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J J Lipsky

Publications and source records attributed to J J Lipsky.

At least 37 records · Page 2Linked to original sources
PubMed1999

Systemic and intestinal pharmacokinetics of methotrexate in patients with inflammatory bowel disease.

BACKGROUND: The pharmacokinetics of low-dose subcutaneous methotrexate have not been determined throughout the standard weekly dosing interval. It is not known whether methotrexate concentrations in the gastrointestinal tract are sufficient for pharmacologic activity in inflammatory bowel disease. METHODS: Ten patients with inflammatory bowel disease participated in the study. After the patients started taking 15 or 25 mg subcutaneous methotrexate once a week, erythrocyte methotrexate concentration was measured every 2 weeks. The absorption, rectal distribution, metabolism, and elimination of methotrexate were measured. The effect of methotrexate on proliferation of an intestinal epithelial cell line was determined. RESULTS: After weekly subcutaneous administration of methotrexate was begun, trough erythrocyte concentration rose to reach a plateau after 6 to 8 weeks, ranging from 150 to 300 nmol/L. More than 90% of subcutaneously administered methotrexate was rapidly excreted in the urine. The methotrexate plasma time course after subcutaneous administration fit a 2-compartment first-order model with biphasic elimination and trough concentration of about 1 nmol/L. Trough and peak methotrexate concentrations (mean value +/- SD) were 64 +/- 33 and 206 +/- 64 fmol/mg in the rectal mucosa and 4 +/- 3 and 51 +/- 26 nmol/L in the rectal lumen. These methotrexate concentrations were in the range found to be pharmacologically active against Caco-2 cell growth, that is, a 50% inhibitory concentration from 10 to 46 nmol/L. CONCLUSION: Subcutaneous methotrexate was well absorbed and distributed to the site of the lesions in patients with inflammatory bowel disease. Methotrexate was concentrated intracellularly in blood and in the rectum. The methotrexate concentration in the rectal mucosa remained within a pharmacologically active range throughout the dosing interval. The findings represent a pharmacologic explanation for the sustained efficacy of weekly methotrexate therapy.

Adult↗
PubMed1999

Lack of effect of intravenous administration on time to respond to azathioprine for steroid-treated Crohn's disease. North American Azathioprine Study Group.

BACKGROUND & AIMS: Azathioprine is effective for Crohn's disease but acts slowly. A loading dose may decrease the time to response. METHODS: A placebo-controlled study was conducted in patients with active Crohn's disease despite prednisone treatment. Patients were randomized to a 36-hour infusion of azathioprine, 40 mg/kg (51 patients), or placebo (45 patients) followed by oral azathioprine, 2 mg/kg, for 16 weeks. Prednisone was tapered over 5 weeks. The primary outcome measure was complete remission at week 8, defined by discontinuation of prednisone and a Crohn's Disease Activity Index of </=150 points. Erythrocyte concentrations of the azathioprine active metabolite, 6-thioguanine nucleotide, were measured. RESULTS: At week 8, 13 patients (25%) were in complete remission in the azathioprine-loaded group compared with 11 patients (24%) in the placebo group. The frequency of complete remission did not increase after 8 weeks in either group. Both groups achieved steady state of 6-thioguanine nucleotide by week 2, and no differences were found in mean concentrations between the groups. There were no significant differences in the frequency of adverse events between the groups. CONCLUSIONS: A loading dose does not decrease the time to response in patients with steroid-treated Crohn's disease beginning azathioprine therapy. Steady state of erythrocyte 6-thioguanine nucleotide and complete response occurred earlier than previously reported.

Administration, Oral↗
PubMed1999

A randomized dose-response and pharmacokinetic study of methotrexate for refractory inflammatory Crohn's disease and ulcerative colitis.

