Mycophenolate mofetil: pharmacokinetic strategies for optimizing immunosuppression.
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Dose-Response Relationship, Drug↗
Biomedical subjects
L M Shaw
Publications and source records attributed to L M Shaw.
The integrin alpha 6 beta 1 promotes the survival of metastatic human breast carcinoma cells in mice.
The role of the integrin alpha 6 beta 1 in breast carcinoma progression was studied by targeted elimination of this integrin in MDA-MB-435 cells, a human breast carcinoma cell line that is highly metastatic in athymic mice. The strategy used is based on the finding that expression of a cytoplasmic domain deletion mutant of the beta 4-integrin subunit (beta 4-delta CYT) in MDA-MB-435 cells eliminates formation of the alpha 6 beta 1 heterodimer. MDA-MB-435 cells that lacked alpha 6 beta 1 expression (beta 4-delta CYT transfectants) formed tumors in athymic mice that were suppressed in their growth and that exhibited a significant increase in apoptosis in comparison to the control tumors. Unlike the control MDA-MB-435 cells, the beta 4-delta CYT transfectants were unable to establish metastatic foci in the lungs. Also, the control transfectants grew substantially better than the beta 4-delta CYT transfectants in the liver after intrahepatic injection because of extensive apoptosis in the beta 4-delta CYT transfectants. These data suggest that a major function of the alpha 6 beta 1 integrin in breast carcinoma is to facilitate tumorigenesis and promote tumor cell survival in distant organs.
Animals↗
Activation of Jak/STAT proteins involved in signal transduction pathway mediated by receptor for interleukin 2 in malignant T lymphocytes derived from cutaneous anaplastic large T-cell lymphoma and Sezary syndrome.
Signaling through the interleukin 2 receptor (IL-2R) involves phosphorylation of several proteins including Jak3, STAT5, and, in preactivated cells, STAT3. In the present study, we examined the functional status of the IL-2R-associated Jak/STAT pathway in malignant T lymphocytes from advanced skin-based lymphomas: anaplastic large T-cell lymphoma (ALCL) and Sezary syndrome (SzS). Proliferation of three ALCL cell lines (PB-1, 2A, and 2B) was partially inhibited by rapamycin, a blocker of some of the signals mediated by IL-2R, but not by cyclosporin A, FK-506, and prednisone, which suppress signals mediated by the T-cell receptor. All the cell lines expressed on their surface the high-affinity IL-2R (alpha, beta, and gamma c chains). They showed basal, constitutive phosphorylation, and coassociation of Jak3, STAT5, and STAT3. Weak basal phosphorylation of IL-2R gamma c was also detected. In regard to SzS, peripheral blood mononuclear cells from 10 of 14 patients showed basal phosphorylation of Jak3, accompanied by phosphorylation of STAT5 in 9 patients, and STAT3 in 4 patients. However, in vitro overnight culture of SzS cells without exogenous cytokines resulted in markedly decreased Jak3 and STAT5 phosphorylation, which could be reversed by stimulation with IL-2. This indicates that the basal phosphorylation of Jak3 and STAT5 in freshly isolated SzS cells is induced rather than constitutive. The basal activation of the Jak/STAT pathway involved in IL-2R signal transduction in ALCL and SzS cells reported here suggests that this pathway may play a role in the pathogenesis of cutaneous T-cell lymphomas, although the mechanism (induced versus constitutive) may vary between different lymphoma types.
Cell Membrane↗
Function of the integrin alpha 6 beta 1 in metastatic breast carcinoma cells assessed by expression of a dominant-negative receptor.
The involvement of the alpha 6 beta a integrin, a laminin receptor, in breast carcinoma progression needs to be addressed rigorously. We report that a human breast carcinoma cell line, MDA-MB-435, known to be highly invasive and metastatic, expresses three potential integrin laminin receptors: alpha 2 beta 1, alpha 3 beta 1, and alpha 6 beta 1, but uses only alpha 6 beta 1 to mediate adhesion and migration on laminin matrices. To investigate the contribution of alpha 6 beta 1 to the aggressive behavior of these cells, we developed a dominant-negative strategy for knocking out alpha 6 beta 1 function that involved expression of a cytoplasmic domain deletion mutant of the beta 4 integrin subunit by cDNA transfection. Stable transfectants of MDA-MB-435 cells that expressed this mutant beta 4 subunit were inhibited dramatically in their ability to adhere and migrate on laminin matrices, and their capacity to invade Matrigel was reduced significantly. These findings support the hypothesis that alpha 6 beta 1 is important for breast cancer progression. Moreover, this approach is a powerful method that should be useful in assessing the role of alpha 6 beta 1 in other cells.
