Combined array-comparative genomic hybridization and single-nucleotide polymorphism-loss of heterozygosity analysis reveals complex genetic alterations in cervical cancer

BACKGROUND: Cervical carcinoma develops as a result of multiple genetic alterations. Different studies investigated genomic alterations in cervical cancer mainly by means of metaphase comparative genomic hybridization (mCGH) and microsatellite marker analysis for the detection of loss of heterozygosity (LOH). Currently, high throughput methods such as array comparative genomic hybridization (array CGH), single nucleotide polymorphism array (SNP array) and gene expression arrays are available to study genome-wide alterations. Integration of these 3 platforms allows detection of genomic alterations at high resolution and investigation of an association between copy number changes and expression. RESULTS: Genome-wide copy number and genotype analysis of 10 cervical cancer cell lines by array CGH and SNP array showed highly complex large-scale alterations. A comparison between array CGH and SNP array revealed that the overall concordance in detection of the same areas with copy number alterations (CNA) was above 90%. The use of SNP arrays demonstrated that about 75% of LOH events would not have been found by methods which screen for copy number changes, such as array CGH, since these were LOH events without CNA. Regions frequently targeted by CNA, as determined by array CGH, such as amplification of 5p and 20q, and loss of 8p were confirmed by fluorescent in situ hybridization (FISH). Genome-wide, we did not find a correlation between copy-number and gene expression. At chromosome arm 5p however, 22% of the genes were significantly upregulated in cell lines with amplifications as compared to cell lines without amplifications, as measured by gene expression arrays. For 3 genes, SKP2, ANKH and TRIO, expression differences were confirmed by quantitative real-time PCR (qRT-PCR). CONCLUSION: This study showed that copy number data retrieved from either array CGH or SNP array are comparable and that the integration of genome-wide LOH, copy number and gene expression is useful for the identification of gene specific targets that could be relevant for the development and progression in cervical cancer

MLPAinter for MLPA interpretation: An integrated approach for the analysis, visualisation and data management of Multiplex Ligation-dependent Probe Amplification

Background: Multiplex Ligation-Dependent Probe Amplification (MLPA) is an application that can be used for the detection of multiple chromosomal aberrations in a single experiment. In one reaction, up to 50 different genomic sequences can be analysed. For a reliable work-flow, tools are needed for administrative support, data management, normalisation, visualisation, reporting and interpretation.Results: Here, we developed a data management system, MLPAInter for MLPA interpretation, that is windows executable and has a stand-alone database for monitoring and interpreting the MLPA data stream that is generated from the experimental setup to analysis, quality control and visualisation. A statistical approach is applied for the normalisation and analysis of large series of MLPA traces, making use of multiple control samples and internal controls.Conclusions: MLPAinter visualises MLPA data in plots with information about sample replicates, normalisation settings, and sample characteristics. This integrated approach helps in the automated handling of large series of MLPA data and guarantees a quick and streamlined dataflow from the beginning of an experiment to an authorised report

A genetic perspective on cetacean evolution

Studies of cetacean evolution using genetics and other biomolecules have come a long way—from the use of allozymes and short sequences of mitochondrial or nuclear DNA to the assembly of full nuclear genomes and characterization of proteins and lipids. Cetacean research has also advanced from using only contemporary samples to analyzing samples dating back thousands of years, and to retrieving data from indirect environmental sources, including water or sediments. Combined, these studies have profoundly deepened our understanding of the origin of cetaceans; their adaptation and speciation processes; and of the past population change, migration, and admixture events that gave rise to the diversity of cetaceans found today

A comparison of weekly versus 3-weekly cisplatin during adjuvant radiotherapy for high-risk head and neck cancer

SummaryObjectivesTo compare cumulative cisplatin dose and toxicity between patients who received 3-weekly versus weekly cisplatin during adjuvant radiotherapy for high-risk head and neck squamous cell carcinoma (HNSCC).Materials and methodsConsecutive HNSCC patients with involved resection margins and/or extra-capsular extension in two tertiary cancer centers with different institutional practices were identified. Cumulative cisplatin dose was calculated and information on toxicity reviewed and compared between patients who received 3-weekly versus weekly cisplatin.ResultsOf 270 high risk patients, 60 received 3-weekly 100mg/m2 and 48 received weekly 50mg/m2 cisplatin during adjuvant radiotherapy (60–66Gy in 30–33 fractions). Fourteen patients received other chemotherapy schedules and 148 received no chemotherapy. Mean cumulative cisplatin dose was 199.4mg/m2 (standard error (SE) 5.4) in 3-weekly versus 239.8mg/m2 (SE 11.0, P=0.001) in weekly treated patients. Cumulative cisplatin ⩾200mg/m2 was given to 67.7% of patients in the 3-weekly cohort and 85.2% (P=0.039) in the weekly cohort. The rate of feeding tube dependency 6months after treatment, osteoradionecrosis, neutropenic fever, and persistent renal function decline were not statistically different.ConclusionsAbout one half of high-risk HNSCC patients are not eligible for cisplatin during postoperative radiotherapy. Patients treated with weekly 50mg/m2 cisplatin received a higher cumulative dose with comparable toxicity as patients who received 3-weekly 100mg/m2 cisplatin. Efficacy and applicability to the frequently used weekly 40mg/m2 schedule remains to be evaluated

