EU surveys insights: analytical tools, future directions, and the essential requirement for reference materials in wastewater monitoring of SARS-CoV-2, antimicrobial resistance and beyond.

BACKGROUND: Wastewater surveillance (WWS) acts as a vigilant sentinel system for communities, analysing sewage to protect public health by detecting outbreaks and monitoring trends in pathogens and contaminants. To achieve a thorough comprehension of present and upcoming practices and to identify challenges and opportunities for standardisation and improvement in WWS methodologies, two EU surveys were conducted targeting over 750 WWS laboratories across Europe and other regions. The first survey explored a diverse range of activities currently undertaken or planned by laboratories. The second survey specifically targeted methods and quality controls utilised for SARS-CoV-2 surveillance. RESULTS: The findings of the two surveys provide a comprehensive insight into the procedures and methodologies applied in WWS. In Europe, WWS primarily focuses on SARS-CoV-2 with 99% of the survey participants dedicated to this virus. However, the responses highlighted a lack of standardisation in the methodologies employed for monitoring SARS-CoV-2. The surveillance of other pathogens, including antimicrobial resistance, is currently fragmented and conducted by only a limited number of laboratories. Notably, these activities are anticipated to expand in the future. Survey replies emphasise the collective recognition of the need to enhance the accuracy of results in WWS practices, reflecting a shared commitment to advancing precision and effectiveness in WWS methodologies. CONCLUSIONS: These surveys identified a lack of standardised common procedures in WWS practices and the need for quality standards and reference materials to enhance the accuracy and reliability of WWS methods in the future. In addition, it is important to broaden surveillance efforts beyond SARS-CoV-2 to include other emerging pathogens and antimicrobial resistance to ensure a comprehensive approach to protecting public health.

SHOX triggers the lysosomal pathway of apoptosis via oxidative stress.

The SHOX gene encodes for a transcription factor important for normal bone development. Mutations in the gene are associated with idiopathic short stature and are responsible for the growth failure and skeletal defects found in the majority of patients with Léri-Weill dyschondrosteosis (LWD) and Langer mesomelic dysplasia. SHOX is expressed in growth plate chondrocytes where it is supposed to modulate the proliferation, differentiation and cell death of these cells. Supporting this hypothesis, in vitro studies have shown that SHOX expression induces cell cycle arrest and apoptosis in both transformed and primary cells. In this study, we further characterized the cell death mechanisms triggered by SHOX and compared them with the effects induced by one clinically relevant mutant form of SHOX, detected in LWD patients (SHOX R153L) and a SHOX C-terminally truncated version (L185X). We show that SHOX expression in U2OS osteosarcoma cells leads to oxidative stress that, in turn, induces lysosomal membrane rupture with release of active cathepsin B to the cytosol and subsequent activation of the intrinsic apoptotic pathway characterized by mitochondrial membrane permeabilization and caspase activation. Importantly, cells expressing SHOX R153L or L185X did not display any of these features. Given the fact that many of the events observed in SHOX-expressing cells also characterize the complex cell death process occurring in the growth plate during endochondral ossification, our findings further support the hypothesis that SHOX may play a central role in the regulation of the cell death pathways activated during long bone development.

Oncolytic parvoviruses: from basic virology to clinical applications.

Accumulated evidence gathered over recent decades demonstrated that some members of the Parvoviridae family, in particular the rodent protoparvoviruses H-1PV, the minute virus of mice and LuIII have natural anticancer activity while being nonpathogenic to humans. These studies have laid the foundations for the launch of a first phase I/IIa clinical trial, in which the rat H-1 parvovirus is presently undergoing evaluation for its safety and first signs of efficacy in patients with glioblastoma multiforme. After a brief overview of the biology of parvoviruses, this review focuses on the studies which unraveled the antineoplastic properties of these agents and supported their clinical use as anticancer therapeutics. Furthermore, the development of novel parvovirus-based anticancer strategies with enhanced specificity and efficacy is discussed, in particular the development of second and third generation vectors and the combination of parvoviruses with other anticancer agents. Lastly, we address the key challenges that remain towards a more rational and efficient use of oncolytic parvoviruses in clinical settings, and discuss how a better understanding of the virus life-cycle and of the cellular factors involved in virus infection, replication and cytotoxicity may promote the further development of parvovirus-based anticancer therapies, open new prospects for treatment and hopefully improve clinical outcome.

