Chemotherapy-induced infiltration of neutrophils promotes pancreatic cancer metastasis via Gas6/AXL signalling axis.

OBJECTIVE: Pancreatic ductal adenocarcinoma (PDAC) is a highly metastatic disease and cytotoxic chemotherapy is the standard of care treatment for patients with advanced disease. Here, we investigate how the microenvironment in PDAC liver metastases reacts to chemotherapy and its role in metastatic disease progression post-treatment, an area which is poorly understood. DESIGN: The impact of chemotherapy on metastatic disease progression and immune cell infiltrates was characterised using flow and mass cytometry combined with transcriptional and histopathological analysis in experimental PDAC liver metastases mouse models. Findings were validated in patient derived liver metastases and in an autochthonous PDAC mouse model. Human and murine primary cell cocultures and ex vivo patient-derived liver explants were deployed to gain mechanistical insights on whether and how chemotherapy affects the metastatic tumour microenvironment. RESULTS: We show that in vivo, chemotherapy induces an initial infiltration of proinflammatory macrophages into the liver and activates cytotoxic T cells, leading only to a temporary restraining of metastatic disease progression. However, after stopping treatment, neutrophils are recruited to the metastatic liver via CXCL1 and 2 secretion by metastatic tumour cells. These neutrophils express growth arrest specific 6 (Gas6) which leads to AXL receptor activation on tumour cells enabling their regrowth. Disruption of neutrophil infiltration or inhibition of the Gas6/AXL signalling axis in combination with chemotherapy inhibits metastatic growth. Chemotherapy increases Gas6 expression in circulating neutrophils from patients with metastatic pancreatic cancer and recombinant Gas6 is sufficient to promote tumour cell proliferation ex vivo, in patient-derived metastatic liver explants. CONCLUSION: Combining chemotherapy with Gas6/AXL or neutrophil targeted therapy could provide a therapeutic benefit for patients with metastatic pancreatic cancer.

Impact on blood loss and transfusion rates following administration of tranexamic acid in major oncological abdominal and pelvic surgery: A systematic review and meta-analysis.

BACKGROUND AND OBJECTIVES: Major bleeding and receiving blood products in cancer surgery are associated with increased postoperative complications and worse outcomes. Tranexamic acid (TXA) reduces blood loss and improves outcomes in various surgical specialities. We performed a systematic review and meta-analysis to investigate TXA use on blood loss in elective abdominal and pelvic cancer surgery. METHODS: A literature search was performed for studies comparing intravenous TXA versus placebo/no TXA in patients undergoing major elective abdominal or pelvic cancer surgery. RESULTS: Twelve articles met the inclusion criteria, consisting of 723 patients who received TXA and 659 controls. Patients receiving TXA were less likely to receive a red blood cell (RBC) transfusion (p < 0.001, OR 0.4 95% CI [0.25, 0.63]) and experienced less blood loss (p < 0.001, MD -197.8 ml, 95% CI [-275.69, -119.84]). The TXA group experienced a smaller reduction in haemoglobin (p = 0.001, MD -0.45 mmol/L, 95% CI [-0.73, -0.18]). There was no difference in venous thromboembolism (VTE) rates (p = 0.95, OR 0.98, 95% CI [0.46, 2.08]). CONCLUSIONS: TXA use reduced blood loss and RBC transfusion requirements perioperatively, with no significant increased risk of VTE. However, further studies are required to assess its benefit for cancer surgery in some sub-specialities.

Failure of nonoperative management in patients with acute diverticulitis complicated by abscess: a systematic review.

