| Background:Distinct to the western countries,the incidence of rectal cancer in China is higher than that of colon cancer.About 60 to 75% of rectal cancer is low rectal cancer.Most cancers can be touched during directeral rectun examination.In addition,young people under the age of30 have a high proportion of rectal cancer,which is up to 10%-15%.Because a considerable part of the patients are already in the advanced stage at the time of consultation,the surgical trauma is large,the urination function and sexual function are obviously impaired,and the rate of permanent stoma and local recurrence is high.Neoadjuvant chemoradiotherapy(n CRT)therapy for rectal cancer can reduce tumor volume,increase the safety margin of surgery,reduce surgical trauma,increase the rate of anus preservation,and reduce local recurrence.However,a lot of rectal cancers are not sensitive to n CRT,and even tumor progression occurs during preoperative radiotherapy and chemotherapy.However,there are no reliable indicators that can be used to clinically predict the radiation sensitivity of rectal cancer.Therefore,it is important to find a new radiosensitivity biomarkers for the individualized treatment of patients with rectal cancer.The purpose of this study is to determine the impact of postoperative pathological tumor regression grading(TRG)on subsequent treatment and prognosis of patients through the analysis of clinical data,and to search for biomarkers related to radiotherapy sensitivity by proteomics to achieve pre-n CRT predict of radiosensitivity to rectal cancer.Methods:This is a retrospective cohort study.Inclusion criteria: rectal adenocarcinoma confirmed by pathology before n CRT without distant metastasis;MRI diagnosed as low-medium rectal cancer before n CRT,clinical staging of the tumor was c T3 ? 4N + and c T2N0 stage of n CRT for anal preservation(3)Total mesorectal excision(TME)was performed after n CRT was completed.Exclusion criteria:(1)short-course radiotherapy before surgery;recurrent rectal cancer or other tumors history;(3)the interval from the end of radiotherapy to surgery is longer than 20 weeks.According to the above criteria,from January 2010 to June 2018,377 patients enrolled in our hospital who underwent n CRT and TME surgery were collected,of which 15 had distant metastases before surgery,and 3 of the remaining 362 patients had less than 2 months follow-up time and they were excluded,and the final 359 cases were analyzedfor prognosis.All patients received long-term intensity-modulated radiation therapy(IMRT),2 Gy 5 times a week,for a total of 50 Gy for 5 weeks;and capecitabine was administered orally at the same time;158 of them underwent CAPEOX or FOLFOX regimens after n CRT.All patients underwent surgery 4 to 20 weeks after n CRT.Follow-up to understand tumor recurrence,metastasis and survival,including disease-free survival(DFS),distant metastasis-free survival(DMFS),local recurrence-free survival(LRFS)and overall survival(OS),and radiotherapy sensitivity Logistic regression model was used for multivariate analysis,Kaplan-Meier method and Log-rank test were used for survival analysis,and Cox proportional hazard model was used for prognostic factor analysis.A total of 70 pre-n CRT adenocarcinoma tissue samples confirmed by pathological consultation were collected,and the DDA data acquisition mode was used in combination with the stable isotope labeling strategy(TMT10)to find biomarkers closely related to the sensitivity of rectal cancer radiotherapy.By controlling data quality,protein false positive rate and other indicators,a total of 5341 trusted proteins were quantified in the proteome data.HCA and PCA clustering methods were used to analyze the differences in protein between the radiosensitive group(TRG 0 + 1)and the radiotherapy resistant group(TRG 2 + 3),and the David website was used to perform biological function annotation and pathway enrichment of the differentially changed proteins.Use the pathview software package in R software to display the enriched part of the pathway.Results:Postoperative pathological TRG of 0,1,2,and 3 in 377 patients were 20.4%,17.2%,33.%,and 28.9%,respectively.Tumor circumference(P = 0.014),CEA before n CRT(P =0.006),degree of differentiation(P <0.001),c T(P = 0.030),and consolidation chemotherapy(P = 0.004)are related to TRG,and consolidation chemotherapy(OR = 1.854,95% CI: 1.052? 3.266,P = 0.033),and CEA elevation before n CRT(OR = 0.482,95% CI: 0.275 ? 0.847,P= 0.011)were independent influencing factors of TRG.The 5-year DFS(78.8% vs.64.0%,P= 0.005)of the patients in the TRG 0 + 1 group was significantly higher than that in the TRG2 + 3 group,and there was no significant difference in OS between the two groups(88.0% vs.73.9%,P = 0.098).Poor differentiation(HR = 1.873,95% CI: 1.111 ? 