Preoperative molecular testing in thyroid nodules with Bethesda VI cytology: Clinical experience and review of the literature.

Risk assessment is critical to determine the timing of elective surgeries and preserve valuable resources in time of pandemic. This study was undertaken to better understand the potential value of molecular testing to risk-stratify thyroid nodules with malignant cytology (Bethesda VI). Systematic review of the literature contributed 21 studies representing 2036 preoperative specimens.

Analytical and clinical validation of pairwise microRNA expression analysis to identify medullary thyroid cancer in thyroid fine-needle aspiration samples.

Background: Medullary thyroid carcinoma (MTC) is an aggressive malignancy originating from the parafollicular C cells. Preoperatively, thyroid nodule fine-needle aspiration cytology (FNAC) and pathogenic gene mutations are definitive in approximately one-half of cases. MicroRNAs (miRNAs) are endogenous, noncoding, single-stranded RNAs that regulate gene expression, a characteristic that confers the potential for identifying malignancy.

miRNA expression can classify pediatric thyroid lesions and increases the diagnostic yield of mutation testing.

Background: Genetic alterations in multiple cell signaling pathways are involved in the molecular pathogenesis of thyroid cancer. Oncogene mutation testing and gene-expression profiling are routinely used for the preoperative risk management of adult thyroid nodules. In this study, we evaluated the potential value of miRNA biomarkers for the classification of pediatric thyroid lesions.

Molecular Testing for Oncogenic Gene Alterations in Pediatric Thyroid Lesions.

Background: Thyroid nodules are less common in pediatric patients (i.e., those ≤18 years) than they are in adults.

Molecular classification of thyroid lesions by combined testing for miRNA gene expression and somatic gene alterations.

Multiple molecular markers contribute to the pathogenesis of thyroid cancer and can provide valuable information to improve disease diagnosis and patient management. We performed a comprehensive evaluation of miRNA gene expression in diverse thyroid lesions (n = 534) and developed predictive models for the classification of thyroid nodules, alone or in combination with genotyping. Expression profiling by reverse transcription-quantitative polymerase chain reaction in surgical specimens (n = 257) identified specific miRNAs differentially expressed in 17 histopathological categories.

Utility and cost-effectiveness of molecular testing in thyroid nodules with indeterminate cytology.

Context: Molecular testing on biopsies from thyroid nodules with indeterminate cytology can improve patient management by preventing unnecessary surgeries on benign nodules.

Objective: The aim of the study was to determine the health outcome benefits and cost-effectiveness of molecular testing in nodules with AUS/FLUS or FN/SFN cytology.

Design: The initial diagnosis and treatment of a hypothetical cohort of adult U.

Molecular Testing for miRNA, mRNA, and DNA on Fine-Needle Aspiration Improves the Preoperative Diagnosis of Thyroid Nodules With Indeterminate Cytology.

Context: Molecular testing for oncogenic mutations or gene expression in fine-needle aspirations (FNAs) from thyroid nodules with indeterminate cytology identifies a subset of benign or malignant lesions with high predictive value.

Objective: This study aimed to evaluate a novel diagnostic algorithm combining mutation detection and miRNA expression to improve the diagnostic yield of molecular cytology.

Setting: Surgical specimens and preoperative FNAs (n = 638) were tested for 17 validated gene alterations using the miRInform Thyroid test and with a 10-miRNA gene expression classifier generating positive (malignant) or negative (benign) results.

Conversion, correction, and International Scale standardization: results From a Multicenter External Quality Assessment Study for BCR-ABL1 testing.

Context: Monitoring BCR-ABL1 expression levels relative to clinically validated response criteria on the International Scale (IS) is vital in the optimal management of patients with chronic myeloid leukemia, yet significant variability remains across laboratories worldwide.

Objective: To assess method performance, interlaboratory precision, and different IS standardization modalities in representative laboratories performing routine BCR-ABL1 testing.

