An Integrative Genomic Approach to Examine Mutations and Biological Pathways Associated with Hematological Malignancy Development in DDX41 Mutated Families

Abstract

DEAD-Box helicase 41 (DDX41) is one of the most commonly reported familial hematological malignancy (HM) genes, first reported in 2015. Mutated, it predisposes to both MDS/AML and lymphoma with an age of diagnosis similar to that of sporadic cases. Consequently, unrecognized DDX41 mutated cases are present in 'sporadic' cohorts with families often identified this way. Individuals with DDX41 mutation or deficiency have generally poor outcome with no effective targeted therapies available. The biological mechanism by which mutant DDX41 predisposes to HM is poorly understood making risk assessment for asymptomatic carriers difficult. In over 50% of HM affected individuals with germline DDX41 mutations, an acquired somatic mutation is identified on the other DDX41 allele with R525H being most common. Analysis of other somatic mutations in germline DDX41 mutation carriers reveals a minimal fingerprint of acquired somatic alterations in known myeloid malignancy driver genes with an absence of NPM1 and FLT3 mutations and an under-representation of mutations in DNMT3A, TET2, IDH1/2 and RUNX1. This may indicate a mechanism of leukemia development unique to germline DDX41 cases. Through an integrative combinatorial genomics approach of whole exome sequencing, leukemic panel sequencing and RNA-sequencing, we are in the process of comprehensively assessing six independent DDX41 germline mutated families. We and others have identified that DDX41 mutations are recurrently associated with MDS with erythroid dysplasia and erythroid subtype leukemia consistent with functional in vitro experiments showing that DDX41 mutations including loss-of-function can impact erythroid differentiation. Our preliminary RNA-Seq analysis on peripheral blood mononuclear cells of DDX41 carriers with HM reveals altered expression of genes involved in hemoglobin complex and innate immunity. DDX41 is reported to bind RNA splicing components such as SF3B1 and PRPF8 which are recurrently mutated in MDS/AML and intriguingly, the R525H mutation disrupts this interaction. Further analysis of our RNA-Seq data interrogating alternative splicing, gene expression analysis and biological pathways to uncover mechanisms for mutant DDX41 oncogenicity is currently ongoing. We have used CRISPR-Cas9 technology to generate Ddx41M1I mice as a model for one of the most common germline mutations in familial HM to investigate the direct effect of DDX41 mutation on hematopoiesis. Preliminary results indicate a sub-clinical reduction in hemoglobin levels in mice carrying the M1I mutation in heterozygosity when compared to their wildtype littermates. We are further investigating these mice with particular emphasis on characterizing hematopoiesis longitudinally to better understand mechanisms of disease onset and progression. This integrative genomic and functional approach to evaluate both mutations and biological pathways affected in DDX41 mutated malignancies will provide biological insight facilitating advancement in diagnosis, risk assessment, monitoring and personalized treatment. Disclosures Scott: Celgene: Honoraria.

A novel bi-alleleic DDX41 mutations in B-cell lymphoblastic leukemia: case report

“…We found altered gene expression levels of genes involved in the process essential for red blood cells, which is consistent with previous observations that DDX41 mutations, including LOF, can affect erythroid differentiation [ 12 ]. For myeloid cell differentiation, Ph + B-ALL IKZF1 +/ DDX41dm had significantly altered levels of expression compared to Ph + B-ALL IKZF1 +/ DDX41− .…”

DDX41 regulates the expression and alternative splicing of genes involved in tumorigenesis and immune response

“…Polprasert et al ( 16 ) identified somatic DDX41 mutations in myeloid neoplasms that resulted in loss of tumor suppressor function due to altered pre-mRNA splicing and RNA processing. RNA-sequencing (seq) analysis on peripheral bold mononuclear cells of DDX41 mutation carriers with hematological malignancy (HM) revealed altered expression levels of genes involved in hemoglobin complex and innate immunity ( 17 ). The majority of germline mutations are frameshift mutations, suggesting loss of function, with DDX41 serving as a tumor suppressor, which may impact initiation, maintenance or progression of tumorigenesis ( 18 ).…”