Postmortem Minimally Invasive Autopsy in Critically Ill COVID-19 Patients at the Bedside: A Proof-of-Concept Study at the ICU.

BACKGROUND: Economic restrictions and workforce cuts have continually challenged conventional autopsies. Recently, the COVID-19 pandemic has added tissue quality and safety requirements to the investigation of this disease, thereby launching efforts to upgrade autopsy strategies. METHODS: In this proof-of-concept study, we performed bedside ultrasound-guided minimally invasive autopsy (US-MIA) in the ICU of critically ill COVID-19 patients using a structured protocol to obtain non-autolyzed tissue. Biopsies were assessed for their quality (vitality) and length of biopsy (mm) and for diagnosis. The efficiency of the procedure was monitored in five cases by recording the time of each step and safety issues by swabbing personal protective equipment and devices for viral contamination. FINDINGS: Ultrasound examination and tissue procurement required a mean time period of 13 min and 54 min, respectively. A total of 318 multiorgan biopsies were obtained from five patients. Quality and vitality standards were fulfilled, which not only allowed for specific histopathological diagnosis but also the reliable detection of SARS-CoV-2 virions in unexpected organs using electronic microscopy and RNA-expressing techniques. INTERPRETATION: Bedside multidisciplinary US-MIA allows for the fast and efficient acquisition of autolytic-free tissue and offers unappreciated potential to overcome the limitations of research in postmortem studies.

PARP1-targeted fluorescence molecular endoscopy as novel tool for early detection of esophageal dysplasia and adenocarcinoma.

BACKGROUND: Esophageal cancer is one of the 10 most common cancers worldwide and its incidence is dramatically increasing. Despite some improvements, the current surveillance protocol with white light endoscopy and random untargeted biopsies collection (Seattle protocol) fails to diagnose dysplastic and cancerous lesions in up to 50% of patients. Therefore, new endoscopic imaging technologies in combination with tumor-specific molecular probes are needed to improve early detection. Herein, we investigated the use of the fluorescent Poly (ADP-ribose) Polymerase 1 (PARP1)-inhibitor PARPi-FL for early detection of dysplastic lesions in patient-derived organoids and transgenic mouse models, which closely mimic the transformation from non-malignant Barrett's Esophagus (BE) to invasive esophageal adenocarcinoma (EAC). METHODS: We determined PARP1 expression via immunohistochemistry (IHC) in human biospecimens and mouse tissues. We also assessed PARPi-FL uptake in patient- and mouse-derived organoids. Following intravenous injection of 75 nmol PARPi-FL/mouse in L2-IL1B (n = 4) and L2-IL1B/IL8Tg mice (n = 12), we conducted fluorescence molecular endoscopy (FME) and/or imaged whole excised stomachs to assess PARPi-FL accumulation in dysplastic lesions. L2-IL1B/IL8Tg mice (n = 3) and wild-type (WT) mice (n = 2) without PARPi-FL injection served as controls. The imaging results were validated by confocal microscopy and IHC of excised tissues. RESULTS: IHC on patient and murine tissue revealed similar patterns of increasing PARP1 expression in presence of dysplasia and cancer. In human and murine organoids, PARPi-FL localized to PARP1-expressing epithelial cell nuclei after 10 min of incubation. Injection of PARPi-FL in transgenic mouse models of BE resulted in the successful detection of lesions via FME, with a mean target-to-background ratio > 2 independently from the disease stage. The localization of PARPi-FL in the lesions was confirmed by imaging of the excised stomachs and confocal microscopy. Without PARPi-FL injection, identification of lesions via FME in transgenic mice was not possible. CONCLUSION: PARPi-FL imaging is a promising approach for clinically needed improved detection of dysplastic and malignant EAC lesions in patients with BE. Since PARPi-FL is currently evaluated in a phase 2 clinical trial for oral cancer detection after topical application, clinical translation for early detection of dysplasia and EAC in BE patients via FME screening appears feasible.

[Multimodal ultrasound techniques for the differential diagnosis of splenic lesions - A diagnostic challenge]. / Multimodale Ultraschalltechniken zur Differenzialdiagnose von Milzherden ­ eine diagnostische Herausforderung.

