Bert A. Bonsing

Causes of fecal and urinary incontinence after total mesorectal excision for rectal cancer based on cadaveric surgery: a study from the Cooperative Clinical Investigators of the Dutch total mesorectal excision trial

PURPOSE Total mesorectal excision (TME) for rectal cancer may result in anorectal and urogenital dysfunction. We aimed to study possible nerve disruption during TME and its consequences for functional outcome. Because the levator ani muscle plays an important role in both urinary and fecal continence, an explanation could be peroperative damage of the nerve supply to the levator ani muscle. METHODS TME was performed on cadaver pelves. Subsequently, the anatomy of the pelvic floor innervation and its relation to the pelvic autonomic innervation and the mesorectum were studied. Additionally, data from the Dutch TME trial were analyzed to relate anorectal and urinary dysfunction to possible nerve damage during TME procedure. RESULTS Cadaver TME surgery demonstrated that, especially in low tumors, the pelvic floor innervation can be damaged. Furthermore, the origin of the levator ani nerve was located in close proximity of the origin of the pelvic splanchnic nerves. Analysis of the TME trial data showed that newly developed urinary and fecal incontinence was present in 33.7% and 38.8% of patients, respectively. Both types of incontinence were significantly associated with each other (P = .027). Low anastomosis was significantly associated with urinary incontinence (P = .049). One third of the patients with newly developed urinary and fecal incontinence also reported difficulty in bladder emptying, for which excessive perioperative blood loss was a significant risk factor. CONCLUSION Perioperative damage to the pelvic floor innervation could contribute to fecal and urinary incontinence after TME, especially in case of a low anastomosis or damage to the pelvic splanchnic nerves.

Magnetic Resonance Imaging Surveillance Detects Early-Stage Pancreatic Cancer in Carriers of a p16-Leiden Mutation

BACKGROUND & AIMS Surveillance of high-risk groups for pancreatic cancer might increase early detection and treatment outcomes. Individuals with germline mutations in p16-Leiden have a lifetime risk of 15% to 20% of developing pancreatic cancer. We assessed the feasibility of detecting pancreatic cancer at an early stage and investigated the outcomes of patients with neoplastic lesions. METHODS Individuals with germline mutations in p16-Leiden (N = 79; 31 male; mean age, 56 years; range, 39-72 years) were offered annual surveillance by magnetic resonance imaging (MRI) and magnetic resonance cholangiopancreatography (MRCP). Those found to have neoplastic lesions were offered options for surgery or intensive follow-up. Individuals found to have possible neoplastic lesions were examined again by MRI/MRCP within 2 to 4 months. RESULTS After a median follow-up period of 4 years (range, 0-10 years), pancreatic cancer was diagnosed in 7 patients (9%). The mean age at diagnosis was 59 years (range, 49-72 years). Three of the tumors were present at the first examination, and 4 were detected after a negative result in the initial examination. All 7 patients had a resectable lesion; 5 underwent surgery, 3 had an R0 resection, and 2 had lymph node metastases. Possible precursor lesions (ie, duct ectasias, based on MRCP) were found in 9 individuals (11%). CONCLUSIONS MRI/MRCP detects small, solid pancreatic tumors and small duct ectasias. Although surveillance increases the rate of resectability, carriers of a p16-Leiden mutation develop aggressive tumors.

Specificity of seven monoclonal antibodies against p53 evaluated with Western blotting, immunohistochemistry, confocal laser scanning microscopy, and flow cytometry

p53 immunostaining of histological sections shows inter- and intratumor variability in distribution and staining intensity which are usually scored semiquantitatively. In order to investigate the variation in p53 expression more accurately and its possible relation to other cellular parameters (e.g., DNA content), we have studied the possibility to measure p53 accumulation by multiparameter flow cytometry. To this end we have evaluated seven, commercially available, monoclonal antibodies (MAbs) against p53 (MAbs 1801, 240, 246, 421, 1620, Do1, and Do7) on five tumor cell lines with known p53 gene status: MCF-7 (wild-type p53 gene), COV362.cl4 and T47d (mutated p53 genes), and SAOS-2 and HL60 (no p53 mRNA). Localization of immunofluorescence was investigated with confocal laser scanning microscopy, immunofluorescence signal intensity with flow cytometry, and antibody specificity with Western blotting. Subsequently, single cell suspensions from two breast carcinomas were flow cytometrically analyzed after triple staining for p53, cytokeratin 8/18, and DNA, and compared to immunohistochemical staining. MAbs Do1 and Do7, and to a lesser extent MAb 421, accurately discriminated p53 positive from p53 negative cell lines. Even at high concentrations these MAbs yielded nuclear immunofluorescence, whereas with MAbs 1801, 240, and 246 strong cytoplasmic signals in both the p53 accumulating and p53 negative cell lines were seen. By using lower antibody concentrations the cytoplasmic immunofluorescence disappeared, but simultaneously the nuclear p53 immunostaining intensity in p53 accumulating cell lines decreased, resulting in false negative nuclei. With MAb 1620 only weak intranuclear spots were obtained in all cell lines tested. Western blotting yielded results with MAbs 1801, Do1, and Do7 in the 53 kD region of the p53 accumulating cell lines. The signal intensity obtained with MAb 1801 was much less compared to MAbs Do1 and Do7. Although all three MAbs are also described as wild-type p53 specific, only MAbs, Do1 and Do7 showed bands in the 53 kD region of cell line MCF-7. With MAb 1801 ascites and MAb 1801 supernatant an additional approximately 80 kD band was present in all cell lines tested, including SAOS-2, indicating cross reactivity of this MAb. Immunohistochemical staining of two clinical breast carcinomas confirmed the results obtained in the cell lines. Multiparameter flow cytometric analysis of these breast carcinomas with MAbs Do1 and Do7 showed intratumor heterogeneity for p53 accumulation, which was independent of DNA index heterogeneity. We conclude that MAbs Do1 and Do7 enable quantitative analysis of p53 accumulation in a multiparameter flow cytometric analysis.

