CD44v3 is a marker of invasive cancer stem cells driving metastasis in gastric carcinoma.

BACKGROUND: Cancer stem cells (CSCs) are at the origin of tumour initiation and progression in gastric adenocarcinoma (GC). However, markers of metastasis-initiating cells remain unidentified in GC. In this study, we characterized CD44 variants expressed in GC and evaluated the tumorigenic and metastatic properties of CD44v3+ cells and their clinical significance in GC patients. METHODS: Using GC cell lines and patient-derived xenografts, we evaluated CD44+ and CD44v3+ GC cells molecular signature and their tumorigenic, chemoresistance, invasive and metastatic properties, and expression in patients-derived tissues. RESULTS: CD44v3+ cells, which represented a subpopulation of CD44+ cells, were detected in advanced preneoplastic lesions and presented CSCs chemoresistance and tumorigenic properties in vitro and in vivo. Molecular and functional analyses revealed two subpopulations of gastric CSCs: CD44v3+ CSCs with an epithelial-mesenchymal transition (EMT)-like signature, and CD44+/v3- CSCs with an epithelial-like signature; both were tumorigenic but CD44v3+ cells showed higher invasive and metastatic properties in vivo. CD44v3+ cells detected in the primary tumours of GC patients were associated with a worse prognosis. CONCLUSION: CD44v3 is a marker of a subpopulation of CSCs with metastatic properties in GC. The identification of metastasis-initiating cells in GC represents a major advance for further development of anti-metastatic therapeutic strategies.

Management of Pathogenic CDH1 Variant Carriers Within the FREGAT Network: A Multicentric Retrospective Study.

OBJECTIVE: To describe the management of pathogenic CDH1 variant carriers (pCDH1vc) within the FREGAT (FRench Eso-GAsTric tumor) network. Primary objective focused on clinical outcomes and pathological findings, Secondary objective was to identify risk factor predicting postoperative morbidity (POM). BACKGROUND: Prophylactic total gastrectomy (PTG) remains the recommended option for gastric cancer risk management in pCDH1vc with, however, endoscopic surveillance as an alternative. METHODS: A retrospective observational multicenter study was carried out between 2003 and 2021. Data were reported as median (interquartile range) or as counts (proportion). Usual tests were used for univariate analysis. Risk factors of overall and severe POM (ie, Clavien-Dindo grade 3 or more) were identified with a binary logistic regression. RESULTS: A total of 99 patients including 14 index cases were reported from 11 centers. Median survival among index cases was 12.0 (7.6-16.4) months with most of them having peritoneal carcinomatosis at diagnosis (71.4%). Among the remaining 85 patients, 77 underwent a PTG [median age=34.6 (23.7-46.2), American Society of Anesthesiologists score 1: 75%] mostly via a minimally invasive approach (51.9%). POM rate was 37.7% including 20.8% of severe POM, with age 40 years and above and low-volume centers as predictors ( P =0.030 and 0.038). After PTG, the cancer rate on specimen was 54.5% (n=42, all pT1a) of which 59.5% had no cancer detected on preoperative endoscopy (n=25). CONCLUSIONS: Among pCDH1vc, index cases carry a dismal prognosis. The risk of cancer among patients undergoing PTG remained high and unpredictable and has to be balanced with the morbidity and functional consequence of PTG.

Neoadjuvant treatment for borderline resectable pancreatic adenocarcinoma is associated with higher R0 rate compared to upfront surgery.

