Source: Marketing Authorisation Holder Revision Year: 2015
DUOVENT UDV nebulizer solution is a combination of the anticholinergic bronchodilator ipratropium bromide and the β2 adrenergic bronchodilator fenoterol hydrobromide. Ipratropium bromide is a quaternary ammonium derivative of atropine and is an anticholinergic drug which has bronchodilator properties. Each unit dose vial contains a total of 0.5 mg ipratropium bromide and 1.25 mg fenoterol hydrobromide in 4 mL of normal saline.
Ipratropium bromide is a quaternary ammonium compound with anticholinergic (parasympatholytic) properties. In preclinical studies, it inhibits vagally mediated reflexes by antagonising the action of acetylcholine, the transmitter agent released from the vagus nerve.
Anticholinergics prevent the increase in intracellular concentration of Ca++ which is caused by interaction of acetylcholine with the muscarinic receptor on bronchial smooth muscle. Ca++ release is mediated by the second messenger system consisting of IP3 (inositol triphosphate) and DAG (diacylglycerol).
The bronchodilatation following inhalation of ipratropium bromide is primarily a local, sitespecific effect, not a systemic one.
Preclinical and clinical evidence suggest no deleterious effect of ipratropium bromide on airway mucous secretion, mucociliary clearance or gas exchange.
Fenoterol hydrobromide is a direct acting sympathomimetic agent, selectively stimulating beta2-receptors in the therapeutic dose range. The stimulation of beta1-receptors comes into effect at a higher dose range. Occupation of beta2-receptors activates adenyl cyclase via a stimulatory Gs-protein.
The increase in cyclic AMP activates protein kinase A which then phosphorylates target proteins in smooth muscle cells. This in turn leads to the phosphorylation of myosin light chain kinase, inhibition of phosphoinositide hydrolysis, and the opening of large-conductance calcium-activated potassium channels.
Fenoterol hydrobromide relaxes bronchial and vascular smooth muscle and protects against bronchoconstricting stimuli such as histamine, methacholine, cold air, and allergen (early response). After acute administration the release of bronchoconstricting and pro-inflammatory mediators from mast cells is inhibited. Further, an increase in mucociliary clearance has been demonstrated after administration of doses of fenoterol (0.6 mg).
The bronchodilating effect of fenoterol hydrobromide is produced primarily by stimulation of β2 receptors in the bronchial smooth muscles. When administered by inhalation, fenoterol exerts a significant increase in pulmonary function 5 minutes after administration with a maximal effect in 30 to 60 minutes. This effect remains at the same level for 2-3 hours before gradually declining. A significant degree of bronchodilation has been reported in some studies for 6-8 hours.
The concurrent administration of ipratropium bromide and fenoterol hydrobromide results in dilatation of the bronchi by affecting different pharmacologic sites of action. The two active substances thus complement each other in their spasmolytic action on the bronchial muscles. The complementary action is such that only a very low proportion of the ß-adrenergic component is needed to obtain the desired effect, facilitating individual dosage suited to each patient with a minimum of adverse reactions.
Large single inhaled doses of ipratropium bromide have been given to man without any signs of toxicity. After administration of 400 µg by inhaler (10 times the recommended single dose) to 10 normal subjects, no changes were detected in pulse rate, blood pressure, intraocular pressure, salivary secretion, visual accommodation or electrocardiograms. Likewise, in another study, no changes in pulse rate or salivary secretion were seen when cumulative doses up to 1.2 mg were administered by inhalation to 12 normal volunteers.
Higher plasma concentrations, which are more frequently achieved with oral, or even more so, with intravenous administration inhibit uterine motility. Also at higher doses, metabolic effects are observed: lipolysis, glycogenolysis, hyperglycaemia and hypokalaemia, the latter caused by increased K+-uptake primarily into skeletal muscle.
Beta-adrenergic effects on the heart such as increase in heart rate and contractility, are caused by the vascular effects of fenoterol, cardiac beta2-receptor stimulation, and at supratherapeutic doses by beta1-receptor stimulation. As with other beta-adrenergic agents, QTc prolongations have been reported. For fenoterol pressurized inhalation solution these were discrete and observed at doses higher than recommended. However, systemic exposure after administration with nebulizers (nebulizer solution, nebulizer solution in UDV) might be higher than with recommended pressurized inhalation solution doses. The clinical significance has not been established. Tremor is a more frequently observed effect of beta-agonists. Unlike the effects on the bronchial smooth muscle, the systemic effects on skeletal muscle of ß-agonists are subject to the development of tolerance.
Special studies utilizing therapeutic doses in asthmatic and chronic bronchitic patients, again did not reveal any systemic anticholinergic effects. In one study, 14 patients were treated for 45 days with either Atrovent inhaler 40 µg qid or Atrovent inhaler 40 µg plus oral Berotec 5 mg qid. No changes in visual acuity, intraocular pressure, pupil size or accommodation of vision occurred. Micturition function studies in 20 male patients showed no differences in urinary flow, total flow time and time until maximum flow between placebo and ipratropium bromide inhaler 40 µg tid administered for 3 days.
Deterioration in pulmonary function in patients treated in all clinical trials with therapeutic doses of Atrovent solution was examined. The following table shows the number of patients who showed a 15% or greater fall in FEV1 at any time within 2 hours following the administration of the drug. Also shown are the figures for comparative agents used.
| Treatment | Incidence |
|---|---|
| Normal saline | 15/90 (16.7%) |
| Atrovent Solution | 14/214 (6.5%) |
| Atrovent Inhaler | 4/78 (5.1%) |
| Berotec Solution | 4/83 (4.8%) |
| Atrovent Solution + Berotec Solution | 1/81 (1.2%) |
Dose titration studies in stable asthmatic patients with Atrovent solution have indicated that maximal improvement in pulmonary function occurs at approximately 250 µg for adults and 125 µg for children over 5 years.
