JOURNAL OF PHYSIOLOGY AND PHARMACOLOGY 2007, 58, Suppl 5, 603614 www.jpp.krakow.pl
G. SCHEUCH1, P. BRAND2, T. MEYER2, C. HERPICH2, B. MÜLLINGER1, J. BROM3, G. WEIDINGER3, M. KOHLHÄUFL4, K. HÄUSSINGER4, M. SPANNAGL5, W. SCHRAMM5, R. SIEKMEIER6
ANTICOAGULATIVE EFFECTS OF THE INHALED LOW MOLECULAR WEIGHT HEPARIN CERTOPARIN IN HEALTHY SUBJECTS Activaero GmbH, Gemuenden/Wohra, Germany; 2Inamed Research GmbH & Co. KG, Gauting, Germany; 3Novartis Pharma, Nürnberg, Germany; 4Asklepios-Fachkliniken, Gauting, Germany; 5Ludwig-Maximilians-Universität München, Abt. Hämostaseologie, München Germany; 6Bundesinstitut für Arzneimittel und Medizinprodukte (BfArM) Bonn, Germany
1
Inhalation of heparin results in local antiinflammatory and antifibrotic effects and an inhibition of blood coagulation. A number of experimental and clinical studies demonstrated that inhalant administration of heparin or low molecular weight heparin (LMWH) is a feasible and save tool for anticoagulative treatment. However, heparin and LMWH differ in respect to their molecular weight, pulmonary absorption, and principle of their anticoagulative pattern. In our study we investigated the anticoagulative effect of different doses of the LMWH certoparin after inhalation (3000 IU-9000 IU) and subcutaneous injection (3000 IU) in healthy individuals in a cross-over design. Inhalations were performed using a new device allowing inhalations with optimized and standardized breathing patterns. The anticoagulative effect was determined by measurement of the anti-factor-Xa (antiFXa) activity. Lung function parameters were measured before and after drug inhalation. Analysis of the anti-FXa activity as a function of the time after administration revealed values of the area under the curve (AUC) of 5.70 ±1.58 U·hour/ml and 8.43 ± 1.31 U·hour/ml (mean ±SD) with interindividual coefficients of variation of 28% and13 % after injection of 3000 IU and inhalation of 9000 IU, respectively. The AUC after inhalation of 9000 IU was significantly higher (P=0.0007) compared with subcutaneous injection of 3000 IU. In consequence, in order to obtain plasma anti-FXa activities of above 0.2 U/ml, which is considered sufficient for prophylaxis of venous thrombosis, 9000 IU LMWH have to be inhaled. Compared with the subcutaneous administration, the action of certoparin is longer after inhalation than after injection. Apparently, the drug is released rapidly according to a two-compartment kinetics, and its anticoagulant activity lasts over a long time without a marked plasma peak after administration. In detail, an elevation of plasma anti-FXa activity is achieved for 12 hours to 24 hours without a distinct peak shortly after inhalation. Inhalation of LMWH does not result in any changes in lung function or other side effects. The administration of LMWH by inhalation bears
604 the following: the non-invasive route of drug application, the low interindividual variability of the anticoagulative effect, and a long-time pharmacological effect. These properties suggest that controlled inhalation of heparin is an attractive alternative to subcutaneous administration. K e y w o r d s : aerosol therapy, inhalation, low molecular weight heparin, certoparin, coagulation
INTRODUCTION
Heparin, an acidic sulfated mucopolysaccharide, is isolated from tissues (especially gut and lung) rich in glycosaminoglycans and mast cells (1). The molecular weight of unfractionated heparin is between 2750 Da and 57500 Da (mean value: 15000 Da). However, its anticoagulative activity is restricted to a small specifically sulphated subunit and a linear molecular structure. It binds to antithrombin III (AT-III), a plasma glycoprotein, and to a small extent also to heparin cofactor II. Binding of heparin to AT-III results in a change of the conformation and a strong increase of the thrombin inhibitory effect, which becomes about 1000-times higher than before. Other targets of heparin in blood coagulation are: inhibition and reduced activation of factors V, VIII and IX and inhibition of thrombocyte function due to an unspecific binding of platelet factor IV. However, beside of its role in anticoagulation heparin has a lot of other properties (e.g., interaction with growth