BACKGROUND: The aim of this study was to assess failure rates following nonoperative management of acute diverticulitis complicated by abscess and trends thereof. METHOD: Pubmed, MEDLINE, EMBASE, CINAHL, Cochrane Library, and Web of Science were systematically searched. Nonoperative management was defined as a combination of nil per os, IV fluids, IV antibiotics, CT scan-guided percutaneous drainage, and total parenteral nutrition. The primary endpoint was failure of nonoperative management defined as persistent or worsening abscess and/or sepsis, development of new complications, such as peritonitis, ileus, or colocutaneous fistula, and urgent surgery within 30-90 days of index admission. Data were stratified by three arbitrary time intervals: 1986-2000, 2000-2010, and after 2010. The primary outcome was calculated for those groups and compared. RESULTS: Thirty-eight of forty-four eligible studies published between 1986 and 2019 were included in the quantitative synthesis of data (n = 2598). The pooled rate of failed nonoperative management was 16.4% (12.6%, 20.2%) at 90 days. In studies published in 2000-2010 (n = 405), the pooled failure rate was 18.6% (10.5%, 26.7%). After 2000 (n = 2140), the pooled failure rate was 15.3% (10.7%, 20%). The difference was not statistically significant (p = 0.725). After controlling for heterogeneity in the definition of failure of nonoperative management, subgroup analysis yielded the pooled rate of failure of 21.8% (16.1%, 27.4%). CONCLUSION: This meta-analysis found that failure rates following nonoperative management of acute diverticulitis complicated by abscess did not significantly decrease over the past three decades. The general quality of published data and the level and certainty of evidence produced were low.

Intra-operative assessment of left-sided colorectal anastomotic integrity: a systematic review of available techniques.

AIM: Anastomotic leak (AL) after colorectal resection is associated with increased rates of morbidity and mortality: potential permanent stoma formation, increased local recurrence, reduced cancer-related survival, poor functional outcomes and associated quality of life. Techniques to reduce leak rates are therefore highly sought. METHOD: A literature search was performed for published full text articles using PubMed, Cochrane and Scopus databases with a focus on colorectal surgery 1990-2020. Additional papers were detected by scanning references of relevant papers. RESULTS: A total of 53 papers were included after a thorough literature search. Techniques assessed included leak tests, endoscopy, perfusion assessment and fluorescence studies. Air-leak testing remains the most commonly used method across Europe, due to ease of reproducibility and low cost. There is no evidence that this reduces the leak rate; however, identification of a leak intra-operatively provides the opportunity for either suture reinforcement or formal takedown with or without re-do of the anastomosis and consideration of diversion. Suture repair alone of a positive air-leak test is associated with an increased AL rate. The use of fluorescence studies to guide the site of anastomosis has demonstrated reduced leak rates in distal anastomoses, is safe, feasible and has a promising future. CONCLUSION: Although over reliance on any assessment tool should be avoided, intra-operative techniques with the aim of reducing AL rates are increasingly being employed. Standardization of these methods is imperative for routine use. However, in the interim it is recommended that all anastomoses should be assessed intra-operatively for mechanical failure, particularly distal anastomoses.

Management of benign anastomotic strictures following rectal resection: a systematic review.

AIM: Benign anastomotic strictures following colorectal surgical resection are a commonly under-reported complication in up to 30% of patients, with a significant impact upon quality of life. In this systematic review, we aim to assess the utility of endoscopic techniques in avoiding the need for surgical reintervention. METHOD: A literature search was performed for published full text articles using the PubMed, Cochrane and Scopus databases. Additional papers were found by scanning the references of relevant papers. RESULTS: A total of 34 papers were included, focusing upon balloon dilatation, endoscopic stenting, electroincision, stapler stricturoplasty and cortiocosteroids alone and in combination, with success rates varying from 20% to 100%. The most challenging strictures were reported as those with a narrow lumen, frequently observed following neoadjuvant chemoradiotherapy or an anastomotic leak. Endoscopic balloon dilatation was the most commonly used first-line method; however, repeated dilatations were often required and this was associated with an increased risk of perforation. Although initial success rates for stents were good, patients often experienced stent migration and local symptoms. Only a small number of patients experienced endoscopic management failure and progressed to surgical intervention. CONCLUSION: Following identification of an anastomotic stricture and exclusion of underlying malignancy, endoscopic management is both safe and feasible as a first-line option, even if multiple treatment exposures or multimodal management is required. Surgical resection or a defunctioning stoma should be reserved for emergency or failed cases. Further research is required into multimodal and novel therapies to improve quality of life for these patients.

Measuring sarcopenia on pre-operative CT in older adults undergoing emergency laparotomy: a comparison of three different calculations.

