The grade of obesity affects the noninvasive diagnosis of advanced fibrosis in individuals with MASLD.

OBJECTIVE: Metabolic dysfunction-associated steatotic liver disease (MASLD) is closely associated with obesity. We aimed to assess the impact of obesity on the performance of different noninvasive tests, including liver stiffness measurement (LSM) and Agile3+ (A3+), to detect advanced fibrosis (AF) in a population of patients with MASLD encompassing a wide range of BMI values. METHODS: A total of 479 patients with MASLD were consecutively included (Lyon Hepatology Institute). Clinical data and noninvasive tests, including FibroTest, LSM, A3+, Fibrosis-4 (FIB-4), magnetic resonance elastography, and liver biopsies, were collected. AF was determined by a composite endpoint, i.e., histological stage ≥ F3, overt diagnosis of cirrhosis by magnetic resonance elastography, or concordant LSM ≥ 9.6 kPa and FibroTest ≥ F3. RESULTS: The median BMI was 35.0 kg/m2, and the prevalence of AF was 28.6%. Patients with BMI ≥ 35 versus <35 had a lower proportion of AF, i.e., 19.3% versus 38.1% (p < 0.001), but higher indeterminate status for AF (34.2% vs. 15.4%; p < 0.001). In the case of BMI ≥ 35, LSM had lower specificity to rule in AF (77.9%) versus A3+ (90.4%), but A3+ had decreased sensitivity to rule out AF. A sequential LSM/A3+ strategy achieved high specificity to rule in AF and lowered the proportion of indeterminate cases in patients with BMI ≥ 35. CONCLUSIONS: The grade of obesity affects the detection of MASLD-related AF. A sequential use of LSM/A3+ could improve AF detection in patients with BMI ≥ 35.

Phenotypic characterisation of SMAD4 variant carriers.

BACKGROUND: Both hereditary haemorrhagic telangiectasia (HHT) and juvenile polyposis syndrome (JPS) are known to be caused by SMAD4 pathogenic variants, with overlapping symptoms for both disorders in some patients. Additional connective tissue disorders have also been reported. Here, we describe carriers of SMAD4 variants followed in an HHT reference centre to further delineate the phenotype. METHODS: Observational study based on data collected from the Clinical Investigation for the Rendu-Osler Cohort database. RESULTS: Thirty-three participants from 15 families, out of 1114 patients with HHT, had an SMAD4 variant (3%).Regarding HHT, 26 out of 33 participants (88%) had a definite clinical diagnosis based on Curaçao criteria. Complication frequencies were as follows: epistaxis (n=27/33, 82%), cutaneous telangiectases (n=19/33, 58%), pulmonary arteriovenous malformations (n=17/32, 53%), hepatic arteriovenous malformations (AVMs) (n=7/18, 39%), digestive angiodysplasia (n=13/22, 59%). No cerebral AVMs were diagnosed.Regarding juvenile polyposis, 25 out of 31 participants (81%) met the criteria defined by Jass et al for juvenile polyposis syndrome. Seven patients (21%) had a prophylactic gastrectomy due to an extensive gastric polyposis incompatible with endoscopic follow-up, and four patients (13%) developed a digestive cancer.Regarding connective tissue disorders, 20 (61%) had at least one symptom, and 4 (15%) participants who underwent echocardiography had an aortic dilation. CONCLUSION: We describe a large cohort of SMAD4 variant carriers in the context of HHT. Digestive complications are frequent, early and diffuse, justifying endoscopy every 2 years. The HHT phenotype, associating pulmonary and hepatic AVMs, warrants systematic screening. Connective tissue disorders broaden the phenotype associated with SMAD4 gene variants and justify systematic cardiac ultrasound and skeletal complications screening.

Neuropilin 2 and soluble neuropilin 2 in neuroendocrine neoplasms.