BACKGROUND: Neoadjuvant treatment (NAT) is debated for borderline resectable pancreatic cancer (BRPC). This retrospective study assessed the impact of NAT on R0 rate and survival for BRPC patients in comparison with upfront surgery (US). MATERIAL AND METHODS: Between 2010 and 2017 patient records for all consecutive patients treated for BRPC according to NCCN 2017 were reviewed. The endpoints analysed were R0 rate, recurrence-free-survival (RFS) and overall survival (OS). RESULTS: Seventy-nine patients were included: 63 (79.7%) patients received NAT and 16 (20.3%) were upfront operated. NAT consisted in FOLFIRINOX (median cycles: 5, range 4-8) followed by chemoradiation (n = 55, 87.3%, median dose: 54 Gy). Thirty-nine (61.9%) patients had resection. R0 rate was higher in the NAT group considering a margin clearance of 0 mm (94.9%) or 1 mm (89.7%) compared to the US group (68.8% and 43.8% respectively). In the whole population, median RFS was 12.6 [95%CI: 10.5-22.1] in the NAT group vs 7.7 [95%CI: 4.4-14] months in the US group (p < 0.01). Median OS was 29.0 [95%CI: 23.5-63.1] and 27.2 [95%CI: 11.6-38.8] months in the NAT and US groups respectively (p = 0.06). In operated patients the NAT group achieved better RFS and OS than the US group (p < 0.01 for both). In multivariate analysis NAT, surgical resection and age <65 (p < 0.01 for both) were prognostic of RFS. NAT, surgical resection and adjuvant chemotherapy were prognostic of OS (p < 0.05 for all). In operated patients (n = 55) multivariate analysis showed that N1 status was associated with decreased RFS; age < 65 and NAT were associated with a longer RFS. Receiving a NAT, an adjuvant chemotherapy and achieving a ypT0-1N0 status were associated with better OS. NAT was well tolerated with 14.3% grade ≥ 3 toxicities. CONCLUSION: NAT permitted a high R0 rate with a 0- or 1-mm clearance margin and was associated with better RFS and OS for patients with BRPC.

Colonic MicroRNA Profiles, Identified by a Deep Learning Algorithm, That Predict Responses to Therapy of Patients With Acute Severe Ulcerative Colitis.

BACKGROUND & AIMS: Acute severe ulcerative colitis (ASUC) is a life-threatening condition managed with intravenous steroids followed by infliximab, cyclosporine, or colectomy (for patients with steroid resistance). There are no biomarkers to identify patients most likely to respond to therapy; ineffective medical treatment can delay colectomy and increase morbidity and mortality. We aimed to identify biomarkers of response to medical therapy for patients with ASUC. METHODS: We performed a retrospective analysis of 47 patients with ASUC, well characterized for their responses to steroids, cyclosporine, or infliximab, therapy at 2 centers in France. Fixed colonic biopsies, collected before or within the first 3 days of treatment, were used for microarray analysis of microRNA expression profiles. Deep neural network-based classifiers were used to derive candidate biomarkers for discriminating responders from non-responders to each treatment and to predict which patients would require colectomy. Levels of identified microRNAs were then measured by quantitative PCR analysis in a validation cohort of 29 independent patients-the effectiveness of the classification algorithm was tested on this cohort. RESULTS: A deep neural network-based classifier identified 9 microRNAs plus 5 clinical factors, routinely recorded at time of hospital admission, that associated with responses of patients to treatment. This panel discriminated responders to steroids from non-responders with 93% accuracy (area under the curve, 0.91). We identified 3 algorithms, based on microRNA levels, that identified responders to infliximab vs non-responders (84% accuracy, AUC = 0.82) and responders to cyclosporine vs non-responders (80% accuracy, AUC = 0.79). CONCLUSION: We developed an algorithm that identifies patients with ASUC who respond vs do not respond to first- and second-line treatments, based on microRNA expression profiles in colon tissues.

[Technical recommendations and best practice guidelines for May-Grünwald-Giemsa staining: literature review and insights from the quality assurance]. / Recommandations techniques et règles de bonne pratique pour la coloration de May-Grünwald-Giemsa : revue de la littérature et apport de l'assurance qualité.

May-Grünwald-Giemsa (MGG) stain is a Romanowsky-type, polychromatic stain as those of Giemsa, Leishman and Wright. Apart being the reference method of haematology, it has become a routine stain of diagnostic cytopathology for the study of air-dried preparations (lymph node imprints, centrifuged body fluids and fine needle aspirations). In the context of their actions of promoting the principles of quality assurance in cytopathology, the French Association for Quality Assurance in Anatomic and Cytologic Pathology (AFAQAP) and the French Society of Clinical Cytology (SFCC) conducted a proficiency test on MGG stain in 2013. Results from the test, together with the review of literature data allow pre-analytical and analytical steps of MGG stain to be updated. Recommendations include rapid air-drying of cell preparations/imprints, fixation using either methanol or May-Grünwald alone for 3-10minutes, two-step staining: 50% May-Grünwald in buffer pH 6.8 v/v for 3-5minutes, followed by 10% buffered Giemsa solution for 10-30minutes, and running water for 1-3minutes. Quality evaluation must be performed on red blood cells (RBCs) and leukocytes, not on tumour cells. Under correct pH conditions, RBCs must appear pink-orange (acidophilic) or buff-coloured, neither green nor blue. Leukocyte cytoplasm must be almost transparent, with clearly delineated granules. However, staining may vary somewhat and testing is recommended for automated methods (slide stainers) which remain the standard for reproducibility. Though MGG stain remains the reference stain, Diff-Quik(®) stain can be used for the rapid evaluation of cell samples.

