Common genetic variation at human 8q23.3 is significantly associated with colorectal cancer (CRC) risk. To elucidate the basis of this association we compared the frequency of common variants at ...8q23.3 in 1,964 CRC cases and 2,081 healthy controls. Reporter gene studies showed that the single nucleotide polymorphism rs16888589 acts as an allele-specific transcriptional repressor. Chromosome conformation capture (3C) analysis demonstrated that the genomic region harboring rs16888589 interacts with the promoter of gene for eukaryotic translation initiation factor 3, subunit H (EIF3H). We show that increased expression of EIF3H gene increases CRC growth and invasiveness thereby providing a biological mechanism for the 8q23.3 association. These data provide evidence for a functional basis for the non-coding risk variant rs16888589 at 8q23.3 and provides novel insight into the etiological basis of CRC.
Objective
To evaluate our institutional experience with molecular diagnostics (MD) on thyroid cytology smears, evaluate the costs and describe MD guided clinical management of indeterminate Bethesda ...III/V thyroid nodules.
Methods
We performed a retrospective review of 164 Bethesda III or V thyroid cytopathology reports subjected to MD from 2013 to 2020, that altered Bethesda classification or management. MD consisted of mutation and gene fusion analysis by next‐generation sequencing (NGS) of morphologically analysed and selected cytological slides. Findings were modelled to nationwide data on Bethesda incidences from ‘the Dutch Pathology Registry’ PALGA, and costs were estimated.
Results
82 of 164 cases received an upgrade in Bethesda class. Twenty cases changed from Bethesda III to IV/V, 62 from Bethesda III or V to VI, and 72 remained unaltered. We estimate net savings with implementing MD, by preventing 454 repeat cytology and 326 (diagnostic) hemithyroidectomies, to be at least 2 million Euro annually in the Netherlands. Per Bethesda III and V patient, net savings would be about 100 Euro and 4100 Euro, respectively.
Conclusion
NGS‐based MD on nucleic acids extracted directly from cytology slides is a feasible and cost saving tool for personalized management in indeterminate Bethesda III/V thyroid cytology. Based on the interpretation of our retrospective data, we assume that this approach results in less disease burden for the patient, reduced surgical interventions and complication risks, reduced sick leave, among others. Further evaluation of structural implementation of the presented approach in routine thyroid Bethesda III/V cytology in a prospective setting is warranted.
We evaluated our institutional experience with molecular diagnostics (MD) on thyroid cytology smears, evaluated the costs and described MD guided clinical management of indeterminate Bethesda III/V thyroid nodules. NGS‐based MD on nucleic acids extracted directly from cytology slides is a feasible and cost saving tool for personalized management in indeterminate Bethesda III/V thyroid cytology.
Germline mutations in POLE and POLD1 have been shown to cause predisposition to colorectal multiple polyposis and a wide range of neoplasms, early-onset colorectal cancer being the most prevalent. In ...order to find additional mutations affecting the proofreading activity of these polymerases, we sequenced its exonuclease domain in 155 patients with multiple polyps or an early-onset colorectal cancer phenotype without alterations in the known hereditary colorectal cancer genes. Interestingly, none of the previously reported mutations in POLE and POLD1 were found. On the other hand, among the genetic variants detected, only two of them stood out as putative pathogenic in the POLE gene, c.1359 + 46del71 and c.1420G > A (p.Val474Ile). The first variant, detected in two families, was not proven to alter correct RNA splicing. Contrarily, c.1420G > A (p.Val474Ile) was detected in one early-onset colorectal cancer patient and located right next to the exonuclease domain. The pathogenicity of this change was suggested by its rarity and bioinformatics predictions, and it was further indicated by functional assays in Schizosaccharomyces pombe. This is the first study to functionally analyze a POLE genetic variant outside the exonuclease domain and widens the spectrum of genetic changes in this DNA polymerase that could lead to colorectal cancer predisposition.
Small intestine-neuroendocrine tumors (SI-NETs) are one of the most common tumors of the small bowel. Despite an increasing incidence, the exact mechanisms driving underlying pathology remain to be ...determined. Interestingly, recent studies linked the development of (SI-)NETs to both Lynch syndrome (LS) and MUTYH variants. If confirmed, these associations would have important consequences for treatment. In this study we therefore investigated the prevalence of mismatch repair (MMR) deficiency and MUTYH variants in 64 primary resected SI-NETs. Immunohistochemistry was used to assess the expression of the MMR genes, and competitive allele-specific PCR (KASPar) targeting two hotspot MUTYH variants p.(Tyr179Cys), p.(Gly396Asp) was performed to determine their prevalence in SI-NETs. Strikingly, all 64 SI-NETs stained positive for MSH6 and PMS2, indicating MMR proficiency. In addition, no MUTYH hotspot variant was found in any of the 64 SI-NETs. As such, these results do not support an association between SI-NET development and LS or MUTYH variants. In order to gain insight into SI-NET pathogenesis and optimally manage patients, future research should therefore focus on other candidate genes.
•In a cohort of 64 primary resected small intestine-neuroendocrine tumors, no mismatch repair deficiency was identified.•In a cohort of 64 primary resected small intestine-neuroendocrine tumors, no MUTYH hotspot variants were found.•Universal screening of small intestine-neuroendocrine tumors for mismatch repair deficiency and MUTYH variants is not recommended.
