Engineered adeno-associated virus 3 vector with reduced reactivity to serum antibodies

Ito, Mika; Takino, Naomi; Nomura, Takeshi; Kan, Akihiko; Muramatsu, Shin‐ichi

doi:10.1038/s41598-021-88614-9

Cited by 16 publications

(15 citation statements)

References 44 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…A recent report showed that the CRISPR system was successfully delivered into human hepatocytes from humanized liver chimeric mice using genetically modified AAV, [ 17 ] which could be a potential HBV‐CRISPR delivery option in patients. In particular, a genetically modified AAV vector (AAV.GT5) has been reported to have high transduction efficiency in human hepatocytes and low reactivity with neutralizing antibodies, [ 10 ] which would enable repeated administration to deliver CRISPR into all HBV‐infected hepatocytes in the liver. A potential concern about CRISPR treatment is that it may induce DSBs of the HBV DNA integrated in the host genome and cause genome instability and cytotoxicity.…”

Section: Discussionmentioning

confidence: 99%

“…[ 6 ] Thus, it is difficult to eradicate cccDNA from infected hepatocytes with currently available antiviral therapies. Several potential new therapies, including capsid assembly modulators, [ 7 , 8 , 9 , 10 ] nucleic acid–based polymers, and antisense oligonucleotides tested in clinical trials, also do not directly target cccDNA. Therefore, further development of therapeutics targeting cccDNA is awaited.…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Inhibition of nonhomologous end joining‐mediated DNA repair enhances anti‐HBV CRISPR therapy

et al. 2022

View full text Add to dashboard Cite

Current anti–hepatitis B virus (HBV) therapies have little effect on covalently closed circular DNA (cccDNA) and fail to eliminate HBV. The clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9 system has been reported to directly target cccDNA and exert antiviral effects. In this study, we hypothesized that the inhibition of the DNA repair machinery, which is important for the repair of CRISPR‐induced double‐strand breaks, may enhance the effect of CRISPR targeting cccDNA, and we investigated the antiviral effect of potential combination therapy. The antiviral effect of CRISPR targeting cccDNA (HBV‐CRISPR) was evaluated in HBV‐susceptible HepG2‐hNTCP‐C4 cells expressing Cas9 (HepG2‐hNTCP‐C4‐iCas9) or primary human hepatocytes (PHHs) expressing Cas9. Following HBV infection, HBV‐CRISPR reduced cccDNA levels, accompanied by decreases in pregenomic RNA (pgRNA) levels and supernatant HBV DNA, hepatitis B surface antigen and hepatitis B e antigen levels in HepG2‐hNTCP‐C4‐iCas9 cells, and PHHs. HBV‐CRISPR induced indel formation in cccDNA and up‐regulated poly(adenosine diphosphate ribose) polymerase (PARP) activity in HBV‐infected HepG2‐hNTCP‐C4‐iCas9 cells. The suppression of PARP2‐Histone PARylation factor 1 (HPF1) (involved in the initial step of DNA repair) with small interfering RNA (siRNA) targeting either PARP2 or HPF1 increased the reduction in pgRNA and cccDNA by HBV‐CRISPR in HBV‐infected HepG2‐hNTCP‐C4‐iCas9 cells. The suppression of DNA Ligase 4 (LIG4) (essential for nonhomologous end joining [NHEJ]) but not breast cancer susceptibility gene (BRCA) (essential for homologous recombination) enhanced the antiviral effect of HBV‐CRISPR in HBV‐infected HepG2‐hNTCP‐C4‐iCas9 cells. Finally, the clinically available PARP inhibitor olaparib increased the reductions in pgRNA and cccDNA levels induced by HBV‐CRISPR in HBV‐infected HepG2‐hNTCP‐C4‐iCas9 cells and PHHs. Conclusion : The suppression of the NHEJ‐mediated DNA repair machinery enhances the effect of CRISPR targeting cccDNA. The combination of CRISPR and olaparib may represent a therapy for HBV elimination.

