\n
Spatial proximity evaluation of adenovirus foci and hypoxia<\/h3>\n
Animal experimentation was performed beneath moral approval from the UK Residence Workplace (Challenge license 30\/3391 and private license I5916669E). Feminine CB17-SCID mice (6\u20138 weeks outdated) have been inoculated subcutaneously in the correct flank with 2\u2009\u00d7\u2009106 DLD-1 cells in 50\u2009\u00b5L. On day 1, when xenografts reached 80\u2013120\u2009mm3, mice obtained 100\u2009\u00b5L of chlodronate liposomes (Chlodronateliposomes.org) intravenously to maximise subsequent adenovirus supply.54 EnAd-SA-fLuc reporter virus particles (2\u2009\u00d7\u20091010) have been injected through the tail vein on days 2 and 4. Throughout steady-state an infection, beforehand proven to be 16 days after the final remedy,12 mice have been injected intraperitoneally with pimonidazole at 60\u2009mg\u00b7kg\u22121 and have been culled 2\u2009h later. Tumors have been explanted, formalin-fixed, and paraffin-embedded.<\/p>\n
Eight pairs of serial tissue sections per tumor (4\u2009\u00b5m thick, ca. 40\u2009\u00b5m aside, 4 pairs for mouse 3) have been stained for hexon (horseradish peroxidase\/3,3\u00b4-diaminobenzidine), pimonidazole adducts (alkaline phosphatase\/Quick purple), and nuclei (hematoxylin) as beforehand described.12 Spatial proximity between hexon-positive adenovirus foci and hypoxic areas was analyzed utilizing a customized pipeline in Fiji\/ImageJ.55 Briefly, matched serial sections have been scanned at 20x (Aperio ImageScope, Leica), aligned, cropped to tissue boundaries, and cleaned of staining artifacts. Binary masks have been generated from color-deconvolved alerts utilizing depth thresholding. The pimonidazole masks was skeletonized to render an Euclidean distance map extending 200\u2009\u00b5m outward from the core\/skeleton of pimonidazole staining. Hexon- and hematoxylin-positive areas, representing nuclei in viable tumor areas, have been projected onto this map to extract the normalized pixel fraction as a operate of distance from the pimonidazole skeleton. An in depth framework is offered in Supplementary Fig. 1.<\/p>\n
Cell strains<\/h3>\n
DLD-1, SW480, SK-OV-3, HCT116, HT29 (ATCC), and AD293 (Agilent Applied sciences) have been cultured in DMEM (Sigma Aldrich) with 10% (v\/v) heat-inactivated fetal bovine serum (FBS, Gibco) at 37\u2009\u00b0C and 5% CO2. Cell strains have been maintained in a humidified incubator, routinely examined for mycoplasma (Lonza), and authenticated (Supply Bioscience).<\/p>\n
Hypoxia remedy<\/h3>\n
Hypoxia incubation (1% pO2) was carried out within the In vivo2 400 chamber (Baker Ruskin) utilizing media pre-equilibrated in a single day at 1% pO\u2082. Experiments utilized gas-permeable Lumox\u00ae plates (Sarstedt) on a zig-zag rocker (PMR-30, VWR) at 5 cycles per minute to keep up gasoline equilibrium. Exterior oxygen calibration of the chamber was carried out routinely. Pharmacological hypoxia was induced with 50\u2009\u00b5M FG4592 (Roxadustat, Cayman Chemical).<\/p>\n
Viruses and infections<\/h3>\n