Neuropilin 2 (NRP2), a transmembrane non-tyrosine kinase receptor, has been described as a potential critical player in the tumourigenesis of several solid cancers and particularly in neuroendocrine neoplasms (NENs). A soluble form of NRP2 (sNRP2) has been previously described and corresponds to a truncated splice isoform. Its prognostic value has never been studied in NEN. NRP2 expression was studied by immunochemistry on tissue microarrays (n = 437) and on circulating tumour cells (CTCs, n = 5 patients with neuroendocrine carcinoma, NEC). We described the levels of sNRP2 in 229 patients with NEN using the ELISA method to identify the factors associated with sNRP2 levels and to evaluate its prognostic role; 90 blood donors represented the healthy control group. NRP2 was found in 97% of neuroendocrine tumours (396/410) and in 74% of NEC (20/27). NRP2 was also expressed in CTC of all the studied patients. The receiver operating characteristic (ROC) analysis showed that sNRP2 had a weak capacity to discriminate between NEN patients and healthy controls (area under curve (AUC) = 0.601, P = 0.053). Abnormal sNRP2 levels were associated with inflammatory syndrome, bone and peritoneal metastases, and abnormal chromogranin A levels. Patients with high sNRP2 levels (sNRP2Q3-Q4) had significantly poorer overall survival in multivariate analysis (HR 0.16, 95% CI (0.04-0.67), P = 0.015). In conclusion, the present study found that sNRP2 and NRP2 could represent a new prognostic biomarker and a therapeutic target, respectively, particularly in aggressive NEN.

Histopronostic factors in superficial colorectal adenocarcinomas treated by endoscopy: reproducibility and impact of immunohistochemistry and digital pathology.

Endoscopic dissection is the first-choice treatment for superficial pT1 colorectal adenocarcinoma (sCRC). Complementary surgery decision is influenced by histopronostic factors. Prognostic significance and reproducibility of each factor are not well established. The role of immunohistochemistry (IHC) and digital pathology in this context is unknown. Our aims were (1) to evaluate each histopronostic factor reproducibility comparing HES and IHC ± digital pathology and (2) to evaluate how the different techniques would affect indications for additional surgery. We performed a single-centre retrospective study of 98 patients treated between 2010 and 2019 in Hospices Civils de Lyon, France. We analyzed physical or digital slides of HES and keratin/desmin immunostaining of 98 sCRC dissection specimens. Three pathologists evaluate the histopronostic factors including submucosal invasion depth (SMI) measured using different recommended methods. Assessment of SMI with Ueno or JSCCR methods showed good to excellent interobserver reproducibility (IOR) (ICCs of 0.858 to 0.925) using HES staining and IHC. Assessment of budding on HES sections was poorly reproducible compared to IHC which exhibit moderate IOR (κ = 0.714). IHC increased high-grade budding detection. For lymphovascular invasion and poor differentiation, the IOR was poor (κ = 0.141, 0.196 and 0.313 respectively). IHC gave a better reproducibility for further treatment indication according to JSCCR criteria (κ = 0.763) or forthcoming European guidelines (κ = 0.659). Digital pathology was equivalent to the microscope for all analyses. Histopronostic factor reproducibility in sCRC is moderate. Immunohistochemistry may facilitate the evaluation of certain criteria and improve the reproducibility of treatment decisions.

Comprehensive bile acid pool analysis during ex-vivo liver perfusion in a porcine model of ischemia-reperfusion injury.

Bile acids (BA) are key for liver regeneration and injury. This study aims at analyzing the changes in the BA pool induced by ischemia-reperfusion (IRI) and investigates the impact of hypothermic oxygenated perfusion (HOPE) on the BA pool compared to static cold storage (SCS). In a porcine model of IRI, liver grafts underwent 30 min of asystolic warm ischemia followed by 6 h of SCS (n = 6) ± 2 h of HOPE (n = 6) and 2 h of ex-situ warm reperfusion. The BA pool in bile samples was analyzed with liquid chromatography coupled with tandem mass spectrometry. We identified 16 BA and observed significant changes in response to ischemia-reperfusion, which were associated with both protective and injury mechanisms. Second, HOPE-treated liver grafts exhibited a more protective BA phenotype, characterized by a more hydrophilic BA pool compared to SCS. Key BA, such as GlycoCholic Acid, were identified and were associated with a decreased transaminase release and improved lactate clearance during reperfusion. Partial Least Square-Discriminant Analysis revealed a distinct injury profile for the HOPE group. In conclusion, the BA pool changes with liver graft IRI, and preservation with HOPE results in a protective BA phenotype compared to SCS.

