Anus preservation in low rectal adenocarcinoma based on MMR/MSI status (APRAM): a study protocol for a randomised, controlled, open-label, multicentre phase III trial.

BACKGROUND: Anus preservation has been a challenge in the treatment of patients with low rectal adenocarcinoma (within 5 cm from the anal verge) because it is difficult to spare the anus with its functioning sphincter complex under the safe margin of tumour resection. Patients with dMMR/MSI-H can achieve a favourable complete response (CR) rate by using a single immune checkpoint inhibitor. For patients with pMMR/MSS/MSI-L, intensified neoadjuvant three-drug chemotherapy may be the preferred option for anal preservation. In addition, the watch and wait (W&W) strategy has been proven safe and feasible for patients with rectal cancer who achieve a clinical complete response (cCR). Therefore, we initiated this clinical trial to explore the optimal neoadjuvant treatment pattern for patients with low locally advanced rectal cancer (LARC) with different MMR/MSI statuses, aiming to achieve a higher cCR rate with the W&W strategy and ultimately provide more patients with a chance of anus preservation. METHODS: This is a randomised, controlled, open-label, multicentre phase III trial. Patients with clinical stage T2-4 and/or N + tumours located within 5 cm from the anal verge are considered eligible. Based on the results of pathological biopsy, the patients are divided into two groups: dMMR/MSI-H and pMMR/MSS. Patients in the dMMR/MSI-H group will be randomly allocated in a 1:1 ratio to either arm A (monoimmunotherapy) or arm B (short-course radiotherapy followed by monoimmunotherapy). Patients in the pMMR/MSS group will be initially treated with long-term pelvic radiation with concurrent capecitabine combined with irinotecan. Two weeks after the completion of chemoradiotherapy (CRT), the patients will be randomly allocated in a 1:1 ratio to arm C (XELIRI six cycle regime) or arm D (FOLFIRINOX nine cycle regime). The irinotecan dose will be adjusted according to the UGT1A1-genotype. After treatment, a comprehensive assessment will be performed to determine whether a cCR has been achieved. If achieved, the W&W strategy will be adopted; otherwise, total mesorectal excision (TME) will be performed. The primary endpoint is cCR with the maintenance of 12 months at least, determined using digital rectal examination, endoscopy, and rectal MRI or PET/CT as a supplementary method. DISCUSSION: APRAM will explore the best anus preservation model for low LARC, combining the strategies of consolidation chemotherapy, immunotherapy, and short-course radiotherapy, and aims to preserve the anus of more patients using W&W. Our study provides an accurate individual treatment mode based on the MMR/MSI status for patients with low LARC, and more patients will receive the opportunity for anus preservation under our therapeutic strategy, which would transform into long-term benefits. TRIAL REGISTRATION: Clinicaltrials.gov NCT05669092 (Registered 28th Nov 2022).

Development and external validation of MRI-based RAS mutation status prediction model for liver metastases of colorectal cancer.

BACKGROUND: The mutation status of rat sarcoma viral oncogene homolog (RAS) has prognostic significance and serves as a key predictive biomarker for the effectiveness of antiepidermal growth factor receptor (EGFR) therapy. However, there remains a lack of effective models for predicting RAS mutation status in colorectal liver metastases (CRLMs). This study aimed to construct and validate a diagnostic model for predicting RAS mutation status among patients undergoing hepatic resection for CRLMs. METHODS: A diagnostic multivariate prediction model was developed and validated in patients with CRLMs who had undergone hepatectomy between 2014 and 2020. Patients from Institution A were assigned to the model development group (i.e., Development Cohort), while patients from Institutions B and C were assigned to the external validation groups (i.e., Validation Cohort_1 and Validation Cohort_2). The presence of CRLMs was determined by examination of surgical specimens. RAS mutation status was determined by genetic testing. The final predictors, identified by a group of oncologists and radiologists, included several key clinical, demographic, and radiographic characteristics derived from magnetic resonance images. Multiple imputation was performed to estimate the values of missing non-outcome data. A penalized logistic regression model using the adaptive least absolute shrinkage and selection operator penalty was implemented to select appropriate variables for the development of the model. A single nomogram was constructed from the model. The performance of the prediction model, discrimination, and calibration were estimated and reported by the area under the receiver operating characteristic curve (AUC) and calibration plots. Internal validation with a bootstrapping procedure and external validation of the nomogram were assessed. Finally, decision curve analyses were used to characterize the clinical outcomes of the Development and Validation Cohorts. RESULTS: A total of 173 patients were enrolled in this study between January 2014 and May 2020. Of the 173 patients, 117 patients from Institution A were assigned to the Model Development group, while 56 patients (33 from Institution B and 23 from Institution C) were assigned to the Model Validation groups. Forty-six (39.3%) patients harbored RAS mutations in the Development Cohort compared to 14 (42.4%) in Validation Cohort_1 and 8 (34.8%) in Validation Cohort_2. The final model contained the following predictor variables: time of occurrence of CRLMs, location of primary lesion, type of intratumoral necrosis, and early enhancement of liver parenchyma. The diagnostic model based on clinical and MRI data demonstrated satisfactory predictive performance in distinguishing between mutated and wild-type RAS, with AUCs of 0.742 (95% confidence interval [CI]: 0.651─0.834), 0.741 (95% CI: 0.649─0.836), 0.703 (95% CI: 0.514─0.892), and 0.708 (95% CI: 0.452─0.964) in the Development Cohort, bootstrapping internal validation, external Validation Cohort_1 and Validation Cohort_2, respectively. The Hosmer-Lemeshow goodness-of-fit values for the Development Cohort, Validation Cohort_1 and Validation Cohort_2 were 2.868 (p = 0.942), 4.616 (p = 0.465), and 6.297 (p = 0.391), respectively. CONCLUSIONS: Integrating clinical, demographic, and radiographic modalities with a magnetic resonance imaging-based approach may accurately predict the RAS mutation status of CRLMs, thereby aiding in triage and possibly reducing the time taken to perform diagnostic and life-saving procedures. Our diagnostic multivariate prediction model may serve as a foundation for prognostic stratification and therapeutic decision-making.