EnAd-E1A-cFLAG, used for E1A expression evaluation, was generated utilizing a shuttle plasmid containing the E1A open studying body flanked by NdeI websites within the pColoAd2.4 vector.23 Cloning of this virus is summarized in Supplementary Figs. 3 and 4. EnAd-SA-fLuc, utilized in experiments monitoring viral life cycle, accommodates a firefly luciferase coupled to the main late transcription unit by a splice acceptor sequence.23 EnAd-CMV-BiTE and EnAd-SA-BiTE specific a deca-His-tagged EpCAM\/CD3-BiTE beneath a CMV promoter or built-in into the late transcription unit through a splice acceptor, respectively.25 Ad5-E1A-fLuc is a replication-competent reporter adenovirus encoding firefly luciferase fused to E1A.56 Virus shares have been concentrated and double-purified through cesium chloride gradient centrifugation, then titered by plaque assay on A549 cells as beforehand described.12<\/p>\n
Infections have been carried out in serum-free media for two\u2009h. For virus manufacturing, inocula have been eliminated, and cells have been washed twice with full medium. The inoculum was set to 125\u2009\u03bcL\u00b7cm\u22122 impartial of the seeding format, and MOIs have been based mostly on titers derived by plaque assay (Supplementary Desk 2).<\/p>\n
Quantification of infectious adenovirus particles utilizing bioluminescence<\/h3>\n
A high-dynamic-range technique was established to measure infectious viral particles by correlating gentle emitted by reporter adenoviruses of unknown focus with plaque assay-tittered requirements (Supplementary Fig. 2). Phenol red-free supernatants from virus manufacturing assays with EnAd-SA-fLuc have been collected, weighed, and cleared by centrifugation. Neat or diluted 25\u2009\u00b5L samples, together with a regular collection (5.57\u2009\u00d7\u2009105 PFU\u00b7mL\u22121 2.72\u2009\u00d7\u2009102 PFU\u00b7mL\u22121), have been used to contaminate 1.5\u2009\u00d7\u2009104 A549 cells seeded the day past in 50\u2009\u00b5L phenol red-free DMEM\u2009+\u200910% FBS (Gibco) on strong black 96-well plates (Corning). Plates have been then spun at 600\u00d7gn for 10\u2009min. At 20 to 24 hpi, luminescence was measured utilizing a PolarStar plate reader (BMG Labtech) after injecting 25\u2009\u00b5L of a 1.2\u2009mg\u00b7mL\u22121 D<\/span>-Luciferin resolution into the effectively (Gold Biotechnology). Bioluminescence was recorded over six kinetic cycles at 0.5\u2009s intervals, and virus concentrations have been calculated through four-parameter becoming of built-in luminescence (MARS, BMG Labtech). Not less than three technical replicates have been carried out for every an infection.<\/p>\nAssay for capsid integrity<\/h3>\n
To investigate the protein composition of adenovirus particles in supernatants, the tradition medium was centrifuged (300\u00d7gn, 5\u2009min), filtered (0.22\u2009\u00b5m), and concentrated utilizing a 300\u2009kDa cut-off ultrafiltration unit (Sartorius). The retentate was washed twice with PBS, and protein focus was measured utilizing the QuantiPro BCA Equipment (Thermo Fisher). Two micrograms of lysate in RIPA buffer (Thermo Fisher) have been separated as for immunoblotting, stained with SyproRuby (Thermo Fisher) and imaged on a Biorad Chemidoc imager.<\/p>\n
Absolute quantification of viral mRNA and genomic DNA by qPCR<\/h3>\n
DNA was extracted from contaminated cell pellets or supernatants utilizing the PureLink genomic DNA Mini Equipment (Invitrogen), and whole RNA utilizing RNeasy Mini Equipment (Qiagen). RNA integrity was routinely examined by capillary electrophoresis (Tape Station, Agilent). For cDNA synthesis, 500 to 800\u2009ng of RNA was transcribed utilizing the QuantiTect Equipment (Qiagen). PCR reactions (20\u2009\u00b5L) contained 10\u2009ng of cDNA or DNA with 2x qPCRBIO Probe Combine Hello-ROX (TaqMan assays, PCR Biosystems). Fiber, E1A, and E2B 200\u2009bp-standards for absolute quantification have been synthesized (IDT Applied sciences, Supplementary Fig. 7). Fiber detection was used to quantify genome copy quantity. TaqMan probes have been labeled with JOE or FAM and quenched with BHQ1 (Sigma). Measurements have been carried out on the StepOnePlus cycler (Utilized Biosciences). Copy numbers have been calculated by linear regression on normal curves (Supplementary Fig. 7). Primer sequences can be found in Supplementary Desk 3.<\/p>\n
