The Antiangiogenic Effect Of Metronomic Chemotherapy With Paclitaxel(PTX) On Tumor-bearing Murine Model

Background and ObjectivesSurgical intervention, radiotherapy and chemotherapy are the basic approaches of tumor therapy. Since several decades, tumor therapy has been developed, especially in chemotherapy. A large amount of new drugs targeted on different receptors were applied in clinical therapy. Meanwhile, with the development of mechanism and pharmacokinetics about these drugs, more proper administration was used to kill tumor cells but protect normal tissue. Since the anti-angiogenic therapy was proposed, a new field has been paved. Because of its higher efficacy, lower toxicity, less resistance, so far, the anti-angiogenic therapy has been the focus. Targeted therapy of anti-angiogenesis was first proposed by Folkman in 1971. since then, researchers turned to the antiangiogenic effects of cytoxic drugs. Experiments had shown that some cytotoxic drugs inhibited angiogenesis effectively, especially when administered more frequently and at lower dosage. Compared to MTD chemotherapy with long rest interval between consecutive cycles, drugs given at lower doses and more frequently have stronger targeted effects on tumor vessel endothelial cells. This regimen was called "metronomic" dosing, or "antiangiogenic chemotherapy".Wang Yun, et al. found that tumor microvessel density (MVD) in Lewis lung carcinoma bearing C57BL/6 mice decreased obviously after metronomic chemotherapy with CTX, the intact tumor capsule formed between tumor tissue and normal tissue, infiltration of inflammatory cells and stromal reaction in metronomic group had significant difference with control group and routine chemotherapy. Why did the tumor tissue present these changes in metronomic group? We presumed that metronomic chemotherapy indirectly produced anti-angiogenesis by means of inhibiting localized inflammation.Macrophage is an important element of tumor stroma. Nowadays, papers report that lots of macrophages involved in tumor tissue contribute to tumor growth, angiogenesis, metrical formation and solution. NF-κB is an important molecule, bridging inflammation and tumor, involved in many physiological and pathologic courses by regulating gene transcription and playing a pivotal role in tumor angiogenesis.This experiment was presented on Lewis lung carcinoma bearing C57BL/6 mice aiming to evaluate the effect of paclitaxel as metronomic chemotherapy on angiogenesis and tumor growth of tumor bearing murine model, compare its efficacy and toxicity with MTD chemotherapy, and detect tumor microvessel density (MVD), macrophages(CD68) and the expression of NF-κB in metronomic group to demonstrate that whether metronomic chemotherapy produce indirectly anti-angiogenesis by means of inhibiting localized inflammation. Materials and MethodsC57BL/6 mice bearing Lewis lung carcinoma were randomised into three groups, and ten mice in each group:①metronomic group: administered Paclitaxel 15mg/kg intraperitoneally every other day;②MTD group: administered Paclitaxel 100mg/kg intraperitoneally in the first day, with a 21-day cycle;③control group: administered the normal saline 0,2ml intraperitoneally every other day. Paclitaxel was given from the 6^th day of post inoculation. The general status of mice was observed every day. At the 25^th day of post inoculation, all mice were killed. Then tumor growth and weight loss of mice were monitored to evaluate the efficacy and toxicity of different therapy. At the end of experiment, tumors were removed, weighed and immunohistochemistry was done. Tumor microvessel density (MVD),macrophages(CD68) and nuclear kappa-κB (NF-κB) were detected. The histological characteristics of the tumor were detected by microscopy. Statistical analysis was performed with SPSS10.0 software, using chi-square test, ANOVA and S-N-K test. Pearson correlation between MVD and macrophages was analyzed. The level of significance was set atα=0.05.Results1. The tumor growth curve in metronomic group is near flat and that in control group is very steep, however, no difference in the early stage. In MTD group, the tumor volume decrease in a short time after therapy, but rebounds during the rest interval.2. Mice in metronomic group have no weight loss and in good status. During the first week of the first cycle of chemotherapy, mice in MTD group suffered from side effects, such as apperant weight loss, sluggishness, hyporeflexia and losing skin luster. One mouse died at the 22^nd day of post inoculation. In MTD group, during the rest interval, mice weight rebound. Mice weight in control group increased steadily and decreased at the end because of heavy tumor burden. Two mice died at the 20^th and the 24^th day of post inoculation, respectively. At the end of experiment, the weight of tumor in control group, metronomic group I, metronomic group II and MTD group were (4.25±0.54)g,(1.88±0.42)g and (2.06±0.45)g, respectively.3. Observing the gross tumor samples, we found that the tumor capsule in control group and MTD group was broken and tumor was ruddy and crisp, while in metronomic group, the capsule was intact and tumor was pale and tenacious.4. The microvessel density (MVD) in control group, metronomic group and MTD group were 26.38±2.56,15.10±2.06 and 23.51±2.78 respectively. MVD in metronomic group was lower than that in control group and MTD group, and the difference was statistically significant (P<0.05). But the difference of MVD between MTD group and control group was not statistically significant (P>0.05).5. The macrophage count in control group, metronomic group and MTD group were 50.78±3.72,35.47±4.26 and 48.05±3.86 respectively. MVD in metronomic group was lower than that in control group and MTD group, and the difference was statistically significant (P<0.05). But the difference of MVD between MTD group and control group was not statistically significant (P>0.05). MVD and macrophage count had positive correlation in metronomic group.The correlation coefficient was 0.953(p>0.05)6. Expression of NF-κB in metronomic group was lower than that in control group and MTD group, and the difference was statistically significant (P<0.05). But the difference of NF-κB between MTD group and control group was not statistically significant (P>0.05).Conclusions1. Metronomic chemotherapy with PTX can inhibit the growth of Lewis lung carcinoma effectively, and its efficacy is better than MTD chemotherapy.2. Metronomic chemotherapy with PTX has lower toxicity than MTD chemotherapy. It can improve the general status of the tumor-bearing mice.3. Metronomic chemotherapy with PTX significantly inhibits the angiogenesis of Lewis lung carcinoma.4. Expression of NF-κB decreased after metronomic chemotherapy with PTX, as well as the macrophage count, as may be attributed to the antiangiogenic effect of metronomic chemotherapy.