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Cited by 59Virus-based gene therapy researchRNA Interference and Gene DeliveryImmunotherapy and Immune Responses

Immunomodulation in Administration of RAAV: Preclinical and Clinical Adjuvant Pharmacotherapies

Open access
Chu, Wing Sum, Ng, Joanne·2021·Frontiers in Immunology
DOI 10.3389/fimmu.2021.658038PMC PMC8049138
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Abstract

Recombinant adeno-associated virus (rAAV) has attracted a significant research focus for delivering genetic therapies to target cells. This non-enveloped virus has been trialed in many clinical-stage therapeutic strategies but important obstacle in clinical translation is the activation of both innate and adaptive immune response to the protein capsid, vector genome and transgene product. In addition, the normal population has pre-existing neutralizing antibodies against wild-type AAV, and cross-reactivity is observed between different rAAV serotypes. While extent of response can be influenced by dosing, administration route and target organ(s), these pose concerns over reduction or complete loss of efficacy, options for re-administration, and other unwanted immunological sequalae such as local tissue damage. To reduce said immunological risks, patients are excluded if they harbor anti-AAV antibodies or have received gene therapy previously. Studies have incorporated immunomodulating or suppressive regimens to block cellular and humoral immune responses such as systemic corticosteroids pre- and post-administration of Luxturna® and Zolgensma®, the two rAAV products with licensed regulatory approval in Europe and the United States. In this review, we will introduce the current pharmacological strategies to immunosuppress or immunomodulate the host immune response to rAAV gene therapy.

AI snapshot

Written from the paper’s full text.

TL;DR

This review summarizes licensed and repurposed pharmacotherapies used to immunomodulate host responses to rAAV gene therapy, including corticosteroids, rapamycin, mycophenolate mofetil, calcineurin inhibitors, rituximab, imlifidase, proteasome inhibitors, chemotherapeutics, oxidative-stress agents, and antimalarials. It argues that drug choice, timing, duration, and combination are critical for enabling durable transgene expression and possible re-administration.

Key findings

  • Host immunity to rAAV capsid, genome, and transgene limits efficacy; pre-existing neutralizing antibodies can block transduction even at 1:5, and AAV2 seroprevalence is reported at 60%, leading many trials to exclude seropositive patients.
  • In an rAAV2 hemophilia B trial, factor IX activity was initially >10% for 4 weeks but declined to <1% with self-limiting transaminitis and capsid-specific CTL changes; in a subsequent AAV8 study, steroids maintained factor IX levels albeit with a 50–70% decline from peak.
  • Preclinical/clinical adjuvant strategies include rapamycin with prednisolone reducing pre-existing anti-capsid IgG by up to 93% after 8 weeks in mice; SVP[Rapa] enabling AAV8 re-dosing at 4×10^12 vg/kg in mice and NHP; rituximab producing at least a half-log anti-AAV2 NAb reduction in 9/28 rheumatoid arthritis patients; bortezomib enhancing transgene expression >50% for one year versus ~10% in hemophilia A mice; hydroxychloroquine improving photoreceptor transgene expression 5.9-fold; arsenic trioxide increasing luciferase 3.9-fold; and teniposide producing 2-log higher bioluminescence at 48 hours.
  • Combination regimens show mixed results: adding daclizumab to MMF/rapamycin decreased Treg and caused anti-hFIX antibodies; delayed rATG prevented anti-transgene humoral response; and a 5-drug regimen reduced T-cell response but did not prevent NAb emergence after regimen removal.

Methods

  • Narrative review of current pharmacological strategies to immunosuppress or immunomodulate the host immune response to rAAV gene therapy.
  • Sources are published preclinical and clinical studies and licensed agents; Table 1 lists drug indications, adverse effects, example AAV serotypes, and study type, excluding MnTBAP and teniposide because they are not or no longer licensed in Europe and the US.
  • No systematic search, meta-analysis, or primary data collection is described.

Limitations

  • Stated: lack of fully predictive animal models; exact mechanisms of high-dose hepatotoxicity remain to be elucidated; hydroxychloroquine’s endosomal pH effect may not replicate with systemic application; IdeS re-administration success in one NHP was not replicated in a larger cohort; further studies are required for proteasome inhibitor effectiveness across cell types and serotypes and for teniposide durability.
  • Evident: as a narrative review, it does not systematically assess study quality or risk of bias, and many cited findings are preclinical or from small samples.
  • Evident: Table 1 excludes MnTBAP and teniposide because they are not or no longer licensed in Europe and the US, so coverage is restricted to licensed agents.

