than that for DXR-DSPC/CH liposomes or free DXR, respectively. DXR-GM1/DSPC/CH liposomes gave a decreased DXR concentration in the reticuloendothelial system (RES) of the liver and the spleen. Both liposomal formulations effectively reduced the DXR concentration in the heart as compared with that in the case of free DXR. At 6 hours after i.v. injection, DXR-GM1/DSPC/CH liposomes provided an approximately 3.3- or 9-fold higher peak DXR level in the tumor as compared with DXR-DSPC/CH liposomes or the free drug, respectively. These high tumor levels of DXR appear to reflect the prolonged residence time of the liposomes. The results suggest that encapsulation of DXR in GM1-bearing long-circulating liposomes will be useful for cancer chemotherapy.

Liposome drug delivery system for murine neuroblastoma.

PURPOSE: The effects of liposome-infused doxorubicin on C-1300 murine neuroblastoma were studied. The liposome surface was covered with polyethylene glycol to avoid migration toward the reticuloendothelial system and to prolong its presence in the bloodstream. Liposome-infused doxorubicin hydrochloride (DXR), an anthracycline was used as an anticancer antibiotic substance. METHODS: Each A/J mouse was transplanted with 1 x 10(5) C-1300 murine neuroblastoma cells subcutaneously in the thigh. The experiment was conducted when the maximum tumor dimension was 1 cm. The control group was given only physiological saline solutions, the second group was given DXR alone, and the third group received liposome-infused DXR (Lip-DXR). The survival and doubling times were measured. One, 12, and 24 hours after the injection, the DXR concentration in the cardiac tissues was measured for statistical comparison. RESULTS: The survival time of the mice was found to be 27+/-5.10 days in the control group, 31.40+/-3.15 days in the DXR group, and 43.86+/-2.13 days in the Lip-DXR group. The Lip-DXR group showed the longest survival time. The tumor-doubling time was found to be 9.07+/-2.30, 10.75+/-3.49, and 19.80+/-3.26 days, for each group, respectively. When comparing the DXR concentration in the heart tissues, the Lip-DXR-administered mice showed significantly lower DXR accumulation in the cardiac tissues after 1 and 12 hours than the DXR-administered mice. CONCLUSION: This study proved that liposome-infused DXR could be used effectively on murine neuroblastoma (C-1300 tumor cell model) and may reduce the incidence of cardiac toxicity as compared with DXR alone.

Falsely abnormally elevated blood trough concentration of tacrolimus measured by antibody-conjugated magnetic immunoassay in a renal transplant recipient: a case report.

This report presents a falsely abnormally elevated blood trough concentration (C(t)) of tacrolimus measured by antibody-conjugated magnetic immunoassay (ACMIA) methods in a renal transplant recipient. Because the C(t) of tacrolimus was 78.5 ng/mL at day 2 after a 52-year-old man underwent renal transplantation, we stopped the tacrolimus extended-release formulation. However, because the abnormally elevated blood C(t) continued in the range of 41.1-59.1 ng/mL, we then measured the tacrolimus concentration in a stored blood sample before renal transplantation, it was 43 ng/mL. Consequently, the day-7 blood sample was measured with both ACMIA and enzyme-linked immunoassay, showing C(t) values of 42.8 ng/mL and 0.89 ng/mL, respectively. Because the abnormally elevated C(t) was falsely measured by the ACMIA method, we restarted tacrolimus However, the calcineurin inhibitor was subsequently converted to cyclosporine at day 21 after renal transplantation. Although cyclosporine was also measured by ACMIA, there was not an abnormally elevated C(t). Subsequently, the tacrolimus concentration ratio in plasma and whole blood (P/B-tacrolimus concentration ratio) was measured by ACMIA in a posttacrolimus blood sample. The P/B-tacrolimus concentration ratio was 100%. In contrast, the P/B-tacrolimus concentration ratio was <30% in 2 control patients administered tacrolimus. It has been reported recently that there were cases showing falsely slightly elevated C(t) of tacrolimus within the therapeutic range of concentrations. Therefore, we must be careful not to reduce the tacrolimus dose falsely. We consider confirmatory methods for a falsely abnormally elevated C(t) of tacrolimus measured by ACMIA to (1) measure P/B-tacrolimus concentration ratio, (2) compare ACMIA with another measurement, and (3) evaluate a blood sample stored before tacrolimus administration.