Neoadjuvant Chemotherapy With CAPOX Versus Chemoradiation for Locally Advanced Rectal Cancer With Uninvolved Mesorectal Fascia (CONVERT): Initial Results of a Phase III Trial.

OBJECTIVE: To compare neoadjuvant chemotherapy (nCT) with CAPOX alone versus neoadjuvant chemoradiotherapy (nCRT) with capecitabine in locally advanced rectal cancer (LARC) with uninvolved mesorectal fascia (MRF). BACKGROUND DATA: nCRT is associated with higher surgical complications, worse long-term functional outcomes, and questionable survival benefits. Comparatively, nCT alone seems a promising alternative treatment in lower-risk LARC patients with uninvolved MRF. METHODS: Patients between June 2014 and October 2020 with LARC within 12 cm from the anal verge and uninvolved MRF were randomly assigned to nCT group with 4 cycles of CAPOX (Oxaliplatin 130 mg/m2 IV day 1 and Capecitabine 1000 mg/m2 twice daily for 14 d. Repeat every 3 wk) or nCRT group with Capecitabine 825 mg/m² twice daily administered orally and concurrently with radiation therapy (50 Gy/25 fractions) for 5 days per week. The primary end point is local-regional recurrence-free survival. Here we reported the results of secondary end points: histopathologic response, surgical events, and toxicity. RESULTS: Of the 663 initially enrolled patients, 589 received the allocated treatment (nCT, n=300; nCRT, n=289). Pathologic complete response rate was 11.0% (95% CI, 7.8-15.3%) in the nCT arm and 13.8% (95% CI, 10.1-18.5%) in the nCRT arm ( P =0.33). The downstaging (ypStage 0 to 1) rate was 40.8% (95% CI, 35.1-46.7%) in the nCT arm and 45.6% (95% CI, 39.7-51.7%) in the nCRT arm ( P =0.27). nCT was associated with lower perioperative distant metastases rate (0.7% vs. 3.1%, P =0.03) and preventive ileostomy rate (52.2% vs. 63.6%, P =0.008) compared with nCRT. Four patients in the nCT arm received salvage nCRT because of local disease progression after nCT. Two patients in the nCT arm and 5 in the nCRT arm achieved complete clinical response and were treated with a nonsurgical approach. Similar results were observed in subgroup analysis. CONCLUSIONS: nCT achieved similar pCR and downstaging rates with lower incidence of perioperative distant metastasis and preventive ileostomy compared with nCRT. CAPOX could be an effective alternative to neoadjuvant therapy in LARC with uninvolved MRF. Long-term follow-up is needed to confirm these results.

The re-evaluation of optimal lymph node yield in stage II right-sided colon cancer: is a minimum of 12 lymph nodes adequate?

PURPOSE: Adequate lymphadenectomy is critical for accurate nodal staging and planning adjuvant therapy in colon cancer. However, the optimal lymph node (LN) yield for stage II right-sided colon cancer (RSCC) is still unclear. This population-based study aimed to determine the optimal LN yield associated with survival and LN positivity in patients with stage II RSCC. METHODS: All patients with stage II-III RSCC were identified from the Surveillance, Epidemiology, and End Results database over a 10-year interval (2006-2015). The optimal threshold for LN yield was explored using an outcome-oriented approach based on survival and LN positivity. RESULTS: The median number of LNs examined for all 17,385 patients with stage II RSCC was 17 (IQR 12-23). Nineteen LNs were determined as the optimal cut-off point to maximize survival benefit from lymphadenectomy. Increased LN yield was associated with a gradual increase in the risk of node positivity, with no change after 19 nodes. Compared with patients with 19 or more LNs examined, the group with fewer LNs had a significantly poor cancer-specific survival (< 12 nodes: hazard ratio (HR) 2.26, P < 0.001; 12-18 nodes: HR 1.58, P < 0.001) and overall survival (< 12 nodes: HR 1.80, P < 0.001; 12-18 nodes: HR 1.31, P < 0.001). Similar survival results were found in the validation cohort. Patients with older age, small tumor size, and appendix and transverse colon cancer were more likely to receive inadequate LN harvest. CONCLUSION: A minimum of 19 LNs is needed to be examined for optimal survival and adequate node staging in lymph node-negative RSCC.

