RAS activation via CRLF2 signaling is a widespread mechanism in Down syndrome acute lymphoblastic leukemia regardless of RAS mutations

Abstract

Background: Down syndrome acute lymphoblastic leukemia (DS-ALL) is characterized by the high frequency of CRLF2-rearrangements, JAK2-mutations, or RAS-pathway mutations. Intriguingly, JAK2 and RAS mutations are mutually exclusive in leukemic sub-clones, causing dichotomy in therapeutic target choices.Results: Here we show that in primary leukemic cells from DS-ALL, in the absence of RAS-mutations, wild-type (wt)RAS is active, and/or can be induced by the physiological ligand TSLP of the transmembrane-receptor CRLF2. We show active/inducible RAS in 14/20 (70%) of primary DS-ALL samples analyzed, 8 of which had no RAS-mutations, but 75% of those had either mutated or hyperphosphorylated JAK2. No wtRAS cases with mutated/hyperphosphorylated JAK2 were observed that lacked activated RAS protein. We prove in a cell model that elevated CRLF2 in combination with constitutionally active JAK2 is sufficient to activate wtRAS. We show that TSLP boosts the direct binding of active PTPN11 to wtRAS. Pre-inhibition of RAS or PTPN11, but not of PI3K or JAK signaling, prevented TSLP-induced RAS-GTP boost.Using multivariate-clustering based on RAS-activity/inducibility we show significant separation between standard-risk and high-risk DS-ALL groups. Cox proportional-hazards model showed proteinactivity (but not mutation status) as independently predictive of outcome. Conclusions:Our data indicate that RAS protein activity levels (and not JAK2/RAS mutation profiles), are predictive of outcome. Importantly, our data suggest that inhibition of RAS and direct RASpathway components should be combined with PI3K/mTOR and/or JAK2 inhibitors for high-risk cases.Therapeutically this is relevant for >75% of DS-ALL and our additional data suggest that it warrants further investigation in high-risk non-DS-ALL.wtRAS activation is frequent in DS-ALL