BACKGROUND AND AIMS: The optimum initial dose of methotrexate for steroid-requiring inflammatory bowel disease is not known. AIM: To compare directly the efficacy and toxicity of methotrexate 15 and 25 mg/week, and to explore the value of methotrexate blood levels as predictors of outcome. METHODS: A 16-week randomized single-blind comparison of subcutaneous methotrexate 15 or 25 mg/week was performed in 32 patients with steroid-requiring Crohn's disease or ulcerative colitis. Patients who did not respond to methotrexate 15 mg/week were further studied for an additional 16 weeks on methotrexate 25 mg/week. Blood was drawn every 2 weeks for methotrexate levels. RESULTS: After 16 weeks, 17% of patients in each group achieved remission; 39% of patients randomized to 15 mg/week and 33% of patients randomized to 25 mg/week improved (P=N.S. ). Clinical status improved in four out of 11 patients after methotrexate dose escalation from 15 to 25 mg/week. Toxicity was not different between the treatment groups. Methotrexate blood levels did not predict efficacy or toxicity. CONCLUSIONS: For induction of remission in steroid-requiring inflammatory bowel disease, subcutaneous methotrexate at initial doses of 15 and 25 mg/week are equally efficacious. At these doses, response is not associated with blood methotrexate concentrations.

Adult↗
PubMed1999

Plasma and rectal adenosine in inflammatory bowel disease: effect of methotrexate.

In animal models, the antiinflammatory mechanism of action of methotrexate has been attributed to elevation of the extracellular concentration of the endogenous autocoid, adenosine. Our goal was to determine if methotrexate elevates adenosine concentrations in plasma and at the site of disease in patients with inflammatory bowel disease. In 10 patients with Crohn's disease or ulcerative colitis, rectal adenosine and plasma adenosine concentrations were measured before and immediately after a subcutaneous injection of methotrexate, 15 or 25 mg. The mean predose rectal adenosine concentration of 2.4 mumol/l was not significantly different from the postdose concentration of 2.1 mumol/l, p = 0.17, (paired two-tailed t test). Rectal adenosine concentration tended to correlate with rectal endoscopic disease activity, r = 0.59, p = 0.067 (Spearman rank order correlation). After methotrexate injection, neither the mean daily plasma adenosine concentration, nor the plasma adenosine at any individual time point, were significantly different from preinjection values. In patients with inflammatory bowel disease, an injection of methotrexate in the clinically effective dose range does not raise rectal or plasma adenosine concentrations. A role for adenosine as the mediator of the antiinflammatory action of methotrexate is not supported.

Adenosine↗
PubMed1999

Roles of FMO and CYP450 in the metabolism in human liver microsomes of S-methyl-N,N-diethyldithiocarbamate, a disulfiram metabolite.

BACKGROUND: The conversion of S-methyl-N,N-diethyldithiocarbamate (MeDDC) to MeDDC sulfine is the first step after methylation in the metabolic pathway of disulfiram, an alcohol deterrent, to its ultimate active metabolite. Various isoforms of CYP450 have recently been shown to catalyze this reaction, but the involvement of flavin monooxygenase (FMO) in this metabolism in humans has not been evaluated. In this study we examined the ability of recombinant human FMO3 in insect microsomes to metabolize MeDDC, and investigated the relative roles of FMO and CYP450 in the metabolism of MeDDC in human liver microsomes. METHODS: HPLC-mass spectrometry was used to identify the products of MeDDC formed by human liver microsomes and by recombinant human FMO3. MeDDC metabolism in human liver microsomes was studied by using either heat inactivation to inhibit FMO, or N-benzylimidazole (NBI) or antibodies to the CYP450 NADPH reductase to inhibit CYP450. RESULTS: We confirmed by HPLC-mass spectrometry that MeDDC sulfine was the major product of MeDDC formed by human liver microsomes and by FMO3. Recombinant FMO3 was an efficient catalyst for the formation of MeDDC sulfine (5.3+/-0.2 nmol/min/mg, mean+/-SEM, n = 6). Inhibition studies showed MeDDC was metabolized primarily by CYP450 in human liver microsomes at pH 7.4, with a 10% contribution from FMO (total microsomal activity 3.1+/-0.2, n = 17). In the course of this work, methyl p-tolyl sulfide (MTS), sulfoxidation of which is used by some investigators as a specific probe for FMO activity, was found to be a substrate for both FMO and CYP450 in human liver microsomes. CONCLUSIONS: Our results prove that MeDDC sulfine is the major product of MeDDC oxidation in human liver microsomes, MeDDC is a good substrate for human FMO3, and MeDDC is metabolized in human liver microsomes primarily by CYP450. We also showed that use of MTS sulfoxidation as an indicator of FMO activity in microsomes is valid only in the presence of a CYP450 inhibitor, such as NBI.

