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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.

Background: Extrapulmonary neuroendocrine carcinoma (EP-NEC) are an aggressive subgroup of neuroendocrine neoplasms (NEN). Advanced EP-NEC is generally treated with platinum-based cytotoxic regimens, but progressive disease occurs rapidly, resulting in a poor prognosis. Genetic alterations in the mammalian target for rapamycin (mTOR) pathway have been identified in NEN, providing a rationale for treatment with the mTOR-inhibitor everolimus. Methods: A prospective phase 2 single-arm study included patients with advanced EP-NEC from three Dutch NEN expertise centres between March 2016 and January 2020. Treatment consisted of cisplatin 75 mg/m2 every 3 weeks in combination with daily everolimus 7.5 mg for a maximum of six cycles, followed by maintenance everolimus until disease progression. Primary endpoint was disease control rate (DCR), defined as the sum of overall response rate (ORR) plus the rate of stable disease according to RECIST 1.1, assessed at 9-week intervals. Toxicity was evaluated according to CTCAE version 5.0. Results: Thirty-nine patients, with a median age of 64 years (range: 28–74), of whom 20 (51%) were male, were enrolled. DCR was 82.1% (95% confidence interval (CI): 66.4–92.4), with an ORR of 58.9% (CI: 42.1–74.4). Median duration of response was 6.4 (CI: 5.8–7.0) months and median progression-free survival was 6.0 (CI: 4.3–7.8) months. Three patients (8%) had durable responses lasting > 12 months. Median overall survival was 8.7 (CI: 7.8–9.6) months. Most common grade 3/4 toxicities were haematological (36%) and renal (21%). Conclusion: Everolimus in combination with cisplatin is an effective first-line treatment option for advanced EP-NEC, especially in highly selected patients. Trial registration: Clinicaltrials.gov, NCT02695459, https://clinicaltrials.gov/ct2/show/NCT02695459 .

Neuroendocrine tumours of the small intestine (SI-NETs) are rare and heterogeneous. There is an unmet need for prognostication of disease course and to aid treatment strategies. A previously developed nomogram based on clinical and tumour characteristics aims to predict disease-specific survival (DSS) in patients with a SI-NET. We aimed to validate the nomogram and identify predictors of survival. Four hundred patients with a grade 1 or 2 SI-NET were included, between January 2000 and June 2016. Predicted 5- and 10-year survival was compared to actual DSS. Multivariable analysis identified predictors for actual DSS. We found that in low-, medium- and high-risk groups 5-year nomogram DSS vs. actual DSS was 0.86 vs. 0.82 (p &lt, 0.001), 0.52 vs. 0.71 (p &lt, 0.001) and 0.26 vs. 0.53 (p &lt, 0.001), respectively. Ten-year nomogram DSS vs. actual DSS was 0.68 vs. 0.69 (p &lt, 0.001), 0.40 vs. 0.50 (p &lt, 0.001) and 0.20 vs. 0.35 (p &lt, 0.001), respectively. Age, WHO-performance score of 2, Ki-67 index &ge, 10, unknown primary tumour, CgA &gt, 6x ULN and elevated liver tests were identified as independent predictors for a worse DSS. This shows that the nomogram was able to differentiate, but underestimated DSS for patients with a SI-NET. Improvement of prognostication incorporating new emerging biomarkers is necessary to adequately estimate survival.

