Preclinical Evaluation of Recombinant Microbial Glycoside Hydrolases in the Prevention of Experimental Invasive Aspergillosis.

Aspergillus fumigatus is a ubiquitous mold that can cause invasive pulmonary infections in immunocompromised patients. Within the lung, A. fumigatus forms biofilms that can enhance resistance to antifungals and immune defenses. Aspergillus biofilm formation requires the production of a cationic matrix exopolysaccharide, galactosaminogalactan (GAG). In this study, recombinant glycoside hydrolases (GH)s that degrade GAG were evaluated as antifungal agents in a mouse model of invasive aspergillosis. Intratracheal GH administration was well tolerated by mice. Pharmacokinetic analysis revealed that although GHs have short half-lives, GH prophylaxis resulted in reduced fungal burden in leukopenic mice and improved survival in neutropenic mice, possibly through augmenting pulmonary neutrophil recruitment. Combining GH prophylaxis with posaconazole treatment resulted in a greater reduction in fungal burden than either agent alone. This study lays the foundation for further exploration of GH therapy in invasive fungal infections. IMPORTANCE The biofilm-forming mold Aspergillus fumigatus is a common causative agent of invasive fungal airway disease in patients with a compromised immune system or chronic airway disease. Treatment of A. fumigatus infection is limited by the few available antifungals to which fungal resistance is becoming increasingly common. The high mortality rate of A. fumigatus-related infection reflects a need for the development of novel therapeutic strategies. The fungal biofilm matrix is in part composed of the adhesive exopolysaccharide galactosaminogalactan, against which antifungals are less effective. Previously, we demonstrated antibiofilm activity with recombinant forms of the glycoside hydrolase enzymes that are involved in galactosaminogalactan biosynthesis. In this study, prophylaxis with glycoside hydrolases alone or in combination with the antifungal posaconazole in a mouse model of experimental aspergillosis improved outcomes. This study offers insight into the therapeutic potential of combining biofilm disruptive agents to leverage the activity of currently available antifungals.

Aerosolized Lipid Amphotericin B for Complementary Therapy and/or Secondary Prophylaxis in Patients with Invasive Pulmonary Aspergillosis: A Single-Center Experience.

BACKGROUND: Experience with aerosolized lipid amphotericin B (aeLAB) as therapy or secondary prophylaxis in patients with invasive pulmonary aspergillosis (IPA) is anecdotal. METHODS: We performed a single-center retrospective cohort study to evaluate the efficacy of systemic antifungal therapy with and without aeLAB in patients with proven or probable IPA. Complete or partial response at 3 months was the primary end-point. Clinical response and mortality at 12 months, occurrence of adverse drug reactions and respiratory fungal colonization were secondary end-point. RESULTS: Eleven patients (39%) received aeLAB in addition to systemic antifungal therapy (group A), and 22 (61%) received systemic antifungal therapy only (group B). The use of aeLAB was not standardized. Amphotericin B lipid complex was used in all patients but one, who received liposomal amphotericin B. Five patients received aeLAB as antifungal complementary therapy and 6 received it as secondary prophylaxis. Except for the requirement of inhaled corticosteroids and home oxygen therapy, more frequent in group A, both groups were similar in baseline conditions. A better (nonsignificant) clinical outcome was observed at 3 months in patients receiving aeLAB. Only uncontrolled baseline condition was associated with one-year mortality in univariate analysis (p = 0.002). A multivariate Cox regression analysis suggests that aeLAB, corrected for uncontrolled underlying disease, reduces mortality at 12 months (HR 0.258; 95% CI 0.072-0.922; p = 0.037). CONCLUSION: Although no significant difference was observed in the main variable (3-month clinical response) and in spite of methodological limitations of the study, the possible survival benefit of aeLAB, adjusted for the control of the underlying disease, could justify the performance of well-controlled studies with a greater number of patients.

A strategy for designing voriconazole dosage regimens to prevent invasive pulmonary aspergillosis based on a cellular pharmacokinetics/pharmacodynamics model.