Universal baleen whale microsatellite panel for individual identification and power to detect parentage

Highly polymorphic single tandem repeat loci (STR, also known as microsatellite loci) remain a familiar, cost efficient class of markers for genetic analyses in ecology, behavior and conservation. We characterize a new universal set of ten STR loci (from 28 potential candidate loci) in seven baleen whale species, which are optimized for PCR amplification in two multiplex reactions along with a Y chromosome marker for sex determination. The optimized, universal set of STR loci provides an ideal starting point for new studies in baleen whales aimed at individual-based and population genetic studies, and facilitates data sharing among research groups. Data from the new STR loci were combined with genotypes from other published STR loci to assess the power to assign parentage (paternity) using exclusion in four species: fin whales, humpback whales, blue whales and bowhead whales. We argue that parentage studies should present a power analysis to demonstrate that the specific data are sufficiently informative to assign parentage with statistical rigor

The PRO-RCC study:a long-term PROspective Renal Cell Carcinoma cohort in the Netherlands, providing an infrastructure for ‘Trial within Cohorts’ study designs

BACKGROUND: Ongoing research in the field of both localized, locally advanced and metastatic renal cell carcinoma has resulted in the availability of multiple treatment options. Hence, many questions are still unanswered and await further research. A nationwide collaborative registry allows to collect corresponding data. For this purpose, the Dutch PROspective Renal Cell Carcinoma cohort (PRO-RCC) has been founded, for the prospective collection of long-term clinical data, patient reported outcome measures (PROMs) and patient reported experience measures (PREMs).METHODS: PRO-RCC is designed as a multicenter cohort for all Dutch patients with renal cell carcinoma (RCC). Recruitment will start in the Netherlands in 2023. Importantly, participants may also consent to participation in a 'Trial within cohorts' studies (TwiCs). The TwiCs design provides a method to perform (randomized) interventional studies within the registry. The clinical data collection is embedded in the Netherlands Cancer Registry (NCR). Next to the standardly available data on RCC, additional clinical data will be collected. PROMS entail Health-Related Quality of Life (HRQoL), symptom monitoring with optional ecological momentary assessment (EMA) of pain and fatigue, and optional return to work- and/or nutrition questionnaires. PREMS entail satisfaction with care. Both PROMS and PREMS are collected through the PROFILES registry and are accessible for the patient and the treating physician.TRIAL REGISTRATION: Ethical board approval has been obtained (2021_218) and the study has been registered at ClinicalTrials.gov (NCT05326620).DISCUSSION: PRO-RCC is a nationwide long-term cohort for the collection of real-world clinical data, PROMS and PREMS. By facilitating an infrastructure for the collection of prospective data on RCC, PRO-RCC will contribute to observational research in a real-world study population and prove effectiveness in daily clinical practice. The infrastructure of this cohort also enables that interventional studies can be conducted with the TwiCs design, without the disadvantages of classic RCTs such as slow patient accrual and risk of dropping out after randomization.</p

Fin whale (Balaenoptera physalus) mitogenomics: A cautionary tale of defining sub-species from mitochondrial sequence monophyly

The advent of massive parallel sequencing technologies has resulted in an increase of studies based upon complete mitochondrial genome DNA sequences that revisit the taxonomic status within and among species. Spatially distinct monophyly in such mitogenomic genealogies, i.e., the sharing of a recent common ancestor among con-specific samples collected in the same region has been viewed as evidence for subspecies. Several recent studies in cetaceans have employed this criterion to suggest subsequent intraspecific taxonomic revisions. We reason that employing intra-specific, spatially distinct monophyly at non-recombining, clonally inherited genomes is an unsatisfactory criterion for defining subspecies based upon theoretical (genetic drift) and practical (sampling effort) arguments. This point was illustrated by a re-analysis of a global mitogenomic assessment of fin whales, Balaenoptera physalus spp., published by Archer et al. (2013), which proposed to further subdivide the Northern Hemisphere fin whale subspecies, B. p. physalus. The proposed revision was based upon the detection of spatially distinct monophyly among North Atlantic and North Pacific fin whales in a genealogy based upon complete mitochondrial genome DNA sequences. The extended analysis conducted in this study (1676 mitochondrial control region, 162 complete mitochondrial genome DNA sequences and 20 microsatellite loci genotyped in 380 samples) revealed that the apparent monophyly among North Atlantic fin whales reported by Archer et al. (2013) to be due to low sample sizes. In conclusion, defining sub-species from monophyly (i.e., the absence of para- or polyphyly) can lead to erroneous conclusions due to relatively 'trivial' aspects, such as sampling. Basic population genetic processes (i.e., genetic drift and migration) also affect the time to the most recent common ancestor and hence the probability that individuals in a sample are monophyletic

