[Show abstract][Hide abstract] ABSTRACT: This article presents selected methodological pitfalls of clinical studies in oncology, which in the authors‘ experience may jeopardize the validity of the trial. We discuss among others the suitability of progression-free survival (PFS) as a primary endpoint and issues to be taken into account when using PFS. Other topics of discussion encompass surveillance of the quality of carrying out the trial, the transparency of documentation involving predefined quality endpoints, sources of errors in sample size calculation leading to target numbers of events which are unachievable in the actual trial and the problem of a lack of funding opportunities for trials without commercial interests.
[Show abstract][Hide abstract] ABSTRACT: Systems biology approaches for mutational (exome analysis and targeted sequencing) and gene expression analysis (transcriptome-wide gene expression profiling) represent a new and growing scientific field in head and neck oncology. In addition to medical biological expertise, bioinformatic assistance is increasingly required. For squamous cell head and neck cancer (HNSCC), the recent molecular genetic single-gene and signal pathway observations represent basic research. Important aspects of this have now been significantly enhanced by systems biology approaches, which have grown into relevant areas of translational clinical research. It is now known that HPV16 is associated with genetic alterations at various locations, but also that it functionally affects genes not altered in their base sequence at the level of methylation. In transcriptome analyses, various consortia found matching clusters of gene expression and HPV16 association with the spectrum of somatic mutations. The differential methylation of gene promoters discovered in HPV16-driven HNSCC proved predictive for survival-even in HNSCC patients without HPV detection. The authors present an overview of some translationally relevant findings and venture an outlook on possible future clinical developments.
[Show abstract][Hide abstract] ABSTRACT: Although Burkitt lymphomas and follicular lymphomas both have features of germinal center B cells, they are biologically and clinically quite distinct. Here we performed whole-genome bisulfite, genome and transcriptome sequencing in 13 IG-MYC translocation-positive Burkitt lymphoma, nine BCL2 translocation-positive follicular lymphoma and four normal germinal center B cell samples. Comparison of Burkitt and follicular lymphoma samples showed differential methylation of intragenic regions that strongly correlated with expression of associated genes, for example, genes active in germinal center dark-zone and light-zone B cells. Integrative pathway analyses of regions differentially methylated in Burkitt and follicular lymphomas implicated DNA methylation as cooperating with somatic mutation of sphingosine phosphate signaling, as well as the TCF3-ID3 and SWI/SNF complexes, in a large fraction of Burkitt lymphomas. Taken together, our results demonstrate a tight connection between somatic mutation, DNA methylation and transcriptional control in key B cell pathways deregulated differentially in Burkitt lymphoma and other germinal center B cell lymphomas.
[Show abstract][Hide abstract] ABSTRACT: Despite the established role of the transcription factor MYC in cancer, little is known about the impact of a new class of transcriptional regulators, the long noncoding RNAs (lncRNAs), on MYC ability to influence the cellular transcriptome. Here, we have intersected RNA-sequencing data from two MYC-inducible cell lines and a cohort of 91 B-cell lymphomas with or without genetic variants resulting in MYC overexpression. We identified 13 lncRNAs differentially expressed in IG-MYC-positive Burkitt lymphoma and regulated in the same direction by MYC in the model cell lines. Among them, we focused on a lncRNA that we named MYC-induced long noncoding RNA (MINCR), showing a strong correlation with MYC expression in MYC-positive lymphomas. To understand its cellular role, we performed RNAi and found that MINCR knockdown is associated with an impairment in cell cycle progression. Differential gene expression analysis after RNAi showed a significant enrichment of cell cycle genes among the genes down-regulated after MINCR knockdown. Interestingly, these genes are enriched in MYC binding sites in their promoters, suggesting that MINCR acts as a modulator of the MYC transcriptional program. Accordingly, MINCR knockdown was associated with a reduction in MYC binding to the promoters of selected cell cycle genes. Finally, we show that down-regulation of Aurora kinases A and B and chromatin licensing and DNA replication factor 1 may explain the reduction in cellular proliferation observed on MINCR knockdown. We, therefore, suggest that MINCR is a newly identified player in the MYC transcriptional network able to control the expression of cell cycle genes.
