Developing bio-orthogonal chemistries to prepare nanocarriers for controlled release
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Abstract
The design of nanocarriers for drug delivery requires tremendous efforts and planning to
ensure the preparation method is compatible with the drug used as the payload, that the
resulting nanocarriers are biocompatible, ideally biodegradable and display in vivo stability.
Furthermore, they need to be able to target specific cells, be uptaken by the cells, and able to
release the payload after cell uptake. All those requirements need to be met with a fabrication
method that can be scaled up to meet the requirements in terms of scale, quality, and
reproducibility associated with the pharmaceutical industry. The main goals of this thesis were
to develop a process that can be used for the large scale synthesis of high-quality nanocarriers
and to develop new crosslinking chemistry possibilities to produce nanocarriers suitable for the
encapsulation of sensitive payloads.
In this thesis, microfluidization was used to prepare large quantities of nanocarriers
synthesized based on the interfacial crosslinking of precursor droplets formed by inverse
miniemulsion. Those nanocarriers were prepared in high quality and large quantity in a
reproducible manner with the possibility to tune the size of the nanocarriers (section 4.1). The
versatility of the microfluidization method was also demonstrated by using different precursor
polymers such as polysaccharide, proteins, and lignin, and by using different crosslinking
strategies (section 4.1).
The microfluidization approach to prepare the precursor droplets was then combined
with new crosslinking strategies. Bio-orthogonal reactions involving the reaction between
reactive carbonyls and hydrazide derivatives were used to prepare stimuli-responsive
nanocarriers (sections 4.2 and 4.3). Rather than using an unselective reaction that can react with
complex and sensitive payloads, a strategy based on the selective reaction between dextran
functionalized either with aldehyde or terminal ketone groups and polyfunctional hydrazide
derivative was used to produce nanocapsules (Section 4.2) or nanogels (section 4.3). This reaction
is highly suitable because it is a selective reaction, has a sufficiently high reaction rate, and since
the stability of the resulting hydrazone linkages is pH-sensitive, those new nanocarriers enabled
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the release of the payloads it in a spatiotemporally controlled manner. The dissociation of the
hydrazone crosslinking points in mildly acidic conditions was responsible for the controlled
release of the cargos.
In the first approach (section 4.2), the hydrazone network was built by interfacial
crosslinking of inverse miniemulsion droplets to form nanocarriers. The water droplets contained
the dextran precursor, and the crosslinker poly(styrene-co-methyl hydrazide) was dissolved in
the continuous toluene phase. These nanocarriers with capsule morphology were able to both
encapsulate model hydrophilic compounds and release them upon changing the acidity of the
environment. Furthermore, they were uptaken by HeLa cells and did not show any noticeable
cytotoxicity even at high concentrations. For this reason, nanocarriers represent a promising
approach for gene and medication delivery and the targeting of many pathological environments
and specific intracellular compartments that are more acidic than the normal physiological
conditions.
The second approach (section 4.3) was based on the reaction between aqueous droplets
containing the functionalized dextran and other aqueous droplets containing the water-soluble
crosslinker. The formation of the hydrazone network led to the formation of nanogels by mixing
the two types of precursor droplets. In addition to bearing two hydrazide groups able to create
the pH-responsive network, those water-soluble crosslinkers also bear other functionality; the
crosslinkers contained either a disulfide bond reactive in the presence of a reducing environment
or a thioketal bonds responsive to the presence of reactive oxygen species. The resulting
nanogels successfully encapsulated large payloads, and the release of the payload could be
triggered by changes in acidity, the addition of dithiothreitol or glutathione as a reducing agent
or by the addition of superoxide as Reactive oxygen species (ROS). The nanogels displayed
limited toxicity and good uptake in HeLa cells. The results gathered and obtained in section (4.3)
could pave the way for building desired multi-stimuli responsive polymer nanogels for specific
tumor-targeting.