Generation and characterization of a novel oncolytic adenovirus expressing OX40L and CD40L

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dc.contributor Helsingin yliopisto, Bio- ja ympäristötieteellinen tiedekunta, Bio- ja ympäristötieteellinen tiedekunta fi
dc.contributor University of Helsinki, Faculty of Biological and Environmental Sciences, Faculty of Biological and Environmental Sciences en
dc.contributor Helsingfors universitet, Bio- och miljövetenskapliga fakulteten, Bio- och miljövetenskapliga fakulteten sv
dc.contributor.author Martins, Beatriz
dc.date.issued 2020
dc.identifier.uri URN:NBN:fi:hulib-202006052630
dc.identifier.uri http://hdl.handle.net/10138/315896
dc.description.abstract According to the latest estimations, cancer is the second leading cause of death worldwide. Despite the significant advances in the range of drugs and treatment modalities to treat cancer, the number of deaths is estimated to continue rising, posing serious challenges for the patients, their families, and the healthcare systems. Conventional treatments tend to be associated with severe adverse side effects and treatment resistance. Consequently, safer and more efficient therapy options are urgently needed, especially for the treatment of metastatic tumors refractory to conventional treatments. A new and revolutionizing field in oncology is immunotherapy, in which oncolytic viruses are included. Oncolytic viruses have an inherent or acquired selectivity to replicate exclusively in tumor cells, ultimately destroying them. Simultaneously, they also activate the dormant host’s immune system to fight against the tumor. Adenoviruses, particularly, have shown to be safe, inducing only mild adverse side effects in clinical trials, making them a great candidate for further clinical development. Adenoviruses can be genetically modified to increase their infectivity or improve the anti-cancer immune responses induced by the virus, e.g., through the expression of immunostimulatory molecules. The focus of this thesis was to develop and characterize several genetically modified oncolytic adenoviruses expressing either OX40L alone or OX40L and CD40L, two co-stimulatory molecules capable of engaging both the innate and adaptive arms of the immune system to fight the tumor. The insertion of the transgenes into the E3B-14.7k region of the Ad5/3-∆24 adenovector plasmid was performed using Gibson Assembly® cloning approach. After successful cloning, the recombinant viral genomes were transfected into A549 cells for viral amplification, followed by CsCl purification to produce a high titer viral preparation. The expression of the transgenes was studied in vitro by ELISA and functional assays, showing promising expression levels of functional OX40L and CD40L. However, when the infectivity and virus killing potency were analyzed, in vitro by immunocytochemistry and MTS assay; and in vivo using an immunodeficient mouse model, the data showed that the cloned viruses performed sub-optimally when compared to the control unarmed virus (Ad5/3-∆24). These findings suggest that the insertion of the two transgenes in place of the E3-14.7k gene was detrimental to the fitness of the virus. en
dc.language.iso eng
dc.publisher Helsingin yliopisto fi
dc.publisher University of Helsinki en
dc.publisher Helsingfors universitet sv
dc.subject cancer en
dc.subject oncolytic adenovirus en
dc.subject OX40L en
dc.subject CD40L en
dc.subject Gibson Assembly en
dc.title Generation and characterization of a novel oncolytic adenovirus expressing OX40L and CD40L en
dc.type.ontasot pro gradu -tutkielmat fi
dc.type.ontasot master's thesis en
dc.type.ontasot pro gradu-avhandlingar sv
dc.subject.discipline biotekniikka fi
dc.subject.discipline Biotechnology en
dc.subject.discipline bioteknik sv
dct.identifier.urn URN:NBN:fi:hulib-202006052630

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