Prognostic Role of Neutrophil-to-Lymphocyte Ratio in Locally Advanced Rectal Cancer Treated with Neoadjuvant Chemoradiotherapy.

BACKGROUND Increasing evidence suggests that cancer-associated inflammation is associated with poorer outcomes. The neutrophil-to-lymphocyte ratio (NLR), considered as a systemic inflammation marker, is thought to predict prognoses in colorectal cancer. In this study, we explored the association between the NLR and prognoses following neoadjuvant chemoradiotherapy (CRT) for locally advanced rectal cancer (LARC). MATERIAL AND METHODS From February 2002 to December 2012, a group of 202 patients diagnosed with LARC and receiving neoadjuvant CRT followed by radical surgery was included in our retrospective study. The associations between the pre-CRT NLR and clinicopathological characteristics, as well as the predictive value of pre-CRT NLR against survival outcomes, were analyzed. RESULTS The average NLR was 2.7±1.5 (median 2.4, range 0.6-12.8). There were 63 (31.2%) patients with NLR ≥3.0, and 139 (68.8%) patients with NLR <3.0. Correlation analyses showed that no clinicopathological characteristics except age were associated with NLR. We did not find an association between NLR and survival outcomes. In multivariate Cox model analyses, the R1/R2 resection, lymph node ratio ≥0.1, and perineural/lymphovascular invasion were independently associated with worse disease-free survival and overall survival. CONCLUSIONS In our cohort, the NLR did not correlate with survival outcomes in LARC patients undergoing neoadjuvant CRT. The prognostic value of NLR should be validated in large-scale prospective studies.

Pan-cancer analysis of Sushi domain-containing protein 4 (SUSD4) and validated in colorectal cancer.

Sushi domain-containing protein 4 (SUSD4) is a complement regulatory protein whose primary function is to inhibit the complement system, and it is involved in immune regulation. The role of SUSD4 in cancer progression has largely remained elusive. SUSD4 was studied across a variety of cancer types in this study. According to the results, there is an association between the expression level of SUSD4 and prognosis in multiple types of cancer. Further analysis demonstrated that SUSD4 expression level was related to immune cell infiltration, immune-related genes, tumor heterogeneity, and multiple cancer pathways. Additionally, we validated the function of SUSD4 in colorectal cancer cell lines and found that knockdown of SUSD4 inhibited cell growth and impacted the JAK/STAT pathway. By characterizing drug sensitivity in organoids, we found that the expression of SUSD4 showed a positive correlation trend with IC50 of Selumetinib, YK-4-279, and Piperlongumine. In conclusion, SUSD4 is a valuable prognostic indicator for diverse types of cancer, and it has the potential to be a target for cancer therapy.

Primary Surgery Followed by Selective Chemoradiotherapy Versus Preoperative Chemoradiotherapy Followed by Surgery for Locally Advanced Rectal Cancer: A Randomized Clinical Trial.

PURPOSE: The aim of this work was to determine whether locally advanced rectal cancer (LARC) with negative mesorectal fascia (MRF) predicted by magnetic resonance imaging (MRI) can be excluded from preoperative radiation therapy treatment. METHODS AND MATERIALS: This multicenter, open-label, non-inferiority, randomized clinical trial enrolled patients with LARC within 6 to 12 cm from the anal verge and with negative MRI-predicted MRF. Participants were randomized to the intervention group (primary surgery, in which the patients with positive pathologic [CRM] circumferential margins were subjected to chemoradiotherapy [CRT] and those with negative CRM underwent adjuvant chemotherapy according to pathologic staging) or the control group (preoperative CRT, in which all patients underwent subsequent surgery and adjuvant chemotherapy). The primary endpoint was 3-year disease-free survival (DFS). RESULTS: A total of 275 patients were randomly assigned to the intervention (n = 140) and control (n = 135) groups, in which 33.57% and 28.15% patients were at clinical T4 stage and 85.92% and 80.45% patients were at "bad" or "ugly" risk in the intervention and control groups, respectively. There were 2 patients (1.52%) and 1 patient (0.77%) with positive CRM in the intervention and control groups, respectively (P > .05). The non-adherence rates for the intervention and control groups were 3.6% and 23.7%, respectively. After a median follow-up of 34.6 months (IQR, 18.2-45.7), 43 patients had positive events (28 patients and 15 patients in the intervention and control groups, respectively). There were 6 patients (4.4%) with local recurrence in the intervention group and none in the control group, which led to the termination of the trial. The 3-year DFS rate was 81.82% in the intervention group (95% CI, 78.18%-85.46%) and 85.37% in the control group (95% CI, 81.75%-88.99%), with a difference of -3.55% (95% CI, -3.71% to -3.39%; hazard ratio, 1.76; 95% CI, 0.94-3.30). In the per-protocol data set, the difference between 3-year DFS rates was -5.44% (95% CI, -5.63% to -5.25%; hazard ratio, 2.02; 95% CI, 1.01-4.06). CONCLUSIONS: Based on the outcomes of this trial, in patients with LARC and MRI-negative MRF, primary surgery could negatively influence their DFS rates. Therefore, primary surgery was an inferior strategy compared with preoperative CRT followed by surgery and cannot be recommended for patients with LARC.

