Brain capillary endothelial cells (BCECs) express transferrin receptors. The uptake of a potential drug vector (OX26, or anti-transferrin receptor antibody IgG2a) conjugated to polyethyleneglycol-coated liposomes by BCECs was studied using in situ perfusion in 18-day-old rats in which the uptake of OX26 is almost twice as high as in the adult rat. Using radio-labeling, the uptake of OX26 by BCECs after 15-minute perfusion was approximately 16 times higher than that of nonimmune IgG2a (Ni-IgG2a). OX26 and OX26-conjugated liposomes selectively distributed to BCECs, leaving choroid plexus epithelium, neurons, and glia unlabeled. Ni-IgG2a and unconjugated liposomes did not reveal any labeling of BCECs. The labeling of BCECs by OX26 was profoundly higher than that of transferrin. Perfusion with albumin for 15 minutes did not reveal any labeling of neurons or glia, thus confirming the integrity of the blood-brain barrier. The failure to label neurons and glia shows that OX26 and OX26-conjugated liposomes did not pass through BCECs. The expression of transferrin receptors by endothelial cells selective to the brain qualifies OX26 as a candidate for blood-to-endothelium transport. A specifically designed formulation of liposomes may allow for their degradation within BCECs, leading to subsequent transport of liposomal cargo further into the brain.

Targeting the tumor vasculature and selectively modifying endothelial functions is an attractive anti-tumor strategy. We prepared polyethyleneglycol modified immunoliposomes (IL) directed against vascular cell adhesion molecule 1 (VCAM-1), a surface receptor over-expressed on tumor vessels, and investigated the liposomal targetability in vitro and in vivo. In vitro, anti-VCAM-1 liposomes displayed specific binding to activated endothelial cells under static conditions, as well as under simulated blood flow conditions. The in vivo targeting of IL was analysed in mice bearing human Colo 677 tumor xenografts 30 min and 24 h post i.v. injection. Whereas biodistribution studies using [3H]-labelled liposomes displayed only marginal higher tumor accumulation of VCAM-1 targeted versus unspecific ILs, fluorescence microscopy evaluation revealed that their localisations within tumors differed strongly. VCAM-1 targeted ILs accumulated in tumor vessels with increasing intensities from 30 min to 24 h, while control ILs accumulated in the tumor tissue by passive diffusion. ILs that accumulated in non-affected organs, mainly liver and spleen, primarily co-localised with macrophages. This is the first morphological evidence for selective in vivo targeting of tumor vessels using ILs. VCAM-directed ILs are candidate drug delivery systems for therapeutic anti-cancer approaches designed to alter endothelial function.

Rats were studied for [ 59 I]transferrin uptake in total brain, and fractions containing brain capillary endothelial cells (BCECs) or neurons and glia. 59 Fe was transported through Throughout life, brain capillary endothelial cells (BCECs) transport iron from blood plasma to the brain interior via transferrin receptor-mediated uptake at their luminal side. This iron uptake ensures an adequate iron supply to neuronal and glial cells, which require iron for mitochondrial respiration, neurotransmitter synthesis and myelin formation. The blood-to-brain iron transport is regulated via BCEC transferrin receptor expression, which yields higher iron uptake during development and conditions with inadequate iron supply (Taylor et al. 1991). Conversely, BCECs downregulate this transport when sufficient neuronal and glial iron prevails, thus preventing iron overload in the brain (Crowe and Morgan 1992). This is also reflected by the absence of significant cellular damage in the brain as opposed to virtually any other organ in the body during systemic iron overload . Low molecular weight iron complexes of non-transferrinbound iron (NTBI) are absent from blood plasma under physiological conditions ( Abbreviations used: Alb, albumin, AUC, area under the plasma radioactivity curve; b/b, homozygous Belgrade rat; BCEC, brain capillary endothelial cell; DAB, diaminobenzidine; DMT1, divalent metal transporter 1; IRE, iron-responsive element; KPBS, potassium phosphate-buffered saline; NTBI, non-transferrin bound iron; +/b, heterozygous Belgrade rat; P, postnatal; PBS, phosphate-buffered saline; PFA, paraformaldehyde; TCA, trichloroacetic acid; Trf, transferrin; V D , volume of distribution.

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