Do non-pathogenic variants of DNA mismatch repair genes modify neurofibroma load in neurofibromatosis type 1?

Non-pathogenic mismatch repair (MMR) gene variants can be associated with decreased MMR capacity in several settings. Due to an increased mutation rate, reduced MMR capacity leads to accumulation of somatic sequence changes in tumour suppressor genes such as in the neurofibromatosis type 1 (NF1) gene. Patients with autosomal dominant NF1 typically develop neurofibromas ranging from single to thousands. Concerning the number of neurofibromas NF1 patients face a situation that is still not predictable. A few studies suggested that germline non-pathogenic MMR gene variants modify the number of neurofibromas in NF1 and by this mechanism may promote the extent of neurofibroma manifestation. This review represents first evidence that specific non-pathogenic single nucleotide variants of MMR genes act as a modifier of neurofibroma manifestation in NF1, highlighting MSH2 re4987188 as the best analysed non-pathogenic variant so far. In summary, besides MSH2 promotor methylation, specific non-pathogenic germline MSH2 variants are associated with the extent of neurofibroma manifestation. Those variants can serve as a biomarker to facilitate better mentoring of NF1 patients at risk.

Novel ratio-metric features enable the identification of new driver genes across cancer types

An emergent area of cancer genomics is the identification of driver genes. Driver genes confer a selective growth advantage to the cell. While several driver genes have been discovered, many remain undiscovered, especially those mutated at a low frequency across samples. This study defines new features and builds a pan-cancer model, cTaG, to identify new driver genes. The features capture the functional impact of the mutations as well as their recurrence across samples, which helps build a model unbiased to genes with low frequency. The model classifies genes into the functional categories of driver genes, tumour suppressor genes (TSGs) and oncogenes (OGs), having distinct mutation type profiles. We overcome overfitting and show that certain mutation types, such as nonsense mutations, are more important for classification. Further, cTaG was employed to identify tissue-specific driver genes. Some known cancer driver genes predicted by cTaG as TSGs with high probability are ARID1A, TP53, and RB1. In addition to these known genes, potential driver genes predicted are CD36, ZNF750 and ARHGAP35 as TSGs and TAB3 as an oncogene. Overall, our approach surmounts the issue of low recall and bias towards genes with high mutation rates and predicts potential new driver genes for further experimental screening. cTaG is available at https://github.com/RamanLab/cTaG.

Hypermethylation involved in DNA profiles of lung cancer specific tumour suppressor genes and epigenetic modification caused by an ultra-highly diluted homeopathic drug, Condurango 30C, in vitro and in vivo

Background and objectives: DNA hyper-methylation is an important aspect involved in carcinogenesis and cancer progression, which affects mainly CpG islands of DNA and causes inactivation of tumour suppressor genes. Therefore DNA hypermethylation status of the genomic DNA in both the transformed cancerous cell lines and in carcinogen-induced lung cancer was ascertained by analysis of expressions of certain major lung cancer specific tumour suppressor genes. The other objective was to examine if ultra highly diluted homeopathic drug, Condurango 30C, had ability to modulate DNA methylation. Methods: DNA methylation activity, if any, has been ascertained in H460-NSCLC cells in vitro and in BaP-induced lung cancer of rats in vivo, in respect of tumour suppressor genes like p15, p16, p18 and p53 by using PCR-SSCP analyses. The ability of modulation of DNA methylation, if any, by Condurango 30C was also verified against placebo control in a blinded manner. Results: Condurango 30C-treated DNA showed significant decrease in band-intensity of p15 and p53 genes especially in methylated condition, in vitro, at the IC50 dose (2.43µl/100µl). SSCP analysis of p15 and p53 genes in Condurango 30C-treated DNA also supported ability of Condurango 30C to modulate methylation state, in vitro. Inhibition of p15 hypermethylation was observed after post cancer treatment of rat with Condurango 30C. SSCP results gave a better indication of differences in band-position and single strand separation of p15 and p53 in Condurango 30C treated samples. Conclusion: Condurango 30C could trigger epigenetic modification in lung cancer via modulation of DNA hypermethylation but placebos could not.

