BIODICA: a computational environment for Independent Component Analysis of omics data.

SUMMARY: We developed BIODICA, an integrated computational environment for application of independent component analysis (ICA) to bulk and single-cell molecular profiles, interpretation of the results in terms of biological functions and correlation with metadata. The computational core is the novel Python package stabilized-ica which provides interface to several ICA algorithms, a stabilization procedure, meta-analysis and component interpretation tools. BIODICA is equipped with a user-friendly graphical user interface, allowing non-experienced users to perform the ICA-based omics data analysis. The results are provided in interactive ways, thus facilitating communication with biology experts. AVAILABILITY AND IMPLEMENTATION: BIODICA is implemented in Java, Python and JavaScript. The source code is freely available on GitHub under the MIT and the GNU LGPL licenses. BIODICA is supported on all major operating systems. URL: https://sysbio-curie.github.io/biodica-environment/.

Glioma oncogenesis in the Constitutional mismatch repair deficiency (CMMRD) syndrome.

Background: Constitutional mismatch repair deficiency (CMMRD) is a cancer predisposition due to biallelic mutations in one of the mismatch repair (MMR) genes associated with early onset of cancers, especially high-grade gliomas. Our aim was to decipher the molecular specificities of these gliomas. Methods: Clinical, histopathological, and whole exome sequencing data were analyzed in 12 children with genetically proven CMMRD and a high-grade glioma. Results: PDL1 expression was present in immunohistochemistry in 50% of the samples. In 9 patients, the glioma harbored an ultra-hypermutated phenotype (104-635 coding single nucleotide variants (SNV) per Mb, median 204). Driver mutations in POLE and POLD1 exonuclease domains were described for 8 and 1 patients respectively and were always present in the mutation burst with the highest variant allele frequency (VAF). The mutational signatures were dominated by MMR-related ones and similar in the different mutation bursts of a same patient without subsequent enrichment of the mutation signatures with POL-driven ones. Median number of coding SNV with VAF above one of the driving polymerase mutation per Mb was 57 (17-191). Our findings suggest that somatic polymerase alterations does not entirely explain the ultra-hypermutant phenotype. SETD2, TP53, NF1, EPHB2, PRKDC, and DICER1 genes were frequently mutated with higher VAF than the deleterious somatic polymerase mutation. Conclusions: CMMRD-associated gliomas have a specific oncogenesis that does not involve usual pathways and mutations seen in sporadic pediatric or adult glioblastomas. Frequent alterations in other pathways such as MAPK may suggest the use of other targeted therapies along with PD1 inhibitors.

Systems Biology Analysis for Ewing Sarcoma.

Ewing sarcoma (EwS) is a highly aggressive pediatric bone cancer that is defined by a somatic fusion between the EWSR1 gene and an ETS family member, most frequently the FLI1 gene, leading to expression of a chimeric transcription factor EWSR1-FLI1. Otherwise, EwS is one of the most genetically stable cancers. The situation when the major cancer driver is well known looks like a unique opportunity for applying the systems biology approach in order to understand the EwS mechanisms as well as to uncover some general mechanistic principles of carcinogenesis. A number of studies have been performed revealing the direct and indirect effects of EWSR1-FLI1 on multiple aspects of cellular life. Nevertheless, the emerging picture of the oncogene action appears to be highly complex and systemic, with multiple reciprocal influences between the immediate consequences of the driver mutation and intracellular and intercellular molecular mechanisms, including regulation of transcription, epigenome, and tumoral microenvironment. In this chapter, we present an overview of existing molecular profiling resources available for EwS tumors and cell lines and provide an online comprehensive catalogue of publicly available omics and other datasets. We further highlight the systems biology studies of EwS, involving mathematical modeling of networks and integration of molecular data. We conclude that despite the seeming simplicity, a lot has yet to be understood on the systems-wide mechanisms connecting the driver mutation and the major cellular phenotypes of this pediatric cancer. Overall, this chapter can serve as a guide for a systems biology researcher to start working on EwS.

A kinome-wide shRNA screen uncovers vaccinia-related kinase 3 (VRK3) as an essential gene for diffuse intrinsic pontine glioma survival.

Diffuse intrinsic pontine glioma (or DIPG) are pediatric high-grade gliomas associated with a dismal prognosis. They harbor specific substitution in histone H3 at position K27 that induces major epigenetic dysregulations. Most clinical trials failed so far to increase survival, and radiotherapy remains the most efficient treatment, despite only transient tumor control. We conducted the first lentiviral shRNA dropout screen in newly diagnosed DIPG to generate a cancer-lethal signature as a basis for the development of specific treatments with increased efficacy and reduced side effects compared to existing anticancer therapies. The analysis uncovered 41 DIPG essential genes among the 672 genes of human kinases tested, for which several distinct interfering RNAs impaired cell expansion of three different DIPG stem-cell cultures without deleterious effect on two control neural stem cells. Among them, PLK1, AURKB, CHEK1, EGFR, and GSK3A were previously identified by similar approach in adult GBM indicating common dependencies of these cancer cells and pediatric gliomas. As expected, we observed an enrichment of genes involved in proliferation and cell death processes with a significant number of candidates belonging to PTEN/PI3K/AKT and EGFR pathways already under scrutiny in clinical trials in this disease. We highlighted VRK3, a gene involved especially in cell cycle regulation, DNA repair, and neuronal differentiation, as a non-oncogenic addiction in DIPG. Its repression totally blocked DIPG cell growth in the four cellular models evaluated, and induced cell death in H3.3-K27M cells specifically but not in H3.1-K27M cells, supporting VRK3 as an interesting and promising target in DIPG.

TP53 Pathway Alterations Drive Radioresistance in Diffuse Intrinsic Pontine Gliomas (DIPG).

