RESUMO
Trastuzumab plays an important role in breast cancer therapy. However, a significant fraction of patients do not respond to therapy or they tend to develop resistance shortly after beginning therapy. Although some resistance mechanisms have been described, it is unclear whether these mechanisms can coexist. In this study, we analyzed the resistance mechanisms in the breast cancer cell line JIMT-1, a model of intrinsic trastuzumab resistance. We compared the JIMT-1 cell line with a panel of eight HER-2 positive breast cancer cell lines. All cell lines were characterized for the phosphatidylinositol 3-kinase (PIK3CA) mutation status, expression levels of the phosphatase and tensin homolog on chromosome 10 (PTEN) and neuregulin-1 (NRG1) mRNA, HER-2 gene copy number, and protein expression. The results were correlated to the sensitivity to trastuzumab and lapatinib as well as the potency of trastuzumab-mediated antibody-dependent cellular cytotoxicity (ADCC) evoked by trastuzumab. JIMT-1 cells showed several co-existing drug resistance mechanisms, including an activating mutation of the PIK3CA gene, low expression of PTEN, high expression of NRG1, and relatively low expression of HER-2 receptor protein (despite gene amplification). All these features were present at variable levels in other cell lines, whereas JIMT-1 was unique in displaying all these factors at the same time. Unexpectedly, ADCC reaction by normal lymphocytes was equally strong in all HER-2 positive cell lines, without any correlation to molecular markers or direct sensitivity to the drugs. Resistance to trastuzumab and lapatinib is probably caused by several co-existing molecular mechanisms. Direct sensitivity to trastuzumab and lapatinib was not correlated with ADCC.
Assuntos
Anticorpos Monoclonais/farmacologia , Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Quinazolinas/farmacologia , Anticorpos Monoclonais Humanizados , Citotoxicidade Celular Dependente de Anticorpos/efeitos dos fármacos , Neoplasias da Mama/genética , Neoplasias da Mama/imunologia , Linhagem Celular Tumoral , Classe I de Fosfatidilinositol 3-Quinases , Resistencia a Medicamentos Antineoplásicos , Feminino , Amplificação de Genes , Humanos , Lapatinib , Mutação , Neuregulina-1/biossíntese , Neuregulina-1/genética , PTEN Fosfo-Hidrolase/biossíntese , PTEN Fosfo-Hidrolase/genética , Fosfatidilinositol 3-Quinases/genética , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Receptor ErbB-2/biossíntese , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismo , TrastuzumabRESUMO
Autoimmune regulator (AIRE) is responsible for the development of organ-specific autoimmune disease in a monogenic fashion. Rare and low levels of tissue expression together with the lack of AIRE-expressing cell lines have hampered a detailed analysis of the molecular dynamics of AIRE. Here we have established cell lines stably transfected with AIRE and studied the regulatory mechanisms for its subcellular expression. We found that nuclear body (NB) formation by AIRE was dependent on the cell cycle. Biochemical fractionation revealed that a significant proportion of AIRE is associated with the nuclear matrix, which directs the functional domains of chromatin to provide sites for gene regulation. Upon proteasome inhibition, AIRE NBs were increased with concomitant reduced expression in the cytoplasm, suggesting that subcellular targeting of AIRE is regulated by a ubiquitin-proteasome pathway. We also found that AIRE NBs compete for cAMP-response element-binding protein-binding protein/p300, a common coactivator of transcription, with the promyelocytic leukemia gene product. These results suggest that the transcriptional regulating activities of AIRE within a cell are controlled and organized in a spatiotemporal manner.