INTRODUCTION: Sarcopenia is associated with outcomes in older-adults undergoing emergency surgery. Psoas major measurement is a surrogate marker of sarcopenia with multiple calculations existing normalising to body size and no consensus as to which is optimal. We compared three different psoas-major calculations to predict outcomes in older adults undergoing emergency laparotomy. METHODS: Consecutive over 65s were identified from the National Emergency Laparotomy Audit(NELA) database at a single centre between 2014 and 2018. Psoas major was measured at the L3 level and normalised to height (psoas muscle index, PMI), L3 vertebral body (psoas muscle:L3 ratio, PML3) or body surface area (psoas:body surface area, PBSA) and each correlated to outcomes. Outcome measures included inpatient, 30-day and 90-day mortality. A comparison of the three calculations was performed using the Mann-Whitney U, chi-squared, receiver operating characteristic curves (ROC) and binary logistic regression. RESULTS: Two hundred and sixty-four older adults underwent emergency laparotomy (median age, 75 years ((IQR, 70-81 years), 50% female)). Inpatient mortality was 19.6%, 30-day mortality was 15.1% and 90-day mortality was 18.5%. A total of 31.1% of males and 30% of females were sarcopenic (30.6% overall). A multivariate analysis confirmed each method of psoas major calculation (p < 0.0001) to be associated with mortality, as was ASA-grade (p < 0.0001). Area under the curve (AUC) was greatest for PML3 in predicting mortality (inpatient: PML3, 0.76; PMI, 0.71; PBSA, 0.70; 30-day: PML3, 0.74; PMI, 0.68; PBSA, 0.68; and 90-day: PML3, 0.78; PMI, 0.71; PBSA, 0.70). ASA-grade, P-POSSUM and PML3 were independently associated with mortality on multivariate analysis. ROC analysis of predictions from logistic regression models demonstrated PML3 to be more closely aligned to mortality than ASA or P-POSSUM (inpatient: AUC:PML3, 0.807; ASA, 0.783; P-POSSUM, 0.762; 30-day:AUC: PML3, 0.799; ASA, 0.784; P-POSSUM, 0.787; and 90-day: AUC:PML3, 0.805; ASA, 0.781; P-POSSUM, 0.756). CONCLUSIONS: Sarcopenia was present in 30.6% of older adults undergoing emergency surgery and is associated with a significantly increased mortality. PML3 is superior to PMI or PBSA and should be considered the method of calculation of choice. Additionally, PML3 compares favourably to ASA and P-POSSUM.

Prospective patient stratification into robust cancer-cell intrinsic subtypes from colorectal cancer biopsies.

Colorectal cancer (CRC) biopsies underpin accurate diagnosis, but are also relevant for patient stratification in molecularly-guided clinical trials. The consensus molecular subtypes (CMSs) and colorectal cancer intrinsic subtypes (CRISs) transcriptional signatures have potential clinical utility for improving prognostic/predictive patient assignment. However, their ability to provide robust classification, particularly in pretreatment biopsies from multiple regions or at different time points, remains untested. In this study, we undertook a comprehensive assessment of the robustness of CRC transcriptional signatures, including CRIS and CMS, using a range of tumour sampling methodologies currently employed in clinical and translational research. These include analyses using (i) laser-capture microdissected CRC tissue, (ii) eight publically available rectal cancer biopsy data sets (n = 543), (iii) serial biopsies (from AXEBeam trial, NCT00828672; n = 10), (iv) multi-regional biopsies from colon tumours (n = 29 biopsies, n = 7 tumours), and (v) pretreatment biopsies from the phase II rectal cancer trial COPERNCIUS (NCT01263171; n = 44). Compared to previous results obtained using CRC resection material, we demonstrate that CMS classification in biopsy tissue is significantly less capable of reliably classifying patient subtype (43% unknown in biopsy versus 13% unknown in resections, p = 0.0001). In contrast, there was no significant difference in classification rate between biopsies and resections when using the CRIS classifier. Additionally, we demonstrated that CRIS provides significantly better spatially- and temporally- robust classification of molecular subtypes in CRC primary tumour tissue compared to CMS (p = 0.003 and p = 0.02, respectively). These findings have potential to inform ongoing biopsy-based patient stratification in CRC, enabling robust and stable assignment of patients into clinically-informative arms of prospective multi-arm, multi-stage clinical trials. © 2018 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Proteomic analysis to identify biomarkers in the primary tumour that predict response to neoadjuvant chemotherapy in liver metastases.

