RESUMO
Due to cancer's genetic complexity, significant advances in the treatment of metastatic disease will require sophisticated, multi-pronged therapeutic approaches. Here we demonstrate the utility of a Drosophila melanogaster cell platform for the production and in vivo delivery of multi-gene biotherapeutic systems. We show that cultured Drosophila S2 cell carriers can stably propagate oncolytic viral therapeutics that are highly cytotoxic for mammalian cancer cells without adverse effects on insect cell viability or gene expression. Drosophila cell carriers administered systemically to immunocompetent animals trafficked to tumors to deliver multiple biotherapeutics with little apparent off-target tissue homing or toxicity, resulting in a therapeutic effect. Cells of this Dipteran invertebrate provide a genetically tractable platform supporting the integration of complex, multi-gene biotherapies while avoiding many of the barriers to systemic administration of mammalian cell carriers. These transporters have immense therapeutic potential as they can be modified to express large banks of biotherapeutics with complementary activities that enhance anti-tumor activity.
Assuntos
Drosophila melanogaster/genética , Terapia Genética/métodos , Neoplasias Pulmonares/terapia , Terapia Viral Oncolítica/métodos , Vírus Oncolíticos/genética , Animais , Chlorocebus aethiops , Drosophila melanogaster/citologia , Drosophila melanogaster/imunologia , Drosophila melanogaster/virologia , Feminino , Regulação Neoplásica da Expressão Gênica , Regulação Viral da Expressão Gênica , Células HT29 , Células HeLa , Humanos , Imunocompetência , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/virologia , Células MCF-7 , Camundongos Endogâmicos BALB C , Vírus Oncolíticos/imunologia , Vírus Oncolíticos/patogenicidade , Fatores de Tempo , Transfecção , Carga Tumoral , Células Vero , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
In our earlier studies, Semliki Forest virus vector VA7 completely eliminated type I interferon (IFN-I)-unresponsive human U87-luc glioma xenografts, whereas interferon-responsive mouse gliomas proved refractory. Here, we describe in two clones of CT26 murine colon carcinoma, opposed patterns of IFN-I responsiveness and sensitivity to VA7. Both CT26WT and CT26LacZ clones secreted biologically active interferon in vitro upon virus infection but only CT26WT cells were protected. Focal infection of CT26WT cultures was self-limiting but could be rescued using IFN-I pathway inhibitor Ruxolitinib or antibody against IFNß. Whole transcriptome sequencing (RNA-Seq) and protein expression analysis revealed that CT26WT cells constitutively expressed 56 different genes associated with pattern recognition and IFN-I signaling pathways, spanning two reported anti-RNA virus gene signatures and 22 genes with reported anti-alphaviral activity. Whereas CT26WT tumors were strictly virus-resistant in vivo, infection of CT26LacZ tumors resulted in complete tumor eradication in both immunocompetent and severe combined immune deficient mice. In double-flank transplantation experiments, CT26WT tumors grew despite successful eradication of CT26LacZ tumors from the contralateral flank. Tumor growth progressed uninhibited also when CT26LacZ inoculums contained only a small fraction of CT26WT cells, demonstrating dominance of IFN responsiveness when heterogeneous tumors are targeted with interferon-sensitive oncolytic viruses.
Assuntos
Neoplasias do Colo/terapia , Terapia Viral Oncolítica , Vírus Oncolíticos/genética , Vírus da Floresta de Semliki/genética , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Efeito Espectador , Linhagem Celular Tumoral , Neoplasias do Colo/imunologia , Neoplasias do Colo/patologia , Vetores Genéticos , Proteínas de Fluorescência Verde/biossíntese , Interferon Tipo I/farmacologia , Interferon Tipo I/uso terapêutico , Interferon beta/metabolismo , Camundongos Endogâmicos BALB C , Necrose , Transplante de Neoplasias , Fator de Transcrição STAT1/metabolismo , Transfecção , Resultado do TratamentoRESUMO
Rhabdoviruses (RVs) are currently being pursued as anticancer therapeutics for various tumor types, notably leukemia. However, modest virion production and limited spread between noncontiguous circulating leukemic cells requires high-dose administration of RVs, which exceeds the maximum tolerable dose of the live virus. Furthermore, in severely immunosuppressed leukemic patients, the potential for uncontrolled live virus spread may compromise the safety of a live virus approach. We hypothesized that the barriers to oncolytic virotherapy in liquid tumors may be overcome by administration of high-dose non-replicating RVs. We have developed a method to produce unique high-titer bioactive yet non-replicating rhabdovirus-derived particles (NRRPs). This novel biopharmaceutical is multimodal possessing direct cytolytic and immunomodulatory activity against acute leukemia. We demonstrate that NRRP resistance in normal cells is mediated by intact antiviral defences including interferon (IFN). This data was substantiated using murine models of blast crisis. The translational promise of NRRPs was demonstrated in clinical samples obtained from patients with high-burden multidrug-resistant acute myeloid leukemia. This is the first successful attempt to eradicate disseminated cancer using a non-replicating virus-derived agent, representing a paradigm shift in our understanding of oncolytic virus-based therapies and their application toward the treatment of acute leukemia.