Impact of centralization of pancreatic cancer surgery on resection rates and survival

Centralization of pancreatic surgery has been shown to reduce postoperative mortality. It is unknown whether resection rates and survival have also improved. The aim of this study was to analyse the impact of nationwide centralization of pancreatic surgery on resection rates and long‐term survival.

Benefit of Surveillance for Pancreatic Cancer in High-Risk Individuals: Outcome of Long-Term Prospective Follow-Up Studies From Three European Expert Centers

PURPOSE Pancreatic ductal adenocarcinoma (PDAC) has a poor prognosis. Hereditary factors play a role in the development of PDAC in 3% to 5% of all patients. Surveillance of high-risk groups, may facilitate detection of PDAC at an early stage. The aim of this study was to assess whether surveillance aids detection of early-stage PDAC or precursor lesions (PRLs) and improves the prognosis. PATIENTS AND METHODS Screening outcomes were collected from three European centers that conduct prospective screening in high-risk groups including families with clustering of PDAC (familial pancreatic cancer [FPC]) or families with a gene defect that predisposes to PDAC. The surveillance program consisted of annual magnetic resonance imaging, magnetic resonance cholangiopancreatography, and/or endoscopic ultrasound. RESULTS Four hundred eleven asymptomatic individuals participated in the surveillance programs, including 178 CDKN2A mutation carriers, 214 individuals with FPC, and 19 BRCA1/2 or PALB2 mutation carriers. PDAC was detected in 13 (7.3%) of 178 CDKN2A mutation carriers. The resection rate was 75%, and the 5-year survival rate was 24%. Two CDKN2A mutation carriers (1%) underwent surgical resection for low-risk PRL. Two individuals (0.9%) in the FPC cohort had a pancreatic tumor, including one advanced PDAC and one early grade 2 neuroendocrine tumor. Thirteen individuals with FPC (6.1%) underwent surgical resection for a suspected PRL, but only four (1.9%) had high-risk lesions (ie, high-grade intraductal papillary mucinous neoplasms or grade 3 pancreatic intraepithelial neoplasms). One BRCA2 mutation carrier was found to have PDAC, and another BRCA2 mutation carrier and a PALB2 mutation carrier underwent surgery and were found to have low-risk PRL. No serious complications occurred as consequence of the program. CONCLUSION Surveillance of CDNK2A mutation carriers is relatively successful, detecting most PDACs at a resectable stage. The benefit of surveillance in families with FPC is less evident.

Meta-analysis of the risk for anastomotic leakage, the postoperative mortality caused by leakage in relation to the overall postoperative mortality.

BACKGROUND Availability of anastomotic leakage rates and mortality rates following anastomotic leakage is essential when informing patients with rectal cancer preoperatively. We performed a meta-analysis of studies describing anastomotic leakage and the subsequent postoperative mortality in relation to the overall postoperative mortality after low anterior resection for rectal cancer. METHODS A systematic search was performed of the published literature. Data on the definition and incidence rate of AL, postoperative mortality caused by AL, and overall postoperative mortality were extracted. Data were pooled and a meta-analysis was performed. RESULTS Twenty-two studies with 10,343 patients in total were analyzed. Meta-analysis of the data showed an average AL rate of 9%, postoperative mortality caused by leakage of 0.7% and overall postoperative mortality of 2%. The studies showed variation in incidence, definition and measurement of all outcomes. CONCLUSION We found a considerable overall AL rate and a large contribution of AL to the overall postoperative mortality. The variability of definitions and measurement of AL, postoperative mortality caused by leakage and overall postoperative mortality may hinder providing reliable risk information. Large-scale audit programs may provide accurate and valid risk information which can be used for preoperative decision making.

