Genetic hallmarks of recurrent/metastatic adenoid cystic carcinoma

Abstract

BACKGROUNDAdenoid cystic carcinoma (ACC) is a rare malignancy arising in salivary glands and other sites, characterized by high rates of relapse and distant spread. Recurrent/metastatic (R/M) ACCs are generally incurable, due to a lack of active systemic therapies. To improve outcomes, deeper understanding of genetic alterations and vulnerabilities in R/M tumors is needed.METHODSAn integrated genomic analysis of 1,045 ACCs (177 primary, 868 R/M) was performed to identify alterations associated with advanced and metastatic tumors. Intratumoral genetic heterogeneity, germline mutations, and therapeutic actionability were assessed.RESULTSCompared with primary tumors, R/M tumors were enriched for alterations in key Notch (NOTCH1, 26.3% vs. 8.5%; NOTCH2, 4.6% vs. 2.3%; NOTCH3, 5.7% vs. 2.3%; NOTCH4, 3.6% vs. 0.6%) and chromatin-remodeling (KDM6A, 15.2% vs. 3.4%; KMT2C/MLL3, 14.3% vs. 4.0%; ARID1B, 14.1% vs. 4.0%) genes. TERT promoter mutations (13.1% of R/M cases) were mutually exclusive with both NOTCH1 mutations (q = 3.3 × 10-4) and MYB/MYBL1 fusions (q = 5.6 × 10-3), suggesting discrete, alternative mechanisms of tumorigenesis. This network of alterations defined 4 distinct ACC subgroups: MYB+NOTCH1+, MYB+/other, MYBWTNOTCH1+, and MYBWTTERT+. Despite low mutational load, we identified numerous samples with marked intratumoral genetic heterogeneity, including branching evolution across multiregion sequencing.CONCLUSIONThese observations collectively redefine the molecular underpinnings of ACC progression and identify further targets for precision therapies.FUNDINGAdenoid Cystic Carcinoma Research Foundation, Pershing Square Sohn Cancer Research grant, the PaineWebber Chair, Stand Up 2 Cancer, NIH R01 CA205426, the STARR Cancer Consortium, NCI R35 CA232097, the Frederick Adler Chair, Cycle for Survival, the Jayme Flowers Fund, The Sebastian Nativo Fund, NIH K08 DE024774 and R01 DE027738, and MSKCC through NIH/NCI Cancer Center Support Grant (P30 CA008748).

Keywords: Head and neck cancer; Oncology.

Figure 1. Comparison of primary vs.

R/M ACC genomic alterations. (A) Oncoprint of primary and R/M ACC. (B) Mutations in cancer genes that are enriched in R/M ACC relative to primary tumors. Statistical comparisons are provided in Supplemental Table 4.

Figure 2. Lollipop plots of mutations in key genes in R/M ACC.

For NOTCH1 in particular, 337 distinct alterations were observed, with 221 (65.6%) found in established hot-spot regions and considered activating mutations. Total number of alterations and incidence by gene are listed in Supplemental Table 4. HD, heterodimerization negative regulatory region; JmjC, Jumonji C; BAF250_C, SWI/SNF chromatin remodeling complex.

Figure 3. Distinct molecular ACC subgroups based on genetic alterations.

(A) Volcano plot illustrating genes correlated or anticorrelated with NOTCH1 mutation in R/M ACC. NOTCH1 alterations were found to be highly mutually exclusive with TERT promoter mutations and highly cooccurrent with chromatin-remodeling pathway gene alterations (KDM6A, CREBB, ARID1A, EP300), where q represents the Benjamini-Hochberg FDR statistic. (B) R/M ACC subgroups based on mutually exclusive genetic alterations, including MYB⁺NOTCH⁺, MYB⁺/other, MYB^WTTERT⁺, and MYB^WTNOTCH⁺ divisions. Displayed cases represent 53.2% (305/573) of R/M cases with available data. (C) OS of R/M ACC subgroups. Triple-negative cases are defined as cases lacking alterations in MYB, NOTCH1, or the TERT promoter (MYB^WTNOTCH1^WTTERT^WT). Comparison of the triple-negative subgroup with other subgroups by log-rank test: MYB⁺NOTCH1⁺ (P = 5.42 × 10^–5), MYB^WTNOTCH1⁺ (P = 0.0269), MYB⁺/other (P = 0.414), MYB^WTTERT⁺ (P = 0.296).

Figure 4. OS comparison for R/M ACC by individual gene.

(A) NOTCH1 mutant vs. NOTCH1 WT, (B) NOTCH1 mutant (activating) vs. NOTCH1 mutant (nonactivating), (C) KDM6A mutant vs. KDM6A WT, and (D) TERT promoter mutant vs. TERT promoter WT.

Figure 5. R/M ACC tumors that harbor gene mutations potentially targetable with available kinase inhibitors.

(A) Oncoprint of R/M ACC genomic alterations stratified by genes potentially targeted by tyrosine kinase inhibitor agents, encompassing 40.3% of R/M ACC cohort. Unpublished data outlined in Supplemental Table 1. (B) Incidence of ACC patients with potentially targetable alterations, derived from OncoKB database. Levels of evidence based upon FDA labeling, NCCN guidelines, expert group recommendations, and biologic response in scientific literature. FMI, Foundation Medicine, Inc.

Figure 6. Clinical response of representative metastatic ACC patients with tumors found to have PIK3CA mutations who were enrolled in PI3K basket trials.

Images represent axial CT scans with volume reduction by RECIST criteria. (A) Baseline imaging of a 43-year-old woman with a spleen metastasis. (B) PR after 2 months of treatment with 30.6% volume reduction. (C) Baseline imaging of a 73-year-old woman with metastatic ACC in the left lung. (D) SD after 2 months of treatment with 23.6% volume reduction. (E) Baseline imaging of a 58-year-old man with metastatic ACC in the left lung. (F) SD after 1 month of treatment with 25.0% volume reduction.

Figure 7. Multiregion sequencing and clonality analysis in a single salivary ACC patient with subsequent lung metastases.

(A) Mutation heatmap demonstrating CCF for each mutation in each site. Primary tumor, 6 subspatial sites, and 8 distant lung metastatic lesions were biopsied and evaluated via high-depth sequencing and orthogonal validation. (B) NMF analysis defines 3 distinct subgroups. Cluster 1 (metastases 6D, 2B, and 4A) diverges significantly from clusters that comprise the primary tumor (bulk) or the tumor subspatial regions. Coph, cophenetic correlation coefficient.