BACKGROUND: Colorectal cancer is the fourth commonest cancer in the UK, and the second commonest cause of cancer-related death. A knowledge of the biological phenotype of colorectal liver metastases would be invaluable to inform clinical decision making; however, deriving this information from the metastatic lesions is not feasible until after resection. We aimed to use proteomic analysis to identify biomarkers in the primary tumour that predict response to neoadjuvant chemotherapy in liver metastases. METHODS: Fresh tissue from both primary colorectal tumour and liver metastases from 17 patients was subjected to proteomic analysis using isobaric tagging for relative quantification. Data were analysed with Protein Pilot (Ab Sciex, Framingham, MA, USA), with stratification of patients into those showing low or high response to chemotherapy permitting the identification of potential predictive biomarkers. These markers were subsequently validated by immunohistochemistry on a tissue microarray of 63 patients. FINDINGS: We identified 5768 discrete proteins. Five of them predicted histopathological response to fluorouracil-based chemotherapy regimens, of which the FAD binding protein NQO1 was subsequently validated by immunohistochemistry. When compared with the chemotherapeutic agent alone, knockdown of the corresponding gene with small interfering RNA decreased cell viability when co-incubated with fluorouracil (77·1% vs 46·6%, p=0·037) and irinotecan (41·7% vs 24·4%, p=0·006). Similar results were also seen after inhibition of protein activity by pretreating cells with dicoumarol. INTERPRETATION: These results show that proteomic sequencing of matched metastatic colorectal cancer samples is feasible, with high protein coverage. The high degree of similarity between the primary and secondary proteomes suggests that primary tissue is predictive of the metastatic phenotype. NQO1 expression in the primary tumour predicts response to neoadjuvant chemotherapy in the liver metastases, and inhibition of this protein at both genetic and functional levels improves chemosensitivity. FUNDING: Cancer Research UK.

Genetic, epigenetic and environmental factors in diverticular disease: systematic review.

BACKGROUND: Diverticulosis is a normal anatomical variant of the colon present in more than 70% of the westernized population over the age of 80. Approximately 3% will develop diverticulitis in their lifetime. Many patients present emergently, suffer high morbidity rates and require substantial healthcare resources. Diverticulosis is the most common finding at colonoscopy and has the potential for causing a significant morbidity rate and burden on healthcare. There is a need to better understand the aetiology and pathogenesis of diverticular disease. Research suggests a genetic susceptibility of 40-50% in the formation of diverticular disease. The aim of this review is to present the hypothesized functional effects of the identified gene loci and environmental factors. METHODS: A systematic literature review was performed using PubMed, MEDLINE and Embase. Medical subject headings terms used were: 'diverticular disease, diverticulosis, diverticulitis, genomics, genetics and epigenetics'. A review of grey literature identified environmental factors. RESULTS: Of 995 articles identified, 59 articles met the inclusion criteria. Age, obesity and smoking are strongly associated environmental risk factors. Intrinsic factors of the colonic wall are associated with the presence of diverticula. Genetic pathways of interest and environmental risk factors were identified. The COLQ, FAM155A, PHGR1, ARHGAP15, S100A10, and TNFSF15 genes are the strongest candidates for further research. CONCLUSION: There is increasing evidence to support the role of genomics in the spectrum of diverticular disease. Genomic, epigenetic and omic research with demographic context will help improve the understanding and management of this complex disease.

Enhancing radiotherapy outcomes in rectal cancer: A systematic review of targeting hypoxia-induced radioresistance.

Introduction: Neoadjuvant radiotherapy is successfully used in rectal cancer to improve overall survival. However, treatment response is both unpredictable and variable. There is strong evidence to show that the phenomenon of tumour hypoxia is associated with radioresistance, however the mechanism(s) behind this are poorly understood. Consequently, there have only been a small number of studies evaluating methods targeting hypoxia-induced radioresistance. The purpose of this systematic review is to evaluate the potential effectiveness of targeting hypoxia-induced radioresistance in rectal cancer and provide recommendations for future research in this area. Methods: A comprehensive literature search was performed following the PRISMA guidelines. This study was registered on the Prospero database (CRD42023441983). Results: Eight articles met the inclusion criteria. All studies identified were in vitro or in vivo studies, there were no clinical trials. Of the 8 studies identified, 5 assessed the efficacy of drugs which directly or indirectly targeted hypoxia and three that identified potential targets. There was conflicting in vivo evidence for the use of metformin to overcome hypoxia induced radioresistance. Vorinostat, atovaquone, and evofosfamide showed promising preclinical evidence that they can overcome hypoxia-induced radioresistance. Discussion: The importance of investigating hypoxia-induced radioresistance in rectal cancer is crucial. However, to date, only a small number of preclinical studies exist evaluating this phenomenon. This systematic review highlights the importance of further research to fully understand the mechanism behind this radioresistance. There are promising targets identified in this systematic review however, substantially more pre-clinical and clinical research as a priority for future research is needed.

