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Feline immunodeficiency virus (FIV) is a lentivirus in the family Retroviridae that infects domestic cats resulting in an immunodeficiency disease featuring a progressive and profound decline in multiple sets of peripheral lymphocytes. Despite compelling evidence of FIV-associated immunopathology, there are conflicting data concerning the clinical effects of FIV infection on host morbidity and mortality. To explore FIV-associated immunopathogenesis and clinical disease, we experimentally inoculated a cohort of four specific pathogen-free kittens with a biological isolate of FIV clade C and continuously monitored these animals along with two uninfected control animals for more than thirteen years from the time of inoculation to the humane euthanasia endpoint. Here, we report the results obtained during the late asymptomatic and terminal phases of FIV infection in this group of experimentally FIV-infected cats.

The clinical impact of any therapy requires the product be safe and effective. Gammaretroviral vectors pose several unique risks, including inadvertent exposure to replication competent retrovirus (RCR) that can arise during vector manufacture. The US FDA has required patient monitoring for RCR, and the National Gene Vector Biorepository is an NIH resource that has assisted eligible investigators in meeting this requirement. To date, we have found no evidence of RCR in 338 pre-treatment and 1,595 post-treatment blood samples from 737 patients associated with 60 clinical trials. Most samples (75%) were obtained within 1 year of treatment, and samples as far out as 9 years after treatment were analyzed. The majority of trials (93%) were cancer immunotherapy, and 90% of the trials used vector products produced with the PG13 packaging cell line. The data presented here provide further evidence that current manufacturing methods generate RCR-free products and support the overall safety profile of retroviral gene therapy.

Metagenomic next-generation sequencing (mNGS) has enabled the high-throughput multiplexed identification of sequences from microbes of potential medical relevance. This approach has become indispensable for viral pathogen discovery and broad-based surveillance of emerging or re-emerging pathogens. From 2015 to 2019, plasma was collected from 9586 individuals in Cameroon and the Democratic Republic of the Congo enrolled in a combined hepatitis virus and retrovirus surveillance program. A subset (n = 726) of the patient specimens was analyzed by mNGS to identify viral co-infections. While co-infections from known blood-borne viruses were detected, divergent sequences from nine poorly characterized or previously uncharacterized viruses were also identified in two individuals. These were assigned to the following groups by genomic and phylogenetic analyses: densovirus, nodavirus, jingmenvirus, bastrovirus, dicistrovirus, picornavirus, and cyclovirus. Although of unclear pathogenicity, these viruses were found circulating at high enough concentrations in plasma for genomes to be assembled and were most closely related to those previously associated with bird or bat excrement. Phylogenetic analyses and in silico host predictions suggested that these are invertebrate viruses likely transmitted through feces containing consumed insects or through contaminated shellfish. This study highlights the power of metagenomics and in silico host prediction in characterizing novel viral infections in susceptible individuals, including those who are immunocompromised from hepatitis viruses and retroviruses, or potentially exposed to zoonotic viruses from animal reservoir species.

Methods to deliver gene editing agents in vivo as ribonucleoproteins could offer safety advantages over nucleic acid delivery approaches. We report the development and application of engineered DNA-free virus-like particles (eVLPs) that efficiently package and deliver base editor or Cas9 ribonucleoproteins. By engineering VLPs to overcome cargo packaging, release, and localization bottlenecks, we developed fourth-generation eVLPs that mediate efficient base editing in several primary mouse and human cell types. Using different glycoproteins in eVLPs alters their cellular tropism. Single injections of eVLPs into mice support therapeutic levels of base editing in multiple tissues, reducing serum Pcsk9 levels 78% following 63% liver editing, and partially restoring visual function in a mouse model of genetic blindness. In vitro and in vivo off-target editing from eVLPs was virtually undetected, an improvement over AAV or plasmid delivery. These results establish eVLPs as promising vehicles for therapeutic macromolecule delivery that combine key advantages of both viral and nonviral delivery.

Viruses in the family Retroviridae are found in a wide variety of vertebrate hosts. Enveloped virions are 80-100 nm in diameter with an inner core containing the viral genome and replicative enzymes. Core morphology is often characteristic for viruses within the same genus. Replication involves reverse transcription and integration into host cell DNA, resulting in a provirus. Integration into germline cells can result in a heritable provirus known as an endogenous retrovirus. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the family Retroviridae, which is available at ictv.global/report/retroviridae.

