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#viral vectors

4 public questions tagged with this topic.

Gene Therapy

Which viral vector shows very low immune response?

Recombinant AAV produced by triple transfection HEK293 cells with plasmids encoding rep/cap removed from vector, adenoviral helper functions E2A E4 VA RNA, and vector genome flanked by ITRs yields particles depleted viral coding sequences that normally generate PAMPs triggering innate and adaptive immunity. Capsid proteins VP1 VP2 VP3 ratio 1:1:10 from serotypes AAV2 AAV5 AAV9 exhibit low TLR2 mediated NF-kB activation and minimal TLR9 stimulation due to CpG depletion via codon optimization resulting reduced type I interferons, TNF-alpha, IL-12. Consequently transduced hepatocytes, muscle fibers, dorsal root ganglia display prolonged persistence without CTL elimination although capsid-specific CD8 cells can occur at high doses. Humoral neutralizing antibodies predominantly preexisting limits redosing but allows primary treatment. Episomal concatemers persist as circular monomers multimers associated with histones providing transcriptionally active chromatin for years minimal genotoxicity due low random integration

Ref: PMC AAV Low Immunogenicity Review PMCID 10142300; UQ Biomedical Sciences Vector Immunogenicity Comparison; Nature Med AAV Clinical Safety.

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Gene Therapy

Stable integration into host genome is a feature of:

Lentiviral vectors originating from HIV-1 carry two copies ssRNA genome 9 kb encapsidated with reverse transcriptase, integrase, nucleocapsid. Entry mediated by VSV-G pseudotyping binding LDL receptor ubiquitous, membrane fusion releasing core into cytoplasm where reverse transcriptase synthesizes ds cDNA via tRNA lys3 primer and strand transfer forming preintegration complex containing integrase, matrix p17, Vpr, host LEDGF p75 tethering factor. Complex traverses intact nuclear pore via importin alpha/beta pathway recognizing nuclear localization signals allowing transduction nondividing cells such as neurons and quiescent HSC unlike gamma retroviruses dependent on mitotic envelope breakdown. Integrase catalyzes 3 prime processing removing GT dinucleotides and strand transfer inserting provirus preferentially into active transcription units enriched H3K36 trimethylation marked by LEDGF reader ensuring expression. Self-inactivating design deletes U3 enhancer promoter in LTR abolishing LTR transcriptional activity reducing oncogenic transactivation risk. Integrated provirus replicates with host chromosomes providing stable long-term expression utilized in ex vivo hematopoietic correction and CAR-T manufacturing where durability essential despite low insertional risk.

Ref: Danaher Lentiviral Integration Mechanism; PMC Lentiviral Delivery CRISPR Review; NIH AAV vs Lentivirus Integration Safety.

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Gene Therapy

Which viral vector remains episomal and does not integrate into host genome?

Adenoviral vectors derived from nonenveloped icosahedral dsDNA viruses genome 26-36 kb flanked by inverted terminal repeats and packaging signal possess capacity 8 kb in first-generation deleted E1/E3 and up to 30 kb in helper-dependent gutless vectors lacking all viral coding sequences. Infection initiated by fiber knob binding CAR and alphaV beta3 integrin, internalization via clathrin pits, partial capsid disassembly releasing protein VI that lyses early endosomes at pH 6.0 permitting cytosolic transport via dynein to nuclear pore where genome translocates and remains extrachromosomal as linear and circular episome associated with histones without integrase. Expression from strong CMV promoter peaks 24-48 hours achieving high mRNA levels but declines over weeks in dividing tissue due to episomal loss and promoter methylation. Capsid hexon and fiber trigger innate immunity via TLR2, TLR9, NLRP3 inflammasome producing IL-6 and IL-1 beta and adaptive CTL response clearing transduced cells within two weeks. Transient high-level nonintegrating profile favors oncolytic therapy, vaccine delivery, and provision genome-editing enzymes.

Ref: Wiley Bioeng Transl Med Adenovirus Vector Zhao 2022; Danaher Viral Vector Comparison Episomal; NCBI Adenovirus Persistence Review.

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