Model-written summary. Check it against the paper before relying on it.

Integrity signals

Registry

Crossref · 10.3389/fimmu.2021.658038

Cites

115 of 115 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

0 of 114 cited sources checked · 0 supported · 0 flagged

  • Premisecitation

    Pre-existing neutralizing antibody (NAb) can effectively block rAAV transduction even at low levels (1:5).

    “Pre-existing neutralizing antibody (NAb) can effectively block rAAV transduction even at low levels (1:5) (6).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [6] Jiang, 2006, Effects of transient immunosuppression on adenoassociated,virus-mediated, liver-directed gene transfer in rhesus macaques and implications for human genetherapy, Blood source

      Not checked in the free run.

  • Premisenone given

    The extent of immune response to rAAV can be influenced by dosing, administration route, and target organ(s).

    “While extent of response can be influenced by dosing, administration route and target organ(s), these pose concerns over reduction or complete loss of efficacy, options for re-administration, and other unwanted immunological sequalae such as local tissue damage.”
    Quote in the paperSubstantial claim
  • Premisecitation

    Initial exposure to AAV induces humoral and cellular anti-capsid responses that are reactive to rAAV due to capsid similarity.

    “Although AAV infection is non-pathogenic in humans, initial exposure induces humoral and cellular anti-capsid response that are reactive to rAAV due to capsid similarity (4, 5).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [4] Boutin, 2010, Prevalence of serum IgG and neutralizing factors against adeno-associated virus (AAV) types 1, 2, 5, 6, 8, and 9 in the healthy population: Implications for gene therapy using AAV vectors, Hum Gene Ther source

      Not checked in the free run.

    • cites [5] Mingozzi, 2007, CD8+ T-cell responses to adeno-associated virus capsid in humans, Nat Med source

      Not checked in the free run.

  • Premisenone given

    Cross-reactivity is observed between different rAAV serotypes.

    “In addition, the normal population has pre-existing neutralizing antibodies against wild-type AAV, and cross-reactivity is observed between different rAAV serotypes.”
    Quote in the paperNot a substantial claim
  • Premisecitation

    Most rAAV clinical trials exclude seropositive patients.

    “Most rAAV clinical trials exclude seropositive patients; given the high seroprevalence (60% for AAV2), limiting patients suitable for rAAV therapy (7, 8).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [7] Calcedo, 2009, Worldwide Epidemiology of Neutralizing Antibodies to Adeno-Associated Viruses, J Infect Dis source

      Not checked in the free run.

    • cites [8] Li, 2012, Neutralizing antibodies against adeno-associated virus examinedprospectively in pediatric patients with hemophilia, Gene Ther source

      Not checked in the free run.

  • May combine two claims. 41
  • Classified as probably not a substantial claim. 10

  • May combine two claims.

    Interpretationreasoning

    Immune responses to rAAV pose concerns over reduction or complete loss of efficacy, options for re-administration, and other unwanted immunological sequelae such as local tissue damage.

    “While extent of response can be influenced by dosing, administration route and target organ(s), these pose concerns over reduction or complete loss of efficacy, options for re-administration, and other unwanted immunological sequalae such as local tissue damage.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    AAV has a 4.7kb single-stranded DNA genome containing 4 open reading frames (rep, cap, aap, and MAAP) flanked by inverted terminal repeats (ITRs).

    “It is 4.7kb single-stranded DNA genome containing 4 open reading frames (ORFs) (rep, cap, aap, and MAAP) flanked by inverted terminal repeats (ITRs) (1, 2).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [1] Ogden, 2019, Comprehensive AAV capsid fitness landscape reveals a viral gene andenables machine-guided design, Science source

      Not checked in the free run.

    • cites [2] Naso, 2017, Adeno-Associated Virus (AAV) as a Vector for Gene Therapy, BioDrugs source

      Not checked in the free run.

  • Classified as probably not a substantial claim.

    Premisereasoning

    rAAV has emerged as a leading vector to deliver genetic therapies due to its ability to transduce diverse cell types and safety profile.

    “It has emerged as a leading vector to deliver genetic therapies due to its ability to transduce diverse cell types and safety profile.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    AAV is a 26nm, non-enveloped virus of Parvoviridae family.

    “Adeno-associated virus (AAV) is a 26nm, non-enveloped virus of Parvoviridae family.”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [1] Ogden, 2019, Comprehensive AAV capsid fitness landscape reveals a viral gene andenables machine-guided design, Science source

      Not checked in the free run.

