Interleukin 7 (IL-7) and its receptor, formed by IL-7Rα (encoded by IL7R) and γc, are essential for normal T-cell development and homeostasis. Here we show that IL7R is an oncogene mutated in T-cell acute lymphoblastic leukemia (T-ALL). We find that 9% of individuals with T-ALL have somatic gain-of-function IL7R exon 6 mutations. In most cases, these IL7R mutations introduce an unpaired cysteine in the extracellular juxtamembrane-transmembrane region and promote de novo formation of intermolecular disulfide bonds between mutant IL-7Rα subunits, thereby driving constitutive signaling via JAK1 and independently of IL-7, γc or JAK3. IL7R mutations induce a gene expression profile partially resembling that provoked by IL-7 and are enriched in the T-ALL subgroup comprising TLX3 rearranged and HOXA deregulated cases. Notably, IL7R mutations promote cell transformation and tumor formation. Overall, our findings indicate that IL7R mutational activation is involved in human T-cell leukemogenesis, paving the way for therapeutic targeting of IL-7R-mediated signaling in T-ALL.

Graphical Abstract Highlights d H3.3 K27M mutation enhances neural stem cell self-renewal d Neonatal PDGFRa activation and Trp53 loss induces supratentorial and brainstem glioma d H3.3 K27M preferentially accelerates hindbrain tumorigenesis d H3.3 K27M drives bivalent gene activation associated with neurodevelopment in DIPG SUMMARYDiffuse intrinsic pontine gliomas (DIPGs) are incurable childhood brainstem tumors with frequent histone H3 K27M mutations and recurrent alterations in PDGFRA and TP53. We generated genetically engineered inducible mice and showed that H3.3 K27M enhanced neural stem cell self-renewal while preserving regional identity. Neonatal induction of H3.3 K27M cooperated with activating platelet-derived growth factor receptor a (PDGFRa) mutant and Trp53 loss to accelerate development of diffuse brainstem gliomas that recapitulated human DIPG gene expression signatures and showed global changes in H3K27 posttranslational modifications, but relatively restricted gene expression changes. Genes upregulated in H3.3 K27M tumors were enriched for those associated with neural development where H3K27me3 loss released the poised state of apparently bivalent promoters, whereas downregulated genes were enriched for those encoding homeodomain transcription factors.

Histone H3 K27M mutation is the defining molecular feature of the devastating pediatric brain tumor, diffuse intrinsic pontine glioma (DIPG). The prevalence of histone H3 K27M mutations indicates a critical role in DIPGs, but the contribution of the mutation to disease pathogenesis remains unclear. We show that knockdown of this mutation in DIPG xenografts restores K27Mdependent loss of H3K27me3 and delays tumor growth. Comparisons of matched DIPG xenografts with and without K27M knockdown allowed identification of mutation-specific effects on the transcriptome and epigenome. The resulting transcriptional changes recapitulate expression Terms of use and reuse: academic research for non-commercial purposes, see here for full terms. https://www.springer.com/aamterms-v1 *

AS and PYJ contributed equally to this work. Acknowledgments: The authors would like to thank the contribution of patients and the clinical teams involved in providing primary leukemia samples, Dr. Miguel Abecasis for providing thymic specimens, and Dr. Adolfo Ferrando for providing some of the NOTCH sequencing primers. Funding: this work was supported by grants from Fundação para a Ciência e a Tecnologia (FCT; POCI/SAU-OBS/58913 and PTDC/SAU-OBD/69974), and Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP; 08/10034-1), and Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq; 401122/2005-0). AS and PYJ have FCT SFRH and FAPESP PhD scholarships, respectively. Manuscript received on May 27, 2009. Revised version arrived on September 18, 2009. Manuscript accepted on October 8, 2009. Correspondence: João T. Barata, Cancer Biology Unit, Instituto de Medicina Molecular, Lisbon University Medical School, Av. Prof. Egas Moniz, 1649 .pt The Online version of this article has a Supplementary Appendix.T-cell acute lymphoblastic leukemia (T-ALL) patients frequently display NOTCH1 activating mutations and Notch can transcriptionally down-regulate the tumor suppressor PTEN. However, it is not clear whether NOTCH1 mutations associate with decreased PTEN expression in primary T-ALL. Here, we compared patients with or without NOTCH1 mutations and report that the former presented higher MYC transcript levels and decreased PTEN mRNA expression. We recently showed that T-ALL cells frequently display CK2-mediated PTEN phosphorylation, resulting in PTEN protein stabilization and concomitant functional inactivation. Accordingly, the T-ALL samples analyzed, irrespectively of their NOTCH1 mutational status, expressed significantly higher PTEN protein levels than normal controls. To evaluate the integrated functional impact of Notch transcriptional and CK2 post-translational inactivation of PTEN, we treated T-ALL cells with both the gamma-secretase inhibitor DAPT and the CK2 inhibitors DRB/TBB. Our data suggest that combined use of gamma-secretase and CK2 inhibitors may have therapeutic potential in T-ALL. Haematologica. 2010;95:674-678. doi:10.3324/haematol.2009 This is an open-access paper. ABSTRACT 674haematologica | 2010; 95(4) © F e r r a t a S t o r t i F o u n d a t i o nIn some cases, exon 26 and exon 27 were amplified and sequenced from genomic DNA with the intronic primers Notch26F and Notch26R, and Notch27F and Ex27R743, respectively. The region spanning TAD and PEST domains of exon 34 was amplified with primers Notch34TF and Notch34PR, and the resulting 855 bp fragment was sequenced with primers Notch34TF and Ex34PestF. The PEST coding region was also amplified with primers Ex34PestF and Ex34PestR, followed by semi-nested PCR with primers Notch34PF and Ex34PestR, and sequenced with primer Ex34PestR. This strategy covered all mutational hot-spot regions previously reported for NOTCH1. 1,6,7 Primer sequences and the PCR protocol are shown in the Online Supplementary Appendix. All mutations were c...

SummaryLymphotoxin-mediated activation of the lymphotoxin-b receptor (LTbR; LTBR) has been implicated in cancer, but its role in T-cell acute lymphoblastic leukaemia (T-ALL) has remained elusive. Here we show that the genes encoding lymphotoxin (LT)-a and LTb (LTA, LTB) are expressed in T-ALL patient samples, mostly of the TAL/LMO molecular subtype, and in the TEL-JAK2 transgenic mouse model of cortical/mature T-ALL (Lta, Ltb). In these mice, expression of Lta and Ltb is elevated in early stage T-ALL. Surface LTa 1 b 2 protein is expressed in primary mouse T-ALL cells, but only in the absence of microenvironmental LTbR interaction. Indeed, surface LT expression is suppressed in leukaemic cells contacting Ltbr-expressing but not Ltbr-deficient stromal cells, both in vitro and in vivo, thus indicating that dynamic surface LT expression in leukaemic cells depends on interaction with its receptor. Supporting the notion that LT signalling plays a role in T-ALL, inactivation of Ltbr results in a significant delay in TEL-JAK2-induced leukaemia onset. Moreover, young asymptomatic TEL-JAK2;Ltbr À/À mice present markedly less leukaemic thymocytes than agematched TEL-JAK2;Ltbr +/+ mice and interference with LTbR function at this early stage delayed T-ALL development. We conclude that LT expression by T-ALL cells activates LTbR signalling in thymic stromal cells, thus promoting leukaemogenesis.