| Cancer is a main killer which threatens our human life. The traditionaltherapeutic methods consist of operation,chemotherapy,radiotherapy andintervention treatment, all of which are not perfect. Biological treatment to thetumor is more and more emphasized. Gene therapy to the tumor may becomeanother kind of promising treatment method. IL-12 has the strongest immunityagainst tumor in so many cytokines. But, effective dose of interleukin-12 (IL-12)to the tumor often causes serious toxic effects, or even lead to death, therefore,the clinical application of IL-12 was restricted. Gene therapy with IL-12 againsttumor may increase the concentration of IL-12 in local tumor to increase effectand reduce the concentration in system to abate toxic effects. However,according to our experiments and other investigations, it is also found that greattoxicity of IL-12 gene therapy still existed. So, this study is to treat Lewis lungcarcinoma with IL-12 gene therapy and chemotherapy combinatively, then toobserve the effects and toxic effects.Cyclophosphamide(CTX) is a widely used chemotherapeutic agent incancer therapy. The anti-tumor range of CTX is wide, and the cost of CTX ischeap. According to previous reports, CTX can exhibit immunomodulatoryeffects, which may play an important role in the anti-tumor responds;CTX caninduce apoptosis and may have antiangiogenic effect.In this experiment, adenoviruses carrying IL-12 gene was injected intotumor to transfect the cells of Lewis lung carcinoma, and combined withchemotherapy. Different therapy projects were designed. Not only observedwhether the combination treatment was able to increase effect but also observedwhether these two methods had synergistic effect.I. The effect of AdcmvIL-12 gene therapy to Lewis lung carcinoma wasgoodIn this study, 7x108pfu was chosen for IL-12 group. The growth speed oftumor treated in IL-12 group slowed down;survival date prolonged obviously.Obvious difference appeared compared with the control group (P<0.005);therewas no metastasis. Histologically, in early stage, there were a wide range ofhaemorrhage and necroses, plenty of lymphocytes and some neutrophilicgranulocytes infiltrated in the tumor, apoptosis cells increased and tumor MVDdecreased in the IL-12 group while compared to the control group. The aboveresults showed that IL-12 had strong anti-tumor effect.Because the function of IL-12 can only last 7-12 days and no better affectscan achieve in the second injection, we chose the combination therapy withCTX regime.II. Chemotherapy's action to Lewis lung carcinomaThe role of cell immunity is T cell immunity. The ratio of CD4 and CD8may reflect the state of cell immunity of the body in some degree. In ourresearch a single infection of different dose of CTX resulted in the decrease ofspleen weight. The rates of CD4, CD8 and CD4/CD8 decreased significantly in200mg/kg group or higher dose group. We chose the dose of CTX lower than200mg/kg in case it might greatly affected the immune function. Also the lowdose of CTX could improve the immune function of mice.After continuous low dose of CTX injection in mice, the survival rateobviously prolonged and the tumor size was smaller when compared withcontrol group (P<0.05), which showed that continuous low dose regimen ofCTX therapy did have effect to Lewis lung carcinoma. While in aspect ofsurvival date and tumor size, a single injection group had no difference fromcontrol group and the curative effect was obviously poorer than low dose group.Metastasis was found in lung in two groups. It showed from our experiment thata single high dose of CTX could inhibit the tumor growth in earlier period ,butsoon the tumors grew rapidly. Tumor growth was inhibited for a long time afterhigh time low dose of CTX injection and this might be explained by its affect ofantiangiogenesis.Continuous low dose of CTX injection had better anti-tumor effect thansingle infection group, the reason might be the follows: (1) inducing secretion ofIFN-γ, improving the function of body immunity;(2) inducing apoptosis toinhibit tumor;(3) inhibiting angiogenesis of the tumor;(4) low dose of CTX haslittle toxicity, so it could prolong the survival date and improve the quality oflife. Tumor volume had the trend of decreasing in the dose of 40mg/kgcompared to 20mg/kg group, so we chose the dose 40mg/kg of CTX.The continuous low dose regimen of CTX could significantly increase thetherapeutic activity with decreased toxicity and prolonged animal survival forlung cancer. It might act as an antiangiogenic and lead to less drug resistanceand it might induce apoptosis, but the mechanism is not very clear now.III. Treat Lewis lung carcinoma with CTX and IL-12 gene therapycombinativelyThere were some reports on combination therapy with gene therapy andchemotherapy to treat tumor.Pan etal.found that IL-12 plasmid had better effectin inhibiting tumor growth when combined with CTX.But there is no report oncombination therapy with AdCMVIL-12 and CTX to treat Lewis lung cancer.In our experiment there were two combination groups,that werecombination group 1 (firstly injected 40mg/kg CTX for 4 days, the 5th dayinjected AdcmvIL-12 7x108pfu into tumor) and combination group 2(firstlyinjected 40mg/kg CTX for 4 days, the 5th day injected AdcmvIL-12 7x108pfuinto tumor, 8 days later injected 40mg/kg CTX for another 4 days).Incombination group the tumor size was much smaller and survival date prolongedobviously (P<0.005) than the single CTX group and single IL-12 group. We alsofound the combination group 2 was better than the combination group 1. Thetumor volume was smaller, the tumor growth was slower and the survival datewas much longer (P<0.05) in combination group 2 compared with combinationgroup 1. Besides, no toxic effects of combination therapy were seen and nometastasis was found in lung.The tumor volume was the smallest and the survival date was the longest incombination group compared to other group. In combination group the MVDwas significantly decreased and apoptosis tumor cells were the most, so thisgroup had the strongest effect of antiangiogenenisis and induction of apoptosis.The reason might be both AdCMVIL-12 and CTX had the effect of inhibitingangiogenesis and inducing apoptosis in the tumor site;Besides, low dose of CTXcould strengthen the immune function of the mice, promote the anti-tumorimmune effect of AdCMVIL-12 significantly, elevating the treatment effect.Our results showed that combination of AdCMVIL-12 and CTX had thebest effect of inhibiting Lewis lung cancer. It could reduce the dose of CTX sothat it could diminish the side effects of CTX. The combination regiment is anew strategy to tumor therapy and is worthy of our further research.Innovation and signification of this study:It is the first time to treat mouse's Lewis lung carcinoma withAdCMVIL-12 gene therapy and traditional chemotherapy drug CTXcombinatively, and we found the two methods have inter-promotion actions. Wefound for the first time that continuous low dose of CTX injection has betteranti-tumor effect compared to single infection group in Lewis lung carcinomatherapy through antiangiogenesis and inducing apoptosis.
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