A clinical pharmacology study comparing single doses of Atrovent inhaler (80 µg) and Atrovent solution (250 µg) in 16 stable adult asthmatics was performed. No difference between the regimens was found, based on an improvement in pulmonary function over a 2 hour period. A wide variety of challenge studies have been conducted using ipratropium bromide as a protective agent. In pharmacologically induced bronchospasm, ipratropium bromide, in clinical doses, was very effective against metacholine and acetylcholine, moderately effective against propranolol but had little or no effect against histamine or serotonin. Studies in exercise induced bronchospasm have yielded variable results. Some investigations have indicated that ipratropium bromide has little or no effect but other studies have shown that some patients are protected against bronchospasm induced by exercise. Likewise, the protective effects of ipratropium bromide against cold air induced bronchospasm have been variable.
Antigen challenge studies have demonstrated that Atrovent offers some protection against the “early” allergic asthma response, but has no effect on the “late” response.
In patients with asthma and COPD, better efficacy compared to its components ipratropium or fenoterol was demonstrated. Two studies (one with asthma patients, one with COPD patients) have shown that DUOVENT is as efficacious as double the dose of fenoterol administered without ipratropium but was better tolerated in cumulative dose response studies.
In acute bronchoconstriction DUOVENT is effective shortly after administration and is therefore also suitable for treating acute episodes of bronchospasm.
The pharmacokinetics of ipratropium bromide and fenoterol are not altered when the two drugs are administered concurrently.
The therapeutic effect of the combination ipratropium bromide and fenoterol hydrobromide is produced by a local action in the airway. The pharmacodynamics of the bronchodilation are therefore not related to the pharmacokinetics of the active constituents of the preparation.
Following inhalation 10 to 39% of a dose is generally deposited in lungs, depending on the formulation, inhalation technique and device, while the remainder of the delivered dose is deposited in the mouthpiece, mouth and the upper part of the respiratory tract (oropharynx).
There is no evidence that the pharmacokinetics of both ingredients in the combination differ from those of the mono-substance.
Absorption: Ipratropium bromide is absorbed quickly after oral inhalation of a nominal dose of 40 µg administered from a pressurized metered dose inhaler. The peak plasma concentration (mean Cmax = 32 pg/mL) is reached within 5 minutes after inhalation. The therapeutic effect of ipratropium bromide is produced by a local action in the airways. Therefore time courses of bronchodilation and systemic pharmacokinetics do not run in parallel. The time to reach peak plasma concentration was similar to that seen after oral administration, likely reflecting the large fraction of inhaled dose which is deposited in the pharyngeal mucosa and swallowed. The absolute bioavailability after oral administration is approximately 2%.
Distribution: Intravenous administration of 1.0 mg in man showed a rapid distribution into tissues (half-life of alpha phase approximately 5 minutes), and a terminal half-life (beta phase) of 3-4 hours. Plasma concentrations after inhaled ipratropium bromide were 1000 times lower than equipotent oral or intravenous doses (15 and 0.15 mg, respectively).
The half-life of the terminal elimination phase is about 1.6 hours. Ipratropium has a total clearance of 2.3 L/min and a renal clearance of 0.9 L/min. After intravenous administration approximately 60% of a dose is metabolised, the major portion probably in the liver by oxidation.
Radio-labelled technetium was administered with Atrovent (ipratropium bromide) solution in an adult dose finding study. The following table outlines the doses reaching the patient. The figures for Atrovent inhaler are published estimates.
| Dose Available (µg) | Amount Reaching Patient (µg) | Lung Dose (µg) |
|---|---|---|
| 500 | 53 | 17.0 |
| 250 | 27 | 8.5 |
| 125 | 13 | 4.3 |
| 40 (Atrovent Inhaler) | 40 | 4.4 |
The drug is minimally (less than 20%) bound to plasma proteins. The ipratropium ion does not cross the blood-brain barrier, consistent with the quaternary amine structure of the molecule. It is not known if the placental barrier is crossed.
Metabolism: Up to eight metabolites of ipratropium have been detected in man, rat and dog. However, the main metabolites bind poorly to the muscarinic receptor.
Excretion: In man, about 70% of the 14C labelled drug is excreted unchanged after i.v. administration and only one metabolite exceeds 10% of the total radioactivity. The elimination of ipratropium and its metabolites occurs primarily via the kidneys with less than 10% of the total intravenous dose excreted via the biliary or fecal route. After oral or inhaled doses, however, up to 90% of the radiolabelled dose is detectable in the feces, suggesting relatively low lung deposition and poor absorption of the swallowed portion.
Cumulative renal excretion (0-24 hrs) of ipratropium (parent compound) is approximated to 46% of an intravenously administered dose, below 1% of an oral dose and approximately 3 to 13% of an inhaled dose via DUOVENT metered dose inhaler. Based on these data, the total systemic bioavailability of oral and inhaled doses of ipratropium bromide is estimated at 2% and 7 to 28% respectively. Taking this into account, swallowed dose portions of ipratropium bromide do not relevantly contribute to systemic exposure.
Kinetic parameters describing the disposition of ipratropium were calculated from plasma concentrations after i.v. administration. A rapid biphasic decline in plasma concentrations is observed. The apparent volume of distribution at steady-state (Vdss) is approximately 176 L (≈ 2.4 L/kg). The drug is minimally (less than 20%) bound to plasma proteins. Preclinical studies with rats and dogs revealed that the quaternary amine ipratropium does not cross the blood-brain barrier.