BACKGROUND: Invasive pulmonary aspergillosis (IPA) is a life-threatening disease in immunosuppressed patients. Voriconazole is commonly used to prevent and treat IPA in the clinic, but the optimal prophylactic antifungal regimen is unknown. The objective of this study was to clarify the mechanism underlying how voriconazole prevents IPA based on a target cellular pharmacokinetics/pharmacodynamics model, with the aim of identifying a way to design an optimal prophylactic antifungal regimen. METHODS: A nystatin assay was used to establish a target-cells model for A. fumigatus infection. An inhibitory effect sigmoid Emax model was developed to explore the cellular PK/PD breakpoint, and Monte Carlo simulation was used to design the prophylactic antifungal regimen. RESULTS: The intracellular activity of voriconazole in the target cells varied with its concentration, with the minimum inhibitory concentration (MIC) being an important determinant. For A. fumigatus strains AF293 and AF26, voriconazole decreased the intracellular inoculum by 0.79 and 0.84 lg cfu, respectively. The inhibitory effect sigmoid Emax model showed that 84.01% of the intracellular inoculum was suppressed by voriconazole within 24 h, and that a PK/PD value of 35.53 for the extracellular voriconazole concentration divided by MIC was associated with a 50% suppression of intracellular A. fumigatus. The Monte Carlo simulation results showed that the oral administration of at least 200 mg of voriconazole twice daily was yielded estimated the cumulative fraction of response value of 91.48%. Concentration of voriconazole in the pulmonary epithelial lining fluid and the plasma of > 17.77 and > 1.55 mg/L, respectively, would ensure the PK/PD > 35.53 for voriconazole against most isolates of A. fumigatus and may will be benefit to prevent IPA in clinical applications. CONCLUSIONS: This study used a target cellular pharmacokinetics/pharmacodynamics model to reveal a potential mechanism underlying how voriconazole prevents IPA and has provided a method for designing voriconazole prophylactic antifungal regimen in immunosuppressed patients.

Chitinase Induction Prior to Caspofungin Treatment of Experimental Invasive Aspergillosis in Neutropenic Rats Does Not Enhance Survival.

Host chitinases, chitotriosidase and acidic mammalian chitinase (AMCase), improved the antifungal activity of caspofungin (CAS) against Aspergillus fumigatus in vitro These chitinases are not constitutively expressed in the lung. Here, we investigated whether chitosan derivatives were able to induce chitinase activity in the lungs of neutropenic rats and, if so, whether these chitinases were able to prolong survival of rats with invasive pulmonary aspergillosis (IPA) or of rats with IPA and treated with CAS. An oligosaccharide-lactate chitosan (OLC) derivative was instilled in the left lung of neutropenic rats to induce chitotriosidase and AMCase activities. Rats instilled with OLC or with phosphate-buffered saline (PBS) were subsequently infected with A. fumigatus and then treated with suboptimal doses of CAS. Survival, histopathology, and galactomannan indexes were determined. Instillation of OLC resulted in chitotriosidase and AMCase activities. However, instillation of OLC did not prolong rat survival when rats were subsequently challenged with A. fumigatus In 5 of 7 rats instilled with OLC, the fungal foci in the lungs were smaller than those in rats instilled with PBS. Instillation of OLC did not significantly enhance the survival of neutropenic rats challenged with A. fumigatus and treated with a suboptimal dosage of CAS. Chitotriosidase and AMCase activities can be induced with OLC, but the presence of active chitinases in the lung did not prevent the development of IPA or significantly enhance the therapeutic outcome of CAS treatment.

Posaconazole prophylaxis in experimental azole-resistant invasive pulmonary aspergillosis.

We investigated the efficacy of posaconazole prophylaxis in preventing invasive aspergillosis due to azole-resistant Aspergillus fumigatus isolates. Using a neutropenic murine model of pulmonary infection, posaconazole prophylaxis was evaluated using three isogenic clinical isolates, with posaconazole MICs of 0.063 mg/liter (wild type), 0.5 mg/liter (F219I mutation), and 16 mg/liter. A fourth isolate harboring TR34/L98H (MIC of 0.5 mg/liter) was also tested. Posaconazole prophylaxis was effective in A. fumigatus with posaconazole MICs of ≤0.5 mg/liter, where 100% survival was reached. However, breakthrough infection was observed in mice infected with the isolate for which the posaconazole MIC was >16 mg/liter.