14 research outputs found

    Integrative Analysis of Pleomorphic Dermal Sarcomas Reveals Fibroblastic Differentiation and Susceptibility to Immunotherapy

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    Purpose: Pleomorphic dermal sarcoma (PDS) is a rare malignant cutaneous tumor with an unknown cell of origin. Locally defined tumors can be treated by curative excisions, whereas advanced stages of the disease are difficult to treat, using standard regimens. Experimental Design: We performed whole-exome sequencing on a cohort of 28 individuals and corresponding transcriptomic analysis on 21 patients, as well as quantitative IHC image analysis on 27 patients. Results: PDS exhibits a universally high mutational load (42.7 mutations/mega base) with an inflamed, immunogenic tumor microenvironment. Three cases of PDS showed response to immune checkpoint blockade. Local mutation rate variation together with mRNA expression data demonstrate that PDS form a distinct entity, with PDGFRB as a lineage marker. In addition, we found that PDS is of mesenchymal, fibroblastic differentiation. Conclusions: PDS is of fibroblastic differentiation and exhibits a strong susceptibility to immunotherapy, including a high mutational burden and an inflamed tumor microenvironment

    Sex-specific prognostic effect of CD66b-positive tumor-infiltrating neutrophils (TANs) in gastric and esophageal adenocarcinoma

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    Background Tumor-associated neutrophils (TANs) have recently been identified as a relevant component of the tumor microenvironment (TME) in solid tumors. Within the TME TANs mediate either tumor-promoting or tumor-inhibiting activities. So far, their prognostic relevance remains to be determined. This study aims to evaluate the prognostic relevance of TANs in different molecular subtypes of gastric and esophageal adenocarcinoma. Methods We analyzed a total of 1118 Caucasian patients divided into gastric adenocarcinoma (n = 458) and esophageal adenocarcinoma cohort (n = 660) of primarily resected and neoadjuvant-treated individuals. The amount of CD66b + TANs in the tumor stroma was determined using quantitative image analysis and correlated to both molecular, as well as clinical data. Results An accumulation of TANs in the tumor stroma of gastric carcinomas was associated to a significant favorable prognosis (p = 0.026). A subgroup analysis showed that this effect was primarily related to the molecular chromosomal instable subtype (CIN) of gastric carcinomas (p = 0.010). This was only observed in female patients (p = 0.014) but not in male patients (p = 0.315). The same sex-specific effect could be confirmed in adenocarcinomas of the esophagus (p = 0.027), as well as in female individuals after receiving neoadjuvant therapy (p = 0.034). Conclusions Together, we show a sex-specific prognostic effect of TANs in gastric cancer within a Caucasian cohort. For the first time, we showed that this sex-specific prognostic effect of TANs can also be seen in esophageal cancer

    Deep Learning Predicts HPV Association in Oropharyngeal Squamous Cell Carcinomas and Identifies Patients with a Favorable Prognosis Using Regular H&E Stains

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    Purpose: Human papillomavirus (HPV) in oropharyngeal squamous cell carcinoma (OPSCC) is tumorigenic and has been associated with a favorable prognosis compared with OPSCC caused by tobacco, alcohol, and other carcinogens. Meanwhile, machine learning has evolved as a powerful tool to predict molecular and cellular alterations of medical images of various sources. Experimental Design: We generated a deep learning-based HPV prediction score (HPV-ps) on regular hematoxylin and eosin (H&E) stains and assessed its performance to predict HPV association using 273 patients from two different sites (OPSCC; Giessen, n = 163; Cologne, n = 110). Then, the prognostic relevance in a total of 594 patients (Giessen, Cologne, HNSCC TCGA) was evaluated. In addition, we investigated whether four board-certified pathologists could identify HPV association (n = 152) and compared the results to the classifier. Results: Although pathologists were able to diagnose HPV association from H&E-stained slides (AUC = 0.74, median of four observers), the interrater reliability was minimal (Light Kappa = 0.37; P = 0.129), as compared with AUC = 0.8 using the HPV-ps within two independent cohorts (n = 273). The HPV-ps identified individuals with a favorable prognosis in a total of 594 patients from three cohorts (Giessen, OPSCC, HR = 0.55, P < 0.0001; Cologne, OPSCC, HR = 0.44, P = 0.0027; TCGA, non-OPSCC head and neck, HR = 0.69, P = 0.0073). Interestingly, the HPV-ps further stratified patients when combined with p16 status (Giessen, HR = 0.06, P < 0.0001; Cologne, HR = 0.3, P = 0.046). Conclusions: Detection of HPV association in OPSCC using deep learning with help of regular H&E stains may either be used as a single biomarker, or in combination with p16 status, to identify patients with OPSCC with a favorable prognosis, potentially outperforming combined HPV-DNA/p16 status as a biomarker for patient stratification

