SOX2 Expression in Canine Neoplasia

2020 ◽  
pp. 030098582096013
Author(s):  
Ileana C. Miranda ◽  
Andrew D. Miller
Keyword(s):  
Rational Design ◽  
Prognostic Indicator ◽  
Screening Methods ◽  
Tumor Type ◽  
Sox2 Expression ◽  
Formalin Fixed Paraffin ◽  
Normal Expression ◽  
Formalin Fixed Paraffin Embedded ◽  
Stem Cell Pluripotency ◽  
Tumor Types

SOX2 is a major transcriptional regulator of stem cell pluripotency and self-renewability. Its expression in cancer stem cells from several different tumor types in humans and rodent models directly implicates SOX2 in tumorigenicity, metastasis, drug resistance, recurrence, and poor survival. Our objective was to investigate the expression of SOX2 in canine neoplasia. Immunohistochemistry for SOX2 was performed in sets of 10 archived formalin-fixed paraffin-embedded tissues from 45 distinct canine neoplasms. Normal expression of SOX2 was evaluated in a canine tissue microarray. Strong and diffuse SOX2 intranuclear immunolabeling was consistently found in the majority of ectodermal (13/15) and endodermal tumors (5/7). Negative, variable, or inconsistent SOX2 intranuclear immunolabeling was detected in the majority of mesodermal tumors (10/16) and in tumors with dual or uncertain origin (5/7). Although further studies are necessary to understand mechanistically how SOX2 contributes to the biology of each tumor type, this study demonstrates the expression of SOX2 in a wide variety of canine cancers. In the future, screening methods based on cellular plasticity and pluripotency biomarkers may provide avenues for the rational design of therapeutic strategies that target vulnerable signals upstream or downstream of SOX2 in different cancers, and possibly offer novel clinical applications for SOX2 as a prognostic indicator.

scholarly journals Time and tumor type (primary or metastatic) do not influence the detection of BRAF/NRAS mutations in formalin fixed paraffin embedded samples from melanomas

2016 ◽  
Vol 54 (11) ◽  
Author(s):  
Miriam Potrony ◽  
Celia Badenas ◽  
Bénédicte Naerhuyzen ◽  
Paula Aguilera ◽  
Joan Anton Puig-Butille ◽  
...  
Keyword(s):  
Melanoma Patient ◽  
Primary Melanoma ◽  
Sentinel Lymph Nodes ◽  
Tumor Type ◽  
Formalin Fixed Paraffin ◽  
Ffpe Samples ◽  
Formalin Fixed Paraffin Embedded ◽  
Complete Sequencing ◽  
Formalin Fixed ◽  
Tumor Area

AbstractBackground:Methods:DNA was obtained from 144 FFPE samples (62 primary melanoma, 43 sentinel lymph nodes [SLN] and 39 metastasis).Results:Complete sequencing results were obtained from 75% (108/144) of the samples, and at least one gene was sequenced in 89% (128/144) of them.Conclusions:Preserving sufficient tumor area in FFPE blocks is important. It is necessary to keep the FFPE blocks, no matter their age, as they are necessary to decide the best treatment for the melanoma patient.

scholarly journals Apolipoprotein D Expression in Primary Brain Tumors: Analysis by Quantitative RT-PCR in Formalin-fixed, Paraffin-embedded Tissue

2005 ◽  
Vol 53 (8) ◽  
pp. 963-969 ◽  
Author(s):  
Stephen B. Hunter ◽  
Vijay Varma ◽  
Bahig Shehata ◽  
J.D.L. Nolen ◽  
Cynthia Cohen ◽  
...  
Keyword(s):  
Brain Tumors ◽  
Tumor Type ◽  
Rt Pcr ◽  
Who Grade ◽  
Primary Brain Tumors ◽  
Formalin Fixed Paraffin ◽  
Pilocytic Astrocytomas ◽  
Formalin Fixed Paraffin Embedded ◽  
Apolipoprotein D ◽  
Formalin Fixed

