Publications by authors named "Thomas F E Barth"

116 Publications

Molecular features and vulnerabilities of recurrent chordomas.

J Exp Clin Cancer Res 2021 Jul 30;40(1):244. Epub 2021 Jul 30.

Institute of Pathology, University Hospital Ulm, Albert-Einstein-Allee 11, 89081, Ulm, Germany.

Background: Tumor recurrence is one of the major challenges in clinical management of chordoma. Despite R0-resection, approximately 50% of chordomas recur within ten years after initial surgery. The underlying molecular processes are poorly understood resulting in the lack of associated therapeutic options. This is not least due to the absence of appropriate cell culture models of this orphan disease.

Methods: The intra-personal progression model cell lines U-CH11 and U-CH11R were compared using array comparative genomic hybridization, expression arrays, RNA-seq, and immunocytochemistry. Cell line origin was confirmed by short tandem repeat analysis. Inter-personal cell culture models (n = 6) were examined to validate whether the new model is representative. Cell viability after HOX/PBX complex inhibition with small peptides was determined by MTS assays.

Results: Using whole genome microarray analyses, striking differences in gene expression between primary and recurrent chordomas were identified. These expression differences were confirmed in the world's first intra-personal model of chordoma relapse consisting of cell lines established from a primary (U-CH11) and the corresponding recurrent tumor (U-CH11R). Array comparative genomic hybridization and RNA-sequencing analyses revealed profound genetic similarities between both cell lines pointing to transcriptomic reprogramming as a key mechanism of chordoma progression. Network analysis of the recurrence specific genes highlighted HOX/PBX signaling as a common dysregulated event. Hence, HOX/PBX complexes were used as so far unknown therapeutic targets in recurrent chordomas. Treating chordoma cell lines with the complex formation inhibiting peptide HXR9 induced cFOS mediated apoptosis in all chordoma cell lines tested. This effect was significantly stronger in cell lines established from chordoma relapses.

Conclusion: Clearly differing gene expression patterns and vulnerabilities to HOX/PBX complex inhibition in highly therapy resistant chordoma relapses were identified using the first intra-personal loco-regional and further inter-personal chordoma progression models. For the first time, HOX/PBX interference was used to induce cell death in chordoma and might serve as the basic concept of an upcoming targeted therapy for chordomas of all progression stages.
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http://dx.doi.org/10.1186/s13046-021-02037-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8325178PMC
July 2021

A phase II study of the PI3K inhibitor copanlisib in combination with the anti-CD20 monoclonal antibody rituximab for patients with marginal zone lymphoma: treatment rationale and protocol design of the COUP-1 trial.

BMC Cancer 2021 Jun 29;21(1):749. Epub 2021 Jun 29.

Department of Internal Medicine III, University Hospital Ulm, Albert-Einstein-Allee 23, 89081, Ulm, Germany.

Background: Advanced stage marginal zone lymphoma (MZL) is an incurable indolent B-cell lymphoma, for which a wide variety of treatments ranging from single agent rituximab to more dose intense immunochemotherapy exists. One of the major goals in this palliative setting is to develop chemotherapy-free treatments, which approach the efficacy of immunochemotherapies, but avoid chemotherapy associated toxicity in this often elderly patient population. The PI3K inhibitor copanlisib has recently shown remarkable clinical activity in refractory or relapsed indolent B-cell lymphomas, among them MZL. Based on these data, copanlisib monotherapy was granted breakthrough designation by the FDA for the treatment of adult patients with relapsed marginal zone lymphoma who have received at least two prior therapies. However, data are still limited in particular for MZL. Based on this, the COUP-1 trial aims at testing the toxicity and efficacy of copanlisib in combination with rituximab in treatment naive and relapsed MZL.

Methods: COUP-1 is a prospective, multicenter, single-arm, open-label, non-randomized phase II trial of 6 cycles (28 days cycle) of copanlisib (60 mg intravenous day 1, 8, 15) and rituximab (375 mg/m intravenous day 1) in the induction phase followed by a maintenance phase of copanlisib (d1, d15 every 4 weeks for a maximum of 12 cycles) and rituximab (d1 every 8 weeks for a maximum of 12 cycles) in patients aged ≥18 years with previously untreated or relapsed MZL in need of treatment. A total of 56 patients are to be enrolled. Primary endpoint is the complete response (CR) rate determined 12 months after start of induction therapy. Secondary endpoints include the overall response (OR) rate, progression free survival (PFS), overall survival (OS), safety and patient related outcome with quality of life. The study includes a translational bio-sampling program with the prospect to measure minimal residual disease. The study was initiated in November 2019.

Discussion: The COUP-1 trial evaluates the efficacy and toxicity of the treatment of copanlisib in combination with rituximab in patients with MZL and additionally offers the chance for translational research in this heterogenous type of lymphoma.

Trial Registration: ClinicalTrials.gov : NCT03474744 . Registration date: 03/23/2018.
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http://dx.doi.org/10.1186/s12885-021-08464-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8243426PMC
June 2021

Spatial distribution of immune checkpoint proteins in histological subtypes of lung adenocarcinoma.

Neoplasia 2021 06 5;23(6):584-593. Epub 2021 Jun 5.

