In vivo rAAV-mediated human TGF-ß overexpression reduces perifocal osteoarthritis and improves osteochondral repair in a large animal model at one year.

OBJECTIVE: Osteoarthritis (OA) is a serious consequence of focal osteochondral defects. Gene transfer of human transforming growth factor beta (hTGF-ß) with recombinant adeno-associated virus (rAAV) vectors offers a strategy to improve osteochondral repair. However, the long-term in vivo effects of such rAAV-mediated TGF-ß overexpression including its potential benefits on OA development remain unknown. METHOD: Focal osteochondral defects in minipig knees received rAAV-lacZ (control) or rAAV-hTGF-ß in vivo. After one year, osteochondral repair and perifocal OA were visualized using validated macroscopic scoring, ultra-high-field MRI at 9.4 T, and micro-CT. A quantitative estimation of the cellular densities and a validated semi-quantitative scoring of histological and immunohistological parameters completed the analysis of microarchitectural parameters. RESULTS: Direct rAAV-hTGF-ß application induced and maintained significantly improved defect filling and safranin O staining intensity and overall cartilage repair at one year in vivo. In addition, rAAV-hTGF-ß led to significantly higher chondrocyte densities within the cartilaginous repair tissue without affecting chondrocyte hypertrophy and minimized subarticular trabecular separation. Of note, rAAV-hTGF-ß significantly improved the adjacent cartilage structure and chondrocyte density and reduced overall perifocal OA development after one year in vivo. CONCLUSIONS: rAAV-hTGF-ß treatment improves long-term osteochondral repair and delays the progression of perifocal OA in a translational model. These findings have considerable potential for targeted molecular approaches to treat focal osteochondral defects.

Alteration of sarcoplasmic reticulum Ca2+ ATPase expression in lower limb ischemia caused by atherosclerosis obliterans.

Atherosclerosis is a disease caused by a build-up of fatty plaques and cholesterol in the arteries. The lumen of the vessels is obliterated resulting in restricted blood supply to tissues. In ischemic conditions, the cytosolic Ca2+ level of skeletal muscle may increase, indicating the alteration of Ca2+ removal mechanisms. Ca2+ is transported from cytosol into the sarcoplasmic reticulum by Ca2+ ATPase (SERCA), with its 1a isoform expressed in adult, while its 1b isoform in neonatal and regenerating fast-twitch skeletal muscle. To investigate the role of these isoforms in ischemic skeletal muscle, biopsies from musculus biceps femoris of patients who underwent amputation due to atherosclerosis were examined. Samples were removed from the visibly healthy and hypoxia-affected tissue. Significantly increased SERCA1a expression was detected under the ischemic conditions (246 ± 69%; p < 0.05) compared with the healthy tissue. Furthermore, the ratio of SERCA1a-positive fibers was slightly increased (46 ± 4% in healthy tissue and 60 ± 5% in ischemic tissue; p > 0.05), whereas SERCA2a did not change. In addition, in primary cultures derived from hypoxia-affected tissue, the diameter and fusion index of myotubes were significantly increased (30 ± 1.6 µm vs. 41 ± 2.4 µm and 31 ± 4% vs. 45 ± 3%; p < 0.05). We propose that the increased SERCA1a expression indicates the existence and location of compensating mechanisms in ischemic muscle.

[Surgical treatment of acquired, benign esophago-respiratory fistulas]. / Die chirurgische Behandlung gutartiger Fisteln zwischen dem Osophagus und dem Respirationstrakt.

BACKGROUND: Acquired, benign esophago-respiratory fistula occurs as a rare complication of different disorders. STUDY DESIGN: Over a period of 30 years, 37 patients with acquired, benign esophago-respiratory fistulas were treated at the authors' institutions. 35 of the 37 patients were operated upon. Acute operation involved primary repair of the trachea and esophagus, or intubation of the gullet and/or gastrostomy. Elective operations were performed on 13 patients. RESULTS: Complications were observed in 14 patients. There were 4 cases with anastomotic leakage (all of them on the neck). Direct closure was carried out in 2 patients; the remaining 2 cases healed spontaneously. None of the 4 patients died. Seven of the 37 patients died (18.9%). This seems acceptable in view of the severity of the cases and the extension of the operations. CONCLUSIONS: 1. The causes of a majority of the esophago-respiratory fistulas in this material are connected with the complications of corrosive injury and peptic stricture of the esophagus. 2. The well-known fact that most of these fistulas occur in the right chest is confirmed. 3. Gastrostomy and/or intubation of the gullet seems to be the best solution for this lesion in the acute cases and in severely ill patients. 4. In elective cases, reconstruction is suggested, if possible in one step.

