Diagnostic accuracy of endoscopic ultrasonography-guided tissue acquisition prior to resection of pancreatic carcinoma: a nationwide analysis.

INTRODUCTION: Endoscopic ultrasonography guided tissue acquisition (EUS + TA) is used to provide a tissue diagnosis in patients with suspected pancreatic cancer. Key performance indicators (KPI) for these procedures are rate of adequate sample (RAS) and sensitivity for malignancy (SFM). AIM: assess practice variation regarding KPI of EUS + TA prior to resection of pancreatic carcinoma in the Netherlands. PATIENTS AND METHODS: Results of all EUS + TA prior to resection of pancreatic carcinoma from 2014-2018, were extracted from the national Dutch Pathology Registry (PALGA). Pathology reports were classified as: insufficient for analysis (b1), benign (b2), atypia (b3), neoplastic other (b4), suspected malignant (b5), and malignant (b6). RAS was defined as the proportion of EUS procedures yielding specimen sufficient for analysis. SFM was calculated using a strict definition (malignant only, SFM-b6), and a broader definition (SFM-b5+6). RESULTS: 691 out of 1638 resected patients (42%) underwent preoperative EUS + TA. RAS was 95% (range 89-100%), SFM-b6 was 44% (20-77%), and SFM-b5+6 was 65% (53-90%). All centers met the performance target RAS>85%. Only 9 out of 17 met the performance target SFM-b5+6 > 85%. CONCLUSION: This nationwide study detected significant practice variation regarding KPI of EUS + TA procedures prior to surgical resection of pancreatic carcinoma. Therefore, quality improvement of EUS + TA is indicated.

Antibodies Against Immune Checkpoint Molecules Restore Functions of Tumor-Infiltrating T Cells in Hepatocellular Carcinomas.

BACKGROUND & AIMS: Ligand binding to inhibitory receptors on immune cells, such as programmed cell death 1 (PD-1) and cytotoxic T-lymphocyte associated protein 4 (CTLA4), down-regulates the T-cell-mediated immune response (called immune checkpoints). Antibodies that block these receptors increase antitumor immunity in patients with melanoma, non-small-cell lung cancer, and renal cell cancer. Tumor-infiltrating CD4+ and CD8+ T cells in patients with hepatocellular carcinoma (HCC) have been found to be functionally compromised. We analyzed HCC samples from patients to determine if these inhibitory pathways prevent T-cell responses in HCCs and to find ways to restore their antitumor functions. METHODS: We collected HCC samples from 59 patients who underwent surgical resection from November 2013 through May 2017, along with tumor-free liver tissues (control tissues) and peripheral blood samples. We isolated tumor-infiltrating lymphocytes (TIL) and intra-hepatic lymphocytes. We used flow cytometry to quantify expression of the inhibitory receptors PD-1, hepatitis A virus cellular receptor 2 (TIM3), lymphocyte activating 3 (LAG3), and CTLA4 on CD8+ and CD4+ T cells from tumor, control tissue, and blood; we studied the effects of antibodies that block these pathways in T-cell activation assays. RESULTS: Expression of PD-1, TIM3, LAG3, and CTLA4 was significantly higher on CD8+ and CD4+ T cells isolated from HCC tissue than control tissue or blood. Dendritic cells, monocytes, and B cells in HCC tumors expressed ligands for these receptors. Expression of PD-1, TIM3, and LAG3 was higher on tumor-associated antigen (TAA)-specific CD8+ TIL, compared with other CD8+ TIL. Compared with TIL that did not express these inhibitory receptors, CD8+ and CD4+ TIL that did express these receptors had higher levels of markers of activation, but similar or decreased levels of granzyme B and effector cytokines. Antibodies against CD274 (PD-ligand1 [PD-L1]), TIM3, or LAG3 increased proliferation of CD8+ and CD4+ TIL and cytokine production in response to stimulation with polyclonal antigens or TAA. Importantly, combining antibody against PD-L1 with antibodies against TIM3, LAG3, or CTLA4 further increased TIL functions. CONCLUSIONS: The immune checkpoint inhibitory molecules PD-1, TIM3, and LAG3 are up-regulated on TAA-specific T cells isolated from human HCC tissues, compared with T cells from tumor-free liver tissues or blood. Antibodies against PD-L1, TIM3, or LAG3 restore responses of HCC-derived T cells to tumor antigens, and combinations of the antibodies have additive effects. Strategies to block PD-L1, TIM3, and LAG3 might be developed for treatment of primary liver cancer.

GRIN2B encephalopathy: novel findings on phenotype, variant clustering, functional consequences and treatment aspects.

