Abstract
Rationale: Patients with acute lung injury have impaired function of the lung surfactant system. Prior clinical trials have shown that treatment with exogenous recombinant surfactant protein C (rSP-C)-based surfactant results in improvement in blood oxygenation and have suggested that treatment of patients with severe direct lung injury may decrease mortality.
Objectives: Determine the clinical benefit of administering an rSP-C–based synthetic surfactant to patients with severe direct lung injury due to pneumonia or aspiration.
Methods: A prospective randomized blinded study was performed at 161 centers in 22 countries. Patients were randomly allocated to receive usual care plus up to eight doses of rSP-C surfactant administered over 96 hours (n = 419) or only usual care (n = 424).
Measurements and Main Results: Mortality to 28 days after treatment, the requirement for mechanical ventilation, and the number of nonpulmonary organ failure–free days were not different between study groups. In contrast to prior studies, there was no improvement in oxygenation in patients receiving surfactant compared with the usual care group. Investigation of the possible reasons underlying the lack of efficacy suggested a partial inactivation of rSP-C surfactant caused by a step of the resuspension process that was introduced with this study.
Conclusions: In this study, rSP-C–based surfactant was of no clinical benefit to patients with severe direct lung injury. The unexpected lack of improvement in oxygenation, coupled with the results of in vitro tests, suggest that the administered suspension may have had insufficient surface activity to achieve clinical benefit.
Clinical trial registered with www.clinicaltrials.gov (NCT00074906).
Prior studies have suggested that treatment with exogenous surfactant of patients with severe direct lung injury may be beneficial.
In this prospective, blinded, randomized study of 843 patients, delivery of a recombinant surfactant protein C–based surfactant provided no benefit to patients with severe direct lung injury.
Rationale for treatment of patients with ALI with exogenous surfactant includes supportive observations in animal models, improvement in pulmonary gas exchange in most clinical trials of surfactant replacement, and decreased mortality in pediatric patients with ALI treated with exogenous surfactant (2, 4). However, mortality of adults with ALI has been unaffected by treatment with exogenous surfactant (5), although post hoc subgroup analysis showed that adults with severe direct lung injury (from pneumonia or aspiration) might benefit (6). This benefit could result, in part, through reduction of mechanical stress on components of the lung parenchyma and reduction of ventilator-induced lung injury (7).
Improvement in blood oxygenation occurs in almost all trials in which natural surfactants (4, 8–12) or synthetic surfactants (13, 14) have been administered. The exceptions are trials of Exosurf, a nonprotein-containing formulation (15, 16), and of HL-10 surfactant (17).
The synthetic surfactant containing recombinant surfactant protein C (rSP-C), phospholipids, and palmitic acid (rSP-C surfactant; Nycomed GmbH, Konstanz, Germany) has excellent surface activity and markedly improves gas exchange in animal models of lung injury (18–20). All patient groups receiving rSP-C surfactant have had improvement in blood oxygenation (6). We report here a phase III prospective, randomized, parallel-group, double-blind, controlled multinational study to test whether rSP-C surfactant treatment of patients with severe direct lung injury reduces mortality.
Patients between 12 and 85 years of age with severe impairment of gas exchange (PaO2/FiO2 ≤ 170 mm Hg) due to aspiration of gastric contents or pneumonia were eligible for study. Patients were not eligible if they had evidence of a source of infection or sepsis outside the lung. Complete diagnostic criteria and inclusion and exclusion criteria are listed in the online supplement.
Patients were recruited during the period November 2003 to March 2008 from the intensive care units of 161 medical centers in 22 countries. The study, registered with clinicaltrials.gov identifier NCT00074906, was performed in accordance with the declaration of Helsinki (1996), the rules of International Conference on Harmonization of Technical Requirements for the Registration of Pharmaceuticals for Human Use–Good Clinical Practice Consolidated Guideline E6, CPMP/ICH/135/95 and national legal stipulations. All patients or their legal representatives provided written informed consent, and the study protocol was approved by independent ethics committees or institutional review boards at each center. The study was monitored by an independent Data and Safety Monitoring Board.