Preoperative biopsies or imbedded cytological cells will become more and more a primary source of tissue for molecular diagnostic analyses as a result of novel neo-adjuvant treatment regimens for ...several cancer types. Furthermore there is a growing need to examine metastatic cancer tissue. Hence, nucleic acids need to be reliably isolated and analyzed from small amounts of formalin-fixed and paraffin-embedded (FFPE) tissue. The limited numbers of (tumor) cells in these samples make high quality and sensitive DNA isolation challenging. Also demands for faster turnaround times are growing. Therefore, we evaluated a fully automated DNA/RNA isolation system and compared this with a manual, classical routine molecular pathology method. We compared the quality of the isolates from both tissue cores and micro-dissection for detection of hotspot mutations in KRAS, BRAF applying hydrolysis probe assays. In addition we determined whether the automated method decreases the hands-on-time and turnaround times in routine molecular pathology workflow.
In conclusion, the automated method delivers high quality DNA from both small FFPE tissue cores and micro-dissected tissue material. In comparison to classical methods, less than 50% of starting tissue was sufficient as input for micro-dissection. Turnaround times decreased significantly and 50% less hands-on time was needed.
Abstract
Context
DICER1 syndrome is a rare autosomal-dominantly inherited disorder that predisposes to a variety of cancerous and noncancerous tumors of mostly pediatric and adolescent onset, ...including differentiated thyroid carcinoma (DTC). DTC has been hypothesized to arise secondarily to the increased prevalence of thyroid hyperplastic nodules in syndromic patients.
Objective
To determine somatic alterations in DICER1-associated DTC and to study patient outcomes.
Design
Retrospective series.
Setting
Tertiary referral centers.
Patients
Ten patients with germline pathogenic DICER1 variants and early-onset DTC.
Methods
Somatic DICER1 mutation analysis, extensive somatic DNA variant and gene fusion analyses were performed on all tumors.
Results
Median age at DTC diagnosis was 13.5 years and there was no recurrent or metastatic disease (median follow-up, 8 years). All thyroid specimens showed diffuse nodular hyperplasia with at least one focus suspicious of DTC but without infiltrative growth, extrathyroidal extension, vascular invasion, or lymph node metastasis. Most of the individual nodules (benign and malignant) sampled from the 10 tumors harbored distinct DICER1 RNase IIIb hotspot mutations, indicating a polyclonal composition of each tumor. Furthermore, nine of 10 DICER1-related DTCs lacked well-known oncogenic driver DNA variants and gene rearrangements.
Conclusion
On the basis of our clinical, histological, and molecular data, we consider that most DICER1-related DTCs form a low-risk subgroup. These tumors may arise within one of multiple benign monoclonal nodules; thus, hemi-thyroidectomy or, more likely, total thyroidectomy may often be required. However, radioiodine treatment may be unnecessary given the patients’ ages and the tumors’ low propensity for metastases.
Characterization of 10 DICER1-related thyroid carcinomas showed, in most cases, the presence of multiple, distinct, low-risk polyclonal tumors that may alter treatment strategies.
The 2016 World Health Organization classification defines diffuse large B-cell lymphoma (DLBCL) subtypes based on Epstein-Barr virus (EBV) infection and oncogenic rearrangements of
as drivers of ...lymphomagenesis. A subset of DLBCL, however, is characterized by activating mutations in
We investigated whether
mutations could improve the classification and prognostication of DLBCL. In 250 primary DLBCL,
mutations were identified by allele-specific polymerase chain reaction or next-generation-sequencing,
rearrangements were analyzed by fluorescence
hybridization, and EBV was studied by EBV-encoded RNA
hybridization. Associations of molecular features with clinicopathologic characteristics, outcome, and prognosis according to the International Prognostic Index (IPI) were investigated.
and
mutations were identified in 29.6% and 12.3%,
, and
rearrangements in 10.6%, 13.6%, and 20.3%, and EBV in 11.7% of DLBCL, respectively. Prominent mutual exclusivity between EBV positivity, rearrangements, and
mutations established the value of molecular markers for the recognition of biologically distinct DLBCL subtypes.
-mutated DLBCL had a significantly inferior 5-year overall survival than wild-type
DLBCL (log-rank;
=0.019). DLBCL without any of the studied aberrations had superior overall survival compared to cases carrying ≥1 aberrancy (log-rank;
=0.010).
mutations retained their adverse prognostic impact upon adjustment for other genetic and clinical variables by multivariable analysis and improved the prognostic performance of the IPI. This study demonstrates the clinical utility of defining
-mutated DLBCL as a distinct molecular subtype with adverse prognosis. Our data call for sequence analysis of
in routine diagnostics of DLBCL to optimize classification and prognostication, and to guide the development of improved treatment strategies.
Endobronchial Ultrasound Guided Transbronchial Needle Aspiration (EBUS-TBNA) and Trans-esophageal Ultrasound Scanning with Fine Needle Aspiration (EUS-FNA) are important, novel techniques for the ...diagnosis and staging of non-small cell lung cancer (NSCLC) that have been incorporated into lung cancer staging guidelines. To guide and optimize treatment decisions, especially for NSCLC patients in stage III and IV, EGFR and KRAS mutation status is often required. The concordance rate of the mutation analysis between these cytological aspirates and histological samples obtained by surgical staging is unknown. Therefore, we studied the extent to which allele-specific quantitative real-time PCR with hydrolysis probes could be reliably performed on EBUS and EUS fine needle aspirates by comparing the results with histological material from the same patient. We analyzed a series of 43 NSCLC patients for whom cytological and histological material was available. We demonstrated that these standard molecular techniques can be accurately applied on fine needle cytological aspirates from NSCLC patients. Importantly, we show that all mutations detected in the histological material of primary tumor were also identified in the cytological samples. We conclude that molecular profiling can be reliably performed on fine needle cytology aspirates from NSCLC patients.
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