Neuroblastoma is a malignancy of the developing sympathetic nervous system that is often lethal when relapse occurs. We here used whole-exome sequencing, mRNA expression profiling, array CGH and DNA ...methylation analysis to characterize 16 paired samples at diagnosis and relapse from individuals with neuroblastoma. The mutational burden significantly increased in relapsing tumors, accompanied by altered mutational signatures and reduced subclonal heterogeneity. Global allele frequencies at relapse indicated clonal mutation selection during disease progression. Promoter methylation patterns were consistent over disease course and were patient specific. Recurrent alterations at relapse included mutations in the putative CHD5 neuroblastoma tumor suppressor, chromosome 9p losses, DOCK8 mutations, inactivating mutations in PTPN14 and a relapse-specific activity pattern for the PTPN14 target YAP. Recurrent new mutations in HRAS, KRAS and genes mediating cell-cell interaction in 13 of 16 relapse tumors indicate disturbances in signaling pathways mediating mesenchymal transition. Our data shed light on genetic alteration frequency, identity and evolution in neuroblastoma.
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DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SBMB, UILJ, UKNU, UL, UM, UPUK
•Carbon ions induced more severe DNA double-strand breaks than photons.•Carbon ion-induced double-strand breaks were repaired slower and less efficiently.•Carbon ions led to stronger cell cycle ...arrest and more apoptosis than photons.•PTEN−/− cells were more dependent on homologous recombination after carbon ions versus photons.
Carbon ion radiotherapy is a promising therapeutic option for glioblastoma patients due to its high physical dose conformity and greater biological effectiveness than photons. However, the biological effects of carbon ion radiation are still incompletely understood. Here, we systematically compared the biological effects of clinically used carbon ion radiation to photon radiation with emphasis on DNA repair.
Two human glioblastoma cell lines (U87 and LN229) were irradiated with carbon ions or photons and DNA damage response was systematically analyzed, including clonogenic survival, induction and repair of DNA double-strand breaks (DSBs), cell cycle arrest and apoptosis or autophagy. γH2AX foci were analyzed by flow cytometry, conventional light microscopy and 3D superresolution microscopy.
DSBs were repaired delayed and with slower kinetics after carbon ions versus photons. Carbon ions caused stronger and longer-lasting cell cycle delays, predominantly in G2 phase, and a higher rate of apoptosis. Compared to photons, the effectiveness of carbon ions was less cell cycle-dependent. Homologous recombination (HR) appeared to be more important for DSB repair after carbon ions versus photons in phosphatase and tensin homolog (PTEN)-deficient U87 cells, as opposed to PTEN-proficient LN229 cells.
Carbon ions induced more severe DSB damage than photons, which was repaired less efficiently in both cell lines. Thus, carbon ion radiotherapy may help to overcome resistance mechanisms of glioblastoma associated with DNA repair for example in combination with repair pathway-specific drugs in the context of personalized radiotherapy.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Pancreatic acinar cell carcinoma (ACC) is an aggressive exocrine tumor with largely unknown biology. Here, to identify potential targets for personalized treatment, we perform integrative genome-wide ...and epigenome-wide analyses. The results show frequently aberrant DNA methylation, abundant chromosomal amplifications and deletions, and mutational signatures suggesting defective DNA repair. In contrast to pancreatic ductal adenocarcinoma, no recurrent point mutations are detected. The tumor suppressors ID3, ARID1A, APC, and CDKN2A are frequently impaired also on the protein level and thus potentially affect ACC tumorigenesis. Consequently, this work identifies promising therapeutic targets in ACC for drugs recently approved for precision cancer therapy.
Drug resistance is a leading cause for treatment failure in many cancers, including neuroblastoma, the most common solid extracranial childhood malignancy. Previous studies from our lab indicate that ...histone deacetylase 10 (HDAC10) is important for the homeostasis of lysosomes, i.e. acidic vesicular organelles involved in the degradation of various biomolecules. Here, we show that depleting or inhibiting HDAC10 results in accumulation of lysosomes in chemotherapy-resistant neuroblastoma cell lines, as well as in the intracellular accumulation of the weakly basic chemotherapeutic doxorubicin within lysosomes. Interference with HDAC10 does not block doxorubicin efflux from cells via P-glycoprotein inhibition, but rather via inhibition of lysosomal exocytosis. In particular, intracellular doxorubicin does not remain trapped in lysosomes but also accumulates in the nucleus, where it promotes neuroblastoma cell death. Our data suggest that lysosomal exocytosis under doxorubicin treatment is important for cell survival and that inhibition of HDAC10 further induces DNA double-strand breaks (DSBs), providing additional mechanisms that sensitize neuroblastoma cells to doxorubicin. Taken together, we demonstrate that HDAC10 inhibition in combination with doxorubicin kills neuroblastoma, but not non-malignant cells, both by impeding drug efflux and enhancing DNA damage, providing a novel opportunity to target chemotherapy resistance.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Radiotherapy is an efficient tool in cancer treatment, but it brings along the risk of side effects such as fibrosis in the irradiated healthy tissue thus limiting tumor control and impairing quality ...of life of cancer survivors. Knowledge on radiation-related fibrosis risk and therapeutic options is still limited and requires further research. Recent studies demonstrated that epigenetic regulation of diacylglycerol kinase alpha (DGKA) is associated with radiation-induced fibrosis. However, the specific mechanisms are still unknown. In this review, we scrutinized the role of DGKA in the radiation response and in further cellular functions to show the potential of DGKA as a predictive marker or a novel target in fibrosis treatment. DGKA was reported to participate in immune response, lipid signaling, exosome production, and migration as well as cell proliferation, all processes which are suggested to be critical steps in fibrogenesis. Most of these functions are based on the conversion of diacylglycerol (DAG) to phosphatidic acid (PA) at plasma membranes, but DGKA might have also other, yet not well-known functions in the nucleus. Current evidence summarized here underlines that DGKA activation may play a central role in fibrosis formation post-irradiation and shows a potential of direct DGKA inhibitors or epigenetic modulators to attenuate pro-fibrotic reactions, thus providing novel therapeutic choices.
