Multicenter study on the incidence and treatment of mediastinal leaks after esophagectomy (MuMeLe 2).

BACKGROUND: Management of mediastinal anastomotic leaks (MALs) after Ivor Lewis esophagectomy includes conservative, endoscopic, or surgical management. Endoscopic vacuum therapy (EVAC) is becoming a routine approach for MALs, although the outcomes have not been defined. This study aimed to describe the incidence, treatment, and outcomes of MALs in patients who underwent esophagectomy in 3 Italian high-volume centers that routinely use EVAC for MAL. METHODS: Patients who underwent Ivor Lewis esophagectomy between September 2018 and March 2023 were included. RESULTS: A total of 681 patients underwent Ivor Lewis esophagectomy, of whom 88 had MAL. The MAL rates for open, minimally invasive, and robotic esophagectomies were 11.5%, 13.4%, and 14.8%, respectively. Global and specific 30- and 90-day mortality rates for MAL were 0.9% and 2.1% and 6.8% and 15.9%, respectively. Nonoperative management (NOM) as the primary treatment was chosen for 62 patients. EVAC was the most common NOM (62.9%), and the most common operative management (OM) was anastomotic redo (53.8%). Diversion was the OM for 7 patients, of whom 3 patients died. Primary treatment proved successful in 40 patients. Among them, EVAC alone was successful in 35.9% of patients. Globally, endoscopic treatment, including EVAC, was successful in 79.0% of NOM and 55.7% of MALs. NOM and OM were chosen as secondary treatments for 27 and 10 patients, respectively. Secondary treatment proved successful in 21 patients. CONCLUSION: The incidence of MALs after Ivor Lewis esophagectomy is approximately 13%. Endoscopic techniques have a success rate of almost 80%, with EVAC representing a significant part of this treatment process.

Clinical predictors of postoperative complications in the context of enhanced recovery (ERAS) in patients with esophageal and gastric cancer.

The overall frequency of postoperative complications in patients with esophageal and gastric cancer diverges between studies. We evaluated the frequency and assessed the relationship between complications and demographic and clinical features. For this observational study, data were extracted from the ERAS Registry managed by the University of Verona, Italy. Patients were evaluated and compared for postoperative complications according to the consensus-based classification and the Clavien-Dindo scale. The study population was 877 patients: 346 (39.5%) with esophageal and 531 (60.5%) with gastric cancer; 492 (56.2%) reported one or more postoperative complications, 213 (61.6%) of those with esophageal and 279 (52.5%) of those with gastric cancer. When stratified by consensus-based classification, patients with esophageal cancer reported general postoperative complications more frequently (p < 0.001) than those with gastric cancer, but there was no difference in postoperative surgical complications between the two groups. Multiple logistic regression models revealed an association between postoperative complications and the Charlson Comorbidity Index (adjusted odds ratio [OR] 1.22; 95% confidence interval [CI] 1.08-1.36), operation time (adjusted OR, 1.08; 95% CI 1.00-1.15), and days to solid diet intake (adjusted OR, 1.39; 95% CI 1.20-1.59). Complications in patients with esophageal and gastric cancer are frequent, even in those treated according to ERAS principles, and are often associated with comorbidities, longer operative time, and longer time to solid diet intake.

Mutational analysis of gastrointestinal stromal tumors (GISTs): procedural approach for diagnostic purposes.

BACKGROUND: Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumors in the digestive tract characterized, in the majority of cases, by activating mutations in the KIT (v-kit Hardy-Zuckerman 4 feline sarcoma viral oncogene homolog) or PDGFRA (platelet-derived growth factor receptor, alpha polypeptide) genes. Mutations affecting these tyrosine kinase receptors are also responsible for the mechanisms of primary and secondary drug resistance during the treatment with tyrosine kinase inhibitors. We performed mutational analysis to evaluate the pharmacotherapy susceptibility of GISTs, adopting a comprehensive procedural approach, in order to optimize the identification of mutations that may result in cellular resistance to conventional therapy. MATERIALS AND METHODS: DNA from paraffin-embedded tumor sections from 40 GISTs were analyzed using microdissection, direct sequencing analysis and allelic separation by cloning. RESULTS: KIT mutations were found in 55.0% of the tumor samples. PDGFRA mutations were present in 5.0% of cases. Allelic cloning assay allowed for better definition of the extent of the mutations and clarification of the exact nucleotidic position of complex mutations. CONCLUSION: Our experience suggests that sequential microdissection, direct sequencing and allelic separation by PCR cloning of large variants may improve the approach to mutational analysis and interpretation of sequence data of KIT and PDGFRA in patients with GIST.