BACKGROUND: Splenic tumors are rare and can pose a differential diagnostic challenge, especially as an incidental imaging finding. Due to a lack of large scale biopsy studies the available literature is limited with respect to clear imaging criteria for dignity. OBJECTIVE: The present work is intended to show the chances of a targeted elicitation of the medical history as well as the possibilities and limitations of multimodal sonography in order to achieve the correct diagnosis of a splenic lesion using simple and gentle methods. MATERIAL AND METHODS: Selective literature search and clinical case studies. RESULTS: In the differential diagnostics of focal splenic lesions, information about pre-existing hemato-oncological or inflammatory rheumatological diseases is essential in order to correctly classify incidental findings in particular. In addition to B­mode ultrasound (B-US) and color-coded Doppler ultrasound (CD-US), contrast-enhanced ultrasound (CEUS) in particular provides crucial differential diagnostic information. While hyperechoic foci in B­US or arterially hypervascularized splenic foci in CD-US/CEUS are usually benign, hypoechoic and arterially hypoperfused foci in CD-US/CEUS must be further clarified. Although the ultrasound-guided biopsy of the spleen has a higher risk of bleeding than a liver biopsy, it is still the gentlest and most effective method for achieving the histological clarification of splenic lesions when the indications are correct. CONCLUSION: Through the combination of the medical history and multimodal ultrasound methods, if necessary supplemented by an ultrasound-guided biopsy, focal splenic lesions can be successfully classified in most cases with a direct impact on further clinical procedures.

High RIPK3 expression is associated with a higher risk of early kidney transplant failure.

Renal ischemia-reperfusion injury (IRI) is associated with reduced allograft survival, and each additional hour of cold ischemia time increases the risk of graft failure and mortality following renal transplantation. Receptor-interacting protein kinase 3 (RIPK3) is a key effector of necroptosis, a regulated form of cell death. Here, we evaluate the first-in-human RIPK3 expression dataset following IRI in kidney transplantation. The primary analysis included 374 baseline biopsy samples obtained from renal allografts 10 minutes after onset of reperfusion. RIPK3 was primarily detected in proximal tubular cells and distal tubular cells, both of which are affected by IRI. Time-to-event analysis revealed that high RIPK3 expression is associated with a significantly higher risk of one-year transplant failure and prognostic for one-year (death-censored) transplant failure independent of donor and recipient associated risk factors in multivariable analyses. The RIPK3 score also correlated with deceased donation, cold ischemia time and the extent of tubular injury.

Low microsatellite instability: A distinct instability type in gastric cancer?

PURPOSE: We recently showed that low microsatellite instability (MSI-L) is associated with a good response to platinum/5-fluorouracil (5-FU) neoadjuvant chemotherapy (CTx) in gastric cancer. The purpose of this study was to characterize the instability pattern and to investigate an association of MSI-L tumors with mutations in genes of DNA repair pathways and with total tumor mutation burden (TMB). METHODS: MSI patterns were compared between 67 MSI high (-H) and 35 MSI-L tumors. Whole-exome sequencing was performed in 34 microsatellite stable (MSS) and 20 MSI-L tumors after or without neoadjuvant CTx. RESULTS: Of the 35 MSI-L tumors, 33 tumors had instability at a dinucleotide repeat marker. In the homologous recombination (HR) pathway, 10 of the 34 (29%) MSS and 10 of the 20 (50%) MSI-L tumors showed variants (p = 0.154). In the DNA damage tolerance pathway, 6 of the 34 (18%) MSS and 7 of the 20 (35%) MSI-L tumors had variants (p = 0.194). The HR deficiency score was similar in both tumor groups. TMB was significantly higher in MSI-L compared to MSS tumors after CTx (p = 0.046). In the MSS and MSI-L tumors without CTx no difference was observed (p = 1.00). CONCLUSION: MSI-L due to instability at dinucleotide repeat markers was associated with increased TMB after neoadjuvant CTx treatment, indicating sensitivity to platinum/5-FU CTx. If confirmed in further studies, this could contribute to refined chemotherapeutic options including immune-based strategies for GC patients with MSI-L tumors.

Proteogenomic analysis reveals RNA as a source for tumor-agnostic neoantigen identification.

