Periodic Reporting for period 1 - HsrELPnano (Healthcare applications of Stimuli-Responsive Elastin-Like Polypeptide Nanocarriers)
Período documentado: 2021-08-01 hasta 2023-07-31
Traditional chemotherapy might be effective but the associated side-effects and toxicity cause a huge negative impact in patient's life or even death. The development of nanocarriers to host those drugs until they reach the target tissue helps to increase the benefit-risk balance and reduce the side effects. Elastin-like polypeptides (ELPs) are an excellent opportunity to build nanocarriers due to their biocompatibility, non-immunogenicity, drug encapsulation and localization possibilities. In the other hand, these systems are disassembled upon dilution and thus their stability can be compromised when injected into the bloodstream or a tissue. These ELP-NC hybrids are especially interesting because they add the inorganic features (fluorescence, magnetic or pH/redox responsive properties) to the peptide-based assemblies. The contribution of this proposal rely on the development of new possibilities and tools in the biomedical field, enhancing the quality of future drug formulations and therefore contributing in a small way to ensure healthy lives and promote well-being for all at all ages. The objectives of this project are focused on the understanding of how ELP nanocarriers can be stabilized by using metal-based nanoparticles, the characterization of the so-called hybrid ELP nanoparticles chemo-physical properties, and their capability to load a cargo that can deliver a treatment in vivo.
We have designed different ELPs with inorganic nanoparticle binding properties to achieve the growth or binding of metal-based nanocrystals (NCs) as a way to form nanoparticles. The design consists in a hydrophilic ELP block followed by a more hydrophobic block with small peptide sequence at the end which is capable to specifically bind gold surfaces. We performed the assembly of ELP nanoparticles through the binding to Au nanoclusters, which can stabilize the ELP NPs assembly upon dilution and temperatures below the transition temperature. This approach is based on the generation of pristine Au nanoclusters (without functionalization of the surface) and further mixing with the diblock ELPs equipped with a Au peptide binding motif. Since ELP nanoparticle assembly is dependent on both the concentration and temperature, a set of experiments was designed to observe the AuNC-ELP hybrid behavior and understand its characteristics. In summary, the results showed that it is possible to generate stable hybrid nanoparticles with different amounts of AuNKNCs and different AuELP concentrations. The assemblies are notably sensitive to the protocol used, where the route in which the assembly is done by slow heating showed the most reliable size measurements over time. The stability of the hybrids upon dilution and cooling below transition temperature supports the hypothesis in which AuNCs can act as crosslinking points to hold the NPs together. Finally, the absence (or smoothness) of the transition temperature of the ELP hydrophobic block and the drop of the transition temperature of the hydrophilic block could reveal that the metal binding event brings ELP chains in close proximity causing the assembly of the particles even at temperatures below the transition temperature. TEM images confirmed the formation of nanoparticles with 200 nm in size that contain gold nanoclusters.
In a second approach, we made use of a hydrophilic block ELP together with a redox block able to reduce inorganic salts and achieve the formation of the ELP hybrid nanoparticles. We performed different synthetic routes to achieve the reduction of potassium permanganate to form manganese oxide nanoparticles, which can be dissolved in biologically relevant concentrations of glutathione (GHS) and hydrogen peroxide. The results showed that the synthesis route determined the long-term stability of manganese oxide-ELP hybrids, where a co-solvent synthesis has demonstrated the best stability in physiological conditions. The particles generated kept the thermoresponsive behavior of the hydrophilic block, which proves that tha surface of the particles is stabilized by this block. In addition, the formation of the maganese oxide and its dissolution was monitored, showing the possibility of triggering the disassembly of the particles by cancer cells. The hybrid particles were tested for their competence in magnetic resonance imaging, showing a successful performance as a MRI contrast agents.
In a second approach, we made use of a hydrophilic block ELP together with a redox block able to reduce inorganic salts and achieve the formation of the ELP hybrid nanoparticles. We performed different synthetic routes to achieve the reduction of potassium permanganate to form manganese oxide nanoparticles, which can be dissolved in biologically relevant concentrations of glutathione (GHS) and hydrogen peroxide. The results showed that the synthesis route determined the long-term stability of manganese oxide-ELP hybrids, where a co-solvent synthesis has demonstrated the best stability in physiological conditions. The particles generated kept the thermoresponsive behavior of the hydrophilic block, which proves that tha surface of the particles is stabilized by this block. In addition, the formation of the maganese oxide and its dissolution was monitored, showing the possibility of triggering the disassembly of the particles by cancer cells. The hybrid particles were tested for their competence in magnetic resonance imaging, showing a successful performance as a MRI contrast agents.
The project has reveal new insights in the formation of ELP hybrids. In this time, we have unveil new aspects in the balance between the presence (or growth) of inorganic nanoparticles and ELP assembly properties, which is highly relevant in the field and will increase the impact of R&I output of elastin-like polypeptides. We have make a useful progress in the understanding of the mechanism behind the formation of these hybrid particles and how to control their stability and properties by synthetic means. The results are expected to be published in two research articles in highly ranked peer reviewed international journals. Moreover, the knowledge I cultivated during this action has given me the chance to publish an expert opinion in Frontiers in Materials journal. The publication of the results will enhance the application possibilities of ELPs as platform for other authors and inventors and thus it may have a future impact in the biomedical field.