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Background: MicroRNAs (miRNAs), small RNA molecules of approximately 22 nucleotides, have been shown to be up- or downregulated in specific cell types and disease states. These molecules have become recognized as one of the major regulatory gatekeepers of coding genes in the human genome. Content: We review the structure, nomenclature, mechanism of action, technologies used for miRNA detection, and associations of miRNAs with human cancer. miRNAs are produced in a tissue-specific manner, and changes in miRNA within a tissue type can be correlated with disease status. miRNAs appear to regulate mRNA translation and degradation via mechanisms that are dependent on the degree of complementarity between the miRNA and mRNA molecules. miRNAs can be detected via several methods, such as microarrays, bead-based arrays, and quantitative real-time PCR. The tissue concentrations of specific miRNAs have been associated with tumor invasiveness, metastatic potential, and other clinical characteristics for several types of cancers, including chronic lymphocytic leukemia, and breast, colorectal, hepatic, lung, pancreatic, and prostate cancers. Summary: By targeting and controlling the expression of mRNA, miRNAs can control highly complex signal-transduction pathways and other biological pathways. The biologic roles of miRNAs in cancer suggest a correlation with prognosis and therapeutic outcome. Further investigation of these roles may lead to new approaches for the categorization, diagnosis, and treatment of human cancers.

Completion of the Human Genome Project, in conjunction with dramatic reductions in the cost of DNA sequencing and advances in translational research, is gradually ushering genomic discoveries and technologies into the practice of medicine. The rapid pace of these advances is opening up a gap between the knowledge available about the clinical relevance of genomic information and the ability of clinicians to include such information in their medical practices. This educational gap threatens to be rate limiting to the clinical adoption of genomics in medicine. Solutions will require not only a better understanding of the clinical implications of genetic discoveries but also training in genomics at all levels of professional development, including for individuals in formal training and others who long ago completed such training. The National Human Genome Research Institute has convened the Inter-Society Coordinating Committee for Physician Education in Genomics (ISCC) to develop and share best practices in the use of genomics in medicine. The ISCC has developed a framework for development of genomics practice competencies that may serve as a starting point for formulation of competencies for physicians in various medical disciplines. Genet Med 16 11, 804–809.

Opportunity is the essence of a career in science or medicine and this continues to be a major source of satisfaction for the many scientists and health care providers worldwide. Having trained as a PhD scientist, the world of clinical medicine seemed to be a galaxy away. I could not have been more wrong in that assumption. Here, I tell the story of my career trajectory so that those new to the sciences understand the potential and the opportunities afforded by a career in clinical laboratory medicine.

Evidence contends lower levels of physical exertion reduce blood pressure (BP) as effectively as more rigorous levels. We compared the effects of low (40% peak oxygen consumption, V o 2peak), moderate (60% V o 2peak), and vigorous (100% V o 2peak) exercise intensity on the BP response immediately following aerobic exercise. We also examined clinical correlates of the BP response. Subjects were 45 men (mean ± SEM, 43.9 ± 1.4 years) with elevated awake ambulatory BP (ABP, 144.5 ± 1.5/85.4 ± 1.2 mm Hg). Men completed four randomly assigned experiments: non-exercise control and three exercise bouts at low, moderate, and vigorous intensity. All experiments began with a baseline period of seated rest. Subjects left the laboratory wearing an ABP monitor. Systolic ABP increased 2.8 ± 1.6 mm Hg less after low, 5.4 ± 1.4 mm Hg less after moderate, and 11.7 ± 1.5 mm Hg less after vigorous than control over 9 h ( P < .001). Diastolic ABP decreased 1.5 ± 1.2 mm Hg more after low, 2.0 ± 1.0 mm Hg more after moderate, and 4.9 ± 1.3 mm Hg more after vigorous versus control over 9 h ( P < .010). Baseline correlates of the systolic ABP post-exercise response to vigorous were fasting glucose ( r = -0.415), C-reactive protein ( r = -0.362), renin ( r = −0.348), fasting insulin ( r = 0.310), and fasting low density lipoprotein ( r = −0.298) ( R 2 = 0.400, P = .002). Baseline correlates of the diastolic ABP post-exercise response to vigorous were V o 2peak ( r = −0.431), fasting low density lipoprotein ( r = -0.431), renin ( r = -0.411), fibrinogen ( r = 0.369), and fasting glucose ( r = −0.326) ( R 2 = 0.429, P < .001). The antihypertensive effects of exercise intensity occurred in dose response fashion. Clinicians should weigh the benefits and risks of prescribing vigorous exercise intensity for those with hypertension on an individual basis.

