The impact of molecular and mismatch repair status on the survival outcomes of surgically treated patients with colorectal peritoneal metastases.

Background: Stratification of patients with colorectal peritoneal metastases (CRPM) using RAS/BRAF mutational status may refine patient selection for cytoreductive surgery (CRS) and hyperthermic intraperitoneal chemotherapy (HIPEC). This study aimed to analyse the association of RAS/BRAF status and their variants, with clinicopathological variables and survival outcomes in patients who have undergone CRS ± HIPEC.

Methods: A single centre, peritonectomy database was interrogated for patients with CRPM who underwent peritonectomy procedures between 2010 and 2020.

Clinical validation and implementation of droplet digital PCR for the detection of BRAF mutations from cell-free DNA.

Droplet digital PCR (ddPCR) has been demonstrated in many research studies to be a sensitive method in the analysis of circulating tumour DNA (ctDNA) for identifying mutations and tracking disease. The transition of ddPCR into the diagnostic setting requires a number of critical steps including the assessment of accuracy and precision and ultimately implementation into clinical use. Here we present the clinical validation of ddPCR for the detection of BRAF mutations (V600E and V600K) from plasma.

Identification of a recurrent mosaic variant in brain tissue from an individual with nevus sebaceous syndrome.

Nevus sebaceous syndrome (NSS) is a rare, multisystem neurocutaneous disorder, characterized by a congenital nevus, and may include brain malformations such as hemimegalencephaly or focal cortical dysplasia, ocular, and skeletal features. It has been associated with several eponyms including Schimmelpenning and Jadassohn. The isolated skin lesion, nevus sebaceous, is associated with postzygotic variants in or in all individuals studied.

ROS1 rearrangements in non-small cell lung cancer: screening by immunohistochemistry using proportion of cells staining without intensity and excluding cases with MAPK pathway drivers improves test performance.

Therapeutically actionable ROS1 rearrangements have been described in 1-3% of non-small cell lung cancer (NSCLC). Screening for ROS1 rearrangements is recommended to be by immunohistochemistry (IHC), followed by confirmation with fluorescence in situ hybridisation (FISH) or sequencing. However, in practise ROS1 IHC presents difficulties due to conflicting scoring systems, multiple clones and expression in tumours that are wild-type for ROS1.

Towards Routine Implementation of Liquid Biopsies in Cancer Management: It Is Always Too Early, until Suddenly It Is Too Late.

Blood-based liquid biopsies are considered a new and promising diagnostic and monitoring tool for cancer. As liquid biopsies only require a blood draw, they are non-invasive, potentially more rapid and assumed to be a less costly alternative to genomic analysis of tissue biopsies. A multi-disciplinary workshop ( = 98 registrations) was organized to discuss routine implementation of liquid biopsies in cancer management.

Breast ductal carcinoma in situ carry mutational driver events representative of invasive breast cancer.

The spectrum of genomic alterations in ductal carcinoma in situ (DCIS) is relatively unexplored, but is likely to provide useful insights into its biology, its progression to invasive carcinoma and the risk of recurrence. DCIS (n=20) with a range of phenotypes was assessed by massively parallel sequencing for mutations and copy number alterations and variants validated by Sanger sequencing. PIK3CA mutations were identified in 11/20 (55%), TP53 mutations in 6/20 (30%), and GATA3 mutations in 9/20 (45%).

Blood-based detection of RAS mutations to guide anti-EGFR therapy in colorectal cancer patients: concordance of results from circulating tumor DNA and tissue-based RAS testing.

An accurate blood-based RAS mutation assay to determine eligibility of metastatic colorectal cancer (mCRC) patients for anti-EGFR therapy would benefit clinical practice by better informing decisions to administer treatment independent of tissue availability. The objective of this study was to determine the level of concordance between plasma and tissue RAS mutation status in patients with mCRC to gauge whether blood-based RAS mutation testing is a viable alternative to standard-of-care RAS tumor testing. RAS testing was performed on plasma samples from newly diagnosed metastatic patients, or from recurrent mCRC patients using the highly sensitive digital PCR technology, BEAMing (beads, emulsions, amplification, and magnetics), and compared with DNA sequencing data of respective FFPE (formalin-fixed paraffin-embedded) tumor samples.

