title: Experimental evolution of an oncolytic vesicular stomatitis virus with increased selectivity for p53-deficient cells.
abstract: Experimental evolution has been used for various biotechnological applications including protein and microbial cell engineering, but less commonly in the field of oncolytic virotherapy. Here, we sought to adapt a rapidly evolving RNA virus to cells deficient for the tumor suppressor gene p53, a hallmark of cancer cells. To achieve this goal, we established four independent evolution lines of the vesicular stomatitis virus (VSV) in p53-knockout mouse embryonic fibroblasts (p53-/- MEFs) under conditions favoring the action of natural selection. We found that some evolved viruses showed increased fitness and cytotoxicity in p53-/- cells but not in isogenic p53+/+ cells, indicating gene-specific adaptation. However, full-length sequencing revealed no obvious or previously described genetic changes associated with oncolytic activity. Half-maximal effective dose (EC50) assays in mouse p53-positive colon cancer (CT26) and p53-deficient breast cancer (4T1) cells indicated that the evolved viruses were more effective against 4T1 cells than the parental virus or a reference oncolytic VSV (MΔ51), but showed no increased efficacy against CT26 cells. In vivo assays using 4T1 syngeneic tumor models showed that one of the evolved lines significantly delayed tumor growth compared to mice treated with the parental virus or untreated controls, and was able to induce transient tumor suppression. Our results show that RNA viruses can be specifically adapted typical cancer features such as p53 inactivation, and illustrate the usefulness of experimental evolution for oncolytic virotherapy.
authors.affiliation: Instituto Cavanilles de Biodiversidad y Biologia Evolutiva, Universidad de Valencia, Valencia, Spain; Center for Innovative Cancer Research, Ottawa Hospital Research Institute, Ottawa, ON, Canada.
authors.forename: Raquel
authors.initials: R
authors.lastname: Garijo
authors.affiliation: Instituto Cavanilles de Biodiversidad y Biologia Evolutiva, Universidad de Valencia, Valencia, Spain.
authors.forename: Pablo
authors.initials: P
authors.lastname: Hernández-Alonso
authors.affiliation: Departamento de Biología Molecular y Celular, Centro Nacional de Biotecnología, Madrid, Spain; Centro de Investigación en Medicina Molecular (CIMUS) and Instituto de Investigaciones Sanitarias (IDIS), Universidade de Santiago de Compostela, Santiago de Compostela, Spain.
authors.forename: Carmen
authors.initials: C
authors.lastname: Rivas
authors.affiliation: Center for Innovative Cancer Research, Ottawa Hospital Research Institute, Ottawa, ON, Canada.
authors.forename: Jean-Simon
authors.initials: JS
authors.lastname: Diallo
authors.affiliation: Instituto Cavanilles de Biodiversidad y Biologia Evolutiva, Universidad de Valencia, Valencia, Spain; Department of Genetics, Universidad de Valencia, Valencia, Spain.
authors.forename: Rafael
authors.initials: R
authors.lastname: Sanjuán
mesh_terms: D000818:Animals; D001943:Breast Neoplasms / Q000235:genetics* / Q000473:pathology; D045744:Cell Line, Tumor; D003110:Colonic Neoplasms / Q000235:genetics* / Q000473:pathology; D019020:Directed Molecular Evolution; D005260:Female; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D050130:Oncolytic Virotherapy; D050504:Oncolytic Viruses / Q000235:genetics*; D016159:Tumor Suppressor Protein p53 / Q000235:genetics* / Q000378:metabolism; D054243:Vesicular Stomatitis / Q000235:genetics / Q000821:virology; D018116:Vesiculovirus / Q000235:genetics* / Q000378:metabolism
chemical_list: D016159:Tumor Suppressor Protein p53