factors, regulation of cell proliferation and angiogenesis, modulation of proteases and antiproteases) (2-4) making it an interesting subject of investigations in the field of inflammation research, allergy and immunology (2, 4-7), interstitial lung fibrosis (8), and oncology (3). About 20 years ago a number of low molecular weight heparins (LMWH) were developed and introduced into therapy (9). LMWH are produced from heparin by controlled depolymerization and bear a molecular weight of 60% of the polysaccharides between 2000 Da and 6000 Da (mean values between 4000 Da and 6000 Da) (9). Compared with unfractionated heparin, LMWH are characterized by a longer plasma half-life time, a lower effect on platelets and endothelium, a higher bioavailability even at lower doses, and a lower rate of haemorrahic diathesis even at a similar anticoagulative effect (9-13). However, there are no or small differences between unfractionated heparin and LMWH in respect to the requirement of parenteral administration and the side effects, such as heparin-induced osteoporosis and thrombocytopenia (1, 9, 12, 14). As a consequence, LMWH is routinely used for prophylaxis and therapy of thromboembolic disorders. Even though heparin has been developed in 1915, it has been introduced in clinical therapy many years later and first reports on a possible relevance in patients with venous thrombosis or thromboembolia are from 1939 (1). Several studies were performed to investigate alternative routes of drug application.
605 However, although some feasibility and experimental data from animal studies exist (15-17), oral and nasal application of heparins have not been studied extensively in humans because the major problems of a low bioavailability and a considerable intraindividual variance were not resolved. Another approach is the inhalant application which has been firstly described about 40 years ago (18). Up to now, several hundred subjects inhaled unfractionated heparin and LMWH in dose finding studies for anticoagulation but also to prevent thrombosis in single patients at risk. Even long-time application of inhaled heparin for up to 485 days did not result in any considerable side effects (4). Only after the inhalation of high doses of unfractionated heparin (more than 400000 international units (IU) minor bleeding was observed (4). These studies indicate that inhalation of heparin is an effective, safe, and convenient procedure to obtain sufficient and long lasting anticoagulant action (18-25). Further studies have demonstrated local antiinflammatory effects of inhaled heparin, which might be beneficial for patients with asthma (2, 5, 7, 26, 27) and interstitial lung disease (8). Prior studies for optimization of aerosol inhalation techniques have shown that the variability of drug deposition in the lungs is mostly caused by suboptimal inhalation techniques and interindividual differences of breathing patterns of the subjects (28). If inhalation is performed using optimized and standardized breathing patterns, peripheral drug deposition is much higher and its variability is much lower as compared with uncontrolled inhalations (29-33). Therefore, we present a dosefinding study with inhalation of the LMWH certoparin in volunteers by means of a novel inhalation device allowing an optimized and standardized breathing pattern. MATERIAL AND METHODS
Study design In the first part of the study, in which 5 randomly selected healthy subjects participated, the dose of certoparin was assessed, which has to be inhaled to obtain a peak plasma anticoagulative activity of above 0.2 U/ml, which is considered to be sufficient for prophylaxis of venous thromboembolism. In a second study part, this dose was applied to 5 more healthy individuals to assess further pharmacological parameters of the inhaled LMWH in comparison with subcutaneous application of 3000 IU certoparin (a dose used in clinical routine for prophylaxis of venous thromboembolism), so that in this study part a total of 10 subjects was investigated in a cross-over design. Informed written consent was obtained from each subject. The study protocol was approved by the ethics committee of the Ludwig-Maximilians-University in Munich, Germany.