To quantify encapsidated genomes, 200\u2009\u00b5L of contaminated supernatants have been handled with 100U Benzonase (Millipore) for 3\u2009h at 37\u2009\u00b0C. Earlier than DNA extraction and fiber quantification by qPCR, benzonase was inactivated with EDTA (0.1\u2009M remaining focus) and heat-inactivation at 80\u2009\u00b0C for 10\u2009min.<\/p>\n
Movement cytometry<\/h3>\n
Cells have been indifferent utilizing cell dissociation buffer (GIBCO), stained for 10\u2009min with LIVE\/DEAD fixable near-infrared staining equipment (1:5000, Thermo Fisher), after which mounted in 2% neutral-buffered formalin for 10\u2009min (Sigma). Antigens have been blocked in 2\u2009mM EDTA\u2009+\u20090.5% BSA with Fc-block (1:100, Biolegend) in PBS for 20\u2009min. Cells have been then stained with anti-CD46 antibody (1:200, clone TRA-2-10, Biolegend) or the corresponding isotype management for 30\u2009min in PBS\u2009+\u20092\u2009mM EDTA\u2009+\u20090.5% BSA. Stained cells have been measured on the FACSCalibur stream cytometer (BD Biosciences). Evaluation was carried out on cell populations excluding particles, doublets and lifeless cells.<\/p>\n
Actual-time monitoring of cytopathic impact<\/h3>\n
Lack of cell adhesion as a result of cytopathic impact was monitored in real-time utilizing the xCELLigence RTCA DP instrument (Roche). xCELLigence plates have been equilibrated with 50\u2009\u00b5L of medium for 1\u2009h to determine a baseline. Then, 10,000 cells have been plated in 100\u2009\u00b5L of development medium. Infections have been initiated with 50\u2009\u00b5L, and impedance (cell index) was recorded each 15\u2009min. Measurements have been normalized 4\u2009h earlier than an infection.<\/p>\n
Immunoblotting<\/h3>\n
Immunoblotting was carried out as described beforehand, with blot improvement on X-ray movies (GE Healthcare) or the Biorad Chemidoc imager.12 Dot blots have been carried out to quantify the secretion of deca-His-tagged EpCAM\/CD3-BiTE, as described beforehand.57 Nuclear and cytoplasmic fractionation was carried out utilizing the NE-PER\u2122 Equipment in keeping with the producer\u2019s directions (Thermo Scientific). Antibody dilutions and blocking circumstances are laid out in Supplementary Desk 4. Semi-quantitative evaluation of band depth was carried out by densitometry utilizing the ImageLab software program (Biorad), with depth normalization to the loading management of the identical run.<\/p>\n
Cell cycle synchronization and evaluation<\/h3>\n