Plasma timolol concentrations of timolol maleate: timolol gel-forming solution (TIMOPTIC-XE) once daily versus timolol maleate ophthalmic solution twice daily.

PURPOSE: The objective of this study was to compare plasma concentrations of timolol following multiple dosing of the therapeutic regimens of timolol maleate ophthalmic gel-forming solution (Timolol GS; TIMOPTIC-XE) and timolol maleate ophthalmic solution. Timolol maleate ophthalmic gel-forming solution is also referred to as Timolol GS, i.e. gel-forming solution. METHODS: This was a masked observer, two-period crossover study in six normal male subjects randomized to receive either Timolol GS, 0.5% (TIMOPTIC-XE,) once daily (0530 hours) or timolol maleate ophthalmic solution (0.5% TIMOPTIC) twice daily (0530 and 1730 hours) for 8 days, in both eyes. On Day 8, a blood sample was obtained prior to treatment, as well as 1, 2, 4, 8, 10, 12, 13, 14, 16, and 24 hours following the morning instillation. After a 7-day inter-period washout interval, subjects received the opposite treatment. RESULTS: Timolol GS (TIMOPTIC-XE): Plasma concentrations of timolol rarely exceeded 0.375 ng/ml (the lower limit of assay quantification). For all subjects, peak plasma concentrations of timolol averaged <0.3 ng/ml within 4 hours after the last dose. The highest single observation was 0.49 ng/ml in one subject (at hour 2). Timolol solution: For all subjects, peak plasma concentrations of timolol averaged about 0.5 ng/ml and 0.3 ng/ml within 4 hours following the first and second dose, respectively, on Day 8. The highest single observation was 0.95 ng/ml in one subject (at hour 2). CONCLUSIONS: The data suggest that there is less systemic exposure to timolol following once-daily therapy with Timolol GS 0.5% compared with twice daily therapy with timolol maleate ophthalmic solution 0.5%.

Influence of beta-adrenoceptor antagonists on the pharmacokinetics of rizatriptan, a 5-HT1B/1D agonist: differential effects of propranolol, nadolol and metoprolol.

AIMS: Patients with migraine may receive the 5-HT1B/1D agonist, rizatriptan (5 or 10 mg), to control acute attacks. Patients with frequent attacks may also receive propranolol or other beta-adrenoceptor antagonists for migraine prophylaxis. The present studies investigated the potential for pharmacokinetic or pharmacodynamic interaction between beta-adrenoceptor blockers and rizatriptan. METHODS: Four double-blind, placebo-controlled, randomized crossover investigations were performed in a total of 51 healthy subjects. A single 10 mg dose of rizatriptan was administered after 7 days' administration of propranolol (60 and 120 mg twice daily), nadolol (80 mg twice daily), metoprolol (100 mg twice daily) or placebo. Rizatriptan pharmacokinetics were assessed. In vitro incubations of rizatriptan and sumatriptan with various beta-adrenoceptor blockers were performed in human S9 fraction. Production of the indole-acetic acid-MAO-A metabolite of each triptan was measured. RESULTS: Administration of rizatriptan during propranolol treatment (120 mg twice daily for 7.5 days) increased the AUC(0, infinity) for rizatriptan by approximately 67% and the Cmax by approximately 75%. A reduction in the dose of propranolol (60 mg twice daily) and/or the incorporation of a delay (1 or 2 h) between propranolol and rizatriptan administration did not produce a statistically significant change in the effect of propranolol on rizatriptan pharmacokinetics. Administration of rizatriptan together with nadolol (80 mg twice daily) or metoprolol (100 mg twice daily) for 7 days did not significantly alter the pharmacokinetics of rizatriptan. No untoward adverse experiences attributable to the pharmacokinetic interaction between propranolol and rizatriptan were observed, and no subjects developed serious clinical, laboratory, or other significant adverse experiences during coadministration of rizatriptan with any of the beta-adrenoceptor blockers. In vitro incubations showed that propranolol, but not other beta-adrenoceptor blockers significantly inhibited the production of the indole-acetic acid metabolite of rizatriptan and sumatriptan. CONCLUSIONS: These results suggest that propranolol increases plasma concentrations of rizatriptan by inhibiting monoamine oxidase-A. When prescribing rizatriptan to migraine patients receiving propranolol for prophylaxis, the 5 mg dose of rizatriptan is recommended. Administration with other beta-adrenoceptor blockers does not require consideration of a dose adjustment.