Apolipoprotein D (apoD) expression has been shown to correlate both with cell cycle arrest and with prognosis in several types of malignancy, including central nervous system astrocytomas and medulloblastomas. ApoD expression was investigated by real-time quantitative RT-PCR using RNA extracted from 68 formalin-fixed, paraffin-embedded brain specimens. Glyceraldehyde phosphate dehydrogenase was used as an internal control. Quantitation was achieved on all specimens. Sixteen poorly infiltrating WHO grade I glial neoplasms (i.e., pilocytic astrocytomas and gangliogliomas) showed an average 20-fold higher apoD expression level compared with the 20 diffusely infiltrating glial neoplasms (i.e., glioblastoma, anaplastic astrocytoma, oligodendrogliomas; p=0.00004). A small number of exceptions (i.e., two high-expressing glioblastomas and three low-expressing gangliogliomas) were identified. Analyzed as individual tumor groups, poorly infiltrating grade I pilocytic astrocytomas and gangliogliomas differed significantly from each tumor type within the diffusely infiltrating higher-grade category ( p<0.05 for each comparison) but not from each other ( p>0.05). Conversely, each individual tumor type within the diffusely infiltrating category differed significantly from both pilocytic astrocytomas and gangliogliomas ( p<0.05) but did not vary from other infiltrating tumors ( p>0.05). Ependymomas, non-infiltrating grade II neoplasms, expressed levels of apoD similar to or lower than levels expressed by the diffusely infiltrating gliomas. Ten medulloblastomas with survival longer than 3 years averaged slightly higher apoD expression than four fatal medulloblastomas; however, this result was not statistically significant and individual exceptions were notable. In 17 of the medulloblastomas, MIB-1 proliferation rates quantitated by image cytometry did not correlate with apoD expression. In addition, apoD expression was 5-fold higher in the slowly proliferating grade I glial neoplasms compared with non-proliferating normal brain tissue ( p=0.01), suggesting that apoD expression is not simply an inverse measure of proliferation. ApoD expression measured by quantitative RT-PCR may be useful in the differential diagnosis of primary brain tumors, particularly pilocytic astrocytomas and gangliogliomas.

Frequency of actionable genomic alterations in early-stage lung adenocarcinoma (LA) detected by next-generation sequencing (NGS).

2012 ◽  
Vol 30 (15_suppl) ◽  
pp. e17541-e17541
Author(s):  
Kai Wang ◽  
Philip Stephens ◽  
Roman Yelensky ◽  
Jeffrey S. Ross ◽  
Vincent A. Miller ◽  
...  
Keyword(s):  
Early Stage ◽  
Mtor Inhibitors ◽  
Driver Mutations ◽  
Tumor Type ◽  
Genomic Alterations ◽  
Genomic Libraries ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Ffpe Tissues ◽  
Risk Patients

e17541 Background: Early stage LA (≤T1N0) has a variable natural history. Although many patients are cured (~2/3) with surgery, many tumors recur and better methods are needed to distinguish these subsets and tailor therapy accordingly. We undertook a pilot study of NGS in such tumors to characterize the frequency and diversity of actionable genomic alterations in this setting. Methods: Formalin-fixed paraffin embedded (FFPE) tissues from 21 archival samples collected from 2007 to 2011 were obtained commercially with accompanying clinical and pathologic data. Clinical characteristics: female (17)/male (4); T1N0 (18), AIS (2), MIA (1). Pathologic LA subtype: acinar (n=8), mucinous (n=4), lepidic (n=4), and mixed (n=2) in addition to AIS (n=2) and MIA (n=1). Targeted NGS performed in a CLIA laboratory (Foundation Medicine) gave evaluable results in all cases. Genomic libraries were captured for 3230 exons in 182 cancer-related genes plus 37 introns from 14 genes often rearranged in cancer and sequenced to average median depth of 640X with 99% of bases covered >100X. Results: Forty-one driver mutations were identified in 17 genes among the 21 patients (avg= 1.9) including 28 base substitutions, 5 indels, 4 amplifications, 3 homozygous deletions and 1 rearrangement. Notably, 90% of cases (19/21) harbored at least one alteration that can be classified as actionable-linked to an approved therapy in LA or another tumor type or a clinical trial. Examples include alterations in EGFR (n=6), NF1 (n=2, PI3K/mTOR inhibitors), CDNK2A (n=1, CDK inhibitors), EML4-ALK (fusion, n=1, ALK inhibitor), ERBB2 (n=1, TKI inhibitors), STK11 (n=1, mTOR inhibitors), and MDM2 (n=1, nutlins). Conclusions: NGSidentified drivergenomic alterations in T1N0 LA, MIA and AIS. On average, two known driver mutations were identified per tumor, many of which are associated with existing or emerging targeted therapies. This provides rationale for future investigation of larger series of T1N0 LA, AIS and MIA, using NGS to evaluate the prognostic value of these changes and could presage adjuvant trials in higher risk patients.