Systemic pan-tumor analyses may reveal the significance of common features implicated in cancer immunogenicity and patient survival. Here, we provide a comprehensive multi-omics data set for 32 patients across 25 tumor types for proteogenomic-based discovery of neoantigens. By using an optimized computational approach, we discover a large number of tumor-specific and tumor-associated antigens. To create a pipeline for the identification of neoantigens in our cohort, we combine DNA and RNA sequencing with MS-based immunopeptidomics of tumor specimens, followed by the assessment of their immunogenicity and an in-depth validation process. We detect a broad variety of non-canonical HLA-binding peptides in the majority of patients demonstrating partially immunogenicity. Our validation process allows for the selection of 32 potential neoantigen candidates. The majority of neoantigen candidates originates from variants identified in the RNA data set, illustrating the relevance of RNA as a still understudied source of cancer antigens. This study underlines the importance of RNA-centered variant detection for the identification of shared biomarkers and potentially relevant neoantigen candidates.

Evolutionary and immune microenvironment dynamics during neoadjuvant treatment of oesophagael adenocarcinoma.

Locally advanced oesophageal adenocarcinoma (EAC) remains difficult to treat because of common resistance to neoadjuvant therapy and high recurrence rates. The ecological and evolutionary dynamics responsible for treatment failure are incompletely understood. Here, we performed a comprehensive multi-omic analysis of samples collected from EAC patients in the MEMORI clinical trial, revealing major changes in gene expression profiles and immune microenvironment composition that did not appear to be driven by changes in clonal composition. Multi-region multi-timepoint whole exome (300x depth) and paired transcriptome sequencing was performed on 27 patients pre-, during and after neoadjuvant treatment. EAC showed major transcriptomic changes during treatment with upregulation of immune and stromal pathways and oncogenic pathways such as KRAS, Hedgehog and WNT. However, genetic data revealed that clonal sweeps were rare, suggesting that gene expression changes were not clonally driven. Additional longitudinal image mass cytometry was performed in a subset of 15 patients and T-cell receptor sequencing in 10 patients, revealing remodelling of the T-cell compartment during treatment and other shifts in microenvironment composition. The presence of immune escape mechanisms and a lack of clonal T-cell expansions were linked to poor clinical treatment response. This study identifies profound transcriptional changes during treatment with limited evidence that clonal replacement is the cause, suggesting phenotypic plasticity and immune dynamics as mechanisms for therapy resistance with pharmacological relevance.

Microsatellite instability and sex-specific differences of survival in gastric cancer after neoadjuvant chemotherapy without and with taxane: An observational study in real world patients.

PURPOSE: To investigate the prognostic role of microsatellite instability (MSI) in association with sex of patients treated with platinum/fluoropyrimidine neoadjuvant chemotherapy (CTx) with or without a taxane-containing compound. METHODS: Of the 505 retrospectively analyzed patients with gastric or gastroesophageal adenocarcinoma, 411 patients were treated without taxane and 94 patients with a taxane-containing compound. MSI was determined using standard assays. RESULTS: Females demonstrated a better overall survival (OS) than males in the non-taxane group (HR, 0.59; 95% CI 0.41-0.86; p = 0.005), whereas no significant difference was found in the taxane group (HR 1.22; 95% CI 0.55-2.73, p = 0.630). MSI-High (-H) was associated with a better prognosis in both groups (without taxane: HR 0.56; 95% CI 0.33-0.97; p = 0.038; with taxane: HR 0.28; 95% CI 0.04-2.02, p = 0.204). In the non-taxane group, female MSI-H patients showed the best OS (HR 0.18, 95% CI 0.05-0.73; p = 0.016), followed by the female microsatellite stable (MSS) (HR 0.67, 95% CI 0.46-0.98, p = 0.040) and the male MSI-H group (HR 0.76; 95% CI 0.42-1.37, p = 0.760) taken the male MSS group as reference. In the taxane group, female and male MSI-H patients demonstrated the best OS (female MSI-H: HR 0.05, 95% CI 0.00-240.46; male MSI-H: HR 0.45, 95% CI 0.61-3.63, p = 0.438), whereas the female MSS group showed a decreased OS (HR 1.39 95% CI 0.62-3.12, p = 0.420) compared to male MSS patients. CONCLUSION: OS in gastric/gastroesophageal cancer after CTx might depend on sex and MSI status and may differ between patients treated with or without a taxane compound in the chemotherapeutic regimen.