Bovine leukemia virus (BLV) is a member of the Retroviridae family and belongs to the Deltaretrovirus genus. It has a close relationship with human T-cell leukemia virus type I. BLV is responsible for causing enzootic bovine leukosis (EBL), a contagious disease that affects the bovine lymphatic system. This virus poses challenges for the global cattle industry, as it impacts cattle populations all over the world. Despite being widespread and impactful, BLV often goes unnoticed, with many researchers unaware of its presence and the potential consequences it carries. BLV demonstrates varying levels of pathogenicity. The majority of cattle (around 70%) become seropositive asymptomatic carriers, displaying no noticeable clinical symptoms. However, a smaller proportion of infected animals experience persistent lymphocytosis, characterized by an elevated number of lymphocytes in the bloodstream. If not monitored and managed, a subset of these persistently infected cattle may advance to lymphosarcoma. This condition typically presents as tumors in different lymphoid tissues, impacting various organs and overall health and productivity. Furthermore, recent research has highlighted the potential association between the occurrence of breast and lung cancer in humans and the presence of BLV. This review will delve into the recent discoveries concerning BLV, specifically exploring its epidemiology, the economic impact it has on the global cattle industry, its implications for human medicine, and the association between different alleles of the major histocompatibility complex (MHC) and susceptibility or resistance to BLV. [ABSTRACT FROM AUTHOR]

Host antiviral proteins engage in evolutionary arms races with viruses, in which both sides rapidly evolve at interaction interfaces to gain or evade immune defense. For example, primate TRIM5α uses its rapidly evolving v1 loop to bind retroviral capsids, and single mutations in this loop can dramatically improve retroviral restriction. However, it is unknown whether such gains of viral restriction are rare, or if they incur loss of pre-existing function against other viruses. Using deep mutational scanning, we comprehensively measured how single mutations in the TRIM5α v1 loop affect restriction of divergent retroviruses. Unexpectedly, we found that the majority of mutations increase weak antiviral function. Moreover, most random mutations do not disrupt potent viral restriction, even when it is newly acquired via a single adaptive substitution. Our results indicate that TRIM5αs adaptive landscape is remarkably broad and mutationally resilient, maximizing its chances of success in evolutionary arms races with retroviruses.

Introduction. The urgent problem of modern medicine is the fight against acute respiratory viral infections (ARVI). To combat ARVI, drugs of wide antiviral potency are needed, as well as immunomodulating drugs. Such antiviral and immunomodulatory effects has sodium deoxyribonucleate (DNA-Na) and its complex with iron (DNA-Na-Fe) developed on the basis of double-stranded DNA of natural origin. Aim of the study: To assess antiviral and virucidal activity of DNA-Na and DNA-Na-Fe against viruses of different kingdoms and families. Materials and methods. Antiviral and virucidal activity of DNA-Na and DNA-Na-Fe was assessed in cell cultures infected with viruses. Results and discussion. DNA-Na and DNA-Na-Fe had antiviral activity against adenovirus at concentrations of 2501000 mcg/ml. Antiviral effect of both drugs was not detected in case of poliovirus. DNA-Na and DNA-Na-Fe had antiviral activity against coronavirus in all administration schemes. EC50 for DNA-Na ~ 2500 mcg/ml, for DNA-Na-Fe ~ 1000 mcg/ml. In cells treated with DNA-Na-Fe, secretion of following proinflammatory cytokines was detected: Interleukin (IL) 1, IL-2, IL-6, IL-18, interferon- (IFN-), IFN-, as well as anti-inflammatory cytokines: IL-4, IL-10, antagonist of IL-1 receptor. Evidently, DNA-Na and DNA-Na-Fe have antiviral effect, but mechanism of action does not seem to be associated with specific effect on viral replication. Presence of virucidal activity of drugs against representatives of Coronaviridae, Adenoviridae, Picornaviridae, Retroviridae, Herpesviridae in vitro test in range of 1.03.0 lg TCID50 was identified. Conclusion. Presence of simultaneous antiviral and virucidal activity of DNA-Na and DNA-Na-Fe against adeno- and coronaviruses shows their prospects for prevention and treatment of ARVI.

Retroviruses evolved from long terminal repeat (LTR) retrotransposons by acquisition of envelope functions, and subsequently reinvaded host genomes. Together, endogenous retroviruses and LTR retrotransposons represent major components of animal, plant, and fungal genomes. Sequences from these elements have been exapted to perform essential host functions, including placental development, synaptic communication, and transcriptional regulation. They encode a Gag polypeptide, the capsid domains of which can oligomerize to form a virus-like particle. The structures of retroviral capsids have been extensively described. They assemble an immature viral particle through oligomerization of full-length Gag. Proteolytic cleavage of Gag results in a mature, infectious particle. In contrast, the absence of structural data on LTR retrotransposon capsids hinders our understanding of their function and evolutionary relationships. Here, we report the capsid morphology and structure of the archetypal Gypsy retrotransposon Ty3. We performed electron tomography (ET) of immature and mature Ty3 particles within cells. We found that, in contrast to retroviruses, these do not change size or shape upon maturation. Cryo-ET and cryo-electron microscopy of purified, immature Ty3 particles revealed an irregular fullerene geometry previously described for mature retrovirus core particles and a tertiary and quaternary arrangement of the capsid (CA) C-terminal domain within the assembled capsid that is conserved with mature HIV-1. These findings provide a structural basis for studying retrotransposon capsids, including those domesticated in higher organisms. They suggest that assembly via a structurally distinct immature capsid is a later retroviral adaptation, while the structure of mature assembled capsids is conserved between LTR retrotransposons and retroviruses.