The half-life of the terminal elimination phase is approximately 1.6 hours. Ipratropium has a total clearance of 2.3 L/min and a renal clearance of 0.9 L/min. After intravenous administration approximately 60% of a dose is metabolised probably mainly in the liver by oxidation.
In an excretion balance study cumulative renal excretion (6 days) of drug-related radioactivity (including parent compound and all metabolites) accounted for 72.1% after intravenous administration, 9.3% after oral administration and 3.2% after inhalation. Total radioactivity excreted via the faeces was 6.3% following intravenous application, 88.5% following oral dosing and 69.4% after inhalation. Regarding the excretion of drug-related radioactivity after intravenous administration, the main excretion occurs via the kidneys. The half-life for elimination of drugrelated radioactivity (parent compound and metabolites) is 3.6 hours. Binding of the main urinary metabolites to the muscarinic receptor is negligible and the metabolites have to be regarded as ineffective.
Absorption: In man, fenoterol is rapidly absorbed from the gastrointestinal tract, with an absorption level of 60%. After administration of tritium labelled fenoterol, peak plasma levels (2.5% of the oral dose) are reached in two hours, the half-life of radioactivity being 6 to 7 hours. When given from a pressured container, absorption proceeds in two phases: the first one is essentially independent of the dose and apparently takes place between the first and fourth subdivisions of the bronchial tree. The second phase appears to be identical to oral absorption. After inhalation, blood levels remain almost unchanged for 7 hours (0.3-0.4 ng/mL fenoterol).
Following intravenous administration, three phases were observed, whereby the half-life of the terminal phase was approximately 3 hours.
Distribution: Fenoterol is very rapidly taken up by the tissues, where it conjugated to the extent of 99% (as sulphates).
Metabolism: Unlike isoproterenol, fenoterol is not metabolized by catechol-O-methyl transferase.
After intravenous administration, free fenoterol and conjugated fenoterol are approximated to 15% and 27% of the administered dose in the cumulative 24-hour urine. After inhalation via DUOVENT metered dose inhaler approximately 1% of an inhaled dose is excreted as free fenoterol in the 24-hour urine. Based on these data, the total systemic bioavailability of inhaled doses of fenoterol hydrobromide is estimated at 7%.
Kinetic parameters describing the disposition of fenoterol were calculated from plasma concentrations after i.v. administration. Following intravenous administration, plasma concentrationtime profiles can be described by a 3-compartment model, whereby the terminal half-life is approximately 3 hours. In this 3-compartment model the apparent volume of distribution of fenoterol at steady state (Vdss) is approximately 189 L (≈ 2.7 L/kg).
Excretion: The resulting metabolites are excreted via the kidneys (40% within the first 48 hours after oral administration) and the bile (fecal excretion: 40% of the oral dose).
About 40% of the drug is bound to plasma proteins.
Preclinical studies with rats revealed that fenoterol and its metabolites do not cross the blood-brain barrier. Fenoterol has a total clearance of 1.8 L/min and a renal clearance of 0.27 L/min.
In an excretion balance study cumulative renal excretion (2 days) of drug-related radioactivity (including parent compound and all metabolites) accounted for 65% of dose after intravenous administration and total radioactivity excreted in faeces was 14.8% of dose. Following oral administration, total radioactivity excreted in urine was approximately 39% of dose and total radioactivity excreted in faeces was 40.2% of dose within 48 hours.
Ipratropium bromide is an anticholinergic agent which, when delivered by aerosol, exerts its effect primarily in the bronchial tree. It abolishes acetylcholine induced bronchospasm in the guinea pig and dog after intravenous administration of ED50 of 0.15-0.40 µg/kg with a transient effect on blood pressure. By inhalation, approximately 25 µg ipratropium bromide produces a 50% inhibition of acetylcholine-induced bronchospasm in the dog with no detectable effect on blood pressure but with an increased duration of action compared to intravenous administration. Histologic evaluation of human bronchial mucosa following chronic inhalation of ipratropium bromide showed no alterations of epithelial, cilated or goblet cells. Short term mucociliary clearance in normal and bronchitic subjects was not adversely affected by 200 µg of inhaled ipratropium bromide.
The anticholinergic effects of ipratropium bromide were evaluated in several other organ systems following oral, subcutaneous, intravenous and inhalation administration. In dogs, a 50% increase in heart rate resulted from a s.c. dose of about 0.011 mg/kg, equipotent to atropine, but the equieffective oral dose of ipratropium was 58 times greater. By inhalation, no increase in heart rate or pathologic changes in ECG pattern were recorded at dose up to 8 mg. In another study, blood pressure and heart rate in the dog could be modulated after i.v. administration of low doses of ipratropium but metered aerosol administration of 100 puffs (40 µg/puff) was required to produce an 11% increase in heart rate.
Salivary secretion in the rat, mouse and dog was effectively inhibited by low parenteral doses of ipratropium bromide (0.001 to 0.032 µg/kg) but when given by the oral route, the effective dose increased over 100-fold. Aerosol administration to dogs of about 65 puffs (40 µg/puff) produced a 50% decrease in salivary flow. Similarly, effects on gastric secretion in the rat showed at least a 100-fold difference between effective enteral and subcutaneous doses.
Mydriatic effects of ipratropium bromide in mice were approximately equipotent to atropine after s.c. doses but were 10-20 times less after oral administration. Tests of doses of ipratropium bromide up to 100 mg/kg in the rabbit showed no effect on the central nervous system.
