Functional identification of genes associated with mutant IL7Ra in T-ALL

• Grant number: 15/21149-8
• Support Opportunities: Scholarships abroad - Research Internship - Doctorate
• Start date: January 29, 2016
• End date: January 28, 2017
• Field of knowledge: Biological Sciences - Genetics - Human and Medical Genetics
• Principal Investigator: José Andrés Yunes
• Grantee: Gisele Olinto Libanio Rodrigues
• Supervisor: Scott Kenneth Durum
• Host Institution: Centro Infantil de Investigações Hematológicas Dr Domingos A Boldrini (CIB). Campinas , SP, Brazil
• Institution abroad: National Cancer Institute (NCI), Frederick, United States
• Associated to the scholarship: 12/10284-3 - Identification of mutations functionally associated with the mutant IL7Ra in T-ALL, BP.DR

Abstract

The IL7/IL7R mediated signaling is essential for normal development and homeostasis of T cell precursors. Early studies have shown that around 10% of patients with T-cell Acute Lymphoblastic Leukemia (T-ALL) have mutations in the alpha chain of the receptor for IL7 (IL-7R±). A considerable number of IL7R mutations occur in exon 6 and, 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-7R± subunits, thereby driving constitutive signaling via JAK1 and independently of IL-7 and the gamma-chain. Some genetic changes are important factors to initiate leukemia, but in many cases these changes are insufficient to generate a full leukemic phenotype, suggesting that collaborative oncogenic mutations may be present. Our SNP/CNV-Array and exome sequencing analysis of patient samples carrying the mutant IL7R (L7Rmut/T-ALL) showed some recurrent gene mutations that may potentially collaborate with the mutant IL7R. Silencing of these genes in murine BaF3 pro-B-lymphocyte cells, stably expressing the mutant human IL7R, showed a beneficial effect in proliferation and clonogenic assays. This suggests candidate tumor suppressor genes that cooperate with gain-of-function IL-7R mutations. In this project we propose to validate these findings in a cellular context closer to T-ALL cells, i.e. by using D1 cells, a murine TP53 knockout thymocyte cell line, or immature thymocytes. D1 cells will be used to investigate the proliferative/survival effects in vitro and engraftment/progression (time-to-leukemia) in immunocompromised mice, caused by mutation (knockout) of individual candidate genes in conjunction with ectopic expression of the mutant IL7R. Experiments with primary thymocytes, cultured onto OP9-DL4 cells feeders with cytokines, will be used to investigate effects on T-cell differentiation and leukemogenesis in vivo. (AU)