Biomedical Engineering Reference
In-Depth Information
4.4 ENZYME-RESPONSIVE SYSTEMS
Like their pH-responsive counterparts, systems that respond to specific enzymes exhibit great
potential due to the disease-specific nature in which the expression of particular enzymes is upregu-
lated. In order for cancerous cells to continue growing, promote angiogenesis, and metastasize to
other tissues, extensive remodeling of the extracellular matrix is required (Forsyth et al., 1999).
This means that various enzymes, such as the matrix metalloproteases (MMPs), must be secreted
in exceedingly large quantities. This overexpression of MMPs makes them a potential target for use
in a stimuli-responsive drug delivery system, which can be designed such that part of the system
must be cleaved by the MMPs in order to allow drug release. This is exactly the strategy that was
employed by Tauro et al
.
who linked platinum to a short six amino acid peptide sequence that was
a known substrate for MMPs (Tauro et al., 2008). These platinum-peptide complexes were then
loaded into hydrogels such that the rate of platinum release was dependent on MMP cleavage of
the peptide. After demonstrating successful MMP-controlled platinum release
in vitro
, the peptide
complexes were evaluated for biocompatibility against malignant glioma cells (HTB-14) using the
MTT assay. Following 24 h incubation, the intact peptide complex did not show any cytotoxicity
against the HTB-14 cells at any of the concentrations tested (0.5-200 μM). Likewise, the products
of peptide cleavage by MMPs did not show any cytotoxic effects when studied under the same
conditions as the intact peptides. Further studies were conducted to evaluate the biocompatibility of
the peptide-loaded hydrogels. These studies, evaluated using the MTT assay following 48 and 96 h
incubations, also showed no cytotoxic effects induced by the hydrogel or the peptides contained
within. Follow-up studies were conducted in which exogenous MMPs (MMP-2 and MMP-9) were
added to the cell culture media. These studies demonstrated that the platinum-peptide complexes
contained within the hydrogel showed no cytotoxicity at 24 or 48 h unless exogenous MMP-2 and
MMP-9 were added to the media. Taken together, the data indicate that the system is biocompatible
until the peptide is cleaved and the cytotoxic platinum component is released.
A similar approach was utilized by Pedersen et al. in the development of a liposomal formula-
tion composed of a retinoid-phospholipid prodrug (Pedersen et al., 2010). This prodrug was able
to form liposomes due to the incorporation of the phospholipid component. The cytotoxic retinoid
portion was only released once it was cleaved off of the phospholipid by secretory phospholipase A
2
IIA (sPLA
2
). Using the MTT assay, the biocompatibility of the prodrug was assessed using two cell
lines: HT-29 (human colon carcinoma) and Colo-205 (human colon adenocarcinoma). The HT-29
cells do not secrete sPLA
2
, while the Colo-205 cells do secrete this enzyme. This allowed the inves-
tigators to evaluate the cytotoxicity of their prodrug in the presence and absence of the activating
enzyme in quantities that closely mimic the natural expression level of sPLA
2
seen clinically in
colon adenocarcinoma. Further control over the system was demonstrated through the addition of
exogenous sPLA
2
(5 nM). Following 24 and 48 h incubations, the prodrug was not able to induce
significant cytotoxicity in the absence of the sPLA
2
enzyme (HT-29 cells). However, when sPLA
2
was present, the cytotoxicity of the prodrug increased over 20-fold; with IC
50
values below 10 μM
for HT-29 cells when exogenous sPLA
2
was coincubated, and below 20 μM for Colo-205 cells (no
exogenous sPLA
2
added). This data indicated that the prodrug formulation was biocompatible until
cleaved by the activating enzyme sPLA
2
, and that cells that naturally secrete this enzyme (Colo-205
cells) are able to cleave the prodrug into its active form within 24 h.
4.5 FACTORS INFLUENCING BIOCOMPATIBILITY OF STIMULI-RESPONSIVE
NANOMATERIALS
Many factors affect the body's response to the introduction of foreign materials, including mor-
phology, size, presence of complement activation groups (e.g., hydroxyl groups), and the surface
chemistry of the foreign material. The surface chemistry (hydrophilic/hydrophobic nature and sur-
face charge) is possibly the most important parameter in determining how the body will react to