Institute of Pathology, Ulm University, Ulm, Germany. Electronic address:

The most prevalent histological type of non-small cell lung cancer (NSCLC) is adenocarcinoma. The WHO classifies this tumor into subtypes according to the predominant growth pattern such as lepidic, acinar, papillary, solid or micropapillary, each harboring specific molecular features. NSCLC adenocarcinoma heterogeneity is discussed to be a reason for therapy failure using targeted therapy or immune checkpoint inhibitors. For successful therapy of immune checkpoint inhibitors the expression and distribution of the involved immune checkpoint proteins is essential. Therefore, we aimed to investigate the distribution of five prominent immune checkpoint proteins in regard of the histological growth patterns of lung adenocarcinoma. We performed immunohistochemical staining of 84 tumor segments from 22 resected tumor samples to evaluate the expression of PD-L1, PD-1, Nectin-2, PVR, and TIGIT in distinct growth patterns of lung adenocarcinoma. We determined a distinct heterogeneity between and within different tumor segments regarding morphological growth patterns. Furthermore, expression of immune checkpoint proteins varied between different growth pattern areas as well as within one distinct growth pattern. Expression of PVR was significantly higher in solid compared to acinar growth pattern (p= 0.00736). Of note, we detected TIGIT not only on tumor infiltrating lymphocytes but also on tumor cells, whereas non-neoplastic lung tissue was consistently TIGIT-negative. The immune checkpoint protein distribution in histologic subtypes of pulmonary adenocarcinoma displays an considerable intra- and intertumoral heterogeneity implying the requirement of either a multiregion or an adjusted analysis when determining the expression status of PD-1:PD-L1 and the TIGIT:PVR/Nectin-2 checkpoint proteins as predictive markers.
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http://dx.doi.org/10.1016/j.neo.2021.05.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8190489PMC
June 2021

A Prospective Feasibility Trial to Challenge Patient-Derived Pancreatic Cancer Organoids in Predicting Treatment Response.

Cancers (Basel) 2021 May 21;13(11). Epub 2021 May 21.

Department of Internal Medicine, University Hospital Ulm, 89081 Ulm, Germany.

Real-time isolation, propagation, and pharmacotyping of patient-derived pancreatic cancer organoids (PDOs) may enable treatment response prediction and personalization of pancreatic cancer (PC) therapy. In our methodology, PDOs are isolated from 54 patients with suspected or confirmed PC in the framework of a prospective feasibility trial. The drug response of single agents is determined by a viability assay. Areas under the curves (AUC) are clustered for each drug, and a prediction score is developed for combined regimens. Pharmacotyping profiles are obtained from 28 PDOs (efficacy 63.6%) after a median of 53 days (range 21-126 days). PDOs exhibit heterogeneous responses to the standard-of-care drugs, and are classified into high, intermediate, or low responder categories. Our developed prediction model allows a successful response prediction in treatment-naïve patients with an accuracy of 91.1% for first-line and 80.0% for second-line regimens, respectively. The power of prediction declines in pretreated patients (accuracy 40.0%), particularly with more than one prior line of chemotherapy. Progression-free survival (PFS) is significantly longer in previously treatment-naïve patients receiving a predicted tumor sensitive compared to a predicted tumor resistant regimen (mPFS 141 vs. 46 days; = 0.0048). In conclusion, generation and pharmacotyping of PDOs is feasible in clinical routine and may provide substantial benefit.
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http://dx.doi.org/10.3390/cancers13112539DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8196829PMC
May 2021

H3F3A-mutated giant cell tumour of bone without giant cells-clinical presentation, radiology and histology of three cases.

Histopathology 2021 Nov 21;79(5):720-730. Epub 2021 Jul 21.

Bone Tumour Reference Centre at the Institute of Pathology, University Hospital Basel, Basel, Switzerland.

Aims: Giant cell tumour of bone (GCTB) is histologically defined as a lesion containing reactive giant cells and a neoplastic mononuclear cell population; in up to 92% of cases, GCTB is characterised by a specific mutation of the histone gene H3F3A. The cellular composition ranges from giant-cell-rich to giant-cell-poor. The diagnosis of GCTB can be challenging, and several other lesions need to be excluded, e.g. aneurysmal bone cysts, non-ossifying fibromas, chondroblastomas, brown tumours, and giant-cell-rich osteosarcomas. Our aim was to analyse the clinical history, imaging, molecular pathology and histology of three H3F3A-mutated bone tumours without detectable giant cells. None of the patients received denosumab therapy.

Methods And Results: Diagnostic material was obtained by curettage or resection and/or biopsy. Common histomorphological features of all three reported lesions were fibrocytic, oval cells in a background of osteoid and an absence of multinuclear giant cells as confirmed with CD68 immunohistochemistry. We used immunohistochemistry and Sanger sequencing to demonstrate positivity for the H3.3 p.G34W mutation. Differential diagnoses were systematically excluded on the basis of histomorphology, immunohistochemistry, and fluorescence in-situ hybridisation. The imaging (radiography, computed tomography, and magnetic resonance imaging) for all three cases is presented and discussed.

Conclusions: We believe that these GCTBs without giant cells expand one end of the heterogeneous range of GCTB. Because of the lack of giant cells, correct diagnosis of GCTB is challenging or even impossible on histological grounds alone. In these cases, detection of the characteristic H3F3A mutation (G34W-specific antibody RM263 or sequencing) is extremely helpful for diagnosing those lesions without giant cells as giant cell tumours of bone.
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http://dx.doi.org/10.1111/his.14401DOI Listing
November 2021

Alveolar Echinococcosis of the Parotid Gland-An Ultra Rare Location Reported from Western Europe.

Pathogens 2021 Apr 3;10(4). Epub 2021 Apr 3.

Department of Otorhinolaryngology, Head and Neck Surgery, University Medical Center Bonn, Venusberg-Campus 1, 53127 Bonn, Germany.