BACKGROUND: We aimed for a comprehensive delineation of genetic, functional and phenotypic aspects of GRIN2B encephalopathy and explored potential prospects of personalised medicine. METHODS: Data of 48 individuals with de novo GRIN2B variants were collected from several diagnostic and research cohorts, as well as from 43 patients from the literature. Functional consequences and response to memantine treatment were investigated in vitro and eventually translated into patient care. RESULTS: Overall, de novo variants in 86 patients were classified as pathogenic/likely pathogenic. Patients presented with neurodevelopmental disorders and a spectrum of hypotonia, movement disorder, cortical visual impairment, cerebral volume loss and epilepsy. Six patients presented with a consistent malformation of cortical development (MCD) intermediate between tubulinopathies and polymicrogyria. Missense variants cluster in transmembrane segments and ligand-binding sites. Functional consequences of variants were diverse, revealing various potential gain-of-function and loss-of-function mechanisms and a retained sensitivity to the use-dependent blocker memantine. However, an objectifiable beneficial treatment response in the respective patients still remains to be demonstrated. CONCLUSIONS: In addition to previously known features of intellectual disability, epilepsy and autism, we found evidence that GRIN2B encephalopathy is also frequently associated with movement disorder, cortical visual impairment and MCD revealing novel phenotypic consequences of channelopathies.

Cumulative sum learning curves guiding multicenter multidisciplinary quality improvement of EUS-guided tissue acquisition of solid pancreatic lesions.

Background and study aims In this study, we evaluated the performance of community hospitals involved in the Dutch quality in endosonography team regarding yield of endoscopic ultrasound (EUS)-guided tissue acquisition (TA) of solid pancreatic lesions using cumulative sum (CUSUM) learning curves. The aims were to assess trends in quality over time and explore potential benefits of CUSUM as a feedback-tool. Patients and methods All consecutive EUS-guided TA procedures for solid pancreatic lesions were registered in five community hospitals between 2015 and  2018. CUSUM learning curves were plotted for overall performance and for performance per center. The American Society of Gastrointestinal Endoscopy-defined key performance indicators, rate of adequate sample (RAS), and diagnostic yield of malignancy (DYM) were used for this purpose. Feedback regarding performance was provided on multiple occasions at regional interest group meetings during the study period. Results A total of 431 EUS-guided TA procedures in 403 patients were included in this study. The overall and per center CUSUM curves for RAS improved over time. CUSUM curves for DYM revealed gradual improvement, reaching the predefined performance target (70 %) overall, and in three of five contributing centers in 2018. Analysis of a sudden downslope development in the CUSUM curve of DYM in one center revealed temporary absence of a senior cytopathologist to have had a temporary negative impact on performance. Conclusions CUSUM-derived learning curves allow for assessment of best practices by comparison among peers in a multidisciplinary multicenter quality improvement initiative and proved to be a valuable and easy-to-interpret means to evaluate EUS performance over time.

Do endosonographers agree on the presence of bile duct sludge and the subsequent need for intervention?

Background and study aims Endoscopic ultrasonography (EUS) is a tool widely used to diagnose bile duct lithiasis. In approximately one out of five patients with positive findings at EUS, sludge is detected in the bile duct instead of stones. The objective of this study was to establish the agreement among endosonographers regarding: 1. presence of common bile duct (CBD) stones, microlithiasis and sludge; and 2. the need for subsequent treatment. Patients and methods 30 EUS videos of patients with an intermediate probability of CBD stones were evaluated by 41 endosonographers. Experience in EUS and endoscopic retrograde cholangiopancreatography, and the endosonographers' type of practices were recorded. Fleiss' kappa statistics were used to quantify the agreement. Associations between levels of experience and both EUS ratings and treatment decisions were investigated using mixed effects models. Results A total of 1230 ratings and treatment decisions were evaluated. The overall agreement on EUS findings was fair (Fleiss' κ 0.32). The agreement on presence of stones was moderate (κ 0.46). For microlithiasis it was fair (κ 0.25) and for sludge it was slight (κ 0.16). In cases with CBD stones there was an almost perfect agreement for the decision to subsequently perform an ERC + ES. In case of presumed microlithiasis or sludge an ERC was opted for in 78 % and 51 % of cases, respectively. Differences in experience and types of practice appear unrelated to the agreement on both EUS findings and the decision for subsequent treatment. Conclusions There is only slight agreement among endosonographers regarding the presence of bile duct sludge. Regarding the need for subsequent treatment of bile duct sludge there is no consensus.

Circulating levels of PD-L1 and Galectin-9 are associated with patient survival in surgically treated Hepatocellular Carcinoma independent of their intra-tumoral expression levels.