Ipratropium bromide, subcutaneously, inhibited the secretory effects of the cholinergic agonist, oxtremorine, in mice. It also inhibited spasmolytic effects equivalent to or greater than atropine in isolated guinea pig gut. In vitro tests with isolated rectum of the guinea pig demonstrated the effectiveness of ipratropium bromide in suppressing the spasmogenic effects of acetylcholine and pilocarpine. It was ineffective against histamine or barium chloride induced spasm. Ipratropium bromide exerted anticholinergic effects on the in situ bladder and intestine preparations of the dog. Intravenous doses were 500 times more potent than oral or intraduodenal administration.
Ipratropium bromide was administered by inhalation in combination with a β~2~ sympathomimetic agent (fenoterol hydrobromide). In both the dog and guinea pig, these agents were additive in antagonizing acetylcholine induced bronchospasm with ED50 being 19.8 µg (ipratropium), 49.25 µg (fenoterol) and 11.05 µg + 27.63 µg (ipratropium + fenoterol). In the dog, 50 µg of fenoterol by inhalation produced an 8% increase in heart rate and a 16% increase in left ventricular dp/dt. When 20 µg ipratropium was added to the above, the corresponding increases were 8% and 9%.
Fenoterol has been shown by pharmacologic studies in animals to exert a preferential effect on β~2~ adrenergic receptors, such as those located in bronchial smooth muscles.
Studies with isolated guinea pig trachea and atria were performed to evaluate the effects of isoproterenol, salbutamol and fenoterol using cumulative concentration-effect curves. In vitro, the agonists were similar in potency in relaxing guinea pig trachea but the order of potency for the chronotropic response of guinea pig trachea was isoproterenol, > fenoterol, > salbutamol.
Effect on acetylcholine-induced bronchoconstriction:
Guinea Pig:
a) Simultaneous recordings of the effects of intravenous administration of several adrenergic agents on bronchomotor tone and on cardiac rate were made following bronchospastic challenge with acetylcholine. In these experiments the β selectivity of fenoterol was superior to isoproterenol and orciprenaline and comparable to salbutamol and terbutaline.
b) Bronchospasm induced with i.v. acetylcholine was counteracted by sympathomimetic agents previously administered in aerosol form in varying concentrations. The maximum protective effect declined by 50% at 12 minutes (isoproterenol), 14 minutes (orciprenaline), 18 minutes (terbutaline), 25 minutes (salbutamol) and 27 minutes (fenoterol). The associated increases in heart rate at doses producing equal protection were respectively, 25 beats/min (isoproterenol), 15 beats/min (orciprenaline), 13 beats/min (terbutaline), 10 beats/min (fenoterol) and 1 beat/min (salbutamol).
Effect of Histamine-induced Bronchoconstriction:
Guinea Pig: The heart rate and protective effect of sympathomimetic agents were measured in guinea pigs continuously exposed to histamine aerosol (0.01%) for 10 minutes. Two hours after intraperitoneal administration of 200 µg/kg of drug, the time to collapse following histamine provocation were 6.3 minutes for fenoterol, 3.2 minutes for salbutamol, 3.1 minutes for isoproterenol and 2.2 minutes for saline. When given 15 minutes before histamine, all beta agonists at low aerosol concentrations (0.3 mg/mL) delayed collapse time significantly and only minor changes in heart rate were produced.
At higher aerosol concentrations (6 mg/mL), isoproterenol produced greater tachycardia than either of the other two agonists. Finally, when given by the oral route the dose required to completely prevent collapse during histamine administration was 10 mg/kg for fenoterol, 5 mg/kg for salbutamol and 10 mg/kg for isoproterenol. The tachycardia produced by salbutamol or fenoterol was of short duration, while that produced by isoproterenol persisted over the 90-minute period of the experiment.
Dog: In the anesthetized dog with histamine-induced bronchospasm, isoproterenol, salbutamol and fenoterol were given i.p. and measurements of pulmonary resistance, as well as of heart rate, were made. The bronchoconstrictor effects of histamine were significantly reduced by i.v. administration of the adrenergic agents tested. Isoproterenol-induced bronchodilatation was shorter than that of either fenoterol or salbutamol. Fenoterol and salbutamol produced longlasting bronchodilatation with less cardiac stimulation than did isoproterenol.
| % PROTECTION IN RESISTANCE | ||||
|---|---|---|---|---|
| DRUG | DOSE μg/kg i.p. | (15 min) | (60 min) | HEART RATE (beats/min) |
| Saline | 8.7 ± 6.2 | 9.2 ± 7.0 | -0.2 ± 1.11 | |
| Isoproterenol | 3.12 6.25 12.50 | 38.3 ± 7.6* 71.8 ± 3.2* 82.0 ± 6.9* | 7.7 ± 7.7 41.0 ± 15.0 29.5 ± 5.5 | 41.4 ± 11.3* 46.5 ± 7.3* 43.1 ± 11.1* |
| Salbutamol | 0.39 1.56 3.12 6.25 | 33.0 ± 16.6 74.0 ± 3.3* 85.0 ± 3.4* 82.3 ± 2.8* | 7.7 ± 7.7 32.0 ± 1.1* 48.1 ± 10.0* 64.1 ± 6.9* | 1.0 ± 1.0 6.0 ± 6.0 17.4 ± 4.6* 28.0 ± 6.9* |
| Fenoterol | 0.39 0.78 1.56 3.12 | 70.2 ± 7.2* 67.6 ± 10.4* 76.5 ± 4.9* 85.9 ± 4.1* | 53.0 ± 15.2* 63.9 ± 10.4* 79.8 ± 5.7* 83.7 ± 6.6* | 7.0 ± 7.0 11.5 ± 6.0 36.0 ± 14.5* 45.6 ± 14.2* |
* Significantly different from saline treated group, p <0.05
Metabolic Effects: In common with other β adrenergic agents, fenoterol exerts glycolytic, lipolytic, hypoglycemic effects and hypokalemic effects.