Programmed death-ligand 1 (PD-L1) expression in cured and not cured testicular and other germ cell tumors (GCT).

2016 ◽  
Vol 34 (2_suppl) ◽  
pp. 485-485 ◽  
Author(s):  
Brandon David Bernard ◽  
Laurence Albiges ◽  
Sabina Signoretti ◽  
Jesse Novak ◽  
Michelle S. Hirsch ◽  
...  
Keyword(s):  
Immune Cell ◽  
Therapeutic Option ◽  
Germ Cell Tumors ◽  
Unknown Origin ◽  
Mediastinal Tumors ◽  
Formalin Fixed Paraffin ◽  
Programmed Death ◽  
Formalin Fixed Paraffin Embedded ◽  
Tumor Types ◽  
Formalin Fixed

485 Background: PD-L1 is involved in immune regulation and is prognostic in many tumor types. The aim of this study was to assess PD-L1 expression in orchiectomy and metastases (mets) from GCT and to assess for an association with outcome. Methods: Immunohistochemistry (IHC) on whole sections from archival formalin-fixed paraffin-embedded tissue from Dana-Farber Cancer Institute (DFCI) and Gustave Roussy (IGR) was performed using anti-PD-L1 antibody E1L3N. Stained slides were scored semi-quantitatively and assessed independently. At DFCI, PD-L1 was scored as percent of positive tumor cells (TC) and extent of positive immune cell (IC) infiltrate; at IGR, a cut-off of 5% defined positive TC and IC. PD-L1 status was assessed by site, histology and chemotherapy (CT) status. PD-L1 status was correlated with clinical outcome. Most samples at DFCI were pre-CT (74.2%) whereas most at IGR were post-CT (85.4%). Results: IHC from 171 patients with GCT (89 DFCI, 82 IGR) from 1987-2014 was reviewed. The DFCI cohort included 69 orchiectomy, 15 mets and 5 unknown origin with 28 total deaths; the IGR cohort included 26 lung mets, 36 mediastinal or retroperitoneal lymph node mets and 20 primary mediastinal tumors. DFCI had 38 seminoma (S), 50 nonseminoma (NS) and 1 unknown; at IGR, 13 S and 69 NS. In both cohorts, PD-L1 staining in IC was higher in S (DFCI 33/38 moderate (mod)-high (86.8%); IGR 12/13 positive (92.3%)) vs. NS (DFCI 26/50 mod-high (52.0%); IGR 35/68 positive (51.5%)) (DFCI p = 0.003; IGR p = 0.006). At DFCI, 5 samples had PD-L1 positive TC (2 mixed GCT, 2 pure choriocarcinomas (CC) and 1 S); at IGR, 14 had positive TC (all NS, including 11 pure CC). More PD-L1 positive TC were CC than other GCT types (DFCI p = 0.003; IGR p < 0.001). In 36 S orchiectomy samples at DFCI, 0/4 that scored none-mild died and 2/31 (6.5%) that scored mod-high died (p = 1.00); in 30 NS orchiectomy samples, 4/9 (44.4%) that scored none-mild died and 2/16 (12.5%) that scored mod-high died (p = 0.14). Conclusions: Higher PD-L1 status in orchiectomy is not associated with GCT survival. PD-L1 positive TC may serve as a histological marker for CC among NS. Higher PD-L1 expression on IC in S and on TC in CC may point to immunotherapy as a therapeutic option in these tumors.