Patients were prescreened at each study site; the methods of prescreening were not stipulated. Final screening was performed by a Scientific Enrollment Coordinating Board (SECB), composed of an international panel of trained intensivists, to which investigators submitted a prescreened study patient via internet or telephone (21). A member of the SECB then discussed the patient with the site investigator and approved or rejected the patient for enrollment based on compliance with study inclusion and exclusion criteria. Patients could be approved conditionally, contingent on meeting study criteria during the defined protocol enrollment period. Approved patients for whom consent had been obtained were enrolled in a 2-hour baseline observation period during which blood samples and clinical observations were obtained. At the conclusion of this baseline period, patients who continued to meet inclusion criteria were randomized to a usual care group or a usual care plus surfactant treatment group (Figure 1). Although investigators were explicitly advised to use the Acute Respiratory Distress Syndrome (ARDS) Network lung protective ventilation strategy (7), patients were not required to have bilateral pulmonary opacities on the chest radiograph, and thus not all patients fulfilled the criteria for ARDS.
Figure 1. Flow of patients through the trial. In the analysis of patients receiving at least one study treatment, 419 patients receiving surfactant and 424 patients not receiving surfactant were available for analysis. SEBC = Scientific Enrollment Coordinating Board.
[More] [Minimize]Patients who were approved by the SECB were immediately randomized, using a computer-based approach (22), on a 1:1 basis to one of the two groups. Randomization was stratified by center and concealed from the investigators. After collection of data during a 2-hour baseline period, patients were treated with 1 ml rSP-C surfactant per kg lean body weight (each ml containing 1 mg rSP-C and 50 mg phospholipids) exactly as described previously (14). Blinding was accomplished using a weighted blinding bag as previously described (14). Patients received a maximum of seven additional administrations at 6, 12, 24, 36, 48, 72, and 96 hours after the initial treatment provided they remained intubated and mechanically ventilated with a positive end-expiratory pressure (PEEP) greater than or equal to 5 cm H2O and with a PaO2/FiO2 value in the range of 60 to 170 mm Hg.
Before each administration, the dry rSP-C surfactant powder was reconstituted with sterile saline, drawn into a syringe, and, in a step not used in prior trials, passed forcefully six times through a Luer lock adapter with a 3.8-mm internal diameter orifice to a second syringe so that shear forces generated in the narrow channel would augment suspension of the surfactant. Nonsheared as well as sheared rSP-C surfactant had been demonstrated by the manufacturer to have a surface pressure less than 40 mN/m measured in vitro at a concentration of 25 mg phospholipids/ml, a surface age of 0.1 seconds, at 37°C in a maximum bubble pressure tensiometer of the manufacturer's design. In vivo testing using the lavaged rat lung (23) showed that nonsheared as well as sheared rSP-C surfactant, at a concentration of 100 mg phospholipid/kg, restored the PaO2 to not less than 400 mm Hg 120 minutes after administration.
The primary study outcome variable was the percentage of patients alive at Day 28. Secondary outcome variables of major interest included: ventilator-free days (VFD) at Day 28 (assigning 0 VFD to nonsurvivors), measures of oxygenation and ventilation, and the number of nonpulmonary organ failure–free days (the definitions of organ failure are provided in the online supplement).
Sample size was calculated based on expected 28-day mortality. A group sequential design with two interim and one final analysis was planned, and a one-sided test procedure was chosen with α = 0.025 and the power set to 80%. Based on results of the prior pooled analysis (6), we estimated an overall mortality difference of 7.6% (25.8 vs. 33.4%). A sample size of 1,132 patients was needed for 80% power based on the chi-square test. Because a group-sequential design was chosen, the sample size was slightly higher, and final plans called for randomization of 1,200 patients.
As a result of a first planned interim analysis after enrollment of 400 patients, the study Data and Safety Monitoring Committee recommended continuation of the study. After a second planned interim analysis after enrollment of 800 patients, the Committee recommended termination of the study for futility.