Effects of systemic inflammation and frailty on survival in elderly cancer patients: Results from the INSCOC study.

Background: Frailty and systemic inflammation are parameters, which are easy to evaluate, can be used to predict disease outcomes, and are potentially modifiable. The combination of frailty and inflammation-based data may help identify elderly cancer patients predisposed to adverse clinical outcomes. The aim of this study was to examine the association of systemic inflammation and frailty at admission, and to determine whether these risk factors interact and may predict the survival of elderly cancer patients. Methods: A prospective Investigation on Nutrition Status and Clinical Outcome of Common Cancers (INSCOC) with 5,106 elderly cancer patients admitted from 2013 through 2020 was included in this study. The primary marker of inflammation was the neutrophil-to-lymphocyte ratio (NLR), with the reference group having NLR<3, which indicated no inflammation. Frailty was assessed using the FRAIL scale, and patients with≥3 positives out of a total of five components were assumed to be frail. The primary outcome was all-cause mortality. We classified participants according to the presence (or absence) of frailty and high inflammation and assessed their association with overall survival using the Cox proportional hazards models adjusted for demographic, tumor, and treatment factors. Results: Among the 5,106 patients enrolled in the study, 3396 individuals (66.51%) were male and the mean( ± SD) age at diagnosis was 70.92( ± 5.34). Over a median of 33.5 months follow-up, we observed 2,315 deaths. Increasing NLR was associated with frailty (compared with NLR<3, odds ratio=1.23, 95%CI=1.08-1.41 for NLR≥3). An NLR≥3 and frailty independently predicted the overall survival [hazard ratio(HR)=1.35, 95%CI=1.24-1.47 and HR=1.38, 95%CI=1.25-1.52, respectively). Patients with both frailty and NLR≥3 had the lowest overall survival(HR=1.83, 95%CI=1.59-2.04) than patients with no risk factors. The mortality rate increased with the presence of the frailty components. Conclusions: Systemic inflammation was positively associated with frailty. Frail elderly cancer patients with elevated systemic inflammation had low survival rate.

Distinct profiles of epigenetic evolution between colorectal cancers with and without metastasis.

Liver metastasis is a fatal step in the progression of colorectal cancer (CRC); however, the epigenetic evolution of this process is largely unknown. To decipher the epigenetic alterations during the development of liver metastasis, the DNA methylation status of 12 genes, including 5 classical CpG island methylator phenotype (CIMP) markers, was analyzed in 62 liver metastases and in 78 primary CRCs (53 stage I-III; 25 stage IV). Genome-wide methylation analysis was also performed in stage I-III CRCs and in paired primary and liver metastatic cancers. Methylation frequencies of MGMT and TIMP3 increased progressively from stage I-III CRCs to liver metastasis (P = 0.043 and P = 0.028, respectively). The CIMP-positive cases showed significantly earlier recurrence of disease than did CIMP-negative cases with liver metastasis (P = 0.030), whereas no such difference was found in stage I-III CRCs. Genome-wide analysis revealed that more genes were methylated in stage I-III CRCs than in paired stage IV samples (P = 0.008). Hierarchical cluster analysis showed that stage I-III CRCs and stage IV CRCs were clustered into two distinct subgroups, whereas most paired primary and metastatic cancers showed similar methylation profiles. This analysis revealed distinct methylation profiles between stage I-III CRCs and stage IV CRCs, which may reflect differences in epigenetic evolution during progression of the disease. In addition, most methylation status in stage IV CRCs seems to be established before metastasis.

[Detection and clinical significance of apoptosis related protein in local advanced rectal cancer patients with preoperative neoadjuvant therapy].