Cell synchronization was achieved utilizing a double thymidine block: 2\u2009mM thymidine (Sigma) was added for 16\u2009h, adopted by an 8-h launch, and one other 16-h block. For cell cycle evaluation, cells have been pulsed with 20\u2009\u00b5M bromodeoxyuridine (BrdU, Sigma) for 10\u2009min earlier than harvest. Cells have been subsequently indifferent utilizing trypsin, mounted in ice-cold 70% ethanol for 30\u2009min, and handled with 2\u2009M HCl + 1\u2009mg\u00b7mL\u22121 pepsin (Sigma) for 20\u2009min. After two PBS washes, cells have been stained with a major mouse anti-BrdU antibody (1:100, 90\u2009min, BD Biosciences) and a secondary goat anti-mouse-488 Fab-fragment (1:500, 60\u2009min, Thermo Fisher) in DPBS\u2009+\u20092% FBS. After washing, cells have been stained with 50\u2009\u00b5g\u00b7mL\u22121 propidium iodide and 400\u2009\u00b5g\u00b7mL\u22121 RNase A. Movement cytometry was carried out on an Attune stream cytometer (Thermo Fisher), and knowledge have been processed with FlowJo v10.0.7r2 software program (TreeStar Inc., USA).<\/p>\n
Assay for promoter exercise<\/h3>\n
In whole, 5\u2009\u00d7\u200910\u2074 cells have been seeded in 24-well plates. The following day, 200\u2009ng of plasmid per response (~600\u2013800\u2009pmol) and a pair of.5\u2009\u00b5L Lipofectamine 2000 per 1000\u2009ng DNA have been every diluted in 150\u2009\u00b5L Optimem (Thermo Fisher), mixed, and incubated for 30\u2009min earlier than including to cells. After in a single day incubation, the medium was changed to start out hypoxic publicity. Co-transfection was carried out with one twentieth of the transfected plasmid mass of a \u03b2-galactosidase-encoding plasmid (SV40-betaGal, Promega) to normalize for transfection effectivity. Cells have been washed with PBS, lysed, and subjected to at least one freeze-thaw cycle earlier than measuring luciferase exercise in black 96-well plates with luciferin reagent (Promega). Luciferase expression was normalized to \u03b2-galactosidase exercise, which was measured by incubating lysates with 2\u00d7 ONPG buffer (137\u2009mM HNa2PO4\u2009+\u200963\u2009mM H2NaPO4\u2009+\u20092\u2009mM MgCl2\u2009+\u20090.7% (v\/v) \u03b2-mercaptoethanol + 1.33\u2009mg\u2219mL\u22121 ortho-Nitrophenyl-\u03b2-galactoside, pH 7.2) at 37\u2009\u00b0C for 10\u2009min. Reactions have been stopped with 1\u2009M Na\u2082CO\u2083, and OD was measured at 410\u2009nm.<\/p>\n
Puromycin incorporation assay<\/h3>\n
World translation was assessed by pulsing cells with 1\u2009\u00b5g\u2219mL\u22121 puromycin (Sigma) for 30\u2009min. Pre-treatment with cycloheximide (100\u2009\u00b5M) served as a adverse management. Lysates have been then subjected to immunoblotting.<\/p>\n
Reverse transfection of siRNA<\/h3>\n
siRNA transfections have been performed at a ratio of 10 pmol siRNA to 1\u2009\u00b5L RNAiMax (Thermo Fisher) in Optimem (Thermo Fisher). After mixing, the options have been incubated for 30\u2009min earlier than plating 3\u2009\u00d7\u200910\u2074 cells (24-well) or 1.2\u2009\u00d7\u200910\u2075 cells (6-well). After in a single day incubation, the medium was changed with full medium to start out hypoxic remedy. siRNA sequences are listed in Supplementary Desk 5.<\/p>\n
Sequence alignment of adenovirus genomes<\/h3>\n
Putative HREs have been recognized by handbook scanning of annotated promoters of Ad11p (GenBank: AY598970.1). Corresponding areas from 59 adenovirus genomes (Supplementary Desk 1) have been aligned utilizing the CLC Genomics Workbench (Qiagen). Nucleotide frequencies of the HRE motif throughout all genomes have been visualized as a sequence emblem.<\/p>\n
Chromatin immunoprecipitation-qPCR (ChIP-qPCR)<\/h3>\n
In whole, 6\u2009\u00d7\u2009106 cells have been plated onto 15-cm plates. Chromatin crosslinking was achieved with 1% methanol-free formaldehyde (v\/v) for 10\u2009min with light rocking, adopted by quenching with 125\u2009mM glycine for 10\u2009min. Cells have been washed twice on ice with PBS, scraped into 5\u2009mL of PBS, and pelleted at 500\u00d7\u2009g. Pellets have been resuspended in SDS lysis buffer (1% SDS, 10\u2009mM EDTA, 50\u2009mM Tris pH 8.1) with protease inhibitors (1:30, Thermo Fisher).<\/p>\n