    • cites [2] Naso, 2017, Adeno-Associated Virus (AAV) as a Vector for Gene Therapy, BioDrugs source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    The seroprevalence of AAV2 is high (60%), limiting patients suitable for rAAV therapy.

    “Most rAAV clinical trials exclude seropositive patients; given the high seroprevalence (60% for AAV2), limiting patients suitable for rAAV therapy (7, 8).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [7] Calcedo, 2009, Worldwide Epidemiology of Neutralizing Antibodies to Adeno-Associated Viruses, J Infect Dis source

      Not checked in the free run.

    • cites [8] Li, 2012, Neutralizing antibodies against adeno-associated virus examinedprospectively in pediatric patients with hemophilia, Gene Ther source

      Not checked in the free run.

  • May combine two claims.

    Premisereasoning

    Patients are excluded from rAAV therapy if they harbor anti-AAV antibodies or have received gene therapy previously.

    “To reduce said immunological risks, patients are excluded if they harbor anti-AAV antibodies or have received gene therapy previously.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Mycophenolate mofetil (MMF), prodrug of mycophenolic acid, preferentially inhibits type II IMPDH, suppressing T and B cells proliferation.

    “Mycophenolate mofetil (MMF), prodrug of mycophenolic acid, preferentially inhibits type II IMPDH, suppressing T and B cells proliferation (74).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [74] Allison, 2005, Mechanisms of action of mycophenolate mofetil, Lupus source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Ciclosporin and tacrolimus were found to inhibit Treg proliferation and activity in vitro, and similar effects were observed in tacrolimus-treated allograft patients ex vivo.

    “Ciclosporin and tacrolimus were found to inhibit Treg proliferation and activity in vitro (79), and similar effects were observed in tacrolimus-treated allograft patients ex vivo (80); this could be detrimental in inhibiting the development of peripheral tolerance following rAAV administration.”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [79] Miroux, 2012, In vitro effects of cyclosporine a and tacrolimus on regulatoryt-cell proliferation and function, Transplantation source

      Not checked in the free run.

    • cites [80] Akimova, 2012, Differing effects of rapamycin or calcineurin inhibitor onT-Regulatory cells in pediatric liver and kidney transplant recipients, Am J Transplant source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Unmethylated CpG motifs in rAAV vector genome have been suggested as the cause of loss of expression in a rAAV8 hemophilia B trial.

    “and has been suggested as the cause of loss of expression in a rAAV8 hemophilia B trial (21).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [21] Konkle, 2021, BAX 335 hemophilia B gene therapy clinical trial results: potential impact of CpG sequences on gene expression, Blood source

      Not checked in the free run.

  • May combine two claims.

    Premisenone given

    Rapamycin binds to the same intracellular target (immunophilin) as tacrolimus; however, rapamycin/FKPB12 complex inhibits a crucial cell-cycle kinase known as mammalian target of rapamycin (mTOR).

    “Rapamycin is a macrolide immunosuppressant that binds to the same intracellular target (immunophilin) as tacrolimus; however, rapamycin/FKPB12 complex inhibits a crucial cell-cycle kinase known as mammalian target of rapamycin (mTOR).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    In the rAAV2 hemophilia B clinical trial, initial stable therapeutic factor IX (FIX) expression (>10% activity) lasted for 4 weeks, then FIX levels gradually declined to baseline (<1%).

    “Despite initial stable therapeutic factor IX (FIX) expression (>10% activity) for 4 weeks, FIX levels gradually declined to baseline (<1%).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [12] Manno, 2006, Successful transduction of liver in hemophilia by AAV-Factor IX and limitations imposed by the host immune response, Nat Med source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Rapamycin has beneficial downstream effects including Treg generation, suppressing CTL and T helper (TH) activation and at higher doses, B-cell proliferation and differentiation.

    “Beneficial downstream effects include Treg generation, suppressing CTL and T helper (TH) activation and at higher doses, B-cell proliferation and differentiation (69–71).”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [69] Limon, 2014, MTOR kinase inhibitors promote antibody class switching via mTORC2 inhibition source

      Not checked in the free run.

    • cites [70] Delgoffe, 2009, The mTOR Kinase Differentially Regulates Effector and Regulatory T Cell Lineage Commitment

      Not checked in the free run.

    • cites [71] Zeiser, 2008, Differential impact of mammalian target of rapamycin inhibition on CD4 +CD25+Foxp3+ regulatory T cells compared with conventional CD4+ T cells, Blood source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    No generalized toxicity was reported with daily tacrolimus in NHP, but T-cell and macrophages infiltrations were observed.