OBJECTIVE: To discuss the mechanism of rectal cancer apoptosis induced by preoperative chemoradiotherapy and evaluate its effect by detection of apoptosis related proteins in locally advanced colorectal cancer patients who had received preoperative chemoradiation. METHODS: To detect Bcl-XL and Bax expression in rectal cancer before and after chemoradiotherapy by EnVision method, combined with patients clinical and pathological index, statistically analysis and evaluation their relationship and clinical significance. RESULTS: Patients with or without tumor shrinkage after preoperative chemoradiotherapy was 13 cases and 21 cases. While the positive rate of Bcl-XL in rectal cancer before and after chemoradiotherapy were 58.8% (20/34) and 52.9% (18/34), respectively. There were significant difference between Bcl-XL change before and after chemoradiation with tumor size, tumor cells shrinkage and operation pattern. The positive rate of Bax in rectal cancer before and after chemoradiotherapy were 32.4% (11/34) and 44.1% (15/34), respectively. There were no significant difference between Bax change before and after chemoradiotherapy with tumor cells shrinkage. There were statistically significant difference between Bax ratio (χ(2) = 9.607, P = 0.048) before and after chemoradiation while there were no significant difference between Bcl-XL/Bax ratio before and after chemoradiation with tumor shrinkage. According to layered analysis with preoperative therapy, there were statistically significant difference (χ(2) = 13.964, P = 0.007) between Bcl-XL change with operation pattern while the same of significant difference between Bax change with tumor infiltration and tumor shrinkage (χ(2) = 10.806 and 10.455, both P < 0.05). CONCLUSIONS: Preoperative chemoradiation can influence rectal cancer cell's apoptosis and treatment effect by changing Bcl-XL and Bax expression. Bcl-XL downregulation and Bax upregulation have shown important function in colorectal cancer cell apoptosis which induced by preoperative chemoradiation, it can also improve the effection of chemoradiation in rectal cancer.

Patterns of Lymph Node Metastasis and Optimal Surgical Strategy in Small (≤20 mm) Gastroenteropancreatic Neuroendocrine Tumors.

Background: Regional lymph node metastasis (LNM) is crucial for planning additional lymphadenectomy, and is directly correlated with poor prognosis in gastroenteropancreatic neuroendocrine tumors (GEP-NETs). However, the patterns of LNM for small (≤20 mm) GEP-NETs remain unclear. This population-based study aimed at evaluating LNM patterns and identifying optimal surgical strategies from the standpoint of lymph node dissemination. Methods: This retrospective cohort study retrieved data from the Surveillance, Epidemiology, and End Results (SEER) 18 registries database for 17,308 patients diagnosed as having localized well-differentiated GEP-NETs ≤ 20 mm between January 1, 2004, and December 31, 2017. The patterns of LNM were characterized in 6,622 patients who underwent extended resection for adequate lymph node harvest. Results: Of 6,622 patients with localized small GEP-NETs in the current study, 2,380 (36%) presented with LNM after regional lymphadenectomy. Nodal involvement was observed in approximately 7.4%, 49.1%, 13.6%, 53.7%, 13.8%, 7.8%, and 15.4% of gastric (g-), small intestinal (si-), appendiceal (a-), colonic (c-), rectal (r-), non-functional pancreatic (nfp-), and functional pancreatic (fp-) NETs ≤ 20 mm. Patients with younger age, larger tumor size, and muscularis invasion were more likely to present with LNM. Additional lymphadenectomy conferred a significant survival advantage in NETs (≤10 mm: HR, 0.47; 95% CI, 0.33-0.66; p < 0.001; 11-20 mm: HR, 0.54; 95% CI, 0.34-0.85; p = 0.008) and fp-NETs ≤ 20 mm (HR, 0.08; 95% CI, 0.02-0.36; p = 0.001), as well as g-NETs (HR, 0.39; 95% CI, 0.16-0.96; p = 0.041) and c-NETs of 11-20 mm (HR, 0.07; 95% CI, 0.01-0.48; p = 0.007). Survival benefits of additional lymphadenectomy were not found in a-NETs, r-NETs, and nfp-NETs with a small size. Conclusions: Given the increased risk for nodal metastasis, primary tumor resection with regional lymphadenectomy is a potential optimal surgical strategy for si-NETs and fp-NETs ≤ 20 mm, as well as g-NETs and c-NETs of 11-20 mm. Local resection is an appropriate and reliable surgical approach for a-NETs, r-NETs, and nfp-NETs ≤ 20 mm.

A Comprehensive Prognostic Model for Colorectal Cancer Liver Metastasis Recurrence After Neoadjuvant Chemotherapy.

Background: For patients with colorectal cancer liver metastases (CRLMs), it is important to stratify patients according to the risk of recurrence. This study aimed to validate the predictive value of some clinical, imaging, and pathology biomarkers and develop an operational prognostic model for patients with CRLMs with neoadjuvant chemotherapy (NACT) before the liver resection. Methods: Patients with CRLMs accompanied with primary lesion and liver metastases lesion resection were enrolled into this study. A nomogram based on independent risk factors was identified by Kaplan-Meier analysis and multivariate Cox proportional hazard analysis. The predictive ability was evaluated by receiver operating characteristic (ROC) curves and decision curve analysis (DCA). Calibration plot were also used to explore the consistency between prediction and reality. Results: A total of 118 patients were enrolled into the study. Multivariable Cox analysis found that histopathological growth patterns (HGPs) [Hazard Rate (HR) = 2.130], radiology response (stable disease vs. partial response, HR = 2.207; progressive disease vs. partial response, HR = 3.824), lymph node status (HR = 1.442), and age (HR = 0.576) were independent risk factors for disease-free survival (DFS) (p < 0.05). Corresponding nomogram was constructed on the basis of the above factors, demonstrating that scores ranging from 5 to 11 presented better prognosis than the scores of 0-4 (median DFS = 14.3 vs. 4.9 months, p < 0.0001). The area under ROC curves of the model for 1-, 2-, and 3-year DFS were 0.754, 0.705, and 0.666, respectively, and DCA confirmed that the risk model showed more clinical benefits than clinical risk score. Calibration plot for the probability of DFS at 1 or 3 years verified an optimal agreement between prediction and actual observation. In the course of our research, compared with pure NACT, a higher proportion of desmoplastic HGP (dHGP) was detected in patients treated with NACT plus cetuximab (p = 0.030), and the use of cetuximab was an independent factor for decreased replacement HGP (rHGP) and increased dHGP (p = 0.049). Conclusion: Our model is concise, comprehensive, and high efficient, which may contribute to better predicting the prognosis of patients with CRLMs with NACT before the liver resection. In addition, we observed an unbalanced distribution of HGPs as well.

