title: The positive is inside the negative: HER2-negative tumors can express the HER2 intracellular domain and present a HER2-positive phenotype.
abstract: Overexpression of human epithelial growth factor receptor 2 (HER2) is a poor prognostic factor in breast cancer. HER2 is a transmembrane receptor comprising an extracellular domain (ECD), a single transmembrane domain, and an intracellular domain (ICD) with tyrosine-kinase activity. Receptor dimerization triggers pivotal effector pathways in cancer, such as phosphatidylinositol 3-kinase (PI3K) signaling. Currently, screening of HER2 in breast tumors for prognostic and therapeutic purposes involves immunohistochemical (IHC) phenotyping for the ECD, in which tumors with IHC scores below 2+ are reported as HER2-negative. We used a label-free liquid chromatography-mass spectrometry (LC-MS) proteomic approach to compare plasma samples from patients with HER2-positive breast tumors and patients with HER2-negative tumors. Patients with HER2-negative tumors expressed higher circulating levels of calpain-10 than patients with HER2-positive tumors. Calpains cleave HER2, releasing its ECD and transforming phenotypically positive tumors into phenotypically negative tumors. Therefore, we investigated the expression of the ICD in HER2-negative samples that overexpressed calpain-10. We found that 16% of HER2-negative tumors were positive for HER2-ICD, which was associated with circulating HER2-ECD. HER2 gene amplification was also observed in some HER2-negative tumors. Positive staining for the PI3K pathway was observed in the HER2-negative, ICD-positive tumors, similar to the HER2-positive cohort. Microarray analysis revealed that HER2-negative, ICD-positive samples clustered between HER2-positive tumors and triple-negative tumors. Survival analysis revealed that outcome in women with HER2-negative, ICD-positive tumors was better than in women bearing HER2-negative, ICD-negative (triple negative) tumors but was quite similar to HER2-positive tumors and worse than women with luminal A tumors. Moreover, in vitro analyses revealed that MDA-MB 231, a triple negative cell line, possesses calpain-10 and HER2-p-ICD up-regulation and blockage of calpain-10 activity promoted an increase in HER2-p-ICD and p-AKT levels, suggesting an increase in these pathways signaling. These data indicate that HER2-negative tumors with HER2-ICD positivity exhibit clinical behavior closer to that of HER2-positive tumors. This indicates a need for HER2-ICD screening when determining the molecular profile of breast tumors. These findings further indicate that lapatinib should be investigated as a target therapy for HER2-ICD-positive breast tumors.
authors.affiliation: Laboratório de Células Tronco, Instituto Nacional do Câncer, INCA, Rio de Janeiro, Brazil; Laboratório de Mediadores Inflamatórios, Universidade Estadual do Oeste do Paraná, Unioeste, Francisco Beltrão, Paraná, Brazil. Electronic address: carolpanis@sercomtel.com.br.
authors.forename: Carolina
authors.initials: C
authors.lastname: Panis
authors.affiliation: Laboratório de Células Tronco, Instituto Nacional do Câncer, INCA, Rio de Janeiro, Brazil; INCT para o Controle do Câncer, Laboratório de Células Tronco, Instituto Nacional do Câncer, INCA, Rio de Janeiro, Brazil; Departamento de Bioquímica, Instituto de Química, Universidade Federal do Rio de Janeiro, UFRJ, Rio de Janeiro, Brazil.
authors.forename: Luciana
authors.initials: L
authors.lastname: Pizzatti
authors.affiliation: Laboratório de Células Tronco, Instituto Nacional do Câncer, INCA, Rio de Janeiro, Brazil; INCT para o Controle do Câncer, Laboratório de Células Tronco, Instituto Nacional do Câncer, INCA, Rio de Janeiro, Brazil.
authors.forename: Stephany
authors.initials: S
authors.lastname: Corrêa
authors.affiliation: Laboratório de Células Tronco, Instituto Nacional do Câncer, INCA, Rio de Janeiro, Brazil; INCT para o Controle do Câncer, Laboratório de Células Tronco, Instituto Nacional do Câncer, INCA, Rio de Janeiro, Brazil.
authors.forename: Renata
authors.initials: R
authors.lastname: Binato
authors.affiliation: Laboratório de Células Tronco, Instituto Nacional do Câncer, INCA, Rio de Janeiro, Brazil; INCT para o Controle do Câncer, Laboratório de Células Tronco, Instituto Nacional do Câncer, INCA, Rio de Janeiro, Brazil.
authors.forename: Gabriela Ferreira
authors.initials: GF
authors.lastname: Lemos
authors.affiliation: Instituto do Câncer de Londrina, Londrina, Paraná, Brazil.
authors.forename: Ana Cristina
authors.initials: AC
authors.lastname: da Silva do Amaral Herrera
authors.affiliation: Laboratório de Células Tronco, Instituto Nacional do Câncer, INCA, Rio de Janeiro, Brazil.
authors.forename: Teresa Fernandes
authors.initials: TF
authors.lastname: Seixas
authors.affiliation: Laboratório de Fisiopatologia de Radicais Livres, Universidade Estadual de Londrina, Londrina, Paraná, Brazil.
authors.forename: Rubens
authors.initials: R
authors.lastname: Cecchini
authors.affiliation: Laboratório de Células Tronco, Instituto Nacional do Câncer, INCA, Rio de Janeiro, Brazil; INCT para o Controle do Câncer, Laboratório de Células Tronco, Instituto Nacional do Câncer, INCA, Rio de Janeiro, Brazil.
authors.forename: Eliana
authors.initials: E
authors.lastname: Abdelhay
mesh_terms: D014408:Biomarkers, Tumor / Q000235:genetics / Q000378:metabolism; D001943:Breast Neoplasms / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D002853:Chromatography, Liquid; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D013058:Mass Spectrometry; D008875:Middle Aged; D010641:Phenotype; D011379:Prognosis; D011446:Prospective Studies; D040901:Proteomics; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000096:biosynthesis / Q000172:deficiency / Q000378:metabolism*; D015398:Signal Transduction; D016019:Survival Analysis
chemical_list: D014408:Biomarkers, Tumor; D018719:Erb-b2 Receptor Tyrosine Kinases; C508053:ERBB2 protein, human
keywords: Breast cancer; Calpain-10; Human epithelial growth factor receptor 2 (HER2); Intracellular domain; Proteomics