    “No generalized toxicity was reported but T-cell and macrophages infiltrations were observed.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [47] Ishii, 2020, rAAV8 and rAAV9-Mediated Long-Term Muscle Transduction with Tacrolimus (FK506) in Non-Human Primates, Mol Ther - Methods Clin Dev source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    In mice MMF reduced rAAV transduction efficiency by depleting guanosine triphosphate required for vector genome second strand synthesis, but this was not observed in higher animals.

    “In mice MMF reduced rAAV transduction efficiency by depleting guanosine triphosphate required for vector genome second strand synthesis (75), but this was not observed in higher animals.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [75] Montenegro-Miranda, 2011, Mycophenolate mofetil impairs transduction of single-stranded adeno-associated viral vectors, Hum Gene Ther source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    CCS are adopted in approved gene therapies for inherited retinal dystrophy and spinal muscle atrophy (SMA).

    “They are also adopted in approved gene therapies for inherited retinal dystrophy (25) and spinal muscle atrophy (SMA) (26).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [25] Russell, 2017, Efficacy and safety of voretigene neparvovec (AAV2-hRPE65v2) in patients with RPE65-mediated inherited retinal dystrophy: a randomised, controlled, open-label, phase 3 trial, Lancet source

      Not checked in the free run.

    • cites [26] Mendell, 2017, Single-Dose Gene-Replacement Therapy for Spinal Muscular Atrophy, N Engl J Med source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    rAAV capsid-targeting TLR2, various DNA sensors, and complement activation may also play a role.

    “Furthermore, rAAV capsid-targeting TLR2, various DNA sensors, and complement activation may also play a role (22).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [22] Muhuri, 2021, Overcoming innate immune barriers that impede AAV gene therapy vectors, J Clin Invest source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    SVP[Rapa] provided sufficient reduction of B and T cell activation in an antigen-selective manner, inhibited CTL liver infiltration, and efficiently blocked memory T cell response.

    “SVP [Rapa] provided sufficient reduction of B and T cell activation in an antigen-selective manner, inhibited CTL liver infiltration, and efficiently blocked memory T cell response.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [39] Meliani, 2018, Antigen-selective modulation of AAV immunogenicity with tolerogenicrapamycin nanoparticles enables successful vector re-administration, Nat Commun source

      Not checked in the free run.

  • May combine two claims.

    Interpretationcitation

    Endosomal acidification is essential for rAAV escape, and hydroxychloroquine increases endosomal and lysosomal pH, so the effect may not be replicated or consistent with systemic application.

    “However, endosomal acidification is essential for rAAV escape (84), and hydroxychloroquine increases endosomal and lysosomal pH (60), this effect may not be replicated or consistent with systemic application.”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [84] Dhungel, 2020, Journey to the Center of the Cell: Tracing the Path of AAV Transduction, Trends Mol Med source

      Not checked in the free run.

    • cites [60] Schrezenmeier, 2020, Mechanisms of action of hydroxychloroquine and chloroquine:implications for rheumatology, Nat Rev Rheumatol source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Rituximab depletes B cells by inducing apoptosis, antibody dependent cell-mediated cytotoxicity and complement dependent cytotoxicity, thereby limiting antibody production and epitope presentation via MHC class II to TH cells.

    “It depletes B cells by inducing apoptosis, antibody dependent cell-mediated cytotoxicity and complement dependent cytotoxicity, thereby limiting antibody production and epitope presentation via MHC class II to TH cells (81).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [81] Smith, 2003, Rituximab (monoclonal anti-CD20 antibody): Mechanisms of action and resistance, Oncogene source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Proteasome inhibitors (PIs) are licensed for multiple myeloma.

    “Proteasome inhibitors (PIs) are licensed for multiple myeloma.”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [54] Merin, 2014, Clinical use of proteasome inhibitors in the treatment of multiple myeloma, Pharmaceuticals source

      Not checked in the free run.

    • cites [55] Schlafer, 2017, Safety of proteasome inhibitors for treatment of multiple myeloma, Expert Opin Drug Saf source

      Not checked in the free run.

  • Classified as probably not a substantial claim. May combine two claims.

    Interpretationcitation

    It is possible by postponing rATG lowers the Th17/Treg ratio, allowing peripheral tolerance to the transgene product.

