RESUMEN
Neurofibromatosis type 2 (NF2) is a rare disorder that causes vestibular schwannomas (VS), meningiomas and ependymomas. To date, there is no FDA approved drug-based treatment for NF2. We have previously identified that BET inhibition can selectively reduce growth of the NF2-null schwannoma and Schwann cells in vitro and tumorigenesis in vivo and, separately, reported that inhibition of Focal Adhesion Kinase 1 (FAK1) via crizotinib has antiproliferative effects in NF2-null Schwann cells. The current study was aimed at determining whether combined BET and FAK inhibition can synergize and to identify the mechanisms of action. A panel of normal and NF2-null Schwann and schwannoma cell lines were used to characterize the effects of combined BET and FAK inhibition in vitro and in vivo using pharmacological and genetic approaches. The mechanism of action was explored by chromatin immunoprecipitation, ChIP-PCR, western blotting, and functional approaches. We find that combined BET and FAK inhibition are synergistic and inhibit the proliferation of NF2-null schwannoma and Schwann cell lines in vitro and in vivo, by arresting cells in the G1/S and G2/M phases of the cell cycle. Further, we identify the mechanism of action through the downregulation of FAK1 transcription by BET inhibition, which potentiates inhibition of FAK by 100-fold. Our findings suggest that combined targeting of BET and FAK1 may offer a potential therapeutic option for the treatment of NF2-related schwannomas.
Asunto(s)
Proliferación Celular , Quinasa 1 de Adhesión Focal , Neurilemoma , Neurofibromina 2 , Neuroma Acústico , Animales , Humanos , Ratones , Quinasa 1 de Adhesión Focal/metabolismo , Quinasa 1 de Adhesión Focal/antagonistas & inhibidores , Quinasa 1 de Adhesión Focal/genética , Neuroma Acústico/patología , Neuroma Acústico/genética , Neuroma Acústico/tratamiento farmacológico , Neuroma Acústico/metabolismo , Neurilemoma/patología , Neurilemoma/genética , Neurilemoma/tratamiento farmacológico , Neurilemoma/metabolismo , Neurofibromina 2/genética , Neurofibromina 2/metabolismo , Proliferación Celular/efectos de los fármacos , Línea Celular Tumoral , Sinergismo Farmacológico , Neurofibromatosis/tratamiento farmacológico , Neurofibromatosis/genética , Neurofibromatosis/patología , Neurofibromatosis 2/genética , Neurofibromatosis 2/tratamiento farmacológico , Neurofibromatosis 2/patología , Neurofibromatosis 2/metabolismo , Células de Schwann/efectos de los fármacos , Células de Schwann/metabolismo , Células de Schwann/patología , Neoplasias Cutáneas/patología , Neoplasias Cutáneas/tratamiento farmacológico , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Background: Neurofibromatosis type 2 (NF2) is an autosomal dominant genetic disease characterized by development of schwannomas on the VIIIth (vestibular) cranial nerves. Bromodomain and extra-terminal domain (BET) proteins regulate gene transcription and their activity is required in a variety of cancers including malignant peripheral nerve sheath tumors. The use of BET inhibitors as a therapeutic option to treat NF2 schwannomas has not been explored and is the focus of this study. Methods: A panel of normal and NF2-null Schwann and schwannoma cell lines were used to characterize the impact of the BET inhibitor JQ1 in vitro and in vivo. The mechanism of action was explored by chromatin immunoprecipitation of the BET BRD4, phospho-kinase arrays and immunohistochemistry (IHC) of BRD4 in vestibular schwannomas. Results: JQ1 inhibited proliferation of NF2-null schwannoma and Schwann cell lines in vitro and in vivo. Further, loss of NF2 by CRISPR deletion or siRNA knockdown increased sensitivity of cells to JQ1. Loss of function experiments identified BRD4, and to a lesser extent BRD2, as BET family members mediating the majority of JQ1 effects. IHC demonstrated elevated levels of BRD4 protein in human vestibular schwannomas. Analysis of signaling pathways effected by JQ1 treatment suggests that the effects of JQ1 treatment are mediated, at least in part, via inhibition of PI3K/Akt signaling. Conclusions: NF2-deficient Schwann and schwannoma cells are sensitive to BET inhibition, primarily mediated by BRD4, which is overexpressed in human vestibular schwannomas. Our results suggest BRD4 regulates PI3K signaling and likely impedes NF2 schwannoma growth via this inhibition. These findings implicate BET inhibition as a therapeutic option for NF2-deficient schwannomas.