(1) Background: (AE) is restricted to the northern hemisphere with high endemic regions in Central Europe, North and Central Asia as well as Western China. The larval stage of causes AE with tumor-like growth. Humans are accidental hosts. This report is on the first case of AE becoming clinically manifested in the parotic gland. (2) Case presentation: A 52-year-old male patient presented with progressive and painful swelling of the right parotid gland persisting for one year. We performed a partial parotidectomy. The histological examination and immunohistological staining revealed larval stage of . (3) Conclusion: is known to infect animals and humans coincidentally, and leads to AE. It is one of the most life-threatening zoonoses in Europe. It typically manifests in the liver (50-77%), with further spreading to other organs being a rare phenomenon. Echinococcosis should be considered in the differential diagnosis of lesions of the parotid gland in endemic areas, but AE has not been described so far in the parotid gland as the sole manifestation and, therefore, impedes the correct diagnosis. A complete resection should be the aim, however, preservation of the facial nerve and adjuvant albendazole therapy is mandatory.
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http://dx.doi.org/10.3390/pathogens10040426DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8067166PMC
April 2021

Modeling plasticity and dysplasia of pancreatic ductal organoids derived from human pluripotent stem cells.

Cell Stem Cell 2021 06 28;28(6):1105-1124.e19. Epub 2021 Apr 28.

Institute of Neuroanatomy & Developmental Biology (INDB), Eberhard Karls University Tübingen, Tübingen, Germany.

Personalized in vitro models for dysplasia and carcinogenesis in the pancreas have been constrained by insufficient differentiation of human pluripotent stem cells (hPSCs) into the exocrine pancreatic lineage. Here, we differentiate hPSCs into pancreatic duct-like organoids (PDLOs) with morphological, transcriptional, proteomic, and functional characteristics of human pancreatic ducts, further maturing upon transplantation into mice. PDLOs are generated from hPSCs inducibly expressing oncogenic GNAS, KRAS, or KRAS with genetic covariance of lost CDKN2A and from induced hPSCs derived from a McCune-Albright patient. Each oncogene causes a specific growth, structural, and molecular phenotype in vitro. While transplanted PDLOs with oncogenic KRAS alone form heterogenous dysplastic lesions or cancer, KRAS with CDKN2A loss develop dedifferentiated pancreatic ductal adenocarcinomas. In contrast, transplanted PDLOs with mutant GNAS lead to intraductal papillary mucinous neoplasia-like structures. Conclusively, PDLOs enable in vitro and in vivo studies of pancreatic plasticity, dysplasia, and cancer formation from a genetically defined background.
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http://dx.doi.org/10.1016/j.stem.2021.03.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8461636PMC
June 2021

Profiling of three H3F3A-mutated and denosumab-treated giant cell tumors of bone points to diverging pathways during progression and malignant transformation.

Sci Rep 2021 Mar 11;11(1):5709. Epub 2021 Mar 11.

Institute of Pathology, University of Ulm, Albert-Einstein-Allee 11, 89081, Ulm, Germany.

Giant cell tumor of bone (GCTB) is a locally aggressive lesion of intermediate malignancy. Malignant transformation of GCTB is a rare event. In 2013, the humanized monoclonal antibody against receptor activator of nuclear factor-κb-Ligand (RANKL) denosumab was approved for treatment of advanced GCTB. Since then, several reports have questioned the role of denosumab during occasional malignant transformation of GCTB. We report on three patients with H3F3A-mutated GCTBs, treated with denosumab. The tissue samples were analysed by histomorphology, immunohistochemistry, and in two instances by next generation panel sequencing of samples before and after treatment. One patient had a mutation of ARID2 in the recurrence of the GCTB under treatment with denosumab. One patient developed a pleomorphic sarcoma and one an osteoblastic osteosarcoma during treatment. Sequencing revealed a persisting H3F3A mutation in the osteosarcoma while the pleomorphic sarcoma lost the H3F3A mutation; however, a FGFR1 mutation, both in the recurrence and in the pleomorphic sarcoma persisted. In addition, the pleomorphic sarcoma showed an AKT2 and a NRAS mutation. These data are inconclusive concerning the role denosumab plays in the event of malignant progression/transformation of GCTB and point to diverging pathways of tumor progression of GCTB associated with this treatment.
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http://dx.doi.org/10.1038/s41598-021-85319-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7952552PMC
March 2021

Genome-wide DNA methylation analysis along the progression of gastric marginal zone B-cell lymphoma of mucosa-associated lymphoid tissue (MALT) type.

Br J Haematol 2021 04 16;193(2):369-374. Epub 2021 Feb 16.

Institute of Pathology, Ulm University, Ulm, Germany.

Extra-nodal marginal zone B-cell lymphoma (MZBL) of mucosa-associated lymphoid tissue is an indolent lymphoma mostly affecting the gastrointestinal tract. The lymphoma initially has small-cell morphology (SC-MZBL) and often arises in the background of Helicobacter pylori-induced gastritis. In some cases, a clonal malignant progression to large-cell morphology (LC-MZBL) is observed. Here, we studied the DNA methylation profile of 30 gastric MZBLs along their progression. Genome-wide DNA methylation profiling, identified 7698 significantly differentially methylated loci during gastric MZBL progression (σ/σ ≥0·4, q ≤ 0·001). LC-MZBL showed hypermethylation in comparison to SC-MZBL with an enrichment of regions involved in transcriptional regulation. In conclusion, our present data show that the morphological distinction between SC- and LC-MZBL is reflected by characteristic DNA methylation profiles.
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http://dx.doi.org/10.1111/bjh.17193DOI Listing
April 2021

Emerging human alveolar echinococcosis in Hungary (2003-2018): a retrospective case series analysis from a multi-centre study.

BMC Infect Dis 2021 Feb 10;21(1):168. Epub 2021 Feb 10.

WHO Collaborating Centre for the epidemiology, detection and control of cystic and alveolar echinococcosis, Department of Infectious Diseases, Istituto Superiore di Sanità, Rome, Italy.

Background: Human alveolar echinococcosis (AE) caused by Echinococcus multilocularis is an underreported, often misdiagnosed and mistreated parasitic disease mainly due to its low incidence. The aim of this study was to describe the epidemiological and clinical characteristics of human AE patients in Hungary for the first time.

