Construction Of Intelligent Targeted Nanogel And Its Antitumor Mechanism

Cancer is one of the diseases that serious threaten to human health today, andtraditional therapy methods (surgical resection, chemotherapy, radiation therapy) haveserious side effects due to the lack of targeting and the normal organs and tissues are ofteninjured. Furthermore, the metastasis rate of malignant tumor is high at late stage, thus theireffectiveness is difficult to further improve, and the prognosis is poor. In the field of cancertargeted therapy, gene therapy research is increasingly attracting widespread concern andattention by scientists. Thanks to the development of gene manipulation and DNArecombinant technology, gene therapy for cancer is a breakthrough in the field of tumortherapy. First of all, the exogenous genes (gene fragments, pDNA, RNA, etc.) weretransfected into target cells, through correction, compensation, replacement or inhibit theabnormal expression of genes in the tumor cells, so as to achieve the purpose of treatment,gene therapy is also the hot spot in the study of tumor treatment.The primary technical issues facing by cancer gene therapy is to select a targetingvector of the safe, efficient and can introduce the negatively charged gene into the targetcells. Although the transfection efficiency are relatively high by traditional physicaltechnologies (such as microinjection, electroporation) they are limited by high technologyand equipment requirements, and only can be used in limited tissues such as the skin andmuscle. In addition, these traditional physical transfection technologies have some extendof damage to the target cells, and it can not be performed in vivo.Although the transfection efficiency are relative high using viral vectors (retrovirusvector, adenovirus vector and adenovirus-associated virus vector or the like), there areobvious defects of non-targeted, highly immunogenic. Furthermore, the security problemof the viral vector has been controversial. Although cationic liposomes overcome securityproblems of the viral vector, but the transfection efficiency is low, due to the poor serumstability, the composition of the carrier system complexity and wide structural sizedistribution. Although adding a variety of auxiliary components (cholesterol, emulsifierand dispersing agent) can improve the efficiency of cationic liposomes transfection,introduction of adjuvant also reduce their clinical value. Therefore, designing and makingnovel specific targeting non-viral nanocarriers which are simple in structure, high intransfection efficiency and stable in vitro and in vivo is the key of gene therapy.Safe cationic vectors with high transfection efficiency and serum stability are highly desired in practical gene delivery. Herein, poly (ethylenimine)(PEI,25KD) wasmodified by copolymerization with N-isopropylacrylamide (NIPAM) to form amphiphilicgraft copolymer. A thermosensitive PNIPAM/PEI core shell nanolgel was in-suit obtainedby selfmulsification from this copolymer with volume phase transition temperature(VPTT)~32℃. Both the narrow size (~300nm) and well-defined core shell structurewas clearly confirmed by TEM images. Its cytotoxicity assay against293T cells wasextremely lower than that of PEI was the surface potential decreased to~20mV. But in theserum containing condition, the transfection efficiency was ampilfied about2times whichwas attributed to the uniform and stable PNIPAM/PEI polyplex (~300nm) with weaksurface potential (~2mV). Such transfection was further promoted about2times by thetemperature regulated cellular targeting as T> VPTT. As observed by IVIS LuminaImaging System (Caliper Life Sciences, Hokpinton, MA), the intratumor accumulation ofsuch stable and uniform nanogel polyplex was obviously promoted24h post i.v.administration to Balb/c nude mice bearing breast cancer. The in vivo xenografted breasttumor growth was significantly inhibited by pRA-EGFP gene encoding ricin A (RA)protein as delivered by the thermosensive PNIPAM/PEI nanogel due to its high serumstability, enhanced cellular targeting and intracellular proton-buffering capability, Therelative tumor volume was about~2.3times (i.v.) and5times (i.t.) smaller than that ofthe control group. These results presented in this paper show that the non-cytotoxicitycationic thermosensitive PNIPAM/PEI core shell nanogels could be effective plasmid DNAcarriers for gene therapy.This thesis mainly focuses on the tumor microenviroment and targets to the obstaclesappeared in the clinical antitumor therapy. By polymer design, intelligent nanogelpreparation, systemically investigation and comparison, we found that the tumor can besuccessfully suppressed by the intelligent nanogel based gene delivery system. We do hopeour work would be referable for the gene therapy in clinical oncology.