Biomedical Engineering Reference
In-Depth Information
metastasis model suggesting a novel role of the fusion
protein as a therapeutic intervention in cancer metastasis
[71]. A lytic peptide (L12) that has been shown to exhibit
anticancer activity was further fused to the C-terminus of
Tat-ELP and the resulting fusion protein possessed cytotoxic
properties to cancer cells in vitro indicating that Tat-ELP-
L12 has the potential to present an effective vehicle to
thermally target tumors [72]. A similar strategy makes
use of the cell penetrating peptide Bac-7. Bac-7 was
N-terminally fused to ELP and the cell-cycle inhibitory
protein p21 was C-terminally fused. The fusion protein
Bac-ELP-p21 revealed antiproliferative effects on SKOV-3
cells specifically after hyperthermia-induced biopolymer
formation [70]. Paclitaxel is a hydrophobic antitumor
drug interfering with the cell cycle by inhibiting the destruc-
tion of microtubules. Paclitaxel was fused to ELP with an
acid-labile linker and increased the overall solubility of
Paclitaxel by forming of ELP micelles (Figure 14.3B).
Hyperthermia induced ELP biopolymer formation increased
the local concentration of ELP-Paclitaxel and forced the
uptake into MCF-7 cells. The acid-labile linker resulted in
Paclitaxel release from ELP within the lysosome and induc-
tion of MCF-7 cell apoptosis [73]. Furthermore, ELP was
conjugated to a therapeutic radionuclide and injected into
tumors in mice (Figure 14.3C). Intratumoral drug delivery
might be an attractive alternative to systemic drug delivery,
which is, however, hampered by rapid clearance of the drug.
ELPs might serve as general framework for local drug
depot formation. Here, a thermally sensitive ELP-conjugate
was compared to a thermally insensitive ELP-conjugate,
with only the first resulting in depot formation, leading to
longer residence time in the tumor and improved antitumor
efficacy of iodine-131. Interestingly, ELP-polymer forma-
tion protected the radionucleotide from cleavage by
dehalogenation [74].
The self-assembly of ELPs into basic forms such as
capsules and aggregates might be utilized to bridge the
gap from biomaterials to nanodevices [75]. ELPs were
cross-linked to form matrices with mechanical properties
comparable to those of native elastin [76]. With a better
understanding of the 3D architecture and structural trans-
formations of ELP aggregation, it will be possible to produce
tailor-made ELPs for biotechnological applications. A first
example for this development is soluble ELP (sELP), which
combines physical and mechanical features of silkworm silk
and ELPs in one polypeptide. sELPs formed temperature
and pH responsive hydrogels [77] and were spun into fibers
[78,79] with a high tensile strength and high deformability
[80]. A second example is the formation of an ELP layer
with pH-controllable nanopore formation [81]. Considering
the responsiveness of ELPs to temperature, it should be
possible to improve this feature to literally build nano-
structures with switch-mechanisms, for example, to open
or close nanopores at will.
14.4 MOLECULAR PHARMING: A NEW
APPLICATION FOR ELPYLATION
For centuries, plant products have been used with different
intention, as part of foods and powders to cure and prevent
diseases or, as plant raw materials. The twenty-first century
brought plants a new function, namely, the synthesis of
heterologous proteins. Traditional production systems for
recombinant proteins, microbial fermentation, insect or
mammalian cell cultures, and transgenic animals, hold
drawbacks in terms of production costs, scalability, and
product safety. These limitations can be addressed through
the use of plants as production hosts. In developing countries
with a low level of technological infrastructure, where the
demand for pharmaceuticals is greatest, but where the
financial resources to pay for them are the most limited
the production of recombinant proteins in transgenic plants
is probably more realistic than the current alternatives of
bacterial, yeast, insect, and mammalian cells [82-84].
Neutralizing and infection-preventing antibodies against
HIV-1 as components of microbicides [85] or recombinant
vaccines against avian influenza [86] are promising candi-
date proteins and currently under investigation. The term
molecular pharming refers to the large-scale production and
purification of pharmaceutical proteins in transgenic plants
or plant-based expression systems. Four systems for plant-
based expression of recombinant proteins have been devel-
oped so far; plant cell cultures (including duck weed and
mosses), transient expression systems using Agrobacterium
tumefaciens or plant viruses for introducing the expression
cassette into plant leaves, expression in transplastomic
plants, and expression in stably transformed transgenic
plants [87]. Several plant species are used as expression
systems [88]. Since the successful expression of assembled
immunoglobulins in transgenic tobacco plants [89] and the
first report of plant-based vaccine production [90], a large
number of different vaccines, antibodies, as well as antibody
fragments have been produced in plants for both medical and
veterinary purposes and have entered clinical trials [91].
However, only two plant-produced vaccine-related products
have gone all the way through the production and regulatory
hurdles, and only one, a plant made single-chain variable
fragment (scFv) is used in the production of a recombinant
hepatitis B virus (HBV) vaccine in Cuba [92]. A major step
was made at the beginning of 2006 by Dow AgroSciences
(DAS, Indianapolis, USA). Their plant cell expressed veter-
inary vaccine against the Newcastle disease virus (NDV),
produced in a suspension-cultured tobacco cell line, has
gained regulatory approval by the U.S. Department of
Agriculture (USDA) Center for Veterinary Biologics—the
final authority for veterinary vaccines in the United States
[93]. However, this vaccine has not been introduced to the
market yet. Dow AgroSciences apparently wished to dem-
onstrate that their Concert
TM
Plant-Cell-Produced system
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