A Study On The Effect And Mechanisms Of Inhibiting Tumor Angiogenesis After Embolization In The Treatment Of Liver Cancer With Apatinib Loaded PIB Nanogels

?Objective?: Transcatheter arterial embolization(TAE)is playing a major role in the treatment of advanced liver cancer.However,there are still exist two defects to influence the long-term effect of TAE.On one hand,the angiogenesis of hepatocellular carcinoma(HCC)induced by hypoxia in tumor after TAE has been proved to play an important role in the malignant biological behavior of HCC,on the other hand,embolization of tumor peripheral blood vessels can also be a key factor affecting the curative effect of TAE.Our previous studies have showed that PIB has an excellent peripheral vascular embolization effect.Meanwhile,Apatinib mesylate,as a single-target inhibitor of vascular endotheliar growth factor receptor(VEGFR),can efficient inhibit tumor angiogenesis by targeting the VEGF/VEGFR pathway.Therefore,in this study we will investigate the treatment effect and mechanism of combining transcatheter arterial embolization with Apatinib-loaded PIB on suppressing rabbit VX2 liver tumor growth.And the effect of anti-tumor and anti-angiogenesis after embolization will be evaluated.A new targeted drug delivery system would be developed through this research,which will be meaningful to improve the interventional treatment effect of liver cancer.The mechanism of inhibition of VEGF/VEGFR pathway was elucidated,and antiangiogenic properties in anoxic environment or not and the role in the treatment of liver cancer will be explored as well.We hope that our effort could provide a new therapeutic strategy for interventional therapy of liver cancer.?Materials and Methods?: We simulated the tumor microenvironment before and after TAE both in vitro and in vivo.In vivo study,65 tumor-bearing rabbits were randomly divided into five groups and respectively treated transarterially with Apatinib-loaded PIB(Group PA)0.4 ml(n=13);PIB alone(Group P)0.4ml(n=13);iodized oil alone(Group I)0.4ml(n=13);Apatinib solution(Group A)0.4ml(n=13);saline(Group NS)0.4 ml(n=13).The dose of apatinib was 2 mg/kg.Then,an enhanced CT scan was performed 1 day before surgery and 7 days after surgery.The preoperative volume V1 and postoperative volume V7 were measured,and the tumor growth rates of each group were calculated.The survival time of the tumor-bearing animals was recorded.Tumor harvest,sectioning and Immuno-histochemistry on 7 days after surgery,and the expressions of microvessel density(MVD)were measured and analyzed in five groups of tumor at each time point.And in vitro experiment,HUVECs were cultured with different concentration gradients Apatinib(0,1,10,50?mol/L,24h)in the culture medium of Hep G2 with normal oxygen or hypoxia.The effect and mechanism of Apatinib was explored by bioassays,CCK-8 method was used to determine the effect of apatinib on HCC growth,the migration impact of Apatinib on human umbilical vein endothelial cells was detected by scratch test and Transwell assay in vitro testing.Apatinib in vitro angiogenesis on Matrigel-based tubele formation.In addition,the PI3K-AKT,Raf-MEK-ERK and P38 MAPK pathways were detected and verified by immunofluorescence.?Results?: The tumor growth rate of Group PA was significantly lower than the other four groups(P=0.000<0.01),and the survival time was significantly prolonged(P=0.000<0.01).The result was further confirmed by CCK-8 assays that Apatinib has the ability to inhibit the growth of HCC.According to the results of histopathological,we found that a large tumor necrosis area was observed after embolization.The result of immunohistochemistry showed that CD31 staining of Group PA was significantly lower than that of the other four groups(P=0.000<0.01).Further research into therapeutic mechanisms we found that Apatinib has a stronger inhibitory effect of multiplication,migration and invasion of HUVECs in anoxic environment rather than in normoxia and Apatinib inhibits HUVECs proliferation by down-regulating the PI3K-AKT,Raf-MEK-ERK and P38 MAPK pathways.?Conculsions ? : The Apatinib loaded PIB can be used as an effective liquid embolic material for the interventional treatment of VX2 liver cancer.Apatinib is targeted to liver cancer tissues through PIB,and achieves a slow release of the drug in the tumor.In hypoxic environments,Apatinib inhibits angiogenesis and migration of HCC after interventional embolization by inhibiting PI3K-AKT,Raf-MEK-ERK and P38 MAPK pathways,which in turn showed a significant prolonging survival time and inhibitory effect for tumor growth.This study is expected to provide new targets and therapeutic strategies for interventional treatment of HCC,and this way has a good application prospect.