Masters Thesis

Efficacy and Pharmacokinetics of Aerosolized and Intravenous Ambisome® Therapy For Pulmonary Aspergillosis

Invasive Pulmonary Aspergillosis (IPA) causes 50-90% mortality with present forms of oral and intravenous (IV) antifungal drugs, emphasizing the need for improved treatments. To explore this, we built an aerosol system to deliver inhalable, antifungal therapy to mice. We investigated the in vivo efficacy of inhalable liposomal Amphotericin B, AmBisomeⓇ (AmBi), given as monotherapy or in combination with IV AmBi for the treatment of IPA in Aspergillus fumigatus infected mice. Swiss-Webster female mice were immunosuppressed with the steroid triamcinolone acetonide or cyclophosphamide and challenged d0 with 9 x 106 A. fumigatus (ATCC#13073). For all in vivo studies, mice were placed in a 12-compartment chamber (one mouse/compartment) and exposed to aerosol (aero) AmBi. In study 1, we compared the efficacy of IV vs. aero AmBi. Mice were treated 1X daily with either IV AmBi (7.5 mg/kg), 20 mins of aero AmBi (1.33 µg/mL in nebulizer) or IV phosphate buffered saline (PBS) for a total of 3 treatments. In study 2, we varied the aero AmBi monotherapy dosing regimen, starting 4h post-challenge and given 5X, once daily for 10 or 20 minutes/treatment or given every other day and compared it to IV AmBi monotherapy given three times, once daily. In study 3, we investigated treatment regimens that would be more clinically applicable since IPA is often diagnosed late. We did this by delaying aero AmBi monotherapy to 8h or combining aero AmBi at 8h with IV AmBi therapy starting at 12h to treat an established, systemic infection. In study 4, we used the same treatment schedule as in study 3 to test the efficacy of aero AmBi monotherapy vs. aero AmBi combined with IV AmBi in a cyclophosphamide immunosuppressed IPA mouse model. Finally, we tested the efficacy of aero AmBi monotherapy or its combination with IV AmBi to treat IPA caused by different strains of A. fumigatus (strains V079 and V080). In study 1 aero AmBi monotherapy was more effective than IV AmBi monotherapy in reducing lung CFU/g, but less effective at reducing liver, kidney or spleen CFU/g, indicating that aero AmBi monotherapy may need further optimization to reduce the fungal burden in the internal organs. Even though reduction of CFU/g in these internal organs was limited, mouse survival was very good with aero AmBi monotherapy (75%) compared to IV AmBi monotherapy (43%) and PBS (33%) suggesting that aero AmBi was preventing fungal dissemination from the lungs to the internal organs. In study 2, longer (20 minutes vs 10 minutes/treatment) aero AmBi monotherapy had exceptional survival (100%, p ≤ 0.05 vs all other groups). Intermittent every other day aero AmBi and daily IV AmBi monotherapy were less effective (43% survival). In study 3, when treatment started later at 8h. The combination of 3 IV AmBi treatments with 5 aero AmBi treatments was as effective as earlier aero AmBi monotherapy beginning at 4h (86% survival for both treatments). The delayed combination treatment was also significantly better in reducing organ fungal burden. In study 4, in the cyclophosphamide immunosuppressed IPA model (A. fumigatus ATCC 13073), the combination treatment yielded 100% survival. In study 5, combination treatment in the steroid suppressed IPA model, yielded 100% survival following infection with A. fumigatus strain V079 and 86% with V080, with significant reduction in organ fungal burden (p ≤ 0.05 vs. IV AmBi alone). Compared to IV AmBi monotherapy, aero AmBi monotherapy given early or combined with IV AmBi for delayed therapy significantly increased survival, reduced lung fungal burden and decreased dissemination to the internal organs. These results suggest that aero AmBi may be a promising treatment option to reduce the high mortality associated with IPA.

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