Subjects In total, 10 nonsmoking healthy subjects, 4 women and 6 men, aged 18-60 years, participated in the study. Body height and weight ranged from 1.63 m to 1.89 m and 60.0 kg to 82.0 kg, respectively, demonstrating a normal body mass index (BMI). Anthropometric data and baseline lung function parameters of body plethysmography and spirometry of the study population are shown in Table 1. From this group, 5 randomly selected subjects participated in the first part of the
606 Table 1. Anthropometric data and baseline values of lung function parameters of the study participants (n=10). Mean ±SD Age (yr) Height (m) Weight (kg)
Range
34.60±12.23 18.00-60.0 1.76±0.08 1.63-1.89 71.80±7.74 60.00-82.00
FEV1 (l) FEV1 (%) MEF25 (l/s) MEF25 (%) MEF50 (l/s) MEF50 (%) MEF75 (l/s) MEF75 (%) PEF (l/s) PEF (%)
3.73±0.73 97±10 1.60±0.57 69±21 4.30±1.44 85±25 7.42±2.33 98±25 9.38±1.67 108±11
2.50-4.65 82-117 0.68-2.37 36-107 1.92-6.58 45-129 3.73-10.17 49-123 6.73-11.64 89-126
VC (l) VC (%) TLC (l) TLC (%) ITGV (l) ITGV (%) Raw (kPa•l/s)
4.85±0.79 103±5 6.71±1.14 103±4 3.62±0.80 112±20 0.16±0.05
3.77-5.78 98-113 5.16-8.06 98-110 2.40-4.73 90-146 0.09-0.25
study, which served for evaluating the dose to be inhaled to achieve a plasma anti-FXa activity in the plasma of above 0.2 U/ml to 0.3 U/ml.
Safety analysis Lung function tests, body plethysmography, and spirometry were performed before and 30 min after inhalation, using a Jäger-Masterlab (Erich Jaeger GmbH, Würzburg, Germany). The following parameters were measured: total lung capacity (TLC), vital capacity (VC), intrathoracic gas volume (ITGV), airway resistance (Raw), peak expiratory flow (PEF), forced expiratory volume in one second (FEV1), and midexpiratory flow rates at 75%, 50%, and 25% VC (MEF75, MEF50 and MEF25, respectively). Measured lung function parameters were normalized to the reference values proposed by the European Community for Coal and Steel (34). Lung function was measured before the inhalant application of heparin and 30 min and 48 hours afterwards. In addition, clinical examination and blood cell counts were performed before and after drug application.
Drug application The pharmacological effect of a single dose inhalation of the LMWH certoparin (MonoEmbolex ®, Novartis Pharma) was investigated. Certoparin was used for inhalation as provided by the manufacturer. For the inhalant administration a standardized breathing pattern (volume: 1000
607 cm3, inhalation flow: 200 cm3/s) was chosen for which a peripheral lung deposition of 60% had been experimentally established in previous studies (29, 35). In detail, after inhalation of 800 cm3 aerosol, 200 cm3 of clean air were inhaled in order to optimize the alveolar deposition and to keep aerosol deposition in the respiratory dead-space as small as possible. Since the output of the nebulizer connected with the AKITA® inhalation-system (Pari LC+) is known from the predicted deposition, the drug amount deposited peripherally per breath was estimated. This calculation led to the conclusion that inhaling 54 breaths with the breathing pattern employed resulted in a peripheral deposition of 0.5 ml certoparin (3000 IU). The randomly selected test group from the first part of the study inhaled 3 different doses of certoparin, resulting in peripherally deposited doses of 3000 IU, 6000 IU, and 9000 IU on different days. Time intervals between certoparin inhalations were 5 days. In the second part of the study, the other five subjects inhaled the highest certoparin dose which resulted in a plasma anti-FXa activity which was above 0.2 U/ml. For comparison, all ten subjects received 3000 IU certoparin by subcutaneous administration on a separate study day five days after inhalation.
Pharmacodynamic analysis For the assessment of anti-FXa activity, venous blood samples were taken before and 15 min, 30 min, 1 h, 2 h, 4 h, 6 h, 24 h, and 48 h after drug application by both inhalation and subcutaneous injection. Certoparin concentration in plasma was not determined but the anti-FXa activity, which is closely correlated, was. The latter was assessed by means of the chromogenic Berichrom® assay (Dade Behring Marburg GmbH, Germany). The calibrators supplied by the manufacturer were declared in international units (IU) in relation to the international standard for LMWH. Based on the time course of anti-FXa activity the area under the curve (AUC) was calculated for all individuals at all study days. Other parameters determined in our study were the time to peak of the anticoagulative effect (tmax) and the maximum of the antiocoagulative effect (corresponding to Cmax).
