Biomedical subjects

T Burckin

Publications and source records attributed to T Burckin.

6 recordsLinked to original sources

PubMed2001

Fully modified 2' MOE oligonucleotides redirect polyadenylation.

Many genes have been described and characterized that have alternative polyadenylation signals at the 3'-end of their pre-mRNAs. Many of these same messages also contain destabilization motifs responsible for rapid degradation of the mRNA. Polyadenylation site selection can thus determine the stability of an mRNA. Fully modified 2'-O:-methoxy ethyl/phosphorothioate oligonucleotides that hybridize to the 3'-most polyadenylation site or signal of E-selectin were able to inhibit polyadenylation at this site and redirect it to one of two upstream cryptic sites. The shorter transcripts produced after antisense treatment have fewer destabilization sequences, increased mRNA stability and altered protein expression. This study demonstrates that antisense oligonucleotides can be successfully employed to redirect polyadenylation. This is the first demonstration of the use of oligonucleotides to increase, rather than decrease, abundance of a message.

3' Untranslated Regions↗

PubMed2001

Comparison of pharmacokinetics and tissue disposition of an antisense phosphorothioate oligonucleotide targeting human Ha-ras mRNA in mouse and monkey.

The plasma pharmacokinetics and tissue disposition of ISIS 2503 were studied in mice following single and multiple bolus intravenous (iv) injections of 1-50 mg/kg, and in monkeys following single and multiple 2-h iv infusions of 1-10 mg/kg and bolus iv injections of 1 mg/kg of ISIS 2503. ISIS 2503 and its metabolites were measured in plasma, urine, and tissues using solid-phase extraction followed by capillary gel electrophoresis (CGE). In both species, the plasma clearance of ISIS 2503 was characterized by rapid distribution to tissues, and to a lesser extent, metabolism. The plasma clearance in mice was at least two-fold more rapid than in monkeys at equivalent doses. The plasma disposition (t1/2) increased with dose. The highest concentrations of oligonucleotide were consistently observed in the kidney and liver in both species. At equivalent doses, tissue concentrations in monkeys were much higher than tissue concentrations in mice. Urinary excretion of total oligonucleotide was a minor elimination pathway in both species at doses < 10 mg/kg. However, urinary excretion of total oligonucleotide in mice was increased to 12-29% as dose increased from 20 to 50 mg/kg.

Animals↗

PubMed2001

Absolute bioavailability of 2'-O-(2-methoxyethyl)-modified antisense oligonucleotides following intraduodenal instillation in rats.

Three modified 20-mer antisense oligonucleotides targeted to human intercellular adhesion molecule-1 mRNA were characterized for their presystemic stability and oral bioavailability compared with a first-generation phosphorothioate oligodeoxynucleotide (PS ODN), ISIS 2302. The three modified oligonucleotides contained 2'-O-(2-methoxyethyl) (2'-O-MOE) ribose sugar modifications on a portion, or on all of the nucleotides in the antisense sequence. In vitro metabolism studies conducted in various gastrointestinal and digestive tissue preparations indicated substantial improvement in stability of 2'-O-MOE-modified oligonucleotides. In addition, in vivo presystemic stability of these oligonucleotides was monitored in rats following intraduodenal administration. By 8 h after administration, only chain-shortened metabolites of the PS ODN were recovered in the gastrointestinal contents. In contrast, approximately 50% of the 2'-O-MOE ribose-modified (partial) compound remained intact (20-mer) by 8 h following administration. Both of the fully modified compounds (2'-O-MOE PO and PS) were completely stable with no measurable metabolites observed within 8 h of administration. The rank order of bioavailability was ISIS 11159 (full PS, full MOE) < ISIS 2302 (PS ODN) < ISIS 16952 (full PO, full MOE) < ISIS 14725 (full PS, partial MOE); the absolute plasma concentration bioavailability was measured in reference to intravenous dosing in the rat and was estimated at 0.3, 1.2, 2.1, and 5.5%, respectively. The optimal oligonucleotide chemistry for improved permeability and resulting bioavailability was the partially modified 3' hemimer 2'-O-MOE phosphorothioate oligonucleotide (ISIS 14725). Improved presystemic stability coupled with improved permeability were likely responsible for the remarkable improvement in the oral bioavailability of this compound.

Animals↗

PubMed2000

Comparison of the pharmacokinetics of subcutaneous and intravenous administration of a phosphorothioate oligodeoxynucleotide in cynomolgus monkeys.