Allelotype analysis of flow‐sorted breast cancer cells demonstrates genetically related diploid and aneuploid subpopulations in primary tumors and lymph node metastases

Flow cytometric DNA content measurements have demonstrated extensive DNA ploidy heterogeneity in primary breast carcinomas. However, little is known at the molecular level about the clonal relationship between these tumor cell subpopulations, or about the molecular genetic changes associated with aneuploidization. We have used flow cytometric cell sorting to dissect some of this complexity by isolating clonal subpopulations in breast carcinomas for comparative molecular genetic analysis. Clonal subpopulations were isolated from 12 primary breast carcinomas and 5 lymph node metastases from 4 cases based on DNA content and cytokeratin 8/18 labeling. DNA from these clones was screened for allelic imbalances with 92 polymorphic microsatellite markers mapped to 39 different chromosome arms. Diploid and aneuploid populations were concurrently present in 11 out of 12 primary tumors. The DNA ploidy status of primary tumors was identical to that of the related lymph node metastases. Allelic imbalance was present in 10 out of 11 diploid clones (mean, 3.4 ± 4.2). All allelic imbalances observed in the diploid clones recurred in the cognate aneuploid clones, but were, in the latter, accompanied by additional allelic imbalances at other loci and/or chromosome arms (mean, 10.9 ± 5.8). In only two of the four metastatic cases did the allelotypes of metastatic clones show small differences relative to their cognate primary tumors. The primary diploid tumor clone recurred in all lymph node metastases. This study indicates that the majority of allelic imbalances in breast carcinomas are established during generation of DNA ploidy diversity. Recurrence of the allelic imbalances in diploid clones in the aneuploid clones suggests linear tumor progression, whereas the simultaneous presence of early diploid and advanced aneuploid clones in both primary and metastatic tumor sites suggests that acquisition of metastatic propensity can be an early event in the genetic progression of breast cancer. Genes Chromosomes Cancer 28:173–183, 2000.

Image-guided hepatopancreatobiliary surgery using near-infrared fluorescent light

BackgroundImproved imaging methods and surgical techniques have created a new era in hepatopancreatobiliary (HPB) surgery. Despite these developments, visual inspection, palpation, and intraoperative ultrasound remain the most utilized tools during surgery today. This is problematic, though, especially in laparoscopic HPB surgery, where palpation is not possible. Optical imaging using near-infrared (NIR) fluorescence can be used for the real-time assessment of both anatomy (e.g., sensitive detection and demarcation of tumours and vital structures) and function (e.g., assessment of luminal flow and tissue perfusion) during both open and minimally invasive surgeries.MethodsThis article reviews the published literature related to preclinical development and clinical applications of NIR fluorescence imaging during HPB surgery.ResultsNIR fluorescence imaging combines the use of otherwise invisible NIR fluorescent contrast agents and specially designed camera systems, which are capable of detecting these contrast agents during surgery. Unlike visible light, NIR fluorescent light can penetrate several millimetres through blood and living tissue, thus providing improved detectability. Applications of this technique during HPB surgery include tumour imaging in liver and pancreas, and real-time imaging of the biliary tree.ConclusionsNIR fluorescence imaging is a promising new technique that may someday improve surgical accuracy and lower complications.

Near-Infrared Fluorescence Imaging in Patients Undergoing Pancreaticoduodenectomy

Background: Intraoperative visualization of pancreatic tumors has the potential to improve radical resection rates. Intraoperative visualization of the common bile duct and bile duct anastomoses could be of added value. In this study, we explored the use of indocyanine green (ICG) for these applications and attempted to optimize injection timing and dose. Methods: Eight patients undergoing a pancreaticoduodenectomy were injected intravenously with 5 or 10 mg ICG. During and after injection, the pancreas, tumor, common bile duct and surrounding organs were imaged in real time using the Mini-FLARE™ near-infrared (NIR) imaging system. Results: No clear tumor-to-pancreas contrast was observed, except for incidental contrast in 1 patient. The common bile duct was clearly visualized using NIR fluorescence, within 10 min after injection, with a maximal contrast between 30 and 90 min after injection. Patency of biliary anastomoses could be visualized due to biliary excretion of ICG. Conclusion: No useful tumor demarcation could be visualized in pancreatic cancer patients after intravenous injection of ICG. However, the common bile duct and biliary anastomoses were clearly visualized during the observation period. Therefore, these imaging strategies could be beneficial during biliary surgery in cases where the surgical anatomy is aberrant or difficult to identify.

High levels of DNA index heterogeneity in advanced breast carcinomas. Evidence for DNA ploidy differences between lymphatic and hematogenous metastases

Background. The aim of this study was to investigate DNA ploidy status and DNA index heterogeneity of lymphatic and hematogenous metastases of advanced breast carcinomas and the relations among the various tumor sites.