Chantal Pauli

Aβ-Induced Inflammatory Processes in Microglia Cells of APP23 Transgenic Mice

A microglial response is part of the inflammatory processes in Alzheimers disease (AD). We have used APP23 transgenic mice overexpressing human amyloid precursor protein with the Swedish mutation to characterize this microglia response to amyloid deposits in aged mice. Analyses with MAC-1 and F4/80 antibodies as well as in vivo labeling with bromodeoxyuridine demonstrate that microglia in the plaque vicinity are in an activated state and that proliferation contributes to their accumulation at the plaque periphery. The amyloid-induced microglia activation may be mediated by scavenger receptor A, which is generally elevated, whereas the increased immunostaining of the receptor for advanced glycation end products is more restricted. Although components of the phagocytic machinery such as macrosialin and Fc receptors are increased in activated microglia, efficient clearance of amyloid is missing seemingly because of the lack of amyloid-bound autoantibodies. Similarly, although up-regulation of major histocompatibility complex class II (IA) points toward an intact antigen-presenting function of microglia, lack of T and B lymphocytes does not indicate a cell-mediated immune response in the brains of APP23 mice. The similar characteristics of microglia in the APP23 mice and in AD render the mouse model suitable to study the role of inflammatory processes during AD pathogenesis.

A patient with BRAF V600E lung adenocarcinoma responding to vemurafenib.

Journal of Thoracic Oncology • Volume 7, Number 10, October 2012 Vemurafenib is active against metastatic melanoma with BRAF V600E mutation. BRAF mutations were reported in 3% to 5% of non–small-cell lung cancers (NSCLC) by the US Lung Cancer Mutation Consortium and by Italian investigators. In vitro, MEK inhibition in lung cancer cell lines with V600E induced apoptosis, but clinical experience with drugs targeting the RAS/RAF/MEK pathway in V600E-NSCLC is limited.

Whole-Exome Sequencing of Metastatic Cancer and Biomarkers of Treatment Response

IMPORTANCE Understanding molecular mechanisms of response and resistance to anticancer therapies requires prospective patient follow-up and clinical and functional validation of both common and low-frequency mutations. We describe a whole-exome sequencing (WES) precision medicine trial focused on patients with advanced cancer. OBJECTIVE To understand how WES data affect therapeutic decision making in patients with advanced cancer and to identify novel biomarkers of response. DESIGN, SETTING, AND PATIENTS Patients with metastatic and treatment-resistant cancer were prospectively enrolled at a single academic center for paired metastatic tumor and normal tissue WES during a 19-month period (February 2013 through September 2014). A comprehensive computational pipeline was used to detect point mutations, indels, and copy number alterations. Mutations were categorized as category 1, 2, or 3 on the basis of actionability; clinical reports were generated and discussed in precision tumor board. Patients were observed for 7 to 25 months for correlation of molecular information with clinical response. MAIN OUTCOMES AND MEASURES Feasibility, use of WES for decision making, and identification of novel biomarkers. RESULTS A total of 154 tumor-normal pairs from 97 patients with a range of metastatic cancers were sequenced, with a mean coverage of 95X and 16 somatic alterations detected per patient. In total, 16 mutations were category 1 (targeted therapy available), 98 were category 2 (biologically relevant), and 1474 were category 3 (unknown significance). Overall, WES provided informative results in 91 cases (94%), including alterations for which there is an approved drug, there are therapies in clinical or preclinical development, or they are considered drivers and potentially actionable (category 1-2); however, treatment was guided in only 5 patients (5%) on the basis of these recommendations because of access to clinical trials and/or off-label use of drugs. Among unexpected findings, a patient with prostate cancer with exceptional response to treatment was identified who harbored a somatic hemizygous deletion of the DNA repair gene FANCA and putative partial loss of function of the second allele through germline missense variant. Follow-up experiments established that loss of FANCA function was associated with platinum hypersensitivity both in vitro and in patient-derived xenografts, thus providing biologic rationale and functional evidence for his extreme clinical response. CONCLUSIONS AND RELEVANCE The majority of advanced, treatment-resistant tumors across tumor types harbor biologically informative alterations. The establishment of a clinical trial for WES of metastatic tumors with prospective follow-up of patients can help identify candidate predictive biomarkers of response.

Anterior cruciate ligament changes in the human knee joint in aging and osteoarthritis.