Adult↗
PubMed1998

Inhibition of recombinant human mitochondrial aldehyde dehydrogenase by two intermediate metabolites of disulfiram.

Disulfiram is used in aversion therapy for alcoholism. S-Methyl-N,N-diethylthiocarbamate (MeDTC) sulfoxide, a potent inhibitor of the target enzyme mitochondrial aldehyde dehydrogenase (ALDH2), is thought to be the principal active metabolite of disulfiram in vivo. We examined the effects on recombinant human ALDH2 of two intermediate metabolites of disulfiram, S-methyl-N,N-diethyldithiocarbamate (MeDDC) sulfoxide and MeDDC sulfine. MeDDC sulfoxide was a potent inhibitor of ALDH2 with an IC50 of 2.2 +/- 0.5 microM (mean +/- SD, N = 4) after preincubation with enzyme for 30 min. MeDDC sulfine was a relatively weak inhibitor of ALDH2 under the same conditions with an IC50 value of 62 +/- 14 microM. The inhibition of ALDH2 by both compounds was irreversible and did not require the cofactor NAD. The latter finding demonstrates that inactivation of ALDH2 is independent of the dehydrogenase activity of the enzyme. GSH blocked almost completely the inhibition by 20 microM of MeDDC sulfoxide and greatly diminished the inhibition by 200 microM of MeDDC sulfine. Inactivation by MeDDC sulfoxide was time dependent. MeDTC sulfoxide was a more potent inhibitor of recombinant human ALDH2 (IC50 = 1.4 +/- 0.3 microM after preincubation for 15 min) than either of the intermediate metabolites, and its inhibition was unaffected by GSH. Our results suggest that these newer intermediate metabolites of disulfiram, especially the more potent MeDTC sulfoxide, have the potential to inhibit the target enzyme ALDH2 in patients receiving disulfiram. However, until the significance of the interactions of the inhibitors with GSH is more fully understood, the contribution of MeDDC sulfine and MeDDC sulfoxide to the pharmacological effects of disulfiram in vivo is uncertain.

Alcohol Deterrents↗
PubMed1998

Diadenosine 5',5"-P1,P5-pentaphosphate harbors the properties of a signaling molecule in the heart.

Dinucleotide polyphosphates (ApnA) have emerged as signaling molecules in rapidly dividing cells. The presence and role of Ap5A in the heart remain unknown. Here, we report that the myocardium contains abundant amounts of diadenosine 5',5"-P1,P5-pentaphosphate (Ap5A), a member of the ApnA family. Ischemia induced 10-fold decrease in the myocardial concentration of Ap5A. A target of Ap5A action was identified to be the cardiac ATP-sensitive K+ (K(ATP)) channel, a metabolism-sensitive ion conductance activated in ischemia. At levels found in hearts prior to ischemia, Ap5A maintained a low probability of K(ATP) channel opening, but at levels found in hearts following ischemia, Ap5A allowed a high probability of K(ATP) channel opening. Taken together, the present data suggest that Ap5A harbors the properties of a signaling molecule involved in the cardiac response to metabolic stress.

Animals↗
PubMed1998

Theophylline: recent advances in the understanding of its mode of action and uses in clinical practice.