No preview · Article · Sep 2015 · Proceedings of the National Academy of Sciences
[Show abstract][Hide abstract] ABSTRACT: The Three-Factor-Eating-Questionnaire (TFEQ) is an established instrument to assess eating behaviour. Analysis of the TFEQ-factor structure was based on selected, convenient and clinical samples so far. Aims of this study were (I) to analyse the factor structure of the German version of the TFEQ and (II)-based on the refined factor structure-to examine the association between eating behaviour and the body mass index (BMI) in a general population sample of 3,144 middle-aged and older participants (40-79 years) of the ongoing population based cohort study of the Leipzig Research Center for Civilization Diseases (LIFE Health Study). The factor structure was examined in a split-half analysis with both explorative and confirmatory factor analysis. Associations between TFEQ-scores and BMI values were tested with multiple regression analyses controlled for age, gender, and education. We found a three factor solution for the TFEQ with an 'uncontrolled eating', a 'cognitive restraint' and an 'emotional eating' domain including 29 of the original 51 TFEQ-items. Scores of the 'uncontrolled eating domain' showed the strongest correlation with BMI values (partial r = 0.26). Subjects with scores above the median in both 'uncontrolled eating' and 'emotional eating' showed the highest BMI values (mean = 29.41 kg/m²), subjects with scores below the median in all three domains showed the lowest BMI values (mean = 25.68 kg/m²; F = 72.074, p<0.001). Our findings suggest that the TFEQ is suitable to identify subjects with specific patterns of eating behaviour that are associated with higher BMI values. Such information may help health care professionals to develop and implement more tailored interventions for overweight and obese individuals.
[Show abstract][Hide abstract] ABSTRACT: Background
The LIFE-Adult-Study is a population-based cohort study, which has recently completed the baseline examination of 10,000 randomly selected participants from Leipzig, a major city with 550,000 inhabitants in the east of Germany. It is the first study of this kind and size in an urban population in the eastern part of Germany. The study is conducted by the Leipzig Research Centre for Civilization Diseases (LIFE). Our objective is to investigate prevalences, early onset markers, genetic predispositions, and the role of lifestyle factors of major civilization diseases, with primary focus on metabolic and vascular diseases, heart function, cognitive impairment, brain function, depression, sleep disorders and vigilance dysregulation, retinal and optic nerve degeneration, and allergies.
The study covers a main age range from 40-79 years with particular deep phenotyping in elderly participants above the age of 60. The baseline examination was conducted from August 2011 to November 2014. All participants underwent an extensive core assessment programme (5-6 h) including structured interviews, questionnaires, physical examinations, and biospecimen collection. Participants over 60 underwent two additional assessment programmes (3-4 h each) on two separate visits including deeper cognitive testing, brain magnetic resonance imaging, diagnostic interviews for depression, and electroencephalography.
The participation rate was 33 %. The assessment programme was accepted well and completely passed by almost all participants. Biomarker analyses have already been performed in all participants. Genotype, transcriptome and metabolome analyses have been conducted in subgroups. The first follow-up examination will commence in 2016.
Full-text · Article · Jul 2015 · BMC Public Health
[Show abstract][Hide abstract] ABSTRACT: Microsatellite instability (MSI) is detected in approximately 15% of all colorectal cancers (CRC) and virtually in all cases with Lynch syndrome. The MSI phenotype is caused by dysfunctional mismatch repair (MMR) and leads to accumulation of DNA replication errors. Sporadic MSI CRC often harbours BRAF(V600E); however, no consistent data exist regarding targeted treatment approaches in BRAF(wt) MSI CRC.
Mutations and quantitative MSI were analysed by deep sequencing in 196 formalin fixed paraffin embedded (FFPE) specimens comprising Lynch and Lynch-like CRCs from the German Hereditary Nonpolyposis Colorectal Cancer registry. Functional relevance of recurrent ERBB2/HER2 mutations was investigated in CRC cell lines using reversible and irreversible HER-targeting inhibitors, EGFR-directed antibody cetuximab, HER2-directed antibody trastuzumab and siRNA-mediated ERBB2/HER2 knockdown.
Quantification of nucleotide loss in non-coding mononucleotide repeats distinguished microsatellite status with very high accuracy (area under curve=0.9998) and demonstrated progressive losses with deeper invasion of MMR-deficient colorectal neoplasms (p=0.008). Characterisation of BRAF(wt) MSI CRC revealed hot-spot mutations in well-known oncogenic drivers, including KRAS (38.7%), PIK3CA (36.5%), and ERBB2 (15.0%). L755S and V842I substitutions in ERBB2 were highly recurrent. Functional analyses in ERBB2-mutated MSI CRC cell lines revealed a differential response to HER-targeting compounds and superiority of irreversible pan-HER inhibitors.
We developed a high-throughput deep sequencing approach for concomitant MSI and mutational analyses in FFPE specimens. We provided novel insights into clinically relevant alterations in MSI CRC and a rationale for targeting ERBB2/HER2 mutations in Lynch and Lynch-like CRC.
Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.
[Show abstract][Hide abstract] ABSTRACT: Cerebral gliomas of World Health Organization (WHO) grade II and III represent a major challenge in terms of histological classification and clinical management. Here, we asked whether large-scale genomic and transcriptomic profiling improves the definition of prognostically distinct entities. We performed microarray-based genome- and transcriptome-wide analyses of primary tumor samples from a prospective German Glioma Network cohort of 137 patients with cerebral gliomas, including 61 WHO grade II and 76 WHO grade III tumors. Integrative bioinformatic analyses were employed to define molecular subgroups, which were then related to histology, molecular biomarkers, including isocitrate dehydrogenase 1 or 2 (IDH1/2) mutation, 1p/19q co-deletion and telomerase reverse transcriptase (TERT) promoter mutations, and patient outcome. Genomic profiling identified five distinct glioma groups, including three IDH1/2 mutant and two IDH1/2 wild-type groups. Expression profiling revealed evidence for eight transcriptionally different groups (five IDH1/2 mutant, three IDH1/2 wild type), which were only partially linked to the genomic groups. Correlation of DNA-based molecular stratification with clinical outcome allowed to define three major prognostic groups with characteristic genomic aberrations. The best prognosis was found in patients with IDH1/2 mutant and 1p/19q co-deleted tumors. Patients with IDH1/2 wild-type gliomas and glioblastoma-like genomic alterations, including gain on chromosome arm 7q (+7q), loss on chromosome arm 10q (-10q), TERT promoter mutation and oncogene amplification, displayed the worst outcome. Intermediate survival was seen in patients with IDH1/2 mutant, but 1p/19q intact, mostly astrocytic gliomas, and in patients with IDH1/2 wild-type gliomas lacking the +7q/-10q genotype and TERT promoter mutation. This molecular subgrouping stratified patients into prognostically distinct groups better than histological classification. Addition of gene expression data to this genomic classifier did not further improve prognostic stratification. In summary, DNA-based molecular profiling of WHO grade II and III gliomas distinguishes biologically distinct tumor groups and provides prognostically relevant information beyond histological classification as well as IDH1/2 mutation and 1p/19q co-deletion status.
Full-text · Article · Mar 2015 · Acta Neuropathologica
[Show abstract][Hide abstract] ABSTRACT: Prognostically-relevant risk factors in patients with diffuse large B-cell lymphoma (DLBCL) have predominantly been evaluated in elderly populations. We tested whether previously described risk factors are also valid in younger, poor-prognosis DLBCL patients. Paraffin-embedded samples from 112 patients with de novo DLBCL, enrolled in the R-MegaCHOEP trial of the German High Grade Non-Hodgkin Lymphoma Study Group (DSHNHL) were investigated using immunohistochemistry (MYC, FOXP1, LMO2, GCET1, CD5, CD10, BCL2, BCL6, IRF4/MUM1) and fluorescence in situ hybridization (MYC, BCL2, BCL6). MYC, BCL2 and BCL6 breaks occurred in 14%, 21% and 31%, respectively. In the majority of cases, MYC was simultaneously rearranged with BCL2 and/or BCL6. The adverse impact of MYC rearrangements was confirmed, but the sole presence of BCL2 breaks emerged as a novel prognostic marker associated with inferior overall survival (OS) (p=0.002). Combined over-expression of MYC and BCL2 showed only limited association with inferior OS. All immunohistochemical cell of origin (COO) classifiers applied failed to predict survival time. DLBCL tumors with significant proportion of immunoblastic and/or immunoblastic-plasmacytoid cells had inferior OS, independently from from BCL2 break. Younger, poor-prognosis DLBCL patients, therefore, display different biological risk factors compared to an elderly population, with BCL2 translocations emerging as a powerful negative prognostic marker.Leukemia accepted article preview online, 17 February 2015. doi:10.1038/leu.2015.43.