scholarly journals Cell Line Macroarray

2016 ◽  
Vol 64 (12) ◽  
pp. 739-751 ◽  
Author(s):  
Alberto La Spada ◽  
Simona Baronchelli ◽  
Linda Ottoboni ◽  
Francesca Ruffini ◽  
Gianvito Martino ◽  
...  
Keyword(s):  
Stem Cell ◽  
Cell Line ◽  
Induced Pluripotent Stem Cell ◽  
Screening Methods ◽  
Image Analyzer ◽  
Biomarker Validation ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Bona Fide ◽  
Induced Pluripotent

In the past decade, tissue microarray (TMA) technology has evolved as an innovative tool for high-throughput proteomics analysis and mainly for biomarker validation. Similarly, enormous amount of data can be obtained from the cell line macroarray (CLMA) technology, which developed from the TMA using formalin-fixed, paraffin-embedded cell pellets. Here, we applied CLMA technology in stem cell research and in particular to identify bona fide neogenerated human induced pluripotent stem cell (hiPSC) clones suitable for down the line differentiation. All hiPSC protocols generate tens of clones, which need to be tested to determine genetically stable cell lines suitable for differentiation. Screening methods generally rely on fluorescence-activated cell sorting isolation and coverslip cell growth followed by immunofluorescence; these techniques could be cumbersome. Here, we show the application of CLMA to identify neogenerated pluripotent cell colonies and neuronal differentiated cell products. We also propose the use of the automated image analyzer, TissueQuest, as a reliable tool to quickly select the best clones, based upon the level of expression of multiple pluripotent biomarkers.

scholarly journals Development and Validation of an ERCC1 Immunohistochemistry Assay for Solid Tumors

2016 ◽  
Vol 140 (12) ◽  
pp. 1397-1403 ◽  
Author(s):  
Brittany N. Bahamon ◽  
Feng Gao ◽  
Hadi Danaee
Keyword(s):  
Dynamic Range ◽  
Excision Repair ◽  
Critical Role ◽  
Therapeutic Modalities ◽  
Sensitivity Assessment ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Key Enzyme ◽  
Ercc1 Expression ◽  
Tumor Types

Context.— Excision repair cross-complementation 1 (ERCC1) is a key enzyme in nuclear excision repair pathway and has a critical role in helping remove DNA adducts caused by cross-linking agents, such as platinum-containing cancer chemotherapies and other DNA-damaging therapeutic modalities. ERCC1 expression, evaluated by techniques such as immunohistochemistry, has been associated with clinical response; ERCC1+ tumors are more resistant to cisplatin treatment than are ERCC1− tumors. Although several immunohistochemistry, anti-ERCC1 antibodies are available, the 8F1 clone, in particular, has been used in many studies. Recent evidence has suggested that the 8F1 antibody cross-reacts with at least one other protein, raising concerns about the specificity of this clone. Objective.— To design an immunohistochemistry assay to detect ERCC1 levels that show dynamic range and consistent analytic performance. Design.— Two different primary antibodies to ERCC1, clones 4F9 and D6G6, were evaluated on formalin-fixed, paraffin-embedded tissue. We then performed a fit-for-purpose assay validation with the 4F9 clone, which included sensitivity assessment across several solid tumor types and evaluation of analytic parameters, such as precision and reproducibility. Results.— The 4F9 clone was consistently superior to the D6G6 clone in the optimization phase. A range of expression was seen in ovarian, head and neck, non–small cell lung, and esophageal cancer samples when tested with the 4F9 clone. The antibody showed acceptable reproducibility (31.02%) and precision (16.06%). Conclusions.— This assay can be used to assess ERCC1 levels during clinical studies of patient tumors from a variety of tumor types.

Quantitative Evaluation of Protein Expression as a Function of Tissue Microarray Core Diameter: Is a Large (1.5 mm) Core Better Than a Small (0.6 mm) Core?