Method: Between 2003 and 2018, epidemiological and clinical data of suspected AE patients were collected retrospectively from health database management systems.

Results: This case series included a total of 16 AE patients. The mean age of patients was 53 years (range: 24-78 years). The sex ratio was 1:1. Four patients (25%) revealed no recurrence after radical surgery and adjuvant albendazole (ABZ) therapy. For five patients (31.3%) with unresectable lesions, a stabilization of lesions with ABZ treatment was achieved. In seven patients (43.8%), progression of AE was documented. The mean diagnostic delay was 33 months (range: 1-122 months). Three AE related deaths (fatality rate 18.8%) were recorded.

Conclusions: AE is an emerging infectious disease in Hungary with a high fatality rate since based on our results, almost every fifth AE patient died in the study period. Differential diagnosis and appropriate surgical and medical therapy for AE is an urging challenge for clinicians in Hungary, as well as in some other European countries where E. multilocularis is prevalent.
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http://dx.doi.org/10.1186/s12879-021-05859-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7877032PMC
February 2021

SARS-CoV-2 infects and replicates in cells of the human endocrine and exocrine pancreas.

Nat Metab 2021 02 3;3(2):149-165. Epub 2021 Feb 3.

Institute of Diabetes and Regeneration Research, Helmholtz Zentrum München, Neuherberg, Germany.

Infection-related diabetes can arise as a result of virus-associated β-cell destruction. Clinical data suggest that the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), causing the coronavirus disease 2019 (COVID-19), impairs glucose homoeostasis, but experimental evidence that SARS-CoV-2 can infect pancreatic tissue has been lacking. In the present study, we show that SARS-CoV-2 infects cells of the human exocrine and endocrine pancreas ex vivo and in vivo. We demonstrate that human β-cells express viral entry proteins, and SARS-CoV-2 infects and replicates in cultured human islets. Infection is associated with morphological, transcriptional and functional changes, including reduced numbers of insulin-secretory granules in β-cells and impaired glucose-stimulated insulin secretion. In COVID-19 full-body postmortem examinations, we detected SARS-CoV-2 nucleocapsid protein in pancreatic exocrine cells, and in cells that stain positive for the β-cell marker NKX6.1 and are in close proximity to the islets of Langerhans in all four patients investigated. Our data identify the human pancreas as a target of SARS-CoV-2 infection and suggest that β-cell infection could contribute to the metabolic dysregulation observed in patients with COVID-19.
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http://dx.doi.org/10.1038/s42255-021-00347-1DOI Listing
February 2021

Immunohistological detection of small particles of Echinococcus multilocularis and Echinococcus granulosus in lymph nodes is associated with enlarged lymph nodes in alveolar and cystic echinococcosis.

PLoS Negl Trop Dis 2020 12 28;14(12):e0008921. Epub 2020 Dec 28.

Institute of Pathology, University Ulm, Ulm, Germany.

Background: Alveolar (AE) and cystic echinococcosis (CE) in humans are caused by the metacestode of the tapeworms Echinococcus multilocularis and Echinococcus granulosus sensu lato (s.l.). Immunohistochemistry with the monoclonal antibodies (mAb) Em2G11, specific for AE, and the mAb EmG3, specific for AE and CE, is an important pillar of the histological diagnosis of these two infections. Our aim was to further evaluate mAb EmG3 in a diagnostic setting and to analyze in detail the localization, distribution, and impact of small particles of Echinococcus multilocularis (spems) and small particles of Echinococcus granulosus s.l. (spegs) on lymph nodes.

Methodology/principal Findings: We evaluated the mAb EmG3 in a cohort of formalin-fixed, paraffin embedded (FFPE) specimens of AE (n = 360) and CE (n = 178). These samples originated from 156 AE-patients and 77 CE-patients. mAb EmG3 showed a specific staining of the metacestode stadium of E. multilocularis and E. granulosus s.l. and had a higher sensitivity for spems than mAb Em2G11. Furthermore, we detected spegs in the surrounding host tissue and in almost all tested lymph nodes (39/41) of infected patients. 38/47 lymph nodes of AE showed a positive reaction for spems with mAb EmG3, whereas 29/47 tested positive when stained with mAb Em2G11. Spegs were detected in the germinal centers, co-located with CD23-positive follicular dendritic cells, and were present in the sinuses. Likewise, lymph nodes with spems and spegs in AE and CE were significantly enlarged in size in comparison to the control group.

Conclusions/significance: mAb EmG3 is specific for AE and CE and is a valuable tool in the histological diagnosis of echinococcosis. Based on the observed staining patterns, we hypothesize that the interaction between parasite and host is not restricted to the main lesion since spegs are detected in lymph nodes. Moreover, in AE the number of spems-affected lymph nodes is higher than previously assumed. The enlargement of lymph nodes with spems and spegs points to an immunological interaction with the small immunogenic particles (spems and spegs) of Echinococcus spp.
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http://dx.doi.org/10.1371/journal.pntd.0008921DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7769273PMC
December 2020

U-RT1 - A new model for Richter transformation.

Neoplasia 2021 01 11;23(1):140-148. Epub 2020 Dec 11.

Institute of Pathology, University Hospital Ulm, Ulm, Germany.

The advent of highly effective treatments targeting the disease biology of chronic lymphocytic leukemia (CLL) has transformed the therapeutic field tremendously. However, transformation into an aggressive B-cell lymphoma, called Richter syndrome (RS), remains highly challenging since the treatment options for this condition are still insufficient. Exploratory drug testing and experimental studies are restricted by the lack of satisfactory models. We have established U-RT1, a cell line derived from a highly proliferating RS clonally related to the patient's underlying CLL. The cell line shows morphological features and an immunophenotype of RS-DLBCL (non-GCB). Molecular analysis revealed a complex karyotype with driver aberrations characteristic for RS such as loss of TP53 and CDKN2A. Furthermore, U-RT1 displays a chromosomal gain of the NOTCH1 gene locus and strong immunoreactivity for BCL-2. These features suggest that U-RT1 is the first eligible model system for investigations on the pathogenesis of RS and novel treatment options.
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http://dx.doi.org/10.1016/j.neo.2020.11.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7736907PMC
January 2021

Successful hematopoietic stem cell transplantation in a 4-1BB deficient patient with EBV-induced lymphoproliferation.