Computational Dissection of the Role of Trp305 in the Regulation of the Death-Associated Protein Kinase-Calmodulin Interaction.

Death-associated protein kinase 1 (DAPK1), as a calcium/calmodulin (CaM) regulated serine/threonine kinase, functions in apoptotic and autophagy pathways and represents an interesting drug target for inflammatory bowel disease and Alzheimer's disease. The crystal structure of the DAPK1 catalytic domain and the autoregulatory domain (ARD) in complex with CaM provides an understanding of CaM-dependent regulation of DAPK1 activity. However, the molecular basis of how distinct Trp305 (W305Y and W305D) mutations in the ARD modulate different DAPK1 activities remains unknown. Here, we performed multiple, µs-length molecular dynamics (MD) simulations of the DAPK1-CaM complex in three different (wild-type, W305Y, and W305D) states. MD simulations showed that the overall structural complex did not change significantly in the wild-type and W305Y systems, but underwent obvious conformational alteration in the W305D system. Dynamical cross-correlation and principal component analyses revealed that the W305D mutation enhanced the anti-correlated motions between the DAPK1 and CaM and sampled a broader distribution of conformational space relative to the wild-type and W305Y systems. Structural and energetical analyses further exhibited that CaM binding was unfavored in response to the W305D mutation, resulting in the decreased binding of CaM to the W305D mutant. Furthermore, the hydrogen bonds and salt bridges responsible for the loss of CaM binding on the interface of the DAPK1-CaM complex were identified in the W305D mutant. This result may provide insights into the key role of Trp305 in the regulation of CaM-mediated DAPK1 activity.

Short-term and long-term outcomes of natural orifice specimen extraction surgeries (NOSES) in rectal cancer: a comparison study of NOSES and non-NOSES.

Background: Natural orifice specimen extraction surgeries (NOSES) have been applied to colorectal cancer (CRC). Different types of NOSES have been proposed. Traditional laparoscopic CRC surgeries (non-NOSES) have been widely adopted in clinical practice. Therefore, the safety and feasibility of NOSES could be clarified by comparing with non-NOSES. Methods: Consecutive cases who underwent NOSE or non-NOSE rectal surgeries were retrospectively collected at the Second Affiliated Hospital of Harbin Medical University between 1 January 2013 and 31 December 2018. Other inclusion criteria included patients with adenocarcinoma of the rectum within 15 cm of the anal verge, over the age of 18 and undergoing primary laparoscopic rectal resection. Patients who were lost to follow-up or had incomplete information were excluded. Basic characteristics including gender, tumor location, age, staging, treatment, and Body Mass Index (BMI) were analyzed. Short-term outcomes including comorbidities, intra-operative blood loss, hospital stay, gas exhaust time were compared between different NOSES and non-NOSES groups. Long-term outcomes including overall survival (OS) and disease-free survival (DFS) were also analyzed. Patients were followed-up during the inpatient period, at an outpatient clinic, or by phone call. Results: A total of 196 NOSES cases and 243 non-NOSES cases were included. There was a sex difference between the two groups and other factors were comparable. Cases were divided into NOSES groups [including extra-abdominal resection (EVER), specimen extraction and extra-abdominal resection (EXER), and intra-abdominal resection and specimen extraction (IREX)] and non-NOSES groups. Differences in sex (P=0.016), BMI (with mean of 22.08, 22.00, 22.53, and 23.26 kg/m2, P=0.003), and staging (P=0.008) were observed between the four groups. There was a difference in the intra-operative blood loss between NOSES and non-NOSES groups (57.05±62.78, 52.65±68.19, 36.52±43.99 vs. 76.12±90.11 mL, P=0.002), in which NOSES groups had less blood loss. Furthermore, NOSES groups showed a better post-operative gas exhaust time (54.68±37.80, 45.06±24.69, 47.91±28.93 vs. 56.94±27.69 hours, P=0.012). NOSES groups also had fewer ileostomies (17 vs. 37, P=0.003). There was no difference in the long-term DFS and OS between the two groups. Conclusions: NOSES in rectal cancer showed better short-term outcomes and had comparable long-term outcomes compared with non-NOSE surgeries.

