Enhanced oral bioavailability of an etoposide multiple nanoemulsion incorporating a deoxycholic acid derivative–lipid complex

Figures & data

Figure 1. (A) Schematic illustration of an etoposide/low-molecular-weight methylcellulose (ETP/LMC)-loaded water-in-oil-in-water (w/o/w) nanoemulsion incorporating an ionic complex of N^α-deoxycholyl-l-lysyl-methylester and 1,2-didecanoyl-sn-glycero-3-phosphate (sodium salt) (DCK–PA) (ELNE#7). (B) Transmission electron micrographic image of ELNE#7. Scale bar, 100 nm.

Table 1. Effective and apparent permeabilities of ETP, ETP/LMC, ETP emulsion, ENE, and ELNE formulations.

Formulation	Effective permeability (Pe, ×10^–6 cm/s)	Apparent permeability (Papp, ×10^–6 cm/s)
ETP dispersed in water	0.439 ± 0.079	1.86 ± 0.736
ETP/LMC solution	1.85 ± 0.920	4.14 ± 0.619
ETP dispersed in 0.3% NaCMC	1.61 ± 0.180	3.73 ± 0.265
ETP/LMC in 0.3% NaCMC	2.68 ± 0.252	4.32 ± 0.742
ETP in 5% DMSO	0.387 ± 0.091	2.92 ± 0.271
ENE	5.50 ± 1.41	4.82 ± 0.586
ETP emulsion	2.79 ± 0.487	4.25 ± 0.926
ELNE#1	9.10 ± 0.407	6.42 ± 1.58
ELNE#2	9.50 ± 0.393	6.63 ± 1.76
ELNE#3	9.39 ± 1.220	6.76 ± 0.396
ELNE#4	5.90 ± 0.120	6.26 ± 1.03
ELNE#5	9.35 ± 0.482	6.79 ± 0.408
ELNE#6	10.1 ± 0.303	7.93 ± 1.17
ELNE#7	11.4 ± 0.267	9.60 ± 1.10

Each value represents the mean ± standard deviation (n= 6).

Figure 2. Confocal laser scanning microscopic images of the cellular uptake of different coumarin-6-co-loaded etoposide (ETP) vehicles. (A) Cellular uptake of coumarin-6-co-loaded ETP/LMC solution, coumarin-6-co-loaded ETP/LMC in 0.3% NaCMC, coumarin-6-loaded ENE and ELNE#1, and coumarin-6-loaded ELNE incorporating an ionic complex of N^α-deoxycholyl-l-lysyl-methylester and 1,2-didecanoyl-sn-glycero-3-phosphate (sodium salt) (ELNE#7) by Caco-2 cells. Cellular uptake of coumarin-6-co-loaded ETP/LMC solution, coumarin-6-co-loaded ETP/LMC in 0.3% NaCMC, coumarin-6-loaded ENE and ELNE#1, and coumarin-6-loaded ELNE incorporating an ionic complex of N^α-deoxycholyl-l-lysyl-methylester and 1,2-didecanoyl-sn-glycero-3-phosphate (sodium salt) (ELNE#6 and #7) by (B) MDCK cells or (C) ASBT-transfected MDCK cells. Scale bar, 20 µm.

Figure 3. Relative apparent permeability (P_app) values of ELNE#7 without inhibitors (control) and in the presence of various biochemical inhibitors of specific pathways across Caco-2/HT-29-MTX-E12 monolayers. (A) P_app of ELNE#7 across Caco2/HT-29-MTX-E12 monolayers after incubation with various transport inhibitors. (B) P_app of ELNE#7 in the presence of actinomycin D, clofazimine, or both (actinomycin D + clofazimine). Each value represents the mean ± standard deviation (n= 4). **p<.01, ***p<.001 compared with the P_app of ELNE#7 in the absence of all inhibitors (untreated control). (C) P_app ratio of ETP/LMC in water and ELNE#7 in both apical to basolateral (AB) and basolateral to apical (BA) directions in the absence of all inhibitors (net efflux ratio; P_{app, BA, w/o inhibitors}/P_{app, AB, w/o inhibitors}). (D) Relative P_app ratios of ELNE#7 in the presence of cyclosporine A (Cys A) (net P_app ratio; P_{app, AB, with Cys A}/P_{app, AB, w/o inhibitors}). Each value represents the mean ± standard deviation (n= 4). ***p<.001 compared with the P_app of ETP/LMC solution. (E) P_app of ELNE#7 with or without EGTA in the presence or absence of all inhibitors except Cys A. Each value represents the mean ± standard deviation (n= 4). *p<.05, ***p<.001 compared with the P_app of ELNE#7 without EGTA in the absence of all inhibitors (untreated control); ^#p<.05, ^###p<.001 compared with the P_app of ELNE#7 without EGTA in the presence of all inhibitors except Cys A; ^$$$p<.001 compared with the P_app of ELNE#7 with EGTA in the absence of all inhibitors except Cys A.

Figure 4. In vitro cytotoxic effects of ETP dispersed in water, ETP/LMC solution, ETP-IV, ETP emulsion, and ELNE#7 on (A) LLC and (B) A549 cells after incubation for 24 hours. Each value represents the mean ± standard deviation (n= 4).

Figure 5. Mean plasma concentration–time profiles of etoposide (ETP) in rats after (A) a single intravenous (IV) administration of 20 mg/kg ETP (ETP-IV) and (B) oral administration of an aqueous suspension of 20 mg/kg ETP (ETP dispersed in water); 5% DMSO solution of 20 mg/kg ETP (ETP in 5% DMSO); aqueous solution of 20 mg/kg ETP/LMC (ETP/LMC solution); ETP-incorporating emulsion (ETP emulsion); and ETP/LMC-loaded nanoemulsion (ELNE) incorporating an ionic complex of N^α-deoxycholyl-l-lysyl-methylester and 1,2-didecanoyl-sn-glycero-3-phosphate (sodium salt) (DCK–PA) (ELNE#7) as 20 mg/kg ETP. Each value represents the mean ± standard deviation (n= 4 for each group).