OBJECTIVE The development and patterns of spontaneous age-related changes in the anterior cruciate ligament (ACL) and their relationship to articular cartilage degeneration are not well characterized. This study was undertaken to investigate the types and temporal sequence of age-related ACL changes and to determine their correlation with cartilage lesion patterns at all stages of osteoarthritis (OA) development in human knee joints without prior joint trauma. METHODS Human knee joints (n = 120 from 65 donors ages 23-92) were obtained at autopsy, and ACLs and cartilage were graded macroscopically and histologically. Inflammation surrounding the ACL was assessed separately. RESULTS Histologic ACL substance scores and ligament sheath inflammation scores increased with age. Collagen fiber disorganization was the earliest and most prevalent change. The severity of mucoid degeneration and chondroid metaplasia in the ACL increased with the development of cartilage lesions. A correlation between ACL degeneration and cartilage degeneration was observed, especially in the medial compartment of the knee joint. CONCLUSION Our findings indicate that ACL degeneration is highly prevalent in knees with cartilage defects and may even precede cartilage changes. Hence, ACL deficiencies may not only be important in posttraumatic OA, but may also be a feature associated with knee OA pathogenesis in general.

Chromosomal instability drives metastasis through a cytosolic DNA response

Chromosomal instability is a hallmark of cancer that results from ongoing errors in chromosome segregation during mitosis. Although chromosomal instability is a major driver of tumour evolution, its role in metastasis has not been established. Here we show that chromosomal instability promotes metastasis by sustaining a tumour cell-autonomous response to cytosolic DNA. Errors in chromosome segregation create a preponderance of micronuclei whose rupture spills genomic DNA into the cytosol. This leads to the activation of the cGAS–STING (cyclic GMP-AMP synthase–stimulator of interferon genes) cytosolic DNA-sensing pathway and downstream noncanonical NF-κB signalling. Genetic suppression of chromosomal instability markedly delays metastasis even in highly aneuploid tumour models, whereas continuous chromosome segregation errors promote cellular invasion and metastasis in a STING-dependent manner. By subverting lethal epithelial responses to cytosolic DNA, chromosomally unstable tumour cells co-opt chronic activation of innate immune pathways to spread to distant organs.

Ultrashort–Echo Time MR Imaging of the Patella with Bicomponent Analysis: Correlation with Histopathologic and Polarized Light Microscopic Findings

PURPOSE To correlate short and long T2* water fractions, derived from ultrashort-echo time (TE) magnetic resonance (MR) imaging, with semiquantitative histopathologic and polarized light microscopic (PLM) assessment of human cadaveric patellae cartilage. MATERIALS AND METHODS Twenty human cadaveric patellae were evaluated by using ultrashort-TE imaging, spin-echo imaging, histopathologic analysis, and PLM, with institutional review board approval. Short and long T2* water components were evaluated for each patella by using bicomponent fitting of ultrashort-TE signal decay. Four to six regions of interest (ROIs) within each patella were chosen for correlation between ultrashort-TE bicomponent analysis, histopathologic grading (Mankin score), and PLM grading (Vaudey score). RESULTS Ultrashort-TE imaging with bicomponent analysis showed two distinct water components with a short T2* and a longer T2* in all patellae. ROI analysis showed that the short T2* fraction was correlated significantly with the Mankin (ρ = 0.66, P < .001) and Vaudey (ρ = 0.68, P < .001) scores. The Mankin scores were weakly positively correlated with T2 (ρ = 0.28, P = .13) and short T2* (ρ = 0.24, P = .14) but were negatively correlated with long T2* (ρ = -0.55, P < .01). The Vaudey scores were weakly positively correlated with T2 (ρ = 0.18, P = .16) and short T2* (ρ = 0.22, P = .14) but were negatively correlated with long T2* (ρ = -0.55, P < .01). CONCLUSION Short T2* water fraction derived from ultrashort-TE imaging with bicomponent analysis correlates significantly with both the Mankin and Vaudey scores and may serve as a biomarker of cartilage degeneration.

Zone-specific Gene Expression Patterns in Articular Cartilage

OBJECTIVE To identify novel genes and pathways specific to the superficial zone (SZ), middle zone (MZ), and deep zone (DZ) of normal articular cartilage. METHODS Articular cartilage was obtained from the knees of 4 normal human donors. The cartilage zones were dissected on a microtome. RNA was analyzed on human genome arrays. The zone-specific DNA array data obtained from human tissue were compared to array data obtained from bovine cartilage. Genes differentially expressed between zones were evaluated using direct annotation for structural or functional features, and by enrichment analysis for integrated pathways or functions. RESULTS The greatest differences in genome-wide RNA expression data were between the SZ and DZ in both human and bovine cartilage. The MZ, being a transitional zone between the SZ and DZ, thereby shared some of the same pathways as well as structural/functional features of the adjacent zones. Cellular functions and biologic processes that were enriched in the SZ relative to the DZ included, most prominently, extracellular matrix-receptor interactions, cell adhesion molecule functions, regulation of actin cytoskeleton, ribosome-related functions, and signaling aspects such as the IFN, IL4, Cdc42/Rac, and JAK/STAT signaling pathways. Two pathways were enriched in the DZ relative to the SZ, including PPARG and EGFR/SMRTE. CONCLUSION These differences in cartilage zonal gene expression identify new markers and pathways that govern the unique differentiation status of chondrocyte subpopulations.