Immunotherapeutic Strategies for the Treatment of Glioblastoma: Current Challenges and Future Perspectives.

Despite decades of research and the best up-to-date treatments, grade 4 Glioblastoma (GBM) remains uniformly fatal with a patient median overall survival of less than 2 years. Recent advances in immunotherapy have reignited interest in utilizing immunological approaches to fight cancer. However, current immunotherapies have so far not met the anticipated expectations, achieving modest results in their journey from bench to bedside for the treatment of GBM. Understanding the intrinsic features of GBM is of crucial importance for the development of effective antitumoral strategies to improve patient life expectancy and conditions. In this review, we provide a comprehensive overview of the distinctive characteristics of GBM that significantly influence current conventional therapies and immune-based approaches. Moreover, we present an overview of the immunotherapeutic strategies currently undergoing clinical evaluation for GBM treatment, with a specific emphasis on those advancing to phase 3 clinical studies. These encompass immune checkpoint inhibitors, adoptive T cell therapies, vaccination strategies (i.e., RNA-, DNA-, and peptide-based vaccines), and virus-based approaches. Finally, we explore novel innovative strategies and future prospects in the field of immunotherapy for GBM.

Biomarker screen for efficacy of oncolytic virotherapy in patient-derived pancreatic cancer cultures.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is a tumour entity with unmet medical need. To assess the therapeutic potential of oncolytic virotherapy (OVT) against PDAC, different oncolytic viruses (OVs) are currently investigated in clinical trials. However, systematic comparisons of these different OVs in terms of efficacy against PDAC and biomarkers predicting therapeutic response are lacking. METHODS: We screened fourteen patient-derived PDAC cultures which reflect the intra- and intertumoural heterogeneity of PDAC for their sensitivity to five clinically relevant OVs, namely serotype 5 adenovirus Ad5-hTERT, herpes virus T-VEC, measles vaccine strain MV-NIS, reovirus jin-3, and protoparvovirus H-1PV. Live cell analysis, quantification of viral genome/gene expression, cell viability as well as cytotoxicity assays and titration of viral progeny were conducted. Transcriptome profiling was employed to identify potential predictive biomarkers for response to OV treatment. FINDINGS: Patient-derived PDAC cultures showed individual response patterns to OV treatment. Twelve of fourteen cultures were responsive to at least one OV, with no single OV proving superior or inferior across all cultures. Known host factors for distinct viruses were retrieved as potential biomarkers. Compared to the classical molecular subtype, the quasi-mesenchymal or basal-like subtype of PDAC was found to be more sensitive to H-1PV, jin-3, and T-VEC. Generally, expression of viral entry receptors did not correlate with sensitivity to OV treatment, with one exception: Expression of Galectin-1 (LGALS1), a factor involved in H-1PV entry, positively correlated with H-1PV induced cell killing. Rather, cellular pathways controlling immunological, metabolic and proliferative signaling appeared to determine outcome. For instance, high baseline expression of interferon-stimulated genes (ISGs) correlated with relative resistance to oncolytic measles virus, whereas low cyclic GMP-AMP synthase (cGAS) expression was associated with exceptional response. Combination treatment of MV-NIS with a cGAS inhibitor improved tumour cell killing in several PDAC cultures and cells overexpressing cGAS were found to be less sensitive to MV oncolysis. INTERPRETATION: Considering the heterogeneity of PDAC and the complexity of biological therapies such as OVs, no single biomarker can explain the spectrum of response patterns. For selection of a particular OV, PDAC molecular subtype, ISG expression as well as activation of distinct signaling and metabolic pathways should be considered. Combination therapies can overcome resistance in specific constellations. Overall, oncolytic virotherapy is a viable treatment option for PDAC, which warrants further development. This study highlights the need for personalised treatment in OVT. By providing all primary data, this study provides a rich source and guidance for ongoing developments. FUNDING: German National Science Foundation (Deutsche Forschungsgemeinschaft, DFG), German Cancer Aid (Deutsche Krebshilfe), German National Academic Scholarship Foundation (Studienstiftung des deutschen Volkes), Survival with Pancreatic Cancer Foundation.

Generation of an adenovirus-parvovirus chimera with enhanced oncolytic potential.

