Study On Antitumor Activity Of Hydroxycamptothecin Combined With Quercetin Nano-suspension In Vivo And Vitro

Chemotherapy is one of the most effective means of cancer treatment at present,while chemotherapeutic drugs generally have a common defect,which has great toxic and side effect and often prone to multi-drug resistance.Therefore,reducing the toxic and side effects of chemotherapeutic drugs while ensuring the efficacy has become a new direction in the field of tumor treatment.In order to solve the multi-drug resistance of tumor cells,the combination of drugs is usually used in clinic.In this context,HCPT and QUR were used as model drugs to prepare a nano-suspension.In this study,the combination of HCPT and QUR was screened by MTT assay,the cell activity of A549/PTX cells was studied when cultured in different drug ratios for 24 hours under the condition of controlling the same total dosage.The results showed that quercetin and HCPT had a good killing effect when the mass ratio of QUR to HCPT was 2:3.So the optimum ratio of QUR to HCPT was determined to be 2:3.Amphiphilic triblock copolymer PBLA-PEG-PBLA with different block ratios was prepared by ring-opening condensation of BLA-NCA with polyethylene glycol amine as initiator.The successful synthesis of block copolymer was confirmed by NMR and IR.The combined drug nano-suspension was prepared by micro-precipitation-high pressure homogenization method which Use the copolymer as stabilizer,HCPT and QUR as model drugs.The Optimum process were investigated by single factor method.The optimum conditions were determined as follows: 1:15 stabilizer 40 mg,QUR 8 mg,HCPT 12 mg,distilled water 40 mL,Homogeneous pressure: 250 bar 5 times,500 bar 5 times and 900 bar 25 times.HCPT-NPs,QUR-NPs and HQ-NPs were prepared according to the optimal process.The results showed that the average particle size was about 200 nm and PDI was about 0.2.The results of storage stability,dilution stability and plasma stability showed that the stability of the three nano-suspensions was relatively good under different conditions.DSC and XRD studies showed that the crystal form may have changed when HCPT and QUR were prepared into nano-suspension.In addition,it was confirmed by hemolysis experiment that the preparation of nano-suspension could be administered intravenously and could not produce hemolysis.The cytotoxicity of the blank material to normal hepatocyte HL-7702 was investigated by MTT assay.The results showed that the blank material had good biocompatibility and no obvious toxicity.Through the cytotoxicity of HCPT-NPs,QUR-NPs,HQ-NPs and combined free drugs on drug-resistant tumor cells A549/PTX and non-drug-resistant tumor cells HepG2 for 24 hours,it can be seen that the combination of drugs has a good killing effect on drug-resistant tumor cells.It is speculated that the better anti-tumor effect is that quercetin inhibits the overexpression of P-gp in drug-resistant tumor cells,thus reducing the excretion of drugs.In order to confirm this conjecture,the content of HCPT in A549/PTX was further determined after HQ-NPs and HCPT-NPs treated A549/PTX cells for the same time.It was found that the increase of hydroxycamptothecin intake in HQ-NPs group from 0.5h to 2h was about twice as much as that in HCPT-NPs group.Finally,through the changes of tumor volume in mice after administration,it can be seen that the effect of QUR-NPs is relatively weak.HCPT-NPs and HQ-NPs have a better effect on tumor,but there is no significant difference between the two.However,during the treatment of HCPT-NPs,the weight loss of mice was significant and the mortality rate of mice reached 50%,while no mice died in other groups.The results of thymus index,spleen index showed that The thymus index and spleen index in HCPT-NPs group were 4.31 ±0.29 and 1.38 ±0.09,respectively,which were lower than those in saline group(6.50 ±0.82 and 2.52 ±0.67,respectively).The thymus index and spleen index in HQ-NPs group were 7.25 ±0.82 and 3.07 ±0.55,respectively,which were higher than those in saline group.The results showed that HCPT-NPs group caused immune tissue injury and led to low spleen index,while HQ-NPs group had certain protective effect on immune organs,because quercetin had a certain protective effect on immune organs at the same dose.The H&E staining section also confirmed that HQ-NPs group reduced the tissue damage caused by drugs.In this study,hydroxycamptothecin was combined with quercetin to inhibit the overexpression of P-gp in drug-resistant tumor cells and reduce the excretion of hydroxycamptothecin,so as to achieve a better anti-tumor effect.In addition,quercetin can play a certain protective effect on tissues and organs,and the combination of quercetin and quercetin can not only achieve a better anti-tumor effect,but also reduce the toxicity and side effects of the drug.