Open Access

Tumor heterogeneity encompasses genetic, epigenetic, and phenotypic diversity, impacting treatment response and resistance. Spatial heterogeneity occurs both inter- and intra-lesionally, while temporal heterogeneity results from clonal evolution. High-throughput technologies like next-generation sequencing (NGS) enhance tumor characterization, but conventional biopsies still do not adequately capture genetic heterogeneity. Liquid biopsy (LBx), analyzing circulating tumor DNA (ctDNA), provides a minimally invasive alternative, offering real-time tumor evolution insights and identifying resistance mutations overlooked by tissue biopsies. This study evaluates the capability of LBx to capture tumor heterogeneity by comparing genetic profiles from multiple metastatic lesions and LBx samples. Eight patients from the Augsburger Longitudinal Plasma Study with various types of cancer provided 56 postmortem tissue samples, which were compared against pre-mortem LBx-derived circulating-free DNA sequenced by NGS. Tissue analyses revealed significant mutational diversity (4-12 mutations per patient, VAFs: 1.5-71.4%), with distinct intra- and inter-lesional heterogeneity. LBx identified 51 variants (4-17 per patient, VAFs: 0.2-31.1%), which overlapped with mutations from the tissue samples by 33-92%. Notably, 22 tissue variants were absent in LBx, whereas 18 LBx-exclusive variants were detected (VAFs: 0.2-2.8%). LBx effectively captures tumor heterogeneity, but should be used in conjunction with tissue biopsies for comprehensive genetic profiling.

Objective: The utility of intraoperative radiotherapy (IORT) has been demonstrated in many and diverse surgical fields. However, data regarding the application in patients with brain metastases is scares. Methods: A retrospective study of nonconsecutive patients following surgical resection and IORT of brain metastasis between January 2013 and October 2020 was performed. Surgical and Oncological outcome was evaluated. Results: 42 patients were identified. 55% were Female. Median Age was 61 (range 39 – 83). Breast cancer and malignant melanoma were seen the most common pathologies and were found in 19% an 17% of cases, respectively. The laterality was distributed equal and 86% were located supratentorial. 14% of the metastasis were seen in eloquent regions. IORT delivered a mean of 20 Gy (range 13 – 20). Mean Follow up was 36 Months (range 1 -180). Local tumor control was seen in 88% of all cases. Surgical complications were seen in 7% of cases (2 wound healing disorder and one case of postoperative hemorrhage). Conclusions: IORT is a safe and effective method to achieve sustainable local tumor control for patients with brain metastases.

We recently proposed Stroma AReactive Invasion Front Areas (SARIFA), defined as direct tumor-adipocyte interaction at the invasion front, as a novel hematoxylin-and-eosin (H&E)-based histopathological prognostic biomarker in various cancers. Given that microsatellite instability, BRAF, and RAS mutation status are routinely tested for colorectal cancers (CRC), studying SARIFA’s additional prognostic value within these molecular subgroups is crucial. In addition, exploring whether the survival benefit from adjuvant therapy differs according to SARIFA-status may enhance patient treatment and outcome. SARIFA-status, BRAF, RAS, and DNA mismatch repair (MMR) status were available for 1726 CRC patients from the prospective Netherlands Cohort Study (NLCS, 1986–2006). In this study, we investigated (1) the relationship between SARIFA-status and CRC molecular characteristics, (2) the prognostic value of SARIFA-status within these molecular subgroups, and (3) whether SARIFA-status was associated with survival benefit from adjuvant therapy. SARIFA-positive CRCs more frequently showed a BRAF mutation compared to SARIFA-negative CRCs (P<0.001). BRAF-mutant/MMR-proficient CRCs were enriched in SARIFA-positive cases. SARIFA-positivity was associated with poor CRC-specific (HRrange: 1.47 to 1.78) and overall survival (HRrange: 1.35 to 1.70) within all molecular subgroups except MMR-deficient CRCs. Patients with SARIFA-positive CRC showed a CRC-specific survival benefit from adjuvant therapy compared to surgery alone (HRCRC-specific: 0.59; 95% CI: 0.44-0.79), while no CRC-specific survival benefit was observed for patients with SARIFA-negative CRC. To conclude, our results indicate that SARIFA-positivity is more common in the aggressive subset of BRAF-mutant and BRAF-mutant/MMR-proficient CRCs. Moreover, SARIFA-positivity provides additional prognostic value within molecular subgroups based on BRAF, RAS, and MMR status, suggesting that it may enhance prognostic stratification of CRC patients.

