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PubMed2001

Pharmacokinetics and pharmacodynamics of an antisense phosphorothioate oligonucleotide targeting Fas mRNA in mice.

ISIS 22023 is a modified phosphorothioate antisense oligonucleotide targeting murine Fas mRNA. Treatment of mice with ISIS 22023 reduced Fas expression in liver in a concentration-dependent and sequence-specific manner, which completely protected mice from fulminant death induced by agonistic Fas antibody. In this study, we characterized the relationships in mice between total dose administered, dose to the target organ, and ultimately, the intracellular concentration within target cell types to the pharmacologic activity of ISIS 22023. After subcutaneous injection, ISIS 22023 distributed to the liver rapidly and remained in the liver with the t(1/2) ranging from 11 to 19 days, depending on dose. There were apparent differences in patterns of uptake and elimination in different types of liver cells. Oligonucleotide appeared within hepatocytes rapidly, whereas the peak concentrations in Kupffer cells were delayed until 2 days after dose administration. Hepatocytes cleared oligonucleotide the most rapidly, whereas Kupffer cells appeared to retain oligonucleotide longer. The reduction of Fas mRNA levels (pharmacodynamic response) paralleled the increase of oligonucleotide concentration in mouse liver with maximum mRNA reduction of 90% at 2 days after a single 50 mg/kg subcutaneous administration. Moreover, the pharmacodynamics of ISIS 22023 correlated better with the pharmacokinetics in hepatocytes, supporting the concept that the presence of oligonucleotide in target cells results in reductions in mRNA and, ultimately, pharmacologic activity. These results provide a comprehensive understanding of the kinetics of an antisense drug at the site of action and demonstrate that the reductions in mRNA induced by this antisense oligonucleotide correlate with its concentrations in cell targets.

Algorithms↗
PubMed1998

In vivo delivery of phosphorothioate oligonucleotides into murine retina.

OBJECTIVES: To determine the fate of phosphorothioate oligonucleotides (S-ODNs), which are commonly used for antisense strategy, in murine retina in vivo with the use of fluorescein isothiocyanate (FITC)-labeled S-ODNs, and to evaluate our fusogenic liposome system that may facilitate the delivery of S-ODNs. METHODS: The FITC-labeled S-ODNs were encapsulated in liposomes, which were then coated with the envelope of inactivated hemagglutinating virus of Japan (HVJ; Sendai virus) by fusion (HVJ liposomes). Intravitreal injection of naked FITC-labeled S-ODNs or of the HVJ liposomes was done in ICR mice. After fixation, cryosections and flat-mounted retinas were prepared and examined by fluorescence microscopy. RESULTS: Injection of naked FITC-labeled S-ODNs at 3 micromol/L exhibited weak fluorescence in 13% of the cells in the ganglion cell layer. When the concentration was increased to 30 micromol/L, high fluorescence was seen in 59% of cells in the ganglion cell layer at this time. This fluorescence diminished within a day. In contrast, injection of HVJ liposomes containing FITC-labeled S-ODNs at 3 micromol/L resulted in high fluorescence in 44% of the cells in the ganglion cell layer at 1 hour, and this fluorescence lasted for up to 3 days. This treatment also resulted in high fluorescence within retinal vessel walls, and weak fluorescence in photoreceptor cells. CONCLUSIONS: Intravitreally injected S-ODNs were rapidly eliminated from neural retina, and the use of HVJ liposomes could improve the delivery of S-ODNs. This method may be a potentially useful system for the antisense-based therapies for retinal diseases.

Animals↗
PubMed1996

Transforming growth factor-beta-mediated autocrine growth regulation of gliomas as detected with phosphorothioate antisense oligonucleotides.