Evaluation of amino acid-based linkers in potent macrocyclic inhibitors of farnesyl-protein transferase.

A series of amino acid-based linkers was used to investigate the effects of various substituents upon the potency, pharmacokinetic properties, and conformation of macrocyclic farnesyl-protein transferase inhibitors (FTIs). As a result of the studies described herein, highly potent FTIs with improved pharmacokinetic profiles have been identified.

Oxo-piperazine derivatives of N-arylpiperazinones as inhibitors of farnesyltransferase.

The evaluation of SAR associated with the insertion of carbonyl groups at various positions of N-arylpiperazinone farnesyltransferase inhibitors is described herein. 1-Aryl-2,3-diketopiperazine derivatives exhibited the best balance of potency and pharmacokinetic profile relative to the parent 1-aryl-2-piperazinones.

Mechanistic investigation of ionization suppression in electrospray ionization.

We show results from experiments designed to determine the relative importance of gas phase processes and solution phase processes into ionization suppression observed in biological sample extracts. The data indicate that gas phase reactions leading to the loss of net charge on the analyte is not likely to be the most important process involved in ionization suppression. The results point to changes in the droplet solution properties caused by the presence of nonvolatile solutes as the main cause of ionization suppression in electrospray ionization of biological extracts.

Synthesis of conformationally constrained 5,6,7, 8-Tetrahydroimidazo[1,5-a]pyridine inhibitors of farnesyltransferase.

[reaction: see text] Synthesis of the 8-amino-5,6,7,8-tetrahydroimidazo[1,5-a]pyridine ring system was accomplished by intramolecular cyclization of an iminium ion, derived from condensation of an amine and a substituted gamma-(1-imidazolyl)butyraldehyde. The reaction was used to produce conformationally restricted farnesyltransferase inhibitor analogues which exhibit improved in vivo metabolic stability.

Selective alpha1a adrenergic receptor antagonists based on 4-aryl-3,4-dihydropyridine-2-ones.

A series of alpha1a receptor antagonists derived from a 4-aryl-3,4-dihydropyridine-2-one heterocycle is disclosed. Potency in the low nanomolar to picomolar range along with high selectivity was obtained. In vivo efficacy in a prostate contraction model in rats was observed with a few derivatives.

In vitro and in vivo evaluation of dihydropyrimidinone C-5 amides as potent and selective alpha(1A) receptor antagonists for the treatment of benign prostatic hyperplasia.

alpha(1) Adrenergic receptors mediate both vascular and lower urinary tract tone, and alpha(1) receptor antagonists such as terazosin (1b) are used to treat both hypertension and benign prostatic hyperplasia (BPH). Recently, three different subtypes of this receptor have been identified, with the alpha(1A) receptor being most prevalent in lower urinary tract tissue. This paper explores 4-aryldihydropyrimidinones attached to an aminopropyl-4-arylpiperidine via a C-5 amide as selective alpha(1A) receptor subtype antagonists. In receptor binding assays, these types of compounds generally display K(i) values for the alpha(1a) receptor subtype <1 nM while being greater than 100-fold selective versus the alpha(1b) and alpha(1d) receptor subtypes. Many of these compounds were also evaluated in vivo and found to be more potent than terazosin in both a rat model of prostate tone and a dog model of intra-urethral pressure without significantly affecting blood pressure. While many of the compounds tested displayed poor pharmacokinetics, compound 48 was found to have adequate bioavailability (>20%) and half-life (>6 h) in both rats and dogs. Due to its selectivity for the alpha(1a) over the alpha(1b) and alpha(1d) receptors as well as its favorable pharmacokinetic profile, 48 has the potential to relieve the symptoms of BPH without eliciting effects on the cardiovascular system.

Combinatorial diversification of indinavir: in vivo mixture dosing of an HIV protease inhibitor library.

An efficient combination solution-phase/solid-phase route enabling the diversification of the P1', P2', and P3 subsites of indinavir has been established. The synthetic sequence can facilitate the rapid generation of HIV protease inhibitors possessing more favorable pharmacokinetic properties as well as enhanced potencies. Multiple compound dosing in vivo may also accelerate the identification of potential drug candidates.

Lipoma of the liver: a differential-diagnostic problem.