Histone 3 Lysine 27 Trimethylation Signature in Breast Cancer

Cancer development and progression rely on complicated genetic and also epigenetic changes which regulate gene expression without altering the DNA sequence. Epigenetic mechanisms such as DNA methylation, histone modifications, and regulation by lncRNAs alter protein expression by either promoting gene transcription or repressing it. The presence of so-called chromatin modification marks at various gene promoters and gene bodies is associated with normal cell development but also with tumorigenesis and progression of different types of cancer, including the most frequently diagnosed breast cancer. This review is focused on the significance of one of the abundant post-translational modifications of histone 3- trimethylation of lysine 27 (H3K27me3), which was shown to participate in tumour suppressor genes’ silencing. Unlike other reviews in the field, here the overview of existing evidence linking H3K27me3 status with breast cancer biology and the tumour outcome is presented especially in the context of diverse breast cancer subtypes. Moreover, the potential of agents that target H3K27me3 for the treatment of this complex disease as well as H3K27 methylation in cross-talk with other chromatin modifications and lncRNAs are discussed.

Epigenetic Regulation of miR-92a and TET2 and Their Association in Non-Hodgkin Lymphoma

MicroRNAs (miRNAs) are well known for their ability to regulate the expression of specific target genes through degradation or inhibition of translation of the target mRNA. In various cancers, miRNAs regulate gene expression by altering the epigenetic status of candidate genes that are implicated in various difficult to treat haematological malignancies such as non-Hodgkin lymphoma by acting as either oncogenes or tumour suppressor genes. Cellular and circulating miRNA biomarkers could also be directly utilised as disease markers for diagnosis and monitoring of non-Hodgkin lymphoma (NHL); however, the role of DNA methylation in miRNA expression regulation in NHL requires further scientific inquiry. In this study, we investigated the methylation levels of CpGs in CpG islands spanning the promoter regions of the miR-17–92 cluster host gene and the TET2 gene and correlated them with the expression levels of TET2 mRNA and miR-92a-3p and miR-92a-5p mature miRNAs in NHL cell lines, tumour samples, and the whole blood gDNA of an NHL case control cohort. Increased expression of both miR-92a-3p and miR-92a-5p and aberrant expression of TET2 was observed in NHL cell lines and tumour tissues, as well as disparate levels of dysfunctional promoter CGI methylation. Both miR-92a and TET2 may play a concerted role in NHL malignancy and disease pathogenesis.

Aberrant integration of Hepatitis B virus DNA promotes major restructuring of human hepatocellular carcinoma genome architecture

Most cancers are characterized by the somatic acquisition of genomic rearrangements during tumour evolution that eventually drive the oncogenesis. Here, using multiplatform sequencing technologies, we identify and characterize a remarkable mutational mechanism in human hepatocellular carcinoma caused by Hepatitis B virus, by which DNA molecules from the virus are inserted into the tumour genome causing dramatic changes in its configuration, including non-homologous chromosomal fusions, dicentric chromosomes and megabase-size telomeric deletions. This aberrant mutational mechanism, present in at least 8% of all HCC tumours, can provide the driver rearrangements that a cancer clone requires to survive and grow, including loss of relevant tumour suppressor genes. Most of these events are clonal and occur early during liver cancer evolution. Real-time timing estimation reveals some HBV-mediated rearrangements occur as early as two decades before cancer diagnosis. Overall, these data underscore the importance of characterising liver cancer genomes for patterns of HBV integration.

