The 16th international oncolytic virotherapy conference: from to clinical studies, novel approaches in cancer therapy meeting report.

The 16 International Oncolytic Virotherapy Conference, which was held in the vibrant city of Rotterdam, Netherlands, on 27-30 November 2024, featured a variety of novel strategies for improving oncolytic virus therapy. A reflective keynote highlighted industry challenges and lessons. Presentations covered the cutting-edge developments in oncolytic virotherapy from scientific, clinical, and industry perspectives.

Tumor Colonization and Therapy by Nissle 1917 Strain in Syngeneic Tumor-Bearing Mice Is Strongly Affected by the Gut Microbiome.

In the past, different bacterial species have been tested for cancer therapy in preclinical and clinical studies. The success of bacterial cancer therapy is mainly dependent on the ability of the utilized bacteria to overcome the host immune defense system to colonize the tumors and to initiate tumor-specific immunity. In recent years, several groups have demonstrated that the gut microbiome plays an important role of modulation of the host immune response and has an impact on therapeutic responses in murine models and in cohorts of human cancer patients.

Structural basis of the complete poxvirus transcription initiation process.

Poxviruses express their genes in the cytoplasm of infected cells using a virus-encoded multi-subunit polymerase (vRNAP) and unique transcription factors. We present cryo-EM structures that uncover the complete transcription initiation phase of the poxvirus vaccinia. In the pre-initiation complex, the heterodimeric early transcription factor VETFs/l adopts an arc-like shape spanning the polymerase cleft and anchoring upstream and downstream promoter elements.

Efficient Selection of Recombinant Fluorescent Vaccinia Virus Strains and Rapid Virus Titer Determination by Using a Multi-Well Plate Imaging System.

Engineered vaccinia virus (VACV) strains are used extensively as vectors for the development of novel cancer vaccines and cancer therapeutics. In this study, we describe for the first time a high-throughput approach for both fluorescent rVACV generation and rapid viral titer measurement with the multi-well plate imaging system, IncuCyteS3. The isolation of a single, well-defined plaque is critical for the generation of novel recombinant vaccinia virus (rVACV) strains.

Canine Adipose-Derived Mesenchymal Stem Cells (cAdMSCs) as a "Trojan Horse" in Vaccinia Virus Mediated Oncolytic Therapy against Canine Soft Tissue Sarcomas.

Several oncolytic viruses (OVs) including various human and canine adenoviruses, canine distemper virus, herpes-simplex virus, reovirus, and members of the poxvirus family, such as vaccinia virus and myxoma virus, have been successfully tested for canine cancer therapy in preclinical and clinical settings. The success of the cancer virotherapy is dependent on the ability of oncolytic viruses to overcome the attacks of the host immune system, to preferentially infect and lyse cancer cells, and to initiate tumor-specific immunity. To date, several different strategies have been developed to overcome the antiviral host defense barriers.

Structural Basis of Poxvirus Transcription: Vaccinia RNA Polymerase Complexes.

Poxviruses encode a multisubunit DNA-dependent RNA polymerase (vRNAP) that carries out viral gene expression in the host cytoplasm. We report cryo-EM structures of core and complete vRNAP enzymes from Vaccinia virus at 2.8 Å resolution.

Structural Basis of Poxvirus Transcription: Transcribing and Capping Vaccinia Complexes.

Poxviruses use virus-encoded multisubunit RNA polymerases (vRNAPs) and RNA-processing factors to generate mG-capped mRNAs in the host cytoplasm. In the accompanying paper, we report structures of core and complete vRNAP complexes of the prototypic Vaccinia poxvirus (Grimm et al., 2019; in this issue of Cell).

First-in-human study of TK-positive oncolytic vaccinia virus delivered by adipose stromal vascular fraction cells.

Background: ACAM2000, a thymidine kinase (TK)-positive strain of vaccinia virus, is the current smallpox vaccine in the US. Preclinical testing demonstrated potent oncolytic activity of ACAM2000 against several tumor types. This Phase I clinical trial of ACAM2000 delivered by autologous adipose stromal vascular fraction (SVF) cells was conducted to determine the safety and feasibility of such a treatment in patients with advanced solid tumors or acute myeloid leukemia (AML).

Vaccinia virus injected human tumors: oncolytic virus efficiency predicted by antigen profiling analysis fitted boolean models.