Background: The diagnosis of hematologic neoplasms is usually based on a synopsis of the peripheral blood (PB) and bone marrow findings. Morphology continues to be the cornerstone, but genetic analysis plays an increasingly important role. In routine workup, molecular genetic analysis is performed from a bone marrow aspirate (BMA). In the event of inadequate aspiration, PB is used. Not infrequently, however, PB only partially represents the disease. In this situation, molecular genetic analysis of formalin-fixed and paraffin-embedded (FFPE) bone marrow core biopsy (BMCB) could be a better alternative than PB. However, no systematic correlation of genetic findings from BMCB with results from BMA and PB has been reported. Methods: Therefore, BMCB obtained during routine diagnostics were subjected to post hoc molecular genetic analysis using next generation sequencing (NGS). The identified molecular genetic alterations were then compared with data within routine diagnostics of the corresponding BMA and/or PB. Results: In total, 29 BMCB and corresponding BMA samples were analyzed, and in 12/29 cases PB was additionally available. The analysis of BMCB and BMA showed identical results in 17 cases, but BMCB revealed a gain of information in 11, while in only 1 case, BMCB failed to identify the mutations in comparison to BMA. Conclusions: Despite the small numbers, molecular genetic analysis of bone marrow core biopsy using next generation sequencing could detect relevant additional gene mutations compared to bone marrow aspirate and/or peripheral blood.

Delta-like ligand 3 (DLL3) is frequently expressed in pulmonary small cell neuroendocrine carcinoma (SCNEC) and has emerged as a promising therapeutic target. However, limited data on DLL3 expression in other neuroendocrine neoplasms (NEN), such as extrapulmonary SCNEC, large cell neuroendocrine carcinomas (LCNEC), mixed neuroendocrine-non-neuroendocrine neoplasms (MiNEN), gastroenteropancreatic neuroendocrine tumours (GEP-NET), and pulmonary carcinoids, impedes an estimation if other types of NEN might be suitable candidates for anti-DLL3 therapies. We evaluated DLL3 expression in 1294 NEN and 479 non-neuroendocrine carcinomas, correlating the findings with histological subtypes, tumour localisation, and overall survival (OS). Furthermore, we explored the concordance of DLL3 expression during metastatic progression in 67 paired primary NEN and metastases. DLL3 expression was significantly higher in NEC (64.0%) compared to GEP-NET and pulmonary carcinoids (10.1%, p < 0.001), particularly in SCNEC (80.4%), followed by LCNEC (62.6%) and MiNEN (28.6%). DLL3 was common in pulmonary carcinoids (41.5%), but rare in GEP-NET (5.1%) and non-neuroendocrine carcinomas (1.3%). Overall DLL3 expression was highly concordant between metastases and corresponding primary NEN (92.5%, p < 0.001). In univariable analyses, DLL3-expressing pulmonary carcinoids (p = 0.005) and GEP-NET (p = 0.018) were associated with decreased OS, but this was not retained in multivariable analyses adjusting for stage and grade (p = n. s.). No prognostic impact was observed in pulmonary (p = 0.708) or GEP-NEC (p = 0.87). Our study highlights significant differences in DLL3 expression across NEN subtypes and localisations, with largely concordant expression in metastases. DLL3-based therapies may be effective in many NEC and pulmonary carcinoids, while DLL3 appears to be a minor therapeutic target for GEP-NET and non-neuroendocrine carcinomas.

Despite the addition of immune checkpoint blockade to first-line chemotherapy, the prognosis for patients with small cell lung cancer (SCLC) is still devastating. For the subset of SCLC with somatostatin receptor (SSTR) overexpression, radiopharmaceutical therapy (RPT) might be an effective future treatment option. Methods: Here, we present the case of a heavily pretreated stage IV SCLC patient showing an exceptional response to SSTR-directed RPT. A comprehensive translational work-up consisting of histopathologic, immunohistochemical, and molecular pathology analyses at different time points during treatment and especially of lesions with discordant tracer uptake was performed. Results: Besides a promising response to RPT, interesting signs of clonal dynamics under therapy and, most importantly, SSTR downregulation of some lesions as a potential evasion mechanism to SSTR-directed RPT could be identified. Conclusion: This unique investigation for a clinical–molecular understanding of novel treatment paradigms in SCLC may provide the basis for future treatment designs.

