CRISPR-Cas9 is a revolutionary genome-editing technology that has enormous potential for the treatment of genetic diseases. However, the lack of efficient and safe, non-viral delivery systems has hindered its clinical application. Here, we report on the application of polymeric and hybrid microcarriers, made of degradable polymers such as polypeptides and polysaccharides and modified by silica shell, for delivery of all CRISPR-Cas9 components. We found that these microcarriers mediate more efficient transfection than a commercially available liposome-based transfection reagent (>70% vs. <50% for mRNA, >40% vs. 20% for plasmid DNA). For proof-of-concept, we delivered CRISPR-Cas9 components using our capsules to dTomato-expressing HEK293T cells-a model, in which loss of red fluorescence indicates successful gene editing. Notably, transfection of indicator cells translated in high-level dTomato knockout in approx. 70% of transfected cells. In conclusion, we have provided proof-of-principle that our micro-sized containers represent promising non-viral platforms for efficient and safe gene editing.
An
important area in modern malignant tumor therapy is the optimization
of antitumor drugs pharmacokinetics. The use of some antitumor drugs
is limited in clinical practice due to their high toxicity. Therefore,
the strategy for optimizing the drug pharmacokinetics focuses on the
generation of high local concentrations of these drugs in the tumor
area with minimal systemic and tissue-specific toxicity. This can
be achieved by encapsulation of highly toxic antitumor drug (vincristine
(VCR) that is 20–50 times more toxic than widely used the antitumor
drug doxorubicin) into nano- and microcarriers with their further
association into therapeutically relevant cells that possess the ability
to migrate to sites of tumor. Here, we fundamentally examine the effect
of drug carrier size on the behavior of human mesenchymal stem cells
(hMSCs), including internalization efficiency, cytotoxicity, cell
movement, to optimize the conditions for the development of carrier-hMSCs
drug delivery platform. Using the malignant tumors derived from patients,
we evaluated the capability of hMSCs associated with VCR-loaded carriers
to target tumors using a three-dimensional spheroid model in collagen
gel. Compared to free VCR, the developed hMSC-based drug delivery
platform showed enhanced antitumor activity regarding those tumors
that express CXCL12 (stromal cell-derived factor-1 (SDF-1)) gene,
inducing directed migration of hMSCs via CXCL12 (SDF-1)/CXCR4 pathway.
These results show that the combination of encapsulated antitumor
drugs and hMSCs, which possess the properties of active migration
into tumors, is therapeutically beneficial and demonstrated high efficiency
and low systematic toxicity, revealing novel strategies for chemotherapy
in the future.
Mesenchymal stem cells (MSCs) are widely used in cell therapy due to their convenience, multiline differentiation potential, reproducible protocols, and biological properties. The potential of MSCs to impregnate magnetic microcapsules and their possible influence on cell function and ability to response to magnetic field have been explored. Interestingly, the cells suspended in media show much higher ability in internalization of microcapsules, then MSCs adhere into the surface. There is no significant effect of microcapsules on cell toxicity compared with other cell line-capsule internalization reported in literature. Due to internalization of magnetic capsules by the cells, such cell engineering platform is responsive to external magnetic field, which allows to manipulate MSC migration. Magnetically sorted MSCs are capable to differentiation as confirmed by their conversion to adipogenic and osteogenic cells using standard protocols. There is a minor effect of capsule internalization on cell adhesion, though MSCs are still able to form spheroid made by dozen of thousand MSCs. This work demonstrates the potential of use of microcapsule impregnated MSCs to carry internalized micron-sized vesicles and being navigated with external magnetic signaling.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.