    “It is possible by postponing rATG lowers the Th17/Treg ratio, allowing peripheral tolerance to the transgene product (41).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [41] Samelson-Jones, 2020, Timing of Intensive Immunosuppression Impacts Risk of TransgeneAntibodies after AAV Gene Therapy in Nonhuman Primates, Mol Ther - Methods Clin Dev source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    In NHP, rAAV2-hFIX (8×10^12vg/kg) delivered intrahepatically alongside 2-drug regimen of MMF and rapamycin compared to 3-drug adding Daclizumab, the addition of daclizumab resulted in decreased CD4+CD25+FoxP3+ Treg and consistent formation of inhibitory antibodies to hFIX; this was not observed in the 2-drug group.

    “rAAV2-hFIX (8×1012vg/kg) was delivered intrahepatically to NHP alongside 2-drug regimen of MMF and rapamycin compared to 3-drug adding Daclizumab (40). The addition of daclizumab resulted in decreased CD4+CD25+FoxP3+ Treg and consistent formation of inhibitory antibodies to hFIX; this was not observed in the 2-drug group.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [40] Mingozzi, 2007, Modulation of tolerance to the transgene product in a nonhuman primate model of AAV-mediated gene transfer to liver, Blood source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Intraperitoneal ATO 5μg/g/day from day -2 to 2 showed 3.9-fold increase in luciferase assay 12 days after rAAV8 retro-orbital injection, with dose-dependent increase of intracellular reactive oxygen species that inhibit vector degradation pathways.

    “Intraperitoneal ATO 5μg/g/day from day -2 to 2 showed 3.9-fold increase in luciferase assay 12 days after rAAV8 retro-orbital injection, with dose-dependent increase of intracellular reactive oxygen species that inhibit vector degradation pathways (59).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [59] Mitchell, 2013, Arsenic Trioxide Stabilizes Accumulations of Adeno-Associated Virus Virions at the Perinuclear Region, Increasing Transduction In Vitro and In Vivo, J Virol source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Both bortezomib and carfilzomib enhance rAAV2 transduction in vitro, but bortezomib is more efficacious than carfilzomib in vivo when administered by retro-orbital injection with rAAV2.

    “Both bortezomib and carfilzomib enhance rAAV2 transduction in vitro, but bortezomib is more efficacious than carfilzomib in vivo when administered by retro-orbital injection with rAAV2 (56).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [56] Mitchell, 2013, Mechanistic Insights into the Enhancement of Adeno-Associated Virus Transduction by Proteasome Inhibitors, J Virol source

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    Drug selection, initiation and duration of suppression, and role of global immunosuppression are important considerations.

    “This suggests that drug selection, initiation and duration of suppression, and role of global immunosuppression are important considerations.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationcitation

    Careful selection of immunosuppressive agents is necessary as Treg play a critical role in regulating immune response to rAAV products, particularly observed in liver and muscle gene transfer.

    “This indicates careful selection of immunosuppressive agents is necessary as Treg play a critical role in regulating immune response to rAAV products, particularly observed in liver and muscle gene transfer (97).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [97] Biswas, 2018, Gene therapy with regulatory T cells: A beneficial alliance, Front Immunol source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Bortezomib has been shown to dose-dependently decrease cell surface MHC class I antigen presentation and inhibit CTL-mediated lysis after rAAV administration in vitro.

    “Bortezomib has been shown to dose-dependently decrease cell surface MHC class I antigen presentation and inhibit CTL-mediated lysis after rAAV administration in vitro (87).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [87] Finn, 2010, Proteasome inhibitors decrease AAV2 capsid derived peptide epitope presentation on mhc class i following transduction, Mol Ther source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    In the first IdeS re-administration study (n=1), no induction of anti-capsid IgG and NAb, along with lower IgM and increased transgene level was observed for 21 days after second rAAV8-hFIX administration.

    “In the first study (n=1), no induction of anti-capsid IgG and NAb, along with lower IgM and increased transgene level was observed for 21 days after second rAAV8-hFIX administration.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [53] Leborgne, 2020, IgG-cleaving endopeptidase enables in vivo gene therapy in the presence of anti-AAV neutralizing antibodies, Nat Med source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Daclizumab is a humanized monoclonal antibody targeting CD25 present on interleukin-2 receptor commonly found in activated T cells and CD4+CD25+FoxP3+.

    “Daclizumab is a humanized monoclonal antibody targeting CD25 present on interleukin-2 receptor commonly found in activated T cells and CD4+CD25+FoxP3+ (96).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [96] Milo, 2014, The efficacy and safety of daclizumab and its potential role in thetreatment of multiple sclerosis, Ther Adv Neurol Disord source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Using a laboratory version of IdeS with rAAV8, significant reductions in anti-AAV8 IgG and NAb levels, with enhanced liver transduction and transgene expression, were observed in passively immunized murine models and naturally immunized NHP.