Effect on Ciliary Activity of the Respiratory Tract: In the rat airway preparation, fenoterol and isoproterenol were shown to augment, in a dose-dependent fashion, the frequency of ciliary movement, with a concomitant increase in the rate of mucus transport.
Ipratropium, fenoterol and the combination (1:2.5) inhalation aerosols were evaluated against acetylcholine-induced bronchospasm in the dog. The inhalation ED50 values were 19.8 µg (1.98 Duovent UDV Product Monograph Page 27 of 40 puffs of 10 µg ipratropium), 49.25 µg (1.97 puffs of 25 µg fenoterol) and 38.68 µg (2.21 puffs of 5 µg ipratropium + 12.5 µg fenoterol) for ipratropium, fenoterol and the combination, respectively. These results indicated the presence of an additive effect of the combination.
| LD50 Values for Ipratropium | ||
|---|---|---|
| Species | Route | LD50(mg/kg) |
| Mice | IV | 13.5 |
| Mice-males | IV | 12.3 |
| Mice-females | IV | 15.0 |
| Mice | SC | 322 |
| Mice | SC | 300 |
| Mice | Oral | 2010 |
| Mice | Oral | 1038 |
| Rats | IV | 15.8 |
| Rats | SC | 1500 |
| Rats | Oral | >4000 |
| Rats | Oral | 1722 |
The signs of toxicity were apathy, reduced mobility, ataxia, paralysis of skeletal muscle, clonic convulsions and death from respiratory failure. Toxic signs persisted for 3 hours after i.v. administration and for 8 days after oral administration.
Acute dose tolerance studies were performed in dogs. No deaths occurred at doses of up to 400 mg/kg oral or 50 mg/kg s.c. Signs of toxicity were mydriasis, dryness of oral, nasal and optic mucosa, vomiting, ataxia, increased heart rate, decreased body temperature and death from respiratory failure.
An acute inhalation toxicity study of ipratropium bromide administered as a 4% and 8% solution to guinea pigs was performed. No toxic signs were observed with the 4% solution and death occurred five hours after administration of the 8% solution (approximately 200 mg/kg).
An acute inhalation tolerance study in rats with benzalkonium chloride (0.025%) or benzalkonium chloride (0.025%) plus ipratropium bromide (0.025%) administered over 8 hours was performed. No clinical signs of intolerance were observed. Necropsy and histological findings (16 hours and 14 days after administration) were also negative.
Anesthetized normal and hypoventilated dogs tolerated doses up to 200 puffs (4 mg) of ipratropium bromide without ECG changes or heart failure. Reductions in heart rate were observed. Similar findings were seen in dogs given IV infusions (10 mg/kg/min) up to 1550 mg/kg or 1000 mg/kg plus 200 puffs from a placebo inhaler. Blood pressure reductions were also seen in these experiments.
An acute inhalation dose tolerance study in rats using doses up to 160 puffs (3.2 mg) from an Atrovent inhaler was performed. No deaths occurred. A combination of ipratropium bromide (up to 3.2 mg/kg) with fenoterol hydrobromide (up to 8 mg/kg) was administered by inhaler (up to 320 puffs) to rats. There were no deaths or clinical signs observed.
| TOXICITY | ||
|---|---|---|
| Acute Toxicity Species | Route | LD50 (mg/kg) |
| Rat (adult) 21 days observation | Oral | 2500 |
| Rat (adult) 14 days observation | Oral | 3750 |
| Rat (newborn) | Oral | 1360 |
| Rat (adult) | i.v. | 65 |
| Mouse | i.v. | 60.0 |
The signs of toxicity consisted of irritability, tachycardia, hyperpnea, ataxia, coma or convulsion and death.
An acute dose tolerance study in dogs was performed using oral and intravenous administration. No deaths occurred with doses up to 300 mg/kg orally or 35 mg/kg i.v. At higher doses, death was attributable to cardiac failure, as documented by ECG and pathologic findings.
| IPRATROPIUM BROMIDE + FENOTEROL HYDROBROMIDE (RATIO 1:2.5) | |||
|---|---|---|---|
| Species | Sex | Route | LD50 (mg/kg) |
| Mouse | M | i.v. | 23.6 |
| Mouse | F | i.v. | 26.2 |
| Mouse | M | oral | 630 |
| Mouse | F | oral | 650 |
| Rat | M | i.v. | 32.5 |
| Rat | F | i.v. | 32.5 |
| Rat | M | oral | 3200 |
| Rat | F | oral | 2450 |
The signs of toxicity were spasmodic breathing, tonic, clonic and saltatory convulsions, sedation, ataxia, spasms, exophthalmus, chromolacryorrhoea, reduced motility, tremor and positive sliding test. Late mortality occurred only after oral administration.
Single-dose toxicity studies with the combination ipratropium bromide and fenoterol hydrobromide in a ratio of 1/2.5 (ipratropium bromide/fenoterol hydrobromide) in mice and rats after oral, intravenous and inhalation administration revealed a low level of acute toxicity. In comparison to the individual components, the LD50 values of the combination were determined more by the ipratropium bromide component than by fenoterol hydrobromide without any indication of potentiation.
A subacute toxicity study of nine weeks duration in rats, utilizing doses of 10, 100 and 500 mg/kg, revealed no pathologic findings apart from a dose related decrease in food consumption and growth rate.