    Predicting HPV association using deep learning and regular H&E stains allows granular stratification of oropharyngeal cancer patients

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    Abstract Human Papilloma Virus (HPV)-associated oropharyngeal squamous cell cancer (OPSCC) represents an OPSCC subgroup with an overall good prognosis with a rising incidence in Western countries. Multiple lines of evidence suggest that HPV-associated tumors are not a homogeneous tumor entity, underlining the need for accurate prognostic biomarkers. In this retrospective, multi-institutional study involving 906 patients from four centers and one database, we developed a deep learning algorithm (OPSCCnet), to analyze standard H&E stains for the calculation of a patient-level score associated with prognosis, comparing it to combined HPV-DNA and p16-status. When comparing OPSCCnet to HPV-status, the algorithm showed a good overall performance with a mean area under the receiver operator curve (AUROC) = 0.83 (95% CI = 0.77-0.9) for the test cohort (n = 639), which could be increased to AUROC = 0.88 by filtering cases using a fixed threshold on the variance of the probability of the HPV-positive class - a potential surrogate marker of HPV-heterogeneity. OPSCCnet could be used as a screening tool, outperforming gold standard HPV testing (OPSCCnet: five-year survival rate: 96% [95% CI = 90–100%]; HPV testing: five-year survival rate: 80% [95% CI = 71–90%]). This could be confirmed using a multivariate analysis of a three-tier threshold (OPSCCnet: high HR = 0.15 [95% CI = 0.05–0.44], intermediate HR = 0.58 [95% CI = 0.34–0.98] p = 0.043, Cox proportional hazards model, n = 211; HPV testing: HR = 0.29 [95% CI = 0.15–0.54] p < 0.001, Cox proportional hazards model, n = 211). Collectively, our findings indicate that by analyzing standard gigapixel hematoxylin and eosin (H&E) histological whole-slide images, OPSCCnet demonstrated superior performance over p16/HPV-DNA testing in various clinical scenarios, particularly in accurately stratifying these patients

    Influenza virus infected EpiSPC are impaired in their regenerative response due to restricted Fgfr2b expression.

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    <p>(A) Wt mice were infected with 500pfu of the indicated influenza virus strains, or mock-infected, and lung sections were stained with hematoxylin-eosin at d21 pi (arrows indicate areas of non-epithelialized tissue). (B) Infected and non-infected epithelial cell subsets of PR/8 infected wt mice were quantified by flow cytometry for their proliferative response. (C) Quantification of Fgfr2b expression in infected (NP<sup>+</sup>) and non-infected (NP<sup>-</sup>) EpiSPC by flow cytometry at d4 pi. (D) Flow-sorted EpiSPC were <i>ex vivo</i> infected with the indicated MOI of PR/8 and seeded in matrix for 3D cultures. At d6 of culture, the number of formed organoids was quantified. (E) Infected (hemagglutinin<sup>+</sup>; HA<sup>+</sup>) or non-infected (hemagglutinin<sup>neg</sup>; HA<sup>-</sup>) EpiSPC or control HA<sup>-</sup> SAEC were flow-sorted from the lungs of PR/8-infected tdtomato<sup>+</sup> mice at d4 pi for intrapulmonary transplantation into 7d PR/8-infected wt mice. Lung sections were obtained at d7 and d14 after transplantation. Representative micrographs show overlays of brightfield and red staining of tdtomato<sup>+</sup> transplanted cells. Overlay of tdtomato<sup>+</sup> transplanted cells (red) and the type I AEC cell marker T1α (green) is shown in the right panels (arrows indicate co-expression of T1α and tdtomato); bars = 100μm. Bar graphs represent means ± SD of n = 3–4 independent experiments; * <i>p</i><0.05; **<i>p</i><0.01; ***<i>p</i><0.001; HA, hemagglutinin; Tx, transplantaion.</p