Data analysis Differences between the average areas under the curves of the measured time courses of the anti-FXa activity in plasma were tested for statistical significance using the Student-t-test (SAS version 8e for Windows). RESULTS
Lung function parameters determined by body plethysmography and spirometry 30 min and 48 h after drug administration showed no significant differences when compared with the baseline values (P>0.05 for all comparisons, except TLC with P=0.045). In addition, clinical examination and blood cell counts were normal before and after drug application. The maximum anti-FXa activity in plasma was 0.3 U/ml after subcutaneous application of 3000 IU certoparin (n=10), and it was 0.082 U/ml, 0.18 U/ml, and 0.32 U/ml after inhalation of 3000 IU (n=5), 6000 IU (n=5), and 9000 IU (n=10), respectively (Fig. 1). After subcutaneous administration the maximum of the antiFXa activity was found after 2 h. However, different time dependent courses of the anti-FXa activity were observed after inhalation. In detail, after inhalation of 3000 IU two maxima were observed - one after 30 min and the other after 24 h. Inhalation
608
Fig. 1. Time course of the anti-FXa activity after subcutaneous application (3000 IU; in all panels as reference) or inhalation (3000 IU, 6000 IU, and 9000 IU; top, middle, and bottom panels) of certoparin.
of 6000 IU resulted in an increase of anti-FXa activity for 24 h before the maximum was reached demonstrating a relatively high and stable anti-FXa activity over a long period of time (up to 48 hours after inhalation). Finally, after inhalation of 9000 IU a maximum of anti-FXa activity was observed after 4 h (Fig. 1). After subcutaneous administration of certoparin, the AUC was 5.70 ±1.58 U/h/ml with an interindividual variability of 28% (Table 2 and Fig. 2). Compared with the subcutaneous route of administration, inhalation of 3000 IU revealed a lower AUC and a higher interindividual variability (2.60 ±1.74 U/h/ml). Inhalation of 6000 IU was followed by an AUC which was not significantly different when compared with subcutaneous administration (7.17 ±3.0 U/h/ml).
609
Fig. 2. Individual values of the areas under the curve (AUC) after subcutaneous application (3000 IU, n=10) or inhalation (3000 IU, 6000 IU, and 9000 I.; n=5, n=5 and n=10, respectively) of certoparin. Table 2. Values of the area under the curve (AUC) after subcutaneous application (3000 IU) or inhalation (3000 IU-9000 IU) of certoparin. Treatment 3000 IU s.c.2)
n
Mean ±SD of the AUC (U•hours/ml)
CV-value of the AUC (%)
P-value1)
10
5.70 ±1.58
28
-----
3)
5
2.60 ±1.74
66
0.01
3)
6000 IU inhal.
5
7.17 ±3.0
41
0.28
9000 IU inhal.3)
10
9.43 ±1.31
13
0.0007
3000 IU inhal.
for the comparison with the subcutaneous administration subcutaneous administration 3) inhalant administration 1) 2)
However, inhalation of certoparin doses of 3000 IU and 6000 IU was performed in only 5 individuals as the first part of the study and, therefore, the results cannot be directly compared with those obtained after subcutaneous administration. The highest inhaled dose of 9000 IU, which was administered to the same 10 subjects, was followed by an AUC of 9.43 ±1.31 U/h/ml (P=0.0007 compared with subcutaneous administration) and showed the lowest interindividual variability (13%) (Table 2 and Fig. 2). DISCUSSION
Since 1965, a number of studies had been published in which the effect of inhaled unfractionated heparin or LWWH on blood coagulation was investigated.