BACKGROUND AND AIMS: Lipomatous tumors of the liver are extremely rare; most of them are found incidentally at autopsy. Appropriate methods of the diagnosis of these lesions are ultrasonography (US), computed tomography (CT) and biopsy. CASE REPORT: The case of a 65-year-old man who had undergone an upper-rectum resection because of exulcerated adenocarcinoma of the rectum is described. Six months later, routine control examination revealed a solid tumor in the left lobe of the liver. The tumor was not demonstrated by either sonography or CT before the operation. Repeated US, CT scan and fine-needle biopsy could not exclude the possibility of a metastatic tumor secondary to previous malignancy. The lesion was removed by wedge resection and was proven to be a lipoma. CONCLUSION: Despite adequate preoperative examination (CT, biopsy and US), an anamnestic adenocarcinoma of the rectum can lead to inappropriate therapy. The inadequate result of our therapeutic policy can be explained by the extremely rare incidence of lipoma of the liver.

Rizatriptan, a novel 5-HT1B/1D agonist for migraine: single- and multiple-dose tolerability and pharmacokinetics in healthy subjects.

Rizatriptan is a novel 5-HT1D/1B agonist for relief of migraine headache. The pharmacokinetics, metabolite profiles, and tolerability of rizatriptan were examined in a multiple-dose study in healthy subjects. Rizatriptan (N = 24) (or placebo, N = 12) was administered as a single 10 mg dose, followed 48 hours later by administration of one 10 mg dose every 2 hours for three doses on 4 consecutive days, corresponding to the maximum daily dose for a migraine attack. The AUC of rizatriptan and its active N-monodesmethyl metabolite after three 10 mg doses was approximately threefold greater than the plasma concentrations following a single 10 mg dose. Metabolite profiles were similar after single and multiple doses. Adverse events during rizatriptan were mild and transient; similar events occurred during placebo, with a somewhat reduced incidence. Diastolic blood pressure tended to increase compared with placebo (approximately 5 mmHg), particularly on the first multiple-dose day (p < .01 vs. placebo). In conclusion, rizatriptan is well tolerated by healthy subjects during multiple-dose administration, with no unexpected accumulation of drug in plasma.

[Surgical treatment of Boerhaave's syndrome]. / A Boerhaave-szindróma sebészi kezelése.

The authors treated four patients with spontaneous oesophageal rupture between 1995 and 1999. The interval from injury to surgical treatment was less than 24 h in two cases, and longer than 24 h in the other two cases. The surgical treatment was primary closure of the rupture and mediastinal drainage with gastrostomy within 24 h after the onset. Septic complications occurred in lately presented patients. In these cases, the surgical treatment consisted of oesophagectomy, mediastinal drainage and feeding gastrostomy. The alimentary tract was reconstructed with interposed ileocolon eight weeks later. Each patients was fully recovered. The authors underline the importance of early diagnosis which obviously determine the prognosis and the choice of surgical treatment. Primary repair is proposed after early diagnosis, however oesophageal resection is recommended 24 h after the injury.

Pharmacokinetics and tolerability of oral rizatriptan in healthy male and female volunteers.

AIMS: The pharmacokinetics and dose proportionality of rizatriptan single oral doses from 2.5 to 15 mg administered as solutions to healthy volunteers were studied. METHODS: In a randomized, crossover study with four periods, twenty-four healthy volunteers (12 males and 12 females) took single oral doses of 2.5, 5, 10, and 15 mg rizatriptan in Periods 1-4. In a fifth period, subjects received 4 mg intravenous (i.v.) rizatriptan as a reference. Plasma and urine rizatriptan concentrations were determined at several timepoints/intervals for 12 and 24 h, respectively. RESULTS: The arithmetic mean AUC values following single oral doses of 2.5, 5, 10, and 15-mg rizatriptan were 16, 33, 72, and 127 ng ml-1 h, respectively, in males; and 19, 42, 97, and 161 ng ml-1 h, respectively, in females. The overall bioavailability (F ) of rizatriptan was approximately 40% in males. Following the 4 mg reference i.v. dose, the apparent plasma clearance (CL) and renal clearance (CLr ) were 1042 and 225 ml min-1, respectively, in males; and 821 and 174 ml min-1, respectively, in females. CONCLUSIONS: The disposition kinetics of oral rizatriptan were linear for doses of 2. 5-10 mg in males, and for doses of 2.5-5 mg in females. However, the degree of nonlinearity for higher doses was minor for both genders. The plasma concentrations of rizatriptan were slightly greater in women compared to men but the difference was not considered to be clinically meaningful. Also, the clearance of rizatriptan appeared to be mainly nonrenal.

[Esophago-pericardial fistula caused by recurrent esophageal tumor]. / Recidív nyelócsótumor által okozott oesophago-pericardialis fistula.