Design: Fifteen blinded test specimens with 5-level nominal BCR-ABL1 to ABL1 IS percentage ratios ranging from 5% to 0.

Molecular testing for oncogenic gene mutations in thyroid lesions: a case-control validation study in 413 postsurgical specimens.

Molecular testing for oncogenic gene alterations provides clinically actionable information essential for the optimal management of follicular cell thyroid cancer. We aimed to establish the distribution and frequency of common oncogenic gene mutations and chromosomal rearrangements in a comprehensive set of benign and malignant thyroid lesions. A case-control study was conducted in 413 surgical cases comprising 17 distinct histopathologic categories, 244 malignant, 169 benign, and 304 double-blinded specimens.

Centralized molecular testing for oncogenic gene mutations complements the local cytopathologic diagnosis of thyroid nodules.

Background: Molecular testing for oncogenic gene mutations and chromosomal rearrangements plays a growing role in the optimal management of thyroid nodules, yet lacks standardized testing modalities and systematic validation data. Our objective was to assess the performance of molecular cytology on preoperative thyroid nodule fine-needle aspirates (FNAs) across a broad range of variables, including independent collection sites, clinical practices, and anatomic pathology interpretations.

Methods: Single-pass FNAs were prospectively collected from 806 nodules 1 cm or larger by ultrasonography at five independent sites across the United States.

A microRNA-based test improves endoscopic ultrasound-guided cytologic diagnosis of pancreatic cancer.

Background & Aims: Endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) in combination with cytopathology is the optimal method for diagnosis and staging of pancreatic ductal adenocarcinoma (PDAC) and other pancreatic lesions. Its clinical utility, however, can be limited by high rates of indeterminate or false-negative results. We aimed to develop and validate a microRNA (miRNA)-based test to improve preoperative detection of PDAC.

A multiplex technology platform for the rapid analysis of clinically actionable genetic alterations and validation for BRAF p.V600E detection in 1549 cytologic and histologic specimens.

Context: Current clinicopathologic assessment of malignant neoplastic diseases entails the analysis of specific genetic alterations that provide diagnostic, prognostic, or therapy-determining information.

Objective: To develop and validate a robust molecular method to detect clinically relevant mutations in various tissue types and anatomic pathology specimens.

Design: Genes of interest were amplified by multiplex polymerase chain reaction and sequence variants identified by liquid bead array cytometry.

Establishment and validation of analytical reference panels for the standardization of quantitative BCR-ABL1 measurements on the international scale.

Background: Current guidelines for managing Philadelphia-positive chronic myeloid leukemia include monitoring the expression of the BCR-ABL1 (breakpoint cluster region/c-abl oncogene 1, non-receptor tyrosine kinase) fusion gene by quantitative reverse-transcription PCR (RT-qPCR). Our goal was to establish and validate reference panels to mitigate the interlaboratory imprecision of quantitative BCR-ABL1 measurements and to facilitate global standardization on the international scale (IS).

Methods: Four-level secondary reference panels were manufactured under controlled and validated processes with synthetic Armored RNA Quant molecules (Asuragen) calibrated to reference standards from the WHO and the NIST.

An optimized technology platform for the rapid multiplex molecular analysis of genetic alterations associated with leukemia.

Molecular methods play a critical role in the accurate diagnosis of leukemia by complementing morphologic, cytochemical, immunophenotypic, and cytogenetic analyses. We developed a multiplex reverse transcription-polymerase chain reaction (RT-PCR) method combined with liquid bead array cytometry for the rapid detection of genetic alterations associated with leukemia. Fusion transcripts corresponding to the most common recurrent chromosomal translocations were reproducibly detected in as low as 0.

Human native lipoprotein-induced de novo DNA methylation is associated with repression of inflammatory genes in THP-1 macrophages.

Background: We previously showed that a VLDL- and LDL-rich mix of human native lipoproteins induces a set of repressive epigenetic marks, i.e. de novo DNA methylation, histone 4 hypoacetylation and histone 4 lysine 20 (H4K20) hypermethylation in THP-1 macrophages.