Locally advanced pancreatic adenocarcinoma: reassessment of response with CT after neoadjuvant chemotherapy and radiation therapy.

PURPOSE: To prospectively evaluate the utility of computed tomography (CT) for determination of tumor response and prediction of resectability after neoadjuvant combined chemotherapy and radiation therapy (CRT) in patients with nonmetastatic locally advanced pancreatic cancer. MATERIALS AND METHODS: This study received institutional review board approval, and all participants provided written informed consent. Consecutive patients with cephalic locally advanced pancreatic cancer who underwent surgical exploration and/or resection following neoadjuvant CRT were prospectively enrolled from June 2009 to May 2013. Two radiologists independently analyzed the baseline and post-CRT CT scans for the size, attenuation, and circumferential vascular contacts of the tumor. Associations between the postoperative histologic grade of the tumor response (pTNM) and the clinical, biologic, and CT criteria were assessed by using Spearman correlation coefficients. CT criteria related to the presence of complete (ie, R0) resection were assessed by using logistic regression. RESULTS: Forty-seven patients were included, 33 with an R0 resection and 14 with positive margins (ie, R1) or no resection. Variables demonstrating a significant correlation with the histologic tumor classification of tumor response were post-CRT carbohydrate antigen 19-9 level (r = 0.46), post-CRT largest tumor axis (r = 0.44), post-CRT sum of the largest and smallest tumor axes (r = 0.46), change in the largest axis (r = -0.31), change in the sum of the largest and smallest axes (r = -0.39), change in superior mesenteric vein (SMV) and/or portal vein (hereafter, SMV/portal vein) contact (r = -0.38), and post-CRT superior mesenteric artery contact (r = 0.34). Partial regression of tumor contact with the SMV/portal vein was associated in all cases with R0 resection (10 of 10 patients, positive predictive value = 100%), and partial regression of tumor contact with any peripancreatic vascular axis was associated with R0 resection in 91% of cases (20 of 22 patients, positive predictive value = 91%). Persistence of SMV/portal vein stenosis after CRT was not predictive of R1 resection. CONCLUSION: Partial regression of tumor-vessel contact indicates suitability for surgical exploration, irrespective of the degree of decrease in tumor size or the degree of residual vascular involvement.

Droplet digital polymerase chain reaction detection of KRAS mutations in pancreatic FNA samples: Technical and practical aspects for routine clinical implementation.

BACKGROUND: Pancreatic adenocarcinoma (PDAC) is associated with a 5-year survival rate of less than 6%, and current treatments have limited efficacy. The diagnosis of PDAC is mainly based on a cytologic analysis of endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) samples. However, the collected specimens may prove noncontributory in a significant number of cases, delaying patient management and treatment. The combination of EUS-FNA sample examination and KRAS mutation detection can improve the sensitivity for diagnosis. In this context, the material used for molecular analysis may condition performance. METHODS: The authors prospectively compared the performance of cytologic analysis combined with a KRAS droplet digital polymerase chain reaction (ddPCR) assay for PDAC diagnosis using either conventional formalin-fixed, paraffin-embedded cytologic samples or needle-rinsing fluids. RESULTS: Molecular testing of formalin-fixed, paraffin-embedded cytologic samples was easier to set up, but the authors observed that the treatment of preanalytic samples, in particular the fixation process, drastically reduced ddPCR sensitivity, increasing the risk of false-negative results. Conversely, the analysis of dedicated, fresh needle-rinsing fluid samples appeared to be ideal for ddPCR analysis; it had greater sensitivity and was easily to implement in clinical use. In particular, fluid collection by the endoscopist, transportation to the laboratory, and subsequent freezing did not affect DNA quantity or quality. Moreover, the addition of KRAS mutation detection to cytologic examination improved diagnosis performance, regardless of the source of the sample. CONCLUSIONS: Considering all of these aspects, the authors propose the use of an integrated flowchart for the KRAS molecular testing of EUS-FNA samples in clinical routine.