A four week study in dogs using doses of 3, 30 and 150 mg/kg (for three weeks) increased to 300 mg/kg, showed mydriasis, inhibition of lacrimal and salivary secretion, tracheal and ocular inflammation, decreased food intake and weight loss at the medium and high doses. Three of six dogs died when the dose was increased from 150 to 300 mg/kg.
A supplementary study of 13 weeks using doses of 1.5, 3.0 and 15 mg/kg revealed no pathologic changes apart from a dose related inhibition of lacrimal secretion and associated keratoconjunctivitis and dryness of the mouth.
A 32 day study in rats was conducted with the combination of ipratropium bromide and fenoterol hydrobromide at doses of 1.32 + 3.32 µg/kg (Group 1), 8 + 20 µg/kg (Group 2) and 24 + 60 µg/kg (Group 3) respectively. Fenoterol 60 µg/kg (Group 4) and ipratropium 24 µg/kg (Group 5) were also administered. Increases in heart rate (dose related in all treated animals) and dry mouth and nose (Groups 3 and 5) were seen. Increases in LDH (Groups 3 and 4), creatine kinase (all treated Groups), potassium (Groups 2, 3 and 4) and cholesterol (Groups 3 and 4) were observed. Myocardial scars were seen in one animal in Groups 3 and fatty changes in the liver were noted in one animal in Group 4.
Rats were treated with subcutaneous injections of 1, 10 and 100 mg/kg. One death occurred in the 10 mg/kg group from paralytic ileus. Inflammatory changes were noted at the injection site.
A 4 week study in dogs using doses of 10, 20 and 30 mg/kg (increased to 40 mg/kg on the last five days) was conducted. Dryness of oral and nasal mucosal membranes and mydriasis were noted along with conjunctivitis and keratitis associated with decreased lacrimal secretions. A decrease in food intake and body weight also occurred. One dog died in the high dose group. Signs of liver damage were noted in 2 of the high dose dogs. Low testicular weights, which have not been observed in other subsequent studies, were also observed.
Twelve rats were exposed to aerosolized ipratropium bromide at a concentration of 11.5 µg/L for 1 hour, 4 times per day for 7 days. No drug toxicity was observed.
In another study, administration of ipratropium bromide in doses of 128, 256 and 384 µg per rat per day for 30 days showed no signs of toxicity apart from a low grade inflammatory response and areas of fibrosis and hemorrhage in the parametrium of 2 of 9 females in the high dose group. This finding has not been observed in subsequent studies.
Four rhesus monkeys inhaled 500 µg of ipratropium bromide twice a day (total dose 1 mg/day) for 7 days without the appearance of any drug induced toxicity.
In another rhesus monkey study, the animals were given ipratropium bromide at doses of 200, 400 and 800 µg/day of inhalation for 6 weeks. Included in the tests were measurements of mucociliary transport rate and ciliary beat frequency. No signs of drug toxicity were found.
A 13 week study was carried out in 120 rats. The animals being dose by gavage five days per week at 0.5 mg/kg, 5 mg/kg, 50 mg/kg and 150 mg/kg. Of the animals, 109 survived; mortality showed no drug or dosage correlation, with the highest death rate occurring in the control group. The males in the highest dosage group showed a slower weight gain in comparison to the control. The high dose group showed increased, water consumption, and the animals became quite agitated for a short while following each administration of the drug. Out of the 40 animals in this group, 23 had focal necrosis of the myocardium or myocardial scars; one animal had hemorrhage in the adrenal gland, three had focal necrosis and another three animals central globular atrophy in the liver.
The drug was also given for 13 weeks to 18 dogs, seven days a week, in capsules at doses of 0.3 mg/kg, 3 mg/kg and 30 mg/kg. All animals survived and no weight alterations occurred. There was a slight drop in the hemoglobin and hematocrit values in both the middle and high level groups. Elevation of serum potassium levels occurred in the middle and high dosage group. There was also a lowering of non-esterified fatty acids. Post-dose tachycardia was common. A consistent shortening of the P-T interval was seen shortly after administration. In animals of the mid and high dose group, drug-related changes in the heart included: dilatation and/or hypertrophy with numerous subendocardial foci of myocardial necrosis, as well as areas of recent and old scars.
A four week study in dogs was performed using 0.003 mg/kg and 0.03 mg/kg intravenously. In the high dose group there was an elevation of blood urea and non-esterified fatty acids were lowered. Post-dose tachycardia and shortening of the P-T interval also occurred in this group. In two high dose animals there was moderate hypertrophy of the left ventricle associated with small areas of necrosis. Three high dose animals demonstrated hypertrophy of the myocardial fibers.
Monkeys were administered fenoterol hydrobromide 0.5 mg/kg, 2.5 mg/kg, and 5.0 mg/kg as an aerosol 6 hours daily for 6 weeks. All the animals survived and of all the parameters studied, only focal lesions of myocarditis were seen in one or two (low dose) animals per group including the control. In a second similar study, no lesions of myocarditis were observed.
Rats were exposed to a combination of fenoterol and ipratropium twice, 4 times and 8 times per day. Metered dose inhalers containing 50 µg fenoterol and 20 µg ipratropium per actuation were discharged into the exposure chamber at a rate of 6 doses per minute for 25 minutes over a 7 day period. No changes apart from a reduction in food consumption in the first 2 days in the high dose group were noted.
A 28 day study in dogs was conducted using fenoterol and ipratropium in the following doses respectively: 350 + 140 µg (Group 3); 1050 + 420 µg (Group 4); 3150 + 1260 µg (Group 5). Vasodilation occurred in Groups 4 and 5 and heart rate was increased in the treated animals. Potassium levels were raised in Group 5. Liver glycogen content was raised in 4 (of 6) animals in Group 5 and 2 in Group 4.
