© 2004 American Thoracic Society
Aerosolized Urokinase in Pulmonary FibrosisTo the Editor:In a recent issue, we have with great interest noted the editorial by Dr. Idell (1) and the report by Günther and colleagues (2). Idell and also Günther and colleagues suggest that alveolar administration of a plasminogen activator has great potential in treatment of diseases associated with evolution of pulmonary fibrosis—a statement that agrees well with the conclusions from our own published studies (3–6). In his editorial, Idell (1) mentions that scu-PA can be a more effective blocker of lung fibrosis than the two-chain form of urokinase used by Günther and colleagues (2). Idell does not mention that we have shown in a double-blinded, placebo-controlled study that aerosolization of single-chain urokinase plasminogen activator increases the fibrinolytic activity in bronchoalveolar lavage fluid significantly in pigs with acute lung injury caused by severe gunshot trauma (4). We also have demonstrated that a jet nebulizer generates a more favorable particle profile of scu-PA for lower respiratory tract deposition (MMD 2.98 µm) compared with the ultrasonic nebulizer (MMD 3.69 µm) (5). In addition, the ultrasonic nebulization denatures scu-PA, causing a 50% loss of enzymatic activity (5). Thus, the potential positive effects of urokinase reported by Günther and colleagues (2) are not accurate (dose/response) as these authors used ultrasonic nebulization in their study. In the METHODS section, Günther and colleagues (2) describe that the pulmonary deposition of urokinase using radioactively labeled (99mTc) material was 7.8%. We wonder whether they used a direct labeling of urokinase with 99mTc or "the soup method." The latter assumes that the radioactivity follows the drug into the aerosol particle, but this assumption is dubious. Regarding the direct labeling with 99mTc, it is our experience that the stability following a direct labeling was critically low and variable (pretinning method), ranging from 82 to 72% of 99mTc bound to scu-PA. We believe that direct labeling of scu-PA with 99mTc is very difficult, if not impossible (6). Despite these critical comments, the study of Günther and colleagues (2) is of great interest and confirms our published results demonstrating that aerosolized plasminogen activator can be administrated without systemic effects or local bleeding complications (3, 4, 6).
University of Southern Denmark Esbjerg, Denmark FOOTNOTES Conflict of Interest Statement: A-M.B.M., J.G., and J.S. have no declared conflict of interest. REFERENCES
From the Authors: Münster and colleagues address three important aspects of our article (1) and set up questions concerning the accuracy of our data, especially in view of the nebulization of urokinase (u-PA). We would like to respond to these three items. First, as has been stated in the online repository of our manuscript, the currently used ultrasonic device (PulmoSonic; DeVilbiss, Langen, Germany) produces a mean mass aerodynamic diameter (MMAD) of 2.73 µm and is thus even more favorable as compared with the MMAD indicated by Münster and colleagues (2) for their jet nebulizer (2.98 µm). We doubt that it may be possible at all to make any general statement as to the MMAD of different types of nebulizers. Instead, each specific device has to be checked for aerosol properties. This was done in our case and the results are given. Second, denaturation of urokinase is likely to occur during ultrasonic delivery. To some reasonable extent it even occurs under conditions of jet nebulization (Günther and colleagues, personal observations). We believe, however, that there is no true alternative approach for inhalative delivery of u-PA under spontaneous breathing conditions in small animals. Therefore, this loss of activity of u-PA was accepted and openly indicated in the online repository (1) and was overcome by using appropriately high dosages of u-PA. The effective dose of urokinase, as calculated with respect to the loss of activity caused by ultrasonic nebulization, has been given in both the print version of the manuscript and the online repository. Moreover, we have proven a local increase in fibrinolytic activity on u-PA delivery under conditions of bleomycin-induced lung injury, and thus ascertained that the effective activity being delivered is suitable for correction of the altered fibrinolysis balance. Therefore, we cannot see any missing accuracy in our report; instead, our data are very clearcut and comprehensive. Third, Münster and colleagues correctly state that direct labeling of u-PA with 99mTc is somewhat difficult and prone to pitfalls. It is also for this reason that our deposition studies were performed primarily by mixing the u-PA with the 99mTc stock solution. Having in mind the potential pitfalls of direct labeling, such an approach is—at least in our eyes—the most reasonable one and allows the assessment of the pulmonary deposition rate.
Justus-Liebig-University Giessen Giessen, Germany FOOTNOTES Conflict of Interest Statement: A.G. has no declared conflict of interest. REFERENCES
From the Author: Münster and colleagues have indeed provided valuable contributions to the field and confirmed the feasibility of using aerosolized single-chain urokinase (scu-PA) to increase fibrinolytic activity in bronchoalveolar lavage fluid (1, 2). On the other hand, Günther and colleagues (3) used a different preparation—two-chain urokinase rather than scu-PA—to extend this work in a novel interventional context. They acknowledge that the ultrasonic nebulization technique they used results in an approximate loss of 75% of the enzymatic activity (see online data supplement for Reference 3). These observations are consistent with the findings of decreased scu-PA activity using a similar aerosol delivery strategy, as noted by Münster and colleagues (2). Although the technical issues posed by Münster and colleagues broach the question of precisely how much fibrinolysin was delivered to the lung in the Günther study (3), they do not obviate the salient findings relating to the effect of the fibrinolysin (or heparin) aerosols on fibrotic outcome after acute lung injury. Günther and colleagues (3) not only were able to increase fibrinolytic activity in the bronchoalveolar lavage of rabbits with bleomycin-induced lung injury using ultrasonic nebulization and a dosing strategy predicated on prior studies in which two-chain u-PA was aerosolized in isolated lung preparations, but they also were able to demonstrate protection against the clinically relevant endpoint of pulmonary fibrosis (3, 4). On a finer point, the notion that scu-PA is a more effective blocker of lung fibrosis than the two-chain form was presented in the editorial as speculation rather than fact (5). The feasibility and safety profile of aerosolized administration of scu-PA noted by Münster and colleagues buttress the editorial position that further preclinical interventional trials are warranted. Such trials, using comparable aerosolized delivery of scu-PA or two-chain urokinase, are needed to resolve the issue as to whether either agent provides better protection against the development of pulmonary fibrosis after acute lung injury.
University of Texas Health Center at Tyler Tyler, Texas FOOTNOTES Conflict of Interest Statement: S.I. has no declared conflict of interest. REFERENCES
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