Reducing reflex first-line prescriptions in a surgical pathology laboratory: toward sustainable practice with positive economic and clinical effects.

In surgical pathology departments, reflex first-line techniques (RFLTs) are aimed at reducing workloads and addressing recent shortages of medical personnel. However, the impacts thereof on economic and diagnostic factors have been poorly addressed. Also, in the era of global warming, environmental considerations are crucial. This study assessed the economic and diagnostic efficacies of routine pathological RFLT and the quality of care and sustainability. Ten RFLTs of the Bordeaux University Hospital pathology department (six special stains, one cytology technique, and three immunohistochemical tests) were studied. First, a retrospective economic analysis evaluated the average cost of these RFLTs per slide and per year. Second, diagnostic relevance was prospectively surveyed. Third, the effects of changes made were analyzed over 2 years. The ten RFLTs were associated with average annual costs of €46,708. Diagnostic relevance analysis indicated that most stains were unnecessary; only 17% were requested as second-line techniques. Elimination of 7/10 tests afforded annual cost savings of €22,522 and reduced the workload by 5568 tests/year, without compromising the workflow or diagnostic quality. Seven of ten RFLTs could be eliminated without compromising diagnostic quality or the workflow. This afforded not only financial benefits but also positive social and environmental impacts. We offer valuable insights into appropriate practices in surgical pathology laboratories. Collaboration between the medical and technical teams was crucial; other healthcare sectors would also benefit from our approach.

Targeting metastasis-initiating cancer stem cells in gastric cancer with leukaemia inhibitory factor.

Gastric cancer's (GC) bad prognosis is usually associated with metastatic spread. Invasive cancer stem cells (CSC) are considered to be the seed of GC metastasis and not all CSCs are able to initiate metastasis. Targeting these aggressive metastasis-initiating CSC (MIC) is thus vital. Leukaemia inhibitory factor (LIF) is hereby used to target Hippo pathway oncogenic members, found to be induced in GC and associated with CSC features. LIF-treated GC cell lines, patient-derived xenograft (PDX) cells and/or CSC tumourspheres underwent transcriptomics, laser microdissection-associated proteomics, 2D and 3D invasion assays and in vivo xenograft in mice blood circulation. LIFR expression was analysed on tissue microarrays from GC patients and in silico from public databases. LIF-treated cells, especially CSC, presented decreased epithelial to mesenchymal transition (EMT) phenotype and invasion capacity in vitro, and lower metastasis initiation ability in vivo. These effects involved both the Hippo and Jak/Stat pathways. Finally, GC's high LIFR expression was associated with better clinical outcomes in patients. LIF treatment could thus represent a targeted anti-CSC strategy to fight against metastatic GC, and LIFR detection in primary tumours could constitute a potential new prognosis marker in this disease.

FGFR3 has tumor suppressor properties in cells with epithelial phenotype.

BACKGROUND: Due to frequent mutations in certain cancers, FGFR3 gene is considered as an oncogene. However, in some normal tissues, FGFR3 can limit cell growth and promote cell differentiation. Thus, FGFR3 action appears paradoxical. RESULTS: FGFR3 expression was forced in pancreatic cell lines. The receptor exerted dual effects: it suppressed tumor growth in pancreatic epithelial-like cells and had oncogenic properties in pancreatic mesenchymal-like cells. Distinct exclusive pathways were activated, STATs in epithelial-like cells and MAP Kinases in mesenchymal-like cells. Both FGFR3 splice variants had similar effects and used the same intracellular signaling. In human pancreatic carcinoma tissues, levels of FGFR3 dropped in tumors. CONCLUSION: In tumors from epithelial origin, FGFR3 signal can limit tumor growth, explaining why the 4p16.3 locus bearing FGFR3 is frequently lost and why activating mutations of FGFR3 in benign or low grade tumors of epithelial origin are associated with good prognosis. The new hypothesis that FGFR3 can harbor both tumor suppressive and oncogenic properties is crucial in the context of targeted therapies involving specific tyrosine kinase inhibitors (TKIs). TKIs against FGFR3 might result in adverse effects if used in the wrong cell context.

