TMOD-05MODELING ACVR1 AND HIST1H3B MUTATIONS IN DIFFUSE INTRINSIC PONTINE GLIOMAIN VIVOUSING THE SLEEPING BEAUTY SYSTEM

Abstract

Diffuse intrinsic pontine glioma (DIPG) is a brainstem tumor that occurs in the pons in children; median age 6-7 years. The most frequent DIPG mutations affect the N-terminal tail of histone H3.3 and histone H3.1 and result in the change of a lysine to methionine at residue 27. In addition, six recurrent somatic activating mutations in ACVR1, which encodes for the bone morphogenetic protein (BMP) receptor kinase, have also been identified in DIPG tumors. We aim to use the sleeping beauty transposase system to generate spontaneous in vivo tumor models that will allow us to test the contributions of these genes to the pathogenesis of DIPG tumors. We have cloned ACVR1 G328V, and HIST1H3B K27M sequences into a plasmid that contains flanking inverted repeats (IR) which contain binding sequences for the sleeping beauty transposase, which will excise DNA between the IR sites and insert it into the host genome. Plasmids encoding a short hair pin for p53, oncogenic NRAS, the sleeping beauty transposase/luciferase were injected with group 1: ACVR1 G328V; group 2: HIST1H3B K27M; or group 3: ACVR1 G328V and HIST1H3B K27M into the subventricular zone of neonatal mice. Tumor development in mice is monitored by measuring luciferase activity in vivo; animals that have reached moribund stage are perfused and processed for histology. Our long-term goal is to understand the interplay between the ACVR1 activating mutation and the histone H3K27M mutation. The ACVR1 mutations result in constitutive BMP pathway activation and the H3K27M mutation has been shown to globally alter H3K27 methylation. Through this study we hope to elucidate how ACVR1 mutations and H3K27 mutations contribute to tumor progression in order to uncover novel therapeutic targets for DIPG which would improve prognosis for DIPG patients.