In this study, our goal was to generate a chimeric adenovirus-parvovirus (Ad-PV) vector that combines the high-titer and efficient gene transfer of adenovirus with the anticancer potential of rodent parvovirus. To this end, the entire oncolytic PV genome was inserted into a replication-defective E1- and E3-deleted Ad5 vector genome. As we found that parvoviral NS expression inhibited Ad-PV chimera production, we engineered the parvoviral P4 early promoter, which governs NS expression, by inserting into its sequence tetracycline operator elements. As a result of these modifications, P4-driven expression was blocked in the packaging T-REx-293 cells, which constitutively express the tetracycline repressor, allowing high-yield chimera production. The chimera effectively delivered the PV genome into cancer cells, from which fully infectious replication-competent parvovirus particles were generated. Remarkably, the Ad-PV chimera exerted stronger cytotoxic activities against various cancer cell lines, compared with the PV and Ad parental viruses, while being still innocuous to a panel of tested healthy primary human cells. This Ad-PV chimera represents a novel versatile anticancer agent which can be subjected to further genetic manipulations in order to reinforce its enhanced oncolytic capacity through arming with transgenes or retargeting into tumor cells.

Retargeting of rat parvovirus H-1PV to cancer cells through genetic engineering of the viral capsid.

The rat parvovirus H-1PV is a promising anticancer agent given its oncosuppressive properties and the absence of known side effects in humans. H-1PV replicates preferentially in transformed cells, but the virus can enter both normal and cancer cells. Uptake by normal cells sequesters a significant portion of the administered viral dose away from the tumor target. Hence, targeting H-1PV entry specifically to tumor cells is important to increase the efficacy of parvovirus-based treatments. In this study, we first found that sialic acid plays a key role in H-1PV entry. We then genetically engineered the H-1PV capsid to improve its affinity for human tumor cells. By analogy with the resolved crystal structure of the closely related parvovirus minute virus of mice, we developed an in silico three-dimensional (3D) model of the H-1PV wild-type capsid. Based on this model, we identified putative amino acids involved in cell membrane recognition and virus entry at the level of the 2-fold axis of symmetry of the capsid, within the so-called dimple region. In situ mutagenesis of these residues significantly reduced the binding and entry of H-1PV into permissive cells. We then engineered an entry-deficient viral capsid and inserted a cyclic RGD-4C peptide at the level of its 3-fold axis spike. This peptide binds α(v)ß(3) and α(v)ß(5) integrins, which are overexpressed in cancer cells and growing blood vessels. The insertion of the peptide rescued viral infectivity toward cells overexpressing α(v)ß(5) integrins, resulting in the efficient killing of these cells by the reengineered virus. This work demonstrates that H-1PV can be genetically retargeted through the modification of its capsid, showing great promise for a more efficient use of this virus in cancer therapy.

The XVIII International Parvovirus Workshop Rimini, Italy, 14-17 June 2022.

The XVIII International Parvovirus Workshop took place in Rimini, Italy, from 14 to 17 June 2022 as an on-site event, continuing the series of meetings started in 1985 and continuously held every two years. The communications dealt with all aspects of research in the field, from evolution and structure to receptors, from replication to trafficking, from virus-host interactions to clinical and veterinarian virology, including translational issues related to viral vectors, gene therapy and oncolytic parvoviruses. The oral communications were complemented by a poster exhibition available for view and discussion during the whole meeting. The XVIII International Parvovirus Workshop was dedicated to the memory of our dearest colleague Mavis Agbandje-McKenna (1963-2021).

New RT-PCR Assay for the Detection of Current and Future SARS-CoV-2 Variants.

Multiple lineages of SARS-CoV-2 have been identified featuring distinct sets of genetic changes that confer to the virus higher transmissibility and ability to evade existing immunity. The continuous evolution of SARS-CoV-2 may pose challenges for current treatment options and diagnostic tools. In this study, we have first evaluated the performance of the 14 WHO-recommended real-time reverse transcription (RT)-PCR assays currently in use for the detection of SARS-CoV-2 and found that only one assay has reduced performance against Omicron. We then developed a new duplex real-time RT-PCR assay based on the amplification of two ultra-conserved elements present within the SARS-CoV-2 genome. The new duplex assay successfully detects all of the tested SARS-CoV-2 variants of concern (including Omicron sub-lineages BA.4 and BA.5) from both clinical and wastewater samples with high sensitivity and specificity. The assay also functions as a one-step droplet digital RT-PCR assay. This new assay, in addition to clinical testing, could be adopted in surveillance programs for the routine monitoring of SARS-CoV-2's presence in a population in wastewater samples. Positive results with our assay in conjunction with negative results from an Omicron-specific assay may provide timely indication of the emergence of a novel SARS-CoV-2 variant in a certain community and thereby aid public health interventions.