A further 13 week combination study was done in dogs using doses of 23 + 9 µg (Group 1), 160 + 64 µg (Group 2) and 1100 + 440 (Group 3) fenoterol + ipratropium respectively. Peripheral hyperaemia and dry mucous membranes were observed in all treated animals. Increases in heart rate were seen in Groups 1 to 3, and 5 of 6 dogs in Group 3 had disturbances of impulse formation and conduction. Slight increases in GPT in Groups 2 and 3, as well as increases in AP in individual animals of Groups 1 to 3 were noted. Histological findings consisted of a scar in the papillary muscle of the left ventricle of one dog in Group 3 as well as centrolobular fatty infiltration of hepatocytes in dogs of Groups 2 and 3.
Repeat-dose toxicity studies with the combination ipratropium bromide and fenoterol hydrobromide were performed in rats (oral, inhalation) and dogs (intravenous, inhalation) for up to 13 weeks. Only minor toxic effects at concentrations up to several hundred times greater than that recommended in man were observed. Left ventricular myocardial scars were seen only in one animal from the highest treatment group (84µg/kg/day) of the 4-week intravenous study in dogs. The 13-week oral study in rats and the 13-week inhalation study in dogs did not show any toxicological changes beyond that proportional to the individual components.
There was no indication of potentiation with the combination in comparison to the individual components. All of the adverse effects observed are well known for fenoterol hydrobromide and ipratropium bromide.
A 6 month and a 1 year study in rats using doses of 6, 30 and 150 mg/kg were performed. The high dose was increased to 200 mg/kg after 14 weeks. Reductions in food consumption and growth rates were observed in the highest dose group. A dose dependent constipation which caused severe coprostasis and dilatation of the intestines was observed in the highest dose group. A toxic hepatosis was observed in some animals of the highest dose group.
Ipratropium bromide was administered to dogs at doses of 1.5, 3.0, 15.0 and 75.0 mg/kg for 1 year. A decrease in body weight development was seen in the highest dose group and food consumption was reduced in the dogs receiving 3 mg/kg and above. Emesis was seen in all treated groups. A dose dependent decrease (3 mg/kg and above) in nasal, oral and lacrimal secretions, the latter leading to keratoconjunctivitis, was observed. Increases in SGPT and SGOT (15 and 75 mg/kg) and alkaline phosphatase (75 mg/kg) were noted. Localized gastric necrosis was found in two dogs at the highest dose and a non-dose-dependent fatty degeneration of the liver which varied from animal to animal, was also seen.
A 6 month study in rats was performed using doses of 128, 256 and 384 µg per rat per day. Measurements included ciliary beat frequency, lung mechanics and blood gas. The only finding was a dose related decrease in growth rate of the male animals.
A 6 month inhalation toxicity study was performed in rhesus monkeys utilizing daily doses of 20, 800 and 1600 µg. All findings were negative including measurements of lung mechanics, ciliary beat frequency and blood gases.
A 78-week chronic toxicity study was performed with 400 SPF rats, receiving oral daily doses of 0.4 mg/kg, 2 mg/kg, 10 mg/kg or 50 mg/kg of fenoterol. Because of the absence of toxic symptoms, the dosage in the high-dose group of animals was gradually increased to 100 mg/kg. No toxic symptoms were observed in any of the groups. An excessive increase in body weight, particularly in the females, was dose-dependent and correlated with an increased in food and water consumption. A reduction in the glycogen was seen in the liver (high dose) and muscles (mid and high dose).
A one year toxicity study was performed in 24 dogs at dosages of 0.3 mg/kg, 1.5 mg/kg or 7.5 mg/kg daily. A retardation of body weight gain was noticeable in the high dose male animals. A reduction of non-esterified fatty acids was seen in the mid and high dose groups. ECG, hearing and eye examinations remained within normal limits. Gross findings were essentially normal and microscopic examination of only two different sections of myocardium revealed no abnormality.
Three Ames tests, a micronucleus test in mice, a cytogenic study in Chinese hamsters, and a dominant lethal test were performed to assess the mutagenic potential of ipratropium bromide. Two positive tests (one Ames and the micronucleus study) were apparently spurious as they could not be reproduced with subsequent exhaustive experimentation. In the cytogenic study, a doserelated increase in the number of chromatid gaps, but not of other aberrations, was seen. The significance of this finding is not known. All other test results were negative.
A number of short term in vitro and in vivo mutagenicity studies were conducted with fenoterol including several Ames tests (with and without metabolic activation), a HGPRT-test with V79 Chinese hamster cells (with and without metabolic activation), a mouse lymphoma L5178Y test (with and without metabolic activation), one chromosomal aberration study using V79 hamster cells and two chromosomal aberration studies using cultured human lymphocytes (with and without metabolic activation), a micronucleus test in mice and an unscheduled DNA synthesis assay in Hela S3 cells (with and without metabolic activation).
In the mouse lymphoma L5178Y assays, marginal increases in mutation frequency were observed at cytotoxic concentrations of fenoterol hydrobromide (between 2,000 and 3,000 µg/mL) in the absence of S9 activation.
The clastogenic potential of fenoterol hydrobromide was evaluated in cultured human lymphocytes in vitro. In the absence of metabolic activation, increases in the incidence of aberration frequency were recorded following a 45 hour exposure period. A repeat of this study in the same concentrations ranges and at the same exposure levels (up to 45 hours) demonstrated no clastogenic activity of fenoterol hydrobromide in the absence or in the presence of metabolic activation.