Clinicopathological characteristics associated with BRAF(K601E) and BRAF(L597) mutations in melanoma.

BRAF mutations at codons L597 and K601 occur uncommonly in melanoma. Clinical and pathological associations of these mutations were investigated in a cohort of 1119 patients with known BRAF mutation status. A BRAF mutation was identified in 435 patients; Mutations at L597 and the K601E mutation were seen in 3.

Transcriptional profiling of the postnatal brain of the Ts1Cje mouse model of Down syndrome.

The Ts1Cje mouse model of Down syndrome (DS) has partial trisomy of mouse chromosome 16 (MMU16), which is syntenic to human chromosome 21 (HSA21). It develops various neuropathological features demonstrated by DS patients such as reduced cerebellar volume [1] and altered hippocampus-dependent learning and memory [2,3]. To understand the global gene expression effect of the partially triplicated MMU16 segment on mouse brain development, we performed the spatiotemporal transcriptome analysis of Ts1Cje and disomic control cerebral cortex, cerebellum and hippocampus harvested at four developmental time-points: postnatal day (P)1, P15, P30 and P84.

Functional transcriptome analysis of the postnatal brain of the Ts1Cje mouse model for Down syndrome reveals global disruption of interferon-related molecular networks.

Background: The Ts1Cje mouse model of Down syndrome (DS) has partial triplication of mouse chromosome 16 (MMU16), which is partially homologous to human chromosome 21. These mice develop various neuropathological features identified in DS individuals. We analysed the effect of partial triplication of the MMU16 segment on global gene expression in the cerebral cortex, cerebellum and hippocampus of Ts1Cje mice at 4 time-points: postnatal day (P)1, P15, P30 and P84.

High frequency of germline TP53 mutations in a prospective adult-onset sarcoma cohort.

Sarcomas are a key feature of Li-Fraumeni and related syndromes (LFS/LFL), associated with germline TP53 mutations. Current penetrance estimates for TP53 mutations are subject to significant ascertainment bias. The International Sarcoma Kindred Study is a clinic-based, prospective cohort of adult-onset sarcoma cases, without regard to family history.

Quantitative threefold allele-specific PCR (QuanTAS-PCR) for highly sensitive JAK2 V617F mutant allele detection.

Background: The JAK2 V617F mutation is the most frequent somatic change in myeloproliferative neoplasms, making it an important tumour-specific marker for diagnostic purposes and for the detection of minimal residual disease. Sensitive quantitative assays are required for both applications, particularly for the monitoring of minimal residual disease, which requires not only high sensitivity but also very high specificity.

Methods: We developed a highly sensitive probe-free quantitative mutant-allele detection method, Quantitative Threefold Allele-Specific PCR (QuanTAS-PCR), that is performed in a closed-tube system, thus eliminating the manipulation of PCR products.

A multisite blinded study for the detection of BRAF mutations in formalin-fixed, paraffin-embedded malignant melanoma.

Melanoma patients with BRAF mutations respond to treatment with vemurafenib, thus creating a need for accurate testing of BRAF mutation status. We carried out a blinded study to evaluate various BRAF mutation testing methodologies in the clinical setting. Formalin-fixed, paraffin-embedded melanoma samples were macrodissected before screening for mutations using Sanger sequencing, single-strand conformation analysis (SSCA), high resolution melting analysis (HRM) and competitive allele-specific TaqMan® PCR (CAST-PCR).

Proteomic and metabolomic analyses of mitochondrial complex I-deficient mouse model generated by spontaneous B2 short interspersed nuclear element (SINE) insertion into NADH dehydrogenase (ubiquinone) Fe-S protein 4 (Ndufs4) gene.

Eukaryotic cells generate energy in the form of ATP, through a network of mitochondrial complexes and electron carriers known as the oxidative phosphorylation system. In mammals, mitochondrial complex I (CI) is the largest component of this system, comprising 45 different subunits encoded by mitochondrial and nuclear DNA. Humans diagnosed with mutations in the gene NDUFS4, encoding a nuclear DNA-encoded subunit of CI (NADH dehydrogenase ubiquinone Fe-S protein 4), typically suffer from Leigh syndrome, a neurodegenerative disease with onset in infancy or early childhood.

