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Inhibitory effect of CDK2 gene silencing combined with dacarbazine on the growth of B16-F1 melanoma / 中华皮肤科杂志
Chinese Journal of Dermatology ; (12): 658-663, 2017.
Article in Chinese | WPRIM | ID: wpr-607542
ABSTRACT
Objective To evaluate the antitumor effect of dacarbazine (DTIC) on B16-F1 melanoma after CDK2 gene silencing.Methods Cultured B16-F1 melanoma cells were divided into 4 groupscontrol group receiving no treatment,CDK2-shRNA group infected with a recombinant lentivirus pUL-CDK2-shRNA,DTIC group cultured in 96-well plates followed 12 hours later by the treatment with 250 μmol/L DTIC,CDK2-shRNA + DTIC group infected with pUL-CDK2-shRNA followed 12 hours later by the treatment with 250 μmol/L DTIC.MTT assay was performed to evaluate the growth inhibition of B16-F1 melanoma cells,and coefficient of drug interaction (CDI) was calculated.AnnexinV-FITC/PI double staining was conducted to detect cell apoptosis.C57BL/6 mice were subcutaneously injected with B16-F1 cells at exponential growth phase into the right groin to establish melanoma-bearing mouse models.Twenty mouse models were randomly and equally divided into 4 groupscontrol mouse group injected with phosphate-buffered solution (PBS) into tumors,CDK2-shRNA mouse group injected with pUL-CDK2-shRNA into tumors,DTIC mouse group injected with DTIC into the abdominal cavity,and CDK2-shRNA + DTIC mouse group treated with pUL-CDK2-shRNA and DTIC.The animal experiment lasted 18 days,and the tumor growth curve was drawn.After 18-day treatment,all the mice were sacrificed,and tumors were isolated and weighed.The tumor growth inhibition rate was calculated,and the tumor cell apoptosis was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick-end-labeling (TUNEL).Results After 72-hour culture,compared with the control group,the CDK2-shRNA group,DTIC group,and CDK2-shRNA + DTIC group showed significantly decreased relative cell survival rates (40.6% ± 2.8%,45.2% ± 3.7%,28.7% ± 2.1%,respectively;F =458.04,P < 0.05),but significantly increased cell apoptosis rates (25.1% ± 3.3%,15.6% ± 2.2%,45.6% ± 3.5%,respectively;F =115.46,P < 0.05).Additionally,CDK2-shRNA + DTIC group showed significantly lower relative cell survival rates (P < 0.01),but higher cell apoptosis rates (P < 0.01) compared with the DTIC group.The CDI value was less than 0.7.On the sixth day after the in vivo treatment,the tumor volumes in the control mouse group,CDK2-shRNA mouse group,DTIC mouse group,and CDK2-shRNA + DTIC mouse group were (185.44 ± 68.97) mm3,(83.91 ± 14.33) mm3,(123.70 ± 20.85) mm3,and (34.54 ± 10.72) mm3 respectively.From then on,the CDK2-shRNA mouse group,DTIC mouse group,and CDK2-shRNA + DTIC mouse group showed significantly decreased tumor growth rates compared with the control mouse group (F =11.819,P < 0.05),and the tumor growth rate was significantly lower in the CDK2-shRNA + DTIC mouse group than in the DTIC mouse group (P =0.04).The calculated tumor growth inhibition rates in the CDK2-shRNA mouse group,DTIC mouse group and CDK2-shRNA + DTIC mouse group were 52.2%,41.2% and 86.4% respectively.Compared with the control mouse group,the CDK2-shRNA mouse group,DTIC mouse group,and CDK2-shRNA + DTIC mouse group showed significantly increased tumor cell apoptosis indice (32.93% ± 3.72%,21.62% ± 3.54%,63.29% ± 4.74% respectively;F =222.25,P < 0.05).Moreover,the tumor cell apoptosis index was significantly higher in the CDK2-shRNA + DTIC mouse group than in the DTIC mouse group (P < 0.01).Conclusion CDK2 gene silencing can enhance the inhibitory effect of DTIC on the growth of melanoma,and show a synergistic effect with DTIC,likely by increasing the apoptosis of tumor cells.

Full text: Available Index: WPRIM (Western Pacific) Language: Chinese Journal: Chinese Journal of Dermatology Year: 2017 Type: Article

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Full text: Available Index: WPRIM (Western Pacific) Language: Chinese Journal: Chinese Journal of Dermatology Year: 2017 Type: Article