Theophylline, a drug that has been used for several decades, has several different actions at a cellular level, including inhibition of phosphodiesterase isoenzymes, antagonism of adenosine, enhancement of catecholamine secretion, and modulation of calcium fluxes. Recently, theophylline was found to have several immunomodulatory and anti-inflammatory properties, and thus interest in its use in patients with asthma has been renewed. The use of theophylline in the treatment of asthma and chronic obstructive pulmonary disease has diminished with the advent of new medications, but theophylline remains beneficial, especially in the patient with difficult refractory symptoms. In the future, theophylline may be used as treatment for bradyarrhythmias after cardiac transplantation, prophylactic medication to reduce the severity of nephropathy associated with intravenous administration of contrast material, therapy for breathing problems during sleep, and treatment for leukemias.

Apnea↗
PubMed1998

Dialysis of the rectum for sampling drug concentrations in the luminal extracellular fluid of the gut: technique and precision.

BACKGROUND: It is useful to measure the luminal concentration of drugs which act in the gut. Dialysis of the rectum has not previously been used or validated for this purpose. AIM: To determine the precision of rectal dialysis for measuring rectal drug concentrations. METHODS: To establish the duration of dialysis required to approach equilibrium, the rate of methotrexate diffusion into dialysis bags was first determined in vitro. The precision of rectal dialysis for sampling the methotrexate concentration of colonic lumen extracellular fluid was determined in seven subjects who underwent two consecutive dialysis procedures. Subjects treated with subcutaneous methotrexate for refractory inflammatory bowel disease were studied. RESULTS: Methotrexate crossed the dialysis membrane by a first-order process, and after a 2 h in vitro dialysis, equilibration was 74 +/- 2% (mean +/- s.d.) complete. Rectal dialysis was well tolerated by all subjects. The mean +/- s.e. methotrexate concentration of 3.6 +/- 1.1 nmol/L in the first dialysate was not significantly different from 3.6 +/- 0.9 nmol/L in the second dialysate. P = 0.99 (paired two-tailed t-test). Similar precision was obtained for an endogenous molecule, potassium, secreted by the rectal mucosa. CONCLUSIONS: Dialysis of the rectum is a well tolerated and precise technique for sampling the colonic lumen extracellular fluid for quantitative analyses of exogenous and endogenous substances.

Antirheumatic Agents↗
PubMed1997

Inhibition of human mitochondrial aldehyde dehydrogenase by the disulfiram metabolite S-methyl-N,N-diethylthiocarbamoyl sulfoxide: structural characterization of the enzyme adduct by HPLC-tandem mass spectrometry.

S-Methyl-N,N-diethylthiocarbamoyl sulfoxide (MeDTC-SO) is a known metabolite of the aversion therapy drug disulfiram (DSF). MeDTC-SO is also a potent inhibitor of human mitochondrial aldehyde dehydrogenase (hmALDH) with an IC50 of 1.5 microM. Inhibition of the enzyme by MeDTC-SO resulted in the addition of approximately 100 Da to the molecular mass of the intact protein, as determined by on-line HPLC-electrospray ionization MS (LC-MS). Dialysis of the inhibited protein did not reverse the inhibition, and the molecular mass of 54,533 Da (+/- 0.01%) remained unchanged, indicating that a covalent modification of the protein had occurred. Proteolytic digestion of hmALDH under basic conditions using trypsin at pH 7.8 revealed that the adduct was base labile. However, treating the adducted protein with endopeptidase-Glu-C at pH 3.7 produced a peptide adduct at MH+ = 4924, tentatively attributable to a carbamoylated peptide. This peptide contains three adjacent cysteines, one of which has been implicated as a key amino acid in the highly conserved active site region of ALDH. A pepsin digestion of hmALDH carried out at pH 3.7 and subsequent LC-MS analysis revealed an ion at MH2(2+) = 501.5, corresponding to the carbamoylated peptide FNQGQC1C2C3. This peptide contains the same adjacent active site cysteines. This latter peptide was subjected to LC-MS/MS, which enabled us to determine that the site of carbamoylation was at Cys2. The MS/MS product ion data also confirmed the presence of a carbamoyl group as the adduct species.

