Preoperative Evaluation For Colorectal Liver Metastasis-mircoRNA Maker And Clinicopathological Evaluation

BackgroundApproximately,15-25%of patients with colorectal cancer (CRC) are diagnosed with synchronous liver metastases at first diagnose, and50%of patients will develop metachronous liver metastases within first two years after the radical primary colorectal tumor resection.The outcome of patients with colorectal liver metastases (CRLM) have obviously improved due to comprehensive treatment, with5-year overall survival of50%. Several clinical prognostic models for resectable CRLM were proposed, however, the more precise preoperative evaluation remains to be explored. More than40%of suspected resectable CRLM are underwent the unnecessary laparotomy, due to insufficient preoperative evaluation. The issue how to enhance the accuracy of preoperative evaluation for resectable CRLM draws more attentions.Not only over-expression of oncogene and loss of anti-oncogene but also the epigenetics (such as methylation, acelylation, non-code RNA) are relation with CRLM. Systematic review of published studies showed the regulation of microRNAs played an important role in process of liver metastasis. A lot of microRNAs related with metastasis are identified and defined as "metastamiR".AimsThe microarray analysis was used to identify the microRNAs profile related with CRLM. The microRNAs screened by microarray analysis were validated by qRT-PCR using the formalin-fixed paraffin-embedded (FFPE) sample. Specially, the validated miR494was selected to identify the mechanism of liver metastasis in vivo through constructing stable transfect cell line.The meta-analysis was performed to confirm whether the PET-CT can provide additional information for preoperative evaluation in patients with suspected resectable CRLM. The SEER data was acquired to access the impact of lymph node on prognosis of patients with resectable metastatic colorectal cancer.Methods1. Screening microRNA makers correlated with CRLMThe total RNA from primary tumor frozen tissues of31pairs of colorectal patients with or without liver metastasis was abstracted. The different profiles of microRNA and mRNA between two groups were acquired by the Agilent microarray to identify the microRNA makers correlated with CRLM.2. Clinical validation of screened microRNAs correlated with CRLM Combining the result of microarray analysis and conclusions of published studies, seven microRNA(miR15b, miR19a, miR20a, miR181a, miR181d, miR223, miR494) were validated by quantity RT-PCR using FFEP samples of126patients with CRC. The impact of above listed microRNAs on the occurrence of liver metastasis and prognosis were analyzed.3. Molecular mechanism of liver metastasis induced by miR494in colonic cell lineThe cell line of Lovo, lowest expression of miR494, was screened by qRT-PCR. In vitro, stably over-expression miR494cell line (miR494-Lovo) was constructed to elucidate the function of miR494. The target gene of miR494was analyzed by the method of bioinformatics4. The value of status of lymph node used to stratify the CRLM-analysis based on SEER data.From SEER data, the information of2007patients with resectable metastatic CRC who received primary tumor and metastatic lesions resection was acquired. The impact of lymph node on prognosis was analyzed by univariate analysis and multivariate analysis.5. The additional value of PET in preoperative evaluation for patients with resectable CRLM-a meta-analysis.Out of213related studies screened,13studies were selected. The accuracy of PET in diagnosis with the hepatic lesion and extra-hepatic lesion, treatment change rate and unnecessary laparotomy rate was estimated to confirm the values of PET on preoperative evaluation for resectable CRLM ResultsThe analysis of microarray showed that16microRNAs had different expression between with and without liver metastasis groups. Furthermore, the upregulation of miR15b, miR223, miRl9a and down-regulation of miR494in liver metastatic group were validated by qRT-PCR using FFPE sample.As for colorectal cancer, high expression of miR494predicted poor prognosis. In vitro, MiR494can increase the ratio of S-phase and promote cell migration.The positive of lymph node, either the number of metastatic lymph node or the metastatic rate, was related with poor prognosis in resectable metastatic CRC.As for patients with resectable CRLM, the PET can improve the sensitivity of diagnosis in extra hepatic lesion and avoid unnecessary laparotomy.Conclusion1.miRl5b, miR223, miR19a, miR494are correlated with liver metastasis, furthermore, high expression of miR494can predict poor prognosis in CRC.2. miR494promotes Love cell line migration, and should be considered as a new makers to precisely classify the resectable CRLM.3. In clinical practice, both application of PET-CT and status of lymph node can precisely access the patients with resectable CRLM.