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BACKGROUND/AIM: PD-L1 inhibitors have been approved for cisplatin-ineligible urothelial cancer patients relapsing after radical cystectomy. A prerequisite for therapy is a positive PD-L1 expression in the tumor tissue, whereas no options are available for patients with negative PD-L1 status. However, studies revealed that many PD-L1-negative patients also responded to PD-L1 therapy. This study investigated the feasibility of PD-L1 mRNA complementary RNA in situ hybridization (RNAish) analysis to detect PD-L1-responders independent of PD-L1 protein status. MATERIALS AND METHODS: Immunohistochemistry and RNA in situ hybridization were used to assess PD-L1 protein and mRNA in radical cystectomy tissue from patients with advanced and metastasized urothelial cancer. RESULTS: In this study, PD-L1 protein and mRNA were detected in ≥90% of the examined tissue. Positive PD-L1 mRNA expression (≥1%) on TC and IC could be evaluated in 77% and 31% of the cases, respectively. Moreover, scatterplot analysis revealed a PD-L1 mRNA positive and PD-L1 protein negative subpopulation. According to the CPS score, positive PD-L1 protein expression could be evaluated in 88% and positive PD-L1 mRNA expression in 71% of the cases. Scatterplot analysis of the CPS scores revealed a CPS protein negative but CPS mRNA positive small subpopulation. CONCLUSION: The feasibility of RNAish on formalin-fixed tissue could be proven. Moreover, complementary PD-L1 RNAish identified a sub-population of PD-L1 protein-negative and PD-L1 mRNA-positive patients, which may benefit from PD-L1 therapy.
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Carcinoma de Células de Transição , Neoplasias da Bexiga Urinária , Humanos , Antígeno B7-H1/metabolismo , RNA Mensageiro/genética , Recidiva Local de Neoplasia , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/terapia , Neoplasias da Bexiga Urinária/metabolismo , Carcinoma de Células de Transição/patologia , Biomarcadores Tumorais/metabolismoRESUMO
Malignant transformation of oral lichen planus (OLP) into oral squamous cell carcinoma is considered as one of the most serious complications of OLP. For the early detection of oral cancer in OLP follow-up, accurate localization of the OLP center is still difficult but often required for confirmatory biopsy with histopathological examination. Optical coherence tomography (OCT) offers the potential for more reliable biopsy sampling in the oral cavity as it is capable of non-invasively imaging the degenerated oral layer structure. In this case-series study with 15 patients, features of clinically classified forms of OLP in OCT cross-sections were registered and correlated with available histologic sections. Besides patients with reticular, atrophic, erosive and plaque-like OLP, two patients with leukoplakia were included for differentiation. The results show that OCT yields information about the epithelial surface, thickness and reflectivity, as well as the identifiability of the basement membrane and the vessel network, which could be used to complement the visual clinical appearance of OLP variants and allow a more accurate localization of the OLP center. This forms the basis for further studies on OCT-assisted non-invasive clinical classification of OLP, with the aim of enabling decision support for biopsy sampling in the future.
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Background: Within the bone marrow (BM), mature T cells are maintained under homeostatic conditions to facilitate proper hematopoietic development. This homeostasis depends upon a peculiar elevated frequency of regulatory T cells (Tregs) and immune regulatory activities from BM-mesenchymal stem cells (BM-MSCs). In response to BM transplantation (BMT), the conditioning regimen exposes the BM to a dramatic induction of inflammatory cytokines and causes an unbalanced T-effector (Teff) and Treg ratio. This imbalance negatively impacts hematopoiesis, particularly in regard to B-cell lymphopoiesis that requires an intact cross-talk between BM-MSCs and Tregs. The mechanisms underlying the ability of BM-MSCs to restore Treg homeostasis and proper B-cell development are currently unknown. Methods: We studied the role of host radio-resistant cell-derived CD40 in restoring Teff/Treg homeostasis and proper B-cell development in a murine model of BMT. We characterized the host cellular source of CD40 and performed radiation chimera analyses by transplanting WT or Cd40-KO with WT BM in the presence of T-reg and co-infusing WT or - Cd40-KO BM-MSCs. Residual host and donor T cell expansion and activation (cytokine production) and also the expression of Treg fitness markers and conversion to Th17 were analyzed. The presence of Cd40+ BM-MSCs was analyzed in a human setting in correlation with the frequency of B-cell precursors in patients who underwent HSCT and variably developed acute graft-versus-host (aGVDH) disease. Results: CD40 expression is nearly undetectable in the BM, yet a Cd40-KO recipient of WT donor chimera exhibited impaired B-cell lymphopoiesis and Treg development. Lethal irradiation promotes CD40 and OX40L expression in radio-resistant BM-MSCs through the induction of pro-inflammatory cytokines. OX40L favors Teff expansion and activation at the expense of Tregs; however, the expression of CD40 dampens OX40L expression and restores Treg homeostasis, thus facilitating proper B-cell development. Indeed, in contrast to dendritic cells in secondary lymphoid organs that require CD40 triggers to express OX40L, BM-MSCs require CD40 to inhibit OX40L expression. Conclusions: CD40+ BM-MSCs are immune regulatory elements within BM. Loss of CD40 results in uncontrolled T cell activation due to a reduced number of Tregs, and B-cell development is consequently impaired. GVHD provides an example of how a loss of CD40+ BM-MSCs and a reduction in B-cell precursors may occur in a human setting.
