The Synergistic Effect of Tacrolimus (FK506) or Everolimus and Azoles Against Scedosporium and Lomentospora Species In Vivo and In Vitro

Abstract

Scedosporium and Lomentospora infections in humans are generally chronic and stubborn. The use of azoles alone cannot usually inhibit the growth of these fungi. To further explore the combined effect of multiple drugs and potential mechanisms of action, we tested the antifungal effects of tacrolimus (FK506) and everolimus in combination with azoles in vitro and in vivo on 15 clinical strains of Scedosporium/Lomentospora species and detected the level of Rhodamine 6G, ROS activity, and apoptosis. The in vitro results showed that the combinations of tacrolimus with itraconazole, voriconazole, and posaconazole showed synergistic effects on 9 strains (60%), 10 strains (73%), and 7 strains (47%), respectively, and the combinations of everolimus with itraconazole, voriconazole, and posaconazole showed synergistic effects on 8 strains (53%), 8 strains (53%), and 7 strains (47%), respectively. The synergistic effects might correspond to the elevated ROS activity (the tacrolimus + itraconazole group compared to the itraconazole group, (P < 0.05)), early apoptosis (itraconazole (P < 0.05) and voriconazole (P < 0.05) combined with everolimus), and late apoptosis (the tacrolimus + itraconazole group compared to the itraconazole group, (P < 0.01); the tacrolimus + posaconazole group compared to the posaconazole group, (P < 0.05)), but not inhibition of efflux pump activity. Our in vitro results suggested that a combination of tacrolimus or everolimus and azoles have a synergistic effect against Scedosporium/Lomentospora. The synergistic mechanisms of action might be triggering excessive ROS activity and apoptosis. In vivo, the survival rate of G. mellonella (sixth instar larvae) was significantly improved by tacrolimus alone, everolimus alone, azoles alone, and tacrolimus and everolimus combined with azoles separately (P < 0.05 for the tacrolimus group; P < 0.01 for the everolimus group and the itraconazole group; P = 0.0001 for the tacrolimus and posaconazole group; P < 0.0001 for other groups except the everolimus and itraconazole group, everolimus and posaconazole group, and tacrolimus and itraconazole group). From the results, we infer that the combination of tacrolimus or everolimus with azoles has obvious synergistic effect on Scedosporium/Lomentospora, and might enhance the level of apoptosis and necrosis. However, the synergistic effects were not related to the efflux pump. In conclusion, from our in vitro and in vivo study, tacrolimus and everolimus combined with azoles may have a synergistic effect in the treatment against Scedosporium/Lomentospora, improving the drug activity of azoles and promoting a better prognosis for patients.

Case report: Disseminated Scedosporium apiospermum infection with invasive right atrial mass in a heart transplant patient

“…Our patient was on tacrolimus, which was previously shown to have synergistic effect with azoles in vitro and in a Galleria mellonella larvae model ( 14 ). However, this synergistic effect is obtained at high concentrations of tacrolimus (i.e., beyond the therapeutic range) and is counterbalanced by the strong immunosuppressive effect of this calcineurin inhibitor which may favor invasive fungal infections.…”

Analysis of the synergistic antifungal activity of everolimus and antifungal drugs against dematiaceous fungi

“…In precious several studies, the EVL-induced cell survival rate in HaCat, Caki-1, and HepG2 cells were greater than 70% at 30 mM (≈28.75 mg/mL), especially up to 100 mM in HepG2 cells (Yamamoto et al, 2013;Navarro-Villarań et al, 2016). Recent studies have also demonstrated the activity of EVL against Aspergillus, Scedosporium, and Lomentospora species (Wang et al, 2022). However, there have not been any published studies assessing the effects of EVL against dematiaceous fungi.…”

Scedosporium species and Lomentospora prolificans fungaemia is uniformly fatal in patients with haematological malignancy

“…In an Australian laboratory‐based study of 30 L. prolificans isolates, the MIC 50 values for voriconazole, posaconazole and isavuconazole were 8, 16 and 16 mg/L respectively 11 . Synergistic effects have been noted in vitro when tacrolimus, minocycline or terbinafine are added to triazole antifungals 12–14 . Although many of these studies only included a small number of patients, olorofim yielded an MIC 90 value of 0.25 mg/L against 30 L. prolificans isolates 11 .…”

“…11 Synergistic effects have been noted in vitro when tacrolimus, minocycline or terbinafine are added to triazole antifungals. [12][13][14] Although many of these studies only included a small number of patients, olorofim yielded an MIC 90 value of 0.25 mg/L against 30 L. prolificans isolates. 11 Fosmanogepix (APX001) also has demonstrated efficacy in vitro and in a mouse model.…”