Clin Immunol 2021 01 28;222:108639. Epub 2020 Nov 28.

Department of Pediatrics and Adolescent Medicine, University Medical Center Ulm, Germany.

Complete remission from recurrent EBV-positive lymphoma is not mandatory before HSCT to achieve long-term cure in a patient suffering from a recently described immunodeficiency affecting the T-cell coactivation molecule 4-1BB.
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http://dx.doi.org/10.1016/j.clim.2020.108639DOI Listing
January 2021

Drug Inhibition of SARS-CoV-2 Replication in Human Pluripotent Stem Cell-Derived Intestinal Organoids.

Cell Mol Gastroenterol Hepatol 2021 10;11(4):935-948. Epub 2020 Nov 10.

Department of Internal Medicine I, Ulm University Hospital, Ulm, Germany. Electronic address:

Background And Aims: The COVID-19 pandemic has spread worldwide and poses a severe health risk. While most patients present mild symptoms, descending pneumonia can lead to severe respiratory insufficiency. Up to 50% of patients show gastrointestinal symptoms like diarrhea or nausea, intriguingly associating with prolonged symptoms and increased severity. Thus, models to understand and validate drug efficiency in the gut of COVID-19 patients are of urgent need.

Methods: Human intestinal organoids derived from pluripotent stem cells (PSC-HIOs) have led, due to their complexity in mimicking human intestinal architecture, to an unprecedented number of successful disease models including gastrointestinal infections. Here, we employed PSC-HIOs to dissect SARS-CoV-2 pathogenesis and its inhibition by remdesivir, one of the leading drugs investigated for treatment of COVID-19.

Results: Immunostaining for viral entry receptor ACE2 and SARS-CoV-2 spike protein priming protease TMPRSS2 showed broad expression in the gastrointestinal tract with highest levels in the intestine, the latter faithfully recapitulated by PSC-HIOs. Organoids could be readily infected with SARS-CoV-2 followed by viral spread across entire PSC-HIOs, subsequently leading to organoid deterioration. However, SARS-CoV-2 spared goblet cells lacking ACE2 expression. Importantly, we challenged PSC-HIOs for drug testing capacity. Specifically, remdesivir effectively inhibited SARS-CoV-2 infection dose-dependently at low micromolar concentration and rescued PSC-HIO morphology.

Conclusions: Thus, PSC-HIOs are a valuable tool to study SARS-CoV-2 infection and to identify and validate drugs especially with potential action in the gut.
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http://dx.doi.org/10.1016/j.jcmgh.2020.11.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7655023PMC
April 2021

Maintenance Therapy for ATM-Deficient Pancreatic Cancer by Multiple DNA Damage Response Interferences after Platinum-Based Chemotherapy.

Cells 2020 09 16;9(9). Epub 2020 Sep 16.

Department of Internal Medicine 1, University Medical Center Ulm, Albert-Einstein-Allee 23, 89081 Ulm, Germany.

Personalized medicine in treating pancreatic ductal adenocarcinoma (PDAC) is still in its infancy, albeit PDAC-related deaths are projected to rise over the next decade. Only recently, maintenance therapy with the PARP inhibitor olaparib showed improved progression-free survival in germline -mutated PDAC patients after platinum-based induction for the first time. Transferability of such a concept to other DNA damage response (DDR) genes remains unclear. Here, we conducted a placebo-controlled, three-armed preclinical trial to evaluate the efficacy of multi-DDR interference (mDDRi) as maintenance therapy vs. continuous FOLFIRINOX treatment, implemented with orthotopically transplanted ATM-deficient PDAC cell lines. Kaplan-Meier analysis, cross-sectional imaging, histology, and in vitro analysis served as analytical readouts. Median overall survival was significantly longer in the mDDRi maintenance arm compared to the maintained FOLFIRINOX treatment. This survival benefit was mirrored in the highest DNA-damage load, accompanied by superior disease control and reduced metastatic burden. In vitro analysis suggests FOLFIRINOX-driven selection of invasive subclones, erased by subsequent mDDRi treatment. Collectively, this preclinical trial substantiates mDDRi in a maintenance setting as a novel therapeutic option and extends the concept to non-germline -mutant PDAC.
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http://dx.doi.org/10.3390/cells9092110DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7563330PMC
September 2020

Clostridial C3 Toxins Enter and Intoxicate Human Dendritic Cells.

Toxins (Basel) 2020 09 1;12(9). Epub 2020 Sep 1.

Institute of Pharmacology and Toxicology, University of Ulm Medical Center, 89081 Ulm, Germany.