Aldehyde Dehydrogenase↗
PubMed1997

Inhibition of recombinant human mitochondrial and cytosolic aldehyde dehydrogenases by two candidates for the active metabolites of disulfiram.

We expressed recombinant human cytosolic (ALDH1, high Km) and mitochondrial aldehyde dehydrogenase (ALDH2, low Km) in Escherichia coli and purified the enzymes to homogeneity to examine the nature of inhibition of human ALDH by disulfiram, its confirmed metabolite S-methyl N,N-diethylthiocarbamate (MeDTC) sulfoxide, and its proposed metabolite MeDTC sulfone. Disulfiram, MeDTC sulfoxide, and MeDTC sulfone, respectively, were potent inhibitors with IC50 values of 0.15 +/- 0.02 microM, 0.27 +/- 0.04 microM, and 0.12 +/- 0.02 microM for ALDH1, and 1.45 +/- 0.40 microM, 1.16 +/- 0.56, and 0.40 +/- 0.10 microM for ALDH2. Extensive dialysis did not restore the activity of the inactivated enzyme, indicating irreversible inhibition. Both the esterase and dehydrogenase activities of ALDH2 were inhibited to the same extent by MeDTC sulfone and sulfoxide, suggesting that both catalytic sites are closely linked. The time course of inhibition of ALDH appeared to be first-order for both MeDTC sulfone and MeDTC sulfoxide. Kitz and Wilson plots of the half-life of inactivation versus 1/[inhibitor] indicated that the reactions between ALDH and inhibitors were bimolecular. The pseudobimolecular rate constants (k3/KI) for the ALDH-inhibitor reactions were 1 x 10(5), 1 x 10(4), 3 x 10(3), and 1 x 10(3) s-1 M-1 ALDH1-sulfone, ALDH1-sulfoxide, ALDH2-sulfone, and ALDH2-sulfoxide, respectively. ALDH2 was not significantly protected from inactivation from either MeDTC sulfoxide or MeDTC sulfone by NAD alone, but high concentrations of NAD and acetaldehyde completely prevented inhibition. Since disulfiram is rapidly metabolized in vivo, it is believed that disulfiram is too short-lived to inhibit ALDH directly. The results of our study indicate that MeDTC sulfoxide and sulfone are potent inhibitors of human ALDH and are reasonable candidates for the proximal inhibitors of ALDH following disulfiram administration.

Aldehyde Dehydrogenase↗
PubMed1997

Cloning of a cDNA encoding a constitutively expressed rat liver cytosolic aldehyde dehydrogenase.

The presence of a constitutively expressed aldehyde dehydrogenase (ALDH) in the rat liver cytosol is controversial (Tottmar et al., 1973; Lindahl and Evces, 1984; Berger and Weiner, 1977; Tank et al., 1981; Truesdale-Mahoney et al., 1981; Cao et al., 1989). A cDNA encoding a constitutively expressed rat liver cytosolic class 1 ALDH was cloned using a PCR-based strategy. The open reading frame consisted of 1503 nucleotides which encoded a protein of 501 amino acids. In order to compare the rat and human nucleotide sequences, we sequenced the entire open reading frame of a human liver cytosolic ALDH cDNA clone (Zheng et al., 1993). Rat liver constitutively expressed cytosolic ALDH was 99.7, 91.8, 89.0, and 83.8% identical to rat kidney, mouse liver, rat liver phenobarbital-inducible, and human liver cytosolic class 1 ALDH cDNAs, respectively. Northern blot analysis indicated that constitutively expressed rat cytosolic ALDH mRNA is expressed in lung, kidney, liver, skeletal muscle, and testis, with weak expression in heart and brain. These results strongly suggest that a constitutively expressed ALDH is present in rat liver cytosol.

Aldehyde Dehydrogenase↗