Virotherapy on the basis of oncolytic vaccinia virus (VACV) strains is a promising approach for cancer therapy. Recently, we showed that the oncolytic vaccinia virus GLV-1h68 has a therapeutic potential in treating human prostate and hepatocellular carcinomas in xenografted mice. In this study, we describe the use of dynamic boolean modeling for tumor growth prediction of vaccinia virus-injected human tumors.

Delivery of oncolytic vaccinia virus by matched allogeneic stem cells overcomes critical innate and adaptive immune barriers.

Background: Previous studies have identified IFNγ as an important early barrier to oncolytic viruses including vaccinia. The existing innate and adaptive immune barriers restricting oncolytic virotherapy, however, can be overcome using autologous or allogeneic mesenchymal stem cells as carrier cells with unique immunosuppressive properties.

Methods: To test the ability of mesenchymal stem cells to overcome innate and adaptive immune barriers and to successfully deliver oncolytic vaccinia virus to tumor cells, we performed flow cytometry and virus plaque assay analysis of ex vivo co-cultures of stem cells infected with vaccinia virus in the presence of peripheral blood mononuclear cells from healthy donors.

The ratio of ADSCs to HSC-progenitors in adipose tissue derived SVF may provide the key to predict the outcome of stem-cell therapy.

Background: Stromal vascular fraction (SVF) represents an attractive source of adult stem cells and progenitors, holding great promise for numerous cell therapy approaches. In 2017, it was reported that 1524 patients received autologous SVF following the enzymatic digestion of liposuction fat. The treatment was safe and effective and patients showed significant clinical improvement.

Phase I Trial of Intravenous Oncolytic Vaccinia Virus (GL-ONC1) with Cisplatin and Radiotherapy in Patients with Locoregionally Advanced Head and Neck Carcinoma.

Preclinical models have shown that the effectiveness of GL-ONC1, a modified oncolytic vaccinia virus, is enhanced by radiation and chemotherapy. The purpose of this study was to determine the safety of GL-ONC1 when delivered intravenously with chemoradiotherapy to patients with primary, nonmetastatic head and neck cancer. Patients with locoregionally advanced unresected, nonmetastatic carcinoma of the head/neck, excluding stage III-IVA p16-positive oropharyngeal cancers, were treated with escalating doses and cycles of intravenous GL-ONC1, along with radiotherapy and chemotherapy.

Humanized Mice with Subcutaneous Human Solid Tumors for Immune Response Analysis of Vaccinia Virus-Mediated Oncolysis.

Oncolytic vaccinia virus (VACV) therapy is an alternative cancer treatment modality that mediates targeted tumor destruction through a tumor-selective replication and an induction of anti-tumor immunity. We developed a humanized tumor mouse model with subcutaneous human tumors to analyze the interactions of VACV with the developing tumors and human immune system. A successful systemic reconstitution with human immune cells including functional T cells as well as development of tumors infiltrated with human T and natural killer (NK) cells was observed.

Colonization of xenograft tumors by oncolytic vaccinia virus (VACV) results in enhanced tumor killing due to the involvement of myeloid cells.

Background: The mechanisms by which vaccinia virus (VACV) interacts with the innate immune components are complex and involve different mechanisms. iNOS-mediated NO production by myeloid cells is one of the central antiviral mechanisms and this study aims to investigate specifically whether iNOS-mediated NO production by myeloid cells, is involved in tumor eradication following the virus treatment.

Methods: Human colon adenocarcinoma (HCT-116) xenograft tumors were infected by VACV.

Molecular network, pathway, and functional analysis of time-dependent gene changes associated with pancreatic cancer susceptibility to oncolytic vaccinia virotherapy.

Background: Pancreatic cancer is a fatal disease associated with resistance to conventional therapies. This study aimed to determine changes in gene expression patterns associated with infection and susceptibility of pancreatic cancer cells to an oncolyticvaccinia virus, GLV-1h153, carrying the human sodium iodide symporter for deep tissue imaging of virotherapy.

Methods: Replication and susceptibility of pancreatic adenocarcinoma PANC-1 cells to GLV-1h153 was confirmed with replication and cytotoxicity assays.

Recombinant vaccinia virus GLV-1h68 is a promising oncolytic vector in the treatment of cholangiocarcinoma.

Although early stage cholangiocarcinoma (CC) can be cured by surgical extirpation, the options for treatment of advanced stage CC are very few and suboptimal. Oncolytic virotherapy using replication-competent vaccinia virus (VACV) is a promising new strategy to treat human cancers. The ability of oncolytic VACV GLV-1h68 to infect, replicate in, and lyse three human CC cell lines was assayed in vitro and in subcutaneous flank xenografts in athymic nude mice.

