R-loop Dysregulation in Myeloid Neoplasms (360G-Wellcome-200703_Z_16_Z)

Insights gleaned from pre-neoplastic disorders have reinvigorated interest in the cancer-specific properties that trigger and respond to DNA damage. Myeloproliferative neoplasms (MPNs) are pre-leukaemic disorders that reflect an early-stage malignancy with the potential to progress to acute leukaemia. We have previously demonstrated that MPN-derived tumour cells exhibit perturbations in DNA replication that lead to DNA damage. We now have exciting preliminary data implicating nucleic acid structures called "R-loops" as the primary cause of MPN-associated genomic instability. R-loops are essential for multiple normal cellular processes but can also promote DNA damage if inappropriately regulated. This seed award application is focused on a central question: why do R-loops promote DNA damage in MPN cells but not normal cells? To address this issue, we will undertake two thematically-related but non-overlapping streams of investigations. For Aim 1, we will use an unbiased proteomics-based approach to characterise any differences in protein composition of R-loops between normal and MPN cells. For Aim 2, we will perform a pooled CRISPR/Cas9-based genetic screen to identify genes that modulate how MPN cells regulate R-loop-induced recombination. Cumulatively, we hope to generate preliminary data that starts to deconvolute the structural and functional complexity of R-loops in leukaemia development.