Prostate cancer is a dominant health concern calling for advanced diagnostic tools. Utilizing digital pathology and artificial intelligence, this study explores the potential of 11 deep neural network architectures for automated Gleason grading in prostate carcinoma focusing on comparing traditional and recent architectures. A standardized image classification pipeline, based on the AUCMEDI framework, facilitated robust evaluation using an in-house dataset consisting of 34,264 annotated tissue tiles. The results indicated varying sensitivity across architectures, with ConvNeXt demonstrating the strongest performance. Notably, newer architectures achieved superior performance, even though with challenges in differentiating closely related Gleason grades. The ConvNeXt model was capable of learning a balance between complexity and generalizability. Overall, this study lays the groundwork for enhanced Gleason grading systems, potentially improving diagnostic efficiency for prostate cancer.

Borna disease, which is a severe encephalitis that primarily affects horses and sheep, has been recognised for over two centuries. Borna disease virus 1 (BoDV-1) has been identified as a cause of a predominantly fatal encephalitis in humans. Little scientific data exist regarding the virus' transmission, entry portal, and excretion routes. Lesional patterns, immunological responses, and pathogenetic mechanisms remain largely unexplored in both reservoir and dead-end hosts. This systematic review compiles current knowledge on these aspects and provides guidance for future research. PubMed, ScienceDirect, and EBSCO were searched for publications from Jan 1, 2000, to April 30, 2024. 823 records were found, of which 41 studies were included. This systematic review discusses BoDV-1 transmission, pathogenesis, histopathological changes, and immunology in both reservoir and dead-end hosts, with special regard for humans. The exact propagation mechanisms, entry portal, and viral spread within the CNS are not entirely clear in humans. Although more data exist in animals, much remains hypothetical. Future research should focus on identifying potential entry sites and viral spread in dead-end hosts, which could help to clarify the pathogenesis and lesion distribution in the CNS, thereby contributing to a better understanding of BoDV-1 infection in humans and parallels with animal infections.

Introduction Whole Exome Sequencing (WES) has emerged as an efficient tool in clinical cancer diagnostics to broaden the scope from panel-based diagnostics to screening of all genes and enabling robust determination of complex biomarkers in a single analysis. Methods To assess concordance, six formalin-fixed paraffin-embedded (FFPE) tissue specimens and four commercial reference standards were analyzed by WES as matched tumor-normal DNA at 21 NGS centers in Germany, each employing local wet-lab and bioinformatics investigating somatic and germline variants, copy-number alteration (CNA), and different complex biomarkers. Somatic variant calling was performed in 494 diagnostically relevant cancer genes. In addition, all raw data were re-analyzed with a central bioinformatic pipeline to separate wet- and dry-lab variability. Results The mean positive percentage agreement (PPA) of somatic variant calling was 76% and positive predictive value (PPV) 89% compared a consensus list of variants found by at least five centers. Variant filtering was identified as the main cause for divergent variant calls. Adjusting filter criteria and re-analysis increased the PPA to 88% for all and 97% for clinically relevant variants. CNA calls were concordant for 82% of genomic regions. Calls of homologous recombination deficiency (HRD), tumor mutational burden (TMB), and microsatellite instability (MSI) status were concordant for 94%, 93%, and 93% respectively. Variability of CNAs and complex biomarkers did not increase considerably using the central pipeline and was hence attributed to wet-lab differences. Conclusion Continuous optimization of bioinformatic workflows and participating in round robin tests are recommend.

Many pancreatic neuroendocrine tumors (PanNETs) fall into 2 major prognostic subtypes based on DAXX/ATRX-induced alternative lengthening of telomerase phenotype and alpha- and beta-cell-like epigenomic profiles. However, these PanNETs are still flanked by other PanNETs that do not fit into either subtype. Furthermore, despite advanced genotyping, PanNETs are generally not well-characterized in terms of their histologic and hormonal phenotypes. We aimed to identify new subgroups of PanNETs by extending the currently used transcription factor signatures and investigating their correlation with histologic, hormonal, molecular, and prognostic findings. One hundred eighty-five PanNETs (nonfunctioning 165 and functioning 20), resected between 1996 and 2023, were classified into 5 subgroups (A1, A2, B, C, and D) by cluster analysis based on ARX, PDX1, islet-1 (ISL1), and CDX2 expressions and correlated with trabecular vs solid histology, expression of insulin, glucagon, polypeptide (PP), somatostatin, serotonin, gastrin, calcitonin, adrenocorticotropic hormone (ACTH), DAXX/ATRX, MEN1, and alternative lengthening of telomerase status by fluorescence in situ hybridization, and disease-free survival. A1 (46%, ARX+/ISL1+/PDX1−/CDX2−) and A2 (15%, ARX+/ISL1+/PDX1+/CDX2−) showed trabecular histology and glucagon/PP expression, with A2 also showing gastrin expression. B (18%, PDX1+/ISL1+/ARX−/CDX2−) showed solid histology, insulin, and somatostatin expression (P < .001). It included all insulinomas and had the best outcome (P < .01). C (15%, ARX−/PDX1−/ISL1−/CDX2−) showed solid histology and frequent expression of serotonin, calcitonin, and ACTH. D (5%, PDX1+/CDX2+/ISL1−/ARX−) showed solid histology, expressed ACTH/serotonin, and was an independent poor prognosticator (P < .01). Differential expressions of ARX, PDX1, ISL1, and CDX2 stratified PanNETs into 5 subgroups with different histologies, hormone expressions, and outcomes. Subgroups A1 and A2 resembled the alpha-cell-like type, and subgroup B, the beta-cell-like type. Subgroup C with almost no transcription factor signature was unclear in cell lineage, whereas the PDX+/CDX2+ signature of subgroup D suggested a pancreatic/intestinal cell lineage. Assigning PanNETs to the subgroups may help establish the diagnosis, predict the outcome, and guide the treatment.