[Show abstract][Hide abstract] ABSTRACT: Background
Although the growth-factor G-CSF is widely used to prevent granulotoxic side effects of cytotoxic chemotherapies, its optimal use is still unknown since treatment outcome depends on many parameters such as dosing and timing of chemotherapies, pharmaceutical derivative of G-CSF used and individual risk factors. We showed in the past that a pharmacokinetic and ¿dynamic model of G-CSF and human granulopoiesis can be used to predict the performance of yet untested G-CSF schedules. However, only a single chemotherapy was considered so far.In the present paper, we propose a comprehensive model of chemotherapy toxicity and combine it with our cell kinetic model of granulopoiesis. Major assumptions are: proportionality of cell numbers and cell loss, delayed action of chemotherapy, drug, drug-dose and cell stage specific toxicities, no interaction of drugs and higher toxicity of drugs at the first time of application. Correspondingly, chemotherapies can be characterized by a set of toxicity parameters which can be estimated by fitting the predictions of our model to clinical time series data of patients under therapy. Data were either extracted from the literature or were received from cooperating clinical study groups.ResultsModel assumptions proved to be feasible in explaining granulotoxicity of 10 different chemotherapeutic drugs or drug-combinations applied in 33 different schedules with and without G-CSF. Risk groups of granulotoxicity were traced back to differences in toxicity parameters.Conclusion
We established a comprehensive model of combined G-CSF and chemotherapy action in humans which allows us to predict and compare the outcome of alternative G-CSF schedules. We aim to apply the model in different clinical contexts to optimize and individualize G-CSF treatment.
Preview · Article · Dec 2014 · BMC Systems Biology
[Show abstract][Hide abstract] ABSTRACT: Objective:
To test for a possible effect of the apolipoprotein E epsilon 4 (APOE ε4) allele on memory performance and executive functioning (EF) in cognitively intact elderly.
The authors studied 202 randomly selected and cognitively intact older adults (65+ years) of the Leipzig Research Center for Civilization Diseases Health Care Study. Intact global cognitive functioning was defined using a Mini-Mental Status Examination (MMSE) score ≥ 28. Performance in memory was assessed with the CERAD Word List and Constructional Praxis Recall, performance in EF with the Trail Making Test Part B (TMT-B). Multivariable linear regressions were used to evaluate the association between cognitive performance and APOE status, controlled for covariates.
Among the cognitively intact older adults, 21.3% (n = 43) were carriers of the APOE ε4 allele. Carriers did not differ significantly from noncarriers in terms of age, gender, intelligence level, or performance in memory but showed a significantly lower TMT-B performance as a measure of EF (TMT-B M time/SD = 105.6/36.2 vs. 91.9/32.7 s; Mann-Whitney U = 4,313.000; p = .009). The association between lower TMT-B performance and APOE ε4 genotype remained significant in multivariable linear regression analysis. Similar findings were found for the subsample of those 78 elderly, who reached a perfect MMSE-score of 30.
A lower EF performance in cognitively intact older APOE ε4 allele carriers might be related to an early Alzheimer's dementia (AD) prodrome. In this case, a stronger focus on first subtle changes in EF may help to improve early AD detection in those being at genetic risk.
[Show abstract][Hide abstract] ABSTRACT: BACKGROUND: A relevant fraction of patients does not receive salvage therapy at recurrence of glioblastoma initially treated
according to current standards of care. Primary treatment may be of higher relevance in this patient group. Risk factors to
prospectively determine which patients have a higher probability to receive salvage therapy have not been prospectively studied.
METHODS: Patients (n = 1004) with first recurrence of a glioblastoma received no salvage treatment (n = 374; 37.3%, group
N) or ≥1 line of postprogression therapy consisting of surgical resection +/- second radio- and/or chemotherapy (n = 630;
62.7%, group Y) in the prospectively assembled cohort of the German Glioma Network. Age, gender, Karnofsky Performance Status
(KPS), extent of resection, O6-methylguanine methyltransferase (MGMT) and isocitrate dehydrogenase (IDH) 1/2 status as well
as first line therapy were assessed as risk factors not to receive salvage therapy. RESULTS: Median overall survival in the
whole cohort was 14.2 (95% confidence interval 13.4-15.0) months. It was 8.7 (95% CI 8.1-9.2) months in group N as opposed
to 18.5 (95% CI 17.2-19.8) months in group Y patients. Positive prognostic factors to receive salvage treatment were age ≤
60 years (odds ratio (OR), 2.9, p < 0.001), complete resection at first-line treatment (OR 1.8, p = 0.011) and KPS >70 (OR
1.7, p = 0.035), but not initial treatment with radiochemotherapy (versus radio- or chemotherapy) (OR 1.6, p = 0.051), gender
(OR 1.1, p = 0.718), or MGMT (OR 1.5, p = 0.087). A multivariate analysis with test and validation set will be presented.
CONCLUSION: In this analysis, the administration of salvage treatment expectedly was associated with prolonged survival. Older
patients with incomplete resections or biopsies with lower KPS may particularly benefit from improved first-line treatment
since these patients are less likely to receive salvage therapy. A concept of the EORTC and NOA proposes to assess the addition
of bevacizumab to such patients with tumors lacking MGMT promoter methylation.