Retroviral replicating vectors (RRVs) have been shown to achieve efficient tumor transduction and enhanced therapeutic benefits in a variety of cancer models. In the present study, we evaluated a possible combinatorial effect of prodrug activator genes delivered by two different RRVs derived from amphotropic murine leukemia virus (AMLV) and gibbon ape leukemia virus (GALV) on human hepatocellular carcinoma Hep3B cells. Both RRVs showed efficient replicative spread in culture and can overcame superinfection resistance each other. Notably, the replication and spread of each RRV in culture remained unaffected by pretransduction with the counterpart RRV. We further transduced cells with RRVs which individually possessed the prodrug activator genes yeast cytosine deaminase (CD) and herpes simplex virus thymidine kinase (TK) alone or in combination, and evaluated the cytotoxic effects of RRV-mediated gene therapy with CD and TK in the presence of the respective prodrugs, 5-fluorocytosine and ganciclovir. All combinations of the two prodrug activator genes produced synergistic cytocidal effects, but the combined effects of the different genes were significantly greater than those of the same genes when delivered by two different vectors. The present findings indicate the potential utility of dual-vector gene therapy using two different RRVs carrying different prodrug activator genes.

O artigo de opnião de Hector Seaunez Abreu, Chefe do Serviço de Genética do Instituto Nacional de Câncer (INCA), apresenta um breve resumo e atualização sobre a temática: Oncogenes e a Etiologia do Câncer.

Capture of retroviral envelope genes from endogenous retroviruses has played a role in the evolution of mammals, with evidence for the involvement of these genes in the formation of the maternofetal interface of the placenta. It has been shown that the diversity of captured genes is likely to be responsible for the diversity of placental structures, ranging from poorly invasive (epitheliochorial) to highly invasive (hemochorial), with an intermediate state (endotheliochorial) as found in carnivorans. The latter recapitulate part of this evolution, with the hyena being the sole carnivoran with a hemochorial placenta. In this study, we performed RNA sequencing on hyena placental transcripts and searched for endogenous retroviral envelope genes that have been captured specifically in the Hyaenidae clade and are not found in any other carnivoran. We identified an envelope gene that is expressed in the placenta at the level of the maternofetal interface, as evidenced by in situ hybridization/immunohistochemistry. The gene entry is coincidental with the emergence of the Hyaenidae clade 30 million years ago (Mya), being found at the same genomic locus in all 4 extant hyena species. Its coding sequence has further been maintained during all of Hyaenidae evolution. It is not found in any of the 30 other carnivorans-both Felidae and Canidae-that we screened. This envelope protein does not disclose any fusogenic activity in ex vivo assays, at variance with the syncytin-Car1 gene, which is found in all carnivorans, including the hyena, in which it is still present, transcriptionally active in the placenta, and fusogenic. Together, the present results illustrate the permanent renewal of placenta-specific genes by retroviral capture and de facto provide a candidate gene for the endotheliochorial to hemochorial transition of Hyaenidae among carnivorans.IMPORTANCE The placenta is the most diverse organ among mammals, due in part to stochastic capture of retroviral envelope genes. In carnivorans, capture of syncytin-Car1 took place 80 Mya. It is fusogenic, expressed at the syncytialized placental maternofetal interface, and conserved among all carnivorans, consistent with their shared endotheliochorial placenta. Hyenas are a remarkable exception, with a highly invasive hemochorial placenta, as found in humans, where disruption of maternal blood vessels results in maternal blood bathing the syncytial maternofetal interface. In this study, we identified a retroviral envelope gene capture and exaptation that took place about 30 Mya and is coincident with the emergence of the Hyaenidae, being conserved in all extant hyena species. It is expressed at the maternofetal interface in addition to the shared syncytin-Car1 gene. This new env gene, not present in any other carnivoran, is a likely candidate to be responsible for the specific structure of the hyena placenta.