Background Pancreatic ductal adenocarcinoma (PDAC) is a ‘difficult-to-treat’ entity. To forecast its prognosis, we introduced a new biomarker, SARIFA (stroma areactive invasion front areas), which are areas at the tumour invasion front lacking desmoplastic stroma reaction upon malignant invasion in the surrounding tissue, leading to direct contact between tumour cells and adipocytes. SARIFA showed its significance in gastric and colorectal carcinoma, revealing lipid metabolism alternations that promote tumour progression. Methods We reviewed the SARIFA status of 166 PDAC cases on all available H&E-stained tumour slides from archival Whipple-resection specimens. SARIFA positivity was defined as SARIFA detection in at least 66% of the available slides. To investigate alterations in tumour metabolism and microenvironment, we performed immunohistochemical staining for FABP4, CD36 and CD68. To verify and quantify a supposed delipidation of adipocytes, adipose tissue was digitally morphometrised. Results In total, 53 cases (32%) were classified as SARIFA positive and 113 (68%) as SARIFA negative. Patients with SARIFA-positive PDAC showed a significantly worse overall survival compared with SARIFA-negative cases (median overall survival: 11.0 months vs. 22.0 months, HR: 1.570 (1.082–2.278), 95% CI, p = 0.018), which was independent from other prognostic markers (p = 0.014). At the invasion front of SARIFA-positive PDAC, we observed significantly higher expression of FABP4 (p < 0.0001) and higher concentrations of CD68+ macrophages (p = 0.031) related to a higher risk of tumour progression. CD36 staining showed no significant expression differences. The adipocyte areas at the invasion front were significantly smaller, with mean values of 4021 ± 1058 µm2 and 1812 ± 1008 µm2 for the SARIFA-negative and -positive cases, respectively (p < 0.001). Conclusions SARIFA is a promising prognostic biomarker for PDAC. Its assessment is characterised by simplicity and low effort. The mechanisms behind SARIFA suggest a tumour-promoting increased lipid metabolism and altered immune background, both showing new therapeutic avenues.

Purpose Providing Patient Access to Precision Oncology (PO) is a major challenge of clinical oncologists. Here, we provide an easily transferable model from strategic management science to assess the outreach of a cancer center. Methods As members of the German WERA alliance, the cancer centers in Würzburg, Erlangen, Regensburg and Augsburg merged care data regarding their geographical impact. Specifically, we examined the provenance of patients from WERA´s molecular tumor boards (MTBs) between 2020 and 2022 (n = 2,243). As second dimension, we added the provenance of patients receiving general cancer care by WERA. Clustering our catchment area along these two dimensions set up a four-quadrant matrix consisting of postal code areas with referrals towards WERA. These areas were re-identified on a map of the Federal State of Bavaria. Results The WERA Matrix overlooked an active screening area of 821 postal code areas – representing about 50% of Bavaria´s spatial expansion and more than six million inhabitants. The WERA Matrix identified regions successfully connected to our outreach structures in terms of subsidiarity – with general cancer care mainly performed locally but PO performed in collaboration with WERA. We also detected postal code areas with a potential PO backlog – characterized by high levels of cancer care performed by WERA and low levels or no MTB representation. Conclusions The WERA Matrix provided a transparent portfolio of postal code areas, which helped assessing the geographical impact of our PO program. We believe that its intuitive principle can easily be transferred to other cancer centers.