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Inhibition of nonhomologous end joining‐mediated DNA repair enhances anti‐HBV CRISPR therapy

et al. 2022

View full text Add to dashboard Cite

show abstract

“…AAV5 and AAV8, which have been widely used in gene therapy with the liver as the target in clinical trials, and AAV3B, which reportedly possesses a high gene transfer efficiency to the human liver. 12,20,21…”

Section: Comparison Of the Capsid Amino Acid Sequence Among Aav Serot...mentioning

confidence: 99%

Successful Liver transduction by Re-administration of Different Adeno-Associated Virus Vector Serotypes in Mice

Baatartsogt

Kashiwakura

Hiramoto

et al. 2022

Preprint

View full text Add to dashboard Cite

Intravenous administration of adeno-associated virus (AAV) vector is a promising gene therapy approach for monogenic diseases. However, re-administration of the same AAV serotype is impossible due to the induction of anti-AAV neutralizing antibodies (NAbs). Here we examined the feasibility of re-administration of AAV vectors to change the serotypes. We administered AAV3B, AAV5, or AAV8 vectors targeting the liver of C57BL/6 mice intravenously, and then assessed the emergence of NAbs and the transduction efficacy with a second administration. For all serotypes, we confirmed that re-administration with the same serotype was not possible. Although the highest neutralizing activity of NAb was induced by AAV5; however, the NAbs elicited by AAV5 did not react with any other serotypes, resulting in success in re-administration with the other serotypes. The re-administration of AAV5 was also successful in all mice treated with AAV3B and AAV8. The effective secondary administration of AAV3B and AAV8 was observed in most mice treated with AAV8 and AAV3B, respectively. However, few mice developed NAbs cross-reactive with the other serotypes, especially the serotypes with close sequence homology. In summary, AAV vector administration induced NAbs relatively specific to the serotype administrated. Secondary administration of AAVs targeting liver transduction could be successfully achieved by switching AAV serotypes in mice.

show abstract

“…4 AAV.GT5 showed robust transgene expression in human primary hepatocytes as well as Huh-7 and HepG2 (Supplemental figure S2), as reported previously. 16 Transduction efficiency of human primary hepatocytes with AAV.GT5 was more than 100 times higher than AAV-Spark100 and AAV5 (Supplemental figure S2). Conversely, AAV-Spark100 showed the most efficient transduction in murine hepatocyte cell line TLR3 (Supplemental figure S2).…”

Section: Efficient Transgene Expression In Human Hepatocytes By Aavgt...mentioning

confidence: 99%

Efficient Gene Transduction in Pigs and Macaques with the Engineered AAV Vector AAV.GT5 for Hemophilia B Gene Therapy

Kashiwakura

Endo

Ugajin

et al. 2022

Preprint

Self Cite

View full text Add to dashboard Cite

Gene therapy for hemophilia using adeno-associated virus (AAV) vectors allows long-term coagulation factor expression. We examined the potential of a novel engineered liver-tropic AAV3B-based vector AAV.GT5 for hemophilia B gene therapy. In vitro transduction with AAV.GT5 in human hepatocytes was more than 100 times higher than with AAV-Spark100, while in vivo transduction efficacy into the liver and the increase in coagulation factor IX (FIX) antigen following intravenous injection of these vectors were similar in PXB mice (chimeric mice with a humanized liver) and macaques. The discrepancy was due to the low recovery and short half-life of AAV.GT5 in blood, depending on the positive charge of the heparin-binding site in the original AAV3B. The intra-hepatic vascular administration of AAV.GT5, but not AAV-Spark100, enhanced vector transduction into the liver and reduced vector distribution to the kidney in pigs. In macaques, the intra-hepatic artery injection of AAV.GT5 yielded a comparable increase in FIX antigen with a one-third dosage of peripheral venous administration. Two of four macaques who received AAV.GT5 intravenously did not develop neutralizing antibodies (NAbs) against AAV.GT5, while AAV-Spark100 induced serotype-specific NAbs in all four macaques. The NAb produced after the administration was relatively specific to the serotype and less responsive to the other serotype. As a result, the administration of AAV.GT5 successfully boosted FIX expression in one animal previously given AAV-Spark100. Thus, AAV.GT5 has different biodistribution and immunogenic characteristics compared with AAV-Spark100, and the intra-hepatic vascular administration may lessen the vector dose and avoid vector distribution to other organs.

show abstract

Engineered adeno-associated virus 3 vector with reduced reactivity to serum antibodies

Cited by 16 publications

References 44 publications

Inhibition of nonhomologous end joining‐mediated DNA repair enhances anti‐HBV CRISPR therapy

Inhibition of nonhomologous end joining‐mediated DNA repair enhances anti‐HBV CRISPR therapy

Successful Liver transduction by Re-administration of Different Adeno-Associated Virus Vector Serotypes in Mice

Efficient Gene Transduction in Pigs and Macaques with the Engineered AAV Vector AAV.GT5 for Hemophilia B Gene Therapy

Contact Info

Product

Resources

About