Poly(L-ornithine)s As Potential Antibacterial And Anticancer Nanopolymers

Host defense peptides(HDPs)are short peptides found in almost all multicellular organisms and play an important role in the nonspecific defense of microorganisms such as bacteria,fungi and viruses.Natural HDPs show great antibacterial potential in vitro,but there are many obstacles in clinical applications,including complex production procedures,high cost,low bioavailability,and enzymolytic instability.As a promising alternative,cationic peptides that mimic HDPs' structures have unique advantages,including high mass production,cost-effectiveness,high compatibility with drug delivery systems and ease of chemical modification.Based on cationic peptides that mimic HDPs,the present thesis constructs antimicrobial peptides and anticancer peptides with unnatural L-ornithine as the cationic amino acid.The main factors affecting the antibacterial activity of antimicrobial peptides are positive charge density,charge/hydrophobicity balance and molecular conformation.A series of cationic star-shaped antimicrobial peptides PEI-g-PLO,PEI-g-PLL and PEI-g-PLH with polyethyleneimine(PEI)as the core,poly(L-ornithine)(PLO),poly(L-lysine)(PLL)and poly(L-?,?-diaminoheptylic acid)(PLH)as the periphery are synthesized in the present study.Without introducing hydrophobic groups,the charge/hydrophobicity balance of star-shaped antimicrobial peptides is regulated by tailoring the length of methylene segments across the side chains of amino acids,and the antibacterial properties are optimized.Notably,the construction of star-shaped conformation and the introduction of unnatural amino acid structural units not only improve the positive charge density of antibacterial peptides but also enhance their enzymolytic stability.In vitro antimicrobial investigations show that PEI-g-PLO has high enzymolytic stability,strong biofilm disruption and broad-spectrum antibacterial activity,especially for Gram-negative Pseudomonas aeruginosa.In vivo antibacterial studies exhibit that PEI-g-PLO significantly reduces the microbial burden of infection caused by Pseudomonas aeruginosa in burn sites and effectively promotes the healing of burn wounds.Based on the common mechanism of antibacteria and anticancer with cationic peptides,the anti-tumor properties of star-shaped PLO is further examined in the present study.In vitro investigations demonstrate that star-shaped PEI-g-PLO has strong lethality to tumor cells such as Hep G2 and A549.In order to reduce the cytotoxicity of star-shaped PLO to normal cells,anionic polymer PEI-g-PLO(DCA)is prepared by modifying the side chains of L-ornithine with1,2-dicarboxylic-cyclohexene anhydride(DCA).Under physiological p H conditions,PEI-g-PLO(DCA)exhibits minimal toxicity to normal cells,while charge reversion is triggered in the acidic micro-environment of tumor tissue,restoring positive charging state and killing tumor cells.In order to enhance the targeting effect of poly(L-ornithine)-based anti-tumor peptides,amphiphilic poly(L-ornithine)-b-poly(L-phenylalanine)(PLO-b-PLF)block copolypeptides are designed and synthesized.PLO-b-PLF self-assembles in an aqueous solution to form cationic micelles with a particle size of about 100 nm,a size that allows the PLO-b-PLF micelles to achieve passive targeting via the enhanced permeability and retention effect at the tumor site.In vitro studies show that PLO-b-PLF exhibits effective anti-cancer activity towards a variety of tumor cells,including drug-resistant tumor cells.The anionic block polypeptide PLO(DCA)-b-PLF is further prepared by decorating the side chains of L-ornithine with DCA.Low cytotoxic PLO(DCA)-b-PLF under physiological p H is converted into cationic PLO-b-PLF as a result of ?-amide bond fracture induced by the weakly acidic micro-environment in the tumor site,restoring the anti-cancer activity.The investigations on the anti-tumor mechanism show that PLO-b-PLF mainly targets the cell membrane of negatively charged tumor cells,binds tumor cells via electrostatic interactions,destroys the stability of cell membrane,and causes the rapid death of cancer cells.PLO-based cationic peptides kill tumor cells through physical action,which is expected to combat drug-resistant tumors and avoid the development of drug-resistant tumors.The cationic PLO-based anti-tumor peptides presented in this study provide an important reference for the emerging field of drug-free tumor treatment.In summary,star-shaped and block PLO are fabricated with the structural units of L-ornithine in the present study.The introduction of the unnatural L-ornithine significantly improves the enzymolytic stability of peptides and prolongs the circulation time in the body.PLOs can be used as antibacterial peptides for burn infections or as anti-cancer peptides for drug-free tumor treatment.PLOs destroy cell membranes of bacteria or cancer cells via physical action and thus can be employed to combat drug-resistant bacteria or drug-resistant tumors.This thesis demonstrates the great potential of PLOs as ideal biomedical materials.