PURPOSE: Diffuse intrinsic pontine gliomas (DIPG) are the most severe pediatric brain tumors. Although accepted as the standard therapeutic, radiotherapy is only efficient transiently and not even in every patient. The goal of the study was to identify the underlying molecular determinants of response to radiotherapy in DIPG. EXPERIMENTAL DESIGN: We assessed in vitro response to ionizing radiations in 13 different DIPG cellular models derived from treatment-naïve stereotactic biopsies reflecting the genotype variability encountered in patients at diagnosis and correlated it to their principal molecular alterations. Clinical and radiologic response to radiotherapy of a large cohort of 73 DIPG was analyzed according to their genotype. Using a kinome-wide synthetic lethality RNAi screen, we further identified target genes that can sensitize DIPG cells to ionizing radiations. RESULTS: We uncover TP53 mutation as the main driver of increased radioresistance and validated this finding in four isogenic pairs of TP53WT DIPG cells with or without TP53 knockdown. In an integrated clinical, radiological, and molecular study, we show that TP53MUT DIPG patients respond less to irradiation, relapse earlier after radiotherapy, and have a worse prognosis than their TP53WT counterparts. Finally, a kinome-wide synthetic lethality RNAi screen identifies CHK1 as a potential target, whose inhibition increases response to radiation specifically in TP53MUT cells. CONCLUSIONS: Here, we demonstrate that TP53 mutations are driving DIPG radioresistance both in patients and corresponding cellular models. We suggest alternative treatment strategies to mitigate radioresistance with CHK1 inhibitors. These findings will allow to consequently refine radiotherapy schedules in DIPG.

Transcriptomic and epigenetic profiling of 'diffuse midline gliomas, H3 K27M-mutant' discriminate two subgroups based on the type of histone H3 mutated and not supratentorial or infratentorial location.

Diffuse midline glioma (DMG), H3 K27M-mutant, is a new entity in the updated WHO classification grouping together diffuse intrinsic pontine gliomas and infiltrating glial neoplasms of the midline harboring the same canonical mutation at the Lysine 27 of the histones H3 tail.Two hundred and fifteen patients younger than 18 years old with centrally-reviewed pediatric high-grade gliomas (pHGG) were included in this study. Comprehensive transcriptomic (n = 140) and methylation (n = 80) profiling was performed depending on the material available, in order to assess the biological uniqueness of this new entity compared to other midline and hemispheric pHGG.Tumor classification based on gene expression (GE) data highlighted the similarity of K27M DMG independently of their location along the midline. T-distributed Stochastic Neighbor Embedding (tSNE) analysis of methylation profiling confirms the discrimination of DMG from other well defined supratentorial tumor subgroups. Patients with diffuse intrinsic pontine gliomas (DIPG) and thalamic DMG exhibited a similarly poor prognosis (11.1 and 10.8 months median overall survival, respectively). Interestingly, H3.1-K27M and H3.3-K27M primary tumor samples could be distinguished based both on their GE and DNA methylation profiles, suggesting that they might arise from a different precursor or from a different epigenetic reorganization.These differences in DNA methylation profiles were conserved in glioma stem-like cell culture models of DIPG which mimicked their corresponding primary tumor. ChIP-seq profiling of H3K27me3 in these models indicate that H3.3-K27M mutated DIPG stem cells exhibit higher levels of H3K27 trimethylation which are correlated with fewer genes expressed by RNAseq. When considering the global distribution of the H3K27me3 mark, we observed that intergenic regions were more trimethylated in the H3.3-K27M mutated cells compared to the H3.1-K27M mutated ones.H3 K27M-mutant DMG represent a homogenous group of neoplasms compared to other pediatric gliomas that could be further separated based on the type of histone H3 variant mutated and their respective epigenetic landscapes. As these characteristics drive different phenotypes, these findings may have important implication for the design of future trials in these specific types of neoplasms.

Mutation allele burden remains unchanged in chronic myelomonocytic leukaemia responding to hypomethylating agents.

The cytidine analogues azacytidine and 5-aza-2'-deoxycytidine (decitabine) are commonly used to treat myelodysplastic syndromes, with or without a myeloproliferative component. It remains unclear whether the response to these hypomethylating agents results from a cytotoxic or an epigenetic effect. In this study, we address this question in chronic myelomonocytic leukaemia. We describe a comprehensive analysis of the mutational landscape of these tumours, combining whole-exome and whole-genome sequencing. We identify an average of 14±5 somatic mutations in coding sequences of sorted monocyte DNA and the signatures of three mutational processes. Serial sequencing demonstrates that the response to hypomethylating agents is associated with changes in DNA methylation and gene expression, without any decrease in the mutation allele burden, nor prevention of new genetic alteration occurence. Our findings indicate that cytosine analogues restore a balanced haematopoiesis without decreasing the size of the mutated clone, arguing for a predominantly epigenetic effect.

Germline duplication of ATG2B and GSKIP predisposes to familial myeloid malignancies.

No major predisposition gene for familial myeloproliferative neoplasms (MPN) has been identified. Here we demonstrate that the autosomal dominant transmission of a 700-kb duplication in four genetically related families predisposes to myeloid malignancies, including MPN, frequently progressing to leukemia. Using induced pluripotent stem cells and primary cells, we demonstrate that overexpression of ATG2B and GSKIP enhances hematopoietic progenitor differentiation, including of megakaryocytes, by increasing progenitor sensitivity to thrombopoietin (TPO). ATG2B and GSKIP cooperate with acquired JAK2, MPL and CALR mutations during MPN development. Thus, the germline duplication may change the fitness of cells harboring signaling pathway mutations and increases the probability of disease development.