Study Of The Therapeutic Effect Of ³²P-nucleic Acid Targeted Therapy On Transplanted Tumor Of H22 Mice

Objective: Hepatocellular carcinoma（HCC）is one of the most common malignancies in clinical practice,and its mortality rate ranks second in our country.Surgical resection is the first choice for the treatment of early stage hepatocellular carcinoma.Most patients are diagnosed with local progress or distant metastasis,which might be associated with chronic hepatitis and liver cirrhosis,poor liver function and common complications;this is why patients with HHC may not meet the criteria for surgery.The overall prognosis is still poor.Isotope based radiotherapy has unique advantages in the field of HCC therapy because of the broad indications,direct effect to lesions,long exposure time,good safety and repeatable treatment.The basic and clinical studies of 125 I,90Yi,32 P and so on in the treatment of HCC have shown a clear effect,there is a broad prospect of development.32 P is an ideal local radiation isotope.During decay,32 P produce Beta rays,which have a clear killing effect on tumor tissues;with 14.26 days of half-life,it can provide a low dose of continuous irradiation.The isotope also has short range;it causes less damage to normal tissue.The clinical application of ³²P-glass microspheres via hepatic artery interventional therapy for HCC has been studied,although the treatment has certain curative effect,the drugs are difficult to enter into tumor cells and losing efficacy.The interventional treatment with this therapy for HHC has strict indications with limited number of patients can be applied.Recent studies made by Kong Yanping have revealed that some cancer cells,but not nontumorigenic cells,can take DNA fragments directly from the environment without the aid of the transfecting reagent.Using this characteristic of hepatocarcinoma cells,we team made ³²P-labeled DNA fragments into a compound named ISONA,which essentially carries radioactive 32 P into cancer cells to achieve its targeted antitumor effect.Compared with the traditional isotope treatment of HCC,ISONA showed a wider indication,better targeting,more durable and better safety.The study will explore the effects of the ISONA on inhibition of H22 liver cancer growth in vivo,and explore the effects of ISONA on overall survival.This is to provide a theoretical basis for the clinical application of ISONA in the treatment HHC.Method:1 tumor inhibition experiment:Subcutaneous transplanted tumor model of H22 hepatoma was established in mice which were divided into four groups randomly: control group（group C）,ISONA high dose group（IH group）,low dose ISONA group（IL group）,free 32 P group（group P）.C group: saline 0.5ml viatail intravenous injection;IH group: ISONA solution 0.5ml with 90 microcurie viatail intravenous injection,;IL group: ISONA solution 0.5ml with 30 microcurie viatail intravenous injection;P group: free 32 P solution 0.5ml with 30 microcurie viatail intravenous injection.Tow dimension of the subcutaneous transplantation tumors were measured in every mouse at d0,d1,d3,d5 and d7 to calculate the tumor volume,tumor growth inhibition rate and growth curve.The mice body weight was measured at d0 and d7,and all mice were sacrificed after 7 days,stripping out the tumor and spleen to weigh respectively.The tumor’s inhibition rate and spleen index of mice was calculated.Spleen index was studied to evaluate the effect of different approaches on the immune organs.2 Survival experiment:Transplanted ascites tumor model of H22 hepatoma was established in mice and divided into four groups randomly: control group（group C）,ISONA group（group I）,free ³²P（group P）.C group : saline 0.5ml via intraperitoneal injection 0.5ml;I group: ISONA solution 0.5ml with 100 microcurie via intraperitoneal injection;P group: free 32 P solution 0.5ml with 100 microcurie via intraperitoneal injection.Animal’s death was recorded to observe survival.3 Statistical analysis:SPSS21.0 statistical software was used to analyze measurement data.The average number of each group was compared with sing ANOVA test.Kaplan-Meier method was used for survival analysis.P<0.05 is statistically significant.Result:1 Tumor inhibition experiment:1)The subcutaneous transplanted tumor model of H22 hepatoma formation rate was 100%.2)Weight changes: there was no significant difference in the body weight of each group before treatment（P>0.05）.7 days after treatment,the weight in C,IH,IL,P group was:（37.64+2.06）g,（28.97+2.06）g,（32.73+2.11）g,（36.52+2.34）g,the variance analysis showed that the IH group and C group,IL group and C group are statistically significant（P<0.05）,no statistical significance between group C and group P（P>0.05）.3)The analysis of variance showed no significant difference in tumor volume groups before treatment.7 days after different treatment,the volume of subcutaneous transplanted tumor in group C,IH,IL,P was:（4420+494）mm3,（2187+203）mm3,（3624+449）mm3,（4270+376）mm3.Variance analysis showed that the tumor volume of group IH,IL group was lower than that of C group with significant difference（P<0.05）,the tumor volume of group IH is lower than group IL（P<0.05）,the difference between P group and C group had no statistical significance（P>0.05）.4)No significant difference in tumor weight among groups before treatment.7 days after treatment,the weight of subcutaneous transplanted tumor in group C,IH,IL,P was as follows:（6.236+0.838）g,（3.053+0.451）g,（5.172+0.607）g,（6.027+0.798）g.The tumor weight of IH and IL group was lower than that of C group with significant difference（P <0.05）,the tumor weight of group IH is lower than group IL（P<0.05）,the difference between P group and C group had no statistical significance（P>0.05）.The tumor inhibition rate was calculated according to the tumors’ weight: IH group51.4%;IL group: 17.6%;P group: 3.4%.5)Spleen index: spleen index of group P was lower than that of group C（P<0.05）,IH group and IL group had no significant difference with C group（P>0.05）.2 Survival experiment: The average survival time of mice in C group,I group,P group was（9.30±4.79）days,（15.00 ±6.71）days,（8.20±4.37 days）.The life extension rate of I group and P group mice was 61.29% and-11.83%.The mean survival time of I group was higher than that of C group（P<0.05）.The mean survival time of P group was not significantly different from that of C group（P>0.05）.Conclusion: ISONA can effectively inhibit the growth of tumor and prolong the survival time of tumor-bearing mice and improve the quality of life of mice,and free 32 P has some immunosuppressive effects on tumorbearing mice.