2010 ◽  
Vol 134 (4) ◽  
pp. 613-619
Author(s):  
Valsamo K. Anagnostou ◽  
Frank J. Lowery ◽  
Konstantinos N. Syrigos ◽  
Philip T. Cagle ◽  
David L. Rimm
Keyword(s):  
Growth Factor Receptor ◽  
Tissue Microarrays ◽  
Core Size ◽  
Experimental Conditions ◽  
Core Diameter ◽  
Formalin Fixed Paraffin ◽  
Identical Manner ◽  
Formalin Fixed Paraffin Embedded ◽  
Epidermal Growth ◽  
Tumor Types

Abstract Context.—Tissue microarrays (TMAs) have emerged as a high-throughput technology for protein evaluation in large cohorts. This technique allows maximization of tissue resources by analysis of sections from 0.6-mm to 1.5-mm core “biopsies” of standard formalin-fixed, paraffin-embedded tissue blocks and by the processing of hundreds of cases arrayed on a single recipient block in an identical manner. Objective.—To assess the expression of a series of biomarkers as a function of core size. Although pathologists frequently feel better if larger core sizes are used, there is no evidence in the literature showing that large cores are better (or worse) than small cores for assessment of TMAs. Design.—Estrogen receptor, HER2/neu, epidermal growth factor receptor, STAT3, mTOR, and phospho-p70 S6 kinase were measured by immunofluorescence with automated quantitative analysis. One random 0.6-mm field (one 0.6-mm spot) was compared to 6 to 12 fields per spot, representing 1-mm and 1.5-mm cores, for 3 different tumor types. Results.—We show that measurement of a single random 0.6-mm spot was comparable to analysis of the whole 1-mm or 1.5-mm spot (Pearson R coefficient varying from 0.87–0.98) for all markers tested. Conclusions.—Since TMA technology is now being used in all phases of biomarker development, this work shows that TMAs with 0.6-mm cores are as representative as those with any common larger core size for optimization of standardized experimental conditions. Given that a greater number of 0.6-cores can be arrayed in a single master block, use of this core size allows increased throughput and decreased cost.

scholarly journals Higher Mutation Burden in High Proliferation Compartments of Heterogeneous Melanoma Tumors

2021 ◽  
Vol 22 (8) ◽  
pp. 3886
Author(s):  
Tomasz M. Grzywa ◽  
Agnieszka A. Koppolu ◽  
Wiktor Paskal ◽  
Klaudia Klicka ◽  
Małgorzata Rydzanicz ◽  
...  
Keyword(s):  
Single Nucleotide Variants ◽  
Primary Cutaneous Melanoma ◽  
Ki 67 ◽  
Formalin Fixed Paraffin ◽  
Mutation Burden ◽  
Formalin Fixed Paraffin Embedded ◽  
Cause Of Failure ◽  
Tumor Types ◽  
Next Generation Sequencing Ngs

Melanoma tumors are the most heterogeneous of all tumor types. Tumor heterogeneity results in difficulties in diagnosis and is a frequent cause of failure in treatment. Novel techniques enable accurate examination of the tumor cells, considering their heterogeneity. The study aimed to determine the somatic variations among high and low proliferating compartments of melanoma tumors. In this study, 12 archival formalin-fixed paraffin-embedded samples of previously untreated primary cutaneous melanoma were stained with Ki-67 antibody. High and low proliferating compartments from four melanoma tumors were dissected using laser-capture microdissection. DNA was isolated and analyzed quantitatively and qualitatively. Libraries for amplicon-based next-generation sequencing (NGS) were prepared using NEBNext Direct Cancer HotSpot Panel. NGS detected 206 variants in 42 genes in melanoma samples. Most of them were located within exons (135, 66%) and were predominantly non-synonymous single nucleotide variants (99, 73.3%). The analysis showed significant differences in mutational profiles between high and low proliferation compartments of melanoma tumors. Moreover, a significantly higher percentage of variants were detected only in high proliferation compartments (39%) compared to low proliferation regions (16%, p < 0.05). Our results suggest a significant functional role of genetic heterogeneity in melanoma.

scholarly journals Comprehensive NGS Panel Validation for the Identification of Actionable Alterations in Adult Solid Tumors