Quality indicators for colorectal cancer surgery and care according to patient-, tumor-, and hospital-related factors.

BACKGROUND: Colorectal cancer (CRC) care has improved considerably, particularly since the implementation of a quality of care program centered on national evidence-based guidelines. Formal quality assessment is however still needed. The aim of this research was to identify factors associated with practice variation in CRC patient care. METHODS: CRC patients identified from all cancer centers in South-West France were included. We investigated variations in practices (from diagnosis to surgery), and compliance with recommended guidelines for colon and rectal cancer. We identified factors associated with three colon cancer practice variations potentially linked to better survival: examination of ≥ 12 lymph nodes (LN), non-use and use of adjuvant chemotherapy for stage II and stage III patients, respectively. RESULTS: We included 1,206 patients, 825 (68%) with colon and 381 (32%) with rectal cancer, from 53 hospitals. Compliance was high for resection, pathology report, LN examination, and chemotherapy use for stage III patients. In colon cancer, 26% of stage II patients received adjuvant chemotherapy and 71% of stage III patients. 84% of stage US T3T4 rectal cancer patients received pre-operative radiotherapy. In colon cancer, factors associated with examination of ≥ 12 LNs were: lower ECOG score, advanced stage and larger hospital volume; factors negatively associated were: left sided tumor location and one hospital district. Use of chemotherapy in stage II patients was associated with younger age, advanced stage, emergency setting and care structure (private and location); whereas under-use in stage III patients was associated with advanced age, presence of comorbidities and private hospitals. CONCLUSIONS: Although some changes in practices may have occurred since this observational study, these findings represent the most recent report on practices in CRC in this region, and offer a useful methodological approach for assessing quality of care. Guideline compliance was high, although some organizational factors such as hospital size or location influence practice variation. These factors should be the focus of any future guideline implementation.

[Thyroid cytopathology: Bethesda System 2010]. / Cytopathologie thyroïdienne: le système de Bethesda 2010.

When considering the number of thyroid fine-needle aspirations performed not only in France but also in the world, then thyroid pathology, as well as Pap smears, represents the main pathology based on cytological diagnoses. Decisions of clinical follow-up or of surgical treatments will be done based on the cytological results. Until recently, neither official nor consensual terminology has been adopted despite classifications by experts having been proposed. The Bethesda terminology, published in 2010, in English, is linked with a percentage of risk cancer for each of its diagnostic category and with a suitable treatment. Furthermore, this terminology allows the possibility of standardized management for the patients and the opportunity of further assessment of the cytological diagnoses. Therefore, the French Society of Clinical Cytology has decided to publish this official French version of the Bethesda terminology. Its use is highly recommended.

[KRAS analysis management: Process and delays]. / Prise en charge d'une analyse KRAS : processus et délais.

INTRODUCTION: The prescription of some anti-cancer therapies is now based on the detection of specific genetic alterations that should be determined as early as possible not to put patients at a disadvantage. In 2009, the 'Aquitaine platform of molecular tumour genetics' (PGMC) developed a programme to evaluate and to improve the organisation of molecular cancer analyses, particularly the analysis of the KRAS gene. The objective was to describe the analysis process, the organization of pathology laboratories and the delays between the different phases of the process. METHODS: We established a working group to describe the different steps between the prescription of molecular biology analyses and the analysis report. A retrospective study based on the first quarter of 2009 allowed us to measure management delays. In addition, a pathology laboratory organisational questionnaire allowed us to identify organisational features hindering rapid delivery. RESULTS: The median delay between the analysis prescription and the results was 15 days (range: 7-78 days). Practices explaining longer delays were highlighted not only in the pathology laboratories (for example, pending of the prescription before sending the analysis, waiting for several cases before sending the material, sample slicing, sending by standard mail), but also within the PGMC (for example, sample testing by another technique or new extraction for non-contributory samples). CONCLUSION: The results of this study emphasise the necessity of speeding up pre-analytical phases, and of creating an electronic procedure and regional facilities in order to provide results more rapidly to clinicians.

