Survival.

<p>Kaplan-Meier curves represent the 48 hour survival of preterm lambs which underwent surfactant inactivation when treated either with CHF 5633 (dotted line) or Poractant alfa (continuous line). Analysis by Gehan-Breslow-Wilcoxon test indicated a significant longer survival of lambs treated with CHF 5633 compared to Poractant alfa treated lambs (p<0.05).</p

New Surfactant with SP-B and C Analogs Gives Survival Benefit after Inactivation in Preterm Lambs

<div><h3>Background</h3><p>Respiratory distress syndrome in preterm babies is caused by a pulmonary surfactant deficiency, but also by its inactivation due to various conditions, including plasma protein leakage. Surfactant replacement therapy is well established, but clinical observations and <em>in vitro</em> experiments suggested that its efficacy may be impaired by inactivation. A new synthetic surfactant (CHF 5633), containing synthetic surfactant protein B and C analogs, has shown comparable effects on oxygenation in ventilated preterm rabbits versus Poractant alfa, but superior resistance against inactivation <em>in vitro</em>. We hypothesized that CHF 5633 is also resistant to inactivation by serum albumin <em>in vivo</em>.</p> <h3>Methodology/Principal Findings</h3><p>Nineteen preterm lambs of 127 days gestational age (term = 150 days) received CHF 5633 or Poractant alfa and were ventilated for 48 hours. Ninety minutes after birth, the animals received albumin with CHF 5633 or Poractant alfa. Animals received additional surfactant if P_aO₂ dropped below 100 mmHg. A pressure volume curve was done post mortem and markers of pulmonary inflammation, surfactant content and biophysiology, and lung histology were assessed. CHF 5633 treatment resulted in improved arterial pH, oxygenation and ventilation efficiency index. The survival rate was significantly higher after CHF 5633 treatment (5/7) than after Poractant alfa (1/8) after 48 hours of ventilation. Biophysical examination of the surfactant recovered from bronchoalveolar lavages revealed that films formed by CHF 5633-treated animals reached low surface tensions in a wider range of compression rates than films from Poractant alfa-treated animals.</p> <h3>Conclusions</h3><p>For the first time a synthetic surfactant containing both surfactant protein B and C analogs showed significant benefit over animal derived surfactant in an <em>in vivo</em> model of surfactant inactivation in premature lambs.</p> </div

Langmuir Blodgett analysis of bronchoalveolar lavages.

<p>(A) Surface pressure versus % Trough area compression isotherms of Curosurf^® and the sequential broncho-alveolar lavages (BALs). The surfactant films were spread on pure water subphase at 37°C ± 1°C and compressed at a rate of 40 cm²/min. Each curve gives the data from a single experiment representative of three replicates. (B) Volume of Curosurf and BALs spread at the air-water interface to reach a surface pressure of ≈ 15 mN/m. C) Film Compressibility (Cm) of Curosurf and BALs as a function of surface pressure, <i>π</i>. Cm was determined from compression isotherm using the equation: Cm = -1/A*(dA/d<i>π</i>).</p

Respiratory parameters of surfactant-depleted rabbits.

<p>(A) Mean respiratory rate values of the animals managed just with nasal CPAP for 180 min (solid squares) and of the animals treated with 200 mg/kg of surfactant following the InsSurE approach and managed further for 180 min in nasal CPAP (empty circles). (B) Mean pressure values recorded after insufflating 30 ml of air <i>post mortem</i> (V30) in the lungs of the animals managed just with nasal CPAP (control) and of the animals treated with 200 mg/kg of surfactant following the InSurE approach and managed further for 180 min in nasal CPAP are shown.). N = 6 and N = 9 in the Control and InSurE groups, respectively. * P vs. Control < 0.05.</p

Surfactant administration.

<p>Bw – bodyweight. Data expressed as mean±SEM. * p<0.05, unpaired t-test.</p

Gas exchange of surfactant-depleted rabbits.

<p>(A) Mean PaO₂ and (B) PaCO₂ values in animals managed just with nasal CPAP for 180 min (solid squares) and in animals treated with 200 mg/kg of surfactant following the InsSurE approach and managed further for 180 min in nasal CPAP (empty circles). There were no differences between the experimental groups before the induction of respiratory distress (baseline, B in the X-axis). The time point 15ST refers to the 15 min stabilization period established to confirm the respiratory distress induced by repeated broncho-alveolar lavages (BALs). N = 6 and N = 9 in the Control and InSurE groups, respectively. * P vs. Control < 0.05.</p

Ventilation.

<p><b>A</b> The ventilation efficiency index (VEI) of lambs treated with CHF 5633 was elevated at all time points compared to Poractant alfa treated lambs, but only reached significance at 90 minutes after birth, just before inactivation (t = 0). <b>B</b> The partial oxygen pressure in arterial blood (P_aO₂) of lambs treated with CHF 5633 was elevated at most time points compared to Poractant alfa treated lambs, but only reached significance at 18 hours after surfactant inactivation. <b>C</b> The arterial blood pH of lambs treated with synthetic surfactant was elevated at all time points compared to Poractant alfa treated lambs, but only reached significance at 24 hours after surfactant inactivation. Grey spheres = CHF 5633; Black cubes = Poractant alfa. Data expressed as mean±SEM. *p<0.05, two-way repeated-measures analysis of variance.</p

Scheme of the experimental set up.

<p>Scheme of the steps followed during the experimental procedure. Animals were intubated at baseline (Basal) and underwent several broncho-alveolar lavages (BAL) in order to induce surfactant depletion. A 15 min stabilization period was established to confirm the respiratory distress (15ST). In this period, animals were managed on mechanical ventilation (MV). After the respiratory distress was confirmed, animals were randomized to one of the experimental groups. Rabbits in the nCPAP group were maintained on nCPAP for 180 min. Rabbits in the InSurE group received 200 mg/kg of Curosurf^® and were then managed on nCPAP for 180 min. At the end of the experiment all rabbits were re-intubated to determine the pulmonary status. Post mortem a pressure/volume curve (P/V) was performed.</p

Study design.

<p>Preterm lambs received a 200 mg/kg bodyweight intra-tracheal dose of either Poractant alfa or CHF 5633. Ninety minutes after birth a mixture consisting of 9.4 mg/kg human serum albumin and either 100 mg/kg Poractant alfa or 100 mg/kg CHF 5633 was given intra-tracheally to simulate surfactant inactivation. If P_aO₂ dropped below 100 mmHg, the lambs received an additional dose of either 200 mg/kg Poractant alfa or 200 mg/kg CHF 5633 every two hours until the P_aO₂ increased above the 100 mmHg threshold. 48 hours after birth the lambs were euthanized.</p

Parameters defining interfacial performance.

<p>Comparison of parameters defining the interfacial performance in the CBS of films formed by material obtained from the bronchoalveolar lavage of CHF 5633 treated and Poractant alfa treated animals. Data plotted are the minimal surface tension reached after 1 min of initial or post-expansion adsorption (upper panel), and the minimal surface tension, the percent of area reduction required to reach minimal tension and the maximal tension during quasi-static (middle panels) or dynamic (lower panels) compression-expansion cycling. Data expressed as mean±SEM. *p<0.05, two-way repeated-measures analysis of variance.</p