PD-1 plus CTLA-4 blockade is highly effective in advanced-stage, mismatch repair (MMR)-deficient (dMMR) colorectal cancers, yet not in MMR-proficient (pMMR) tumors. We postulated a higher efficacy of neoadjuvant immunotherapy in early-stage colon cancers. In the exploratory NICHE study (ClinicalTrials.gov: NCT03026140), patients with dMMR or pMMR tumors received a single dose of ipilimumab and two doses of nivolumab before surgery, the pMMR group with or without celecoxib. The primary objective was safety and feasibility; 40 patients with 21 dMMR and 20 pMMR tumors were treated, and 3 patients received nivolumab monotherapy in the safety run-in. Treatment was well tolerated and all patients underwent radical resections without delays, meeting the primary endpoint. Of the patients who received ipilimumab + nivolumab (20 dMMR and 15 pMMR tumors), 35 were evaluable for efficacy and translational endpoints. Pathological response was observed in 20/20 (100%; 95% exact confidence interval (CI): 86–100%) dMMR tumors, with 19 major pathological responses (MPRs, ≤10% residual viable tumor) and 12 pathological complete responses. In pMMR tumors, 4/15 (27%; 95% exact CI: 8–55%) showed pathological responses, with 3 MPRs and 1 partial response. CD8+PD-1+ T cell infiltration was predictive of response in pMMR tumors. These data indicate that neoadjuvant immunotherapy may have the potential to become the standard of care for a defined group of colon cancer patients when validated in larger studies with at least 3 years of disease-free survival data. Results from the NICHE study show remarkable pathological responses to neoadjuvant combination immunotherapy in patients with early-stage colon cancer and uncover potential biomarkers of response.

CD151 is a cell surface protein that belongs to the tetraspanin superfamily. It forms complexes with the laminin-binding integrinsα3β1, α6β1 and α6β4 and is codistributed with these integrins in many tissues at sites of cell-matrix interactions. In this study we show that CD151 can also form stable complexes with the laminin-binding integrin α7β1. The strength of this interaction is comparable to that between CD151 and α3β1. Complexes ofα3β1, α6β1 and α7β1 with CD151 are equally well formed with all splice variants of the α3, α6 and α7 subunits, and complex formation is not affected by mutations that prevent the cleavage of the integrin α6 subunit. Like the expression ofα3β1 and α6β1, expression of α7β1 in K562 cells results in increased levels of CD151 at its surface. Two non-integrin laminin receptors, dystroglycan and the polypeptide on which the Lutheran blood group antigens are expressed, are also often colocalized with CD151, but no association with CD151-α3β1 complexes was found with biochemical analysis.The anti-CD151 antibody TS151R detects an epitope at a site at which CD151 interacts with integrins, and therefore it cannot react with CD151 when it is bound to an integrin. Comparison of the straining patterns produced by TS151R with that by of an anti-CD151 antibody recognizing an epitope outside the binding site (P48) revealed that most tissues expressing one or more laminin-binding integrins reacted with P48 but not with TS151R. However,smooth muscle cells that express α7β1 and renal tubular epithelial cells that express α6β1 were stained equally well by TS151R and P48. These results suggest that the interactions between CD151 and laminin-binding integrins are subject to cell-type-specific regulation.

Minimal change nephrosis (MCN) is characterized by massive proteinuria and ultrastructural alterations of glomerular visceral epithelial cells (GVEC). MCN has been associated with elevated production of interleukin (IL)-13 by circulating T lymphocytes and with T helper 2 lymphocyte-dependent conditions. We recently showed that GVEC express IL-4 and IL-13 receptors and that IL-4 and IL-13 increase transcellular ion transport over GVEC monolayers. We therefore hypothesized that IL-13 may directly injure GVEC. Here we demonstrate that IL-4 and IL-13 induce bafilomycin A1-sensitive basolateral proton secretion by cultured GVEC, indicating involvement of vacuolar H+-ATPase. The effects of IL-4 and IL-13 were accompanied by redistribution of the small GTPases Rab5b and Rab7, as shown by confocal immunofluorescence studies. Furthermore, Western blot analysis and assays for cysteine proteinase activity revealed basolateral secretion of the lysosomal proteinase procathepsin L by cultured GVEC, stimulated by IL-4 and IL-13. We speculate that IL-4 and IL-13 influence intracellular trafficking of proteins and promote proteolysis at the basolateral surface of GVEC, which may play a pathogenic role in altered glomerular permeability.