610 These studies demonstrated that inhalative administration of these compounds for anticoagulation was well tolerated and was not followed by relevant pulmonary or systemic side effects (18-25). In addition, studies focusing on the local antiinflammatory and antifibrotic effects of heparin (2, 5, 7, 8, 26, 27) demonstrated the safety of this application technique. The results regarding the safety were also confirmed by the observations in our study in which no acute effects as detected by clinical examination, measurement of lung function parameters, and analysis of blood safety parameters were detectable. Even though long time effects after inhalation of certoparin cannot be excluded based on our study results, they are relatively unlikely because the large number of prior studies confirmed the safety of such type of therapy for heparin and other LMWHs. Previous studies have shown the feasibility of inhalant application of heparin and LMWH (18-25) for anticoagulation, but there was a high variability of the anticoagulative effect among subjects (21). It is likely that the observed high variability of drug action is due to a low reproducibility of the drug amount actually deposited in the lungs, because the principles of pulmonary particle deposition have not been for long well understood (33, 36-40). In consequence, heparin inhalation has not left the experimental stage. However, since about 20 years, there has been large progress due to the development of LMWH (9-13) and advances in aerosol medicine (33, 36-40), promising an improvement of drug inhalation which may help to advance inhalation of heparin and LMWH to clinical feasibility. Recent animal studies show that local deposition of heparin in the lung is of major impact (41). It has been shown that variability of drug deposition in the lungs is mostly due to differences in breathing patterns among subjects and especially due to the use of suboptimal breathing patterns (28). If inhalation is performed using optimized and standardized breathing patterns, peripheral (i.e., alveolar) drug deposition, which is a prerequisite for absorption, is much higher and its variability is lower compared with uncontrolled inhalation (29, 31, 32, 42). Goals of our study were to demonstrate the feasibility of certoparin inhalation, to investigate the pharmacological effect of different doses (3000 IU- 9000 IU), and to minimize the interindividual variability by means of a standardized breathing pattern. For reference we used the subcutaneous administration of 3000 IU, because this dose and way of administration are commonly used in clinical routine for prophylaxis of venous thromboembolism. The intensity and the time course of the anticoagulative effect was determined by measurement of the antiFXa activity in plasma, a method also used in clinical routine for monitoring anticoagulation by means of LMWH. Subcutaneous administration of LMWH leads to a fast increase of plasma antiFXa activity and a high maximum concentration after 2 hours. Thereafter, the anticoagulant activity in plasma rapidly decreased. Kinetics of the anticoagulative effect after inhalation were different from that after subcutaneous administration, and were found to be dose-dependent. Inhalation of 3000 IU, 6000 IU, and 9000 IU were followed by an anticoagulative effect characterized by a rapid onset and
611 prolonged duration. In detail, inhalation of 3000 IU was followed by a first peak of the anticoagulative effect after 30 min, a rapid decay of anti-FXa activity and a broad second peak of the anticoagulative effect, with a maximum after 24 hours. Inhalation of 6000 IU was also followed by a rapid onset of the anticoagulative activity, which was stronger (i.e., higher value of the anti-FXa activity) than after inhalation of 3000 IU certoparin. However, there was no distinct first peak, but instead a slight continuous increase of the anticoagulative activity reaching once more a maximum 24 hours after inhalation. Finally, inhalation of 9000 IU was followed by a delayed peak (maximum 4 hours after inhalation), which was higher than after inhalation of the other doses. Inhalation of certoparin doses of 9000 IU was followed by a sufficient anticoagulative effect for prevention of venous thrombosis (anti-FXa activity above 0.2 U/ml) which lasted until the end of the observation period, which was 48 hours after inhalation. It can be speculated that the observed differences in the intensity and time course of the anticoagulative effect after inhalation compared with subcutaneous administration were caused by another compartment which plays no role after subcutaneous administration (43-46). In this case, the drug fraction which is rapidly released into the blood would be directly absorbed from alveoli, where the drug particles are deposited. However, some amount of the drug may be stored in the