Effect of lactoferrin- and transferrin-conjugated polymersomes in brain targeting: in vitro and in vivo evaluations

Abstract

Aim: To evaluate the effect of lactoferrin (Lf) and transferrin (Tf) in brain targeting.

Methods: Polymersomes (PSs), employed as vectors, were conjugated with Lf or Tf and were characterized by morphology, particle size, zeta potential, and surface densities of the Lf or Tf molecules. In vitro uptake of Lf-PS and Tf-PS by bEnd.3 cells was investigated using coumarin-6 as a fluorescent probe. In vivo tissue distribution and pharmacokinetics of (125)I-Lf-PS and (125)I-Tf-PS were also examined.

Results: The mean particle size of PS, Lf-PS, and Tf-PS was around 150 nm and the zeta potential of the PSs was about -20 mV. Less than 0.12% of the coumarin was released from coumarin-6-loaded PS in 84 h indicating that coumarin-6 was an accurate probe for the PSs' behavior in vitro. It was shown that the uptake of Lf-PS and Tf-PS by bEnd.3 cells was time-, temperature-, and concentration-dependent. Both Lf and Tf could increase the cell uptake of PSs at 37 degrees C, but the uptake of Tf-PS was significantly greater than that of Lf-PS. In vivo tissue distribution and pharmacokinetics in mice revealed higher brain uptake and distribution of Tf-PS than Lf-PS, which was in accordance with in vitro uptake results. The drug targeting index (DTI) of Tf-PS with regard to Lf-PS was 1.51.

Conclusion: Using a PS as the delivery vector and bEnd.3 cells as the model of the blood-brain barrier (BBB), Tf was more effective than Lf in brain targeting.

Figure 1

Transmission electron micrograph of PS negatively stained with uranyl acetate solution. The bar is 50 nm.

Figure 2

In vitro release of 6-coumarin from PS in 0.01 mol/L PBS (pH 7.4 and 4.0).

Figure 3

BCECs uptake (A) 10–600 mg/L PS, Lf-PS, and Tf-PS at 37 ºC and 4 ºC incubation for 1 h, respectively; (B) 100 mg/L PS, Lf-PS, and Tf-PS at 37 ºC incubation for different time. n=3. Mean±SD. ^bP<0.05, ^cP<0.01 vs PS. ^eP<0.05 vs Lf-PS.

Figure 4

Cell uptake of polymersomes by bEND.3 cells. 300 mg/L PS at 37 ºC for 30 min (A), 60 min (D), 120 min (G); 300 mg/L Lf-PS for 30 min (B), 60 min (E), 120 min (H) and 300 mg/L of Tf-PS for 30 min (C), 60 min (F), 120 min (I), respectively, bar 50 μm.

Figure 5

(A) The average PS's concentration-time curve in whole blood followed by iv injection of Lf-PS or Tf-PS in mice. n=6. Mean±SD. (B) The Lf-PS and Tf-PS concentration in brain at different time; (C) The Lf-PS and Tf-PS AUC_0–t in different tissues. ^bP<0.05, ^cP<0.01 vs Lf-PS.