Sonication was carried out at 4\u2009\u00b0C utilizing a Bioruptor (Diagenode) for two:45\u2009min (15-second pulses on\/off, excessive energy) adopted by benzonase remedy (15 U, Milipore) for 25\u2009min for sufficient fragmentation of viral and host chromatin. DNA fragmentation (~300\u2009bp) was confirmed through TapeStation. Protein A agarose beads (40\u2009\u00b5L, Millipore) have been pre-washed with ChIP diluent after which incubated with sonicated DNA for 1\u2009h at 4\u2009\u00b0C on an end-over-end rotator. As beforehand described,58 15\u2009\u00b5L of HIF-1\u03b1 (PM14), HIF-2\u03b1 (PM9), HIF-1\u03b2 (Novus NB100-110), or pre-immune sera (10\u2009\u00b5L) have been used per ChIP response and incubated in a single day at 4\u2009\u00b0C. After bead addition (90\u2009\u00b5L), samples have been incubated for 1.5\u2009h at 4\u2009\u00b0C with end-to-end rotation, after which pelleted at 380\u00d7gn for 8\u2009min. Washes (800\u2009\u00b5L) have been carried out with low- and high-salt buffers and LiCl buffers at 4\u2009\u00b0C for five\u2009min every. The samples have been pelleted at 380\u00d7gn at 4\u2009\u00b0C for five\u2009min, then washed twice in 800\u2009\u00b5L TE. A two-step elution (120\u2009\u00b5L buffer every step) was carried out at room temperature for 15\u2009min with shaking at 1400\u2009rpm. For ChIP buffer parts confer with Supplementary Desk 7.<\/p>\n
Reverse crosslinking was achieved by including 12.5\u2009\u00b5L of 4\u2009M NaCl and heating at 65\u2009\u00b0C in a single day. Proteinase Okay (2\u2009\u00b5l of 20\u2009mg\u00b7mL\u22121) was added for 4\u2009h at 45\u2009\u00b0C, adopted by RNase A (1\u2009\u00b5L) at 37\u2009\u00b0C for 30\u2009min. Precipitated DNA was purified utilizing MinElute columns (Qiagen) and eluted in 20\u2009\u00b5L of DNase-free water. qPCR was carried out utilizing the two\u00d7 SyGreenBlue Hello-ROX in 20\u2009\u00b5L (PCR Biosystems), taking PCR effectivity into consideration (Supplementary Desk 6).<\/p>\n
Statistical evaluation<\/h3>\n
All statistical analyses and plots have been carried out utilizing Prism v.10.6 (Graphpad). Replicates (n) representing organic variance are plotted as single knowledge factors, and technical replicates are indicated the place related. Bars signify the imply\u2009\u00b1\u2009normal deviation, except in any other case acknowledged. For statistical evaluation, log-normal knowledge have been chosen to permit the usage of parametric checks. Two-sided unpaired t checks assuming unequal variance for conservative significance analysis have been carried out (Welch\u00b4s correction). Holm\u2013\u0160id\u00e1k correction was utilized for a number of comparisons.<\/p>\n<\/div>\n
\n
Source link <\/a><\/p>\n","protected":false},"excerpt":{"rendered":"Spatial proximity evaluation of adenovirus foci and hypoxia Animal experimentation was performed beneath moral approval from the UK Residence Workplace (Challenge license 30\/3391 and private license I5916669E). Feminine CB17-SCID mice (6\u20138 weeks outdated) have been inoculated subcutaneously in the correct flank with 2\u2009\u00d7\u2009106 DLD-1 cells in 50\u2009\u00b5L. On day 1, when xenografts reached 80\u2013120\u2009mm3, mice […]<\/p>\n","protected":false},"author":1,"featured_media":3583,"comment_status":"open","ping_status":"open","sticky":false,"template":"","format":"standard","meta":{"fifu_image_url":"https:\/\/media.springernature.com\/m685\/springer-static\/image\/art%3A10.1038%2Fs41392-026-02895-y\/MediaObjects\/41392_2026_2895_Fig1_HTML.png","fifu_image_alt":"","jnews-multi-image_gallery":[],"jnews_single_post":[],"jnews_primary_category":[],"jnews_override_bookmark_settings":[],"jnews_social_meta":[],"jnews_override_counter":[],"footnotes":""},"categories":[10],"tags":[391,3943,3945,2929,3944,3940,3941,3942],"class_list":["post-3581","post","type-post","status-publish","format-standard","has-post-thumbnail","hentry","category-biotechnology","tag-activity","tag-adenoviruses","tag-e1a","tag-expression","tag-hif2stimulated","tag-hypoxia","tag-increases","tag-oncolytic"],"yoast_head":"\n
Hypoxia will increase the exercise of oncolytic adenoviruses via HIF-2\u03b1-stimulated E1A expression - Future News 24<\/title>\n<meta name=\"description\" content=\"Hypoxia, a hallmark of solid tumors, poses a significant challenge in cancer therapy due to its association with poor prognosis and resistance to conventional treatments. Oncolytic viruses represent a promising treatment strategy, as they selectively replicate within cancer cells and lyse them, potentially including those in hypoxic tumor regions. Here, we examined how hypoxic conditions influence the activity of enadenotucirev (EnAd), a clinically relevant group B oncolytic adenovirus previously detected in hypoxic areas of xenograft tumors. We demonstrated that hypoxia enhances virus production by boosting transcription and translation of immediate-early, early, and late adenoviral genes. The immediate-early gene E1A was upregulated within 2 h (17-fold) after virus entry under hypoxia, driven by a conserved hypoxia-response element (HRE) in its promoter. Mechanistic studies revealed that the hypoxia-inducible factor (HIF)-2α and HIF-1β heterodimers bind to this HRE, transactivating E1A. By inducing E1A expression, hypoxia also elevated viral genome synthesis, structural protein production, and therapeutic transgene expression, underscoring the potential of EnAd to target the hypoxic tumor microenvironment. This is the first report of a functional HRE in a human adenovirus, conserved across 59 adenovirus genotypes, and identifies hypoxia as a driver of enhanced oncolytic activity with implications for adenovirus-based therapies in solid tumors.\" \/>\n<meta name=\"robots\" content=\"index, follow, max-snippet:-1, max-image-preview:large, max-video-preview:-1\" \/>\n<link rel=\"canonical\" href=\"https:\/\/futurenews24.com\/index.php\/2026\/08\/11\/s41392-026-02895-y\/\" \/>\n<meta property=\"og:locale\" content=\"en_US\" \/>\n<meta property=\"og:type\" content=\"article\" \/>\n<meta property=\"og:title\" content=\"Hypoxia will increase the exercise of oncolytic adenoviruses via HIF-2\u03b1-stimulated E1A expression - Future News 24\" \/>\n<meta property=\"og:description\" content=\"Hypoxia, a hallmark of solid tumors, poses a significant challenge in cancer therapy due to its association with poor prognosis and resistance to conventional treatments. Oncolytic viruses represent a promising treatment strategy, as they selectively replicate within cancer cells and lyse them, potentially including those in hypoxic tumor regions. Here, we examined how hypoxic conditions influence the activity of enadenotucirev (EnAd), a clinically relevant group B oncolytic adenovirus previously detected in hypoxic areas of xenograft tumors. We demonstrated that hypoxia enhances virus production by boosting transcription and translation of immediate-early, early, and late adenoviral genes. The immediate-early gene E1A was upregulated within 2 h (17-fold) after virus entry under hypoxia, driven by a conserved hypoxia-response element (HRE) in its promoter. Mechanistic studies revealed that the hypoxia-inducible factor (HIF)-2α and HIF-1β heterodimers bind to this HRE, transactivating E1A. By inducing E1A expression, hypoxia also elevated viral genome synthesis, structural protein production, and therapeutic transgene expression, underscoring the potential of EnAd to target the hypoxic tumor microenvironment. This is the first report of a functional HRE in a human adenovirus, conserved across 59 adenovirus genotypes, and identifies hypoxia as a driver of enhanced oncolytic activity with implications for adenovirus-based therapies in solid tumors.\" \/>\n<meta property=\"og:url\" content=\"https:\/\/futurenews24.com\/index.php\/2026\/08\/11\/s41392-026-02895-y\/\" \/>\n<meta property=\"og:site_name\" content=\"Future News 24\" \/>\n<meta property=\"article:published_time\" content=\"2026-08-11T00:00:00+00:00\" \/>\n<meta property=\"article:modified_time\" content=\"2026-08-11T04:59:07+00:00\" \/>\n<meta property=\"og:image\" content=\"https:\/\/media.springernature.com\/m685\/springer-static\/image\/art%3A10.1038%2Fs41392-026-02895-y\/MediaObjects\/41392_2026_2895_Fig1_HTML.png\" \/>\n<meta name=\"author\" content=\"Future News 24\" \/>\n<meta name=\"twitter:card\" content=\"summary_large_image\" \/>\n<meta name=\"twitter:image\" content=\"https:\/\/media.springernature.com\/m685\/springer-static\/image\/art%3A10.1038%2Fs41392-026-02895-y\/MediaObjects\/41392_2026_2895_Fig1_HTML.png\" \/>\n<meta name=\"twitter:label1\" content=\"Written by\" \/>\n\t<meta name=\"twitter:data1\" content=\"Future News 24\" \/>\n\t<meta name=\"twitter:label2\" content=\"Est. reading time\" \/>\n\t<meta name=\"twitter:data2\" content=\"11 minutes\" \/>\n<script type=\"application\/ld+json\" class=\"yoast-schema-graph\">{\"@context\":\"https:\\\/\\\/schema.org\",\"@graph\":[{\"@type\":\"Article\",\"@id\":\"https:\\\/\\\/futurenews24.com\\\/index.php\\\/2026\\\/08\\\/11\\\/s41392-026-02895-y\\\/#article\",\"isPartOf\":{\"@id\":\"https:\\\/\\\/futurenews24.com\\\/index.php\\\/2026\\\/08\\\/11\\\/s41392-026-02895-y\\\/\"},\"author\":{\"name\":\"Future News 24\",\"@id\":\"https:\\\/\\\/futurenews24.com\\\/#\\\/schema\\\/person\\\/cecad1bde21cfc357cf70128144d6c83\"},\"headline\":\"Hypoxia will increase the exercise of oncolytic adenoviruses via HIF-2\u03b1-stimulated E1A expression\",\"datePublished\":\"2026-08-11T00:00:00+00:00\",\"dateModified\":\"2026-08-11T04:59:07+00:00\",\"mainEntityOfPage\":{\"@id\":\"https:\\\/\\\/futurenews24.com\\\/index.php\\\/2026\\\/08\\\/11\\\/s41392-026-02895-y\\\/\"},\"wordCount\":2166,\"commentCount\":0,\"publisher\":{\"@id\":\"https:\\\/\\\/futurenews24.com\\\/#organization\"},\"image\":{\"@id\":\"https:\\\/\\\/futurenews24.com\\\/index.php\\\/2026\\\/08\\\/11\\\/s41392-026-02895-y\\\/#primaryimage\"},\"thumbnailUrl\":\"https:\\\/\\\/media.springernature.com\\\/m685\\\/springer-static\\\/image\\\/art%3A10.1038%2Fs41392-026-02895-y\\\/MediaObjects\\\/41392_2026_2895_Fig1_HTML.png\",\"keywords\":[\"activity\",\"adenoviruses\",\"E1A\",\"expression