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Transplante de Medula Óssea , Medula Óssea/imunologia , Antígenos CD40/genética , Regulação da Expressão Gênica/imunologia , Homeostase/imunologia , Células-Tronco Mesenquimais/imunologia , Ligante OX40/genética , Estresse Fisiológico/imunologia , Adulto , Idoso , Animais , Medula Óssea/fisiologia , Células da Medula Óssea/imunologia , Antígenos CD40/imunologia , Feminino , Homeostase/genética , Humanos , Ativação Linfocitária/imunologia , Masculino , Transplante de Células-Tronco Mesenquimais , Camundongos , Pessoa de Meia-Idade , Ligante OX40/imunologia , Linfócitos T Reguladores/imunologia , Condicionamento Pré-Transplante , Adulto JovemRESUMO
Purpose: Most high-grade serous ovarian cancer (HGSOC) patients develop recurrent disease after first-line treatment, frequently with fatal outcome. This work aims at studying the molecular biology of both primary and recurrent HGSOC.Experimental Design: Gene expression profiles of matched primary and recurrent fresh-frozen tumor tissues from 66 HGSOC patients were obtained by RNA sequencing. Clustering analyses and pairwise comparison of the profiles between matched samples and subsequent functional alignment were used for the identification of molecular characteristics of HGSOC.Results: Both primary and recurrent HGSOC samples presented predominant gene expression differences in their microenvironment, determined by a panel of genes covering all major pathways of immune activation together with a number of genes involved in the remodeling of extracellular matrix and adipose tissues. Stratifying tumor tissues into immune active and silent groups, we further discovered that although some recurrent tumors shared the same immune status as their primary counterparts, others switched the immune status, either from silent to active or active to silent. Interestingly, genes belonging to the B7-CD28 immune checkpoint family, known for their major role as negative regulators of the immune response, were overexpressed in the immune active tumors. Searching for potential tumor antigens, CEACAM21, a member of the carcinoembryonic antigen family, was found to be significantly overexpressed in immune active tissues in comparison with the silent ones.Conclusions: The results illustrate the complexity of the tumor microenvironment in HGSOC and reveal the molecular relationship between primary and recurrent tumors, which have multiple therapeutic implications. Clin Cancer Res; 23(24); 7621-32. ©2017 AACR.
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Antígenos de Neoplasias/genética , Cistadenocarcinoma Seroso/genética , Neoplasias Ovarianas/genética , Microambiente Tumoral/genética , Adulto , Idoso , Antígenos de Neoplasias/imunologia , Linhagem Celular Tumoral , Cistadenocarcinoma Seroso/imunologia , Cistadenocarcinoma Seroso/patologia , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Pessoa de Meia-Idade , Gradação de Tumores , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/patologia , Neoplasias Ovarianas/imunologia , Neoplasias Ovarianas/patologia , Análise de Sequência de RNA , Microambiente Tumoral/imunologiaRESUMO
Patient-derived xenograft (PDX) models have shown to reflect original patient tumors better than any other preclinical model. We embarked in a study establishing a large panel of head and neck squamous cell carcinomas PDX for biomarker analysis and evaluation of established and novel compounds. Out of 115 transplanted specimens 52 models were established of which 29 were characterized for response to docetaxel, cetuximab, methotrexate, carboplatin, 5-fluorouracil and everolimus. Further, tumors were subjected to sequencing analysis and gene expression profiling of selected mTOR pathway members. Most frequent response was observed for docetaxel and cetuximab. Responses to carboplatin, 5-fluorouracil and methotrexate were moderate. Everolimus revealed activity in the majority of PDX. Mutational profiling and gene expression analysis did not reveal a predictive biomarker for everolimus even though by trend RPS6KB1 mRNA expression was associated with response. In conclusion we demonstrate a comprehensively characterized panel of head and neck cancer PDX models, which represent a valuable and renewable tissue resource for evaluation of novel compounds and associated biomarkers.