C3 protein toxins produced by and are mono-ADP-ribosyltransferases, which specifically modify the GTPases Rho A/B/C in the cytosol of monocytic cells, thereby inhibiting Rho-mediated signal transduction in monocytes, macrophages, and osteoclasts. C3 toxins are selectively taken up into the cytosol of monocytic cells by endocytosis and translocate from acidic endosomes into the cytosol. The C3-catalyzed ADP-ribosylation of Rho proteins inhibits essential functions of these immune cells, such as migration and phagocytosis. Here, we demonstrate that C3 toxins enter and intoxicate dendritic cells in a time- and concentration-dependent manner. Both immature and mature human dendritic cells efficiently internalize C3 exoenzymes. These findings could also be extended to the chimeric fusion toxin C2IN-C3lim. Moreover, stimulated emission depletion (STED) microscopy revealed the localization of the internalized C3 protein in endosomes and emphasized its potential use as a carrier to deliver foreign proteins into dendritic cells. In contrast, the enzyme C2I from the binary C2 toxin was not taken up into dendritic cells, indicating the specific uptake of C3 toxins. Taken together, we identified human dendritic cells as novel target cells for clostridial C3 toxins and demonstrated the specific uptake of these toxins via endosomal vesicles.
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http://dx.doi.org/10.3390/toxins12090563DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7551598PMC
September 2020

Rescue of Non-Informative Circulating Tumor DNA to Monitor the Mutational Landscape in NSCLC.

Cancers (Basel) 2020 Jul 16;12(7). Epub 2020 Jul 16.

Institute of Pathology, University Medical Center Ulm, 89070 Ulm, Germany.

In non-small cell lung cancer (NSCLC) the usage of plasma-derived circulating tumor DNA (ctDNA) have come into focus to obtain a comprehensive genetic profile of a given lung cancer. Despite the usage of specific sampling tubes, archived plasma samples as well as inappropriately treated blood samples still cause a loss of information due to cell lysis and contamination with cellular DNA. Our aim was to establish a reliable protocol to rescue ctDNA from such non-informative samples to monitor the mutational landscape in NSCLC. As a proof-of-concept study we used archived plasma samples derived from whole blood EDTA samples of 51 patients suffering from NSCLC. Analysis of the isolated plasma DNA determined only a small fraction of ctDNA in a range of 90-250 bp. By applying a specific purification procedure, we were able to increase the informative ctDNA content and improve in a cohort of 42 patients the detection of driver mutations from 32% to 79% of the mutations found in tissue biopsies. Thus, we present here an easy to perform, time and cost effective procedure to rescue non-informative ctDNA samples, which is sufficient to detect oncogenic mutations in NGS approaches and is therefore a valuable technical improvement for laboratories handling liquid biopsy samples.
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http://dx.doi.org/10.3390/cancers12071917DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7409026PMC
July 2020

Hepatic alveolar echinococcosis: correlation between computed tomography morphology and inflammatory activity in positron emission tomography.

Sci Rep 2020 07 16;10(1):11808. Epub 2020 Jul 16.

Department of Internal Medicine III, Ulm University Hospital, Albert-Einstein-Allee 23, 89081, Ulm, Germany.

Positron emission tomography-computed tomography (PET-CT) with 18F-fluorodesoxyglucose (FDG) is the imaging modality of choice for assessing inflammation surrounding hepatic alveolar echinococcosis (AE) lesions. This study is the first to evaluate FDG uptake in hepatic AE (n = 51) based on the standardized uptake value (SUV) and to correlate the SUVs with primary morphology and calcification patterns, based on the Echinococcus multilocularis Ulm Classification for Computed-Tomography (EMUC-CT). Our results show that the SUVs were increased for lesions with EMUC-CT types I-IV primary morphology, compared to the surrounding healthy liver tissue (SUV = 2.5 ± 0.4; p < 0.05). Type IV lesions included, by far, the highest number of PET-negative lesions. A comparison of lesions with different primary morphologies showed clear differences. The highest SUVs were found for types I and III, and the lowest was found for type IV. Type IV lesions (SUV, 3.8 ± 1.5) showed significantly lower uptake compared to type I (SUV, 6.9 ± 3.5; p = 0.030) and type III (SUV, 7.4 ± 3.9; p = 0.031) lesions. For type II lesions, the results showed only a statistical trend (SUV, 6.1 ± 3.1; p = 0.073). Due to the small number of cases, an evaluation of type V (n = 1) lesions was not possible. The different SUVs of lesions with different primary morphologies, particularly the lower FDG uptake observed in type IV lesions, suggested that these SUVs might reflect different stages of the disease.
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http://dx.doi.org/10.1038/s41598-020-68624-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7366930PMC
July 2020

[The gut: center of immunity : Rare inflammatory bowel diseases caused by immunodeficiencies].

Pathologe 2020 May;41(3):211-223

Institut für Pathologie, Universitätsklinikum Ulm, Albert-Einstein-Allee 8, 89081, Ulm, Deutschland.

The gut is the largest immune organ of the human body with an enormous mucosal interface. By acting as a physical barrier and by hosting many of the body's immune cells and tissues, the gut is the first line of defense against potentially harmful substances. Therefore, diseases leading to impaired immune response or disruption of the epithelial barrier result in autoimmune, infectious, or inflammatory bowel disease, frequently associated with diarrhea, malabsorption, melena, and growth failure. The differential diagnosis represents an interdisciplinary challenge in this group of rare diseases. The diseases are characterized by clinical, immunological, and histopathological features caused by mutations in single genes. In the following, we will focus on histological findings within the various entities of immunodeficiencies.
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http://dx.doi.org/10.1007/s00292-020-00775-yDOI Listing
May 2020

First European Haplotype of Echinococcus multilocularis Identified in the United States: An Emerging Disease?

Clin Infect Dis 2021 04;72(7):1117-1123

Department of Pathology and Laboratory Medicine, University of Vermont Larner College of Medicine, Burlington, Vermont, USA.

Background: Echinococcus multilocularis is one of the most severe and lethal parasitic diseases of humans, most often reported in Europe and Asia. Only 1 previous case has been documented in the contiguous United States from Minnesota in 1977. European haplotypes have been identified in carnivores and domestic dogs as well as recently in patients in western and central Canada.

Methods: We used immunohistochemical testing with the monoclonal antibody Em2G11 and a species-specific enzyme-linked immunosorbent assay affinity-purified antigen Em2, as well as COX1 gene sequencing.