The esophago-pericardial fistula is a rare disease that causes a rather high mortality. Up to now less than 70 cases were published in the literature. The authors report an esophago-pericardial fistula caused by recurrent esophageal tumor one year after resection in the lower third esophagus for esophageal cancer. With conservative treatment the patient was kept alive for a month. No case report can be found in the literature of an esophago-pericardial fistula of the same etiology.

[Drug therapy of gastroesophageal reflux (a prospective controlled clinical trial)]. / A gastrooesophagealis reflux betegség gyógyszeres kezeléséröl (prospektív kontrollált klinikai vizsgálat).

One year follow up(1.5, 3, 6, 12 months) study was established to examine the role of several classes of drugs in the treatment of reflux disease in 40 patients on the basis of objective control parameters (pH-metry, endoscopy, histology). The therapy was initiated, respectively, the different stage of severity (Savary-Miller): in stage 0 sucralfate + domperidone, in stage I and II: ranitidine + domperidone and in stage III-IV omeprazole was introduced. Our results proved that sucralfate + domperidone is curative on reflux oesophagitis in stage 0 cases. In stage I sucralfate and domperidone were effective in 3 of 9 cases, ranitidine + domperidone was optimal in 5 of 9, and omeprazole was required in 1 of 9 patients. In stage II, ranitidine + domperidone was effective only in 4 of 11 patients, and the initial therapy was modified to omeprazole in 7 of 11 patients to find the optimal drug in this stage. In stage III and IV only omeprazole showed curative effect and the doses required were 20 mg in 8 of 13 and 40 mg in 5 of 13 patients. The complaints improved in 34 of 40 patients after 6 weeks treatment, while histological healing of reflux oesophagitis was observed in 12 of 40 cases. After 3 months the endoscopic healing rate was 28/40, but histological healing could be reached after 6 months of optimal treatment in 30 of 40 cases. We can conclude, that the optimal drug selection may result a rapid improvement of complaints, but endoscopic and histological regeneration of the oesophageal mucosa is more graduated with time. The healing process of the reflux oesophagitis requires 3 months. Proton pump inhibitor drugs have an enhanced role in the treatment of gastrooesophageal reflux disease, and our results proved that the efficient and safety treatment of mild form (stage II) of disease requires the administration of proton pump inhibitors.

Metabolism of MK-499, a class III antiarrhythmic agent, in rats and dogs.

MK-499 [(+)-N-[1'-(6-cyano-1, 2, 3, 4-tetrahydro-2(R)-naphthalenyl)-3, 4-dihydro-4(R)-hydroxyspiro(2H-1-benzopyran-2, 4'-piperidin)-6-yl]methanesulfonamide] monohydrochloride is an investigational class III antiarrhythmic agent for treatment of malignant ventricular tachyarrhythmias. The disposition of [3H]MK-499 and [14C]MK-499 was studied in rats and dogs after oral and iv administration. MK-499 was concentrated in organs of excretion and the heart. In the rat, urinary radioactivity elimination values after iv (0.5 mg/kg) and oral (6.25 mg/kg) doses were 21 +/- 3% and 10 +/- 2%, respectively. Corresponding fecal recoveries were 68 +/- 6% and 78 +/- 7%. Similar results were found after corresponding doses of [14C]MK-499. In dogs, urine and feces accounted for 16 +/- 3% and 75 +/- 4% of recovered radioactivity after a [3H]MK-499 iv dose (0.1 mg/kg). Corresponding recoveries after an oral dose (1 mg/kg) were 12 +/- 2% and 76 +/- 3%. Biliary (0-24 hr) excretion accounted for 39 +/- 5% and 41 +/- 18% of [3H] and [14C] oral doses in rats, respectively. Dogs excreted 34% of [3H] oral dose in (0-24 hr) bile. The data indicated that a substantial amount of MK-499 was absorbed by rats and dogs. MK-499, metabolite I (formed by loss of N-substitution), and metabolite II (an acid formed by metabolic scission across the benzopyran ring) each represented 30% of rat urinary label. Rat bile contained MK-499 (10%), II (20%), and IV (10%), which was formed by carbon-4 hydroxylation of the tetralin ring. Additionally, rat bile included glutathione (V) and N-acetyl-1-cysteine (VI) conjugates of a ring-opened metabolite. Metabolite III, a positional isomer of IV, was excreted in rat urine. The major labeled species excreted in dog bile were unchanged MK-499 and its glucuronide (VII), which, respectively, represented 50% and 30% of the biliary radioactivity. MK-499 and a small amount of I represented dog urinary radioactivity. The bioavailability of MK-499 was high in dogs (100%) but low in rats (17%). This difference was probably due to the more extensive presystemic metabolism of MK-499 in rats.