Transforming growth factors-beta 1 and -beta 2 (TGF-beta 1 and -beta 2) are important growth-regulatory proteins for astroglial neoplasms. We analyzed their role in tumor-cell proliferation in 12 glioma cell lines, employing phosphorothioate antisense oligodeoxynucleotides (S-ODNs, 14 mer), specifically targeted against the coding sequences of TGF-beta 1-mRNA and TGF-beta 2-mRNA. TGF-beta 1-S-ODNs inhibited cell proliferation in 5 of 12 gliomas, whereas TGF-beta 2-S-ODNs reduced the cell proliferation in all glioma cell lines, compared to nonsense-S-ODN-treated and S-ODN-untreated cells as controls. The efficacy and specificity of antisense effects was validated by Northern-blot analysis and determination of protein concentrations in culture supernatants (ELISA). Exogenous hrTGF-beta 1 either stimulated or inhibited the cell lines, whereas pnTGF-beta 2 stimulated the proliferation of most glioma cells. Blocking the extracellular pathway of TGF-beta by neutralizing antibodies only slightly inhibited those cell lines, which were markedly stimulated by TGF-betas. As the effects of TGF-beta 2-S-ODNs were much stronger than those of TGF-beta neutralizing antibodies, we postulate that the endogenously produced TGF-beta 2 control glioma-cell proliferation, in part by an intracellular loop.

Base Sequence↗
PubMed1998

Antisense phosphorothioate oligodeoxynucleotide down-regulation of the insulin-like growth factor I receptor in ovarian cancer cells.

The insulin-like growth factors (IGF-I and IGF-II) play a key role in cellular proliferation and are involved in cellular transformation. The expression of the IGF-I receptor has been demonstrated in a variety of human tumor cell lines including ovarian cancer cells. Phosphorothioate antisense oligodeoxynucleotides (S-ODNs) were analyzed for their potential to suppress the IGF-I receptor in the NIH: OVCAR-3 ovarian cancer cell line. The application of the antisense S-ODN reduced potently the cell growth of unstimulated NIH:OVCAR-3 cells, whereas sense and mismatch S-ODNs were without any effect. This effect resembled that of the monoclonal antibody (MAb) alphaIR-3. In contrast to the antisense compound, this MAb only partially inhibited the IGF-I-induced proliferation of ovarian cancer cells. The concentration of the antisense S-ODN to exhibit an identical inhibition of cell proliferation was reduced to 50 nM when the oligonucleotides were delivered by the cationic lipid formulation lipofectin. The specificity of the antisense S-ODN action was confirmed by reduction of the receptor protein and of the receptor mRNA, as assayed by flow cytometry and by Northern blot hybridizations. Our data demonstrate the potency of antisense S-ODNs to target the IGF-I receptor message and show that antisense strategies against the IGF-I receptor may provide new strategies for the therapy of ovarian cancer.

Cell Division↗
PubMed1992

Organ distribution and stability of phosphorothioated oligodeoxyribonucleotides in mice.

Results of recent studies in our laboratory have suggested a potential role for antisense oligodeoxyribonucleotides (oligo(dN)s) as therapeutic agents in the treatment of human hepatitis B virus infection. As a first step towards assessing the potential utility of oligo(dN) in therapy, we have examined the organ distribution, stability and toxicity of a phosphorothioated oligo(dN) (S-oligo) of 20 nucleotides in length which was administered to mice via different routes. Among the various organs analysed, the liver retained the highest amount of S-oligo (1.3-2 per cent of the total injection) at the peak time (10-30 min) regardless of the route of injection. However, the S-oligo appeared to be degraded in the liver to about 40 per cent of its original length within 30 min of injection, presumably by the action of 3' exonucleases. Injection of doses of up to 5 mg kg-1 of S-oligo had no apparent toxic effects on the mice.

Animals↗
PubMed2006

Maternal and fetal distribution of a phosphorothioate oligonucleotide in rats after intravenous infusion.

