P010 Evaluation of antifungal efficacy of two novel cyclic lipopeptides of the class Bacillomycin from Bacillus subtilis RLID 12.1 in a murine model of invasive candidiasis
Authors: Kashyap Nisha , Paul Raees A. , Rudramurthy Shivaprakash , Roy Utpal , Kaur Harsimran , Ghosh Anup , Chakrabarti Arunaloke 
Published: September 2022

Abstract

Poster session 1, September 21, 2022, 12:30 PM - 1:30 PM

Objective
To evaluate the in vivo efficacy of HPLC–purified antifungal lipopeptides (AF4 and AF5) in a murine model of disseminated candidiasis.

Methods
C. albicans AMR16294 isolate was used for all the in vivo experiments. A total of 6-week-old pathogen-free, female BALB/c mice, weighing 20-25 g were used for all animal experiments. For Kaplan-Mier analysis, mice were rendered neutropenic by a loading dose of 200 mg/kg cyclophosphamide three days prior (D-3) to infection and 150 mg/kg (D + 1) maintenance dose on day 1 post-infection (D + 1). A total of 60 mice were randomized into 8 different groups with 5 or 6 animals in each group. Animals were infected with 100μL of ∼ 1 × 10⁵ blastospores (corresponding to LD90) via the lateral tail vein. AF4 and AF5 were formulated in sterile PBS and administered intraperitoneally at doses of 5 mg/kg and 10 mg/kg body weight and compared with a clinically-relevant human equivalent dose of caspofungin. AF4, AF5, caspofungin, or vehicle were administered at 1 h and 24 h post-infection. The survival of the mice was monitored for 14 days post-infection. For organ fungal-burden assessment, mice from each group were euthanized by CO2 inhalation, and the organs were aseptically removed, homogenized, and cultured on SDA.

Results
Both the doses of AF4 significantly reduced the mortality of mice compared to vehicle-treated mice. The survival over 2 weeks in 5 mg/kg, 10 mg/kg, and caspofungin arms were similar and no death was reported in the three groups (P &lt;.01). In contrast, the mortality in-vehicle- administered group was 80% with a median survival of 8 days. A similar survival benefit was observed in AF5-treated mice. While the median survival in the vehicle-treated arm was 5 days, the 2-week survival in 5 mg/kg and 10 mg/kg arms was 80%-100%, comparable to that in the caspofungin arm (P &lt;.01) (Fig. 1).
The median CFU/g kidney tissue in 5 mg/kg arm of AF4 was 1.3 × 10⁴ equivalent to a 4-log reduction compared to the vehicle arm (3.8 × 10⁸ CFU/g kidney, P &lt;.0001). The in vivo efficacy was higher at a higher dose with the kidney homogenates of 10 mg/kg arm yielding sterile cultures comparable to that of CAS arm (Fig. 2). Similar organ fungal-burden reduction was noted in heart and splenic tissues with a median cfu/g tissue of 1.3 × 10⁴ in 10 mg/kg, while CAS arms yielded sterile cultures.
In AF5 treated groups, the median cfu/g kidney tissue in 5 mg/kg arm was 1.3 × 10⁴, however, the heart and splenic tissue homogenates yielded less fungal burden with median (q2) cfu/g as 0, while q3 of 6.7 × 10³and 1.3 × 10⁴, respectively (Fig. 2).

Conclusion
Both the antifungal compounds demonstrated a remarkable in vivo efficacy against C. albicans with a significant improvement in survival and a reduction in the organ-fungal burden.