A qualitative RT-PCR assay for the specific identification of the SARS-CoV-2 B.1.1.529 (Omicron) Variant of Concern.

OBJECTIVES: The aim of this study was to develop a RT-PCR assay for the specific detection of the SARS-CoV-2 Omicron Variant of Concern (VOC) as a rapid alternative to sequencing. METHODS: A RT-PCR was designed in silico and then validated using characterised clinical samples containing Omicron (both BA.1 and BA.2 lineages) and the Omicron synthetic RNA genome. As negative controls, SARS-CoV-2 positive clinical samples collected in May 2020, and synthetic RNA genomes of the isolate Wuhan Hu-1 and of the Alpha (B.1.1.7), Beta (B.1.351), Gamma (P.1), Kappa (B.1.617.1), Iota (B.1.526), Epsilon (B.1.429) and Delta (B.1.617.2) SARS-CoV-2 VOC were used. RESULTS: Experiments performed using as templates the synthetic RNA genomes demonstrate the high specificity of the PCR-method for the SARS-CoV-2 Omicron. Despite the synthetic RNAs were used at high copy numbers, specific signal was mainly detected with the Omicron synthetic genome. Only a non-specific late signal was detected using the Alpha variant genome, but these results were considered negligible as Alpha VOC has been replaced by the Delta and it is not circulating anymore in the world. Using our method, we confirmed the presence of Omicron on clinical samples containing this variant but not of other SARS-CoV-2 lineages. The method is highly sensitive and can detect up to 1 cp of the Omicron virus per µl. CONCLUSIONS: The method presented here, in combination with other methods in use for detection of SARS-CoV-2, can be used for an early identification of Omicron.

Oncolytic H-1 Parvovirus Hijacks Galectin-1 to Enter Cancer Cells.

Clinical studies in glioblastoma and pancreatic carcinoma patients strongly support the further development of H-1 protoparvovirus (H-1PV)-based anticancer therapies. The identification of cellular factors involved in the H-1PV life cycle may provide the knowledge to improve H-1PV anticancer potential. Recently, we showed that sialylated laminins mediate H-1PV attachment at the cell membrane. In this study, we revealed that H-1PV also interacts at the cell surface with galectin-1 and uses this glycoprotein to enter cancer cells. Indeed, knockdown/out of LGALS1, the gene encoding galectin-1, strongly decreases the ability of H-1PV to infect and kill cancer cells. This ability is rescued by the re-introduction of LGALS1 into cancer cells. Pre-treatment with lactose, which is able to bind to galectins and modulate their cellular functions, decreased H-1PV infectivity in a dose dependent manner. In silico analysis reveals that LGALS1 is overexpressed in various tumours including glioblastoma and pancreatic carcinoma. We show by immunohistochemistry analysis of 122 glioblastoma biopsies that galectin-1 protein levels vary between tumours, with levels in recurrent glioblastoma higher than those in primary tumours or normal tissues. We also find a direct correlation between LGALS1 transcript levels and H-1PV oncolytic activity in 53 cancer cell lines from different tumour origins. Strikingly, the addition of purified galectin-1 sensitises poorly susceptible GBM cell lines to H-1PV killing activity by rescuing cell entry. Together, these findings demonstrate that galectin-1 is a crucial determinant of the H-1PV life cycle.

Through its nonstructural protein NS1, parvovirus H-1 induces apoptosis via accumulation of reactive oxygen species.

The rat parvovirus H-1 (H-1PV) attracts high attention as an anticancer agent, because it is not pathogenic for humans and has oncotropic and oncosuppressive properties. The viral nonstructural NS1 protein is thought to mediate H-1PV cytotoxicity, but its exact contribution to this process remains undefined. In this study, we analyzed the effects of the H-1PV NS1 protein on human cell proliferation and cell viability. We show that NS1 expression is sufficient to induce the accumulation of cells in G(2) phase, apoptosis via caspase 9 and 3 activation, and cell lysis. Similarly, cells infected with wild-type H-1PV arrest in G(2) phase and undergo apoptosis. Furthermore, we also show that both expression of NS1 and H-1PV infection lead to higher levels of intracellular reactive oxygen species (ROS), associated with DNA double-strand breaks. Antioxidant treatment reduces ROS levels and strongly decreases NS1- and virus-induced DNA damage, cell cycle arrest, and apoptosis, indicating that NS1-induced ROS are important mediators of H-1PV cytotoxicity.