BACKGROUND: Fetal uptake of an antisense oligonucleotide was evaluated after intravenous (i.v.) dosing of ISIS 2105, a 20-base phosphorothioate oligonucleotide, in timed-pregnant Sprague-Dawley rats. METHODS: To maximize the potential for fetal exposure, ISIS 2105 was administered as a 3-hr infusion at 6.6 mg/kg/hr with a total dose of 20 mg/kg, or as a continuous 7-day infusion at 0.35 mg/kg/hr with a total dose of 59 mg/kg. This dosing regime is higher than a patient would be expected to receive in the clinical use of oligonucleotides. Infusions were delivered through a jugular vein cannula by syringe pump on gestation day (GD) 19 (3-hr exposure) or by osmotic pumps implanted subcutaneously (s.c.) starting on GD 12 (7-day exposures). RESULTS: After a 3-hr infusion, maternal and fetal plasma concentrations of ISIS 2105 were >100 microg/ml and <0.07 microg/ml, respectively with a maternal fetal ratio of >1,000. Maternal regions of the placenta had twice the oligonucleotide concentration compared to fetal regions of the placenta (6 microg/g vs. 3 microg/g). After this acute exposure the concentrations in fetal kidney and liver were approximately 140- and 500-fold less than the maternal kidney and liver respectively. After 7-day infusion maternal plasma concentrations were 0.82 microg/ml and fetal concentrations were <0.22 microg/ml. By capillary gel electrophoresis (CGE) only the fetal liver consistently had quantifiable oligonucleotide concentrations (range=1.01-4.95 microg/g) compared to a mean concentration of 50.11+/-1.71 microg/g in the maternal liver a maternal to fetal ratio of approximately 10:50 after 7 days of infusion. CONCLUSIONS: There was a low level of transfer from dam to fetus, consistent with a slow equilibrium but the permeability of placenta to this 6 kDa polyanionic compound seemed to be limited even at supraclinical doses.

Animals↗
PubMed2004

Complex formation and vectorization of a phosphorothioate oligonucleotide with an amphipathic leucine- and lysine-rich peptide: study at molecular and cellular levels.

Optical spectroscopic techniques such as CD, Raman scattering, and fluorescence imaging allowed us to analyze the complex formation and vectorization of a single-stranded 20-mer phosphorothioate oligodeoxynucleotide with a 15-mer amphipathic peptide at molecular and cellular levels. Different solvent mixtures (methanol and water) and molecular ratios of peptide/oligodeoxynucleotide complexes were tested in order to overcome the problems related to solubility. Optimal conditions for both spectroscopic and cellular experiments were obtained with the molecular ratio peptide/oligodeoxynucleotide equal to 21:4, corresponding to a 7:5 ratio for their respective +/- charge ratio. At the molecular level, CD and Raman spectra were consistent with a alpha-helix conformation of the peptide in water or in a methanol-water mixture. The presence of methanol increased considerably the solubility of the peptide without altering its alpha-helix conformation, as evidenced by CD and Raman spectroscopies. UV absorption melting profile of the oligodeoxynucleotide gave rise to a flat melting profile, corresponding to its random structure in solution. Raman spectra of oligodeoxynucleotide/peptide complexes could only be studied in methanol/water mixture solutions. Drastic changes observed in Raman spectra have undoubtedly shown: (a) the perturbation occurred in the peptide secondary structure, and (b) possible interaction between the lysine residues of the peptide and the oligodeoxynucleotide. At the cellular level, the complex was prepared in a mixture of 10% methanol and 90% cell medium. Cellular uptake in optimal conditions for the oligodeoxynucleotide delivery with low cytotoxicity was controlled by fluorescence imaging allowing to specifically locate the compacted oligonucleotide labeled with fluorescein at its 5'-terminus with the peptide into human glioma cells after 1 h of incubation at 37 degrees C.

Biological Transport↗
PubMed2006

Cellular uptake of phosphorothioate oligonucleotide facilitated by cationic porphyrin: a microfluorescence study.