Diffusion-Weighted MRI of the Liver in Patients With Chronic Liver Disease: A Comparative Study Between Different Fitting Approaches and Diffusion Models.

BACKGROUND: Diffusion-weighted imaging (DWI) has been considered for chronic liver disease (CLD) characterization. Grading of liver fibrosis is important for disease management. PURPOSE: To investigate the relationship between DWI's parameters and CLD-related features (particularly regarding fibrosis assessment). STUDY TYPE: Retrospective. SUBJECTS: Eighty-five patients with CLD (age: 47.9 ± 15.5, 42.4% females). FIELD STRENGTH/SEQUENCE: 3-T, spin echo-echo planar imaging (SE-EPI) with 12 b-values (0-800 s/mm2 ). ASSESSMENT: Several models statistical models, stretched exponential model, and intravoxel incoherent motion were simulated. The corresponding parameters (Ds , σ, DDC, α, f, D, D*) were estimated on simulation and in vivo data using the nonlinear least squares (NLS), segmented NLS, and Bayesian methods. The fitting accuracy was analyzed on simulated Rician noised DWI. In vivo, the parameters were averaged from five central slices entire liver to compare correlations with histological features (inflammation, fibrosis, and steatosis). Then, the differences between mild (F0-F2) or severe (F3-F6) groups were compared respecting to statistics and classification. A total of 75.3% of patients used to build various classifiers (stratified split strategy and 10-folders cross-validation) and the remaining for testing. STATISTICAL TESTS: Mean squared error, mean average percentage error, spearman correlation, Mann-Whitney U-test, receiver operating characteristic (ROC) curve, area under ROC curve (AUC), sensitivity, specificity, accuracy, precision. A P-value <0.05 was considered statistically significant. RESULTS: In simulation, the Bayesian method provided the most accurate parameters. In vivo, the highest negative significant correlation (Ds , steatosis: r = -0.46, D*, fibrosis: r = -0.24) and significant differences (Ds , σ, D*, f) were observed for Bayesian fitted parameters. Fibrosis classification was performed with an AUC of 0.92 (0.91 sensitivity and 0.70 specificity) with the aforementioned diffusion parameters based on the decision tree method. DATA CONCLUSION: These results indicate that Bayesian fitted parameters may provide a noninvasive evaluation of fibrosis with decision tree. EVIDENCE LEVEL: 1 TECHNICAL EFFICACY: Stage 1.

A Single Preservation Solution for Static Cold Storage and Hypothermic Oxygenated Perfusion of Marginal Liver Grafts: A Preclinical Study.

BACKGROUND: Hypothermic oxygenated perfusion (HOPE) improves outcomes of marginal liver grafts. However, to date, no preservation solution exists for both static cold storage (SCS) and HOPE. METHODS: After 30 min of asystolic warm ischemia, porcine livers underwent 6 h of SCS followed by 2 h of HOPE. Liver grafts were either preserved with a single preservation solution (IGL2) designed for SCS and HOPE (IGL2-Machine Perfusion Solution [MPS] group, n = 6) or with the gold-standard University of Wisconsin designed for for SCS and Belzer MPS designed for HOPE (MPS group, n = 5). All liver grafts underwent warm reperfusion with whole autologous blood for 2 h, and surrogate markers of hepatic ischemia-reperfusion injury (IRI) were assessed in the hepatocyte, cholangiocyte, vascular, and immunological compartments. RESULTS: After 2 h of warm reperfusion, livers in the IGL2-MPS group showed no significant differences in transaminase release (aspartate aminotransferase: 65.58 versus 104.9 UI/L/100 g liver; P = 0.178), lactate clearance, and histological IRI compared with livers in the MPS group. There were no significant differences in biliary acid composition, bile production, and histological biliary IRI. Mitochondrial and endothelial damage was also not significantly different and resulted in similar hepatic inflammasome activation. CONCLUSIONS: This preclinical study shows that a novel IGL2 allows for the safe preservation of marginal liver grafts with SCS and HOPE. Hepatic IRI was comparable with the current gold standard of combining 2 different preservation solutions (University of Wisconsin + Belzer MPS). These data pave the way for a phase I first-in-human study and it is a first step toward tailored preservation solutions for machine perfusion of liver grafts.

