The prognosis of metastatic non-small cell lung cancer (NSCLC) is still poor. Activating epithelial growth factor receptor (EGFR) mutations are important genetic alterations with dramatic therapeutical implications. Up to now, in contrast to Asian populations only limited data on the prevalence of those mutations are available from patients with Caucasian and especially European ethnicity. In this multicentre study, 1201 unselected NSCLC patients from Southern Germany were tested in the daily clinical routine for EGFR mutation status. Activating EGFR mutations were found in 9.8% of all tumours. Mutations in exons 18, 19 and 21 accounted for 4.2%, 61.9% and 33.1% of all mutations, respectively. Non-smokers had a significantly higher rate of EGFR mutations than smokers or ex-smokers (24.4% vs 4.2%; P<0.001). Non-lepidic-non-mucinous adenocarcinomas (G2) accounted for 45.5% of all activating EGFR mutations and 3.5% of all squamous cell carcinomas were tested positive. Thyroid transcription factor 1 protein expression was significantly associated with EGFR mutational status. These comprehensive data from clinical routine in Germany add to the knowledge of clinical and histopathological factors associated with EGFR mutational status in NSCLC.

Aberrant DNA methylation at CpG islands is thought to contribute to cancer initiation and progression, but mechanisms that establish and maintain DNA methylation status during tumorigenesis or normal development remain poorly understood. In this study, we used methyl-CpG immunoprecipitation to generate comparative DNA methylation profiles of healthy and malignant cells (acute leukemia and colorectal carcinoma) for human CpG islands across the genome. While searching for sequence patterns that characterize DNA methylation states, we discovered several nonredundant sequences in CpG islands that were resistant to aberrant de novo methylation in cancer and that resembled consensus binding sites for general transcription factors (TF). Comparing methylation profiles with global CpG island binding data for specific protein 1, nuclear respiratory factor 1, and yin-yang 1 revealed that their DNA binding activity in normal blood cells correlated strictly with an absence of de novo methylation in cancer. In addition, global evidence showed that binding of any of these TFs to their consensus motif depended on their co-occurrence with neighboring consensus motifs. In summary, our results had two major implications. First, they pointed to a major role for cooperative binding of TFs in maintaining the unmethylated status of CpG islands in health and disease. Second, our results suggest that the majority of de novo methylated CpG islands are characterized by the lack of sequence motif combinations and the absence of activating TF binding. Cancer Res; 70(4); 1398–407

Microsatellite instability (MSI), a common alteration in endometrial cancers (EC) is known as a biomarker for immune checkpoint therapy response alongside screening for Lynch Syndrome (LS). However, former studies described challenging MSI profiles in EC hindering analysis by using MSI testing methods intensively validated for colorectal cancer (CRC) only. In order to reduce false negatives, this study examined four different PCR-based approaches for MSI testing using 25 EC samples already tested for mismatch repair deficiency (dMMR). In a follow up validation set of 75 EC samples previously tested both for MMR and MSI, the efficiency of a seven-marker system corresponding to the Idylla system was further analyzed. Both Bethesda and Promega marker panels require trained operators to overcome interpretation complexities caused by either hardly visible additional peaks of one and two nucleotides, or small shifts in microsatellite repeat length. Using parallel sequencing adjustment of bioinformatics is needed. Applying the Idylla MSI assay, an evaluation of input material is more crucial for reliable results and is indispensable. Following MMR deficiency testing as a first-line screening procedure, additional testing with a PCR-based method is necessary if inconclusive staining of immunohistochemistry (IHC) must be clarified.

Gene fusions involving the three neurotrophic tyrosine receptor kinase genesNTRK1,NTRK2, orNTRK3were identified as oncogenic drivers in many cancer types. Two small molecule inhibitors have been tested in clinical trials recently and require the detection of aNTRKfusion gene prior to therapeutic application. Fluorescence in situ hybridization (FISH) and targeted next‐generation sequencing (tNGS) assays are commonly used for diagnostic profiling of gene fusions. In the presented study we applied an external quality assessment (EQA) scheme in order to investigate the suitability of FISH and RNA‐/DNA‐based tNGS for detection ofNTRKfusions in a multinational and multicentric ring trial. In total 27 participants registered for this study. Nine institutions took part in the FISH‐based and 18 in the NGS‐based round robin test, the latter additionally subdivided into low‐input and high‐input NGS methods (regarding nucleic acid input). Regardless of the testing method applied, all participants received tumor sections of 10 formalin‐fixed and paraffin‐embedded (FFPE) tissue blocks for in situ hybridization or RNA/DNA extraction, and the results were submitted via an online questionnaire. For FISH testing, eight of nine (88.8%) participants, and for NGS‐based testing 15 of 18 (83.3%) participants accomplished the round robin test successfully. The overall high success rate demonstrates that FISH‐ and tNGS‐basedNTRKtesting can be well established in a routine diagnostic setting. Complementing this dataset, we provide an updated in silico analysis on the coverage of more than 150NTRKfusion variants by several commercially available RNA‐based tNGS panels.

(1) Background: molecular tumor boards (MTBs) are crucial instruments for discussing and allocating targeted therapies to suitable cancer patients based on genetic findings. Currently, limited evidence is available regarding the regional impact and the outreach component of MTBs; (2) Methods: we analyzed MTB patient data from four neighboring Bavarian tertiary care oncology centers in Würzburg, Erlangen, Regensburg, and Augsburg, together constituting the WERA Alliance. Absolute patient numbers and regional distribution across the WERA-wide catchment area were weighted with local population densities; (3) Results: the highest MTB patient numbers were found close to the four cancer centers. However, peaks in absolute patient numbers were also detected in more distant and rural areas. Moreover, weighting absolute numbers with local population density allowed for identifying so-called white spots—regions within our catchment that were relatively underrepresented in WERA MTBs; (4) Conclusions: investigating patient data from four neighboring cancer centers, we comprehensively assessed the regional impact of our MTBs. The results confirmed the success of existing collaborative structures with our regional partners. Additionally, our results help identifying potential white spots in providing precision oncology and help establishing a joint WERA-wide outreach strategy.