Cationic 5,10,15,20-tetrakis (1-methyl-4-pyridyl) porphyrin was tested as a delivery agent for oligonucleotides. By using fluorescence microimaging, it has been shown that complexation of the porphyrin to the phosphorothioate analog of dT(15) labeled by rhodamine enabled its nonendocytic penetration into the cell and regular distribution in the cytoplasm and preferentially into the nucleus. Time-resolved microfluorescence spectroscopy revealed that the oligonucleotide integrity was kept. A small fraction of the porphyrin molecules seems to undergo change of the binding mode after internalization, probably due to duplex formation between the oligonucleotide and its cellular target sequences, or due to dissociation of the porphyrin from the oligonucleotide and subsequent interactions in the cellular environment.

3T3 Cells↗
PubMed1995

Sequence specificity on the growth suppression and induction of apoptosis of chronic myeloid leukemia cells by BCR-ABL anti-sense oligodeoxynucleoside phosphorothioates.

Sequence specificity of inhibitory effects of various BCR-ABL anti-sense oligodeoxynucleoside phosphorothioates (AS PS-ODN) on the proliferation of the chronic myeloid-leukemia cell line BV173 was examined. We confirmed that 26, 18, and 16mer B2A2 AS PS-ODN had strong inhibitory effects on the proliferation of BV173 cells with B2A2 mRNA expression, and that B3A2 AS PS-ODN were equally inhibitory when cultures were initiated at lower cell concentrations. However, at higher cell concentrations, the inhibitory effects by B3A2 AS PS-ODN were reduced and B2A2 AS PS-ODNs could suppress the proliferation of BV173 cells with much more relative sequence specificity. The 26mer B2A2 AS PS-ODN induced apoptosis of BV173 cells following reduction of BCR-ABL mRNA expression and p210 protein synthesis. Various sense (S), reverse order, and random sequences had no inhibitory effects except 16mer B2A2 S and B3A2 S that revealed significant suppressive effects. Furthermore, 26mer B3A2 AS also reduced B2A2 mRNA expression and p210 protein synthesis, while 16mer S sequences did not. These results suggest that B2A2 AS may be cross-reactive with B3A2 AS on the growth suppression of CML cells under certain culture conditions, possibly due to their partial hybridization to the ABL portion of the target mRNA, although other non-sequence-specific mechanisms are also possible.

Base Sequence↗
PubMed1995

c-erbB-2 anti-sense phosphorothioate oligodeoxynucleotides inhibit growth and serum-induced cell spreading of P185c-erbB-2-overexpressing ovarian carcinoma cells.

Overexpression of the c-erbB-2 proto-oncogene product (p185c-erbB-2) occurs frequently in different types of human cancer and is correlated with a significantly decreased survival in ovarian cancer patients. The effect of c-erbB-2 anti-sense phosphorothioate oligodeoxynucleotides (S-ODNs) was examined on the ovarian cancer cell line SK-OV-3. p185c-erbB-2 levels were specifically reduced by a single-dose application of 5 microM c-erbB-2 anti-sense S-ODNs. This was accompanied by a 60% inhibition of anchorage-dependent cell growth. More strikingly, c-erbB-2 anti-sense S-ODNs almost completely abrogated serum-induced cell spreading. A control of complementary sense oligodeoxynucleotides did not show significant inhibitory effects on cell growth or on cell spreading. The inhibition of cell spreading was imitated by a monoclonal antibody (9G6) targeting the extracellular domain of p185c-erbB-2 and by the tyrosine kinase inhibitor erbstatin. The inhibitory activity of these 2 compounds was lost after a few hours, while the inhibition of serum-induced cell spreading by anti-sense S-ODNs was still present after 24 hr. Our results show that c-erbB-2 anti-sense S-ODNs effectively inhibit the mitogenic and spreading activity of p185c-erbB-2 in ovarian cancer cells. Thus, anti-sense strategies have the potential of providing new strategies for the therapy of ovarian cancer.