Detection of BRAF mutations in patients with hairy cell leukemia and related lymphoproliferative disorders.

Hairy cell leukemia has been shown to be strongly associated with the BRAF V600E mutation. We screened 59 unenriched archived bone marrow aspirate and peripheral blood samples from 51 patients with hairy cell leukemia using high resolution melting analysis and confirmatory Sanger sequencing. The BRAF V600E mutation was detected in 38 samples (from 36 patients).

Analysing DNA methylation using bisulphite pyrosequencing.

Bisulphite pyrosequencing is a quantitative methodology for the investigation of DNA methylation of sequences up to 100-bp in length. Biotin-labelled, single-stranded PCR products generated from bisulphite-treated DNA are used as a template with an internal primer to perform the pyrosequencing reaction. Nucleotides are added in a predetermined order in each pyrosequencing cycle and the amount of incorporated nucleotide results in a proportional emission of light.

Investigating the potential role of genetic and epigenetic variation of DNA methyltransferase genes in hyperplastic polyposis syndrome.

Background: Hyperplastic Polyposis Syndrome (HPS) is a condition associated with multiple serrated polyps, and an increased risk of colorectal cancer (CRC). At least half of CRCs arising in HPS show a CpG island methylator phenotype (CIMP), potentially linked to aberrant DNA methyltransferase (DNMT) activity. CIMP is associated with methylation of tumor suppressor genes including regulators of DNA mismatch repair (such as MLH1, MGMT), and negative regulators of Wnt signaling (such as WIF1).

Spatiotemporal regulation of multiple overlapping sense and novel natural antisense transcripts at the Nrgn and Camk2n1 gene loci during mouse cerebral corticogenesis.

Nrgn and Camk2n1 are highly expressed in the brain and play an important role in synaptic long-term potentiation via regulation of Ca(2+)/calmodulin-dependent protein kinase II. We have shown that the gene loci for these 2 proteins are actively transcribed in the adult cerebral cortex and feature multiple overlapping transcripts in both the sense and antisense orientations with alternative polyadenylation. These transcripts were upregulated in the adult compared with embryonic and P1.

Gene network disruptions and neurogenesis defects in the adult Ts1Cje mouse model of Down syndrome.

Background: Down syndrome (DS) individuals suffer mental retardation with further cognitive decline and early onset Alzheimer's disease.

Methodology/principal Findings: To understand how trisomy 21 causes these neurological abnormalities we investigated changes in gene expression networks combined with a systematic cell lineage analysis of adult neurogenesis using the Ts1Cje mouse model of DS. We demonstrated down regulation of a number of key genes involved in proliferation and cell cycle progression including Mcm7, Brca2, Prim1, Cenpo and Aurka in trisomic neurospheres.

A multicenter blinded study to evaluate KRAS mutation testing methodologies in the clinical setting.

Evidence that activating mutations of the KRAS oncogene abolish the response to anti-epidermal growth factor receptor therapy has revolutionized the treatment of advanced colorectal cancer. This has resulted in the urgent demand for KRAS mutation testing in the clinical setting to aid choice of therapy. The aim of this study was to evaluate six different KRAS mutation detection methodologies on two series of primary colorectal cancer samples.

Molecular networks involved in mouse cerebral corticogenesis and spatio-temporal regulation of Sox4 and Sox11 novel antisense transcripts revealed by transcriptome profiling.

Background: Development of the cerebral cortex requires highly specific spatio-temporal regulation of gene expression. It is proposed that transcriptome profiling of the cerebral cortex at various developmental time points or regions will reveal candidate genes and associated molecular pathways involved in cerebral corticogenesis.

Results: Serial analysis of gene expression (SAGE) libraries were constructed from C57BL/6 mouse cerebral cortices of age embryonic day (E) 15.

Hematopoietic defects in the Ts1Cje mouse model of Down syndrome.

Down syndrome (DS) persons are born with various hematopoietic abnormalities, ranging from relatively benign, such as neutrophilia and macrocytosis, to a more severe transient myeloproliferative disorder (TMD). In most cases, these abnormalities resolve in the first few months to years of life. However, sometimes the TMD represents a premalignant disease that develops into acute megakaryocytic leukemia (AMKL), usually in association with acquired GATA1 mutations.