| 【Objective】There are several characteristics in difficulty operation, postoperativecontinence function, high local recurrence rate in advanced rectal cancer, and they havebecome the key and difficult problems in the research field of colorectal surgery in recentyears. Neoadjuvant therapy, also namely the preoperative chemoradiotherapy, isrecommended by international organization as a valid scheme which can significantlyimprove the prognosis of patients with advanced rectal cancer, reduce tumor staging, increasethe anal preservation rate and decrease the local recurrence rate. However, resistance tochemoradiotherapy is difficult and individual sensitivity is relatively difference,chemoradio-resistance is a troublesome problem to clinical treatment, so it is very necessaryto develop the ability to accurately assess the molecular markers in response tochemoradiotherapy, multidisciplinary collaboration on the development of morecomprehensive treatment targets and individualized programs, avoid invalid over treatmentand reduce adverse reaction, in order to further improve the long term prognosis in patientswith advanced rectal cancer. In this study, we aims to clarify the role of multidrugresistance-associated protein3(MRP3) in resistance to neoadjuvant chemoradiotherapy andlong-term prognosis of advanced rectal cancer.【Methods】To retrospectively compare differences of operation time, amount ofbleeding, resection rate, sphincter preserving rate and incidence of postoperativecomplications respectively by analyzing clinicopathologic data of T3and T4patients withmid-low rectal carcinomas in department of colorectal surgery from January2009toDecember2010(experimental group:81cases were treated with preoperativeradiochemotherapy plus operation; contrast group:93cases only underwent operation withoutpreoperative therapy). High-throughput lncRNA/mRNA chip is used to screen lncRNA geneand protein coding gene, whose expressive difference is more than2folds between RKO、LOVO cell lines and RKO cell line receiving2Gy radiation. The main action pathway iscomputed by Gene Ontology Analysis combined with Pathway Analysis in order to explorethe mechanism which induces the effect on radiosensitivity of colorectal carcinoma cell lines.Further experiment on gene expression contents of RKO cell line is confirmed by RealtimePCR. To clarify the relationship between MRP3expression and chemoradiotherapy sensitivityin advanced rectal cancer patients, retrospective analysis of neoadjuvant chemoradiotherapywith clinical and pathological data. Immunohistochemistry was used to measure MRP3expression in biopsy specimens of144stage II–III rectal cancer patients who received preoperative chemoradiotherapy, and evaluate the neoadjuvant treatment tumor pathologicalretraction degree in accordance with the5scores tumor regression grade standards,meanwhile analysis of11clinical pathological factors which may affect the efficacy ofneoadjuvant therapy. The effect of MRP3expression on short-term pathological response andpostoperative long-term prognosis were assessed using the Cox proportional hazards model.Short interfering RNAs targeting MRP3were synthesized and used to transfect humancolorectal carcinoma cell lines. After intervention with graded doses of radiation andchemotherapy drugs, colony forming cell proliferation assay and SF=1-(1-e-D/D0) Nsurvival curve calculate a series of radiation sensitive parameters, flow cytometry andHoechst33342staining was used to detect cell apoptosis, cell cycle arrest function; The effectof MRP3down-regulation on cell proliferation and apoptosis in response to5-fluorouraciland/or irradiation were examined in vitro and in xenograft mouse model, respectively. Furthermechanism research analysis of MRP3in apoptosis signal pathway activation and effectormolecule expression condition in resistance to chemoradiotherapy in advanced rectal cancer.The content of intracellular reactive oxygen species and the activity of caspase-3-dependentapoptotic pathway in response to irradiation were further evaluated.【Results】Both resection rate and sphincter preserving rate are higher in theexperimental group (100%,86.4%) than those in the contrast group (94.6%,73.1%), whilethe difference of sphincter preserving rate is statistically significant（P=0.039）; there are nosignificant differences in the mean operation time (130±15min vs125±20min, P>0.05) andmean amount of bleeding (100±15ml vs95±10ml, P>0.05) between the two groups. Theoverall incidence of postoperative complications is similar (9.9%vs9.7%, P>0.05),meanwhile there is no significant difference of anastomotic leakage rate in both groups (5.7%vs5.9%, P>0.05). High-throughput lncRNA/mRNA chip help us get a total of268lncRNAgenes and770protein coding genes. High expression (immunoreactive score>6) of MRP3significantly predicted poor pathological response to chemoradiotherapy (tumor regressiongrade≤2vs.≥3, p=0.002) in univariate analysis and unfavorable long-term prognosis (5-yearoverall survival: HR=1.612,95%CI,1.094to2.375, p=0.016;5-year disease-free survival:HR=1.513,95%CI,1.041to2.200, p=0.030) in multivariate Cox analysis. MRP3down-regulation significantly increased5-fluorouracil or irradiation-induced cell apoptosisand attenuated tumor growth following irradiation in animal models. MRP3inhibitionsignificantly reduced intracellular reactive oxygen species exporting from cells followingirradiation, and increased expression of cleaved poly ADP-ribose polymerase and caspase-3. 【Conclusion】Preoperative radiochemotherapy could significantly raise sphincterpreserving rate of mid-low rectal carcinomas, meanwhile does not increase the difficulty ofsurgical procedure and the postoperative complications.Aberrant expression of MRP3inrectal cancer confers chemo-radioresistance. MRP3might be a predictive factor and anattractive target in treating advanced rectal cancer. MRP3can be used as an effectivemolecular marker for prospectively screening patients, to provide a theoretical basis for thenext step application of its translational medicine. |
PDF Full Text Request