Combination of GC-MS based metabolomics analysis with mouse xenograft models reveals a panel of dysregulated circulating metabolites and potential therapeutic targets for colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) is a common gastrointestinal tumor with subtle, often undetectable early symptoms, which means that upon diagnosis, patients often present in the middle or late stages of disease. Therefore, the need for an effective biomarker for the early diagnosis and development of novel therapeutic targets is urgent to prolong patient survival time and reduce mortality. METHODS: Twenty mice were randomly divided into patient-derived xenograft (PDX) model (transplantation of fresh CRC tumor samples) and control groups (10 mice in each group). All the animals were euthanized using isoflurane at the end of the experiment. Gas chromatography-mass spectrometry (GC-MS)-based metabolomic profiling was performed to investigate the differential metabolites in the serum, and publicly available gene expression data (GSE106582) were analyzed to determine dysregulated metabolic pathways. Joint pathway analysis was used to identify potential metabolic targets. Immunohistochemistry analysis was performed to confirm the presence of the identified targets at the protein level. RESULTS: A total of 96 differential circulating metabolites were identified, which were predominantly involved in amino acid metabolism. In particular, the serum levels of amino acids such as phenylalanine and aspartic acid were significantly downregulated in the PDX group, suggesting an increased consumption of amino acids in CRC. Moreover, both the mRNA and protein levels of the amino acid transporters, SLC7A5 and SLC1A5, were found to be upregulated in CRC. CONCLUSIONS: By combining GC-MS-based metabolomics profiling with a PDX model of CRC our study successfully identified potential diagnostic circulating metabolites. Dysregulated amino acid metabolism was found to be a significant feature of CRC. The amino acid transporters, SLC7A5 and SLC1A5, were identified as potential metabolic therapeutic targets. This study furthers the understanding of the metabolic processes involved in CRC.

Development and external validation of a novel nomogram for screening Chinese Lynch syndrome: based on a multicenter, population study.

BACKGROUND: This multicenter study aimed to reveal the genetic spectrum of colorectal cancer (CRC) with deficient mismatch repair (dMMR) and build a screening model for Lynch syndrome (LS). METHODS: Through the immunohistochemical (IHC) screening of mismatch repair protein results in postoperative CRC patients, 311 dMMR cases, whose germline and somatic variants were detected using the ColonCore panel, were collected. Univariate and multivariate logistic regression analysis was performed on the clinical characteristics of these dMMR individuals, and a clinical nomogram, incorporating statistically significant factors identified using multivariate logistic regression analysis, was constructed to predict the probability of LS. The model was validated externally by an independent cohort. RESULTS: In total, 311 CRC patients with IHC dMMR included 95 identified MMR germline variant (LS) cases and 216 cases without pathogenic or likely pathogenic variants in MMR genes (non-Lynch-associated dMMR). Of the 95 individuals, approximately 51.6%, 28.4%, 14.7%, and 5.3% cases carried germline MLH1, MSH2, MSH6, and PMS2 pathogenic or likely pathogenic variants, respectively. A novel nomogram was then built to predict the probability of LS for CRC patients with dMMR intuitively. The receiver operating characteristic (ROC) curve informed that this nomogram-based screening model could identify LS with a higher specificity and sensitivity with an area under curve (AUC) of 0.87 than current screening criteria based on family history. In the external validation cohort, the AUC of the ROC curve reached 0.804, inferring the screening model's universal applicability. We recommend that dMMR-CRC patients with a probability of LS greater than 0.435 should receive a further germline sequencing. CONCLUSION: This novel screening model based on the clinical characteristic differences between LS and non-Lynch-associated dMMR may assist clinicians to preliminarily screen LS and refer susceptible patients to experienced specialists.

Exosome-transferred long non-coding RNA ASMTL-AS1 contributes to malignant phenotypes in residual hepatocellular carcinoma after insufficient radiofrequency ablation.

OBJECTIVES: Long non-coding RNAs (lncRNAs) are emerging RNA regulators in cancer progression, including in hepatocellular carcinoma (HCC). Recently, insufficient radiofrequency ablation (RFA) has been reported to lead to recurrence and metastasis of residual HCC tumours. Herein, we aimed to the role of ASMTL-AS1 in residual HCC after insufficient RFA. MATERIALS AND METHODS: In vitro insufficient RFA model was simulated in Huh7 cells and subsequently named Huh7-H cells. In vitro and in vivo assays were conducted to investigate ASMTL-AS1 function in HCC. RESULTS: LncRNA ASMTL-AS1 low expressed in normal human liver was found to be highly expressed in HCC tissues and further increased in tumours after insufficient RFA. ASMTL-AS1 expression was related to stage, metastasis and prognosis in HCC. Huh7-H possessed higher ASMTL-AS1 level and more aggressive than Huh7 cells. ASMTL-AS1 contributed to the malignancy of HCC cells both in vitro and in vivo. Mechanistically, ASMTL-AS1 was trans-activated by MYC and promoted NLK expression to activate YAP signalling via sequestering miR-342-3p in HCC. Interestingly, ASMTL-AS1 could be wrapped by exosomes and then convey malignancy through NLK/YAP axis between cells even in residual HCC after insufficient RFA. CONCLUSIONS: Exosomal ASMTL-AS1 aggravates the malignancy in residual HCC after insufficient RFA via miR-342-3p/NLK/YAP signalling, opening a new road for the treatment of HCC and the prevention of recurrence or metastasis of residual HCC after insufficient RFA.

