Experimental Study Of β1-Integrin Antisense Oligonucleotide In Combination With Cisplatin On Implanted Human Ovarian Cancer In Nude Mice

Objective: To observe the effect and safety of β1-integrin antisense oligonucleotideantiangiogenic in combination with cisplatin on implanted human ovarian cancer innude mice.Reveal the mechanism of inhibiting tumor cell adhension andtransfer,providing a new ideas and methods to treatment patients of ovarian cancer drugresistance.Methods:1.Established nude mice xenograft model and Experiment groups:Established the nude mice subcutaneous xenograft model of transplantation SKOV3tumor of Human ovarian cancer.The32bearing-tumor mice were randomly divided intofour groups as follows: Antisense oligonucleotide group (ASODN group)；Antisenseoligonucleotide conbined with the cisplatin group (A+D group)； Cisplatingroup(CDDP group)；Physiological saline control group(NS group).Drug treatmentswere started after randomization. The drug dosage and administration as follows:Theβ1-integrin ASODN complexed with cationic liposome (1mg/kg/d1,3,6,9,12,15,18,21)and Cisplatin（0.8mg/kg/d1,3,6,9,12,15,18,21）were injected intra-tumorally.2.Theanalysis of efficacy and security: During the experiment period, dynamic measurementmice tumor size, tumor growth curve drawing and observed the tumor-bearing mice. Inthe end time after three days to medicine with cervical death dislocated tumor-burdenedrat and take method, weighing the tumor weight and calculation tumor-suppressing ratesto observe the inhibitory effects of β1-integrin antisense oligonucleotide on implantedhuman ovarian cancer in nude mice. Adopt RT-PCR method to detect beta1integrinmRNA expression on the subcutaneous transplant tumor. Using immunohistochemistryelivision method to detect beta1integrin protein expression on the subcutaneoustransplant tumors. Take each tumor-burdened rat liver and spleen, lungs, kidneys do HEdyed, observed pathology change and tumor organ metastases.Results:1.This studythrough subcutaneous vaccination SKOV3cell levitation liquid can be successful construction nude mice subcutaneous xenograft model.2.β1-integrin ASOND can inhibitthe growth of tumor tissue transplant, the time dependence, opting for treatment of the21st, A+D group transplant tumor growth inhibition rate as high as70.37%. Thetreatment group were significantly less than tumor volume control of saline solution,ASODN group (316.10±21.77mm~3), A+D group (178.70±40.67mm~3), CDDP group(348.52±73.30mm~3), are below NS group (613.17±53.53mm~3)(P <0.05), A+Dgroup of tumor size minimum (P <0.05), ASODN group compared with the group ofCDDP no significant difference (P>0.05). After treatment, stripping tumor block andweighing, ASODN group (0.32±0.07) and A+D group (0.56±0.06) compared withthe NS group(1.08±0.09), was statistically significant difference.(P <0.05).3.RT-PCRdetection four groups transplant tumor tissue in both β1-integrin mRNA expressions,A+D group (0.4636±0.06), ASODN group (0.5765±0.05) and CDDP group (1.2136±0.13) were lower than NS group (1.3247±0.06), especially with A+D group. Themost significant differences between the each group (P <0.05). ASODN group andA+D group compared with NS group have statistically significant difference (P <0.01),A+D group compared with ASODN groups also have statistically significant (P <0.05).4.Immunohistochemical staining to detect four groups are β1-integrin proteinexpression, it’s mainly located in cell membranes.A+D group（0.82±0.11）and ASODNgroup (1.83±0.80) efficiently visible cell dyeing as incomplete membrane expression,the dyeing integral obviously lower NS group（s3.45±0.15）and CDDP group (3.31±0.16)(P﹤0.01), especially with A+D group. The most significant differences between theeach group(P <0.05), A+D group compared with the ASODN group also havestatistically significant compared (P <0.05), the group of CDDP and the group of NSare both complete cell membrane expression and positive rate of100%,the differencewas no statistically significant (P﹥0.05).5.HE stain reveals: the scattered tumor cellswas found in the spleen of nude mice, the more visceral interstitial vascular milddilatation and congestion.Conclusion: ASODN can inhibits the transplant tumor growthof human ovarian cancer implanted in nude mice, and combined with cisplatin havechemotherapy synergy. β1-integrin ASODN through the cut target genes β1-integrinmRNA expressions of reducing β1-integrin protein expression, thus inhibiting tumor cell growth.