Background: The poor prognosis of pancreatic ductal adenocarcinoma (PDAC) is accounted for by the absence of early diagnostic markers and effective treatments. MicroRNAs inhibit the translation of their target mRNAs. The production of microRNAs is strongly altered in cancers, but the causes of these alterations are only partially known. DNA hypermethylation is a major cause of gene inactivation in cancer. Our aims were to identify microRNAs whose gene expression is inactivated by hypermethylation in PDAC and to determine whether this hypermethylation-mediated repression is an early event during pancreatic carcinogenesis. We also sought to investigate whether these differentially methylated regions can serve as a diagnostic marker for PDAC. Methods: MicroRNA production was measured by microarray hybridization and reverse-transcription quantitative PCR. The level of DNA methylation was measured by bisulfite mapping and semiquantitative methylation-specific PCR. Results: We identified 29 microRNAs encoded by genes whose expression is potentially inactivated by DNA hypermethylation. We focused our study on microRNA 148a (miR-148a) and found its production to be repressed, not only in PDAC samples but also in preneoplastic pancreatic intraepithelial neoplasia (PanIN) lesions. More importantly, we found that hypermethylation of the DNA region encoding miR-148a is responsible for its repression, which occurs in PanIN preneoplastic lesions. Finally, we show that the hypermethylated DNA region encoding miR-148a can serve as an ancillary marker for the differential diagnosis of PDAC and chronic pancreatitis (CP). Conclusions: We show that the hypermethylation of the DNA region encoding miR-148a is responsible for its repression in PDAC precursor lesions and can be a useful tool for the differential diagnosis of PDAC and CP.

Digital PCR (dPCR) is emerging as an ideal platform for the detection and tracking of genomic variants in cancer due to its high sensitivity and simple workflow. The growing number of clinically actionable cancer biomarkers creates a need for fast, accessible methods that allow for dense information content and high accuracy. Here, we describe a proof‐of‐concept amplitude modulation‐based multiplex dPCR assay capable of detecting 12 single‐nucleotide and insertion/deletion (indel) variants in EGFR , KRAS , BRAF , and ERBB2 , 14 gene fusions in ALK , RET , ROS1 , and NTRK1 , and MET exon 14 skipping present in non‐small cell lung cancer (NSCLC). We also demonstrate the use of multi‐spectral target‐signal encoding to improve the specificity of variant detection by reducing background noise by up to an order of magnitude. The assay reported an overall 100% positive percent agreement (PPA) and 98.5% negative percent agreement (NPA) compared with a sequencing‐based assay in a cohort of 62 human formalin‐fixed paraffin‐embedded (FFPE) samples. In addition, the dPCR assay rescued actionable information in 10 samples that failed to sequence, highlighting the utility of a multiplexed dPCR assay as a potential reflex solution for challenging NSCLC samples.

Background Central nervous system tumors are among the most lethal types of cancer. A critical factor for tailored neurosurgical resection strategies depends on specific tumor types. However, it is uncommon to have a preoperative tumor diagnosis, and intraoperative morphology-based diagnosis remains challenging. Despite recent advances in intraoperative methylation classifications of brain tumors, accuracy may be compromised by low tumor purity. Copy number variations (CNVs), which are almost ubiquitous in cancer, offer highly sensitive molecular biomarkers for diagnosis. These quantitative genomic alterations provide insight into dysregulated oncogenic pathways and can reveal potential targets for molecular therapies. Methods We develop iSCORED, a one-step random genomic DNA reconstruction method that enables efficient, unbiased quantification of genome-wide CNVs. By concatenating multiple genomic fragments into long reads, the method leverages low-pass sequencing to generate approximately 1–2 million genomic fragments within 1 h. This approach allows for ultrafast high-resolution CNV analysis at a genomic resolution of 50 kb. In addition, concurrent methylation profiling enables brain tumor methylation classification and identifies promoter methylation in amplified oncogenes, providing an integrated diagnostic approach. Results In our retrospective cohort of 26 malignant brain tumors, iSCORED demonstrated 100% concordance in CNV detection, including chromosomal alterations and oncogene amplifications, when compared to clinically validated assays such as Next-Generation Sequencing and Chromosomal Microarray. Furthermore, we validated iSCORED’s real-time applicability in 15 diagnostically challenging primary brain tumors, achieving 100% concordance in detecting aberrant CNV detection, including diagnostic chromosomal gains/losses and oncogene amplifications (10/10). Of these, 14 out of 15 brain tumor methylation classifications aligned with final pathological diagnoses. This streamlined workflow—from tissue arrival to automatic generation of CNV and methylation reports—can be completed within 105 min. Conclusions The iSCORED pipeline represents the first method capable of high-resolution CNV detection within the intraoperative timeframe. By combining CNV detection and methylation classification, iSCORED provides a rapid and comprehensive molecular diagnostic tool that can inform rapid clinical decision. The integrated approach not only enhances the accuracy of tumor diagnosis but also optimizes surgical planning and identifies potential molecular therapies, all within the critical intraoperative timeframe.