The pharmacokinetics of subcutaneous (s.c.) administration of a phosphorothioate oligodeoxynucleotide (PS-ODN) was evaluated in cynomolgus monkeys. In a single dose study, monkeys were injected s.c. or intravenously (i.v.) with doses of either 1 or 5 mg/kg ISIS 2302. The bioavailability of s.c. injection ranged from 26% to 55% and appeared to be dependent on the concentration of the dosing solution rather than the dose. The bioavailability of a subcutaneously administered 5 mg/kg dose of ISIS 2302 was 55% using a 50 mg/ml dosing solution and only 26% using a 10 mg/ml dosing solution. Slow absorption from the s.c. injection site significantly blunted the maximal concentration (Cmax) compared with i.v. administration. The time to peak plasma concentration (Tmax) increased slightly with increasing dose, from 0.5 to 1 hour for the 1 mg/kg dose to 1 to 2.5 hours for the 5 mg/kg dose. Plasma half-lives were prolonged after s.c. administration, indicating more dependence on absorption than elimination. The half-lives after s.c. administration averaged 3 hours, whereas after i.v. administration, the half-lives were <1 hour. Metabolism of the ISIS 2302 after s.c. injection was consistent with exonucleolytic cleavage, as previously observed after i.v. administration. In summary, s.c. administration of PS-ODN resulted in prolonged and extensive absorption of the ODN.

Animals↗

PubMed1997

Pharmacokinetics and metabolism in mice of a phosphorothioate oligonucleotide antisense inhibitor of C-raf-1 kinase expression.

The plasma and tissue disposition of CGP 69846A (ISIS 5132) was characterized in male CD-1 mice following iv bolus injections administered every other day for 28 days (total of 15 doses). The doses ranged from 0.8 mg/kg to 100 mg/kg. Urinary excretion of oligonucleotide was also monitored over a 24-hr period following single dose administration over the same dose range. Pharmacokinetic plasma profiles were determined following single dose administration (dose 1) and after multiple doses (dose 15) at doses of 4 and 20 mg/kg. Concentrations in kidney, liver, spleen, heart, lung, and lymph nodes were characterized following doses 1, 8, and 15 for all doses. Capillary gel electrophoresis was used to quantitate intact (full-length) oligonucleotide and its metabolites (down to N - 11 base deletions) in both plasma and tissue at all time points. The plasma and tissue disposition of CGP 69846A was characterized by a rapid distribution into all tissues analyzed. Rapid plasma clearance of the parent oligonucleotide (9.3-14.3 ml/min/kg) was predominantly the result of distribution to tissue and, to a lesser extent, metabolism. Appearance and pattern of chain-shortened metabolites seen in plasma and tissue were consistent with predominantly exonuclease-mediated base deletion. No measurable accumulation of oligonucleotide was observed in plasma following multiple-dose administration, but both the liver and the kidney exhibited 2-3-fold accumulations. In general, the tissues exhibited half-lives for the elimination of parent oligonucleotide of 16-60 hr compared with plasma half-lives of 30-45 min. After repeated administrations, significant decreases in plasma clearance and volume of distribution at steady state (Vss) were observed following dose 15 at the dose of 20 mg/kg but not at the dose of 4 mg/kg. Changes in tissue accumulation and evidence for saturation of tissue distribution at the high doses may explain the plasma disposition changes observed in the absence of alteration of metabolism or plasma accumulation. Urinary excretion was a minor pathway for elimination of oligonucleotide over the 24-hr period immediately following iv administration. However, the amount of oligonucleotide excreted in the urine increased as a function of dose from less than 1% to approximately 13% of the administered dose over a dose range of 0.8 mg/kg to 100 mg/kg.

Animals↗

PubMedDate not supplied

Evaluation of the renal effects of an antisense phosphorothioate oligodeoxynucleotide in monkeys.

Antisense phosphorothioate oligodeoxynucleotides are therapeutic agents that provide target specificity resulting from Watson-Crick base pairing. However, there are nonspecific effects that in some instances result in toxicity. These compounds accumulate in the kidney and induce renal proximal tubular degeneration at high doses. The relationship between accumulation of phosphorothioate oligodeoxynucleotides in the kidney, indicators of renal toxicity, and histomorphology were investigated in rhesus monkeys. Monkeys received vehicle or an escalating dose regimen of 3, 10, 40, and 80 mg/kg of ISIS 2105 and were then evaluated for changes in clinical pathology indices, urinalysis parameters, and renal histopathology. Urinalysis revealed an increase in protein levels and a slight increase in blood content following the third 40 mg/kg dose and continuing through the 80 mg/kg doses, whereas other urinary markers of renal toxicity were unchanged. Creatinine clearance was slightly decreased in monkeys during the 80 mg/kg dose cycle. Granulation in the cytoplasm of proximal tubular epithelial cells was evident by microscopic examination of kidney and was present at all doses examined and increased with dose. Immunohistochemical staining localized the oligodeoxynucleotide within these granules. Histopathologic changes consisting of minimal to moderate tubular degeneration were present only at the higher doses of 40 and 80 mg/kg and at high tissue concentrations, and these changes occurred concurrent with functional alterations, whereas lower doses (< or = 10 mg/kg) did not affect a pathologic or functional change.

Animals↗