Mutant JAK3 phosphoproteomic profiling predicts synergism between JAK3 inhibitors and MEK/BCL2 inhibitors for the treatment of T-cell acute lymphoblastic leukemia

Abstract

Mutations in the interleukin-7 receptor (IL7R) or the Janus kinase 3 (JAK3) kinase occur frequently in T-cell acute lymphoblastic leukemia (T-ALL) and both are able to drive cellular transformation and the development of T-ALL in mouse models. However, the signal transduction pathways downstream of JAK3 mutations remain poorly characterized. Here we describe the phosphoproteome downstream of the JAK3(L857Q)/(M511I) activating mutations in transformed Ba/F3 lymphocyte cells. Signaling pathways regulated by JAK3 mutants were assessed following acute inhibition of JAK1/JAK3 using the JAK kinase inhibitors ruxolitinib or tofacitinib. Comprehensive network interrogation using the phosphoproteomic signatures identified significant changes in pathways regulating cell cycle, translation initiation, mitogen-activated protein kinase and phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K)/AKT signaling, RNA metabolism, as well as epigenetic and apoptotic processes. Key regulatory proteins within pathways that showed altered phosphorylation following JAK inhibition were targeted using selumetinib and trametinib (MEK), buparlisib (PI3K) and ABT-199 (BCL2), and found to be synergistic in combination with JAK kinase inhibitors in primary T-ALL samples harboring JAK3 mutations. These data provide the first detailed molecular characterization of the downstream signaling pathways regulated by JAK3 mutations and provide further understanding into the oncogenic processes regulated by constitutive kinase activation aiding in the development of improved combinatorial treatment regimens.

CXCR4 allows T cell acute lymphoblastic leukemia to escape from JAK1/2 and BCL2 inhibition through CNS infiltration

“…Further, combining these two inhibitors in vitro, achieved a synergistic effect demonstrating a greater increase in apoptosis and a decrease in proliferation compared to either inhibitor alone. This data confirmed findings previously demonstrated in T-ALL (20,35,36) and other cancers. (37)(38)(39) Ruxolitinib treatment alone has been considered a breakthrough therapy for myeloproliferative neoplasms but long-term studies have shown that this treatment is not curative and does not lead to molecular or pathological remission.…”

JAK/STAT-Activating Genomic Alterations Are a Hallmark of T-PLL

“…JAK3 mutations were the most recurrent genomic aberration affecting JAK/STAT genes in this meta-analysis with a hotspot mutation rate of 36.4% (literature: 21% [12]-71% [17]). The oncogenic potential of the most frequent JAK3 mutation M511I as well as the less frequently occurring mutations A573V and V674A was shown in various systems: M511I, A573V, as well as V674A mutant cell lines demonstrated high phosphorylation of STAT5 and ERK, leading to cytokine independent growth [39,40]. When transplanting mice with bone marrow progenitor cells harboring the M511I or A573V mutation in JAK3, they develop a T-ALL-like disease, characterized by an expansion of immature CD8 + T-cells.…”

Can one target T-cell ALL?

“…Moreover, ruxolitinib inhibited JAK-STAT signaling and abrogated the hyperactivation effect of IL-7 and was highly effective in these preclinical models [41]. Finally, signaling pathways with altered phosphorylation after JAK inhibition (MEK, PI3K) and BCL2 can be pharmacologically inhibited, which results in synergistic antitumor effects in combination with JAK kinase inhibitors in primary T-ALL samples with JAK3 mutations [42]. …”