Release of urokinase plasminogen activator receptor during urosepsis and endotoxemia. Background The urokinase receptor (uPAR; CD87) is a multifunctional molecule involved in fibrinolysis, in proteolysis, in renal tubular functions, and in migration and adhesion of inflammatory cells to the site of infection. Methods To gain insight into systemic and local release of uPAR and into its regulation during urosepsis, which is one of the leading causes of chronic renal failure, uPAR was measured in urine and plasma of healthy human controls ( N = 20), patients with culture-proven urosepsis ( N = 30), and healthy human volunteers intravenously injected with endotoxin ( N = 7). Results Patients had elevated uPAR levels in both plasma and urine. Three hours after endotoxin challenge in volunteers, there was also a significant increase of uPAR in plasma and in urine. The urine/plasma ratio for uPAR was highly elevated during urosepsis and experimental endotoxemia, suggesting local production in the kidney. Accordingly, damaged tubuli strongly expressed uPAR during pyelonephritis. Moreover, tubular epithelial cells produced uPAR in vitro, and this secretion was strongly up-regulated after stimulation with interleukin-1β or tumor necrosis factor-α. Conclusions We found that uPAR is released systemically and in the urinary tract during urosepsis and experimental endotoxemia. This systemic and renal production of uPAR during pyelonephritis may play a central role in eliminating the infection and protecting renal function.

In minimal change nephrosis (MCN), proteinuria is associated with structural changes of the glomerular visceral epithelial cells (GVEC). The occurrence of MCN has been associated with 2 lymphocyte-dependent conditions. To examine a direct role for type 2 cytokines in GVEC injury, the expression of interleukin (IL)-4/IL-13 receptors by GVEC and direct effects of IL-4 and IL-13 on GVEC were studied. Reverse transcription-PCR showed that isolated human and rat glomeruli and cultured human and rat GVEC expressed mRNA for IL-4Ralpha, IL-13Ralpha1, and IL-13Ralpha2. Protein expression of [L-4Ralpha and IL-13Ralpha2 by GVEC in human kidney biopsies and by cultured human GVEC was detected by immunohistochemistry. Western blotting demonstrated phosphorylation of STAT6 in cultured GVEC upon incubation with IL-4 or IL-13. This indicated signal transduction via the heterodimeric receptor complex IL-4R2, which is composed of the IL-4Ralpha and the IL-13Ralpha1. Direct effects on GVEC function were examined in monolayer experiments. IL-4 and IL-13 dose-dependently decreased transepithelial electrical resistance of monolayers of rat GVEC to approximately 30 and 40% of baseline values, respectively. The transepithelial electrical resistance decrease was associated with a significant increase in short-circuit current, whereas no changes were observed in the transmonolayer flux of the macromolecules horseradish peroxidase (molecular weight, 44 kD) and 14C-mannitol (molecular weight, 182 Da). No changes in cell structure were observed with electron microscopy. It is concluded that by binding to specific IL-4/ IL-13 receptors, IL-4 and IL-13 can exert specific effects on GVEC function, which could be of pathogenetic relevance for glomerular injury in MCN.

Contains fulltext : 71431.pdf (Publisher’s version ) (Closed access) Podocyte foot process effacement is characteristic of proteinuric renal diseases. In minimal change nephrotic syndrome (MCNS) foot processes are diffusely effaced whereas the extent of effacement varies in focal segmental glomerulosclerosis (FSGS). Here we measured foot process effacement in FSGS and compared it to that in MCNS and in normal kidneys. A clinical diagnosis was used to differentiate idiopathic FSGS from secondary FSGS. Median foot process width, determined morphometrically by electron microscopy, was 3236 nm in 17 patients with idiopathic FSGS, 1098 nm in 7 patients with secondary FSGS, and 1725 nm in 15 patients with MCNS, as compared to 562 nm in 12 control patients. Multivariate analysis showed that foot process width did not correlate with proteinuria or serum albumin levels but was significantly associated as an independent factor with the type of disease. Foot process width over 1500 nm differentiated idiopathic from secondary FSGS with high sensitivity and specificity. Our results show that quantitative analysis of foot processes may offer a potential tool to distinguish idiopathic from secondary FSGS.

