High Incidence of Notch-1 Mutations in Adult Patients with T-Cell Acute Lymphoblastic Leukaemia.

Notch signalling plays an important role in controlling lymphoid progenitor differentiation, with Notch activation inducing a T-cell fate. In mice, constitutive Notch activation leads to T-cell leukaemia. Acquired activating mutations of Notch-1 involving the heterodimerization (HD) and/or PEST domains have been reported in 56% of paediatric/adolescent patients with T-cell acute lymphoblastic leukaemia (T-ALL). Mutations in the HD domain are predicted to increase sensitivity to ligand activation and truncations of the PEST domain are thought to increase the half-life of intracellular Notch. The presence of mutations in both domains is synergistic. The frequency and nature of Notch-1 mutations in adult patients has not been reported. Therefore, DNA was obtained from diagnostic bone marrow samples of 24 adult T-ALL patients entered into the UK Medical Research Council UKALLXII trial. Fragments of Notch-1 encoding the N-terminal HD domain (HD-N), C-terminal HD domain (HD-C), transcriptional activation domain (TAD) and PEST domain were analysed by denaturing HPLC and products with an abnormal chromatograph sequenced. Seventeen patients (71%) had 1/more mutation(s) of likely pathological significance; 12 in the HD domain only, 1 in the PEST domain only, and 4 in both domains. In the HD domain, 5 patients had point mutations causing leucine to proline substitutions which had been reported previously in paediatric/adolescent patients (L1575P, L1586P, L1594P). One patient had two leucine to proline mutations on different alleles. Of the other HD mutations, four patients had point mutations (L1597H, H1592Q+F1593T in cis, I1617N, A1697D), whilst 7 had in-frame insertions/deletions (of which 4 were complex contiguous deletions and insertions). All of these were novel and without exception involved highly conserved regions of the gene. Five mutations were detected in the PEST domain; all were insertions/deletions that led to premature stop codons, either through a frameshift or through direct insertion of a stop codon, truncating the receptor by 39 to 119 amino acids. Of the 15 patients who had remission DNA samples available, nine had a mutation at disease presentation. In each case the mutation was no longer detectable in remission. Of the four relapse samples available, two patients with mutations at diagnosis relapsed with the same mutation(s). Neither acquired a different mutation. The two patients who were wild type at diagnosis did not acquire a mutation at relapse. Mutations were not simply a feature of the younger adult population; the median age of the mutant-positive patients in this cohort was 35 years (range 16–55) compared to 20 years (range 16–50) for mutant-negative patients. The median presenting white cell count was 51.8 × 10⁹/L and did not differ significantly between the two groups (41.1 × 10⁹/L for positive patients, 108.8 × 10⁹/L for negative patients, P .70). Whilst most of the mutations identified are novel, these data suggest that Notch mutations are at least as common in adult patients, occur in the same domains and are similar in nature to those reported in paediatric patients. Furthermore, the mutations are acquired, recur at relapse, and may be candidate markers for minimal residual disease analysis. Gamma-secretase inhibitors, which have been shown to induce cell cycle arrest in vitro in T-ALL cell lines harbouring Notch-1 mutations, may offer a rational targeted approach for these patients who, in the older age group, have a poor outcome with current therapy.