Radiotherapy is an important weapon in the treatment of cancer, but adverse reactions developing in the co-irradiated normal tissue can be a threat for patients. Early reactions might disturb the ...usual application schedule and limit the radiation dose. Late appearing and degenerative reactions might reduce or destroy normal tissue function. Genetic markers conferring the ability to identify hyper-sensitive patients in advance would considerably improve therapy. Association studies on genetic variation and occurrence of side effects should help to identify such markers. This survey includes published studies and novel data from our own laboratory. It illustrates the presence of candidate polymorphisms in genes involved in the cellular response to irradiation which could be used as predictive markers for radiosensitivity in breast or prostate cancer patients. For other tumor types such as head and neck cancers or brain tumors, the available data are much more limited. In any case, further validation of these markers is needed in large patient cohorts with systematically recorded data on side effects and patient characteristics. Genetic variation contributing to radiosensitivity should be screened on a broader basis using newly developed, more comprehensive approaches such as genome-wide association studies.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
Radiotherapy is a fundamental part of cancer treatment but its use is limited by the onset of late adverse effects in the normal tissue, especially radiation-induced fibrosis. Since the molecular ...causes for fibrosis are largely unknown, we analyse if epigenetic regulation might explain inter-individual differences in fibrosis risk. DNA methylation profiling of dermal fibroblasts obtained from breast cancer patients prior to irradiation identifies differences associated with fibrosis. One region is characterized as a differentially methylated enhancer of diacylglycerol kinase alpha (DGKA). Decreased DNA methylation at this enhancer enables recruitment of the profibrotic transcription factor early growth response 1 (EGR1) and facilitates radiation-induced DGKA transcription in cells from patients later developing fibrosis. Conversely, inhibition of DGKA has pronounced effects on diacylglycerol-mediated lipid homeostasis and reduces profibrotic fibroblast activation. Collectively, DGKA is an epigenetically deregulated kinase involved in radiation response and may serve as a marker and therapeutic target for personalized radiotherapy.
Proteoglycans are often overexpressed in tumors and can be found on several normal and neoplastic stem cells. In this study, we analyzed in‐depth the role of CSPG4 in head and neck squamous cell ...carcinomas (HNSCC). Analysis of CSPG4 in a homogeneous study sample of HPV‐negative stage IVa HNSCCs revealed overexpression of protein and mRNA levels in a subgroup of HNSCC tumors and a significant association of high CSPG4 protein levels with poor survival. This could be validated in three publicly available microarray datasets. As a potential cause for upregulated CSPG4 expression, we identified DNA hypomethylation in a CpG‐island of the promoter region. Accordingly, we found an inverse correlation of methylation and patient outcome. Finally, CSPG4 re‐expression was achieved by demethylating treatment of highly methylated HNSCC cell lines establishing a direct link between methylation and CSPG4 expression. In conclusion, we identified CSPG4 as a novel biomarker in HNSCC on several biological levels and established a causative link between DNA methylation and CSPG4 protein and mRNA expression.
What's New?
Head and neck squamous cell carcinoma (HNSCC) is a frequent cancer characterized by clinical and biological heterogeneity and poor disease outcome. Here, the authors examined the expression level of the stem cell‐associated proteoglycan CSPG4 as a potential novel biomarker in HNSCC. Studies were restricted to stage IVa tumors negative for human papilloma virus infection and showed increased expression of CSPG4 mRNA and protein expression accompanied by decreased promoter methylation. Promoter methylation was inversely correlated with disease outcome and could be reversed with demethylating agents, underscoring the therapeutic potential of CSPG4 targeting in the treatment of HNSCC patients.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
To identify single-nucleotide polymorphisms (SNPs) in oxidative stress-related genes associated with risk of late toxicities in breast cancer patients receiving radiation therapy.
Using a 2-stage ...design, 305 SNPs in 59 candidate genes were investigated in the discovery phase in 753 breast cancer patients from 2 prospective cohorts from Germany. The 10 most promising SNPs in 4 genes were evaluated in the replication phase in up to 1883 breast cancer patients from 6 cohorts identified through the Radiogenomics Consortium. Outcomes of interest were late skin toxicity and fibrosis of the breast, as well as an overall toxicity score (Standardized Total Average Toxicity). Multivariable logistic and linear regression models were used to assess associations between SNPs and late toxicity. A meta-analysis approach was used to summarize evidence.
The association of a genetic variant in the base excision repair gene XRCC1, rs2682585, with normal tissue late radiation toxicity was replicated in all tested studies. In the combined analysis of discovery and replication cohorts, carrying the rare allele was associated with a significantly lower risk of skin toxicities (multivariate odds ratio 0.77, 95% confidence interval 0.61-0.96, P=.02) and a decrease in Standardized Total Average Toxicity scores (-0.08, 95% confidence interval -0.15 to -0.02, P=.016).
Using a stage design with replication, we identified a variant allele in the base excision repair gene XRCC1 that could be used in combination with additional variants for developing a test to predict late toxicities after radiation therapy in breast cancer patients.
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GEOZS, IJS, NUK, OILJ, UL, UM, UPUK
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