The Role And Mechanism Of EIF3a In Colorectal Cancer Metastasis And Irinotecan Resistance

Background: The majority of colorectal cancer-Related deaths are characterized by metastasis,which is the most important focus of clinical treatment of colorectal cancer.For metastasis to occur,malignant cells must first undergo coordinated processes to acquire a pre-migratory phenotype.These processes include cell polarity,cytoskeletal reorganization,cell adhesion alteration,invasive structure formation and surrounding stromal degradation.In addition to the motility of tumor cells,tumor microenvironment also plays a crucial role in tumor metastasis.Cancer occurs and develops in the complex tissue environment,and they depend on the environment for continuous growth,invasion and metastasis.Targeting the processes of aggressive phenotypic transformation and the interaction between tumor cell and microenvironment would effectively inhibit colorectal cancer metastasis.Irinotecan has been approved for the first-line treatment of metastatic colorectal cancer,the application of which greatly improved the clinical therapeutic effect of patients with metastatic disease.However,widespread resistance and individual differences extensively limit the efficiency of chemotherapy medications.There is an urgent need to find more effective markers to identify patients who are more likely to benefit from the treatment and to adopt more tailored approaches to predict and optimize outcomes.Eukaryotic translation initiation factor 3a(eIF3a)is the largest subunit of e IF3,which is widely recognized as a proto-Oncogene correlated with tumor prognosis,occurrence,metastasis and therapeutic response.With the increasing understanding of the physiological and pathological functions of e IF3 a,it is gradually considered as a new potential anticancer target.The role and regulatory mechanism of e IF3 a in colorectal cancer metastasis and irinotecan resistance may provide new and more effective markers for the treatment of metastatic colorectal cancer,which will contribute to identify patients who are more likely to benefit from clinical treatment in advance,and to adopt a more tailored approach to predict and optimize the therapeutic effect.Objective: To investigate the role and molecular mechanism of e IF3 a in the regulation of colorectal cancer metastasis and irinotecan resistance at clinical,animal and cellular levels.Methods: The database analyses,immunohistochemistry and RTPCR assays were used to reveal the clinical characteristics of e IF3 a as well as its role in colorectal cancer metastasis at clinical level.The effects of e IF3 a on the invasion,metastasis,proliferation and irinotecan sensitivity of colorectal cancer cells were investigated in vivo by constructing lung metastasis model,liver metastasis model and subcutaneous transplanted tumor model.The regulatory role of e IF3 a on tumor cell migration,invasion and proliferation was studied in vitro by performing wound healing,Transwell,EDU,CCK8 cell viability and colony formation assay.The immunofluorescence and electron microscopy were conducted to observe the pseudopodia formation and cytoskeleton reorganization.The interaction between tumor cells and vascular endothelial cells was studied by co-culture experiments and tube formation analysis.The CCK8 assay,flow cytometry,TUNEL staining and comet assay were used to investigate the sensitivity and DNA damage response of colorectal cancer cells exposed to irinotecan.Western blot,RT-PCR,RIP,Co-IP and luciferase reporter assays were used to reveal the molecular regulatory mechanism of e IF3 a on downstream genes.Results:(1)We first investigated the clinical significance of e IF3 a and observed an abnormal expression of e IF3 a in colorectal tumor tissue.Patients with high e IF3 a expression are more likely to develop metastasis and have a poor prognosis.(2)Phenotypic experiments revealed that downregulation of e IF3 a significantly inhibited the motility,migration,proliferation and irinotecan sensitivity of tumor cells in vitro and in vivo.(3)Mechanism studies suggested that e IF3 a can not only promote the transformation of colorectal cancer cells to an invasiveness phenotype,but also accelerate tumor metastasis by promoting angiogenesis in the tumor microenvironment.More detailed,e IF3 a translationally activates the expression of CDC42 and RHOA,thus promoting pseudopodia formation and cytoskeleton reorganization of colorectal cancer cells.Besides,e IF3 a also promotes the migration,invasion,proliferation and tube formation abilities of HUVEC cells by regulating YAP/TAZ signal as well as the secretion of ANG,CCL5,PDGF-BB,PLGF and ANGPT2 of colorectal cancer cells,thus promoting angiogenesis and creating a suitable microenvironment for metastasis to occur.(4)The mechanic study of e IF3 a regulation of irinotecan sensitivity of colorectal cancer cells showed that e IF3 a continuously activates ATM/ATR signal by translationally inhibiting PPP2R5 A,a phosphatase that directly dephosphorylates and inactivates ATM/ATR after DNA repair complete.Suppression of PPP2R5 A resulted in chronic ATM/ATR phosphorylation and activation,impairing DNA repair and enhancing irinotecan sensitivity.Conclusions: e IF3 a plays a vital role in the metastasis and irinotecan resistance of colorectal cancer.e IF3 a promotes tumor metastasis not only by accelerating the acquisition of migratory phenotype of colorectal cancer cells,but also promotes angiogenesis in the tumor microenvironment which create a suitable microenvironment for metastasis to occur.Except these,e IF3 a is also involved in irinotecan-induced DNA damage response,which may finally contribute to the treatment outcomes of metastatic colorectal cancer patients.