Objectives Liquid biopsy (LBx) provides diagnostic, prognostic and predictive insights for malignant diseases and offers promising applications regarding tumor burden, tumor heterogeneity and clonal evolution. Methods ALPS is a prospective trial for patients with metastatic cancer that comprises sequential collection of LBx samples, tumor tissue, radiological imaging data, clinical information and patient-reported outcomes. Peripheral blood plasma is collected based on the individual patient’s staging intervals and LBx-derived ctDNA analyses are performed using CAncer Personalized Profiling sequencing (CAPP-seq). Results From April 2021 to October 2023, 419 patients have been enrolled. A total of 1,293 LBx samples were collected, 419 samples (100 %) at the beginning of the study and an average of 3 (range 1–12) during the 30-month follow-up period of the current interim analysis. 380 tissue biopsy (TBx) samples (90.7 %) were available at baseline and 39.6 % had ≥1 TBx samples at follow-up. Lung cancer patients are most prevalent in ALPS (n=147), followed by colorectal (n=38), prostate (n=31) and gastroesophageal cancer (n=28). On average, 12.0 ng/mL plasma cell-free DNA (cfDNA) could be isolated. First CAPP-seq analyses in 60 patients comprised 110 samples and demonstrated a detection sensitivity of 0.1 %. Conclusions The first interim analysis of ALPS confirms feasibility for comprehensive longitudinal evaluation of LBx and demonstrates suitability for ctDNA evaluation.

Background and aims Endoscopic resection (ER) is accepted as standard treatment for intramucosal esophageal adenocarcinoma (EAC) with well or moderate differentiation. Poor differentiation (PD) is judged as a risk factor for lymph node metastasis (LNM) and surgery is recommended. However, the evidence for this recommendation is weak. Study aim was to analyze the clinical course of patients after ER of EAC with PD. Patients and methods Patients undergoing endoscopic submucosal dissection for EAC were included from 16 German centers. Inclusion criteria were PD in the resection specimen, R0 resection and endoscopic follow-up. Primary outcome was the metastasis rate during follow-up. Analysis was performed retrospectively in a prospectively collected database. Results 25 patients with PD as single risk factor (group A) and 15 patients with PD and additional risk factors (submucosal invasion and/or lymphovascular invasion) were included. The metastasis rate was was 1/25 (4.0%; 95%CI 0.4-17.2) in group A and 3/15 (20.0%; 95%CI 6.0-44.4%) in group B, respectively (p=0.293). The rate of EAC-associated deaths was 1/25 (4%; 95%CI 0.4-17.2%) versus 3/15 (20%; 95%CI 6.0-44.4%) in group B (p=0.293) while the overall death rate was 7/25 (28.0%; 95%CI 13.5-47.3%) versus 3/15 (20%; 95%CI 6.0-44.4%) (p=0.715). Median follow-up was 30 months (IQR 15-53). Conclusions During long-term follow-up the risk of metastasis is low after ER of mucosal EAC with PD as single risk factor. A conservative approach seems justified in this small patient group. However, the treatment strategy has to be determined on an individualized basis until further prospective data are available.

As Neuroendocrine Tumors (NET) often present as metastatic lesions, immunohistochemical assignment to a site of origin is one of the most important tasks in their pathological assessment. Since a fraction of NETs eludes the typical expression profiles of their primary localization, additional sensitive and specific markers are required to improve diagnostic certainty. We investigated the expression of the transcription factor Pituitary Homeobox 2 (PITX2) in a large-scale cohort of 909 NET and 248 Neuroendocrine Carcinomas (NEC) according to the Immunoreactive Score (IRS) and correlated PITX2 expression groups with general tumor groups and localization of the primary. PITX2 expression (all expression groups) was highly sensitive (98.1%) for midgut-derived NET, but not perfectly specific, as non-midgut NET (especially pulmonary/duodenal) were quite frequently weak or moderately positive. The specificity rose to 99.5% for a midgut origin of NET if only a strong PITX2 expression was considered, which was found in only 0.5% (one pancreatic/one pulmonary) of non-midgut NET. In metastases of midgut-derived NET, PITX2 was expressed in all cases (87.5% strong, 12.5% moderate), while CDX2 was negative or only weakly expressed in 31.3% of the metastases. In NEC, a fraction of cases (14%) showed a weak or moderate PITX2 expression, which was not associated with a specific tumor localization. Our study independently validates PITX2 as a very sensitive and specific immunohistochemical marker of midgut-derived NET in a very large collective of Neuroendocrine Neoplasms. Therefore, our data argue towards implementation into diagnostic panels applied for NET as a first line midgut marker.