Background: Pancreatic ductal adenocarcinoma (PDAC) has the poorest overall prognosis among gastrointestinal cancers; however, curative resection in early-stage PDAC greatly improves survival rates, indicating the importance of early detection. Because abnormal microRNA production is commonly detected in cancer, we investigated noninvasive precursor pancreatic intraepithelial neoplasia (PanIN) lesions for microRNA production as a potential early biomarker of PDAC. Methods: Pathologists identified and classified ductal lesions. We extracted total RNA from laser-capture microdissected PanIN tissue samples from a conditional KRAS(G12D) mouse model (n = 29) or of human origin (n = 38) (KRAS is v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog). MicroRNA production was quantified by quantitative real-time PCR. Internal controls included 5S and U6 RNAs. Results: Production of microRNAs miR-21, miR-205, and miR-200 paralleled PanIN progression in the KRAS(G12D) mouse model, compared with microRNA production in samples of nonpathologic ducts. miR-21 demonstrated the highest relative concentrations in the precursor lesions. Interestingly, miR-205 and miR-21 up-regulation preceded phenotypic changes in the ducts. The production of microRNAs miR-21, miR-221, miR-222, and let-7a increased with human PanIN grade, with peak production occurring in hyperplastic PanIN-2/3 lesions. In situ hybridization analysis indicated miR-21 production to be concentrated in pathologic ductal cells. miR-21 production was regulated by KRAS(G12D) and epidermal growth factor receptor in PDAC-derived cell lines. Conclusions: Aberrant microRNA production is an early event in the development of PanIN. Our findings indicate that miR-21 warrants further investigation as a marker for early detection of PDAC.

Lynch syndrome (LS) is the most common form of the hereditary colon cancer syndromes. Because of its high prevalence, a nationwide campaign has begun to screen all colorectal cancers for the genetic abnormalities associated with LS. Next to colorectal cancer, endometrial cancer is the most common form of malignancy found in women with LS. Identifying individuals who harbor the well-characterized mismatch-repair gene mutations via immunohistochemistry, microsatellite instability analysis, or direct gene sequencing is critical to managing the LS patient and to surveillance for the development of other associated tumor types. Although many institutions have begun screening all colorectal tumors for LS, the evidence is sufficient to warrant the testing of all endometrial cancers for LS as well. Various testing algorithms, along with genetic-counseling efforts, can lead to a cost-efficient and beneficial screening program.

Some epithelial neoplasms of the appendix, including low-grade appendiceal mucinous neoplasm and adenocarcinoma, can result in pseudomyxoma peritonei (PMP). Little is known about the mutational spectra of these tumor types and whether mutations may be of clinical significance with respect to therapeutic selection. In this study, we identified somatic mutations using the Ion Torrent AmpliSeq Cancer Hotspot Panel v2. Specimens consisted of 3 nonneoplastic retention cysts/mucocele, 15 low-grade mucinous neoplasms (LAMNs), 8 low-grade/well-differentiated mucinous adenocarcinomas with pseudomyxoma peritonei, and 12 adenocarcinomas with/without goblet cell/signet ring cell features. Barcoded libraries were prepared from up to 10 ng of extracted DNA and multiplexed on single 318 chips for sequencing. Data analysis was performed using Golden Helix SVS. Variants that remained after the analysis pipeline were individually interrogated using the Integrative Genomics Viewer. A single Janus kinase 3 (JAK3) mutation was detected in the mucocele group. Eight mutations were identified in the V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) and GNAS complex locus (GNAS) genes among LAMN samples. Additional gene mutations were identified in the AKT1 (v-akt murine thymoma viral oncogene homolog 1), APC (adenomatous polyposis coli), JAK3, MET (met proto-oncogene), phosphatidylinositol-4,5-bisphosphate 3-kinase (PIK3CA), RB1 (retinoblastoma 1), STK11 (serine/threonine kinase 11), and tumor protein p53 (TP53) genes. Among the PMPs, 6 mutations were detected in the KRAS gene and also in the GNAS, TP53, and RB1 genes. Appendiceal cancers showed mutations in the APC, ATM (ataxia telangiectasia mutated), KRAS, IDH1 [isocitrate dehydrogenase 1 (NADP+)], NRAS [neuroblastoma RAS viral (v-ras) oncogene homolog], PIK3CA, SMAD4 (SMAD family member 4), and TP53 genes. Our results suggest molecular heterogeneity among epithelial tumors of the appendix. Next generation sequencing efforts have identified mutational spectra in several subtypes of these tumors that may suggest a phenotypic heterogeneity showing mutations that are relevant for targeted therapies.