Antineoplastic Agents↗
PubMed2003

TGF-beta1-induced Smad 3 binding to the Smad 7 gene: knockout of Smad 7 gene transcription by sense phosphorothioate oligos, autoregulation, and effect on TGF-beta1 secretion: bleomycin acts through TGF-beta1.

Bleomycin produces its fibrogenic effect, at least in part, by TGF-beta1 secretion. Treatment of IMR-90 human embryonic lung fibroblasts with bleomycin at 0.5 microg/ml results in a 1.6-fold increase of TGF-beta1 as determined by a specific ELISA assay for TGF-beta1 after acidification of the conditioned media. This elevation of TGF-beta1 secretion is furthermore enhanced in vivo by TGF-beta1 autoinduction of the TGF-beta1 gene. To demonstrate TGF-beta1 autoinduction, the fibroblasts were pretreated with 12.5 ng/ml TGF-beta1, washed extensively to remove any residual TGF-beta1, and then allowed to incubate for 24 h in AIM V synthetic serum-free media. The media when assayed using the ELISA assay contained a 1.6-fold increase of TGF-beta1. The distal promoter of the human TGF-beta1 gene contains a Smad 3 element (CAGGACA), which is homologous to the Smad 3 binding element motif (CAGA). The nuclear extracts of human embryonic lung fibroblasts treated for either 15 min or 24 h with TGF-beta1 did not demonstrate specificity of binding of a protein(s) to the homologous Smad 3 element as determined by cold wild-type oligodeoxynucleotide competition experiments. However, specific Smad 3 binding to the Smad 3 element (GTCTAGAC) found in proximal promoter of the Smad 7 gene was observed by cold oligo competition and supershift assays using a goat polyclonal Smad 3 antibody in the presence and absence of an N-terminal Smad 3 peptide. To determine the functionality of this Smad 3 binding to the Smad 3 element in the proximal promoter of the Smad 7 inhibitory gene to TGF-beta1 secretion, fibroblasts were transiently pretransfected with double-stranded phosphorothioate oligo "decoys" containing the Smad 7/Smad 3 element in the presence of plasmin to convert latent TGF-beta1 to active TGF-beta1. Under these conditions, which simulate the in vivo situation of 2.2-fold increase of total active TGF-beta1 was observed. Fibroblasts were also pretransfected with these double-stranded oligo "decoys," washed, then treated with TGF-beta1, washed and incubated in AIM V for an additional 24 h. In this latter experiment, a superinduction of TGF-beta1 secretion was observed. We propose that these oligo "decoys" bind Smad 3 preventing this initiation factor from binding to the Smad 7/Smad 3 element thereby decreasing the transcription of the Smad 7 gene. The decrease of the inhibitory Smad 7 would result in less binding of this Smad inhibitor to the Type I TGF-beta receptor and less antagonism of active TGF-beta1, more autoinduction of the TGF-beta1 gene, and more of the fibrogenic effects of TGF-beta1.

Bleomycin↗
PubMed1984

Sequential and dose-dependent alterations in rat bronchiolar epithelium during O,O,S-trimethyl phosphorothioate induced delayed toxicity.

The sequential and dose-dependent effects of O,O,S-trimethyl phosphorothioate (OOS-Me) on rat lung bronchiolar epithelium were investigated using scanning electron microscopy. At 12 h after oral treatment (20 mg/kg), there was a small increase in debris in the bronchioles and, by 24 h, there was much debris as well as phagocytic cells in the bronchioles. After 3 days, there was a decrease in the number of bronchiolar Clara cells as determined by the loss of their characteristic apical bulges. Concomitantly, there was a significant increase in lactate dehydrogenase activity in bronchopulmonary lavage fluid. By day 7, the Clara cells appeared to be reforming and, by day 14, the morphology of bronchiolar epithelium had returned to normal. Dose-dependent studies revealed a threshold dose level of OOS-Me between 10 and 20 mg/kg which produced the observed effects in Clara cells. Both sequential and dose-dependent effects of OOS-Me on Clara cells were correlated with changes in bronchopulmonary lavage lactate dehydrogenase activity.