    “Using a laboratory version of IdeS with rAAV8, significant reductions in anti-AAV8 IgG and NAb levels, with enhanced liver transduction and transgene expression and observed in passively immunized murine models and naturally immunized NHP (53).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [53] Leborgne, 2020, IgG-cleaving endopeptidase enables in vivo gene therapy in the presence of anti-AAV neutralizing antibodies, Nat Med source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    The proteasome degradation pathway results in unsuccessful transduction, and capsid-derived peptides are presented to CTL by MHC class I molecules, provoking elimination of transduced cells and loss of transgene expression.

    “The latter pathway results in unsuccessful transduction, and capsid-derived peptides are presented to CTL by MHC class I molecules, provoking elimination of transduced cells and loss of transgene expression (85).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [85] Wang, 2019, Adeno-associated virus vector as a platform for gene therapy delivery, Nat Rev Drug Discovery source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    For AAV2, 9/28 patients showed at least a half-log reduction, and individuals with NAb titer ≤1:1000 were more likely to respond to RTX.

    “For AAV2, 9/28 patients showed at least a half-log reduction, and inferred individuals with NAb titer ≤1:1000 were more likely to respond to RTX but the contribution of methotrexate is unknown.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [50] Mingozzi, 2013, Prevalence and pharmacological modulation of humoral immunity to AAV vectors in gene transfer to synovial tissue, Gene Ther source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    A single bortezomib dose given with rAAV8 dosing enhanced transgene expression by >50% for one year (compared to ~10%) in hemophilia A mice, and longer in-range clotting time for at least 10 months in hemophilia A dogs.

    “Moreover, a single bortezomib dose given with rAAV8 dosing enhanced transgene expression by >50% for one year (compared to ~10%) in hemophilia A mice, and longer in-range clotting time for at least 10 months in hemophilia A dogs (57).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [57] Monahan, 2010, Proteasome inhibitors enhance gene delivery by AAV virus vectors expressing large genomes in hemophilia mouse and dog models: A strategy for broad clinical application, Mol Ther source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    A 5-drug regimen with rAAV5-PBGD 1×10^13vg/kg infusion resulted in reduced T-cell response in NHP, but did not prevent NAb emergence following regimen removal.

    “This 5-drug regimen with rAAV5-PBGD 1×1013vg/kg infusion resulted in reduced T-cell response in NHP, but did not prevent NAb emergence following regimen removal.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [99] Unzu, 2012, Transient and intensive pharmacological immunosuppression fails to improve AAV-based liver gene transfer in non-human primates, J Transl Med source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Tail vein injection of rAAV2-Luc with teniposide (at doses of 1×10^11vg and 20mg/kg respectively) resulted in bioluminescence 2-log higher 48 hours post-administration without hepatotoxicity.

    “Tail vein injection of rAAV2-Luc with teniposide (at doses of 1×1011vg and 20mg/kg respectively) resulted in bioluminescence 2-log higher 48 hours post-administration without hepatotoxicity.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [94] Nicolson, 2016, Identification and Validation of Small Molecules That Enhance Recombinant Adeno-associated Virus Transduction following High-Throughput Screens, J Virol source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Rituximab (RTX) is a chimeric mouse/human monoclonal antibody targeting CD20 present in pre‐B and mature B cells except plasma cells.

    “Rituximab (RTX) is a chimeric mouse/human monoclonal antibody targeting CD20 present in pre‐B and mature B cells except plasma cells.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [81] Smith, 2003, Rituximab (monoclonal anti-CD20 antibody): Mechanisms of action and resistance, Oncogene source

      Not checked in the free run.

  • May combine two claims.

    Interpretationnone given

    Human immune responses to the viral vector, transgene, and protein product determine the therapeutic efficacy and possibility of re-administration.

    “However, human immune responses to the viral vector, transgene, and protein product determine the therapeutic efficacy and possibility of re-administration.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Studies showed cross-reactive anti-capsid NAb present at 15 years, CTL and Treg infiltrates at injection site after 5 years; and in NHP adverse effects related to high-dosage.

    “Studies showed cross-reactive anti-capsid NAb present at 15 years (100), CTL and Treg infiltrates at injection site after 5 years (101); and in NHP adverse effects related to high-dosage (42, 66).”
    Quote in the paperSubstantial claim0 of 4 sources checked
    • cites [100] George, 2020, Long-Term Follow-Up of the First in Human Intravascular Delivery ofAAV for Gene Transfer: AAV2-hFIX16 for Severe Hemophilia B, Mol Ther source

      Not checked in the free run.

