cysteine groups for drug conjugation. The anticancer drug DOX is attached to the
cysteine residues via an acid-cleavable hydrazone linker. Up to 28 DOX molecules
have been uploaded per protein in a reproducible fashion. The protein backbone
backfolds after drug loading to form DOX-HSA-PEO multishell micelles with
(1) DOX molecules encapsulated inside the core, (2) albumin protein as the
enzyme-degradable protecting shell, and (3) PE as water-soluble and immunesilencing shell. This drug delivery system exhibits a controlled two-step drug
release mechanism in response to proteolysis and acidic pH environment, therefore
reducing undesirable drug leakage. The cell toxicity has been examined in vitro as
well as in vivo. A highly potent toxicity effect in an acute myeloid leukemia (AML)
model has been observed, with an IC50 in the subnanomolar range, and long-term
suppression of AML growth has been observed in a mice model [44]. Furthermore,
by modifying the multiple functional groups of the protein shell, additional
functionalities, such as MRI contrast reagents, could be further attached to this
multishell structure. For instance, the most widely used T1 MRI contrast reagent,
Gd-DOTA, has been conjugated to the lysine residues of the protein backbone, thus
resulting in a theranostic complex that potentially allows therapeutic drug delivery
and diagnostic imaging simultaneously (our unpublished data). In the future, this
system offers great potential for developing personalized anticancer therapy.
1.5 Conclusions
Polyelectrolyte interactions play an important role in cell biology. By introducing
multiple positive charges into the periphery of fluorescently labeled polyphenylene
dendrimers, charge-dependent membrane uptake and cell toxicity was found. In
addition, polycationic dendrimers were able to tightly complex and release DNA
and stain the extracellular matrix by interacting with the natural aggrecane
polyanions. Dendritic polyanions, in contrast, were not able to pass the cellular
membrane; however, in fixed cells, tight binding to histone proteins in the cell nucleus
was observed, where DNA was most probably displaced from the complexes. Both
the polyanionic and polycationic dendritic chromophores specifically stained cellular
structures such as the ECM and the cell nucleus, based on the presence of multiple
charges within a small nanoscopic volume. In addition, protein polyelectrolytes have
been prepared by converting surface-exposed residues into primary amino or carboxylic acid groups. Albumin polycations revealed considerably lower cytotoxicities
than their synthetic analogs as well as efficient cell uptake and intracellular release.
Complex stoichiometries with DNA could be adjusted by the charge densities of the
albumin polycations. Tight DNA binding and efficient transfection was achieved with
the highest cationized cBSA derivative.
Cationization of albumin proteins diminished the accessibility of the albumin
binding pockets. However, the attachment of positively charged dendrons allowed
guest uptake into the hydrophobic binding pockets and efficient delivery of the
cytotoxic drug DOX has been achieved. Denaturing the albumin backbone of
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J. Hedrich et al.
cysteine residues via an acid-cleavable hydrazone linker. Up to 28 DOX molecules
have been uploaded per protein in a reproducible fashion. The protein backbone
backfolds after drug loading to form DOX-HSA-PEO multishell micelles with
(1) DOX molecules encapsulated inside the core, (2) albumin protein as the
enzyme-degradable protecting shell, and (3) PE as water-soluble and immunesilencing shell. This drug delivery system exhibits a controlled two-step drug
release mechanism in response to proteolysis and acidic pH environment, therefore
reducing undesirable drug leakage. The cell toxicity has been examined in vitro as
well as in vivo. A highly potent toxicity effect in an acute myeloid leukemia (AML)
model has been observed, with an IC50 in the subnanomolar range, and long-term
suppression of AML growth has been observed in a mice model [44]. Furthermore,
by modifying the multiple functional groups of the protein shell, additional
functionalities, such as MRI contrast reagents, could be further attached to this
multishell structure. For instance, the most widely used T1 MRI contrast reagent,
Gd-DOTA, has been conjugated to the lysine residues of the protein backbone, thus
resulting in a theranostic complex that potentially allows therapeutic drug delivery
and diagnostic imaging simultaneously (our unpublished data). In the future, this
system offers great potential for developing personalized anticancer therapy.
1.5 Conclusions
Polyelectrolyte interactions play an important role in cell biology. By introducing
multiple positive charges into the periphery of fluorescently labeled polyphenylene
dendrimers, charge-dependent membrane uptake and cell toxicity was found. In
addition, polycationic dendrimers were able to tightly complex and release DNA
and stain the extracellular matrix by interacting with the natural aggrecane
polyanions. Dendritic polyanions, in contrast, were not able to pass the cellular
membrane; however, in fixed cells, tight binding to histone proteins in the cell nucleus
was observed, where DNA was most probably displaced from the complexes. Both
the polyanionic and polycationic dendritic chromophores specifically stained cellular
structures such as the ECM and the cell nucleus, based on the presence of multiple
charges within a small nanoscopic volume. In addition, protein polyelectrolytes have
been prepared by converting surface-exposed residues into primary amino or carboxylic acid groups. Albumin polycations revealed considerably lower cytotoxicities
than their synthetic analogs as well as efficient cell uptake and intracellular release.
Complex stoichiometries with DNA could be adjusted by the charge densities of the
albumin polycations. Tight DNA binding and efficient transfection was achieved with
the highest cationized cBSA derivative.
Cationization of albumin proteins diminished the accessibility of the albumin
binding pockets. However, the attachment of positively charged dendrons allowed
guest uptake into the hydrophobic binding pockets and efficient delivery of the
cytotoxic drug DOX has been achieved. Denaturing the albumin backbone of
224
J. Hedrich et al.