Functional analysis of receptor tyrosine kinase mutations in lung cancer identifies oncogenic extracellular domain mutations of ERBB2

Abstract

We assessed somatic alleles of six receptor tyrosine kinase genes mutated in lung adenocarcinoma for oncogenic activity. Five of these genes failed to score in transformation assays; however, novel recurring extracellular domain mutations of the receptor tyrosine kinase gene ERBB2 were potently oncogenic. These ERBB2 extracellular domain mutants were activated by two distinct mechanisms, characterized by elevated C-terminal tail phosphorylation or by covalent dimerization mediated by intermolecular disulfide bond formation. These distinct mechanisms of receptor activation converged upon tyrosine phosphorylation of cellular proteins, impacting cell motility. Survival of Ba/F3 cells transformed to IL-3 independence by the ERBB2 extracellular domain mutants was abrogated by treatment with small-molecule inhibitors of ERBB2, raising the possibility that patients harboring such mutations could benefit from ERBB2-directed therapy.HER2 | breast cancer | bladder cancer L ung cancer is the leading cause of cancer death, accounting for over 150,000 deaths annually in the United States alone (1). Current treatment options are thus inadequate for the majority of patients and additional therapies are needed. Mutationally activated oncogenes that promote tumorigenesis represent potential drug targets due to frequent dependency of tumor cells on such oncogenes (2, 3), and somatically altered receptor tyrosine kinases in particular have been successfully exploited as therapeutic targets in several cancers.The prototypical therapy targeted to a somatically activated tyrosine kinase oncogene is imatinib mesylate, which targets the BCR-ABL fusion protein in chronic myelogenous leukemia (4). Targeted therapies developed for lung cancer include gefitinib and erlotinib, small-molecule inhibitors of mutationally activated EGFR in lung adenocarcinoma (5-8), and crizotinib, a smallmolecule inhibitor of the EML4-ALK translocation product in lung adenocarcinoma (9). Trastuzumab, a monoclonal antibody inhibitor targeting ERBB2, and the small-molecule EGFR/ ERBB2 inhibitor lapatinib are effective in ERBB2-amplified patients with breast cancer (10, 11).The advent of next-generation sequencing technologies has enabled compilation of large somatic mutation datasets from cancer sequencing studies. Statistical methods that examine differences in gene mutation frequency can reveal evidence of positive selection; however, demonstration of the contribution of a mutated gene to tumorigenesis additionally requires functional validation. To identify new lung cancer oncogenes, we systematically assessed somatic alleles of significantly mutated receptor tyrosine kinase genes reported in patients with lung adenocarcinoma (12) for activity in cellular transformation assays.Although most receptor tyrosine kinase mutations tested failed to score, novel extracellular domain mutations of ERBB2 were oncogenic. Our results indicate a unique therapeutic opportunity for patients with lung and breast cancer who harbor extracellular domain mutations of ERBB2. Resul...

HER2 missense mutations have distinct effects on oncogenic signaling and migration

“…Prior overexpression studies identified that HER2 mutations can lead to transformative changes, including increases in anchorage independent growth and aberrant morphology in 3D culture, even without detectable increases in HER2 phosphorylation (4,7). Therefore, we performed a series of assays to identify transformed properties of our HER2 mutant cell line panel.…”

Analysis of different HER‐2 mutations in breast cancer progression and drug resistance

“…S310F/Y, G309A/E, S335C) 23, 47, 48, 49, 50. Parts of these mutations that cluster in exon 8 are oncogenic and activated by two distinct mechanisms, characterized by elevated C‐terminal tail phosphorylation, such as S310F/Y, or covalent dimerization mediated by intermolecular disulfide bond formation, such as G309E and S335C 47. These extracellular domain mutations are also sensitive to small‐molecule inhibitors of HER‐2, more similar to the kinase domain mutations 23.…”

“…These extracellular domain mutations are also sensitive to small‐molecule inhibitors of HER‐2, more similar to the kinase domain mutations 23. Trastuzumab was shown to be effective against the cells expressing G309 and S310 mutations, giving hope to the patients harbouring these mutations 47.…”

In situ selectivity profiling and crystal structure of SML-8-73-1, an active site inhibitor of oncogenic K-Ras G12C

“…Both target a common activating mutation, G12C, to achieve irreversible binding. K-Ras G12C is present in an estimated 10-20% of Ras-driven cancers and in roughly 50% of Rasmutated lung adenocarcinomas (11)(12)(13). For lung cancer alone, this means that therapeutics targeting the G12C mutation could treat roughly 25,000 people per year in the United States (14).…”