Because of the unexpected observation of complete lack of effect, including lack of effect on blood oxygenation, further scrutiny was applied to all study-related procedures after termination of the study. To examine the effect of the shearing maneuver on rSP-C surfactant, surface tension–lowering activity was evaluated using a pulsating bubble surfactometer as reported previously (24). For this purpose, rSP-C surfactant from a batch used in the clinical trial was resuspended either by gentle swirling (as in prior clinical studies) or using the protocol described for this study in which shearing occurred during passage through a Luer lock adaptor. Because site investigators might introduce small amounts of air into the syringe when aspirating surfactant from the vial in which it is initially resuspended, shearing was performed without the introduction of additional air, or after addition of 0.1, 1, or 2 ml of air to the syringe containing surfactant (to give a final volume in the syringe of 20 ml). After incubation of samples for 30 minutes at room temperature, sheared and nonsheared surfactant was diluted to 0.5 or 1.0 mg phospholipid/ml and tested in a pulsating bubble surfactometer. Three independent resuspensions were made, and at least five samples of each were tested. In addition, the susceptibility of sheared or nonsheared rSP-C surfactant to inhibition by fibrinogen was tested. For this purpose, human fibrinogen (kindly provided by Prof. Heimburger, Behringwerke, Marburg, Germany) was added to the resuspended surfactant preparations at a final concentration of 0, 0.05, 0.1, or 0.5 mg/ml as described previously (25).
All data analysis was performed according to a preestablished analysis plan. The hypothesis that the odds ratio for mortality of treated patients, compared with untreated patients, is greater than 1 was evaluated using a group sequential approach with overall one-sided type I error rate of 0.025. A two-sided 95% confidence interval was calculated for this value and for mortality rates and the difference in mortality rates. Comparison of the control group and the treatment group with regard to the primary efficacy variable was performed using a logistic regression model with pneumonia, aspiration, age, Acute Physiology and Chronic Health Evaluation II score at baseline, and region (North America versus Europe and all other countries) as influencing baseline variables. Secondary variables were tested by a logistic regression approach, a one-sided Wilcoxon test, or a one-sided Fisher exact test.
The SECB screened 1,382 patients. Numbers of patients enrolled, randomized, treated, and completing treatment are shown in Figure 1. Very few patients did not complete the study; only one discontinued the intervention due to an adverse event (hypoxemia). Patients were enrolled from 22 countries (see Table E1 in the online supplement). Baseline demographic data are shown in Table 1, and baseline clinical and physiologic data are shown in Table 2. As illustrated in these tables, the two treatment groups were well matched. Patients to whom surfactant was delivered received 5.2 ± 2.4 (mean ± SD) doses, whereas patients in the control group were administered (into the blinding bag) 5.4 ± 2.3 doses. The numbers of doses administered and information on protocol compliance are detailed in Tables E2 and E3. The percentage of treatment and retreatment surfactant administrations that were compliant with the protocol, as detailed in the online supplement, was greater than 98% for both groups.
Demographic Data | Usual Care + Surfactant (N = 419) | Usual Care (N = 424) | Overall (N = 843) |
|---|---|---|---|
| Age, yr | 57.5 ± 0.8 | 56.5 ± 0.83 | 57.0 ± 0.58 |
| Body height, cm | 171.0 ± 0.51 | 171.4 ± 0.45 | 171.2 ± 0.34 |
| Body weight, kg | 80.4 ± 0.92 | 80.4 ± 0.86 | 80.4 ± 0.63 |
| Ethnic origin, white, % | 377 (90.0) | 390 (92.0) | 767 (91.0) |