Animals↗
PubMed1988

Pulmonary endothelial and alveolar epithelial lesions induced by O,O,S-trimethyl phosphorothioate in rats.

The morphogenesis of pulmonary injury induced by an impurity present in a commercially important organophosphorus insecticide, O,O,S-trimethyl phosphorothioate (OOS-TMP), was studied by combined light and transmission electron microscopy. Weanling female WAG/Rij rats received OOS-TMP dissolved in corn oil by gavage and were studied at intervals from 6 to 168 h after treatment. Sequestration of neutrophils was initially observed at 12 h after treatment and was accompanied by interstitial oedema. Plasmalemma alterations in endothelium lining capillaries and small arteries and veins were observed from 12 to 120 h after treatment and were accompanied by endothelial cell detachment and separation from the basal lamina. Abundant aggregates of fibrin were sequentially observed in intravascular, interstitial, and alveolar spaces. Platelet aggregation and degranulation were occasionally observed in capillaries as early as 12 h after treatment, and frequently observed in capillaries and small vessels from 24 to 96 h after treatment. Significant increases in wet lung weight and lung water content occurred at the same time that morphologic changes were observed in pulmonary endothelium. Alterations in type I alveolar epithelial cells were initially observed at 24 h after treatment. Cell swelling, fragmentation, and necrosis were observed in both type I and type II cells and resulted in a bare basal lamina. Marked attenuation, hypertrophy, and proliferation of type II epithelial cells followed alveolar epithelial cell injury and loss. Minimal changes were observed in non-ciliated bronchiolar epithelial (Clara) cells; predominant changes included the loss of surface microvilli and apical cytoplasmic bulge. The results of this study indicate that the endothelium and alveolar epithelium are the predominant cell types in the rat lung injured following OOS-TMP administration.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Separation and characterization of diastereoisomers of antisense oligodeoxyribonucleoside phosphorothioates.

Diastereoisomers of oligodeoxyribonucleoside phosphorothioates (OPT) up to a tetramer were effectively separated with reversed-phase high-performance liquid chromatography (reversed-phase HPLC) under optimized conditions. The diastereoisomers of OPT resulted in different retention times on the reversed-phase HPLC. From the results, we found that there were certain rules in the elution order of the diastereoisomers. The configurational sequence of the diastereoisomers was determined by digestion with nuclease P1, snake venom phosphodiesterase, and alkaline phosphatase. The diastereoisomers were studied by CD spectroscopy with respect to their conformation in aqueous media. We found that a large variation in the conformation of diastereoisomers exists. Results suggest that much attention should be paid to the diastereoisomerism in antisense molecules having chiral internucleotide linkages such as OPT.

Base Sequence↗
PubMed1996

Development of a novel scintillation proximity competitive hybridization assay for the determination of phosphorothioate antisense oligonucleotide plasma concentrations in a toxicokinetic study.

A novel scintillation proximity competitive hybridization assay was developed for determining plasma concentrations of compound 4003W94, a 15-base phosphorothioate antisense deoxyribonucleotide that is currently under preclinical evaluation for the treatment of restenosis following coronary artery angioplasty. The principle of the assay involves the hybridization binding of antisense (4003W94) to a biotinylated sense oligonucleotide to form a double-stranded nucleic acid complex on the surface of scintillation proximity beads derivatized with streptavidin. As in a competitive radioimmunoassay, there is an inverse relationship between the amount of radioactivity in the final binding complex and the amount of 4003W94 present in the sample being analyzed. Because this is a homogenous assay, no physical separation of bound from free radioligand is necessary. Conventional cross-reactivity studies with either 3'- or 5'-deletion oligomers of 4003W94 indicated that cross-reactivity generally decreased with each base deletion. The assay was used to determine plasma concentrations of 4003W94 equivalents in rhesus monkeys during an exploratory 14-day toxicity study. This method conceivably could be adapted for use as an effective in vitro screening tool for the identification of potential antisense oligonucleotide drug candidates or as a diagnostic tool for the detection of pathological disorders.