A comprehensive procedural approach to genotyping KRAS and BRAF from paraffin embedded tissues for diagnostic purposes.

BACKGROUND: Mutations in the Kirsten Ras 1 (KRAS) and V-Raf Murine Sarcoma Viral Oncogene Homolog B1 (BRAF) genes may be predictive of response to drugs directly linked to the Epidermal Growth Factor Receptor (EGFR) signaling pathway. MATERIALS AND METHODS: A total of 230 samples from patients with metastatic colorectal cancer were analyzed for KRAS exon 1 and 2 and for BRAF exon 15 mutations. DNA from paraffin-embedded tumor sections was analyzed using microdissection, direct sequencing analysis and allelic separation by cloning. RESULTS: KRAS mutations were present in 44.3% of the tumor samples. The mutation frequency at hot-spot codons of exon 1 was 84.2%, whereas non-canonical variants had a frequency of 11.8%. Approximately 4% of the cases exhibited concomitant variations. BRAF mutations were present in 3.9% of the tumor samples. CONCLUSION: Our experience suggests that sequential microdissection, direct sequencing and allelic separation by cloning may improve the approach to mutational analysis of KRAS and BRAF in patients with colorectal cancer.

An AT-rich region in the APC gene may cause misinterpretation of familial adenomatous polyposis molecular screening.

Familial adenomatous polyposis (FAP) is an autosomal-dominant condition mainly due to a mutation of the adenomatous polyposis coli (APC) gene. The present study reports evidence of a technical issue occurring during the mutational analysis of APC exon 4. Genetic conventional direct sequence analysis of a repetitive AT-rich region in the splice acceptor site of APC intron 3 could be misinterpreted as a pathogenetic frameshift result. However, this potential bias may be bypassed adopting a method for random mutagenesis of DNA based on the use of a triphosphate nucleoside analogues mixture. Using this method as a second-level analysis, we also demonstrated the nonpathogenic nature of the variant in the poly A trait in APC exon 4 region (c.423-4delA) that do not result in aberrant splicing of APC exons 3-4; conversely, we did not find a previously reported T deletion/insertion polymorphism.

Preanalytical Procedures for DNA Studies: The Experience of the Interinstitutional Multidisciplinary BioBank (BioBIM).

Preanalytical variables, including the anticoagulants and stabilizing agents, time, storage temperature, and methods of DNA extraction applied to blood samples, may affect quality and quantity of isolated nucleic acids for future genomic applications. Considering the large number of collected samples, standard operating procedure optimization for whole blood preservation before DNA extraction is a crucial step in a biological repository. Moreover, the future application of the biological material may not be known subsequent to its extraction. To define standard operating procedures for whole blood preservation before DNA extraction, we aimed to determine whether different combinations of anticoagulants, blood storage temperatures, and time intervals before storage at -80°C might have an impact on quality and quantity of subsequent extracted DNA. After spectrophotometer quantification, the quality and integrity of DNA were assessed by agarose gel electrophoresis, polymerase chain reaction, and real-time polymerase chain reaction methods. We observed that decrease in DNA recovery during blood storage time was more pronounced at room temperature than at 4°C. Based on our experience, we recommend as anticoagulants of choice sodium citrate and ethylenediaminetetraacetate, whereas sodium citrate theophylline adenosine dipyridamole could represent an alternative choice, excluding a priori lithium heparin and Fluoride-Oxalate. Based on the overall evaluation criteria, we conclude that the procedures necessary to preserve the whole blood before the DNA extraction may have a significant impact on downstream molecular biological applications.