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Carcinoma de Células Escamosas/tratamento farmacológico , Neoplasias de Cabeça e Pescoço/tratamento farmacológico , Sirolimo/análogos & derivados , Serina-Treonina Quinases TOR/antagonistas & inibidores , Ensaios Antitumorais Modelo de Xenoenxerto/métodos , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Antineoplásicos/uso terapêutico , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/virologia , Análise Mutacional de DNA , Everolimo , Feminino , Perfilação da Expressão Gênica , Neoplasias de Cabeça e Pescoço/genética , Neoplasias de Cabeça e Pescoço/virologia , Papillomavirus Humano 16/fisiologia , Humanos , Subunidade gama Comum de Receptores de Interleucina/deficiência , Subunidade gama Comum de Receptores de Interleucina/genética , Masculino , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Pessoa de Meia-Idade , Infecções por Papillomavirus/virologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Sirolimo/uso terapêutico , Serina-Treonina Quinases TOR/genética , Resultado do TratamentoRESUMO
AIM: The sites of haematopoiesis during human ontogeny can be correlated to the sites where haematopoiesis occurs in vertebrate phylogeny. As haematopoiesis has been described in the diencephalon and pituitary gland of water-inhabiting vertebrates we wanted to find out whether such a phenomenon also occurs in human embryos. MATERIAL AND METHODS: Paraffin-embedded specimens from the diencephalon and pituitary gland of human embryos at the 7th to 22nd gestational week and from adults were investigated by conventional histology and immunohistology for the presence of haematopoietic cells. RESULTS: Cellular accumulations predominantly of erythroid and megakaryocytic lineage were identified in the floor of the developing diencephalon of the 7th/8th gestational week. At the older developmental stages of the 18th to 22nd gestational week loose aggregates of haematopoietic cells within the leptomeningeal spaces adjacent to the hypophyseal infundibulum were detected in 2 out of 7 cases analyzed. CONCLUSIONS: As it has been proposed that lymphohaematopoietic clusters occasionally occur within the brain in bone marrow-less vertebrates as a response to noxious agents, we speculate that this temporal appearance of haematopoietic cell clusters within the diencephalon floor in early human ontogeny could also be due to fetal immunomodulations.
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Diencéfalo/embriologia , Hematopoese/fisiologia , Filogenia , Vertebrados/embriologia , Animais , Diferenciação Celular/fisiologia , Sobrevivência Celular/fisiologia , Diencéfalo/citologia , Diencéfalo/fisiologia , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/fisiologia , Sistema Hematopoético/citologia , Sistema Hematopoético/embriologia , Sistema Hematopoético/fisiologia , Humanos , Fatores Imunológicos/fisiologia , Neuro-Hipófise/citologia , Neuro-Hipófise/fisiologia , Tubarões/embriologia , Tubarões/fisiologia , Vertebrados/fisiologiaRESUMO
Monocytoid cells are a subset of B lymphocytes with characteristic morphology and immunophenotype, which proliferate in a broad variety of reactive lymph node conditions. Up to now, no direct infection of these cells by a pathogenic organism has been identified. We found in a series of lymph node specimens from 13 patients having in common a prominent monocytoid B (MCB)-cell reaction in the absence of epithelioid cells or necrosis evidence that Epstein-Barr virus (EBV) can infect MCB cells. Clinical and serologic findings indicative for an acute EBV infection were reported only in 2 cases. By in situ hybridization and immunohistology, a number of MCB cells in all cases was found to contain EBV-encoded small nuclear RNAs and to express the EBV-encoded nuclear antigen (EBNA)-2 and, in lesser extent, the latent membrane protein-1. EBV-transcripts and proteins were also detectable in a number of extrafollicular B blasts and germinal center B cells in 10 of 13 cases. Our findings imply that an EBV infection of MCB cells associated with predominant EBNA-2 expression represents an early sign of primary infection and that it should be included in the differential diagnosis of activated lymph nodes with prominent MCB-cell reaction in the absence of epithelioid cells.