Myoferlin: A Potential Marker of Response to Radiation Therapy and Survival in Locally Advanced Rectal Cancer.

PURPOSE: Patients with locally advanced rectal cancer often require neoadjuvant chemoradiation therapy to downstage the disease, but the response is variable with no predictive biomarkers. We have previously revealed through proteomic profiling that myoferlin is associated with response to radiation therapy. The aims of this study were to further validate this finding and explore the potential for myoferlin to act as a prognostic and/or therapeutic target. METHODS AND MATERIALS: Immunohistochemical analysis of a tissue microarray (TMA) for 111 patients was used to validate the initial proteomic findings. Manipulation of myoferlin was achieved using small interfering RNA, a small molecular inhibitor (wj460), and a CRISPR-Cas9 knockout cell line. Radiosensitization after treatment was assessed using 2-dimensional clonogenic assays, 3-dimensional spheroid models, and patient-derived organoids. Underlying mechanisms were investigated using electrophoresis, immunofluorescence, and immunoblotting. RESULTS: Analysis of both the diagnostic biopsy and tumor resection samples confirmed that low myoferlin expression correlated with a good response to neoadjuvant long-course chemoradiation therapy. High myoferlin expression was associated with spread to local lymph nodes and worse 5-year survival (P = .01; hazard ratio, 3.5; 95% CI, 1.27-10.04). This was externally validated using the Stratification in Colorectal Cancer database. Quantification of myoferlin using immunoblotting in immortalized colorectal cancer cell lines and organoids demonstrated that high myoferlin expression was associated with increased radioresistance. Biological and pharmacologic manipulation of myoferlin resulted in significantly increased radiosensitivity across all cell lines in 2-dimensional and 3-dimensional models. After irradiation, myoferlin knockdown cells had a significantly impaired ability to repair DNA double-strand breaks. This appeared to be mediated via nonhomologous end-joining. CONCLUSIONS: We have confirmed that high expression of myoferlin in rectal cancer is associated with poor response to neoadjuvant therapy and worse long-term survival. Furthermore, the manipulation of myoferlin led to increased radiosensitivity in vitro. This suggests that myoferlin could be targeted to enhance the sensitivity of patients with rectal cancer to radiation therapy, and further work is required.

Sarcopenia estimation using psoas major enhances P-POSSUM mortality prediction in older patients undergoing emergency laparotomy: cross-sectional study.

INTRODUCTION: Emergency laparotomy is a considerable component of a colorectal surgeon's workload and conveys substantial morbidity and mortality, particularly in older patients. Frailty is associated with poorer surgical outcomes. Frailty and sarcopenia assessment using Computed Tomography (CT) calculation of psoas major area predicts outcomes in elective and emergency surgery. Current risk predictors do not incorporate frailty metrics. We investigated whether sarcopenia measurement enhanced mortality prediction in over-65 s who underwent emergency laparotomy and emergency colorectal resection. METHODS: An analysis of data collected prospectively during the National Emergency Laparotomy Audit (NELA) was conducted. Psoas major (PM) cross-sectional area was measured at the L3 level and a ratio of PM to L3 vertebral body area (PML3) was calculated. Outcome measures included inpatient, 30-day and 90-day mortality. Statistical analysis was conducted using Mann-Whitney, Chi-squared and receiver operating characteristics (ROC). Logistic regression was conducted using P-POSSUM variables with and without the addition of PML3. RESULTS: Nine-hundred and forty-four over-65 s underwent emergency laparotomy from three United Kingdom hospitals were included. Median age was 76 years (IQR 70-82 years). Inpatient mortality was 21.9%, 30-day mortality was 16.3% and 90-day mortality was 20.7%. PML3 less than 0.39 for males and 0.31 for females indicated significantly worse outcomes (inpatient mortality 68% vs 5.6%, 30-day mortality 50.6% vs 4.0%,90-day mortality 64% vs 5.2%, p < 0.0001). PML3 was independently associated with mortality in multivariate analysis (p < 0.0001). Addition of PML3 to P-POSSUM variables improved area under the curve (AUC) on ROC analysis for inpatient mortality (P-POSSUM:0.78 vs P-POSSUM + PML3:0.917), 30-day mortality(P-POSSUM:0.802 vs P-POSSUM + PML3: 0.91) and 90-day mortality (P-POSSUM:0.79 vs P-POSSUM + PML3: 0.91). CONCLUSION: PML3 is an accurate predictor of mortality in over-65 s undergoing emergency laparotomy. Addition of PML3 to POSSUM appears to improve mortality risk prediction.