| Sex, female/male, % | 33.9/66.1 | 32.8/67.2 | 33.3/66.7 |
Surfactant Plus Usual Care (N = 419) | Usual Care (N = 424) | Overall (N = 843) | |
|---|---|---|---|
| Time from hospital admission to baseline, h | 100.9 ± 7.48 | 107.4 ± 6.88 | 104.2 ± 5.08 |
| Time from ICU admission to baseline, h | 48.9 ± 1.86 | 48.9 ± 2.01 | 48.9 ± 1.37 |
| Time from intubation to baseline, h | 37.2 ± 0.80 | 37.3 ± 0.81 | 37.3 ± 0.57 |
| Aspiration of gastric contents present | 82 (19.6) | 92 (21.7) | 174 (20.6) |
| Pneumonia present | 359 (85.7) | 369 (87.0) | 728 (86.4) |
| Pneumonia, mode of infection | |||
| Community acquired | 292 (81.3) | 284 (77.0) | 576 (79.1) |
| Hospital acquired | 67 (18.7) | 85 (23.0) | 152 (20.9) |
| APACHE II score | 18.0 ± 0.33 | 17.8 ± 0.32 | 17.9 ± 0.23 |
| Quadrants involved on chest radiograph, n | 2.81 ± 0.05 | 2.85 ± 0.05 | 2.83 ± 0.04 |
| ARDS present at baseline | 245 (58.5) | 249 (58.7) | 494 (58.6) |
| SIRS present at baseline | 373 (89.0) | 366 (86.3) | 739 (87.7) |
| Cardiovascular support with vasopressors present at baseline | 252 (60.1) | 244 (57.5) | 496 (58.8) |
| FiO2 | 0.64 ± 0.007 | 0.64 ± 0.007 | 0.64 ± 0.005 |
| PaO2 | 76.9 ± 3.8 | 76.4 ± 3.7 | 76.7 ± 2.6 |
| PaCO2 | 46.9 ± 2.3 | 46.6 ± 2.3 | 46.7 ± 1.6 |
| PaO2/FiO2 | 123.8 ± 1.30 | 124.1 ± 1.32 | 123.9 ± 0.93 |
| PEEP, cm H2O | 11.1 ± 0.2 | 11.0 ± 0.1 | 11.1 ± 0.1 |
| Pplat, cm H2O | 24.2 ± 0.4 | 24.9 ± 0.4 | 24.6 ± 0.3 |
| Vt, ml/kg PBW | 7.4 ± 0.4 | 7.5 ± 0.4 | 7.5 ± 0.3 |
| Vt, ml | 479 ± 5.7 | 489 ± 5.6 | 484 ± 4.0 |
| Loge IL-6 | 5.38 ± 0.66 | 5.35 ± 0.62 | 5.36 ± 0.45 |
The study did not meet its primary endpoint of showing significantly reduced mortality to Day 28 for the patients receiving surfactant (22.7 vs. 23.8% for standard care alone, P = 0.26; Figure E1). Subgroup analysis showed no difference in mortality for groups defined by mechanism of direct lung injury (aspiration or pneumonia), presence of ARDS, or geographic location (Table 3). Additional analyses, including Cox regression analysis, Cochran-Mantel-Haenszel model analysis, and further logistic regression analysis, failed to show any 28-day mortality reduction for the group receiving surfactant. The proportion of patients alive was similar between the groups at 3 months (64.9% for surfactant plus usual care vs. 65.6% for usual care alone, P = 0.48) and 6 months (62.5 vs. 63.9%, P = 0.57).
Surfactant Plus Usual Care* (N = 419) | Usual Care* (N = 424) | Odds Ratio† (95% CI) | P Value, 1-sided | |
|---|---|---|---|---|
| All cases | 324 (77.3) | 323 (76.2) | 0.90 (0.64–1.25) | 0.26 |
| Patients with aspiration | 66 (80.2) | 75 (81.5) | 0.90 (0.41–1.99) | 0.40 |
| Patients with pneumonia | 276 (76.9) | 280 (75.9) | 0.91 (0.64–1.31) | 0.31 |
| Patients with ARDS | 181 (73.9) | 180 (72.3) | 0.88 (0.58–1.34) | 0.27 |
| Patients from North America | 73 (79.3) | 70 (81.4) | 1.16 (0.51–2.61) | 0.64 |
| Patients from Europe and other regions | 251 (76.8) | 253 (74.9) | 0.86 (0.60–1.24) | 0.21 |
Both groups had improved oxygenation after randomization; the administration of surfactant did not result in more pronounced improvement compared with usual care alone. The mean PaO2/FiO2 ratios for the surfactant plus usual care group and for the usual care alone group were, respectively, 123.8 and 124.1 mm Hg (at baseline), 150.2 and 146.0 mm Hg (at 24 h), 160.7 and 159.9 mm Hg (at 48 h), and 164.8 and 167.0 mm Hg (at 96 h) (Figure 2). Excess area under the PaO2/FiO2 curve from baseline to 48 hours for surfactant plus usual care versus usual care alone was 1,026.5 and 840.1 mm Hg × h (median values, P = 0.34), and thus not different between groups.
Figure 2. PaO2/FiO2 values for patients receiving surfactant plus usual care or usual care only from the time of randomization to 96 hours. Arrows indicate the time of possible surfactant administrations. Differences in PaO2/FiO2 values between groups are not significant.
