RESUMO
Mutations in the GNAO1 gene, which encodes the abundant brain G-protein Gα o, result in neurologic disorders characterized by developmental delay, epilepsy, and movement abnormalities. There are over 50 mutant alleles associated with GNAO1 disorders; the R209H mutation results in dystonia, choreoathetosis, and developmental delay without seizures. Mice heterozygous for the human mutant allele (Gnao1 +/R209H) exhibit hyperactivity in open field tests but no seizures. We developed self-complementary adeno-associated virus serotype 9 (scAAV9) vectors expressing two splice variants of human GNAO1 Gα o isoforms 1 (GoA, GNAO1.1) and 2 (GoB, GNAO1.2). Bilateral intrastriatal injections of either scAAV9-GNAO1.1 or scAAV9-GNAO1.2 significantly reversed mutation-associated hyperactivity in open field tests. GNAO1 overexpression did not increase seizure susceptibility, a potential side effect of GNAO1 vector treatment. This represents the first report of successful preclinical gene therapy for GNAO1 encephalopathy applied in vivo. Further studies are needed to uncover the molecular mechanism that results in behavior improvements after scAAV9-mediated Gα o expression and to refine the vector design. SIGNIFICANCE STATEMENT: GNAO1 mutations cause a spectrum of developmental, epilepsy, and movement disorders. Here we show that intrastriatal delivery of scAAV9-GNAO1 to express the wild-type Gα o protein reduces the hyperactivity of the Gnao1 +/R209H mouse model, which carries one of the most common movement disorder-associated mutations. This is the first report of a gene therapy for GNAO1 encephalopathy applied in vivo on a patient-allele model.
Assuntos
Dependovirus , Subunidades alfa Gi-Go de Proteínas de Ligação ao GTP , Heterozigoto , Animais , Subunidades alfa Gi-Go de Proteínas de Ligação ao GTP/genética , Camundongos , Dependovirus/genética , Humanos , Masculino , Vetores Genéticos/administração & dosagem , Vetores Genéticos/genética , Hipercinese/genética , Mutação , Terapia Genética/métodos , Camundongos Endogâmicos C57BL , Locomoção/genéticaRESUMO
Metastatic cutaneous melanoma is a fatal skin cancer. Resistance to targeted and immune therapies limits the benefits of current treatments. Identifying and adding anti-resistance agents to current treatment protocols can potentially improve clinical responses. Myocardin-related transcription factor (MRTF) is a transcriptional coactivator whose activity is indirectly regulated by actin and the Rho family of GTPases. We previously demonstrated that development of BRAF inhibitor (BRAFi) resistance frequently activates the Rho/MRTF pathway in human and mouse BRAFV600E melanomas. In clinical trials, pretreatment with BRAFi reduces the benefit of immune therapies. We aimed to test the efficacy of concurrent treatment with our MRTF pathway inhibitor CCG-257081 and anti-PD1 in vivo and to examine its effects on the melanoma immune microenvironment. Because MRTF pathway activation upregulates the expression of immune checkpoint inhibitor genes/proteins, we asked whether CCG-257081 can improve the response to immune checkpoint blockade. CCG-257081 reduced the expression of PDL1 in BRAFi-resistant melanoma cells and decreased surface PDL1 levels on both BRAFi-sensitive and -resistant melanoma cells. Using our recently described murine vemurafenib-resistant melanoma model, we found that CCG-257081, in combination with anti-PD1 immune therapy, reduced tumor growth and increased survival. Moreover, anti-PD1/CCG-257081 co-treatment increased infiltration of CD8+ T cells and B cells into the tumor microenvironment and reduced tumor-associated macrophages. Here, we propose CCG-257081 as an anti-resistance and immune therapy-enhancing anti-melanoma agent.
Assuntos
Antígeno B7-H1 , Resistencia a Medicamentos Antineoplásicos , Inibidores de Checkpoint Imunológico , Melanoma , Proteínas Proto-Oncogênicas B-raf , Neoplasias Cutâneas , Microambiente Tumoral , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas B-raf/metabolismo , Animais , Melanoma/tratamento farmacológico , Melanoma/metabolismo , Melanoma/genética , Melanoma/patologia , Camundongos , Humanos , Antígeno B7-H1/antagonistas & inibidores , Antígeno B7-H1/metabolismo , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Neoplasias Cutâneas/tratamento farmacológico , Neoplasias Cutâneas/patologia , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/metabolismo , Microambiente Tumoral/efeitos dos fármacos , Linhagem Celular Tumoral , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/metabolismo , Transativadores/metabolismo , Transativadores/genética , Feminino , Transdução de Sinais/efeitos dos fármacos , Proteínas rho de Ligação ao GTP/metabolismoRESUMO
BACKGROUND: Many hypertension therapeutics were developed prior to major advances in drug receptor theory. Moreover, newer drugs may take advantage of some of the newly understood modalities of receptor function. GOAL: The goal of this review is to provide an up-to-date summary of drug receptor theory. This is followed by a discussion of the drug classes recognized for treating hypertension to which new concepts in receptor theory apply. RESULTS: We raise ideas for mechanisms of potential new antihypertensive drugs and whether they may take advantage of new theories in drug-receptor interaction.