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Acute myeloid leukemia (AML) is an aggressive malignancy of haematopoietic stem cells driven by a well-defined set of somatic mutations.1,2 Identifying the mutations driving individual cases is important for assigning the patient to a recognized World Health Organisation category, establishing prognostic risk and tailoring post-consolidation therapy.3 As a result, AML research and diagnostic laboratories apply diverse methodologies to detect important mutations and many are introducing next-generation sequencing (NGS) approaches to study extended panels of genes in order to refine genomic classification and prognostic category.1 Besides the implications of these developments on costs, expertise and reliance on commercial providers, they also do not capture gene expression data, which have independent prognostic value that cannot be inferred from somatic mutation profiles. The ability to detect AML gene mutations as well as gene expression profiles from a single assay, could provide a holistic tool that accelerates research, simplifies diagnostic work-up and helps develop integrated algorithms to refine individual patient prognosis. Here, we show that AML RNA sequencing (RNA-seq) data can be used to reliably detect all types of clinically important mutations and develop a bespoke fast and easy-to-use software (RNAmut) for this purpose that can be readily used by teams/laboratories without in-house bioinformatic expertise.

Glioma is the most common intrinsic brain tumor and also occurs in the spinal cord. Activating EGFR mutations are common in IDH1 wild-type gliomas. However, the cooperative partners of EGFR driving gliomagenesis remain poorly understood. RESULTS: We explore EGFR-mutant glioma evolution in conditional mutant mice by whole-exome sequencing, transposon mutagenesis forward genetic screening, and transcriptomics. We show mutant EGFR is sufficient to initiate gliomagenesis in vivo, both in the brain and spinal cord. We identify significantly recurrent somatic alterations in these gliomas including mutant EGFR amplifications and Sub1, Trp53, and Tead2 loss-of-function mutations. Comprehensive functional characterization of 96 gliomas by genome-wide piggyBac insertional mutagenesis in vivo identifies 281 known and novel EGFR-cooperating driver genes, including Cdkn2a, Nf1, Spred1, and Nav3. Transcriptomics confirms transposon-mediated effects on expression of these genes. We validate the clinical relevance of new putative tumor suppressors by showing these are frequently altered in patients' gliomas, with prognostic implications. We discover shared and distinct driver mutations in brain and spinal gliomas and confirm in vivo differential tumor suppressive effects of Pten between these tumors. Functional validation with CRISPR-Cas9-induced mutations in novel genes Tead2, Spred1, and Nav3 demonstrates heightened EGFRvIII-glioma cell proliferation. Chemogenomic analysis of mutated glioma genes reveals potential drug targets, with several investigational drugs showing efficacy in vitro. CONCLUSION: Our work elucidates functional driver landscapes of EGFR-mutant gliomas, uncovering potential therapeutic strategies, and provides new tools for functional interrogation of gliomagenesis.

We recently identified the splicing kinase gene SRPK1 as a genetic vulnerability of acute myeloid leukemia (AML). Here, we show that genetic or pharmacological inhibition of SRPK1 leads to cell cycle arrest, leukemic cell differentiation and prolonged survival of mice transplanted with MLL-rearranged AML. RNA-seq analysis demonstrates that SRPK1 inhibition leads to altered isoform levels of many genes including several with established roles in leukemogenesis such as MYB, BRD4 and MED24. We focus on BRD4 as its main isoforms have distinct molecular properties and find that SRPK1 inhibition produces a significant switch from the short to the long isoform at the mRNA and protein levels. This was associated with BRD4 eviction from genomic loci involved in leukemogenesis including BCL2 and MYC. We go on to show that this switch mediates at least part of the anti-leukemic effects of SRPK1 inhibition. Our findings reveal that SRPK1 represents a plausible new therapeutic target against AML.