[More] [Minimize]There was no significant difference in the median number of VFDs at Day 28 (9.0 d for surfactant plus usual care and 10.0 d for usual care alone). The PEEP values for the surfactant plus usual care group and for the usual care alone group were, respectively, 11.1 and 11.0 (at baseline), 10.3 and 10.7 (at 48 h), and 9.7 and 10.1 (at 96 h). The reductions from baseline in PEEP in the surfactant plus usual care group were significantly greater at 48 hours (P = 0.006) and 96 hours (P = 0.024) than in the usual care only group (Figure E2). No other differences in peak inspiratory pressure, plateau pressure, PEEP, or FiO2 between the two groups were significant. The modified lung injury score (26) decreased uniformly and without intergroup differences from a median (range) of 2.67 (1.33, 4.00) at baseline to 2.33 (0.00, 4.00) on Day 6.
The number of patients with nonpulmonary organ failures decreased steadily during the 28-day observation period. On study Day 1, 311 patients (74.2%) in the surfactant plus usual care group and 294 patients (69.3%) in the usual care group had nonpulmonary organ failures. On study Day 28, these values were 139 patients (33.2%) and 146 patients (34.4%), respectively. There was no difference in nonpulmonary organ failure–free days between the groups (19.0 vs. 17.0 d, P = 0.3917).
Serious adverse events were reported for 207 patients (49.4%) who received surfactant plus usual care and for 198 patients (46.7%) who received only usual care. Treatment-related serious adverse events (predominately hypoxia or airway obstruction) were reported for 30 patients (7.2%) receiving surfactant plus usual care and for 5 patients (1.2%) who received only usual care (P < 0.001, chi-square test; Table E4).
To investigate the apparent absence of a surfactant effect on gas exchange, we investigated the effect of the shearing step, which had been added to preparation of the surfactant suspension subsequent to prior trials, on surface tension–lowering activity. Shearing demulsified the surfactant emulsion (Figure 3), impaired function both in the presence or absence of added air when the suspension was diluted to concentrations of less than 2 mg phospholipid/ml (Figure 4), and increased susceptibility to inhibition of surface activity by fibrinogen at the same low concentrations (Figure 5).
Figure 3. Shearing in the presence of air demulsifies the surfactant emulsion. Samples of surfactant were resuspended to a concentration of 50 mg/ml with saline by gentle swirling. Samples were then: (A) not further treated, (B) subjected to shearing after careful exclusion of air from the Luer connector, (C) subjected to shearing without excluding air from the Luer connector, (D) subjected to shearing after addition of 2 ml of air. Samples were allowed to stand for 30 minutes. Surfactant in samples A and B showed minimal sedimentation of the emulsion, whereas surfactant in samples C and D showed marked demulsification.
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Figure 4. Shearing impairs surfactant function. Samples of surfactant were resuspended to a concentration of 50 mg/ml with saline by gentle swirling and subsequently sheared in the absence of air, or in the presence of 0.1, 1, or 2 ml air, or not sheared. Samples were then diluted with saline to concentrations of 0.5 or 1.0 mg/ml phospholipid (PL) and analyzed in a pulsating bubble surfactometer. Minimum surface tension after 5 minutes' pulsation is shown. Samples sheared in the presence of air had significantly impaired surface activity at both concentrations, and the sample sheared in the absence of air had significantly impaired surface activity when tested at 0.5 mg/ml PL. The mean ± SD of five measurements is given. Significance versus unsheared samples (t test) is indicated by *P > 0.05, **P < 0.01, and ***P < 0.001.
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Figure 5. Shearing increases the susceptibility of surfactant to inhibition of surface activity by fibrinogen. surfactant prepared as described in Figure 2 was analyzed in a bubble surfactometer in the presence of various concentrations of fibrinogen or in the absence of fibrinogen.
[More] [Minimize]The results of this study showed that bolus administration of a recombinant SP-C–based surfactant to patients with severe direct lung injury, as performed in this study, did not reduce mortality or improve pulmonary gas exchange. The study was stopped for futility at a planned 800-patient interim analysis, at which time 844 patients had actually been randomized. Overall mortality to Day 28 was 23.3% and did not differ between study groups. This mortality rate is similar to that reported in recent studies from the National Heart, Lung, and Blood Institute ARDS Network (27), although enrollment criteria for ARDS Network studies and for this study differ with respect to requirements, in the ARDS Network studies, for bilateral opacities on the chest radiograph and a PaO2/FiO2 value not greater than 300 mm Hg.