Systematic Resection of the Visible Scar After Incomplete Endoscopic Resection of Rectal Neuroendocrine Tumors.

INTRODUCTION: When initial resection of rectal neuroendocrine tumors (r-NETs) is not R0, persistence of local residue could lead to disease recurrence. This study aimed to evaluate the interest of systematic resection of non-R0 r-NET scars. METHODS: Retrospective analysis of all the consecutive endoscopic revisions and resections of the scar after non-R0 resections of r-NETs. RESULTS: A total of 100 patients were included. Salvage endoscopic procedure using endoscopic submucosal dissection or endoscopic full-thickness resection showed an R0 rate of near 100%. Residual r-NET was found in 43% of cases. DISCUSSION: In case of non-R0 resected r-NET, systematic scar resection by endoscopic full-thickness resection or endoscopic submucosal dissection seems necessary.

Dual alcohol and metabolic-related liver disease: Results from a population of liver transplant patients.

BACKGROUND & AIMS: If alcohol-related liver disease (ALD) and nonalcoholic fatty liver disease (NAFLD) are now the two main indications for liver transplantation (LT), it has been recognized that both conditions can coexist in varying degrees and the concept of dual-aetiology fatty liver disease (DAFLD) has been proposed. This retrospective study aimed to evaluate, in a cohort of patients transplanted for ALD and NAFLD, the prevalence of DAFLD before LT and the impact on liver graft outcome. METHODS: From 1990 to 2010, all patients who underwent LT for the so-called ALD or NAFLD in our centre were included. Before LT, DAFLD was defined as patients with a history of excessive alcohol consumption and obesity associated with either diabetes or hypertension. Before LT, patients were separated into three groups: DAFLD, ALD, and NAFLD. Fatty liver graft disease was classified according to the FLIP algorithm. RESULTS: Out of 907, adult LT recipients were identified: 33 DAFLD patients, 333 ALD patients, and 24 NAFLD patients. After LT, ALD patients experienced significantly more alcohol relapse than DAFLD patients, who had twice more post-LT metabolic syndrome. Out of 926, post-LT biopsies, DAFLD patients had significantly more fatty liver graft disease due to metabolic syndrome features than ALD patients. CONCLUSION: Our results support that DAFLD recently emerged as an indication of LT. In the future, this particular population needs to be identified as a specific entity since post-LT outcome on the graft is different from ALD and more similar to NAFLD patients.

Histological features of liver disease development in the Atp7b-/- mouse: a model of Wilson's disease.

AIMS: Wilson's disease (WD) is caused by mutations in the ATP7B gene, resulting in copper accumulation and toxicity in liver and brain tissues. Due to the initial asymptomatic liver involvement, the progression of liver injuries in WD stays primarily unknown. Atp7b-/- knockout mice have been shown to be an appropriate model of WD for liver involvement. METHODS: A total of 138 Atp7b-/- mice were included and separated into five groups according to age as follows: 6, 20, 39 and 50 weeks without treatment, and 50 weeks with copper chelator treatment from 39 to 50 weeks of age and compared with 101 wild-type (WT) mice at the same stages. The evolution of histological liver lesions was analysed and compared between groups. RESULTS: Significant changes were observed in Atp7b-/- mice compared with WT. Copper deposits in hepatocytes appeared as early as 6 weeks but no significant increase over time was observed. Inflammation appeared as early as 6 weeks and progressed henceforth. Lobular and periportal acidophilic bodies appeared after 20 weeks. Significant atypia was also observed at 20 weeks and increased over time to reach a severe stage at 39 weeks. Fibrosis also became apparent at 20 weeks, progressing subsequently to precirrhotic stages at 50 weeks. Copper content, inflammation and fibrosis scores were significantly reduced in the treated group. No bile duct lesions or dysplastic changes were noted. CONCLUSIONS: Copper accumulation leads to progressive changes in Atp7b-/- mice regarding inflammation, fibrosis and atypia. The severity of liver damage is lessened by chelation therapy.

