A novel functionalized chitosan-based nanoconstruct for targeted subcutaneous delivery of protein therapeutics in squamous cell carcinoma

摘要

Introduction: The significance of both peptides and proteins, as potent therapeutics has increased drastically and administered through the parental route as injectables. Meanwhile, subcutaneous administration has been reported as a robust means for the delivery of many of the approved protein therapeutics. Chitosan based nanoparticles have been investigated and employed as carriers for genes and therapeutic proteins with different levels of effectiveness and set-backs in cancer therapy. Non-targeted delivery and sub-optimal dosage at disease sites are major limitations of most conventional chemotherapeutics. This study aimed to design and characterize novel ligand-enhanced chitosan-based nanoparticles with potential and promising applications as targeted delivery cargos for protein therapeutics in the management of Squamous Cell Carcinoma (SCC). Materials and Methods: Using BSA as a model protein, we have synthesized and characterized a protein loaded PEGYIated chitosan-based polyethyleimine nano-constructs using tripolyphosphate (TPP) as a polyanionic agent. The BSA-loaded chitosan-based nano-construct was characterized for structural and functional modifications, particle size, morphology, zeta potential, thermal stability, protein loading efficiency and subsequent release kinetics. Surface modification of trie synthesized nanoconstructs will be achieved using LyP-1 as a homing ligand for targeted delivery. MTT assay will be performed to access the cytotoxic efficacy of the functionalized nanoconstructs loaded with endostatin in place of BSA. Results and Discussion:H1 NMR and FT-IR experiments both confirmed the successful grafting of the co-polymers to chitosan moiety as presented. The average size and morphology of the BSA loaded nanoparticles as confirmed by TEM, dynamic light scattering (DLS) and SEM results showed that the nanoparticles exhibited a spherical shape and had a narrow particle size distribution with polydispersity index (PDI) of 0.496 with an average size less than 100nm. A positive value zeta potential of 19.8mV was obtained for the nanoparticles with minimal stability with possible mucoadhesive properties. Both the TGA and DSC results showed the synthesized BSA loaded nanoparticles were more stable than the single polymers. The protein loading efficiency was evaluated to be 77.2% with an initial burst release probably due to surface protein desorption and subsequent prolong slow release through diffusion from sublayers only after 6 hours up to 48 hours. Conclusions: In general, this study demonstrated that grafting other biopolymers such as PEI and PEG to chitosan improved its thermal stability and could as well enhances its protein encapsulation and possible release kinetics. However, the initial burst release effect, in releasing large amount of protein molecules remained a major drawback of chitosan-based nanoparticles and its application as potent carrier of protein therapeutics but could be harnessed for cytotoxic effect in tumor cells. Meanwhile, our designed nanoconstmct is intended for targeted delivery of endostatin upon its functionalization for enhanced therapeutic effects in SCC management and represent the first prototype for protein delievry when these polymers are complexed as a single system.