Transplantation of engineered organoids enables rapid generation of metastatic mouse models of colorectal cancer

Colorectal cancer (CRC) is a leading cause of death in the developed world, yet facile preclinical models that mimic the natural stages of CRC progression are lacking. Through the orthotopic engraftment of colon organoids we describe a broadly usable immunocompetent CRC model that recapitulates the entire adenoma–adenocarcinoma–metastasis axis in vivo. The engraftment procedure takes less than 5 minutes, shows efficient tumor engraftment in two-thirds of mice, and can be achieved using organoids derived from genetically engineered mouse models (GEMMs), wild-type organoids engineered ex vivo, or from patient-derived human CRC organoids. In this model, we describe the genotype and time-dependent progression of CRCs from adenocarcinoma (6 weeks), to local disseminated disease (11–12 weeks), and spontaneous metastasis (>20 weeks). Further, we use the system to show that loss of dysregulated Wnt signaling is critical for the progression of disseminated CRCs. Thus, our approach provides a fast and flexible means to produce tailored CRC mouse models for genetic studies and pre-clinical investigation.

Response of an aggressive periosteal aneurysmal bone cyst (ABC) of the radius to denosumab therapy

Aneurysmal bone cyst (ABC), once considered a reactive lesion, has been proven to be a neoplasia characterized by rearrangements of the USP6-gene. Aggressive local growth and recurrences are common and therapeutic options may be limited due to the vicinity of crucial structures. We describe a case of a locally aggressive, multinucleated giant cell-containing lesion of the forearm of a 21-year old woman, treated with denosumab for recurrent, surgically uncontrollable disease. Under the influence of this RANKL inhibitor, the tumor showed a marked reduction of the content of the osteoclastic giant cells and an extensive metaplastic osteoid production leading to the bony containment, mostly located intracortically in the proximal radius. The diagnosis of a periosteal ABC was confirmed by FISH demonstrating USP6 gene rearrangement on the initial biopsy. Function conserving surgery could be performed, enabling reconstruction of the affected bone. Inhibition of RANKL with denosumab may offer therapeutic option for patients not only with giant cell tumors but also with ABCs.

Prognostic value of tumor suppressors in osteosarcoma before and after neoadjuvant chemotherapy

BackgroundPrimary bone cancers are among the deadliest cancer types in adolescents, with osteosarcomas being the most prevalent form. Osteosarcomas are commonly treated with multi-drug neoadjuvant chemotherapy and therapy success as well as patient survival is affected by the presence of tumor suppressors. In order to assess the prognostic value of tumor-suppressive biomarkers, primary osteosarcoma tissues were analyzed prior to and after neoadjuvant chemotherapy.MethodsWe constructed a tissue microarray from high grade osteosarcoma samples, consisting of 48 chemotherapy naïve biopsies (BXs) and 47 tumor resections (RXs) after neoadjuvant chemotherapy. We performed immunohistochemical stainings of P53, P16, maspin, PTEN, BMI1 and Ki67, characterized the subcellular localization and related staining outcome with chemotherapy response and overall survival. Binary logistic regression analysis was used to analyze chemotherapy response and Kaplan-Meier-analysis as well as the Cox proportional hazards model was applied for analysis of patient survival.ResultsNo significant associations between biomarker expression in BXs and patient survival or chemotherapy response were detected. In univariate analysis, positive immunohistochemistry of P53 (P = 0.008) and P16 (P16; P = 0.033) in RXs was significantly associated with poor survival prognosis. In addition, presence of P16 in RXs was associated with poor survival in multivariate regression analysis (P = 0.003; HR = 0.067) while absence of P16 was associated with good chemotherapy response (P = 0.004; OR = 74.076). Presence of PTEN on tumor RXs was significantly associated with an improved survival prognosis (P = 0.022).ConclusionsPositive immunohistochemistry (IHC) of P16 and P53 in RXs was indicative for poor overall patient survival whereas positive IHC of PTEN was prognostic for good overall patient survival. In addition, we found that P16 might be a marker of osteosarcoma chemotherapy resistance. Therefore, our study supports the use of tumor RXs to assess the prognostic value of biomarkers.