Podocyte foot process effacement is not correlated with the level of proteinuria in human glomerulopathies. Background Nephrotic syndromes result from increased glomerular permeability to proteins and are structurally believed to be associated with podocyte foot process effacement. Despite increasing knowledge of the molecular composition of the glomerular filtration barrier, the relationship between proteinuria and foot process effacement is unclear. Methods We conducted a morphologic study on the relationship between podocyte foot process effacement and proteinuria. Electron microscope pictures of glomerular capillaries were randomly taken from 27 cases in various stages of minimal change nephrotic syndrome (MCNS), from six cases of IgA nephropathy (IgAN) with high proteinuria and from seven control kidneys. From each picture, the mean width of the foot processes (FPW) was quantitated. Results In normal kidney the mean FPW was 580 ± 40 nm. In biopsies from patients with MCNS without treatment, foot processes were diffusely effaced, reflected by a FPW of 1600 ± 440 nm. In biopsies from patients with MCNS relapsing under prednisolone treatment, foot processes were significantly less effaced than in untreated MCNS (FPW 920 ± 200 nm). In biopsies displaying IgAN, effacement was significantly more segmental than in untreated MCNS (FPW 800 ± 170 nm). Proteinuria did not differ significantly among the groups. Neither in MCNS nor in IgAN was the extent of foot process effacement correlated with the level of proteinuria. Conclusion Podocyte foot process effacement is not correlated with proteinuria. The differences in podocyte effacement between MCNS, MCNS relapsing under prednisolone treatment, and IgAN may point to different mechanisms of podocyte injury in these diseases.

Interleukin-9 is an immunoregulatory cytokine implicated in the development of asthma and allergy. To investigate the role of IL-9 in vivo, we have generated transgenic mice in which IL-9 is expressed from its own promoter. Strikingly, overexpression of IL-9 resulted in premature mortality associated with a complex phenotype characterized by the development of autoantibodies, hydronephrosis, and T cell lymphoma. By intercrossing IL-9 transgenic mice with a panel of Th2 cytokine-deficient mice, we demonstrate that these disorders represent distinct phenotypes that can be dissociated by their differential dependence on Th2 cytokines. Autoantibody production was ablated in IL-9 transgenic animals with a combined absence of IL-4, IL-5, and IL-13, coincident with a reduction in peritoneal B-1 cells. Hydronephrosis arose in 75% of IL-9 transgenic animals and was dependent on the presence of IL-4 and IL-13. In contrast, T cell lymphomas developed independently of the other Th2 cytokines, with the generation of rapidly proliferating CD8+ or CD4+CD8+ T cell clones that arose in the thymus before infiltrating both lymphoid and nonlymphoid tissues. Our data highlight potentially important new roles for IL-9, through its regulation of downstream Th2 effector cytokines, in autoantibody production and in hydronephrosis.

Expression of urokinase plasminogen activator and its receptor during acute renal allograft rejection. Background In inflammation, urokinase plasminogen activator (uPA) and its receptor (uPAR) play an important role in fibrinolysis and in activation and chemotaxis of neutrophils and lymphocytes. Moreover, the uPA/uPAR system is involved in processes that affect turnover of the extracellular matrix (ECM). The aim of this study was to determine the local and systemic release of uPAR, and the expression of uPA and uPAR in renal tissues during acute renal allograft rejection. Methods Blood, urine, and tissue samples were collected from 33 patients diagnosed with acute allograft rejection and from 14 transplant patients without rejection. From 10 healthy volunteers, blood and urine were collected as a control. In urine and blood samples, the levels of uPAR were determined by enzyme-linked immunosorbent assay (ELISA). Immunostaining and in situ hybridization for uPA and uPAR were performed on renal biopsies. Results uPAR was detectable at low levels in serum and urine of healthy volunteers and was increased in nonrejecting allograft recipients. Serum and urine levels of uPAR were higher in transplant recipients with rejection compared to nonrejectors. The urine and serum levels of uPAR correlated with the renal function. Immunostaining and in situ hybridization showed an up-regulation of both uPA and uPAR in rejection biopsies. Nonrejected grafts displayed no expression of uPA and uPAR by immunostaining, or of uPAR by in situ hybridization. uPA was detected in a limited number of tubular epithelial cells by in situ hybridization. During rejection, lymphocytes as well as tubular epithelial cells showed uPA and uPAR expression. In the vascular types of rejection, strong expression of uPA was also seen in the entire vessel wall, while uPAR was expressed by the endothelium. Conclusion This study shows that ( 1 ) uPA and uPAR are up-regulated during acute renal allograft rejection; ( 2 ) uPAR levels in urine and serum correlate with serum creatinine levels, and ( 3 ) uPA and uPAR are produced by inflammatory cells, tubular epithelium, and vascular endothelium during acute renal allograft rejection.