Background Tumour-associated fat cells without desmoplastic stroma reaction at the invasion front (Stroma AReactive Invasion Front Areas (SARIFA)) is a prognostic biomarker in gastric and colon cancer. The clinical utility of the SARIFA status in oesophagogastric cancer patients treated with perioperative chemotherapy is currently unknown. Methods The SARIFA status was determined in tissue sections from patients recruited into the MAGIC (n = 292) or ST03 (n = 693) trials treated with surgery alone (S, MAGIC) or perioperative chemotherapy (MAGIC, ST03). The relationship between SARIFA status, clinicopathological factors, overall survival (OS) and treatment was analysed. Results The SARIFA status was positive in 42% MAGIC trial S patients, 28% MAGIC and 48% ST03 patients after pre-operative chemotherapy. SARIFA status was related to OS in MAGIC trial S patients and was an independent prognostic biomarker in ST03 trial patients (HR 1.974, 95% CI 1.555–2.507, p < 0.001). ST03 patients with lymph node metastasis (ypN + ) and SARIFA-positive tumours had poorer OS than patients with ypN+ and SARIFA-negative tumours (plogrank < 0.001). Conclusions The SARIFA status has clinical utility as prognostic biomarker in oesophagogastric cancer patients irrespective of treatment modality. Whilst underlying biological mechanisms warrant further investigation, the SARIFA status might be used to identify new drug targets, potentially enabling repurposing of existing drugs targeting lipid metabolism.

Background Inflammatory myofibroblastic tumor (IMTs) are rare mesenchymal neoplasms with slow growth. Resection is considered as therapeutic standard, with chemotherapy being insufficiently effective in advanced disease. ALK translocations are present in 50% of cases, ROS1 fusions (YWHAE::ROS1, TFG::ROS1) are extremely rare. Here, we present a case with TFG::ROS1 fusion and highlight the significance of molecular tumor boards (MTBs) in clinical precision oncology for post-last-line therapy. Case Presentation A 32-year-old woman presented with IMT diagnosed at age 27 for biopsy and treatment evaluation. Previous treatments included multiple resections and systemic therapy with vinblastine, cyclophosphamide, and methotrexate. A computed tomography scan showed extensive tumor infiltration of the psoas muscles and the posterior abdomen. Next generation sequencing revealed an actionable ROS1 fusion (TFG::ROS1) with breakpoints at exon 4/35 including the kinase domain and activating the RAS-pathway. TFG, the Trk-fused gene, exerts functions such as intracellular trafficking and exhibits high sequence homology between species. Based on single reports about efficacy of ROS1-targeting in ROS1 translocation positive IMTs the patient was started on crizotinib, an ATP-competitive small molecule c-MET, ALK and ROS1-inhibitor. With a follow-up of more than 9 months, the patient continues to show a profound response with major tumor regression, improved quality of life and no evidence for severe adverse events. Conclusion This case underscores the importance of the availability of modern molecular diagnostics and interdisciplinarity in precision oncology to identify rare, disease-defining genotypes that make an otherwise difficult-to-treat disease targetable.

Aim Gastric cancer (GC) is a tumor entity with highly variant outcomes. Lymph node metastasis is a prognostically adverse biomarker. We hypothesized that GC primary tissue contains information that is predictive of lymph node status and patient prognosis and that this information can be extracted using Deep Learning (DL). Methods Using three patient cohorts comprising 1146 patients, we trained and validated a DL system to predict lymph node status directly from hematoxylin-and-eosin stained GC tissue sections. We investigated the concordance between the DL-based prediction from the primary tumor slides (aiN score) and the histopathological lymph node status (pN). Furthermore, we assessed the prognostic value of the aiN score alone and when combined with the pN status. Results The aiN score predicted the pN status reaching Area Under the Receiver Operating Characteristic curves (AUROCs) of 0.71 in the training cohort and 0.69 and 0.65 in the two test cohorts. In a multivariate Cox analysis, the aiN score was an independent predictor of patient survival with Hazard Ratios (HR) of 1.5 in the training cohort and of 1.3 and 2.2 in the two test cohorts. A combination of the aiN score and the pN status prognostically stratified patients by survival with p-values <0.05 in log-rank tests. Conclusion GC primary tumor tissue contains additional prognostic information that is accessible using the aiN score. In combination with the pN status, this can be used for personalized management of gastric cancer patients after prospective validation.