Significant Tumor Regression after Neoadjuvant Chemotherapy in Gastric Cancer, but Poor Survival of the Patient? Role of MHC Class I Alterations.

We aimed to determine the clinical and prognostic relevance of allelic imbalance (AI) of the major histocompatibility complex (MHC) class I genes, encompassing the human leukocyte antigen (HLA) class I and beta-2 microglobulin (B2M) genes, in the context of neoadjuvant platinum/fluoropyrimidine chemotherapy (CTx). Biopsies before CTx were studied in 158 patients with adenocarcinoma of the stomach or gastroesophageal junction. The response was histopathologically evaluated. AI was detected by multiplex PCRs analysis of four or five microsatellite markers in HLA and B2M regions, respectively. AI with no marker was significantly associated with response or survival. However, subgroup analysis revealed differences. AI at marker D6S265, close to the HLA-A gene, was associated with an obvious increased risk in responding (HR, 3.62; 95% CI, 0.96-13.68, p = 0.058) but not in non-responding patients (HR, 0.92; 95% CI, 0.51-1.65, p = 0.773). Markers D6S273 and D6S2872 showed similar results. The interaction between AI at D6S265 and response to CTx was significant in a multivariable analysis (p = 0.010). No associations were observed for B2M markers. Our results underline the importance of intact neoantigen presentation specifically for responding patients and may help explain an unexpectedly poor survival of a patient despite significant tumor regression after neoadjuvant platinum/fluoropyrimidine CTx.

Vitronectin promotes immunothrombotic dysregulation in the venular microvasculature.

Microvascular immunothrombotic dysregulation is a critical process in the pathogenesis of severe systemic inflammatory diseases. The mechanisms controlling immunothrombosis in inflamed microvessels, however, remain poorly understood. Here, we report that under systemic inflammatory conditions the matricellular glycoproteinvitronectin (VN) establishes an intravascular scaffold, supporting interactions of aggregating platelets with immune cells and the venular endothelium. Blockade of the VN receptor glycoprotein (GP)IIb/IIIa interfered with this multicellular interplay and effectively prevented microvascular clot formation. In line with these experimental data, particularly VN was found to be enriched in the pulmonary microvasculature of patients with non-infectious (pancreatitis-associated) or infectious (coronavirus disease 2019 (COVID-19)-associated) severe systemic inflammatory responses. Targeting the VN-GPIIb/IIIa axis hence appears as a promising, already feasible strategy to counteract microvascular immunothrombotic dysregulation in systemic inflammatory pathologies.

Ultrasound-based "CEUS-Bosniak"classification for cystic renal lesions: an 8-year clinical experience.

PURPOSE: Renal cysts comprise benign and malignant entities. Risk assessment profits from CT/MRI imaging using the Bosniak classification. While Bosniak-IIF, -III, and -IV cover complex cyst variants, Bosniak-IIF and -III stand out due to notorious overestimation. Contrast-enhanced ultrasound (CEUS) is promising to overcome this deficit but warrants standardization. This study addresses the benefits of a combined CEUS and CT/MRI evaluation of renal cysts. The study provides a realistic account of kidney tumor boards' intricacies in trying to validate renal cysts. METHODS: 247 patients were examined over 8 years. CEUS lesions were graded according to CEUS-Bosniak (IIF, III, IV). 55 lesions were resected, CEUS-Bosniak- and CT/MRI-Bosniak-classification were correlated with histopathological diagnosis. Interobserver agreement between the classifications was evaluated statistically. 105 lesions were followed by ultrasound, and change in CEUS-Bosniak-types and lesion size were documented. RESULTS: 146 patients (156 lesions) were included. CEUS classified 67 lesions as CEUS-Bosniak-IIF, 44 as CEUS-Bosniak-III, and 45 as CEUS-Bosniak-IV. Histopathology of 55 resected lesions revealed benign cysts in all CEUS-Bosniak-IIF lesions (2/2), 40% of CEUS-Bosniak-III and 8% of CEUS-Bosniak-IV, whereas malignancy was uncovered in 60% of CEUS-Bosniak-III and 92% of CEUS-Bosniak-IV. Overall, CEUS-Bosniak-types matched CT/MRI-Bosniak types in 58% (fair agreement, κ = 0.28). CEUS-Bosniak resulted in higher stages than CT/MRI-Bosniak (40%). Ultrasound follow-up of 105 lesions detected no relevant differences between CEUS-Bosniak-types concerning cysts size. 99% of lesions showed the same CEUS-Bosniak-type. CONCLUSION: The CEUS-Bosniak classification is an essential tool in clinical practice to differentiate and monitor renal cystic lesions and empowers diagnostic work-up and patient care.

