Intravenous delivery of a liposomal formulation of voriconazole improves drug pharmacokinetics, tissue distribution, and enhances antifungal activity

Figures & data

Figure 1. Voriconazole liposomal formulation characteristics. TEM micrograph of empty (blank) liposomes (A,B); VCZ liposomes (LVCZ) (C,D); VCZ liposomes (LVCZ) size distribution profiles by intensity obtained by DLS from three independent batches (E); freeze-dried formulation (F); redispersed freeze-dried LVCZ in a transparent infusion bag (G); and representative chromatogram from VCZ liposomal formulation at 2.0 mg/mL by HPLC with UV–VIS detector (H).

Table 1. Minimum inhibitory (MIC) and fungicide (MFC) concentrations (μg/mL) of voriconazole and liposomal voriconazole on Candida species sp. and Aspergillus sp.

	VFEND^®		LVCZ
Yeast isolated	MIC	MFC	MIC	MFC
Candida sp.
C. albicans 77 U	0.03	0.5	0.03	0.12
C. albicans 11 U	0.06	0.12	0.06	0.12
C. albicans ATCC 90028	0.03	>0.5	0.03	>0.5
C. parapsilosis ATCC 22019	0.03	0.03	0.03	0.03
Aspergillus sp.
A. fumigatus L	0.5	1	0.5	0.5
A. fumigatus V1F2	0.25	1	0.25	1
A. flavus V2	0.5	1	0.25	0.25
A. flavus V1F1	1	2	0.5	1

ATCC: American Type Culture Collection; the designations: (77U, 11U, L, V1F2, V2, and V1F1) were used to name clinical isolates originated patients samples.

Figure 2. HPLC-MS/MS and analytical conditions. Full scan mass spectrum of (A) voriconazole and (B) voriconazole N-oxide by electrospray ionization in positive mode of the transitions of m/z 350.2/127.0 and m/z 366.2/224.2, respectively. Chromatographic profiles obtained under following conditions: ACE C18 column (100 × 4.6 mm ×5 µm), at 20 °C; mobile phase consisted of formic acid at 0.025% + ammonium acetate 2 mM and acetonitrile (25:75 v/v), flow rate of 1.0 mL/min. Run time was 2.0 min, and the injection volume was 5.0 µL, with an elution time for VCZ and VNO of 1.43 and 1.19 min, respectively. Calibration curves, ranging 5–5000 and 20–5000 ng/mL for VCZ (C) and VNO (D), respectively.

Table 2. Pharmacokinetic and pharmacodynamic parameters after 10 mg/kg IV administration of two different voriconazole formulations to Balb/c mice.

			Mean ± SD
Parameter	Rule/equation	unit	LVCZ	VFEND^®
Tmax	observed	h	0.25 ± 0.00	0.25 ± 0.00
Cmax	observed	μg/mL	1.23 ± 0.28	0.61 ± 0.15
C0	extrapoled	μg/mL	1.16 ± 0.38	1.01 ± 0.39
AUC0–24	trapezoids	μg/mL*h	4.86 ± 1.01	1.96 ± 0.30
Cl	Dose/AUC	mL/h	52.75 ± 8.88	100.01 ± 20.14
Vd	IV dose/C0	mL	230.18 ± 53.61	314.18 ± 106.24
AUC0–24/MIC	ratio	–	53.51 ± 11.12	21.51 ± 2.88
Cmax/MIC	ratio	–	13.49 ± 31.3	6.73 ± 1.47
T > MIC	observed	h	∼12	∼8

T_max: time to reach C_max; C_max: maximum plasma concentration; C₀: estimated initial (zero-time) drug concentration in blood; AUC_0–24: blood levels of intravenously injected VCZ versus time (0–24 h); Cl: clearance; Vd: volume of distribution; IV: intravenous. Pharmacodynamic parameters calculed, considering estimated voriconazole unbound fraction: AUC_0–24/MIC: ratio between unbound fraction of voriconazole (33% of total AUC_0–24) and MIC; C_max/MIC: ratio between unbound fraction of voriconazole (33% of total C_max) and MIC of C. albicans; T > MIC: duration of time that the voriconazole blood concentration exceeds the MIC; MIC: minimum inhibitory concentration

Figure 3. Pharmacokinetic and biodistribution profile of voriconazole from two different formulations. (A) Blood levels of intravenously injected voriconazole versus time 0–24 h (AUC_0–24) delivered from liposomal formulation (circles) and VFEND^® (squares) at 10 mg/kg. (B) Concentrations versus time of the metabolite (voriconazole-N-oxide) measured within the hour following intravenous administration of the formulations. Data are expressed as mean ± SD (n = 12 animals per group). (C) Tissue distribution of voriconazole 4 h after I.V. administration of liposomal formulation (white bars) and voriconazole complexed in sulfobutyl ether-beta-cyclodextrin so + dium, commercially available as VFEND^® (black bars). Bars represent the standard deviation (n = 6). (*indicates p<.05).

Figure 4. Fungal growth of C. albicans from kidneys of imunossupressed and infected mice treated with placebo (A), VFEND^® (B) and LVCZ (C). (D) Effects of the VCZ treatments on the tissue burden of C. albicans in infected mice and controls (n = 5 per group). Infection was completely resolved in the liver using VFEND^® or LVCZ formulation. Representative micrographs of histological analysis of H&E stained kidney sections of (E) control group, healthy Balb/C mice, (F) infected mice, placebo group, (G) infected mice, treated with VFEND^® and (H) infected mice, treated with LVCZ. PAS stained kidney sections showing fungal invasion in (I) control group, healthy Balb/C mice, (J) infected mice, placebo group, (K) infected mice, treated with VFEND^® and (L) infected mice, treated with LVCZ. Inserts are higher magnification of fungal invasion areas in the kidney tissue, also indicated by black arrows.