Proton beam therapy in rectal cancer: A systematic review and meta-analysis.

INTRODUCTION: Locally advanced rectal cancer is often treated with neoadjuvant chemoradiotherapy and surgery. Radiotherapy carries significant risk of toxicity to organs at risk (OAR). Proton beam therapy (PBT) has demonstrated to be effective in other cancers, delivering equivalent dosimetric radiation but with the benefit of improved sparing of OAR. This review compares dosimetric irradiation of OAR and oncological outcomes for PBT versus conventional photon-based radiotherapy in locally advanced rectal cancer. METHODS: An electronic literature search was performed for studies with comparative cohorts receiving proton beam therapy and photon-based radiotherapy for rectal cancer. RESULTS: Eight articles with a total of 127 patients met the inclusion criteria. There was significantly less irradiated small bowel with PBT compared to three-dimensional conformal radiation therapy (3DCRT) and intensity-modulated radiation therapy (IMRT) (MD -17.01, CI [-24.06, -9.96], p < 0.00001 and MD -6.96, CI [-12.99, -0.94], p = 0.02, respectively). Similar dosimetric results were observed for bladder and pelvic bone marrow. Three studies reported clinical and oncological results for PBT in recurrent rectal cancer with overall survival reported as 43 %, 68 % and 77.2 %, and one study in primary rectal cancer with 100 % disease free survival. CONCLUSION: PBT treatment plans revealed significantly less irradiation of OAR for rectal cancer compared to conventional photon-based radiotherapy. Trials for recurrent rectal cancer and PBT have shown promising results. There are currently no ongoing clinical trials for primary rectal cancer and PBT. More research is required to validate its potential role in dose escalation, higher complete response rate and organ preservation without increasing toxicity.

Targeting Acid Ceramidase to Improve the Radiosensitivity of Rectal Cancer.

Previous work utilizing proteomic and immunohistochemical analyses has identified that high levels of acid ceramidase (AC) expression confers a poorer response to neoadjuvant treatment in locally advanced rectal cancer. We aimed to assess the radiosensitising effect of biological and pharmacological manipulation of AC and elucidate the underlying mechanism. AC manipulation in three colorectal cancer cell lines (HT29, HCT116 and LIM1215) was achieved using siRNA and plasmid overexpression. Carmofur and a novel small molecular inhibitor (LCL521) were used as pharmacological AC inhibitors. Using clonogenic assays, we demonstrate that an siRNA knockdown of AC enhanced X-ray radiosensitivity across all colorectal cancer cell lines compared to a non-targeting control siRNA, and conversely, AC protein overexpression increased radioresistance. Using CRISPR gene editing, we also generated AC knockout HCT116 cells that were significantly more radiosensitive compared to AC-expressing cells. Similarly, two patient-derived organoid models containing relatively low AC expression were found to be comparatively more radiosensitive than three other models containing higher levels of AC. Additionally, AC inhibition using carmofur and LCL521 in three colorectal cancer cell lines increased cellular radiosensitivity. Decreased AC protein led to significant poly-ADP ribose polymerase-1 (PARP-1) cleavage and apoptosis post-irradiation, which was shown to be executed through a p53-dependent process. Our study demonstrates that expression of AC within colorectal cancer cell lines modulates the cellular response to radiation, and particularly that AC inhibition leads to significantly enhanced radiosensitivity through an elevation in apoptosis. This work further solidifies AC as a target for improving radiotherapy treatment of locally advanced rectal cancer.