RESUMO
Primary effusion lymphoma (PEL), which is an aggressive subgroup of B-cell lymphoma associated with Kaposi sarcoma-associated herpes virus/human herpes virus-8, is refractory to the standard treatment, and exhibits a poor survival. Although PU.1 is downregulated in PEL, the potential role of its reduction remains to be elucidated. In this investigation, we analyzed the DNA methylation of PU.1 cis-regulatory elements in PEL and the effect of restoring PU.1 on PEL cells. The mRNA level of PU.1 was downregulated in PEL cells. The methylated promoter and enhancer regions of the PU.1 gene were detected in PEL cells. Suppression of cell growth and apoptosis were caused by the restoration of PU.1 in PEL cells. A microarray analysis revealed that interferon-stimulated genes (ISGs) including pro-apoptotic ISGs were strongly increased in BCBL-1 cells after the induction of PU.1. Reporter assays showed that PU.1 transactivated pro-apoptotic ISG promoters, such as the XAF1, OAS1 and TRAIL promoters. Mutations at the PU.1 binding sequences suppressed its transactivation. We confirmed the binding of PU.1 to the XAF1, OAS1 and TRAIL promoters in a chromatin immunoprecipitation assay. PU.1 suppressed ORF57 activation by inducing IRF7. The reinduction of PU.1 reduced formation of ascites and lymphoma cell infiltration of distant organs in PEL xenograft model mice. Collectively, PU.1 has a role in tumor suppression in PEL and its down-regulation is associated with PEL development. Restoring PU.1 with demethylation agents may be a novel therapeutic approach for PEL.
Assuntos
Interferons/genética , Linfoma de Efusão Primária/genética , Linfoma de Efusão Primária/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Transativadores/genética , Transativadores/metabolismo , Animais , Apoptose/fisiologia , Linhagem Celular Tumoral , Proliferação de Células/fisiologia , Metilação de DNA , Xenoenxertos , Humanos , Fator Regulador 7 de Interferon/biossíntese , Fator Regulador 7 de Interferon/genética , Interferons/farmacologia , Linfoma de Efusão Primária/patologia , Masculino , Camundongos , Camundongos Endogâmicos NOD , Análise em Microsséries , Regiões Promotoras Genéticas , Ativação Transcricional , TransfecçãoRESUMO
Human T-cell leukemia virus type 1 (HTLV-1), which causes adult T-cell leukemia (ATL) in humans, establishes a life-long latent infection. Current therapies are not very effective against HTLV-1-associated disorders. A novel therapeutic approach may help to combat HTLV-1 infection. A molecular therapy that targets the proviral genome is favorable because the therapeutic effect occurs specifically in HTLV-1-infected cells, regardless of whether they express viral genes. In this proof-of-concept study, we developed a therapeutic molecule based on zinc finger nuclease (ZFN) to achieve this goal. We designed a ZFN that specifically recognized conserved region of HTLV-1 long terminal repeat (LTR) and introduced it into various HTLV-1-positive human T-cell lines, including HTLV-1-transformed and ATL-derived cell lines. The ZFN disrupted the promoter function of HTLV-1 LTR and specifically killed HTLV-1-infected cells. We also showed a potential approach of this therapeutic molecule to remove the proviral genome from HTLV-1-infected cells, something that has not been possible before. The therapeutic effect of ZFN was confirmed in an in vivo model of ATL. This strategy may form the basis of a therapy that can eradicate HTLV-1 infection. Similar approaches can be used to target other malignancy-associated viruses.