Histological evaluation of PAXgene tissue fixation in Barrett's esophagus and esophageal adenocarcinoma diagnostics.

The dysplasia grading of Barrett's esophagus (BE), based on the histomorphological assessment of formalin-fixed, paraffin-embedded (FFPE) tissue, suffers from high interobserver variability leading to an unsatisfactory prediction of cancer risk. Thus, pre-analytic preservation of biological molecules, which could improve risk prediction in BE enabling molecular and genetic analysis, is needed. We aimed to evaluate such a molecular pre-analytic fixation tool, PAXgene-fixed paraffin-embedded (PFPE) biopsies, and their suitability for histomorphological BE diagnostics in comparison to FFPE. In a ring trial, 9 GI pathologists evaluated 116 digital BE slides of non-dysplastic BE (NDBE), low-grade dysplasia (LGD), high-grade dysplasia (HGD), and esophageal adenocarcinomas (EAC) using virtual microscopy. Overall quality, cytological and histomorphological parameters, dysplasia criteria, and diagnosis were analyzed. PFPE showed better preservation of nuclear details as chromatin and nucleoli, whereas overall quality and histomorphologic parameters as visibility of basal lamina, goblet cells, and presence of artifacts were scored as equal to FFPE. The interobserver reproducibility with regard to the diagnosis was best for NDBE and EAC (κF = 0.72-0.75) and poor for LGD and HGD (κF = 0.13-0.3) in both. In conclusion, our data suggest that PFPE allows equally confident histomorphological diagnosis of BE and EAC, introducing a novel tool for molecular analysis and parallel histomorphological evaluation.

UICC Staging after Neoadjuvant/Perioperative Chemotherapy Reveals No Significant Survival Differences Compared to Primary Surgery for Locally Advanced Gastric Cancer.

Background: The applicability of UICC TNM staging for gastric cancer (GC) patients treated with neoadjuvant chemotherapy (nCTX) and surgery was not yet analyzed in comparison to patients undergoing primary surgery (PS). The purpose of this analysis was to analyze if the prognostic impact of TNM staging after nCTx is comparable with PS. Methods: Data for patients having been treated for GC with or without nCTx between 1990 and 2016 were analyzed. Uni-(URA) and multivariable regression analyses (MRA) were performed to identify predictors. Survival according to the UICC 8th edition stages was analyzed by the Kaplan−Meier method and cox regression analysis. Propensity score matching (PSM) was performed to balance for confounders. Results: 1149 patients with GC were eligible for primary analysis. URA demonstrated age (p < 0.0001), tumor localization (p < 0.0001), clinical UICC-stage, complications, UICC stage 0, IIB-IIIC, Lauren subtype, grading, and R-stage to be significantly associated with OS. MRA revealed that age, distal tumor localization, more than 25 dissected lymph nodes, UICC stage 0, IIB-IIIC, and Lauren subtype were significantly and independently related to OS. After PSM, survival analyses revealed only a significant difference for pN2/ypN2 (p = 0.03), while all other T and N stages were comparable. Conclusion: UICC dependent survival stages do not change significantly after nCTx treatment for GC. Therefore, UICC staging in its present version is applicable to patients undergoing nCTx.

PET-directed combined modality therapy for gastroesophageal junction cancer: Results of the multicentre prospective MEMORI trial of the German Cancer Consortium (DKTK).