An inflammatory fibroid polyp (IFP) of the gastrointestinal tract is a localized, benign mesenchymal lesion consisting of spindle-shaped stromal cells, eosinophilic granulocytes, and some lymphocytes and plasma cells. The discovery of a frequent mutation of the platelet-derived growth factor receptor A (PDGFRA) gene was the first hint of a gene-regulating process in IFPs. The aim of this study was to investigate the interaction of inflammatory processes and the role of mutation and expression of the PDGFRA gene in the development of IFPs for the first time. We used immunohistochemistry to analyze the composition of inflammatory cells and next generation sequencing (NGS) to provide a broad overview of gene mutations. We report on 29 cases of IFP. The mean age, gender differences, and localization were compatible with the literature. Spindle cell histomorphology was present in 79% of cases showing a typical onion skin-like perivascular arrangement and significantly high CD34 positivity (p = 0.002, Fisher’s exact test). Eosinophilic granulocytes were present in an average density of 60 ± 49/high power field (HPF) (range: 15–200), and there was a significantly higher rate of IFPs larger than 2 cm in size (p = 0.018, Wilcoxon test). All but one cases could be analyzed by NGS. Mutations were observed in 17 cases (60.7%), including 13 (46.4%) mutations in the PDGFRA gene. Among the gastric lesions, mutations were found in exon 18 of the PDGFRA gene with amino acid exchange (Asp842Val) for eight out of 10 cases and in exon 12 in two cases. All three cases in the small intestine revealed mutation of the PDGFRA gene in exon 12. We found no PDGFRA mutation in our colonic cases. PDGFRA expression was significantly correlated with mutations of the same gene (p = 0.005, Fisher’s exact test) and especially with mutations in exon 12 of the same gene (p < 0.001, Fisher’s exact test). Interestingly, three of our cases (10.3%) without mutation or expression of the PDGFRA gene revealed an unusually high concentration of IgG-positive plasma cells (average: 140 ± 26/HPF, range: 110–160) and IgG4-positive plasma cells (average: 87 ± 21/HPF, range: 60–100). For comparison, an IgG4/IgG ratio of more than 0.4 is commonly observed in IgG4-related diseases. Our molecular results were in accordance with 113 genetically analyzed cases published to date. There was a correlation between the IFP site and mutation variants of the PDGFRA gene. IFPs were localized in the stomach in 49.1% of cases, in the small intestine in 47.3%, and in the colon in 3.6%. Exon 12 of the PDGFRA gene was mutated in 41.1% of cases and primarily occurred in the small intestine (82.6%). Exon 18 was mutated in 22.3% of cases and primarily occurred in the stomach (80.0%). The mutated codon interval 566–571 in exon 12 and codon 842 in exon 18 were compatible, as observed in a gastrointestinal stromal tumor. Conclusively, the correlation between mutation and expression of the PDGFRA gene points to different pathways in IFPs. Additionally, our data hint at a morphological but not genetic overlap between IFPs and IgG4-related pseudotumors.

Background The currently available immunotherapies already changed the strategy how many cancers are treated from first to last line. Understanding even the most complex heterogeneity in tumor tissue and mapping the spatial cartography of the tumor immunity allows the best and optimized selection of immune modulating agents to (re-)activate the patient's immune system and direct it against the individual cancer in the most effective way. Recent Findings Primary cancer and metastases maintain a high degree of plasticity to escape any immune surveillance and continue to evolve depending on many intrinsic and extrinsic factors In the field of immune-oncology (IO) immune modulating agents are recognized as practice changing therapeutic modalities. Recent studies have shown that an optimal and lasting efficacy of IO therapeutics depends on the understanding of the spatial communication network and functional context of immune and cancer cells within the tumor microenvironment. Artificial intelligence (AI) provides an insight into the immune-cancer-network through the visualization of very complex tumor and immune interactions in cancer tissue specimens and allows the computer-assisted development and clinical validation of such digital biomarker. Conclusions The successful implementation of AI-supported digital biomarker solutions guides the clinical selection of effective immune therapeutics based on the retrieval and visualization of spatial and contextual information from cancer tissue images and standardized data. As such, computational pathology (CP) turns into “precision pathology” delivering individual therapy response prediction. Precision Pathology does not only include digital and computational solutions but also high levels of standardized processes in the routine histopathology workflow and the use of mathematical tools to support clinical and diagnostic decisions as the basic principle of a “precision oncology”.