In all the other tests performed there was no evidence to show a mutagenic or clastogenic potential of fenoterol hydrobromide.
Carcinogenicity studies in mice (107 weeks duration) and rats (114 weeks duration) utilizing oral doses of up to 6 mg/kg were performed. These studies demonstrated that ipratropium bromide does not have a tumorigenic or carcinogenic effect.
A 78 week toxicity study was performed in male and female Charles-River (France) CD-1 mice with doses of 0, 25, 50 or 100 mg/kg/day of fenoterol administered in drinking water. Throughout the study, there were no overt clinical signs of drug toxicity. Mortality rates were similar among treated and control groups. There was a dose related increase in lung weights in the mid and high dose groups. Heart weights were increased in high dose males and in low and high dose females; there was also an increase of myocarditis in high dose males.
An increased incidence of uterine tumors in the treated female mice was observed although this finding appeared to be unrelated to dose (control, 1%; low, 18%; mid, 22%; high, 7.5%). These tumors were predominantly leiomyomas; 3 of the lesions (one in each dose group) were leiomyosarcomas. It has been postulated that these findings were not related to a directed effect of the drug, but rather to a receptor mediated secondary mechanism; these results are similar to the findings seen in rats with fenoterol and with other β adrenergic drugs. The incidence of bronchoalveolar tumors of the lungs were also increased (15%) in the high dose female mice. These tumors were evident only at necropsy, following a histological examination, and no evidence of decreased latency, multiplicity, or increased incidence of macroscopically recognizable tumors were observed. Neither hyperplasia nor other preneoplastic lesions was present. As historical control data on CD-1 mice indicates a spontaneous incidence of up to 40% for bronchioalveolar lung tumors, the incidences reported in this study were within an expected range for this species at this age.
In a two year peroral toxicity study in Sprague-Dawley rats given 25, 50 or100 mg/kg of fenoterol, there was an increased incidence of mesovarian leiomyomas seen in the female of the low (4%) and high (11%) animals sacrificed at the end of the study. No such tumors were observed in the control and mid dose animals nor in the dosed animals which died or were sacrificed during the study. Similar tumors were seen previously in long term studies carried out in this and other strains of rats using other β adrenergic agonists. No such tumors have been found at the analogous site (cremaster muscle) in male rats with any of these substances. All leiomyomas were histologically benign.
Carcinogenicity studies for the combination were not performed. No tumorigenic or carcinogenic effects were demonstrated in long term studies in mice and rats with ipratropium bromide. For fenoterol hydrobromide, carcinogenicity studies were performed after oral (mouse, 18 months rat, 24 months) and inhalation administration (rat, 24 months). At oral doses of 25 mg/kg/day an increased incidence of uterine leiomyomas with variable mitotic activity in mice and mesovarial leiomyomas in rats were observed. These findings are recognized effects caused by the local action of beta-adrenergic agents on the uterine smooth muscle cell in mice and rats. Taking into account the present level of research, these results are not applicable to man. All other neoplasms found were considered to be common types of neoplasm spontaneously occurring in the strains used and did not show a biologically relevant increased incidence resulting from treatment with fenoterol hydrobromide.
Three teratology studies, one in mice using oral doses of 2 and 10 mg/kg, and two in rats, were performed. The first study used the same doses and the second employed 10 and 20 mg/kg and revealed no drug induced fetal abnormalities.
A similar oral study in rabbits utilizing doses of 2 and 10 mg/kg again demonstrated no teratogenic or embryotoxic effects of ipratropium bromide.
An inhalation teratology study in rabbits using doses of 0.3, 0.9 and 1.8 mg/kg demonstrated no effects on litter parameters and no embryotoxic or teratogenic effects.
A fertility study in rats with oral doses of 5, 50 and 500 mg/kg given 60 days prior to and during early gestation was performed. Fertility was delayed in 8 of 20 couples at the 500 mg/kg dose and spurious pregnancy in 5 of 20 females occurred at this dose. In addition, the conception rate was decreased in 75% of females at this dose. No embryotoxic or teratogenic effects were observed.
Fertility and general reproductive performance have been evaluated in male and female rats at doses of up to 150 mg/kg given orally. Doses above 2.5 mg/kg produced signs of toxicity in the parent generation, but did not influence fertility, reproductive performance or fetal development.
Teratology and peri/postnatal studies have also been performed in rats given oral doses of fenoterol up to a maximum dose of 100 mg/kg. The substance was not shown to be teratogenic. A slight delay in delivery was caused by fenoterol, but F1 generation development, behaviour and reproductive capability were unaffected by the substance.
Teratology studies were performed in rabbits using oral doses up to 100 mg/kg. Decrease in body weight gain of the dose (100 mg/kg), increase in miscarriage rate (100 mg/kg) and an increased number of runts (100 mg/kg) were seen. No teratogenic effects were observed.
After inhalation administration of the combination ipratropium bromide and fenoterol hydrobromide in rats and rabbits no teratogenic effects occurred. Also no teratogenic effects were seen after ipratropium bromide, and after inhalation administration of fenoterol hydrobromide. After oral dosing, at doses >25 mg/kg/day (rabbits) and >38.5 mg/kg/day (mice) fenoterol hydrobromide induced an increase rate of malformations.
The malformations observed are considered a class effect for beta-agonists. Fertility was not impaired in rats at oral doses up to 90 mg/kg/day ipratropium bromide 166 and up to 40 mg/kg/day fenoterol hydrobromide.
Genotoxicity studies for the combination were not performed. In vitro and in vivo assays revealed that neither fenoterol hydrobromide nor ipratropium bromide have a mutagenic potential.
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