epithelium or in alveolar macrophages from which it is slowly released perhaps after partial biochemical degradation. This compartment seems to have a limited capacity, so that a further increase of the deposited drug dose (from 6000 IU to 9000 IU) would not lead to a further increase of the slowly released fraction (as characterized by the second peak after 24 hours). However, in this comparison, it should be noted that inhalation of 6000 I. was performed in 5 subjects only, whereas inhalation of 9000 IU was performed in 10 participants. An analysis of the interindividual variability revealed that the anticoagulative effect after inhalation of 6000 IU was comparable with that after subcutaneous administration of 3000 IU certoparin. For inhalation of 9000 IU, intersubject vartiability was significantly lower. In combination with the data regarding the intensity and duration of the anticoagulative effect described above this suggests that inhalation of a dose of 6000 IU of certoparin might be sufficient for prevention of venous thrombosis. The sustained action observed also implies that a once daily administration of certoparin seems feasible. Our findings may have important implications for the clinical feasibility of heparin inhalation. In the past, it was difficult to judge whether an appropriate antithrombotic activity was achieved after conventional inhalation of heparin. This prevented the application of heparin inhalation outside of clinical studies. Therefore, inhalation of heparin under controlled conditions may lead the way towards clinical feasibility. Furthermore, our data indicate that inhaled LMWH has some advantages which may improve the convenience and logistic requirements of long-term application. First of all, inhalation of LMWH would help to avoid subcutaneous applications, which is more compliant and practical
612 for patients. Another possible advantage is based on the different pharmacological properties. Subcutaneous application of unfractionated heparin has to be administered twice daily to obtain sufficient prophylaxis of thrombosis. In contrast, because of the observed long-time anticoagulative effect of the inhaled LMWH certoparin, an inhalation of this drug may be due only once daily or may be due only every second day. Alternatives to subcutaneous heparin administration may be considered for those patients who require long-term anticoagulant therapy. In these patients, e.g., in oncology or obstetrics, the compliance to frequent subcutaneous injections at home is often limited. Therefore, inhalation of heparin or LMWH would be an attractive alternative. However, the disadvantages and potential side effects of unfractionated heparin and LMWH, like heparin-induced thrombocytopenia or heparin-induced osteoporosis, should be expected until surveillance studies in long-term inhalation of unfractionated heparin or LMWH show a different outcome. Furthermore, up to now there are no data on the bioavailability after inhalant administration in patients with acute or chronic lung disease. Today, the discussion on oral heparin (47, 48) or direct thrombin inhibitors has stalled the development of heparin inhalation. However, it has to be shown that these new drugs are, in respect to bioavailablity and reproducibility, comparable with inhaled LMWH - especially if this is applied through controlled inhalation. Additionally, it should be kept in mind that heparin is not only an anticoagulant drug: The benefit of this drug beyond anticoagulation in patients with asthma (2, 5, 7, 26, 27), the prospect that heparin might slow down the development of interstitial lung disease, like fibrosis (8), and the potential effect in tumor patients, who are also at risk for venous thrombosis (3), may suggest heparin inhalation as a future treatment. In conclusion, onset, intensity, duration, and interindividual variability of the pharmacological effect of inhaled certoparin demonstrate that this type of administration can be applicable in clinical routine for prevention of venous thrombosis if controlled breathing patterns are used. Therefore, this application should be subject of prospective studies in appropriate patients using relevant outcome parameters. REFERENCES 1. Bell WR, Hennebry TA. Heparin and other indirect antithrombin reagents. In Antithrombotics, ACG Uprichard, KP Gallagher (eds). In Handbook of Experimental Pharmacology Vol. 132, GVR Born, P Cuatrecasas, D Ganten, H Herken, K Starke, P Taylor (eds). Berlin, SpringerVerlag, 1999, pp. 259-303. 2. Ahmed T, Gonzalez BJ, Danta I. Prevention of exercise-induced bronchoconstriction by inhaled low-molecular-weight heparin. Am J Respir Crit Care Med 1999; 160: 576-581. 3. Bobek V, Kovarik J. Antitumor and antimetastatic effect of warfarin and heparins. Biomed Pharmacother 2004; 58: 213-219. 4. Köhler D. Aerosolized heparin. J Aerosol Med 1994; 7:307-314.
