RESUMO
A recombinant Newcastle Disease Virus (NDV), encoding either a human (NDVhuGM-CSF, MEDI5395) or murine (NDVmuGM-CSF) GM-CSF transgene, combined broad oncolytic activity with the ability to significantly modulate genes related to immune functionality in human tumor cells. Replication in murine tumor lines was significantly diminished relative to human tumor cells. Nonetheless, intratumoral injection of NDVmuGM-CSF conferred antitumor effects in three syngeneic models in vivo; with efficacy further augmented by concomitant treatment with anti-PD-1/PD-L1 or T-cell agonists. Ex vivo immune profiling, including T-cell receptor sequencing, revealed profound immune-contexture changes consistent with priming and potentiation of adaptive immunity and tumor microenvironment (TME) reprogramming toward an immune-permissive state. CRISPR modifications rendered CT26 tumors significantly more permissive to NDV replication, and in this setting, NDVmuGM-CSF confers immune-mediated effects in the noninjected tumor in vivo Taken together, the data support the thesis that MEDI5395 primes and augments cell-mediated antitumor immunity and has significant utility as a combination partner with other immunomodulatory cancer treatments.
Assuntos
Fator Estimulador de Colônias de Granulócitos e Macrófagos/administração & dosagem , Imunomodulação , Imunoterapia/métodos , Vírus da Doença de Newcastle/genética , Terapia Viral Oncolítica/instrumentação , Microambiente Tumoral , Animais , Apoptose , Proliferação de Células , Neoplasias do Colo/imunologia , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Neoplasias do Colo/terapia , Feminino , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Many natural human proteins have functional properties that make them useful as therapeutic drugs. However, not all these proteins are compatible with large-scale manufacturing processes or sufficiently stable to be stored for long periods prior to use. In this study, we focus on small four-helix bundle proteins and employ ribosome display in conjunction with three parallel selection pressures to favour the isolation of variant proteins with improved expression, solubility and stability. This in vitro evolution strategy was applied to two human proteins with known drug development issues, granulocyte colony-stimulating factor (G-CSF) and erythropoietin (EPO). In the case of G-CSF, the soluble expression levels in Escherichia coli were improved 1000-fold, while for EPO the level of aggregation in an accelerated shelf-life study was reduced from over 80% to undetectable levels. These results exemplify the general utility of our in vitro evolution strategy for improving the drug-like properties of therapeutic proteins.