Iterative antibody-induced bile salt export pump deficiency after successive liver transplantations successfully treated with plasmapheresis and rituximab.

Post-transplantation evolution of progressive familial intrahepatic cholestasis type 2 patients can be complicated by antibody-induced bile salt export pump deficiency (AIBD). There is no consensus on its management. We describe a patient who presented two episodes, 9 years apart. The first episode was refractory to plasmapheresis and intravenous immunoglobulin (IVIG) started 2 months after AIBD onset, leading to graft loss. The second episode responded to plasmapheresis, IVIG and rituximab initiated less than 2 weeks after the beginning of symptoms, allowing for long-term recovery. This case suggests that intensive treatment with minimum delay after symptoms onset could sponsor a better evolution.

Necrosis as a strong independent prognostic factor required in the implementation of pathological reporting for pancreatic adenocarcinoma resection specimens.

INTRODUCTION: Pancreatic ductal adenocarcinoma (PDAC) is a major public health issue with an incidence/mortality ratio reaching 98 %. Only 15-20 % of patients with PDAC can undergo surgery. Following PDAC surgical resection, 80 % of patients will experience local or metastatic recurrence of this disease. pTNM staging is the gold standard for risk stratification but is not sufficient to recapitulate the prognosis. Several prognostic factors are known to impact survival after surgery when uncovered during pathological examination. However, necrosis has been poorly studied in pancreatic adenocarcinoma. MATERIALS & METHODS: We retrieved clinical data and reviewed all tumor slides from patients who had a pancreatic surgery between January 2004 and December 2017, in the Hospices Civils de Lyon, to assess the presence of histopathological prognosis factors associated with poor prognosis. RESULTS: 514 patients with complete clinico-pathological description were included. Necrosis was found in 231 PDAC (44.9 %) and had an important impact on overall survival with a double risk of death when present in tumor samples (HR: 1.871, 95 % CI [1.523; 2.299], p < 0.001). When integrated in the multivariate model, necrosis is the only morphological aggressive feature to retain high statistical significance associated with the TNM staging but independently of it. This effect is independent of the preoperative treatment. CONCLUSIONS: Despites improvement in treatment of PDAC, mortality rates remain relatively stable amongst the last years. There is a desperate need to better stratify patients. Here, we report the strong and prognostic impact of necrosis in surgical PDAC samples and encourage pathologists to report its presence in the future.

[Differential diagnostic between hepatocellular carcinoma and hepatoid carcinoma with bone metastasis revelation: What tools can be used?] / Diagnostic différentiel entre un carcinome hépatocellulaire et un carcinome hépatoïde à révélation métastatique osseuse : quels outils sont à disposition ?

Bone metastases of hepatocellular carcinoma (HCC) are rare and disease-revealing bone metastasis are exceptional. Here, we report the case of a 69-year-old man with a cervical vertebral metastasis of hepatocellular carcinoma. Morphological aspect of a metastatic tumor with eosinophilic and polygonal cells raises the question of the differential diagnosis between a localization of a hepatocellular carcinoma or an hepatoid carcinoma, notably when the metastasis is the first clinical manifestation. The morphological aspect by itself does not provide strong enough arguments for diagnosis. Well selected immunohistochemical markers can sometimes help to orientate towards one of the two hypotheses, in particular SALL4 and LIN28 which are in favour of hepatoid carcinoma when both are positive. Finally, as these two entities have different molecular profiles, molecular study can also be helpful to distinguish them. Indeed, HCCs often present TERT promoter, CTNNB1 mutations and IL-6/JAK/STAT pathway activation while hepatoid adenocarcinoma frequently presents chromosome 20 long arm gain. TP53 mutations are found in both entities and are therefore not discriminating. Differential diagnosis is important because the treatment will be that of the primary. Prognostic data for HCC revealed by bone metastasis are scarce, although they seem to be associated with a poor prognosis, with a 1 to 2 months overall survival. There is currently no data for hepatoid adenocarcinoma with bone metastasis.