Breast Neoplasms↗
Coacaethylene levels in patients who test positive for cocaine.
STUDY OBJECTIVE: To determine the presence of cocaethylene, an active metabolite of the combination of cocaine and ethanol, among trauma patients who test positive for cocaine. METHODS: We assembled a case series of 416 consecutive urban trauma center patients with major trauma. Urine was tested for the presence of the cocaine metabolite benzoylecgonine. Plasma was quantitatively assayed for cocaine, ethanol, and cocaethylene. RESULTS: Of the study subjects, 158 (38%) were positive for benzoylecgonine. Of the 114 of these subjects who had adequate plasma specimens, 68 (60%) tested positive for cocaethylene (mean, 41 +/- 27 ng/mL; range, 3 to 213 ng/mL), all tested positive for cocaine (mean, 92.9 +/- 52 ng/mL), and 56% were positive for ethanol (mean, 175 +/- 85 mg/mL). We found poor correlation between admission levels of cocaethylene and cocaine (R=.02), even when subjects were stratified by ethanol level. The correlation between cocaethylene and ethanol levels was weak (R=.24). Of the 68 patients who tested positive for cocaethylene, 29% tested negative for ethanol. Plasma was also assayed from 94 subjects who tested negative for benzoylecgonine; 9% had detectable levels of cocaine, and 2% had detectable levels of cocaethylene. CONCLUSION: Cocaethylene was present in more than half of the subjects who tested positive for cocaine.
Adolescent↗
Nighttime dosing assures postdistribution sampling for therapeutic drug monitoring of digoxin.
To study the appropriateness of phlebotomy for digoxin therapeutic drug monitoring (TDM) in outpatients, we conducted a retrospective chart review, a computer search of all previous TDM testing, and a questionnaire of all outpatients (n = 86) who had serum digoxin determinations between April 10 and April 28, 1992 (585 tests). In patients who took digoxin at the same time daily (40 patients, 300 tests), 52% of tests were performed on inappropriate samples drawn within 6 h of the last dose. No patient who took digoxin after 1700 had inappropriate tests. Phlebotomy for serum digoxin determinations before distribution of digoxin is complete is a common problem in outpatients, leading to clinically uninterpretable test results. Postdistribution sampling can be assured by nighttime dosing, and this recommendation has been implemented at our hospital.
Adult↗
Toxic effects of immunosuppressive drugs: mechanisms and strategies for controlling them.
Since cyclosporine (CsA) was introduced into clinical practice in late 1983 to prevent rejection in transplant patients, there has been an almost explosive growth in the number and types of transplants and the number of transplant centers, an increase in the life expectancy of the transplanted organ, and substantial decreases in rates of acute rejection and life-threatening infections. Despite these successes, major improvements in immunosuppressive therapy are needed, especially a reduction in toxic side effects and a rigorous definition of the relation between drug concentration and clinical effects. Such improvements may be achievable with the incorporation of new drugs such as tacrolimus and mycophenolate mofetil into immunosuppression protocols and the development of rigorously defined therapeutic drug-monitoring programs.
Cyclosporine↗
Pharmacokinetic profile of amifostine.
This article reviews the chemistry, measurement, metabolism, and pharmacokinetics of the cytoprotective agent amifostine. Validated analytic methodology to measure parent drug and pharmacologically active metabolites and pharmacokinetic studies are essential to the efficient performance and analysis of clinical studies. Well-validated analytic methods developed in the authors' laboratory were used to characterize this agent. Amifostine [s-2-(3-aminopropylamino)ethyl-phosphorothioate] is the phosphorylated pro-drug form of the active free thiol drug WR-1065 [2-(3-aminopropylamino)ethanethiol]. Observations described here support the hypothesis that amifostine is dephosphorylated rapidly by alkaline phosphatase present on the plasma membranes of the arteriolar endothelium of various normal tissues and on the plasma membranes of the kidneys' proximal tubular epithelium to its active thiol metabolite WR-1065, which in turn immediately enters normal tissues. Other metabolites that have been identified are WR-33278, the symmetrical disulfide of WR-1065; the mixed disulfides WR-1065-cysteine and WR-1065-glutathione; and cysteamine. Amifostine was recently approved by the US Food and Drug Administration for use as a cytoprotector in cancer patients receiving chemotherapy. Current pharmacokinetic studies in cancer patients are focusing on establishing a population model as a basis for developing limited sampling strategies for future investigations of the pharmacokinetic-pharmacodynamic behavior of amifostine.
