Abstract
Background: Although several oncolytic viruses have already been tested in early-stage clinical studies of breast cancer, there is still an urgent need to develop patient-derived experimental systems that mimic the response of breast cancer to oncolytic agents in preparation of testing different oncolytic viruses in clinical trials. We addressed this need by developing a protocol to study the effects of oncolytic viruses in stable organoid cell cultures derived from breast cancer tissue. Methods: We used an established three-dimensional organoid model derived from tissue of 10 patients with primary breast cancer. We developed an experimental protocol for infecting organoid cultures with oncolytic viruses and compared the oncolytic effects of a measles vaccine virus (MeV) and a vaccinia virus (GLV) genetically engineered to express either green fluorescent protein (MeV-GFP) and red fluorescent protein (GLV-0b347), respectively, or a suicide gene encoding a fusion of cytosine deaminase with uracil phosphoribosyltransferase (MeV-SCD and GLV-1h94, respectively), thereby enabling enzymatic conversion of the prodrug 5-fluorocytosine (5-FC) into cytotoxic compounds 5-fluorouracil (5-FU) and 5-fluorouridine monophosphate (5-FUMP). Results: The method demonstrated that all oncolytic viruses significantly inhibited cell viability in organoid cultures derived from breast cancer tissue. The oncolytic effects of the oncolytic viruses expressing suicide genes (MeV-SCD and GLV-1h94) were further enhanced by virus-triggered conversion of the prodrug 5-FC to toxic 5-FU and toxic 5-FUMP. Conclusions: We were able to develop a protocol to assess the effects of two different types of oncolytic viruses in stable organoid cell cultures derived from breast cancer tissue. The greatest oncolytic effects were observed when the oncolytic viruses were engineered to express a suicide gene (MeV-SCD and GLV-1h94) in the presence of the prodrug 5-FC. The model therefore provides a promising in vitro method to help further testing and engineering of new generations of virotherapeutic vectors for in vivo use.
Highlights
Breast cancer is the most common cause of cancer-associated death in women aged between 20 and 59 years (Siegel et al, 2021)
In this study we set out to answer the question whether threedimensional cell cultures are suitable for testing oncolytic virotherapy. We addressed this topic by developing a protocol in a stable three-dimensional organoid model derived from patients with primary breast cancer to determine the oncolytic effects of genetically engineered oncolytic viruses, encoding either marker genes for GFP and for red fluorescent protein, or the super cytosine deaminase (SCD)/FCU1 suicide gene on breast cancer organoid cultures
Stable Organoid Cultures Prepared From Breast Cancer Tissues
Summary
Breast cancer is the most common cause of cancer-associated death in women aged between 20 and 59 years (Siegel et al, 2021). Oncolytic viruses are emerging as promising agents for the treatment of cancer because they selectively infect and damage cancerous tissues without causing harm to normal tissue (Russell et al, 2012). They offer an attractive combination of tumorspecific cell lysis coupled with immune stimulation through release of tumor antigens and/or other signals to overcome immunosuppression in the tumor microenvironment. Several oncolytic viruses have already been tested in early-stage clinical studies of breast cancer, there is still an urgent need to develop patient-derived experimental systems that mimic the response of breast cancer to oncolytic agents in preparation of testing different oncolytic viruses in clinical trials. We addressed this need by developing a protocol to study the effects of oncolytic viruses in stable organoid cell cultures derived from breast cancer tissue
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