613 5. Ceyhan BB, Celikel T. Effect of inhaled low molecular weight heparin on methacholineinduced bronchoconstriction. Int J Clin Pharmacol Ther 2000; 38: 446-451. 6. Köhler D, Fleischer W. Medikamente. In Theorie und Praxis der Inhalationstherapie, D Köhler, W Fleischer (eds). München, Arcis Verlag, 2000, pp. 71-99. 7. Tyrrell DJ, Horne AP, Holme KR, Preuss JM, Page CP. Heparin in inflammation: Potential therapeutic applications beyond anticoagulation. Adv Pharmacol 1999; 46: 151-208. 8. Günther A, Lübke N, Ermert M et al. Prevention of bleomycin-induced lung fibrosis by aerosolization of heparin or urokinase in rabbits. Am J Respir Crit Care Med 2003; 168: 1358-1365. 9. Pineo GF, Hull RD. Low molecular weight heparin. In Antithrombotics, ACG Uprichard, KP Gallagher (eds). In Handbook of Experimental Pharmacology, Vol. 132, GVR Born, P Cuatrecasas, D Ganten, H Herken, K Starke, P Taylor (eds). Berlin, Springer-Verlag, 1999, pp. 305-329. 10. Agnelli G, Iorio A, Renga C et al. Prolonged antithrombin activity of low-molecular-weight heparins. Clinical implications for the treatment of thromboembolic diseases. Circulation 1995; 92: 2819-2824. 11. Ansell J, Bergqvist D. Current options in the prevention of thromboembolic disease. Drugs 2004; 64 Suppl 1: 1-5. 12. Warkentin TE, Levine MN, Hirsh J et al. Heparin-induced thrombocytopenia in patients treated with low-molecular-weight heparin or unfractionated heparin. N Engl J Med 1995; 332: 1330-1335. 13. Hirsh J. Low molecular weight heparin. Thromb Haemost 1993; 70: 204-207. 14. Chang P. New anticoagulants for venous thromboembolic disease. Drugs 2004; 7:50-57. 15. Jiao Y, Ubrich N, Hoffart V et al. Anticoagulant activity of heparin following oral administration of heparin-loaded microparticles in rabbits. J Pharm Sci 2002; 91: 760-768. 16. Gonze MD, Salartash K, Sternbergh WC, Baughman RA, Leone-Bay A, Money SR. Orally administered unfractionated heparin with carrier agent is therapeutic for deep venous thrombosis. Circulation 2000; 101: 2658-2661. 17. Arnold J, Ahsan F, Meezan E, Pillion DJ. Nasal administration of low molecular weight heparin. J Pharm Sci 2002; 91: 1707-1714. 18. Rosner SW. Heparin administered as an aerosol. Vasc Dis 1965: 2: 131-134. 19. Bendstrup KE, Gram J, Jensen JI. Effect of inhaled heparin on lung function and coagulation in healthy volunteers. Eur Respir J 2002; 19: 606-610. 20. Bick RL, Ross ES. Clinical use of intrapulmonary heparin. Semin Thromb Hemost 1985; 11: 213-217. 21. Hellgren M, Hägnevik K, Blombäck M. Heparin aerosol - effect on blood coagulation and pulmonary function. Thromb Res 1981; 21: 493-502. 22. Jaques LB, Mahadoo J, Kavanagh LW. Intrapulmonary heparin - A new procedure for anticoagulant therapy. Lancet 1976; 27: 1157-1161. 23. Kavanagh LW, Jaques LB. A new route of heparin administration - The lung. Arzneimittelforschung 1976; 26: 389-392. 24. Lewandowski K, Psuja P, Tokarz A, Zozulinska M, Sowier J, Zawilska K. Antikoagulant activity in the plasma after a single administration of nebulized heparin or LMW Heparin fraction (Fraxiparine) in patients undergoing abdominal surgery. Thromb Res 1990; 58: 525-530. 25. Wright CJ, Mahadoo J, Jaques LB. Anticoagulant activity and operative blood loss after intrapulmonary heparin. Brit J Surg 1979; 66: 844-847. 26. Bendstrup KE, Jensen JI. Inhaled heparin is effective in exacerbations of asthma. Respir Med 2000; 94: 174-175. 27. Diamant Z, Page CP. Heparin and related molecules as a new treatment for asthma. Pulm Pharmacol Ther 2000; 13: 1-4. 28. Brand P, Friemel I, Meyer T, Schulz H, Heyder J, Häußinger K. Total deposition of therapeutic particles during spontaneous and controlled inhalations. J Pharmaceut Sci 2000; 89: 724-731.
