Evaluation of WNT pathway transcriptomes in T-cell Acute Lymphoblastic Leukemia by targeted RNA-Seq

Wnt/Fzd signaling has been implicated in hematopoietic stem cell maintenance and in acute leukemia establishment. We previously described a recurrent rearrangement involving the WNT10B locus (WNT10BR), characterized by expression of the WNT10BIVS1 transcript variant, in acute myeloid leukemia. To determine the occurrence of WNT10BR in T-cell acute lymphoblastic leukemia (T-ALL), we retrospectively analyzed an italian cohort of patients (n=20) and detected the WNT10BIVS1 with a high prevalence (14/20). To address genes involved in WNT10B molecular response, we designed a Wnt targeted RNA sequencing panel. We identify Wnt agonists and antagonists, from which the expression of FZD6, PLCB4, and PROM1 stand out in WNT10BIVS1-positive patients compared to negative ones. Using MOLT4 and MUTZ-2 as cell models, characterized by the expression of WNT10BIVS1, we found that WNT10B drives major Wnt activation through receipt of ligand to the FZD6 receptor complex. The WNT10B/FZD6 interaction were interfered with by short hairpin RNAs (shRNAs)-mediated gene silencing and by small molecules-mediated inhibition of WNTs secretion. We found that WNT10BIVS1 knockdown, or pharmacological interference by the LGK974 porcupine (PORCN) inhibitor, reduces WNT10B/FZD6 protein complex formation and significantly impairs intracellular effectors and leukemic expansion. These results describe the molecular circuit induced by WNT10B and suggest WNT10B/FZD6 as new targets in the T-ALL treatment strategy.