In Vitro Anti-Aspergillus Activity and Antifungal Interactions of Monoclonal Antibodies 1D2 and 4E4

Objective: To determine the in vitro anti-Aspergillus activity of monoclonal antibodies (mAb) 1D2 and 4E4 alone and in combination with voriconazole, posaconazole, amphotericin B and caspofungin and to assess whether selected antimicrobial agents interfere with 1D2/4E4 binding to Aspergillus antigens. Methods: The in vitro antifungal activity of 1D2, 4E4 and all antifungal drugs was determined in a microplate-based assay. The anti-Aspergillus activity of mAb 1D2 and 4E4 in vitro alone and in combination with voriconazole, posaconazole, amphotericin B and caspofungin was determined using a checkerboard assay. Finally, a competitive ELISA was used to assess whether selected antifungal and antibacterial agents interfered with the binding of 1D2 and 4E4 to immobilised Aspergillus antigens. Results: The in vitro interactions of 1D2 with voriconazole, posaconazole, amphotericin B and caspofungin against four different Aspergillus species were all additive, with a fractional inhibitory concentration index (FICI) between 0.5 and 1. Similarly, the in vitro interactions of 4E4 with these same four antifungals against Aspergillus fumigatus (A. fumigatus) were all additive. An additive interaction was also detected for 4E4 and caspofungin against all four Aspergillus species tested. However, no interaction was found for 4E4 combined with voriconazole, posaconazole and amphotericin B against Aspergillus flavus (A. flavus), Aspergillus niger (A. niger) and Aspergillus terreus (A. terreus), with an FICI between 1 and 4. Moreover, none of the tested antifungal or antibacterial agents significantly interfered with 1D2/4E4 binding to Aspergillus antigens in the competitive ELISA. Conclusions: In this study, 1D2 and 4E4 showed in vitro growth-inhibitory activity against the tested Aspergillus isolates. Specifically, 1D2 showed additive interactions with the tested antifungal agents across all tested species, whereas 4E4 showed a more species- and drug-dependent interaction profile. The tested antimicrobial agents did not measurably interfere with 1D2/4E4 binding under the competitive ELISA conditions examined. Overall, these findings provide preliminary in vitro evidence supporting the further evaluation of 1D2 and 4E4.