Establishment Carrier Erythrocytes Drug Delivery Systems And Evaluation Of The Raltitrexed-loaded Erythrocytes In Vitro

Background:Raltitrexed is a thymidylate synthase inhibitor belonging to the antimetabolite class of cytotoxic drugs.It is effective in colorectal cancer(CRC)both as a single agent and in combination with other drugs,in particular in those patients with cardiologic risk factors or previous cardiotoxicity.Additional it is also widely used in esophagus cancer,gastric cancer,malignant pleural mesothelioma and so on.Drug delivery systems have significant clinical impact as they promise sustained systemic delivery and better targeting.It is one of the most effective ways to reduce side effect,decrease drug dose and improve drug efficacy.Carrier erythrocyte has a lot of advantages,such as biocompatibility,biodegradability,long-circulation time and so on.But most of the studies of carrier erythrocytes stay in preclinical because of the drug controlled release ability.Objectives:1.To demonstrate the feasibility of establishing carrier erythrocytes by the technology as described in the literatures,and to find the best drug loading condition and calculate the drug loading ratio.2.To evaluate the damage to the structure and function of erythrocyte membrane during drug loading.To evaluate the low temperature storage stability and sustained-release ability of raltitrexed-loaded erythrocyte.3.To demonstrate tumor cell proliferation inhibition of raltitrexed-loaded erythrocyte by human colorectal cancer cell SW480 in vitro.Method:1.Establishing carrier erythrocytes by a modified hypotonic pre-swelling technique as described in the literatures.Calculating the drug loading ratio by two different methods to find the best raltitrexed concentration and the optimal action time.2.Investigating the modified hypotonic pre-swelling technique by comparing the morphology and osmotic fragility of erythrocyte before and after loading drug.Investigating the low temperature storage stability of raltitrexed-loaded erythrocytes by testing the raltitrexed concentration of the supernatant and observing the erythrocyte morphology every day during raltitrexed-loaded erythrocytes were storaged in NS at 4?.Draw the "Time-Concentration" diagram to investage the sustained-release ability of raltitrexed-loaded erythrocytes.3.Cell proliferation inhibition was measured by MTT to compare the cytotoxicity of raltitrexed-loaded erythrocytes and free raltitrexed.Results:1.The modified hypotonic pre-swelling technique is a effective method to establish carrier erythrocytes and raltitrexed-loaded erythrocytes with the drug loading ratio approximate 69.81%.2.There are no obviously distinguish of morphology and osmotic fragility between carrier erythrocyte and raltitrexed-loaded erythrocyte.Carrier erythrocytes can release raltitrexed sustainably,and approaching 100%raltitrexed was released from carrier erythrocytes at 48h.3.At the same concentration,raltitrexed-loaded erythrocytes appeared better cell proliferation inhibition than free raltitrexed when incubated with SW480 for 24h and 48h.Conclusion:1.The modified hypotonic pre-swelling technique can effectively establish the raltitrexed-loaded erythrocytes with the drug loading ratio approximate 69.81%,and without any damage on the structure and function of the erythrocyte.2.Raltitrexed-loaded erythrocytes can be storaged in NS at 4? steady about 7 days,and raltitrexed can be released approaching 100%when raltitrexed-loaded erythrocytes were storaged in PBS at 37? for 48h.3.Raltitrexed-loaded erythrocytes appeared better cell proliferation inhibition than free raltitrexed on human colorectal cancer cell in vitro,and the cytotoxicity was increased with time longer.