Characterization of prostaglandin F2 alpha production in pregnant and cycling mice.

To clarify the regulation of prostaglandin F2 alpha (PGF2 alpha) production in vivo in mice during pregnancy and the estrous cycle, we injected [3H]PGF2 alpha i.v. into female mice and determined the structures of three urinary metabolites by gas chromatography-mass spectrometry. The compounds were 5,7,11-trihydroxy-tetranor-prosta-9-enoic acid (FUM-I), 5,7,11-trihydroxy-tetranor-prostanoic acid (FUM-II), and 5,7-dihydroxy-11-keto-tetranor-prostanoic acid (FUM-III). The major metabolite, FUM-III, increased four-fold at the term of pregnancy and transiently during diestrus of the estrous cycle. Consistent with the increase in FUM-III, immunoblot analysis with anti-bovine PGF synthase antiserum demonstrated that a 36-kDa protein band corresponding to PGF synthase increased in the uterus at late pregnancy and during diestrus. These results suggest that PGF2 alpha production may increase at term and change during the estrous cycle and is associated with an increase in uterine PGF synthase concentrations.

Antitumor activity of doxorubicin encapsulated in poly(ethylene glycol)-coated liposomes.

The antitumor activity of doxorubicin (DXR) which had been encapsulated in poly(ethylene glycol) (PEP)-coated long-circulating liposomes was examined in mice inoculated with colon 26 carcinoma cells. Six mol% of the distearoylphosphatidylethanolamine derivative of PEGs with different molecular weights was incorporated in liposomes (90-110 nm, mean diameter) composed of distearoylphosphatidylcholine/cholesterol (1/1, molar ratio), and the encapsulating efficiency of DXR in liposomes was more than 98% by the pH gradient method. Each concentration of DXR in blood and tumor tissue was significantly greater after administration of the drug encapsulated in PEG-coated liposomes (DXR-PEG-liposome) compared to the non-coated control liposomes or non-encapsulated free drug. DXR-PEG-liposome prepared with PEG1000 (DXR-PEG1000-liposome) more effectively increased the level of DXR in blood and tumor than did the preparations with PEG5000 or PEG 12000. A single treatment with DXR-PEG1000-liposome (10 mg DXR/kg) resulted in increased survival time. Further therapeutic improvement in terms of tumor growth retardation and prolongation of survival time were observed following multiple treatments with DXR-PEG1000-liposome (3 x 5 mg DXR/kg). Long-circulating liposome coating optimized PEGs should be useful for the delivery of chemotherapeutic agents for the treatment of solid tumors.

Enhanced delivery and antitumor activity of doxorubicin using long-circulating thermosensitive liposomes containing amphipathic polyethylene glycol in combination with local hyperthermia.

Enhanced delivery of doxorubicin (DXR) to a solid tumor subjected to local hyperthermia was achieved by using long-circulating, thermosensitive liposomes (TSL) composed of dipalmitoyl phosphatidylcholine (DPPC)/distearoyl phosphatidylcholine (DSPC) (9:1, m/m) and 3 mol% amphipathic polyethylene glycol (PEG) in colon 26-bearing mice. Inclusion of 3 mol% of distearoyl phosphatidylethanolamine derivatives of PEG (DSPE-PEG, amphipathic PEG) with a mean molecular weight of 1000 or 5000 in DPPC/DSPC liposomes resulted in decreased reticuloendothelial system (RES) uptake and a concomitant prolongation of circulation time, affording sustained increased blood levels of the liposomes. Concomitantly, DXR levels in blood were also kept high over a long period. The presence of amphipathic PEG did not interfere with the encapsulation of DXR by the pH gradient method (> 90% trapping efficiency) or with the temperature-dependent drug release from the liposomes. The optimal size of these liposomes was 180-200 nm in mean diameter for thermosensitive drug release and prolonged circulation time. The DXR levels in the tumor after injection of long-circulating TSL (DXR-PEG1000TSL or DXR-PEG5000TSL, at a dose of 5 mg DXR/kg) with local hyperthermia were much higher than after treatment with DXR-TSL lacking PEG or with free DXR, reaching 7.0-8.5 DXR micrograms/g tumor (approximately 2 times or 6 times higher than that of DXR-TSL or free DXR, respectively). Furthermore, the combination of DXR-PEGTSL and hyperthermia effectively retarded tumor growth and increased survival time.(ABSTRACT TRUNCATED AT 250 WORDS)