CD151 is a cell surface protein that belongs to the tetraspanin superfamily. It forms complexes with the laminin-binding integrins alpha3beta1, alpha6beta1 and alpha6beta4 and is codistributed with these integrins in many tissues at sites of cell-matrix interactions. In this study we show that CD151 can also form stable complexes with the laminin-binding integrin alpha7beta1. The strength of this interaction is comparable to that between CD151 and alpha3beta1. Complexes of alpha3beta1, alpha6beta1 and alpha7beta1 with CD151 are equally well formed with all splice variants of the alpha3, alpha6 and alpha7 subunits, and complex formation is not affected by mutations that prevent the cleavage of the integrin alpha6 subunit. Like the expression of alpha3beta1 and alpha6beta1, expression of alpha7beta1 in K562 cells results in increased levels of CD151 at its surface. Two non-integrin laminin receptors, dystroglycan and the polypeptide on which the Lutheran blood group antigens are expressed, are also often colocalized with CD151, but no association with CD151-alpha3beta1 complexes was found with biochemical analysis. The anti-CD151 antibody TS151R detects an epitope at a site at which CD151 interacts with integrins, and therefore it cannot react with CD151 when it is bound to an integrin. Comparison of the straining patterns produced by TS151R with that by of an anti-CD151 antibody recognizing an epitope outside the binding site (P48) revealed that most tissues expressing one or more laminin-binding integrins reacted with P48 but not with TS151R. However, smooth muscle cells that express alpha7beta1 and renal tubular epithelial cells that express alpha6beta1 were stained equally well by TS151R and P48. These results suggest that the interactions between CD151 and laminin-binding integrins are subject to cell-type-specific regulation.

Background Neuroendocrine tumours (NETs) are rare. However, a rising incidence has been reported over the past decades. For colorectal NETs, this is presumably caused by an increased awareness of colorectal diseases and colonoscopic procedures. This study aims to analyse the change in incidence of colorectal NETs, characteristics and management and evaluate the proportion of colorectal NETs detected in a national colorectal cancer (CRC) screening programme. Methods Histopathological reports on colorectal well-differentiated NETs detected between 2006 and 2016 were collected from the Dutch pathology database (PALGA) containing nationwide histo- and cytopathology reports of all pathology laboratories in the Netherlands. Results Colorectal NETs were detected in 1055 individuals. Increasing incidence rates were observed from 0.36 per 100,000 inhabitants in 2006 to 0.75 per 100,000 inhabitants in 2011 (p value < 0.001), remaining stable afterward. Most NETs were grade I (73.5%) and detected in the rectum (76.4%). The majority (88.2%) were detected by colonoscopy, and the final intervention depended significantly on primary location of the tumour; 94.6% of rectal NETs were endoscopically removed, whereas 61.0% of colonic NETs were removed by surgery. There was an increase in local excision both of rectal and colonic NETs over the years instead of radical resection. Screening for CRC started in 2014 and contributed by detecting 32% of the diagnosed colorectal NETs within the invited age group, of which 94.6% were detected at an early stage. Conclusion The incidence of reported colorectal NETs in the Netherlands doubled over the last decade. The Dutch CRC screening programme had a clear contribution to colorectal NETs incidence among its target population. A shift to more local management of detected lesions was observed over time.