Deciphering the role of lncRNAs in pediatric AML

Acute myeloid leukemia (AML) is a hematopoietic malignancy defined by genetic (and epigenetic) alterations in hematopoietic stem or progenitor cells. The emerging picture portrays a chromatin and transcriptional landscape that precisely controlled in four dimensions (space, time, cell type, developmental status) by a range of regulatory long non-coding RNAs (lncRNAs). Previously, we generated a gene expression atlas uncovering prognostically relevant lncRNA signatures shared between various AML sub-types and healthy hematopoietic stem cells. To elucidate the relevance of lncRNA candidates in leukemogenesis we performed an in vivo CRISPRi-based dropout screen. We targeted leukemic cell lines with various mutational sub-types and patient derived xenografts with stable expression of dCas9-KRAB. In our in vivo experimental design, firstly, we determined frequency of leukemia-initiating cells using a barcode approach. Thereafter we subjected our lncRNA candidate sub-libraries for screening potential candidates. Dropout candidates of the screening are further being validated via proliferation assays and qPCR.