Tumor expression of immune co-inhibitory ligands, such as PD-L1 and Galectin-9, have potential prognostic value in Hepatocellular Carcinoma (HCC). Circulating levels of these molecules, however, have hardly been studied. This study aims to assess the prognostic significance of circulating PD-L1 and circulating Galectin-9 in patients with resected HCC, and to compare their prognostic significance to the intra-tumoral expression of these same molecules. Archived tissues and stored peripheral blood samples from 81 patients who underwent HCC resection or liver transplantation, with curative intent, were used. Immunohistochemistry was performed to determine intra-tumoral expression of PD-L1 and Galectin-9, while ELISA was used to quantify their respective circulating levels. High circulating PD-L1 (HR 0.12, 95%CI 0.16-0.86, p = 0.011) and high circulating Galectin-9 (HR 0.11, 95%CI 0.15-0.85, p = 0.010) levels were both associated with improved HCC-specific survival. Surprisingly, there was no correlation between circulating levels of PD-L1 and Galectin-9 and their intra-tumoral expression levels. In fact, circulating levels of PD-L1 and Galectin-9 were predictive of HCC-specific survival independently of intra-tumoral levels and baseline clinicopathologic characteristics. Combined analysis of circulating levels and intra-tumoral expression of PD-L1 (HR 0.33, 95%CI 0.16-0.68, p = 0.002) and Galectin-9 (HR 0.27, 95%CI 0.13-0.57, p = 0.001) resulted in more confident prediction of survival. In conclusion, circulating PD-L1 and Galectin-9 levels prognostically differentiate resected HCC patients, independently of their intra-tumoral expression. Combining circulating and intra-tumoral expression levels of PD-L1 or Galectin-9 further improves the prognostic values of these immune biomarkers.

PD-L1, Galectin-9 and CD8+ tumor-infiltrating lymphocytes are associated with survival in hepatocellular carcinoma.

Novel systemic treatments for hepatocellular carcinoma (HCC) are strongly needed. Immunotherapy is a promising strategy that can induce specific antitumor immune responses. Understanding the mechanisms of immune resistance by HCC is crucial for development of suitable immunotherapeutics. We used immunohistochemistry on tissue-microarrays to examine the co-expression of the immune inhibiting molecules PD-L1, Galectin-9, HVEM and IDO, as well as tumor CD8+ lymphocyte infiltration in HCC, in two independent cohorts of patients. We found that at least some expression in tumor cells was seen in 97% of cases for HVEM, 83% for PD-L1, 79% for Gal-9 and 66% for IDO. In the discovery cohort (n = 94), we found that lack of, or low, tumor expression of PD-L1 (p < 0.001), Galectin-9 (p < 0.001) and HVEM (p < 0.001), and low CD8+TIL count (p = 0.016), were associated with poor HCC-specific survival. PD-L1, Galectin-9 and CD8+TIL count were predictive of HCC-specific survival independent of baseline clinicopathologic characteristics and the combination of these markers was a powerful predictor of HCC-specific survival (HR 0.29; p <0.001). These results were confirmed in the validation cohort (n = 60). We show that low expression levels of PD-L1 and Gal-9 in combination with low CD8+TIL count predict extremely poor HCC-specific survival and it requires a change in two of these parameters to significantly improve prognosis. In conclusion, intra-tumoral expression of these immune inhibiting molecules was observed in the majority of HCC patients. Low expression of PD-L1 and Galectin-9 and low CD8+TIL count are associated with poor HCC-specific survival. Combining immune biomarkers leads to superior predictors of HCC mortality.

Delineating the GRIN1 phenotypic spectrum: A distinct genetic NMDA receptor encephalopathy.

OBJECTIVE: To determine the phenotypic spectrum caused by mutations in GRIN1 encoding the NMDA receptor subunit GluN1 and to investigate their underlying functional pathophysiology. METHODS: We collected molecular and clinical data from several diagnostic and research cohorts. Functional consequences of GRIN1 mutations were investigated in Xenopus laevis oocytes. RESULTS: We identified heterozygous de novo GRIN1 mutations in 14 individuals and reviewed the phenotypes of all 9 previously reported patients. These 23 individuals presented with a distinct phenotype of profound developmental delay, severe intellectual disability with absent speech, muscular hypotonia, hyperkinetic movement disorder, oculogyric crises, cortical blindness, generalized cerebral atrophy, and epilepsy. Mutations cluster within transmembrane segments and result in loss of channel function of varying severity with a dominant-negative effect. In addition, we describe 2 homozygous GRIN1 mutations (1 missense, 1 truncation), each segregating with severe neurodevelopmental phenotypes in consanguineous families. CONCLUSIONS: De novo GRIN1 mutations are associated with severe intellectual disability with cortical visual impairment as well as oculomotor and movement disorders being discriminating phenotypic features. Loss of NMDA receptor function appears to be the underlying disease mechanism. The identification of both heterozygous and homozygous mutations blurs the borders of dominant and recessive inheritance of GRIN1-associated disorders.