    EpiSPC are resistant to apoptosis and show a high proliferative response after PR/8 infection which is mediated by Fgf10/Fgfr2b signaling.

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    <p>(A) Proliferation rates of the given epithelial cell subsets was analysed in PR/8 infected wt mice by FACS quantification of Ki67<sup>+</sup> cells at the indicated time points pi. (B) Apoptosis of each EpCam<sup>+</sup> subset was quantified by FACS (Annexin V<sup>+</sup> proportions) at d7 post PR/8 infection and of non-infected wt mice. (C) Expression of Fgfr2b on EpiSPC at the given time points post PR/8 or mock infection was quantified by FACS and is given as MFI (median fluorescence intensity) of Fgfr2b ab minus MFI of matched isotype control. The proliferative response of the EpCam<sup>+</sup> cell subsets was quantified by FACS at d7 pi in <i>Rosa26</i><sup><i>rtTA/+</i></sup><i>;tet(O)sFgfr2b/+</i> (D) <i>Rosa26</i><sup><i>rtTA/+</i></sup><i>;tet(O)Fgf10/+</i> mice (E) and <i>Fgf7</i><sup><i>-/-</i></sup> mice (F) compared to non-dox-induced or wt littermates. Bar graphs represent means ± SD of n = 4–6 independent experiments; * <i>p</i><0.05; **<i>p</i><0.01; +dox, doxycycline food; -dox, normal diet.</p

    β-catenin dependent transcription mediates upregulation of <i>Fgfr2b</i> expression, which is inhibited in PR/8-infected, but not in non-infected lung epithelial cells.

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    <p>(A) EpCam<sup>+</sup> lung epithelial cells derived from <i>Rosa26</i><sup><i>ERTCre/ERTCre</i></sup><i>;Ctnnb1</i><sup><i>flox/flox</i></sup> mice were grown to confluency and treated with tamoxifen or DMSO control prior to infection with PR/8 (MOI = 0.1; 24h). mRNA expression of β-catenin (<i>Ctnnb1</i>) (left) or of <i>Fgfr2b</i> (right) was quantified and normalized to values of DMSO-treated control. (B) Wt distal lung epithelial cells in confluent culture were PR/8-infected (MOI 0.1) and treated with an activator (LiCl) or inhibitor (XAV939) of β-catenin signaling. Expression of the viral M segment was quantified at 16 h pi and normalized to LiCl-treated cultures (left). The right plot shows representative photomicrographs of these cultures stained for IV nucleoprotein (NP) after 6 h of PR/8 infection. (C) Wt mice were infected with PR/8 for 7d and infected (IV hemagglutinin<sup>+</sup>, HA<sup>+</sup>) vs. non-infected (HA-) EpCam<sup>+</sup> cells were flow-sorted. Expression of the β-catenin-dependent transcripts <i>Axin2</i>, <i>Fgfr2b</i>, and <i>Ccnd1</i> was quantified in HA- cells and normalized to values from HA<sup>+</sup> cells. All bar graphs represent means ± SD of n = 3–4 independent experiments; * <i>p</i><0.05; **<i>p</i><0.01; Tam, tamoxifen.</p
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