Hemicolectomy versus appendectomy for patients with appendiceal neuroendocrine tumours 1-2 cm in size: a retrospective, Europe-wide, pooled cohort study.

BACKGROUND: Awareness of the potential global overtreatment of patients with appendiceal neuroendocrine tumours (NETs) of 1-2 cm in size by performing oncological resections is increasing, but the rarity of this tumour has impeded clear recommendations to date. We aimed to assess the malignant potential of appendiceal NETs of 1-2 cm in size in patients with or without right-sided hemicolectomy. METHODS: In this retrospective cohort study, we pooled data from 40 hospitals in 15 European countries for patients of any age and Eastern Cooperative Oncology Group performance status with a histopathologically confirmed appendiceal NET of 1-2 cm in size who had a complete resection of the primary tumour between Jan 1, 2000, and Dec 31, 2010. Patients either had an appendectomy only or an appendectomy with oncological right-sided hemicolectomy or ileocecal resection. Predefined primary outcomes were the frequency of distant metastases and tumour-related mortality. Secondary outcomes included the frequency of regional lymph node metastases, the association between regional lymph node metastases and histopathological risk factors, and overall survival with or without right-sided hemicolectomy. Cox proportional hazards regression was used to estimate the relative all-cause mortality hazard associated with right-sided hemicolectomy compared with appendectomy alone. This study is registered with ClinicalTrials.gov, NCT03852693. FINDINGS: 282 patients with suspected appendiceal tumours were identified, of whom 278 with an appendiceal NET of 1-2 cm in size were included. 163 (59%) had an appendectomy and 115 (41%) had a right-sided hemicolectomy, 110 (40%) were men, 168 (60%) were women, and mean age at initial surgery was 36·0 years (SD 18·2). Median follow-up was 13·0 years (IQR 11·0-15·6). After centralised histopathological review, appendiceal NETs were classified as a possible or probable primary tumour in two (1%) of 278 patients with distant peritoneal metastases and in two (1%) 278 patients with distant metastases in the liver. All metastases were diagnosed synchronously with no tumour-related deaths during follow-up. Regional lymph node metastases were found in 22 (20%) of 112 patients with right-sided hemicolectomy with available data. On the basis of histopathological risk factors, we estimated that 12·8% (95% CI 6·5 -21·1) of patients undergoing appendectomy probably had residual regional lymph node metastases. Overall survival was similar between patients with appendectomy and right-sided hemicolectomy (adjusted hazard ratio 0·88 [95% CI 0·36-2·17]; p=0·71). INTERPRETATION: This study provides evidence that right-sided hemicolectomy is not indicated after complete resection of an appendiceal NET of 1-2 cm in size by appendectomy, that regional lymph node metastases of appendiceal NETs are clinically irrelevant, and that an additional postoperative exclusion of metastases and histopathological evaluation of risk factors is not supported by the presented results. These findings should inform consensus best practice guidelines for this patient cohort. FUNDING: Swiss Cancer Research foundation.

Are protocol graft biopsies after pediatric liver transplantation useful? Experience in a single center over 20 years.