To our surprise, and in contrast to prior phase II (13) and phase III (14) studies of rSP-C surfactant, no significant improvement in secondary study outcome variables, including measures of gas exchange and ventilatory parameters, was observed. Although reduction in baseline PEEP values was significantly greater at 48 and 96 hours in the surfactant-treated group, the actual differences between groups were unlikely to be of clinical relevance. Likewise, the incidence of nonpulmonary organ failures and the number of organ failure–free days did not differ between study groups.
The number of serious adverse events also did not differ between study groups and was rather high (48% overall), consistent with the level of critical illness in the study population. Serious adverse events that were likely or definitely related to treatment were more frequent in the group treated with surfactant (7.2%) than in the usual care only group (1.2%) and were due predominately to transient hypoxemia or airway obstruction associated with bolus administration of surfactant. In addition, the rate of adverse events is similar to that seen in the control group of a recently published study of HL-10 surfactant treatment of patients with acute lung injury (55.2%) but considerably lower than seen in the treatment arm of that study (76.6%) (17).
Although the results of this study are consistent with prior reports of surfactant having no overall benefit for patients with severe lung injury (5), they are surprising in light of prior experience with rSP-C surfactant (6, 13, 14). Consistently, in that experience, patients treated with rSP-C surfactant had a significantly greater improvement in blood oxygenation than occurred in untreated patients. Thus, it is puzzling that in the study described in this report, in which the dose and dose volume of rSP-C surfactant were identical to those used previously in two phase III trials, absolutely no evidence for an improvement in gas exchange was detected.
It is unlikely that differences in the study population account for this discrepancy. In contrast to previous trials using rSP-C surfactant, the study population reported here consisted exclusively of patients with severe direct lung injury. Although the previous rSP-C surfactant studies targeted patients with ARDS regardless of predisposing event, the subpopulation of patients with severe direct lung injury who received surfactant had the greatest improvement in blood oxygenation and the least mortality (6) and was therefore chosen as the target population for the current study.
Another potential explanation for the absence of improvement in oxygenation could be systematic errors in surfactant administration (e.g., errors in assignment of the blinding devices). However, the presence of more treatment-related adverse events in the surfactant-treated group than in the usual care only group is strong indirect evidence that treatment assignment and administration were consistent.
To improve dispersion of the rSP-C surfactant during suspension, a shearing step was added to the surfactant preparation protocol for the current study. This step, not previously used, may have resulted in administration of surfactant with impaired surface tension–lowering properties. Indeed, testing of sheared and nonsheared rSP-C surfactant subsequent to termination of the clinical trial provided indication that such shearing results in demulsification of the surfactant emulsion and a partial loss of surface activity when the surfactant suspension is diluted to low concentrations, as may be present in alveoli of an inflamed and edematous lung. This effect was also evident in the presence of inhibitory plasma proteins, such as fibrinogen. Based on these data we propose that shearing-induced loss of surface activity of the rSP-C surfactant may have contributed to the lack of efficacy observed in our study.
One must ask, however, why shearing appeared to have little adverse effect in the in vivo rat model yet clearly inhibited in vitro function in the bubble surfactometer. We consider at least two possibilities. First, the rat lavage model used for release testing was treated with rSP-C surfactant at a concentration of 100 mg phospholipid/kg, whereas patients were treated with 50 mg phospholipid/kg. In the studies performed after the trial was stopped, it became apparent that dilution of the surfactant preparation may be important in uncovering subtle changes in function (as may possibly be produced by shearing). Second, the rat model is an imperfect model of the severe lung injury present in patients, and it is quite possible that results in the model do not perfectly predict results in the clinical setting.
In summary, up to eight doses of an rSP-C surfactant suspension administered as an intratracheal bolus to patients with severe direct lung injury did not reduce 28-day mortality or improve blood oxygenation. No differences in mortality between patients receiving surfactant plus usual care or only usual care were detected. Shearing of the surfactant during resuspension may have resulted in impairment of surface tension–lowering function and increased susceptibility to inhibition by plasma proteins, hence explaining the apparent lack of clinical efficacy.
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