Engraftment of allotransplantated tumour cells in adult rag2 mutant Xenopus tropicalis

Modelling human genetic diseases and cancer in lab animals has been greatly aided by the emergence of genetic engineering tools such as TALENs and CRISPR/Cas9. We have previously demonstrated the ease with which genetically engineered Xenopus models (GEXM) can be generated. This included the induction of autochthonous tumour formation by injection of early embryos with Cas9 recombinant protein loaded with sgRNAs targeting multiple tumour suppressor genes. What has been lacking so far is the possibility to propagate the induced cancers via transplantation. In this paper we describe the generation of a rag2-/- knock-out line in Xenopus tropicalis that is deficient in functional T- and B-cells. This line was validated by means of an allografting experiment with a primary tp53-/- donor tumour. In addition, we optimized available protocols for sub-lethal gamma irradiation of X. tropicalis froglets. Irradiated animals also allowed stable, albeit transient, engraftment of transplanted tp53-/- tumour cells. The novel X. tropicalis rag2-/- line and the irradiated wild type froglets will further expand the experimental toolbox in this diploid amphibian, and help to establish it as a versatile and relevant model for exploring human cancer.

Multidimensional Mutational Profiling of the Indian HNSCC Sub-Population Provides IRAK1, a Novel Driver Gene and Potential Druggable Target

Head and neck squamous cell carcinomas (HNSCC) include heterogeneous group of tumors, classified according to their anatomical site. It is the sixth most prevalent cancer globally. Among South Asian countries, India accounts for 40% of HNC malignancies with significant morbidity and mortality. In the present study, we have performed exome sequencing and analysis of 51 Head and Neck squamous cell carcinoma samples. Besides known mutations in the oncogenes and tumour suppressors, we have identified novel gene signatures differentiating buccal, alveolar, and tongue cancers. Around 50% of the patients showed mutation in tumour suppressor genes TP53 and TP63. Apart from the known mutations, we report novel mutations in the genes AKT1, SPECC1, and LRP1B, which are linked with tumour progression and patient survival. A highly curated process was developed to identify survival signatures. 36 survival-related genes were identified based on the correlation of functional impact of variants identified using exome-seq with gene expression from transcriptome data (GEPIA database) and survival. An independent LASSO regression analysis was also performed. Survival signatures common to both the methods led to identification of 4 dead and 3 alive gene signatures, the accuracy of which was confirmed by performing a ROC analysis (AUC=0.79 and 0.91, respectively). Also, machine learning-based driver gene prediction tool resulted in the identification of IRAK1 as the driver (p-value = 9.7 e-08) and also as an actionable mutation. Modelling of the IRAK1 mutation showed a decrease in its binding to known IRAK1 inhibitors.

P.138 A systematic review of the incidence and prevalence of Neurofibromatosis type 1 and 2

Background: Neurofibromatosis 1 and 2 (NF1 and NF2) are autosomal dominant genetic disorders caused by mutations in tumour suppressor genes. Methods: We conducted a systematic review of the incidence and prevalence of NF1 and NF2 in OVID Medline, OVID Embase, Web of Science, and Cinahl. We included studies until February 19, 2021, that identified cases based on established criteria. Studies were appraised for quality using the Joanna Briggs Institute Prevalence Critical Appraisal tool. Pooled incidence and prevalence rates were estimated through meta-analysis. Results: Of 1,936 studies, 1,866 were irrelevant after title and abstract screening. Sixteen of 69 studies with full text assessment were included for full review: 13 regarding NF1 and 6 regarding NF2. Incidence rates for NF1 and NF2 ranged from 1/11,494 to 1/1,871 and 1/62,185 to 1/33,000 respectively. Prevalence rates for NF1 and NF2 ranged from 1/6,238 to 1/1,001 and 1/600,000 to 1/56,161 respectively. Meta-analysis will be presented at the conference. Conclusions: An accurate estimate of the incidence and prevalence of NF1 and NF2 will offer more insight into health resource allocation. Increased funding and resources for the development of early diagnostic and treatment tools for NF1 and NF2 may improve the quality of life of patients.