Results: Using pathology, immunohistochemical staining, specific immunodiagnostic testing, and COX1 gene sequencing, we were able to definitively identify E. multilocularis as the causative agent of our patient's liver and lung lesions, which clustered most closely with the European haplotype.

Conclusions: We have identified the first case of a European haplotype E. multilocularis in the United States and the first case of this parasitic infection east of the Mississippi River. Given the identification of this haplotype in Canada, this appears to be an emerging infectious disease in North America.
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http://dx.doi.org/10.1093/cid/ciaa245DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8028098PMC
April 2021

Pathology of Echinococcosis: A Morphologic and Immunohistochemical Study on 138 Specimens With Focus on the Differential Diagnosis Between Cystic and Alveolar Echinococcosis.

Am J Surg Pathol 2020 01;44(1):43-54

Department of Pathology and Molecular Pathology.

Infection of humans by the larval stage of the tapeworms Echinococcus granulosus sensu lato or Echinococcus multilocularis causes the life-threatening zoonoses cystic echinococcosis (CE) and alveolar echinococcosis (AE). Although cystic liver lesions are a hallmark of both diseases, course, prognosis, and patients' management decisively differ between the two. The wide and overlapping spectrum of morphologies and the limited availability of ancillary tools are challenges for pathologists to reliably diagnose and subtype echinococcosis. Here, we systematically and quantitatively recorded the pathologic spectrum in a clinically and molecularly defined echinococcosis cohort (138 specimens from 112 patients). Immunohistochemistry using a novel monoclonal antibody (mAbEmG3) was implemented, including its combined application with the mAbEm2G11. Six morphologic criteria sufficiently discriminated between CE and AE: size of smallest (CE/AE: >2/≤2 mm) and largest cyst (CE/AE: >25/≤25 mm), thickness of laminated layer (CE/AE: >0.15/≤0.15 mm) and pericystic fibrosis (CE/AE: >0.6/≤0.6 mm), striation of laminated layer (CE/AE: moderate-strong/weak), and number of cysts (CE/AE: ≤9/>9). Combined immunohistochemistry with mAbEm2G11 (E. multilocularis specific) and mAbEmG3 (reactive in AE and CE) was equally specific as and occasionally more sensitive than polymerase chain reaction. On the basis of these findings, we developed a diagnostic algorithm for the differential diagnosis of echinococcosis. In summary, we have not only identified the means to diagnose echinococcosis with greater certainty, but also defined morphologic criteria, which robustly discriminate between CE and AE. We expect our findings to improve echinococcosis diagnostics, especially of challenging cases, beneficially impacting the management of echinococcosis patients.
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http://dx.doi.org/10.1097/PAS.0000000000001374DOI Listing
January 2020

Simple liver cysts and cystoid lesions in hepatic alveolar echinococcosis: a retrospective cohort study with Hounsfield analysis.

Parasite 2019 30;26:54. Epub 2019 Aug 30.

Department of Diagnostic and Interventional Radiology, University Hospital Ulm, Albert-Einstein-Allee 23, 89081 Ulm, Germany.

Background: Alveolar echinococcosis (AE) is a rare zoonosis caused by the larval stage of the tapeworm Echinococcus multilocularis. AE lesions affect the liver in more than 98% of cases. AE lesions have various morphological characteristics that are described in the Echinococcus multilocularis Ulm classification for computed tomography (EMUC-CT). One of these characteristics is a cystoid portion. The aim of the study was to compare the density of simple hepatic cysts with cystoid portions of AE lesions classified on the basis of the EMUC-CT.

Results: Hounsfield Unit (HU) measurements of the cystoid portions of all EMUC-CT type I-IV AE lesions (n = 155) gave a mean of 21.8 ± 17.6, which was significantly different from that of 2.9 ± 4.5 for the simple hepatic cysts (p < 0.0001). The difference between each of the individual AE types and simple hepatic cysts was also significant. In addition, the HU values of the cystoid portions in types I, II and IIIa/b and simple cysts were each significantly different from type IV (p < 0.0001). The HU measurements in type IV presented by far the highest mean.

Conclusions: The significantly higher density measured in the cystoid portions of hepatic AE lesions offers a good means of differentiation from simple hepatic cysts.
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http://dx.doi.org/10.1051/parasite/2019057DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6716343PMC
December 2019

Echinococcus multilocularis.

Trends Parasitol 2019 09 7;35(9):738-739. Epub 2019 Jun 7.

WHO Collaborating Centre for the Epidemiology, Detection, and Control of Cystic and Alveolar Echinococcosis (in Animals and Humans), Department of Infectious Diseases, Istituto Superiore di Sanità, Rome, Italy; European Reference Laboratory for Parasites, Department of Infectious Diseases, Istituto Superiore di Sanità, Rome, Italy.

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http://dx.doi.org/10.1016/j.pt.2019.05.005DOI Listing
September 2019

Characterization of Three Novel H3F3A-mutated Giant Cell Tumor Cell Lines and Targeting of Their Wee1 Pathway.

Sci Rep 2019 04 23;9(1):6458. Epub 2019 Apr 23.

Institute of Pathology, Ulm University, Ulm, Germany.