\",\"HIF2\u03b1stimulated\",\"Hypoxia\",\"increases\",\"oncolytic\"],\"articleSection\":[\"BioTechnology\"],\"inLanguage\":\"en-US\",\"potentialAction\":[{\"@type\":\"CommentAction\",\"name\":\"Comment\",\"target\":[\"https:\\\/\\\/futurenews24.com\\\/index.php\\\/2026\\\/08\\\/11\\\/s41392-026-02895-y\\\/#respond\"]}]},{\"@type\":\"WebPage\",\"@id\":\"https:\\\/\\\/futurenews24.com\\\/index.php\\\/2026\\\/08\\\/11\\\/s41392-026-02895-y\\\/\",\"url\":\"https:\\\/\\\/futurenews24.com\\\/index.php\\\/2026\\\/08\\\/11\\\/s41392-026-02895-y\\\/\",\"name\":\"Hypoxia will increase the exercise of oncolytic adenoviruses via HIF-2\u03b1-stimulated E1A expression - Future News 24\",\"isPartOf\":{\"@id\":\"https:\\\/\\\/futurenews24.com\\\/#website\"},\"primaryImageOfPage\":{\"@id\":\"https:\\\/\\\/futurenews24.com\\\/index.php\\\/2026\\\/08\\\/11\\\/s41392-026-02895-y\\\/#primaryimage\"},\"image\":{\"@id\":\"https:\\\/\\\/futurenews24.com\\\/index.php\\\/2026\\\/08\\\/11\\\/s41392-026-02895-y\\\/#primaryimage\"},\"thumbnailUrl\":\"https:\\\/\\\/media.springernature.com\\\/m685\\\/springer-static\\\/image\\\/art%3A10.1038%2Fs41392-026-02895-y\\\/MediaObjects\\\/41392_2026_2895_Fig1_HTML.png\",\"datePublished\":\"2026-08-11T00:00:00+00:00\",\"dateModified\":\"2026-08-11T04:59:07+00:00\",\"description\":\"Hypoxia, a hallmark of solid tumors, poses a significant challenge in cancer therapy due to its association with poor prognosis and resistance to conventional treatments. Oncolytic viruses represent a promising treatment strategy, as they selectively replicate within cancer cells and lyse them, potentially including those in hypoxic tumor regions. Here, we examined how hypoxic conditions influence the activity of enadenotucirev (EnAd), a clinically relevant group B oncolytic adenovirus previously detected in hypoxic areas of xenograft tumors. We demonstrated that hypoxia enhances virus production by boosting transcription and translation of immediate-early, early, and late adenoviral genes. The immediate-early gene E1A was upregulated within 2 h (17-fold) after virus entry under hypoxia, driven by a conserved hypoxia-response element (HRE) in its promoter. Mechanistic studies revealed that the hypoxia-inducible factor (HIF)-2α and HIF-1β heterodimers bind to this HRE, transactivating E1A. By inducing E1A expression, hypoxia also elevated viral genome synthesis, structural protein production, and therapeutic transgene expression, underscoring the potential of EnAd to target the hypoxic tumor microenvironment. 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Oncolytic viruses represent a promising treatment strategy, as they selectively replicate within cancer cells and lyse them, potentially including those in hypoxic tumor regions. Here, we examined how hypoxic conditions influence the activity of enadenotucirev (EnAd), a clinically relevant group B oncolytic adenovirus previously detected in hypoxic areas of xenograft tumors. We demonstrated that hypoxia enhances virus production by boosting transcription and translation of immediate-early, early, and late adenoviral genes. The immediate-early gene E1A was upregulated within 2 h (17-fold) after virus entry under hypoxia, driven by a conserved hypoxia-response element (HRE) in its promoter. Mechanistic studies revealed that the hypoxia-inducible factor (HIF)-2α and HIF-1β heterodimers bind to this HRE, transactivating E1A. 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