BACKGROUND: Positron emission tomography (PET) may differentiate responding and non-responding tumours early in the treatment of locally advanced gastroesophageal junction adenocarcinomas. Early PET non-responders (P-NR) after induction CTX might benefit from changing to chemoradiation (CRT). METHODS: Patients underwent baseline 18F-FDG PET followed by 1 cycle of CTX. PET was repeated at day 14-21 and responders (P-R), defined as ≥35% decrease in SUVmean from baseline, continued with CTX. P-NR switched to CRT (CROSS). Patients underwent surgery 4-6 weeks post-CTX/CRT. The primary objective was an improvement in R0 resection rates in P-NR above a proportion of 70%. RESULTS: In total, 160 patients with resectable gastroesophageal junction adenocarcinomas were prospectively investigated by PET scanning. Eighty-five patients (53%) were excluded. Seventy-five eligible patients were enrolled in the study. Based on PET criteria, 50 (67.6%)/24 (32.4%) were P-R and P-NR, respectively. Resection was performed on 46 responders, including one patient who withdrew the ICF, and 22 non-responders (per-protocol population). R0 resection rates were 95.6% (43/45) for P-R and 86.4% (19/22) for P-NR. No treatment related deaths occurred. With a median follow-up time of 24.5 months, estimated 18 months DFS was 75.4%/64.2% for P-R/P-NR, respectively. The estimated 18 months OS was 95.5% for P-R and 68.2% for P-NR. CONCLUSION: The primary endpoint of the study to increase the R0 resection rate in metabolic NR was not met. PET response after induction CTX is prognostic for outcome with a prolonged OS and DFS in PET responders. TRIAL REGISTRATION: NCT00002014-000860-16.

Organ manifestations of COVID-19: what have we learned so far (not only) from autopsies?

The use of autopsies in medicine has been declining. The COVID-19 pandemic has documented and rejuvenated the importance of autopsies as a tool of modern medicine. In this review, we discuss the various autopsy techniques, the applicability of modern analytical methods to understand the pathophysiology of COVID-19, the major pathological organ findings, limitations or current studies, and open questions. This article summarizes published literature and the consented experience of the nationwide network of clinical, neuro-, and forensic pathologists from 27 German autopsy centers with more than 1200 COVID-19 autopsies. The autopsy tissues revealed that SARS-CoV-2 can be found in virtually all human organs and tissues, and the majority of cells. Autopsies have revealed the organ and tissue tropism of SARS-CoV-2, and the morphological features of COVID-19. This is characterized by diffuse alveolar damage, combined with angiocentric disease, which in turn is characterized by endothelial dysfunction, vascular inflammation, (micro-) thrombosis, vasoconstriction, and intussusceptive angiogenesis. These findings explained the increased pulmonary resistance in COVID-19 and supported the recommendations for antithrombotic treatment in COVID-19. In contrast, in extra-respiratory organs, pathological changes are often nonspecific and unclear to which extent these changes are due to direct infection vs. indirect/secondary mechanisms of organ injury, or a combination thereof. Ongoing research using autopsies aims at answering questions on disease mechanisms, e.g., focusing on variants of concern, and future challenges, such as post-COVID conditions. Autopsies are an invaluable tool in medicine and national and international interdisciplinary collaborative autopsy-based research initiatives are essential.

Contamination of personal protective equipment during COVID-19 autopsies.

Confronted with an emerging infectious disease at the beginning of the COVID-19 pandemic, the medical community faced concerns regarding the safety of autopsies on those who died of the disease. This attitude has changed, and autopsies are now recognized as indispensable tools for understanding COVID-19, but the true risk of infection to autopsy staff is nevertheless still debated. To clarify the rate of SARS-CoV-2 contamination in personal protective equipment (PPE), swabs were taken at nine points in the PPE of one physician and one assistant after each of 11 full autopsies performed at four centers. Swabs were also obtained from three minimally invasive autopsies (MIAs) conducted at a fifth center. Lung/bronchus swabs of the deceased served as positive controls, and SARS-CoV-2 RNA was detected by real-time RT-PCR. In 9 of 11 full autopsies, PPE samples tested RNA positive through PCR, accounting for 41 of the 198 PPE samples taken (21%). The main contaminated items of the PPE were gloves (64% positive), aprons (50% positive), and the tops of shoes (36% positive) while the fronts of safety goggles, for example, were positive in only 4.5% of the samples, and all the face masks were negative. In MIAs, viral RNA was observed in one sample from a glove but not in other swabs. Infectious virus isolation in cell culture was performed on RNA-positive swabs from the full autopsies. Of all the RNA-positive PPE samples, 21% of the glove samples, taken in 3 of 11 full autopsies, tested positive for infectious virus. In conclusion, PPE was contaminated with viral RNA in 82% of autopsies. In 27% of autopsies, PPE was found to be contaminated even with infectious virus, representing a potential risk of infection to autopsy staff. Adequate PPE and hygiene measures, including appropriate waste deposition, are therefore essential to ensure a safe work environment.