Amifostine↗
Serum lactate and lactate dehydrogenase in high concentrations interfere in enzymatic assay of ethylene glycol.
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Adult↗
Diurnal variation of cyclosporine clearance in stable renal transplant recipients receiving continuous infusion.
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Circadian Rhythm↗
The alpha 6A beta 1 and alpha 6B beta 1 integrin variants signal differences in the tyrosine phosphorylation of paxillin and other proteins.
Integrin receptors can mediate transmembrane signaling in response to ligand binding. To further examine the role of the integrin alpha subunit in these signaling functions, we assessed the contribution of the alpha 6 cytoplasmic domain variants to the signaling properties of the alpha 6 beta 1 integrin using P388D1 cells that had been transfected with either the alpha 6A or the alpha 6B cDNA. The alpha 6A beta 1 and alpha 6B beta 1 receptors induced marked quantitative differences in the tyrosine phosphorylation of several proteins after binding to laminin. Specifically, the alpha 6A cytoplasmic domain was more effective than the alpha 6B cytoplasmic domain in inducing the tyrosine phosphorylation of three major proteins (molecular mass, 120, 110, and 76 kDa). In addition to these proteins, we also observed that the tyrosine phosphorylation of the cytoskeletal protein paxillin was increased significantly more by alpha 6A beta 1 integrin-mediated adhesion to laminin than by that of alpha 6B beta 1. This differential pattern of tyrosine phosphorylation induction does not appear to be a secondary event initiated by cell shape changes. Also, differences in tyrosine phosphorylation in the alpha 6 transfectants were not evident in response to attachment to other substrates. These findings provide biochemical evidence for functional differences between alpha subunit cytoplasmic domain variants of the same integrin.
Animals↗
Mouse egg integrin alpha 6 beta 1 functions as a sperm receptor.
Binding between sperm and egg plasma membranes is an essential step in fertilization. Whereas fertilin, a mammalian sperm surface protein, is involved in this crucial interaction, sperm receptors on the egg plasma membrane have not been identified. Because fertilin contains a predicted integrin ligand domain, we investigated the expression and function of integrin subunits in unfertilized mouse eggs. Polymerase chain reactions detected mRNAs for alpha 5, alpha 6, alpha v, beta 1, beta 3, and beta 5. Immunofluorescence revealed alpha 6 beta 1 and alpha v beta 3 on the plasma membrane. GoH3, a function-blocking anti-alpha 6 monoclonal antibody, abolished sperm binding, but a nonfunction-blocking anti-alpha 6 monoclonal antibody, a function-blocking anti-alpha v beta 3 polyclonal antibody, and an RGD peptide had no effect. Somatic cells bound sperm avidly, but only if they expressed alpha 6 beta 1. A peptide analog of the fertilin integrin ligand domain inhibited sperm binding to eggs and alpha 6 beta 1+ cells and diminished GoH3 staining of eggs. Our results indicate a novel role for the integrin alpha 6 beta 1 as a cell-cell adhesion receptor that mediates sperm-egg binding.
ADAM Proteins↗
A comparison of the effect of angiotensin converting enzyme inhibition and angiotensin II receptor antagonism on the structural changes associated with hypertension in rat small arteries.
OBJECTIVE: To investigate whether, when angiotensin converting enzyme inhibitors are administered to young, genetically hypertension-prone animals, the demonstrated attenuation of blood pressure development and prevention of the structural changes usually observed in small arteries is attributable to the prevention of angiotensin II production. DESIGN: We have treated spontaneously hypertensive rats (SHR) aged 4-20 weeks with either lisinopril (1 or 10 mg/kg) or the angiotensin II receptor antagonist D 8731 (1, 20 or 50 mg/kg). METHODS: Blood pressure was measured and structural parameters in small arteries from four vascular beds were examined using isometric myography. RESULTS: At age 20 weeks lisinopril had attenuated blood pressure development and prevented cardiac hypertrophy (but not vascular hypertrophy) in a dose-dependent manner. The highest dose of lisinopril had reduced the blood pressure of the SHR to below that of the Wistar-Kyoto (WKY) rats and prevented most structural changes, but there was a slight reduction in body weight in those rats. Comparable blood pressure control with D 8731 was associated with similar structural parameters. CONCLUSION: The prevention of hypertension-associated vascular structural alteration appears to be dependent upon the degree of blood pressure control.
Angiotensin Receptor Antagonists↗
Analytic requirements for immunosuppressive drugs in clinical trials.