    • cites [101] Mueller, 2017, 5 Year Expression and Neutrophil Defect Repair after Gene Therapy in Alpha-1 Antitrypsin Deficiency, Mol Ther source

      Not checked in the free run.

    • cites [42] Hordeaux, 2018, Toxicology Study of Intra-Cisterna Magna Adeno-Associated Virus 9 Expressing Iduronate-2-Sulfatase in Rhesus Macaques source

      Not checked in the free run.

    • cites [66] Hinderer, 2018, Severe Toxicity in Nonhuman Primates and Piglets Following High-Dose Intravenous Administration of an Adeno-Associated Virus Vector Expressing Human SMN, Hum Gene Ther source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    In the delayed rATG study, neither group had cellular response to capsid or transgene, and 2 of 3 NHP in the delayed rATG group did not develop anti-capsid antibodies.

    “Neither group had cellular response to capsid or transgene, and 2 of 3 NHP in the delayed rATG group did not develop anti-capsid antibodies.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [41] Samelson-Jones, 2020, Timing of Intensive Immunosuppression Impacts Risk of TransgeneAntibodies after AAV Gene Therapy in Nonhuman Primates, Mol Ther - Methods Clin Dev source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Although no toxicity was found in the animal models, peripheral neuropathy and myelosuppression are adverse effects observed in humans for PIs.

    “Although no toxicity was found in the animal models, peripheral neuropathy and myelosuppression are adverse effects observed in humans (54).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [54] Merin, 2014, Clinical use of proteasome inhibitors in the treatment of multiple myeloma, Pharmaceuticals source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    After endosomal escape, rAAV particles either enter the nucleus for transgene expression, or become ubiquitylated then degraded by proteasome.

    “After endosomal escape, rAAV particles either enter the nucleus for transgene expression, or become ubiquitylated then degraded by proteasome (84).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [84] Dhungel, 2020, Journey to the Center of the Cell: Tracing the Path of AAV Transduction, Trends Mol Med source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    In a larger cohort (n=5) immunized with rAAV-LK03, anti-capsid IgM and IgG developed, not replicating the first IdeS re-administration result.

    “However, this was not replicated in a larger cohort (n=5) immunized with rAAV-LK03, that developed anti-capsid IgM and IgG.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [53] Leborgne, 2020, IgG-cleaving endopeptidase enables in vivo gene therapy in the presence of anti-AAV neutralizing antibodies, Nat Med source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Second-generation carfilzomib is irreversible and more specifically inhibits chymotrypsin-like activity than bortezomib, the reversible first-generation inhibitor, which also inhibits lysosomal and calcium-activated cellular proteases.

    “Second-generation carfilzomib is irreversible and more specifically inhibits chymotrypsin-like activity than bortezomib, the reversible first-generation inhibitor, which also inhibits lysosomal and calcium-activated cellular proteases (54, 83).”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [54] Merin, 2014, Clinical use of proteasome inhibitors in the treatment of multiple myeloma, Pharmaceuticals source

      Not checked in the free run.

    • cites [83] Fricker, 2020, Proteasome Inhibitor Drugs, Annu Rev Pharmacol Toxicol source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Intraperitoneal MnTBAP 80mg/kg/day from day 0-4 reversibly downregulated CD4 on T cells, inhibiting T cell priming and humoral responses to initial rAAV1 dosing, and allowing re-administration of rAAV1 via a different route 28 days later.

    “Intraperitoneal MnTBAP 80mg/kg/day from day 0-4 reversibly downregulated CD4 on T cells, inhibiting T cell priming and humoral responses to initial rAAV1 dosing, and allowing re-administration of rAAV1 via a different route 28 days later (95).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [95] Da Rocha, 2019, Temporary Reduction of Membrane CD4 with the Antioxidant MnTBAP Is Sufficient to Prevent Immune Responses Induced by Gene Transfer, Mol Ther - Methods Clin Dev source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    CRISPR-Cas9 allows genetic target-specific cleavage and editing.

    “CRISPR-Cas9 is a promising therapeutic tool that allows genetic target-specific cleavage and editing (111).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [111] Charlesworth, 2019, Identification of preexisting adaptive immunity to Cas9 proteins in humans, Nat Med source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Treg in liver and their persistence in muscle fibers after intermuscular delivery have been reported.