The giant cell tumor of bone (GCTB) is a locally aggressive primary bone tumor that is composed of mononuclear stroma cells, scattered macrophages, and multinucleated osteoclast-like giant cells which cause pathologic osteolysis. The stroma cells represent the neoplastic population of the tumor and are characterized by the H3F3A mutation G34W. This point mutation is regarded as the driver mutation of GCTB. We have established three new stable H3F3A mutated GCTB cell lines: U-GCT1, U-GCT2, and U-GCT3M. MK-1775 is a Wee1-kinase inhibitor which has been used for blocking of sarcoma growth. In the cell lines we detected Wee1, Cdk1, Cyclin B1, H3K36me3, and Rrm2 as members of the Wee1 pathway. We analyzed the effect of MK-1775 and gemcitabine, alone and in combination, on the growth of the cell lines. The cell lines showed a significant reduction in cell proliferation when treated with MK-1775 or gemcitabine. The combination of both agents led to a further significant reduction in cell proliferation compared to the single agents. Immunohistochemical analysis of 13 GCTB samples revealed that Wee1 and downstream-relevant members are present in GCTB tissue samples. Overall, our work offers valuable new tools for GCTB studies and presents a description of novel biomarkers and molecular targeting strategies.
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http://dx.doi.org/10.1038/s41598-019-42611-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6478864PMC
April 2019

Analysis of the CDK4/6 Cell Cycle Pathway in Leiomyosarcomas as a Potential Target for Inhibition by Palbociclib.

Sarcoma 2019 21;2019:3914232. Epub 2019 Jan 21.

Institute of Pathology, Ulm University, Ulm, Germany.

Leiomyosarcoma (LMS) is characterized by high genomic complexity, and to date, no specific targeted therapy is available. In a genome-wide approach, we profiled genomic aberrations in a small cohort of eight primary tumours, two relapses, and eight metastases across nine different patients. We identified CDK4 amplification as a recurrent alteration in 5 out of 18 samples (27.8%). It has been previously shown that the LMS cell line SK-LMS-1 has a defect in the p16 pathway and that this cell line can be inhibited by the CDK4 and CDK6 inhibitor palbociclib. For SK-LMS-1 we confirm and for SK-UT-1 we show that both LMS cell lines express CDK4 and that, in addition, strong CDK6 expression is seen in SK-LMS-1, whereas Rb was expressed in SK-LMS-1 but not in SK-UT-1. We confirm that inhibition of SK-LMS-1 with palbociclib led to a strong decrease in protein levels of Phospho-Rb (Ser780), a decreased cell proliferation, and G/G-phase arrest with decreased S/G fractions. SK-UT-1 did not respond to palbociclib inhibition. To compare these findings with patient tissue samples, a p16, CDK4, CDK6, and p-Rb immunohistochemical staining assay of a large LMS cohort (=99 patients with 159 samples) was performed assigning a potential responder phenotype to each patient, which we identified in 29 out of 99 (29.3%) patients. Taken together, these data show that CDK4/6 inhibitors may offer a new option for targeted therapy in a subset of LMS patients.
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http://dx.doi.org/10.1155/2019/3914232DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6360577PMC
January 2019

YAP Activation Drives Liver Regeneration after Cholestatic Damage Induced by Deletion.

Int J Mol Sci 2018 Nov 29;19(12). Epub 2018 Nov 29.

Department of Internal Medicine I, Ulm University, 89081 Ulm, Germany.

Liver cholestasis is a chronic liver disease and a major health problem worldwide. Cholestasis is characterised by a decrease in bile flow due to impaired secretion by hepatocytes or by obstruction of bile flow through intra- or extrahepatic bile ducts. Thereby cholestasis can induce ductal proliferation, hepatocyte injury and liver fibrosis. Notch signalling promotes the formation and maturation of bile duct structures. Here we investigated the liver regeneration process in the context of cholestasis induced by disruption of the Notch signalling pathway. Liver-specific deletion of recombination signal binding protein for immunoglobulin kappa j region (), which represents a key regulator of Notch signalling, induces severe cholestasis through impaired intra-hepatic bile duct (IHBD) maturation, severe necrosis and increased lethality. Deregulation of the biliary compartment and cholestasis are associated with the change of several signalling pathways including a Kyoto Encyclopedia of Genes and Genomes (KEGG) gene set representing the Hippo pathway, further yes-associated protein (YAP) activation and upregulation of SRY (sex determining region Y)-box 9 (SOX9), which is associated with transdifferentiation of hepatocytes. SOX9 upregulation in cholestatic liver injury in vitro is independent of Notch signalling. We could comprehensively address that in vivo depletion is followed by YAP activation, which influences the transdifferentiation of hepatocytes and thereby contributing to liver regeneration.
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http://dx.doi.org/10.3390/ijms19123801DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6321044PMC
November 2018

Boosting Antitumor Drug Efficacy with Chemically Engineered Multidomain Proteins.

Adv Sci (Weinh) 2018 Aug 14;5(8):1701036. Epub 2018 Jun 14.

Max-Planck Institute for Polymer Research Ackermannweg 10 55128 Mainz Germany.

A facile chemical approach integrating supramolecular chemistry, site-selective protein chemistry, and molecular biology is described to engineer synthetic multidomain protein therapeutics that sensitize cancer cells selectively to significantly enhance antitumor efficacy of existing chemotherapeutics. The desired bioactive entities are assembled via supramolecular interactions at the nanoscale into structurally ordered multiprotein complexes comprising a) multiple copies of the chemically modified cyclic peptide hormone somatostatin for selective targeting and internalization into human A549 lung cancer cells expressing SST-2 receptors and b) a new cysteine mutant of the C3bot1 (C3) enzyme from , a Rho protein inhibitor that affects and influences intracellular Rho-mediated processes like endothelial cell migration and blood vessel formation. The multidomain protein complex, SST3-Avi-C3, retargets C3 enzyme into non-small cell lung A549 cancer cells and exhibits exceptional tumor inhibition at a concentration ≈100-fold lower than the clinically approved antibody bevacizumab (Avastin) in vivo. Notably, SST3-Avi-C3 increases tumor sensitivity to a conventional chemotherapeutic (doxorubicin) in vivo. These findings show that the integrated approach holds vast promise to expand the current repertoire of multidomain protein complexes and can pave the way to important new developments in the area of targeted and combination cancer therapy.
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http://dx.doi.org/10.1002/advs.201701036DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6097141PMC
August 2018
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