BACKGROUND: The role of protocol liver biopsies (PLB) in the follow-up of pediatric liver transplant recipients remains questionable. This single-center retrospective study aimed to evaluate their clinical impact on the long-term management of pediatric liver transplant recipients. METHODS: We described histopathological lesions and clinical consequences for patient management of PLB performed 1, 5, 10, 15, 20, and 25 years after pediatric liver transplantation (LT). RESULTS: A total of 351 PLB performed on 133 patients between 1992 and 2021 were reviewed. PLB found signs of rejection in 21.7% of cases (76/351), and moderate to severe fibrosis in 26.5% of cases (93/351). Overall, 264 PLB (75.2%) did not cause any changes to patient care. Immunosuppression was enhanced after 63 PLB, including 23 cases of occult rejection. The 1-year PLB triggered significantly more changes, while biopsies at 15, 20, and 25 years produced the lowest rates of subsequent modifications. PLB had a significantly higher probability of inducing therapeutic changes if the patient had abnormal biological or imaging results (odds ratio [OR] 2.82 and 2.06), or a recent history of rejection or bacterial infection (OR 2.22 and 2.03). CONCLUSION: Our results suggest that, although it often does not prompt any treatment changes, PLB could be performed because of its ability to detect silent rejection requiring an increase in immunosuppression. PLB could be carried out 1, 5, and 10 years after LT and then every 10 years in patients with normal biological and imaging results and no recent complications, while other patients could be kept on a 5-year protocol.

Improved NGS-based detection of microsatellite instability using tumor-only data.

Microsatellite instability (MSI) is a molecular signature of mismatch repair deficiency (dMMR), a predictive marker of immune checkpoint inhibitor therapy response. Despite its recognized pan-cancer value, most methods only support detection of this signature in colorectal cancer. In addition to the tissue-specific differences that impact the sensitivity of MSI detection in other tissues, the performance of most methods is also affected by patient ethnicity, tumor content, and other sample-specific properties. These limitations are particularly important when only tumor samples are available and restrict the performance and adoption of MSI testing. Here we introduce MSIdetect, a novel solution for NGS-based MSI detection. MSIdetect models the impact of indel burden and tumor content on read coverage at a set of homopolymer regions that we found are minimally impacted by sample-specific factors. We validated MSIdetect in 139 Formalin-Fixed Paraffin-Embedded (FFPE) clinical samples from colorectal and endometrial cancer as well as other more challenging tumor types, such as glioma or sebaceous adenoma or carcinoma. Based on analysis of these samples, MSIdetect displays 100% specificity and 96.3% sensitivity. Limit of detection analysis supports that MSIdetect is sensitive even in samples with relatively low tumor content and limited microsatellite instability. Finally, the results obtained using MSIdetect in tumor-only data correlate well (R=0.988) with what is obtained using tumor-normal matched pairs, demonstrating that the solution addresses the challenges posed by MSI detection from tumor-only data. The accuracy of MSI detection by MSIdetect in different cancer types coupled with the flexibility afforded by NGS-based testing will support the adoption of MSI testing in the clinical setting and increase the number of patients identified that are likely to benefit from immune checkpoint inhibitor therapy.

SMAD2/3 mediate oncogenic effects of TGF-ß in the absence of SMAD4.

TGF-ß signaling is involved in pancreatic ductal adenocarcinoma (PDAC) tumorigenesis, representing one of the four major pathways genetically altered in 100% of PDAC cases. TGF-ß exerts complex and pleiotropic effects in cancers, notably via the activation of SMAD pathways, predominantly SMAD2/3/4. Though SMAD2 and 3 are rarely mutated in cancers, SMAD4 is lost in about 50% of PDAC, and the role of SMAD2/3 in a SMAD4-null context remains understudied. We herein provide evidence of a SMAD2/3 oncogenic effect in response to TGF-ß1 in SMAD4-null human PDAC cancer cells. We report that inactivation of SMAD2/3 in SMAD4-negative PDAC cells compromises TGF-ß-driven collective migration mediated by FAK and Rho/Rac signaling. Moreover, RNA-sequencing analyses highlight a TGF-ß gene signature related to aggressiveness mediated by SMAD2/3 in the absence of SMAD4. Using a PDAC patient cohort, we reveal that SMAD4-negative tumors with high levels of phospho-SMAD2 are more aggressive and have a poorer prognosis. Thus, loss of SMAD4 tumor suppressive activity in PDAC leads to an oncogenic gain-of-function of SMAD2/3, and to the onset of associated deleterious effects.