    “Reports on Treg in liver and their persistence in muscle fibers after intermuscular delivery (17) further emphasizes the need for Treg-sparing therapies.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [17] Mueller, 2013, Human treg responses allow sustained recombinant adeno-associatedvirus-mediated transgene expression, J Clin Invest source

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    The timing of immunomodulation, duration, and drug regimen itself have contributed to treatment efficacy.

    “As summarized above, a range of pharmacological agents have been used in clinical and preclinical studies, and the timing of immunomodulation, duration, and drug regimen itself have all contributed to treatment efficacy.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    In mice, binding (non-neutralizing) antibodies have a different biodistribution profile than NAb and higher efficacy in liver transduction.

    “Moreover, binding (non-neutralizing) antibodies in mice seemed to have a different biodistribution profile than NAb and higher efficacy in liver transduction (107).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [107] Fitzpatrick, 2018, Influence of Pre-existing Anti-capsid Neutralizing and Binding Antibodies on AAV Vector Transduction, Mol Ther - Methods Clin Dev source

      Not checked in the free run.

48 of 48

  • The paper relies on this without support.

    The eye is a relatively immunoprivileged site.

    “the eye is a relatively immunoprivileged site”
1 of 1

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • Prednisone, prednisolone, and dexamethasone were used pre-operation (1 to 3 days) and post-surgery (up to three weeks), orally or locally, in multiple clinical trials and with Luxturna.

    “Representatives of this family include prednisone, prednisolone, and dexamethasone, which were used pre-operation (for 1 to 3 days) as well as post-surgery (for longer periods of time, up to three weeks), taken orally or applied locally, in multiple clinical trials as well as in conjunction with EMA-and FDA-approved Luxturna [131,132].”

    From Risk mitigation of immunogenicity: a key to personalized retinal gene therapy (2021) · cites this paper as [131]

  • In recent clinical trials, glucocorticoids have been the most widely used anti-inflammatory drugs.

    “In recent clinical trials, glucocorticoids have been the most widely used anti-inflammatory drugs.”

    From Risk mitigation of immunogenicity: a key to personalized retinal gene therapy (2021) · cites this paper as [131]

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

Each contradiction between two passages listed here was confirmed by an independent check.

  • Mouse rapamycin study is classified as clinical

    Minor

    Table 1 classifies the AAV8 rapamycin combination study cited as reference 37 as clinical, but the text describes that same study as a murine model.

1 of 1

Cites

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3 of the 118 references couldn’t be matched to a known paper.

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Related

Authors
Chu, W. S., & Ng, J.
Affiliations
  • Royal Marsden NHS Foundation Trust (GB)
  • University College London (GB)
Corresponding author
Ng, Joanne
ORCID
Venue
Frontiers in Immunology
Publisher
Frontiers Media SA
Published
2021-04-01
Volume
12
Pages
658038-658038
ISSN
1664-3224 (ISSN-L)
DOI
10.3389/fimmu.2021.658038
PMC
PMC8049138
OpenAlex
W3149013390
Cited by
59
References
118
Citations by year
2026: 5 · 2025: 14 · 2024: 9 · 2023: 16 · 2022: 14 · 2021: 1
FWCI
1.47
Type
journal article
Language
en
Open access
Licence
Locations
  • Frontiers in Immunology (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
  • UCL Discovery (University College London) (submittedVersion)
Funders
  • Rosetrees Trust (MR/R015325/1, V1284)
  • Great Ormond Street Hospital Charity (MR/R015325/1, V1284)
  • Robert Luff Foundation (V1284, MR/R015325/1)
  • Medical Research Council (V1284, MR/R015325/1, MR/K02342X/1)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Virus-based gene therapy research
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Interference and Gene Delivery
  • Life Sciences › Immunology and Microbiology › Immunology › Immunotherapy and Immune Responses
Keywords
recombinant adeno-associated virus, gene therapy, immunomodulation, immunosuppression, innate and adaptive immunity, corticosteroids, Luxturna, Zolgensma, adjuvant pharmacotherapy, pre-existing immunity
MeSH terms
Clinical Studies as Topic, Dependovirus, Animals, Combined Modality Therapy, Drug Evaluation, Preclinical, Genetic Vectors, Humans, Immunity, Cellular, Immunologic Factors, Genetic Therapy, Gene Transfer Techniques, Transgenes, Immunity, Humoral, Immunomodulation, Cell- and Tissue-Based Therapy

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Claims read by deepseek-v4.1-flash under definition version 3 (extract-v3) on 2026-10-06.

Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-06.

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