KRAS gene mutation quantification in the resection or venous margins of pancreatic ductal adenocarcinoma is not predictive of disease recurrence.

Pancreatic ductal adenocarcinoma (PDAC) patients eligible for curative surgery undergo unpredictable disease relapse. Even patients with a good pathological response after neoadjuvant treatment (NAT) remain susceptible to recurrent PDAC. Molecular analysis of R0 margins may identify patients with a worse prognosis. The molecular status of mutant KRAS (exon 2, codon 12/13) was analysed retrospectively by digital droplet PCR in tumour areas, venous and resection margins of resected tumours, either undergoing up-front surgery (UFS) or after NAT with a good pathological response. Expectedly, tumour tissues or remnants from patients who underwent NAT presented lower KRAS mutant allele frequencies (MAF) than patients eligible for UFS. Similarly, ypT1 tumour MAFs were greater than the ypT0 tumour remnant MAFs in the NAT group. Mutant KRAS status in margins did not distinguish NAT subgroups. It was not predictive of shorter recurrence-free or overall survival within or between groups. KRAS-double negativity in both venous and resection margins did not identify patients with a better prognosis, regardless of the groups. The cohorts 'sizes were small due to limited numbers of patients meeting the inclusion criteria, but KRAS-positivity or MAFs in resection and venous margins did not carry prognostic value. Comparison of margins from good versus bad responders receiving NAT may provide better clinical value.

18F-FDG PET for the identification of adrenocortical carcinomas among indeterminate adrenal tumors at computed tomography scanning.

BACKGROUND: 18F-fluorodeoxyglucose positron emission tomography (18F-FDG PET) has been proposed for the evaluation of adrenal tumors. However, only scarce data are available to evaluate its usefulness for the identification of primary adrenal carcinomas in patients with no previous history of cancer and equivocal tumors on computed tomography (CT) scan. The objective of the present study was to evaluate the diagnostic performance of 18F-FDG-PET to predict malignancy in such patients. METHODS AND PATIENTS: This was a retrospective study carried out from 2006 to 2009 in a single university hospital center. Twenty-three consecutive patients without previous history of cancer investigated for adrenal tumors without features of benign adrenocortical adenoma on CT scan but no obvious ACC underwent 18F-FDG PET. All patients underwent adrenalectomy because of CT scan characteristics regardless of the results of 18F-FDG PET. The ratio of maxSUV adrenal tumor on maxSUV liver (adrenal/liver maxSUV ratio) during 18F-FDG PET was compared to Weiss pathological criteria. RESULTS: Seventeen patients had an adrenal adenoma, 2 had small size adrenal carcinomas (<5 cm), 1 had an angiosarcoma, and 3 had noncortical benign lesions. An adrenal/liver maxSUV ratio above 1.6 provided 100% sensitivity, 90% specificity, and 100% negative predictive value for the diagnosis of malignant tumor. CONCLUSIONS: Because of its excellent negative predictive value, 18F-FDG-PET may be of help in avoiding unnecessary surgery in patients with non-secreting equivocal tumors at CT scanning and low 18F-FGD uptake.

Can an intraductal papillary mucinous tumor be a potential indicator of concurrent adenocarcinoma of the pancreas?

CONTEXT: Despite the recent progress of diagnostic and therapeutic modalities, survival rates of pancreatic adenocarcinoma remain poor, mainly due to late diagnosis. CASE REPORT: We report the case of a 56-year-old man who was diagnosed with a symptomatic intraductal papillary mucinous tumor of the pancreas located in the uncus. This tumor was associated with a concurrent stenosis of the isthmic pancreatic duct which resulted in a distal dilation. A Whipple procedure was performed. During the procedure, a concomitant adenocarcinoma was diagnosed 2 cm from the primary intraductal papillary mucinous tumor, causing the isthmic stenosis. A second resection was then performed to the left of the pancreatic isthmus, and adjuvant chemotherapy was performed. The patient is well and without any sign of recurrence 7 months after surgery. CONCLUSION: We discuss the possibility that intraductal papillary mucinous tumors may be a "red flag" enabling earlier diagnosis of a concurrent pancreatic adenocarcinoma arising in another area of the pancreas.