Analysis of Variance↗
PubMed1996

Phosphorothioate-capped antisense oligonucleotides to Ras GAP inhibit cell proliferation and trigger apoptosis but fail to downregulate GAP gene expression.

We have studied the effects of an antisense oligonucleotide to Ras GAP in leukaemia cell lines. When terminal phosphorothioate linkages were introduced into this oligonucleotide, it caused major growth inhibition and apoptosis in the chronic myeloid leukaemia (CML) cell line K562, but had little effect on the promyelocytic leukaemia cell line HL60. Neither the expression of Ras GAP mRNA nor p120 GAP protein was downregulated by the antisense oligonucleotide, suggesting a non-antisense mechanism for growth inhibition. The antisense oligonucleotide contained GGC triplets which have previously been reported to inhibit the activity of p210bcr-abl both in vitro and in vivo. However, cellular phosphotyrosine levels were found to be unaffected, suggesting that the activity of p210bcr-abl was normal and that the antisense oligonucleotide may be interacting aptamerically with a different cellular protein. Since K562 is very resistant to apoptotic cell death, the identity of the putative target molecule would be of considerable interest.

Apoptosis↗
PubMed1998

PAAn-1b and PAAn-E: two phosphorothioate antisense oligodeoxynucleotides inhibit human aromatase gene expression.

Estrogen-dependent diseases, especially breast cancers, are frequently treated with aromatase inhibitors. Another more recent strategy is the antisense technology. In this study, after predicting aromatase mRNA secondary structure, we describe the design, the efficiency, and the toxicity of two antisense phosphorothioate oligodeoxynucleotides (PAAn-1b and PAAn-E) directed toward aromatase mRNA. Indeed, 2 microM PAAn-1b and PAAn-E encapsulated with 54 microM polyethylenimine inhibit aromatase activity by 71 and 79%, respectively, in transfected 293 cells, with IC50 values of 0.2 and 0.6 microM. The mechanism of inhibition appears to be specific after using sense and scramble oligodeoxynucleotides as controls and largely decreases aromatase mRNA and protein amounts. Moreover, PAAn-1b and PAAn-E are not cytotoxic for 293 cells. This study finally provides a new strategy for aromatase inhibition. It offers new tools for studying aromatase gene expression and its role in cancer for instance, and this could be of help for the therapy of estrogen-dependent diseases.

Actins↗
PubMed1997

Inhibition of phenotype modulation, growth, and migration of vascular smooth muscle cells by a guanosine-rich 30-mer phosphorothioate oligodeoxynucleotide.

The testing of a 30-mer dG-rich phosphorothioate oligodeoxynucleotide (LG4PS) for effects on the behaviour of vascular smooth muscle cells (VSMC) in vitro and in vivo is described. LG4PS at 0.3 microM inhibited significantly the phenotype modulation of freshly isolated rabbit VSMC, and cell outgrowth from pig aortic explants was inhibited approximately 80% by 5 microM LG4PS. The growth of proliferating rabbit and pig VSMC was inhibited approximately 70% by 0.3 microM and 5 microM LG4PS, respectively. Though less marked, the antiproliferative effects of LG4PS on human VSMC were comparable to those obtained with heparin. The cytotoxic effects of LG4PS on VSMC in vitro were low. Despite these promising results, adventitial application of 2-200 nmol LG4PS in pluronic gel failed to reduce vascular hyperplasia in balloon-injured rabbit carotid arteries, and the highest dose caused extensive mortality.

Animals↗