Drug-Encoded Biomarkers for Monitoring Biological Therapies.

Blood tests are necessary, easy-to-perform and low-cost alternatives for monitoring of oncolytic virotherapy and other biological therapies in translational research. Here we assessed three candidate proteins with the potential to be used as biomarkers in biological fluids: two glucuronidases from E. coli (GusA) and Staphylococcus sp.

Preclinical Testing Oncolytic Vaccinia Virus Strain GLV-5b451 Expressing an Anti-VEGF Single-Chain Antibody for Canine Cancer Therapy.

Virotherapy on the basis of oncolytic vaccinia virus (VACV) strains is a novel approach for canine cancer therapy. Here we describe, for the first time, the characterization and the use of VACV strain GLV-5b451 expressing the anti-vascular endothelial growth factor (VEGF) single-chain antibody (scAb) GLAF-2 as therapeutic agent against different canine cancers. Cell culture data demonstrated that GLV-5b451 efficiently infected and destroyed all four tested canine cancer cell lines including: mammary carcinoma (MTH52c), mammary adenoma (ZMTH3), prostate carcinoma (CT1258), and soft tissue sarcoma (STSA-1).

Doxycycline Inducible Melanogenic Vaccinia Virus as Theranostic Anti-Cancer Agent.

We reported earlier the diagnostic potential of a melanogenic vaccinia virus based system in magnetic resonance (MRI) and optoacoustic deep tissue imaging (MSOT). Since melanin overproduction lead to attenuated virus replication, we constructed a novel recombinant vaccinia virus strain (rVACV), GLV-1h462, which expressed the key enzyme of melanogenesis (tyrosinase) under the control of an inducible promoter-system. In this study melanin production was detected after exogenous addition of doxycycline in two different tumor xenograft mouse models.

Microglia and astrocytes attenuate the replication of the oncolytic vaccinia virus LIVP 1.1.1 in murine GL261 gliomas by acting as vaccinia virus traps.

Background: Oncolytic virotherapy is a novel approach for the treatment of glioblastoma multiforme (GBM) which is still a fatal disease. Pathologic features of GBM are characterized by the infiltration with microglia/macrophages and a strong interaction between immune- and glioma cells. The aim of this study was to determine the role of microglia and astrocytes for oncolytic vaccinia virus (VACV) therapy of GBM.

Oncolytic gene therapy with recombinant vaccinia strain GLV-2b372 efficiently kills hepatocellular carcinoma.

Background: Hepatocellular carcinoma (HCC) commonly presents at a late stage when surgery is no longer a curative option. As such, novel therapies for advanced HCC are needed. Oncolytic viruses are a viable option for cancer therapy owing to their ability to specifically infect, replicate within, and kill cancer cells.

Gene therapy using therapeutic and diagnostic recombinant oncolytic vaccinia virus GLV-1h153 for management of colorectal peritoneal carcinomatosis.

Background: Peritoneal carcinomatosis (PC) is a terminal progression of colorectal cancer (CRC). Poor response to cytoreductive operation and chemotherapy coupled with the inability to reliably track disease progression by the use of established diagnostic methods, make this a deadly disease. We examined the effectiveness of the oncolytic vaccinia virus GLV-1h153 as a therapeutic and diagnostic vehicle.

Intratumoral INF-γ triggers an antiviral state in GL261 tumor cells: a major hurdle to overcome for oncolytic vaccinia virus therapy of cancer.

Oncolytic vaccinia virus (VACV) therapy is an alternative treatment option for glioblastoma multiforme. Here, we used a comparison of different tumor locations and different immunologic and genetic backgrounds to determine the replication efficacy and oncolytic potential of the VACV LIVP 1.1.

Expression of anti-VEGF antibody together with anti-EGFR or anti-FAP enhances tumor regression as a result of vaccinia virotherapy.

The tumor microenvironment plays an important role in tumor growth and progression. Here we demonstrate that vaccinia virus-mediated, constitutively expressed intratumoral antibodies against vascular endothelial growth factor (VEGF), epidermal growth factor receptor (EGFR), and fibroblast activation protein (FAP) significantly improved tumor regression and oncolytic virotherapy through suppression of angiogenesis, cell proliferation, and stromagenesis in virus-colonized tumors. In contrast to the tumor growth inhibition by the three tumor growth-inhibiting antibodies individually, when two of the three antibodies were expressed simultaneously by single vaccinia virus strains tumor regression was further enhanced.