Background and aims Celiac disease with its endoscopic manifestation of villous atrophy is underdiagnosed worldwide. The application of artificial intelligence (AI) for the macroscopic detection of villous atrophy at routine esophagogastroduodenoscopy may improve diagnostic performance. Methods A dataset of 858 endoscopic images of 182 patients with villous atrophy and 846 images from 323 patients with normal duodenal mucosa was collected and used to train a ResNet 18 deep learning model to detect villous atrophy. An external data set was used to test the algorithm, in addition to six fellows and four board certified gastroenterologists. Fellows could consult the AI algorithm’s result during the test. From their consultation distribution, a stratification of test images into “easy” and “difficult” was performed and used for classified performance measurement. Results External validation of the AI algorithm yielded values of 90 %, 76 %, and 84 % for sensitivity, specificity, and accuracy, respectively. Fellows scored values of 63 %, 72 % and 67 %, while the corresponding values in experts were 72 %, 69 % and 71 %, respectively. AI consultation significantly improved all trainee performance statistics. While fellows and experts showed significantly lower performance for “difficult” images, the performance of the AI algorithm was stable. Conclusion In this study, an AI algorithm outperformed endoscopy fellows and experts in the detection of villous atrophy on endoscopic still images. AI decision support significantly improved the performance of non-expert endoscopists. The stable performance on “difficult” images suggests a further positive add-on effect in challenging cases.

Immunohistochemical analysis of mismatch repair (MMR) protein expression is widely used to identify tumors with a deficient MMR (dMMR). MMR proteins (MLH1/PMS2 and MSH2/MSH6) work as functional heterodimers, which usually leads to the loss of expression in only one functional MMR heterodimer. Recently, there have been studies showing the simultaneous loss of immunoexpression in proteins of both heterodimers. Yet, this phenomenon has been rarely investigated. In this study, we retrospectively considered cases of different digestive system cancers (gastric cancer, ampullary cancer, small bowel cancer, colorectal cancer), which were immunohistochemically tested for dMMR within a 4-year period at our university hospital (n=352). Of the 103 cases showing dMMR, 5 cases (1.4% of all, 5.1% of dMMR cases) showed a concurrent loss of MLH1, PMS2 and MSH6 immunoexpression, whereas in the other 98 dMMR cases only one MMR heterodimer was affected. MLH1-/PMS2-/MSH6- cancer cases almost arose throughout the entire digestive tract: from the gastric antrum to the left colic flexur. To provide a comprehensive molecular characterization of this MLH1-/PMS2-/MSH6- immunophenotype, tumors were analyzed for microsatellite instability, MLH1 promotor hypermethylation and BRAF exon 15 status. Furthermore, we performed next-generation sequencing focusing on genes related to DNA repair. Here, we could detect pathogenic germline variants as well as multiple sporadic mutations in different genes involved in MMR and homologous recombination repair (HRR) respectively. The affected MMR/HRR-related genes were: ATM, BARD1, BRCA1, CDK12, CHEK1, CHEK2, FANCA, MLH1, MSH6, PALB2, TP53. Considering the biologic function of HRR/MMR proteins as potential drug targets and the low frequency of most of these mutations in digestive system cancers in general, their common occurrence in our MLH1-/PMS2-/MSH6- cases seems to be even more noteworthy, highlighting the need for recognition, awareness and further investigation of this unusual IHC staining pattern.

Tumor-infiltrating lymphocytes (TILs) correlate with the number and size of the surrounding lymph nodes in patients with colorectal carcinoma (CRC) and reflect the quality of the antitumor immune response. In this prospective study, we analyzed whether this response correlated with the circulating lymphocytes in peripheral blood (PB). In 47 patients with newly diagnosed CRC, flow cytometry was performed to analyze the B cells, T cells, NK cells, and a variety of their subsets in PB. The results were correlated with TILs in the resected tumor and with the number and size of the surrounding lymph nodes in nodal negative (N- patients (LN5: number of lymph nodes measuring ≥5 mm) and the metastasis-to-lymph node size ratio (MSR) in nodal positive patients (N+). Differences between the number of TILs could be seen between N+ and N- patients, dependent on the LN5 and MSR categories, with higher values in N- cases and in patients with a higher LN5 category or a lower MSR. Additionally, higher values of various circulating lymphocyte subgroups were observed in these patients. For the total PB lymphocytes, CD8 cells, and some of their subgroups, a positive correlation with the TILs was found. This study shows that circulating lymphocytes—in particular, cytotoxic T cells—correlate with the local antitumor immune response displayed by TILs and lymph node activation. Our findings indicate that local and generalized antitumor immune responses are concordant with their different components.