Aug 2026· Human Gene Therapy· Vol 37, pp. 770 - 787· 0 citations· 46 references
Medicine
TL;DR
Improved VSV-G variants with a better on/off-target ratio as attractive tools for in vivo gene therapy applications are presented.
Abstract
Targeted in vivo transduction, entailing direct administration of viral vector preparations to patients, is the next big step in gene therapy. To redirect lentiviral vector (LV) particles selectively to desired cell type(s), different glycoproteins have been engineered to alter their tropism, including the glycoprotein G from Vesicular Stomatitis Virus (VSV-G) as the most commonly employed tropism-defining protein for LV particles. For detargeting from its natural receptor, the low-density lipoprotein receptor (LDLR), a VSV-G variant with two blinding substitutions, K47Q and R354A, is commonly used (VSV-G.pub). We provide first evidence for insufficient blinding of VSV-G.pub in human and murine cell lines and primary cells. In silico modeling pointed toward only slightly reduced LDLR binding affinity of VSV-G.pub. To lower the affinity further, we generated three novel VSV-G variants based on charge-reversing substitutions in two, four, or six key residues. These variants achieved a more stringent blinding compared with VSV-G.pub in the tested cell lines and primary cells. Codisplay of a CD4 binder enabled lentiviral particles pseudotyped with the novel variants to selectively transduce CD4-expressing cells. In summary, we present improved VSV-G variants with a better on/off-target ratio as attractive tools for in vivo gene therapy applications.
Lentiviral vectors have revolutionized gene therapy by efficient and stable transduction of dividing and non-dividing cells, their large packaging capacity, and their compatibility with pseudotyping to alter viral tropism. The vesicular stomatitis virus glycoprotein (VSV-G) is widely used as a viral envelope protein of choice to pseudotype lentiviral vector particles as it confers exceptional particle stability and a broad tropism, due to the ubiquitous nature of the low-density lipoprotein receptor (LDLR). While this broad tropism facilitates transduction of diverse cell types, it precludes accurate in vivo targeting of specific cell populations. Structural insights into VSV-G have made receptor-blinding possible and revealed sites amenable to mutation while preserving fusion capacity. Coupled with targeting moieties, VSV-G pseudotyped lentiviral particles are redirected towards cells expressing target antigens. Such targeted vectors open new possibilities for in vivo gene therapy across oncology, infectious diseases, transplantation medicine, and other diseases. Use of targeted vectors will make in vivo gene therapy more accessible than cost-intensive ex vivo gene therapies. Since targeted vectors will be available as 'off-the-shelf' drugs, they will also drastically reduce time-to-treatment. This review highlights advances in bioengineering to exploit the versatility of VSV-G-pseudotyped lentiviral vectors and explores their vast potential for targeted gene delivery.
Anjali Shrivastava, Felix L. Warnecke, J. Schott et al.· Molecular Therapy· 0 citations
This review provides a comprehensive overview of the current status of viral and non-viral vector systems for in vivo and ex vivo applications, and key comparisons are made across safety, efficacy, scalability, and immune responses.
This efficient engineering process of Iterative Nicking for Synchronous Engineered Reprogramming of T cells (INSERT) establishes a safe, simplified platform for advanced therapeutic CAR T engineering.
Joseph G. Skeate, Nicholas J. Slipek, Walker S. Lahr et al.· Molecular Therapy· 0 citations
The results showed the potential of a combinatorial AAV library for model validation and revealed the human microliver platform-PEG as a reliable system for the development of AAV therapeutics.
Carmen Unzu, Amanda X. Chen, Liliana Mancio-Silva et al.· bioRxiv· 0 citations
An optimized, highly sensitive capillary-based Western method to measure the apparent isoelectric point (pI) and detect charge heterogeneity at the individual VP protein level under reduced denatured conditions is introduced.
Gangadhar Dhulipala, Kun Lu, N. Palackal et al.· Biophysica· 0 citations
PURPOSE OF REVIEW
To summarize current preclinical and clinical efforts to induce CD4bs-directed broadly neutralizing antibodies (bnAbs) against HIV-1 Env using germline-targeting and lineage-based vaccination strategies, and to highlight how differences in immunogen design, particularly along gradients of precursor affinity and structural constraint, shape VRC01-class responses and inform subsequent shaping and polishing steps in vaccine regimens.
RECENT FINDINGS
Phase 1 trials of CD4bs-directed germline-targeting immunogens have shown that VRC01-class precursors can be reproducibly expanded in humans. Smaller gp120-based immunogens exhibit higher precursor affinity and recruit larger, more diverse VRC01-class pools, whereas native-like trimers prime fewer lineages but drive more on-track somatic hypermutation, early glycan accommodation, and neutralization of heterologous viruses. In parallel, preclinical work is centred around the selection of shaping and polishing immunogens, alternative dosing strategies, and controlled-release or mRNA-LNP platforms to extend maturation.
SUMMARY
With proof-of-concept for VRC01-class priming being achieved, the next challenge is to induce strong serum antibodies and memory B cells with breadth and potency. Progress toward a protective CD4bs-directed HIV-1 vaccine will require further optimization of adjuvants, delivery systems, and sequential immunogen design to balance precursor recruitment with structural constraint and to sustain affinity maturation across multidose regimens, ultimately driving rare lineages to durable bnAb responses in diverse populations.
Catarina Mendes Silva, R. Sanders, Tom G. Caniels· Current Opinion in HIV and A...· 0 citations