Drugs that modify the epigenome are powerful tools for treating cancer, but these drugs often have pleiotropic effects, and identifying patients who will benefit from them remains a major clinical challenge. Here we show that medulloblastomas driven by the transcription factor Gfi1 are exquisitely dependent on the enzyme lysine demethylase 1 (Kdm1a/Lsd1). We demonstrate that Lsd1 physically associates with Gfi1, and that these proteins cooperate to inhibit genes involved in neuronal commitment and differentiation. We also show that Lsd1 is essential for Gfi1-mediated transformation: Gfi1 proteins that cannot recruit Lsd1 are unable to drive tumorigenesis, and genetic ablation of Lsd1 markedly impairs tumor growth in vivo. Finally, pharmacological inhibitors of Lsd1 potently inhibit growth of Gfi1-driven tumors. These studies provide important insight into the mechanisms by which Gfi1 contributes to tumorigenesis, and identify Lsd1 inhibitors as promising therapeutic agents for Gfi1-driven medulloblastoma.

BackgroundVocimagene amiretrorepvec (Toca 511) is an investigational gamma-retroviral replicating vector encoding cytosine deaminase that, when used in combination with extended-release 5-fluorocytosine (Toca FC), results preclinically in local production of 5-fluorouracil, depletion of immune-suppressive myeloid cells, and subsequent induction of antitumor immunity. Recurrent high-grade glioma (rHGG) patients have a high unmet need for effective therapies that produce durable responses lasting more than 6 months. In this setting, relapse is nearly universal and most responses are transient.MethodsIn this Toca 511 ascending-dose phase I trial (NCT01470794), HGG patients who recurred after standard of care underwent surgical resection and received Toca 511 injected into the resection cavity wall, followed by orally administered cycles of Toca FC.ResultsAmong 56 patients, durable complete responses were observed. A subgroup was identified based on Toca 511 dose and entry requirements for the follow-up phase III study. In this subgroup, which included both isocitrate dehydrogenase 1 (IDH1) mutant and wild-type tumors, the durable response rate is 21.7%. Median duration of follow-up for responders is 35.7+ months. As of August 25, 2017, all responders remain in response and are alive 33.9+ to 52.2+ months after Toca 511 administration, suggesting a positive association of durable response with overall survival.ConclusionsMultiyear durable responses have been observed in rHGG patients treated with Toca 511 + Toca FC in a phase I trial, and the treatment will be further evaluated in a randomized phase III trial. Among IDH1 mutant patients treated at first recurrence, there may be an enrichment of complete responders.

Toca 511, a retroviral replicating vector (RRV) encoding the yeast cytosine deaminase (yCD) prodrug activator gene, which mediates conversion of the prodrug 5-fluorocytosine (5-FC) to the anticancer drug 5-fluorouracil (5-FU), is currently being evaluated in Phase II/III clinical trials for glioma, and showing highly promising evidence of therapeutic activity. Here we evaluated RRV-mediated prodrug activator gene therapy as a new therapeutic approach for pancreatic ductal adenocarcinoma (PDAC). RRV spread rapidly and conferred significant cytotoxicity with prodrug in a panel of PDAC cells. Efficient intratumoral replication and complete inhibition of tumor growth upon 5-FC administration were observed in both immunodeficient and immunocompetent subcutaneous PDAC models. Biodistribution of RRV was highly restricted in normal tissues, especially in immunocompetent hosts. Tumor growth inhibition by Toca 511 followed by 5-FC was also confirmed in the orthotopic PDAC model. This study provides the first proof-of-concept for application of Toca 511 and Toca FC (extended release 5-FC) to the treatment of human PDAC, and provided support for inclusion of PDAC in a Phase I study evaluating Toca 511 in various systemic malignancies, (NCT02576665), which has recently been initiated.

The production of vertebrate retinal projection neurons, retinal ganglion cells (RGCs), is regulated by cell-intrinsic determinants and cell-to-cell signaling events. The basic-helix-loop-helix (bHLH) protein Atoh7 is a key neurogenic transcription factor required for RGC development. Here, we investigate whether manipulating human ATOH7 expression among uncommitted progenitors can promote RGC fate specification and thus be used as a strategy to enhance RGC genesis. Using the chicken retina as a model, we show that cell autonomous expression of ATOH7 is sufficient to induce precocious RGC formation and expansion of the neurogenic territory. ATOH7 overexpression among neurogenic progenitors significantly enhances RGC production at the expense of reducing the progenitor pool. Furthermore, forced expression of ATOH7 leads to a minor increase of cone photoreceptors. We provide evidence that elevating ATOH7 levels accelerates cell cycle progression from S to M phase and promotes cell cycle exit. We also show that ATOH7-induced ectopic RGCs often exhibit aberrant axonal projection patterns and are correlated with increased cell death during the period of retinotectal connections. These results demonstrate the high potency of human ATOH7 in promoting early retinogenesis and specifying the RGC differentiation program, thus providing insight for manipulating RGC production from stem cell-derived retinal organoids.