Oncolytic Virus Immunotherapy.

Oncolytic viruses (OVs) were originally developed as direct cytotoxic agents but have been increasingly recognised as a form of immunotherapy [...].

Parvovirus-Based Combinatorial Immunotherapy: A Reinforced Therapeutic Strategy against Poor-Prognosis Solid Cancers.

Resistance to anticancer treatments poses continuing challenges to oncology researchers and clinicians. The underlying mechanisms are complex and multifactorial. However, the immunologically "cold" tumor microenvironment (TME) has recently emerged as one of the critical players in cancer progression and therapeutic resistance. Therefore, TME modulation through induction of an immunological switch towards inflammation ("warming up") is among the leading approaches in modern oncology. Oncolytic viruses (OVs) are seen today not merely as tumor cell-killing (oncolytic) agents, but also as cancer therapeutics with multimodal antitumor action. Due to their intrinsic or engineered capacity for overcoming immune escape mechanisms, warming up the TME and promoting antitumor immune responses, OVs hold the potential for creating a proinflammatory background, which may in turn facilitate the action of other (immunomodulating) drugs. The latter provides the basis for the development of OV-based immunostimulatory anticancer combinations. This review deals with the smallest among all OVs, the H-1 parvovirus (H-1PV), and focuses on H-1PV-based combinatorial approaches, whose efficiency has been proven in preclinical and/or clinical settings. Special focus is given to cancer types with the most devastating impact on life expectancy that urgently call for novel therapies.

Oncolytic H-1 parvovirus binds to sialic acid on laminins for cell attachment and entry.

H-1 parvovirus (H-1PV) is a promising anticancer therapy. However, in-depth understanding of its life cycle, including the host cell factors needed for infectivity and oncolysis, is lacking. This understanding may guide the rational design of combination strategies, aid development of more effective viruses, and help identify biomarkers of susceptibility to H-1PV treatment. To identify the host cell factors involved, we carry out siRNA library screening using a druggable genome library. We identify one crucial modulator of H-1PV infection: laminin γ1 (LAMC1). Using loss- and gain-of-function studies, competition experiments, and ELISA, we validate LAMC1 and laminin family members as being essential to H-1PV cell attachment and entry. H-1PV binding to laminins is dependent on their sialic acid moieties and is inhibited by heparin. We show that laminins are differentially expressed in various tumour entities, including glioblastoma. We confirm the expression pattern of laminin γ1 in glioblastoma biopsies by immunohistochemistry. We also provide evidence of a direct correlation between LAMC1 expression levels and H-1PV oncolytic activity in 59 cancer cell lines and in 3D organotypic spheroid cultures with different sensitivities to H-1PV infection. These results support the idea that tumours with elevated levels of γ1 containing laminins are more susceptible to H-1PV-based therapies.

Virotherapy in Germany-Recent Activities in Virus Engineering, Preclinical Development, and Clinical Studies.

Virotherapy research involves the development, exploration, and application of oncolytic viruses that combine direct killing of cancer cells by viral infection, replication, and spread (oncolysis) with indirect killing by induction of anti-tumor immune responses. Oncolytic viruses can also be engineered to genetically deliver therapeutic proteins for direct or indirect cancer cell killing. In this review-as part of the special edition on "State-of-the-Art Viral Vector Gene Therapy in Germany"-the German community of virotherapists provides an overview of their recent research activities that cover endeavors from screening and engineering viruses as oncolytic cancer therapeutics to their clinical translation in investigator-initiated and sponsored multi-center trials. Preclinical research explores multiple viral platforms, including new isolates, serotypes, or fitness mutants, and pursues unique approaches to engineer them towards increased safety, shielded or targeted delivery, selective or enhanced replication, improved immune activation, delivery of therapeutic proteins or RNA, and redirecting antiviral immunity for cancer cell killing. Moreover, several oncolytic virus-based combination therapies are under investigation. Clinical trials in Germany explore the safety and potency of virotherapeutics based on parvo-, vaccinia, herpes, measles, reo-, adeno-, vesicular stomatitis, and coxsackie viruses, including viruses encoding therapeutic proteins or combinations with immune checkpoint inhibitors. These research advances represent exciting vantage points for future endeavors of the German virotherapy community collectively aimed at the implementation of effective virotherapeutics in clinical oncology.