Genome Haploidisation with Chromosome 7 Retention in Oncocytic Follicular Thyroid Carcinoma

Contains fulltext : 108012.pdf (publisher's version ) (Open Access)BACKGROUND: Recurrent non-medullary thyroid carcinoma (NMTC) is a rare disease. We initially characterized 27 recurrent NMTC: 13 papillary thyroid cancers (PTC), 10 oncocytic follicular carcinomas (FTC-OV), and 4 non-oncocytic follicular carcinomas (FTC). A validation cohort composed of benign and malignant (both recurrent and non-recurrent) thyroid tumours was subsequently analysed (n = 20). METHODS: Data from genome-wide SNP arrays and flow cytometry were combined to determine the chromosomal dosage (allelic state) in these tumours, including mutation analysis of components of PIK3CA/AKT and MAPK pathways. RESULTS: All FTC-OVs showed a very distinct pattern of genomic alterations. Ten out of 10 FTC-OV cases showed near-haploidisation with or without subsequent genome endoreduplication. Near-haploidisation was seen in 5/10 as extensive chromosome-wide monosomy (allelic state [A]) with near-haploid DNA indices and retention of especially chromosome 7 (seen as a heterozygous allelic state [AB]). In the remaining 5/10 chromosomal allelic states AA with near diploid DNA indices were seen with allelic state AABB of chromosome 7, suggesting endoreduplication after preceding haploidisation. The latter was supported by the presence of both near-haploid and endoreduplicated tumour fractions in some of the cases. Results were confirmed using FISH analysis. Relatively to FTC-OV limited numbers of genomic alterations were identified in other types of recurrent NMTC studied, except for chromosome 22q which showed alterations in 6 of 13 PTCs. Only two HRAS, but no mutations of EGFR or BRAF were found in FTC-OV. The validation cohort showed two additional tumours with the distinct pattern of genomic alterations (both with oncocytic features and recurrent). CONCLUSIONS: We demonstrate that recurrent FTC-OV is frequently characterised by genome-wide DNA haploidisation, heterozygous retention of chromosome 7, and endoreduplication of a near-haploid genome. Whether normal gene dosage on especially chromosome 7 (containing EGFR, BRAF, cMET) is crucial for FTC-OV tumour survival is an important topic for future research. MICROARRAYS: Data are made available at GEO (GSE31828)

Integrating chromosomal aberrations and gene expression profiles to dissect rectal tumorigenesis

<p>Abstract</p> <p>Background</p> <p>Accurate staging of rectal tumors is essential for making the correct treatment choice. In a previous study, we found that loss of 17p, 18q and gain of 8q, 13q and 20q could distinguish adenoma from carcinoma tissue and that gain of 1q was related to lymph node metastasis. In order to find markers for tumor staging, we searched for candidate genes on these specific chromosomes.</p> <p>Methods</p> <p>We performed gene expression microarray analysis on 79 rectal tumors and integrated these data with genomic data from the same sample series. We performed supervised analysis to find candidate genes on affected chromosomes and validated the results with qRT-PCR and immunohistochemistry.</p> <p>Results</p> <p>Integration of gene expression and chromosomal instability data revealed similarity between these two data types. Supervised analysis identified up-regulation of <it>EFNA1 </it>in cases with 1q gain, and <it>EFNA1 </it>expression was correlated with the expression of a target gene (<it>VEGF</it>). The <it>BOP1 </it>gene, involved in ribosome biogenesis and related to chromosomal instability, was over-expressed in cases with 8q gain. <it>SMAD2 </it>was the most down-regulated gene on 18q, and on 20q, <it>STMN3 </it>and <it>TGIF2 </it>were highly up-regulated. Immunohistochemistry for SMAD4 correlated with <it>SMAD2 </it>gene expression and 18q loss.</p> <p>Conclusion</p> <p>On basis of integrative analysis this study identified one well known CRC gene (<it>SMAD2</it>) and several other genes (<it>EFNA1, BOP1, TGIF2 </it>and <it>STMN3</it>) that possibly could be used for rectal cancer characterization.</p

Baleen whale microsatellite panel for individual identification and parentage assignment in Mysticeti

Highly polymorphic single tandem repeat loci (STR, also known as microsatellite loci) remain a familiar, cost efficient class of genetic markers in genetic studies in ecology, behavior and conservation. Here we characterize a new, universal set of ten STR loci in seven species of baleen whales, optimized for PCR amplification in two multiplex reactions along with a Y chromosome marker for sex determination. The optimized, universal set of STR loci provides a convenient starting point for new genetic studies in baleen whales aimed at identifying individuals and populations. Data from the new STR loci were combined with genotypes from previously published STR loci to assess the power to assign parentage using paternity exclusion in four species: fin whale (Balaenoptera physalus), humpback whale (Megaptera novaeangliae), blue whale (B. musculus) and bowhead whale (Balaena mysticetus). Our results suggest that parentage studies should always be accompanied by a power analysis in order to ascertain that each individual specific study is based upon data with sufficient power to assign parentage with statistical rigor