614 29. Brand P, Beckmann H, Maas Enriquez M et al. G. Peripheral deposition of α1-protease inhibitor using commercial inhalation devices. Eur Resp J 2003; 22: 263-267. 30. Brand P, Maas Enriquez M et al. Neue Strategien zur kontrollierten Inhalation von α1Antitrypsin. Pneumologie 2003; 57: 153-158. 31. Griese M, Ramakers J, Krasselt A et al. Improvement of alveolar glutathione and lung function but not oxidative state in cystic fibrosis. Am J Respir Crit Care Med 2004; 169: 822-828. 32. Scheuch G, Meyer T, Müllinger B et al. Controlled inhalation improves peripheral lung deposition. J Aerosol Med 2001; 14: 388 (Abstract). 33. Scheuch G, Kohlhaeufl MJ, Brand P, Siekmeier R. Clinical perspectives on pulmonary systemic and macromolecular delivery. Adv Drug Deliv Rev 2006; 58: 996-1008. 34. Quanjer PH, Tammeling GJ, Cotes JE, Pedersen OF, Peslin R, Yernault JC. Lung volumes and forced ventilatory flows. Eur Respir J 1993; 6: 5-40. 35. Brand P, Meyer T, Sommerer K, Weber N, Scheuch G. Alveolar deposition of monodisperse aerosol particles in the lung of patients with COPD. Exp Lung Res 2002; 28: 39-54. 36. Heyder J, Gebhart J, Rudolf G, Schiller CF, Stahlhofen W. Deposition of particles in the human respiratory tract in the size range 0.005-15 µm. J Aerosol Sci 1986; 17: 811-825. 37. International Commission on Radiological Protection. Human respiratory model for radiological protection. Ann ICRP 1994; 24: 1-120. 38. Köbrich R, Rudolf G, Stahlhofen W. A mathematical model of mass deposition in man. Ann Occup Hyg 1994; 38: 15-23. 39. Scheuch G, Stahlhofen W. Deposition and dispersion of aerosols in the airways of the human respiratory tract: The effect of particle size. Exp Lung Res 1992; 18: 343-358. 40. Schulz H, Brand P, Heyder J. Particle deposition in the respiratory tract. In Particle-Lung Interactions, P Gehr, J Heyder (eds). In Lung Biology in Health and Disease, Vol. 143, C Lenfant (ed). New York, Basel, Dekker, 2000, pp. 229-290. 41. Qi Y, Zhao G, Liu D, Shriver Z et al. Delivery of therapeutic levels of heparin and low-molecularweight heparin through a pulmonary route. Proc Natl Acad Sci USA 2004; 101: 9867-9872. 42. Siekmeier R. Scheuch G. Systemische Therapie mit Aerosolen. Allgemeine Grundlagen zur pulmonalen Verabreichung von Makromolekülen zur systemischen Therapie. AtemwLungenkrkh 2005; 31: 374-386. 43. Gupta PK, Hickey AJ. Contemporary approaches in aerosolized drug delivery to the lung. J Control Release 1991; 17: 129-148. 44. Niven R.W. Delivery of biotherapeutics by inhalation aerosol. Crit Rev Ther Drug Carrier Syst 1995; 12: 151-231. 45. Patton JS, Platz RM. Routes of delivery: Case studies. (2) Pulmonary delivery of peptides and proteins for systemic action. Adv Drug Deliv Rev 1992; 8: 179-196. 46. Wolff R.K. Safety of inhaled proteins for therapeutic use. J Aerosol Med 1998; 11: 197-219. 47. Gustafsson D. Oral direct thrombin inhibitors in clinical development. J Intern Med 2003; 254: 322-334. 48. Hiebert LM. Oral heparins. Clin Lab 2002; 48: 111-116. Authors address: R. Siekmeier, Bundesinstitut für Arzneimittel und Medizinprodukte (BfArM), Kurt-Georg-Kiesinger-Allee 3, 53175 Bonn; e-mail:
[email protected]