Circulating guanylyl cyclase C (GCC) mRNA is a reliable metastatic predictor and prognostic index of colorectal cancer.

BACKGROUND: Currently, few specific biomarkers or standard cutoff values are available for circulating tumor cells (CTCs) detection and survival prediction in patients with early stage colorectal cancer (CRC). Guanylyl cyclase C (GCC) presents as a specific expression in intestinal tumor cells and during their metastases, indicating its potential application as a metastatic predictor of CRC. METHODS: The circulating GCC mRNA of 160 colorectal cancer patients at stage I-III was detected via quantitative real-time (qRT)-PCR in our study, and the correlation of GCC mRNA level with tumor metastasis and long-term survival was explored. RESULTS: GCC mRNA was found to be positive in 43 out of 160 CRC patients and negative in ten healthy controls. It was found that GCC mRNA over the baseline (>100 copies/µL and 200 copies/µL) showed a significant correlation with disease-free survival (DFS) and overall survival (OS) in the stage II subgroup. It was further revealed that GCC mRNA over 300 copies/µL or higher than the median value of copy numbers was significantly correlated with reduced OS and DFS in CRC patients. A nomogram model based on variables including GCC mRNA copy number was established for predicting the OS of CRC patients (AUC =0.98). CONCLUSIONS: Circulating GCC mRNA over baseline is a reliable predictor for tumor metastasis and can be a prognostic index in CRC patients.

Integrative Analysis of the IQ Motif-Containing GTPase-Activating Protein Family Indicates That the IQGAP3-PIK3C2B Axis Promotes Invasion in Colon Cancer.

BACKGROUND: Colon cancer (CRC) is a common type of tumour, and IQGAP family proteins play an important role in many tumours. However, their roles in CRC remain unclear. METHODS: First, we searched many public databases to comprehensively analyze expression of IQGAPs in CRC. Next, real-time PCR, immunohistochemical (IHC), transwell, siRNA transfection and Western blot assays were used to evaluate relationships among IQGAP3 expression, clinical pathological parameters and CRC prognosis, and the underlying molecular mechanism was investigated. RESULTS: IQGAP3 was elevated in CRC tissues, whereas there was no significant change in expression of IQGAP1 or IQGAP2. Additionally, IQGAP3 expression in CRC tissues was associated with tumour progression, invasion and poor prognosis. In mechanistic studies, we found that IQGAP3 was positively coexpressed with PIK3C2B. In an in vitro assay, the PIK3C2B expression level was increased after exogenous overexpression of IQGAP3, resulting in the promotion of cell invasion, which was blocked by pretransfecting cells with PIK3C2B siRNA. Furthermore, we found that high expression of IQGAP3 and PIK3C2B correlated with tumour stage and vessel invasion in human CRC, whereby patients with high expression of both in tumours had a worse prognosis compared with patients with single-positive or double-negative tumours. CONCLUSION: The results of our current study and corresponding previous studies provide evidence that IQGAP3 is elevated in CRC and promotes colon cancer growth and metastasis by regulating PIK3C2B activation.

O-GlcNAcylation of PGK1 coordinates glycolysis and TCA cycle to promote tumor growth.

Many cancer cells display enhanced glycolysis and suppressed mitochondrial metabolism. This phenomenon, known as the Warburg effect, is critical for tumor development. However, how cancer cells coordinate glucose metabolism through glycolysis and the mitochondrial tricarboxylic acid (TCA) cycle is largely unknown. We demonstrate here that phosphoglycerate kinase 1 (PGK1), the first ATP-producing enzyme in glycolysis, is reversibly and dynamically modified with O-linked N-acetylglucosamine (O-GlcNAc) at threonine 255 (T255). O-GlcNAcylation activates PGK1 activity to enhance lactate production, and simultaneously induces PGK1 translocation into mitochondria. Inside mitochondria, PGK1 acts as a kinase to inhibit pyruvate dehydrogenase (PDH) complex to reduce oxidative phosphorylation. Blocking T255 O-GlcNAcylation of PGK1 decreases colon cancer cell proliferation, suppresses glycolysis, enhances the TCA cycle, and inhibits tumor growth in xenograft models. Furthermore, PGK1 O-GlcNAcylation levels are elevated in human colon cancers. This study highlights O-GlcNAcylation as an important signal for coordinating glycolysis and the TCA cycle to promote tumorigenesis.