Elevated microsatellite instability at selected tetranucleotide (EMAST) repeats in gastric cancer: a distinct microsatellite instability type with potential clinical impact?

We investigated the clinical impact of elevated microsatellite instability at selected tetranucleotide (EMAST) repeats in the context of neoadjuvant chemotherapy (CTx) in gastric/gastro-oesophageal adenocarcinomas. We analysed 583 resected tumours (272 without and 311 after CTx) and 142 tumour biopsies before CTx. If at least two or three of the five tetranucleotide repeat markers tested showed instability, the tumours were defined as EMAST (2+) or EMAST (3+), respectively. Expression of mismatch repair proteins including MSH3 was analysed using immunohistochemistry. Microsatellite instability (MSI) and Epstein-Barr virus (EBV) positivity were determined using standard assays. EMAST (2+) and (3+) were detected in 17.8 and 11.5% of the tumours, respectively. The frequency of EMAST (2+) or (3+) in MSI-high (MSI-H) tumours was 96.2 or 92.5%, respectively, demonstrating a high overlap with this molecular subtype, and the association of EMAST and MSI status was significant (each overall p < 0.001). EMAST (2+ or 3+) alone in MSI-H and EBV-negative tumours demonstrated only a statistically significant association of EMAST (2+) positivity and negative lymph node status (42.3% in EMAST (2+) and 28.8% in EMAST negative, p = 0.045). EMAST alone by neither definition was significantly associated with overall survival (OS) of the patients. The median OS for EMAST (2+) patients was 40.0 months (95% confidence interval [CI] 16.4-63.6) compared with 38.7 months (95% CI 26.3-51.1) for the EMAST-negative group (p = 0.880). The median OS for EMAST (3+) patients was 46.7 months (95% CI 18.2-75.2) and 38.7 months (95% CI 26.2-51.2) for the negative group (p = 0.879). No statistically significant association with response to neoadjuvant CTx was observed (p = 0.992 and p = 0.433 for EMAST (2+) and (3+), respectively). In conclusion, our results demonstrate a nearly complete intersection between MSI-H and EMAST and they indicate that EMAST alone is not a distinct instability type associated with noticeable clinico-pathological characteristics of gastric carcinoma patients.

Genetic alterations of the SUMO isopeptidase SENP6 drive lymphomagenesis and genetic instability in diffuse large B-cell lymphoma.

SUMOylation is a post-translational modification of proteins that regulates these proteins' localization, turnover or function. Aberrant SUMOylation is frequently found in cancers but its origin remains elusive. Using a genome-wide transposon mutagenesis screen in a MYC-driven B-cell lymphoma model, we here identify the SUMO isopeptidase (or deconjugase) SENP6 as a tumor suppressor that links unrestricted SUMOylation to tumor development and progression. Notably, SENP6 is recurrently deleted in human lymphomas and SENP6 deficiency results in unrestricted SUMOylation. Mechanistically, SENP6 loss triggers release of DNA repair- and genome maintenance-associated protein complexes from chromatin thereby impairing DNA repair in response to DNA damages and ultimately promoting genomic instability. In line with this hypothesis, SENP6 deficiency drives synthetic lethality to Poly-ADP-Ribose-Polymerase (PARP) inhibition. Together, our results link SENP6 loss to defective genome maintenance and reveal the potential therapeutic application of PARP inhibitors in B-cell lymphoma.