2021 ◽  
Vol 11 (5) ◽  
pp. 360
Author(s):  
Paula Martínez-Fernández ◽  
Patricia Pose ◽  
Raquel Dolz-Gaitón ◽  
Arantxa García ◽  
Inmaculada Trigo-Sánchez ◽  
...  
Keyword(s):  
Solid Tumors ◽  
Limit Of Detection ◽  
Copy Number Variants ◽  
Hybridization Capture ◽  
Formalin Fixed Paraffin ◽  
Ffpe Tumor ◽  
Formalin Fixed Paraffin Embedded ◽  
Tumor Types ◽  
Next Generation Sequencing Ngs ◽  
Formalin Fixed

The increasing identification of driver oncogenic alterations and progress of targeted therapies addresses the need of comprehensive alternatives to standard molecular methods. The translation into clinical practice of next-generation sequencing (NGS) panels is actually challenged by the compliance of high quality standards for clinical accreditation. Herein, we present the analytical and clinical feasibility study of a hybridization capture-based NGS panel (Action OncoKitDx) for the analysis of somatic mutations, copy number variants (CNVs), fusions, pharmacogenetic SNPs and Microsatellite Instability (MSI) determination in formalin-fixed paraffin-embedded (FFPE) tumor samples. A total of 64 samples were submitted to extensive analytical validation for the identification of previously known variants. An additional set of 166 tumor and patient-matched normal samples were sequenced to assess the clinical utility of the assay across different tumor types. The panel demonstrated good specificity, sensitivity, reproducibility, and repeatability for the identification of all biomarkers analyzed and the 5% limit of detection set was validated. Among the clinical cohorts, the assay revealed pathogenic genomic alterations in 97% of patient cases, and in 82.7%, at least one clinically relevant variant was detected. The validation of accuracy and robustness of this assay supports the Action OncoKitDx’s utility in adult solid tumors.

scholarly journals Targeted RNAseq of Formalin-Fixed Paraffin-Embedded Tissue to Differentiate Among Benign and Malignant Adrenal Cortical Tumors

2020 ◽  
Vol 52 (08) ◽  
pp. 607-613
Author(s):  
Samuel W. Plaska ◽  
Chia-Jen Liu ◽  
Jung Soo Lim ◽  
Juilee Rege ◽  
Nolan R. Bick ◽  
...  
Keyword(s):  
Translational Research ◽  
Transcriptomic Analysis ◽  
Adrenal Tumors ◽  
Tumor Type ◽  
Next Generation ◽  
Reverse Transcription Pcr ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Tissue Specimens ◽  
Formalin Fixed

AbstractLack of routine fresh or frozen tissue is a barrier to widespread transcriptomic analysis of adrenal cortical tumors and an impediment to translational research in endocrinology and endocrine oncology. Our group has previously pioneered the use of targeted amplicon-based next-generation sequencing for archival formalin-fixed paraffin-embedded (FFPE) adrenal tissue specimens to characterize the spectrum of somatic mutations in various forms of primary aldosteronism. Herein, we developed and validated a novel 194-amplicon targeted next-generation RNA sequencing (RNAseq) assay for transcriptomic analysis of adrenal tumors using clinical-grade FFPE specimens. Targeted RNAseq-derived expression values for 27 adrenal cortical tumors, including aldosterone-producing adenomas (APA; n=8), cortisol-producing adenomas (CPA; n=11), and adrenal cortical carcinomas (ACC; n=8), highlighted known differentially-expressed genes (DEGs; i. e., CYP11B2, IGF2, etc.) and tumor type-specific transcriptional modules (i. e., high cell cycle/proliferation transcript expression in ACC, etc.), and a subset of DEGs was validated orthogonally using quantitative reverse transcription PCR (qRT-PCR). Finally, unsupervised hierarchical clustering using a subset of high-confidence DEGs revealed three discrete clusters representing APA, CPA, and ACC tumors with corresponding unique gene expression signatures, suggesting potential clinical utility for a transcriptomic-based approach to tumor classification. Overall, these data support the use of targeted amplicon-based RNAseq for comprehensive transcriptomic profiling of archival FFPE adrenal tumor material and indicate that this approach may facilitate important translational research opportunities for the study of these tumors.

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