Essential to the evaluation of (1) the pharmacokinetics, (2) concentration-effect relationships, and (3) the application of therapeutic drug monitoring, during new immunosuppressive drug clinical trials, is the development of validated analytical methodology for the measurement of pharmacologically active drug and metabolites in biofluids and tissues. The characteristics of analytical methodology developed for cyclosporines A and G, FK-506, mycophenolate mofetil, and rapamycin during clinical trials will be described. The advantages of establishing validated analytical methodology as early as possible during clinical trials include: (a) early identification of metabolites and their quantitative and pharmacological significance; (b) early development of interpretable PK and PK-PD data; (c) accrual of experience that will be directly useful in patient monitoring after drug approval; (d) optimization of analysis conditions; and (e) early development of reference methodology for therapeutic drug monitoring tests. A suggested set of performance criteria for drug analysis during clinical trials and thereafter will be presented.
Clinical Trials as Topic↗
Mycophenolic acid: measurement and relationship to pharmacologic effects.
Validated high-performance liquid chromatographic (HPLC) methods have been developed for the reliable measurement in plasma of mycophenolate mofetil (MMF), mycophenolic acid (MPA), its pharmacologically active metabolite, and mycophenolic acid glucuronide (MPAG), the inactive and primary metabolite of MPA. Using these validated HPLC methods, the pharmacokinetic behavior of MMF has been characterized in renal transplant patients as part of double-blind randomized multicenter clinical trials. The analytical performance characteristics of the HPLC methods are described. Based on investigations of the metabolism of MMF in animals, normal volunteers, and renal transplant patients, it has been established that MMF is rapidly converted to MPA. MPAG is the primary urinary excretion product derived from MPA. In vitro binding studies have revealed that MPA is extensively bound to plasma proteins and that human serum albumin is the primary binding protein. Pharmacokinetic studies have revealed the following: (1) MPA exhibits enterohepatic circulation in humans as a result of biliary excretion of MPAG followed by MPA production in the gastrointestinal tract and the appearance of secondary MPA peaks in plasma; (2) chronic renal impairment produces little change in the clearance of MPA but a marked decrease in plasma MPAG clearance, with a consequent substantial increase in the circulating MPAG concentration; and (3) pharmacokinetic/pharmacodynamic (PK/PD) studies have revealed a good correlation between drug exposure (AUC) and the probability of rejection. The implications of the PK/PD studies for the development of TDM practices are discussed.
Antibiotics, Antineoplastic↗
Mycophenolate mofetil: a report of the consensus panel.
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Animals↗
Topical application of WR-2721 achieves high concentrations in the rectal wall.
Rectal wall injury is an important treatment-related morbidity in patients treated with radiation for prostate cancer. We have undertaken this study to investigate the merits of topical intrarectal application of the radioprotective compound WR-2721. Male Copenhagen rats were injected intrarectally with 2% WR-2721 gel. At 10, 20, 30 and 40 min after application, a laparotomy was performed, and the rectum and prostate were removed. Concentrations of total WR-1065 (the active metabolite of WR-2721) were determined in these samples by an HPLC assay. While the concentration in the rectal wall tended to increase with time, it did not change substantially in the prostate. The concentration in the rectal wall was found to be significantly higher at all times. We conclude that preferential accumulation of WR-2721 in the rectal wall can be achieved by topical application. This is a promising approach to modifying rectal wall tolerance that deserves more study.
Administration, Topical↗
Mycophenolic acid binding to human serum albumin: characterization and relation to pharmacodynamics.
Mycophenolate mofetil, the prodrug form of the immunosuppressive agent mycophenolic acid (MPA), is currently in clinical trials evaluating its effectiveness in transplant recipients. In this study, we validated an ultrafiltration system for the reliable measurement of free MPA. Using this technique, we evaluated factors that might be important in modulating the free fraction of this drug. Human serum albumin (HSA), high concentrations of the primary glucuronide metabolite of MPA, and sodium salicylate significantly affected MPA binding. For HSA the mean +/- SE binding capacity (Bmax) and the dissociation constant (Kd) were 1095 +/- 34 mumol/L and 12.98 +/- 0.93 mumol/L, respectively. The dose for 50% inhibition (IC50) of inosine monophosphate dehydrogenase isoform II by MPA increased 5.4-fold as the concentration of HSA added to the enzyme reaction mixture increased from 0 to 50 g/L (0-724 mumol/L). Furthermore, the IC50 MPA concentration for phytohemagglutinin A-stimulated human peripheral blood mononuclear cells increased 4.8-fold when incubations were performed in the presence of 10 g/L (145 mumol/L) HSA vs no added HSA. These data support the hypothesis that the pharmacological activity of MPA is a function of unbound drug concentration.
Dialysis↗