Establishing a New Platform to Investigate the Efficacy of Oncolytic Virotherapy in a Human Ex Vivo Peritoneal Carcinomatosis Model.

Oncolytic virotherapy constitutes a promising treatment option for many solid cancers, including peritoneal carcinomatosis (PC), which still represents a terminal stage of many types of tumors. To date, the in vitro efficacy of oncolytic viruses is mostly tested in 2D-cultured tumor cell lines due to the lack of realistic 3D in vitro tumor models. We have investigated the feasibility of virotherapy as a treatment option for PC in a human ex vivo peritoneum co-culture model. Human HT-29 cancer cells stably expressing marker genes GFP and firefly luciferase (GFP/luc) were cultured on human peritoneum and infected with two prototypic oncolytic viruses (GLV-0b347 and MeV-DsRed). Both viral constructs were able to infect HT-29 cells in patient-derived peritoneum with high tumor specificity. Over time, both GFP signal and luciferase activity decreased substantially, thereby indicating successful virus-induced oncolysis. Furthermore, immunohistochemistry stainings showed specific virotherapeutic infections of HT-29 cells and effective tumor cell lysis in infected co-cultures. Thus, the PC model established here provides a clinically relevant screening platform to evaluate the therapeutic efficacy of virotherapeutic compounds and also to investigate, in an autologous setting, the immunostimulatory potential of oncolytic viruses for PC in a unique human model system superior to standard 2D in vitro models.

Case report: Immunovirotherapy as a novel add-on treatment in a patient with thoracic NUT carcinoma.

NUT carcinoma (NC) is a rare and extremely aggressive form of cancer, usually presenting with intrathoracic or neck manifestations in adolescents and young adults. With no established standard therapy regimen and a median overall survival of only 6.5 months, there is a huge need for innovative treatment options. As NC is genetically driven by a single aberrant fusion oncoprotein, it is generally characterized by a low tumor mutational burden, thus making it immunologically cold and insusceptible to conventional immunotherapy. Recently, we have demonstrated that oncolytic viruses (OVs) are able to specifically infect and lyse NC cells, thereby turning an immunologically cold tumor microenvironment into a hot one. Here, we report an intensive multimodal treatment approach employing for the first time an OV (talimogene laherparepvec (T-VEC); IMLYGIC®) together with the immune checkpoint inhibitor pembrolizumab as an add-on to a basic NC therapy (cytostatic chemotherapy, radiation therapy, epigenetic therapy) in a patient suffering from a large thoracic NC tumor which exhibits an aberrant, unique BRD3:NUTM1 fusion. This case demonstrates for the first time the feasibility of this innovative add-on immunovirotherapy regimen with a profound, repetitive and durable replication of T-VEC that is instrumental in achieving tumor stabilization and improvement in the patient´s quality of life. Further, a previously unknown BRD3:NUTM1 fusion gene was discovered that lacks the extraterminal domain of BRD3.

A Three-Dimensional Organoid Model of Primary Breast Cancer to Investigate the Effects of Oncolytic Virotherapy.

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.

Clinical Trials of Oncolytic Viruses in Breast Cancer.

Breast cancer is the second most common kind of cancer worldwide and oncolytic viruses may offer a new treatment approach. There are three different types of oncolytic viruses used in clinical trials; (i) oncolytic viruses with natural anti-neoplastic properties; (ii) oncolytic viruses designed for tumor-selective replication; (iii) oncolytic viruses modified to activate the immune system. Currently, fourteen different oncolytic viruses have been investigated in eighteen published clinical trials. These trials demonstrate that oncolytic viruses are well tolerated and safe for use in patients and display clinical activity. However, these trials mainly studied a small number of patients with different advanced tumors including some with breast cancer. Future trials should focus on breast cancer and investigate optimal routes of administration, occurrence of neutralizing antibodies, viral gene expression, combinations with other antineoplastic therapies, and identify subtypes that are particularly suitable for oncolytic virotherapy.

Automated techniques for visualization and mapping of articular cartilage in MR images of the osteoarthritic knee: a base technique for the assessment of microdamage and submicro damage.

The purpose of this paper is to describe automated techniques for the visualization and mapping of articular cartilage in magnetic resonance (MR) images of the osteoarthritic knee. The MR sequences and analysis software which will be described allow the assessment of cartilage damage using a range of standard scanners. With high field strength systems it would be possible, using these techniques, to assess micro-damage. The specific aim of the paper is to develop and validate software for automated segmentation and thickness mapping of articular cartilage from three-dimensional (3-D) gradient-echo MR images of the knee. The method can also be used for MR-based assessment of tissue engineered grafts. Typical values of cartilage thickness over seven defined regions can be obtained in patients with osteoarthritis (OA) and control subjects without OA. Three groups of patients were studied. The first group comprised patients with moderate OA in the age range 45-73 years. The second group comprised asymptomatic volunteers of 50-65 years; the third group, younger volunteers selected by clinical interview, history and X-ray. In this paper, sagittal 3-D spoiled-gradient steady-state acquisition images were obtained using a 1.5-T GE whole-body scanner with a specialist knee coil. For validation bovine and porcine cadaveric knees were given artificial cartilage lesions and then imaged. The animal validations showed close agreement between direct lesion measurements and those obtained from the MR images. The feasibility of semi-automated segmentation is demonstrated. Regional cartilage thickness values are seen as having practical application for fully automated detection of OA lesions even down to the submicrometer level.