FBW7 mutations in leukemic cells mediate NOTCH pathway activation and resistance to γ-secretase inhibitors

Abstract

γ-secretase inhibitors (GSIs) can block NOTCH receptor signaling in vitro and therefore offer an attractive targeted therapy for tumors dependent on deregulated NOTCH activity. To clarify the basis for GSI resistance in T cell acute lymphoblastic leukemia (T-ALL), we studied T-ALL cell lines with constitutive expression of the NOTCH intracellular domain (NICD), but that lacked C-terminal truncating mutations in NOTCH1. Each of the seven cell lines examined and 7 of 81 (8.6%) primary T-ALL samples harbored either a mutation or homozygous deletion of the gene FBW7, a ubiquitin ligase implicated in NICD turnover. Indeed, we show that FBW7 mutants cannot bind to the NICD and define the phosphodegron region of the NICD required for FBW7 binding. Although the mutant forms of FBW7 were still able to bind to MYC, they do not target it for degradation, suggesting that stabilization of both NICD and its principle downstream target, MYC, may contribute to transformation in leukemias with FBW7 mutations. In addition, we show that all seven leukemic cell lines with FBW7 mutations were resistant to the MRK-003 GSI. Most of these resistant lines also failed to down-regulate the mRNA levels of the NOTCH targets MYC and DELTEX1 after treatment with MRK-003, implying that residual NOTCH signaling in T-ALLs with FBW7 mutations contributes to GSI resistance.

It's T-ALL about Notch

“…Although the status of the Notch locus was not examined in tumors from these mice, evidence from human T-ALL samples gives insight into the Fbw7-Notch dynamic in leukemogenesis. Inactivating mutations of Fbw7 were found in 30% of human T-ALL samples examined, and analysis of these tumors revealed that only wild-type or HD mutant Notch forms were expressed (O'Neil et al, 2007;Matsuoka et al, 2008). Whereas, Notch PEST domain mutations were only found in samples expressing wild-type Fbw7, these data imply that in T-ALL, an inactivating Fbw7 mutation would have the same effect as N ICD PEST mutations, increasing Notch activity by decreasing Notch degradation by the proteosome.…”

“…SCF FBW7 -mediated degradation of Notch requires phosphorylation of multiple residues. The first phosphorylation event at S2514 mediates Fbw7 binding, and the second at T2512 is required for polyubiquitination of the protein by SCF FBW7 that results in rapid turnover of N ICD by the proteasome (O'Neil et al, 2007;Thompson et al, 2008). Phosphorylation of Notch at S2514 is mediated by CyclinC:cdk8 after recruitment to the Notch activation complex by Maml (Fryer et al, 2004;O'Neil et al, 2007).…”

Early to Intermediate Steps of Tumor Embolic Formation Involve Specific Proteolytic Processing of E-Cadherin Regulated by Rab7

“…[29][30][31][32][33]. Notch activation involves the proteolytic cleavage of the Notch ligand/receptor complex by g-secretase to release the Notch intracellular domain (NICD) that translocates to the nucleus and upregulates expression of a number of different downstream genes (34). Different Notch receptors are preferentially activated in different breast cancer cell lines.…”

Does Notch play a tumor suppressor role across diverse squamous cell carcinomas?

“…Missense mutations in NRR domain likely destabilize inhibitor state of NRR domain, allowing ligand‐independent Notch activation 90. PEST domain is involved in proteasomal degradation of NOTCH 91, 92, and truncations lacking PEST region likely impair the degradation of NOTCH and augment Notch activation.…”