Preparation,Characterization And Anti-cancer Activity Evaluation Of L-K6 Antimicrobial Peptide-liposomal Drug System

Breast cancer,as a common malignancy of woman,is a serious threat to human health.Traditional treatment on breast cancer using chemotherapeutic drugs is easy to produce drug resistance,which results in poor clinical efficacy or even breast cancer treatment failure.Recently,the research on cationic antimicrobial peptide has indicated that these peptidic molecules also show anticancer activity with good targeted anticancer anctivity,and can overcome drug resistance,therefore are expected to become novel anti-cancer drugs.However,antimicrobial peptides in the body are susceptible to hydrolysis and unstable,which hindered their widely use as a drug in the clinic.To solve this issue,we tried to prepare a cationic antimicrobial peptide-liposomal drug system,and characterize its physical and chemical properties,stability and in vitro targeted anticancer activity.Firstly,the liposomes composed of different proportion of phospholipid component were prepared.bu using different proportion of phospholipid component By comparing the drug encapsulation efficiency,the best preparation procedure of L-K6 liposomes was determined that the ratio of S100 soybean lecithin:cholesterol is 2:1;Additionally,the L-K6 liposomes and PEG-modified long-circulating L-K6 liposomes were also characterized.Experimental results showed that the encapsulation efficiency of L-K6 liposome was(63.29 ± 2.13)%,the particle size was(116.4 ± 1.40)nm,Zeta potential value was(0.133 ± 0.21)mv;Interestingly,while the encapsulation efficiency of the PEG-modified long-circulating L-K6 liposome was similar with that of its non-modified liposomes(60.79±1.56)%,their liposome particle size was decreased to(108.9 ± 1.37)nm,Zeta potential value varied to(-5.04 ± 0.19)mv,suggesting that the PEG-modified liposomes showed a better system stability than the unmodified liposomes.Secondly,we use varies of in vitro experimental methods to investigate the anticancer effects of L-K6,L-K6 liposomes and PEG-modified long-circulating L-K6 liposomes on tumor.The results showed that L-K6,L-K6 liposomes and PEG-modified long-circulating L-K6 liposomes exerted similar anticancer effects to MCF-7 cells after 24h treatment,but did not show significant cytotoxicity against non-cancer HaCaT cells,indicating liposomal drug system preferably retained good cancer-targeting.Finally,the biological stability of drug-loaded liposomes was evaluated both in vitro and in vivo.Experimental results showed that,2 h after L-K6 incubation in serum in vitro,the drug content decreased to 61%(supposing the default drug content is 100%at time point 0h),suggesting that the peptide was susceptible to the degradation enzymes in serum;In contrast,2 h after incubation in serum in vitro,the drug content of long-circulating PEG-modified L-K6 antimicrobial peptide liposomes remained to be 94%,and even after 96h incubation,there was still 34%left,indicating a much better biological stability in vitro.The in vivo assay results further demonstrated that long-circulating PEG-modified L-K6 antimicrobial peptide liposomes have a better biological stability than unprotected L-K6.In summary,PEG-modified long-circulating L-K6 liposomal drug system and L-K6 can selectively bind to tumor cells,and exhibited similar anticancer effects in vitro.In addition,PEG-modified long-circulating L-K6 liposomes showed a better bio-stability both in vivo and in vitro compared to L-K6,and can effectively protect the degradation of L-K6 antimicrobial peptides.All these data provided an important evidence for the preparation and clinical application of antimicrobial peptides as new anticancer drugs in the future.