Biomedical Engineering Reference
In-Depth Information
of cholesterol [40]. Finally, the HDL is taken up by the liver
via SR-BI, where the protein can be recycled or catabolized.
Cholesterol-loaded HDLs therefore reach the liver by the
circulating blood, where cholesterol is internalized by hepa-
tocytes. It has also been demonstrated that after systemic
administration of apo A-I, this exogenous apo A-I is accu-
mulated in the liver [41]. Thus, drug-loaded HDLs have been
used as vehicles for liver-targeted delivery [42,43]. Encap-
sulated drugs are hydrophobic or amphiphilic compounds
incorporated into the hydrophobic nucleus of nonpolar lipids
of HDLs [44], which can be taken up by SR-BI-mediated
mechanisms [45-49]. In addition to the liver, many tumor
cells overexpress SR-BI and therefore HDLs complexes
have proven to be useful as drug targeting for cancer
chemotherapy [50-55].
HDLs present three main advantages over other drug
nanocarriers. First, the structure of HDLs, which consist of a
hydrophobic nucleus enveloped within a hydrophilic sur-
face, provides an ideal vehicle for transporting amphiphilic
drugs, which is a challenge for other drug delivery systems
[56]. Secondly, since they are human-derived nanoparticles,
they are devoid of danger signals that activate the immune
system [56]. Finally, the uptake into cells is mediated by SR-
BI and, therefore, there is no need of additional targeting
strategies [57]. There are two potential disadvantages of
HDLs as drug carriers. The first limitation is the availability
of a large amount of natural HDLs, and the safety issues
because of the use of a human-derived material. This
limitation is circumvented by the development of a form
of HDL derived synthetically and termed reconstituted HDL
(rHDL) [54,58]. These rHDLs are created by a general
method that allows the generation of rHDL particles with
a specified phospholipid and apolipoprotein content. These
rHDLs are analogous to native nascent HDL, displaying all
of its physiological functions. The diameters of these rHDL
disks vary with the number of apo A-I molecules per particle
(2, 3, or 4) and their conformation [58,59].
Secondly, this method is only suitable for hydrophobic
compounds that can be housed inside artificial membranes in
contact with the hydrophobic fraction of the phospholipids.
This category of compound excludes most of the peptides
and proteins of clinical interest. To extend the benefit of the
HDL stabilization and liver targeting to therapeutic peptide
and proteins, fusion protein of apolipoprotein A-I and
the therapeutic protein can be generated. The proof of
principle of this approach was performed with IFN-a [60].
liver disease treatments: IFN-a. This strategy could combine
the advantages of stabilized and liver-targeted formulations.
To confirm that the fusion protein exhibits long half-life
and liver tropism, the mouse IFN-a gene was fused to the
mouse apo A-I gene (IA) (Figure 29.3). A small linker of
three amino acids (GAP) was used, although this is dispens-
able. The chimeric gene was cloned into an eukaryotic
expression plasmid, and the plasmids coding IFN-a or the
fusion protein (IA) were administered by hydrodynamic
injection. The half-life of IA was 2.6-fold that of IFN-a,
in agreement with the fact that apo A-I half-life is about 2-3
times that of IFN-a, and comparable to the half-life of
pegylated IFN-a [10,11,62-65]. Moreover, the half-life of
IA was only 1.6 times less than IFN-a bound to albumin
(albumin-IFN-a), the formulation with the longest half-life
reported to date for an IFN-a conjugate [66].
As a consequence of the superior half-life of IA and the
liver tropism, hepatic expression of ISGs at day 3 following
injection of a plasmid encoding IA or IFN-a was higher with
the former treatment and even at day 3 after hydrodynamic
injection of plasmids encoding IA or albumin-IFN-a, the
expression of ISGs in the liver tended to be higher with IA
treatment, although serum concentration of IAwas half that
of albumin-IFN-a at this time point.
Finally, these properties were formally demonstrated
using recombinant IA (rIA). Recombinant IFN-a (rIFN-
a) presented a sharp decay in mouse plasma levels, while
the concentration of rIA decreased slowly. Moreover, rIA
accumulated in the liver 2.5 h after i.v. administration, while
rIFN-a was undetectable.
29.1.5 Stabilization and Liver Targeting of IFN- a by
Apolipoprotein A-I
Since apo A-I displays a long plasma half-life comparable to
the half-life of PEG-IFN-a [61,62], and has a previously
demonstrated liver-tropism [41], it was chosen as a stabiliz-
ing and liver-targeting moiety for an important cytokine in
FIGURE 29.3 Molecular representation of the fusion protein of
interferon-a (in gray) and apolipoprotein A-I (in black).
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