Preparation Of Bovine Serum Albumin-polymer Conjugates And Their Applications In Tumor Diagnosis And Therapy

As a type of newly emerging biomaterial, the self-assembled nanostructures ofbiomolecules, especially the ones based on albumin, are attracting more attention inthe field of nanomedicine because of their precise geometry, versatile functions, andinherent biocompatibility with biosystems. In this study, the novel protein-polymerconjugates were fabricated using bovine serum albumin (BSA) as substrate. Theprepared protein-polymer conjugates could self-assemble into nanosized aggregatesand a new nano-platform was constructed based on the self-assembly ofprotein-polymer conjugate, which could combine chemotherapy, radionuclide therapyand fluorcent imaging together perfectly. The details are as follows:(1) A novel biodegradable protein-polymer conjugate was prepared by covalentlylinking the tailor made hydrophobic maleimide-functionalized poly(-caprolactone)(PCL) to hydrophilic bovine serum albumin (BSA) via the maleimide sulfhydrylcoupling reaction. The characterization of Mal-PCL and BSA-PCL conjugates werecarried out via1H NMR spectra, MALDI-TOF MS and SDS-PAGE gelelectrophoresis and the results showed that the BSA-PCL conjugates were preparedsuccessfully. The obtained conjugates can self-assemble into nanosized aggregateswith different morphologies.(2) Dox-loaded polymersome was constructed by the self-assembly of amphiphi-lic BSA-PCL conjugate and cetuximab as a targeting ligand was linked to theperiphery of the vesicle. The result of cell uptake suggested that the protein-basedvesicle could perform active targeting of U251cells and MAD-MB-231cells viacetuximab functionalization. MTT results showed that the BSA PCL conjugate had alow cytotoxicity and the DOX-encapsulated cetuximab-functionalized vesicle hadbetter therapeutic efficacy with respect to U251cells and MDA-MB-231cells thanthat of without cetuximab.(3) In order to enhance the cell uptake and retention time in cells of radionuclides,131I labeled BSA-PCL conjugate nanoparticle was prepared via chloramine-T methodusing the residue tyrosine of BSA as the conjugation site. Further, the nanoparticlewas decorated with targeted ligand to enhance the ability to target the tumor cells.And the cetuximab decorated131I labeled BSA-PCL conjugate nanoparticle showedenhanced cell uptake and better therapeutic efficacy with respect to U251cells.(4) A simple, straightforward and reproducible method for direct fabrication ofNIR fluorescence nanoprobe was developed by coating CuInS2/ZnS (CIS/ZnS) QDs with a novel amphiphilic BSA-PCL conjugate for fluorescence imaging. Thedeveloped BSA-PCL conjugate could self-assemble into nano-sized micelle and itcould be used as a platform to achieve the surface engineering of hydrophobicnanoparticle. And the constructed NIR fluorescence nanoprobe exhibited excellentfluorescent properties, good colloidal stability in PBS buffer (pH=7.4) with10%fetalbovine serum, and high biocompatibility and nontoxicity to the living cells.Furthermore, cRGD-decorated nanoprobe can be readily taken up by cells used fortargeted cell imaging. Finally, the feasibility of the NIR fluorescence nanoprobe isverified in vivo imaging. This approach for the construction of NIR fluorescencenanoprobe with novel amphiphilic bio-block copolymer will find wide application inthe preparation of multimodality imaging agents and versatile theranostics.(5) Smart protein–polymer conjugate nanoparticles have potential applications inbiotechnology. These ‘smart’ conjugates are characterized by their ability to changephysico-chemical properties on the stimulus. Herein, protein-reactive ATRP initiatorwas first synthesized and pH-sensitive maleimide-functionalized polymer poly(2-(Diisopropylamino)ethyl methacrylate)(PDPA) was prepared by atom transferradical polymerization (ATRP) using the tailor made ATRP initiator. Then thepH-sensitive BSA-PDPA conjugate was prepared by covalently linking thepH-sensitive maleimide-functionalized PDPA to hydrophilic BSA via themaleimide-sulfhydryl coupling reaction. SDS-PAGE gel electrophoresis resultsshowed that the BSA-PDPA conjugate was prepared successfully. And the Zetapotential of BSA-PDPA conjugate dissolved in the water with different pH value wasmeasured and the result showed BSA-PDPA conjugate was pH-sensitive. Theprepared BSA PDPA conjugate could self-assemble into a spherical nanoparticle with135nm in size. To confirm and realize the pH-sensitivity of the nanoparticle and thepotential of the BSA PDPA conjugate in the drug delivery, further investigations arerequired.In conclusion, the protein-polymer conjugates based on BSA and syntheticpolymers were prepared via maleimide–thiol reactions, and a new nano-platformcolud be constructed using the self-assembly of protein-polymer to combinechemotherapy, radionuclide therapy and fluorcent imaging together perfectly, all ofwhich provide important evidence for the application of nanomaterials and nano-aggregation based on BSA in the field of nano-biomedicine.