3.160,P = 0.019),yp T4(HR = 5.945,95CI: 1.121 ? 31.529,P = 0.036),yp N2(HR = 2.158,95CI: 1.119 ? 4.162,P =0.022)are independent risk factors for DFS.Poor differentiation(HR = 1.985,95% CI: 1.014? 3.866,P = 0.045),yp N2(HR = 3.835,95% CI: 1.665 ? 8.833,P = 0.002)?APR(HR=2.016,95%CI:1.045?3.888,P=0.036)are independent risk factors for OS,and TRG is not an independent prognostic factor of DFS and OS in patients.A total of 121 cases(90 cases in the postoperative chemotherapy group and 31 cases in the non-chemotherapy group)underwent pathological diagnosis of yp T0–2N0 after n CRT in the Low to middle locally advanced rectal cancer.There was no significant difference in DFS and OS between the two groups of patients at 5 years(OS: 79.1% vs.82.9%,P = 0.442;DFS:87.5% vs.78.2%,P = 0.667).COX multivariate analysis showed that c T4(HR = 4.563,95%CI: 1.630?12.771,P = 0.004)is an independent prognostic factor of DFS,and consolidation chemotherapy(HR = 0.253,95% CI: 0.064 ? 0.993,P = 0.049)is an independent protective factor of DFS.CT4(HR = 5.294,95% CI: 1.328 ? 21.105,P = 0.018)and anastomosis(HR =10.169,95% CI: 1.674 ? 61.788,P = 0.012)are independent risk factor of OS.The positive rate of lymph nodes in 76 patients with TRG 0(yp T0N0 ? 2)was 11.8%.The 5-year DFS and OS of the whole group were 86.2% and 94.7%,respectively.The serum CEA before n CRT(HR = 8.395,95% CI: 1.378 ? 51.151,P = 0.021)and yp N+(HR = 6.112,95% CI: 2.289 ? 91.209;P = 0.005)are independent risk factors of DFS,no independent prognostic factor for OS was found.In the proteomics study,there was no significant difference in baseline data between the TRG 0 + 1 and TRG2 + 3 patients.Unsupervised PCA and HCA clustering of the overall protein could not distinguish between the resistance group and the sensitive group,and then selected 19 proteins(P <0.05 and Fold change> 1.2)that were differentially expressed in the sensitive group and the resistance group,and the cluster analysis remained impossible to distinguish whether rectal cancer tissue is sensitive or resistant to radiotherapy.Consistent clustering analysis divided 70 samples into four subtypes.Unsupervised clustering analysis using differential proteins of each subtype can distinguish sensitive and resistant samples from radiotherapy in each subtype.Among the differential proteins,proteins with Fold change> 1.2 were selected.For subtype 1,46 proteins were up-regulated in the sensitive group and 56 proteins were down-regulated;for subtype 2,83 proteins were up-regulated in the sensitive group and 88 proteins were down-regulated;Thirteen proteins of type 3 were up-regulated in the sensitive group and 11 proteins were down-regulated;48 proteins of subtype 4 were up-regulated in the sensitive group and 43 proteins were down-regulated.Different subtypes have different protein enrichment pathways.See Appendix for differential proteins.Conclusions:1.Consolidation chemotherapy and elevated CEA before nc CRT are independent Prognostic factors of TRG after n CRT in rectal cancer.Preoperative consolidation chemotherapy can improve tumor response rate.2.The DFS in the TRG 0 + 1 group was significantly improved compared to the TRG 2+ 3 group.yp N2,yp T4,and poor differentiation were independent risk factors for DFS.yp N2?poor differentiation?APR were independent risk factors for OS,TRG is not an independent prognostic factor of DFS and OS.3.Patients with locally advanced rectal cancer with downstaging after n CRT(yp T0 ?2N0)did not significantly benefit from postoperative adjuvant chemotherapy.CT4 is an independent risk factor for DFS,and consolidation chemotherapy is an independent protective factor for DFS.c T4 and anastomotic leakage are independent risk factors for OS.4.Patients with complete remission of the primary intestinal wall after n CRT have a good prognosis.The yp N + rate is about 11.8%.CEA elevation and yp N + before n CRT are independent risk factors for DFS.5.Based on the proteomics data of pre-n CRT biopsy tissue,rectal cancer is divided into different promics subtypes.Each subtype can distinguish patients with sensitivity and resistance to radiotherapy,and different subtypes are enriched in differential proteins.Pathways are inconsistent,indicating that rectal cancer is highly heterogeneous.From this we reveal the significance of proteomics information of biopsy samples and subtype classification based on omics information for pre-resolution of radiosensitivity in patients with rectal cancer. |
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