A Roadmap for the Success of Oncolytic Parvovirus-Based Anticancer Therapies.

Autonomous rodent protoparvoviruses (PVs) are promising anticancer agents due to their excellent safety profile, natural oncotropism, and oncosuppressive activities. Viral infection can trigger immunogenic cell death, activating the immune system against the tumor. However, the efficacy of this treatment in recent clinical trials is moderate compared with results seen in preclinical work. Various strategies have been employed to improve the anticancer activities of oncolytic PVs, including development of second-generation parvoviruses with enhanced oncolytic and immunostimulatory activities and rational combination of PVs with other therapies. Understanding the cellular factors involved in the PV life cycle is another important area of investigation. Indeed, these studies may lead to the identification of biomarkers that would allow a more personalized use of PV-based therapies. This review focuses on this work and the challenges that still need to be overcome to move PVs forward into clinical practice as an effective therapeutic option for cancer patients.

Suicide gene therapy for the treatment of high-grade glioma: past lessons, present trends, and future prospects.

Suicide gene therapy has represented an experimental cancer treatment modality for nearly 40 years. Among the various cancers experimentally treated by suicide gene therapy, high-grade gliomas have been the most prominent both in preclinical and clinical settings. Failure of a number of promising suicide gene therapy strategies in the clinic pointed toward a bleak future of this approach for the treatment of high-grade gliomas. Nevertheless, the development of new vectors and suicide genes, better prodrugs, more efficient delivery systems, and new combinatorial strategies represent active research areas that may eventually lead to better efficacy of suicide gene therapy. These trends are evident by the current increasing focus on suicide gene therapy for high-grade glioma treatment both in the laboratory and in the clinic. In this review, we give an overview of different suicide gene therapy approaches for glioma treatment and discuss clinical trials, delivery issues, and immune responses.

Oncolytic H-1 Parvovirus Enters Cancer Cells through Clathrin-Mediated Endocytosis.

H-1 protoparvovirus (H-1PV) is a self-propagating virus that is non-pathogenic in humans and has oncolytic and oncosuppressive activities. H-1PV is the first member of the Parvoviridae family to undergo clinical testing as an anticancer agent. Results from clinical trials in patients with glioblastoma or pancreatic carcinoma show that virus treatment is safe, well-tolerated and associated with first signs of efficacy. Characterisation of the H-1PV life cycle may help to improve its efficacy and clinical outcome. In this study, we investigated the entry route of H-1PV in cervical carcinoma HeLa and glioma NCH125 cell lines. Using electron and confocal microscopy, we detected H-1PV particles within clathrin-coated pits and vesicles, providing evidence that the virus uses clathrin-mediated endocytosis for cell entry. In agreement with these results, we found that blocking clathrin-mediated endocytosis using specific inhibitors or small interfering RNA-mediated knockdown of its key regulator, AP2M1, markedly reduced H-1PV entry. By contrast, we found no evidence of viral entry through caveolae-mediated endocytosis. We also show that H-1PV entry is dependent on dynamin, while viral trafficking occurs from early to late endosomes, with acidic pH necessary for a productive infection. This is the first study that characterises the cell entry pathways of oncolytic H-1PV.

H-1 Parvovirus as a Cancer-Killing Agent: Past, Present, and Future.

The rat protoparvovirus H-1PV is nonpathogenic in humans, replicates preferentially in cancer cells, and has natural oncolytic and oncosuppressive activities. The virus is able to kill cancer cells by activating several cell death pathways. H-1PV-mediated cancer cell death is often immunogenic and triggers anticancer immune responses. The safety and tolerability of H-1PV treatment has been demonstrated in early clinical studies in glioma and pancreatic carcinoma patients. Virus treatment was associated with surrogate signs of efficacy including immune conversion of tumor microenvironment, effective virus distribution into the tumor bed even after systemic administration, and improved patient overall survival compared with historical control. However, monotherapeutic use of the virus was unable to eradicate tumors. Thus, further studies are needed to improve H-1PV's anticancer profile. In this review, we describe H-1PV's anticancer properties and discuss recent efforts to improve the efficacy of H-1PV and, thereby, the clinical outcome of H-1PV-based therapies.