Distinct, Aggressive Subgroup of ACC Identified With Activating NOTCH1 Mutations

NOTCH1 mutations have been found to define an aggressive subgroup of patients with adenoid cystic carcinomas (ACC), with a corresponding pattern of metastatic spread, according to the results of a recent study from the Journal of Clinical Oncology.

This distinct, aggressive phenotype is characterized by solid histology, liver and bone metastasis, poor prognosis, and possible responsiveness to Notch1 inhibitors.

ACCs represent a heterogeneous group of chemotherapy refractory tumors, and there is no standard of care treatment for patients with recurrent and/or metastatic disease. The current study investigated the underlying molecular characteristics of this phenotype and evaluated the potential of the Notch1 pathway as a therapeutic target.

The Notch pathway is involved in the maintenance of stem cells, cell fate specification, proliferation, and angiogenesis. Whole exome sequencing (WES) of ACC samples has shed light on the genetic landscape of this disease and has also provided evidence for Notch pathway alterations in 11% to 29% of patients.

There are a total of 4 NOTCH genes that encode transmembrane receptors: NOTCH1, -2, -3, and -4, and 5 membrane-bound ligands: delta-like ligands (DLL1, -3, -4) and Jagged (JAG1, -2).

Notch signaling is typically initiated by receptor-ligand interaction, which then leads to consecutive receptor cleavages, the second cleavage by the gamma-secretase complex that frees the Notch intracellular domain (NICD) to enter the nucleus, displace corepressors such as SPEN, and form a transcriptional activation complex with the DNA-binding factor RBPJ and associated coactivators. The ubiquitin ligase complexes containing FBXW7 regulate the generation and stability of NICD.

NOTCH1 mutations are oncogenic drivers that occur in 50% of T-cell acute lymphoblastic leukemias (T-ALLs). T-ALL-activating mutations concentrate in 2 hotspot regions: in-frame mutations in exons 25 to 28—which disrupt the negative regulatory region and lead to ligand-independent Notch1 activation—and stop-codon or nonsense mutations in exon 34 that result in deletion of the C-terminal degron domain and NICD stabilization.

The researchers conducted genomic profiling in a total of 102 tumors and WES in an additional 46 samples, as well as targeted sequencing for a gene panel that included NOTCH1 in 32 samples. Eighteen NOTCH1 mutations were found in 15 tumors. Two patients harbored more than 1 NOTCH1 mutation. Seventeen mutations in 14 patients occurred in the T-ALL hotspots.

To assess whether these mutations were activating, the investigators evaluated the association between NOTCH1 mutations and NICD immunohistochemical (IHC) staining, which has been established as a marker for Notch1 pathway activation.

Using tumor tissues from 72 patients for NICD staining, the results showed a statistically significant association between NOTCH1 mutations and NICD positivity.

All 10 tumors (100%) with NOTCH1 mutations that were predicted to be activating were NICD positive. Thirty (49%) of 61 NOTCH1 wild-type tumors stained positive (P = .004). The only tumor with a NOTCH1 mutation predicted to be inactivating was NICD negative.

“Our work demonstrates that NICD staining was quite sensitive (100%) in its ability to identify patients with NOTCH1 activating mutations,” the authors wrote in the study. “However, it lacks specificity, because 49% of NOTCH1 wild-type tumors were NICD positive.”

The authors also reported an index patient with NOTCH1-mutant ACC who achieved a partial response after receiving 2 doses of the Notch1-specific monoclonal antibody brontictuzumab during a clinical trial. This patient had at least 2 known NOTCH1 mutations before treatment with brontictuzumab, with a third mutation detected in cell-free DNA. This likely reflects the patient’s tumor heterogeneity. Moreover, the presence of multiple alterations promoting Notch1 signaling supports the role of these mutations as oncogenic drivers.

Although the current data show that NOTCH1 mutations are associated with Notch1 pathway activation, there is also evidence to suggest that pathway activation can occur via alternative routes. In total, 21 patients (20.5%) had mutations in known Notch pathway-related genes. Researchers observed mutations in SPEN in 6 patients, including 3 concurrent with NOTCH1. Two patients were identified with NOTCH2 mutations, 1 with a SPEN comutation.

When compared with patients who have NOTCH1 wild-type, those with mutations were more likely to have solid histology (P <.001), or present with advanced-stage disease (P = .02), or both (solid subtype and stage IV vs others; P = .01).

Although the lungs represented the most common site of metastasis among patients with recurrent ACC, patients with NOTCH1 mutations were significantly less likely to develop such metastasis (odds ratio [OR], 0.24; P = .02). Instead, these individuals were far more likely to develop liver metastasis (OR, 3.8; P = .04).

According to a correlation analysis between NICD-positive (n = 40) and NICD-negative (n = 32) tumors, patients with NICD-positive tumors were more likely to have solid histology (P = .02) and metastasis in the liver (P = .02).

The median relapse-free survival (RFS) and overall survival (OS) in the overall patient population were, respectively, 30 months and 108 months. Median RFS was 12.5 months and 33.9 months, respectively, for NOTCH1-mutant and NOTCH1 wild-type. For patients with NOTCH1 mutations, OS was significantly shorter, with a median of 29.6 months (121.9 months for NOTCH1 wild-type; P = .001).

The NICD-positive patients demonstrated a shorter RFS when compared with those who were NICD-negative (14.6 vs 39 months; P = .03). OS, however, did not significantly differ in a comparison between the 2 NICD groups (44 vs 108 months; P = .2).

Following this analysis of genomic, pathologic, and clinical outcomes, further research is warranted to investigate the activity of Notch1 inhibitors in biomarker-selected patients with ACC, according to the authors.

Ferrarotto R, Mitani Y, Diao L, et al. Activating NOTCH1 mutations define a distinct subgroup of patients with adenoid cystic carcinoma who have poor prognosis, propensity to bone and liver metastasis, and potential responsiveness to Notch1 inhibitors [published online November 21, 2016]. J Clin Oncol. doi: 10.1200/JCO.2016.67.5264.