Sensitive multiplex detection of KRAS codons 12 and 13 mutations in paraffin-embedded tissue specimens.

Background: Colorectal cancer patients harbouring KRAS mutations in codon 12 or 13 do not benefit from current anti-epidermal growth factor receptor (EGFR) monoclonal antibody therapies. Efficient and robust methods are therefore required for routine clinical testing of KRAS mutation status.

Aims: To evaluate a novel multiplex assay for the rapid detection of common KRAS mutations in formalin-fixed paraffin-embedded (FFPE) tissues.

Performance and clinical evaluation of a sensitive multiplex assay for the rapid detection of common NPM1 mutations.

Determination of NPM1 mutation status has become essential for the molecular classification of acute myeloid leukemias (AML). Methods with high clinical sensitivity and specificity adapted to the molecular laboratory workflow are required for the diagnosis, prognosis, and monitoring of AML with normal karyotype. We report here the development and evaluation of a novel, streamlined, RNA-based assay for the rapid multiplex detection of common NPM1 mutations in a 96-well assay format.

International standards and reference materials for quantitative molecular infectious disease testing.

The utility of quantitative molecular diagnostics for patient management depends on the ability to relate patient results to prior results or to absolute values in clinical practice guidelines. To do this, those results need to be comparable across time and methods, either by producing the same value across methods and test versions or by using reliable and stable conversions. Universally available standards and reference materials specific to quantitative molecular technologies are critical to this process but are few in number.

Analysis of microRNAs in pancreatic fine-needle aspirates can classify benign and malignant tissues.

Background: MicroRNAs (miRNAs) are RNA molecules that are involved in the regulation of many cellular processes, including those related to human cancers. The aim of this study was to determine, as a proof of principle, whether specific candidate miRNAs could be detected in fine-needle aspirate (FNA) biopsies of pancreatic ductal adenocarcinoma (PDAC) and could accurately differentiate malignant from benign pancreatic tissues.

Methods: We used TaqMan(R) assays to quantify miRNA levels in FNA samples collected in RNARetain (n = 16) and compared the results with a training set consisting of frozen macrodissected pancreatic samples (n = 20).

Accurate molecular characterization of formalin-fixed, paraffin-embedded tissues by microRNA expression profiling.

Formalin-fixed, paraffin-embedded tissues are an invaluable tool for biomarker discovery and validation. As these archived specimens are not always compatible with modern genomic techniques such as gene expression arrays, we assessed the use of microRNA (miRNA) as an alternative means for the reliable molecular characterization of formalin-fixed, paraffin-embedded tissues. Expression profiling using two different microarray platforms and multiple mouse and human formalin-fixed, paraffin-embedded tissue types resulted in the correlation ratios of miRNA expression levels between frozen and fixed tissue pairs ranging from 0.

Asuragen Inc.

Asuragen is a fully integrated molecular diagnostics company with products and services that incorporate cutting-edge technologies for isolating, detecting and quantifying nucleic acids. Asuragen's Signature product line offers streamlined methods for the detection of important genetic alterations and innovative RNA-based solutions for molecular oncology. The company has also pioneered research on microRNA (miRNA) function, isolation and detection in clinical samples.

Evaluation and validation of total RNA extraction methods for microRNA expression analyses in formalin-fixed, paraffin-embedded tissues.

Histopathology archives of well-annotated formalin-fixed, paraffin-embedded (FFPE) tissue specimens are valuable resources for retrospective studies of human diseases. Since recovery of quality intact mRNA compatible with molecular techniques is often difficult due to degradation, we evaluated microRNA (miRNA), a novel class of small RNA molecules with growing therapeutic and diagnostic potential, as an alternative analyte for gene expression studies of FFPE samples. Analyzing total RNA yield, miRNA recovery, and robustness of real-time polymerase chain reaction for miRNA, mRNA, and rRNA species, we compared the performance of commercially available RNA isolation kits and identified a preferred methodology.