Aug 2026· Frontiers in Molecular Medicine· 0 citations· 39 references
TL;DR
This proof-of-concept study underlines the utility of the Env variant BaEV-TR for the establishment of stable pseudotype VPCLs and the utilization of HT1080 cells as an alternative to frequently used 293T host cells.
Abstract
Lentiviral, α-retroviral, and γ-retroviral vector particles pseudotyped with envelope (Env) proteins of baboon endogenous retrovirus (BaEV) were shown to mediate highly efficient gene transfer into a range of primary human hematopoietic cell types in gene therapeutic strategies. However, the fusogenicity of utilized Env proteins mediated syncytia formation, hindering the establishment of stable viral packaging cell lines (VPCLs). Consequently, we used another BaEV Env variant, termed BaEV-TR—previously reported not to induce cell-to-cell fusion—and assessed its utility in stable producer cell line development. The absence of syncytia formation was reconfirmed in human 293T and HT1080 cell lines. Using HT1080 host cells, the stable VPCL HT/BaEV-TR was established. Conducting a Western blot analysis, BaEV-TR protein incorporation into vector particles was readily detected. Murine leukemia virus (MLV) pseudotype vectors efficiently transduced a variety of human target cell lines, reaching high vector titers of more than 3.0 × 10
6
transducing units per mL (TU/mL) in 293T target cells. In summary, this proof-of-concept study underlines the utility of the Env variant BaEV-TR for the establishment of stable pseudotype VPCLs and the utilization of HT1080 cells as an alternative to frequently used 293T host cells.
The HEK293T model stably expressing membrane-anchored HIV-1 gp160(ΔCT) model provides a robust platform for benchmarking the targeted recognition and cytotoxic activity of antiviral antibodies, agents, and CAR-T therapies in HIV research.
Jiarui Liu, Yang Gao, Changjun Wang et al.· Sheng wu gong cheng xue bao...· 0 citations
An industrial-grade platform based on monoclonal producer cell lines that enables the continuous and scalable generation of engineered virus-like particles (eVLPs) co-packaging Cas9–gRNA ribonucleoproteins (RNPs) and provides a GMP-compliant and broadly adaptable strategy for the streamlined manufacturing of next-generation autologous and allogeneic gene-edited CAR-T/NK therapies.
Wei Lin, Jiaru Shi, Hanyi Chen et al.· Frontiers in Immunology· 0 citations
A virus-like particle (VLP)-based toolkit that delivers diverse CRISPR editing modalities to human monocytes, macrophages and dendritic cells with high efficiency while preserving viability and innate immune responsiveness is presented.
Hyuncheol Jung, Pascal Devant, Carter Ching et al.· Nature Biotechnology· 0 citations
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
Human endogenous retroviruses (HERVs) originate from ancient retroviral integration into the primate germline. Although most are defective proviruses, the most recently endogenized groups, like the HERV-K family, retain intact ORFs encoding retroviral proteins. HERVs usually remain transcriptionally silent, yet this status is reversible. Multiple HIV-HERV interactions, mainly mediated by the HIV Tat protein, lead to HERV transcription and protein production. The present study investigates HERV-K transcription in particular of Human MMTV-like (HML) group-2 and 6 in peripheral blood of people with HIV (PWH). Using different experimental approaches—such as single-cell and plasma transcriptomics-, we found that HERV-K transcripts may be detected during both acute and chronic phases of the infection, with HML-6 showing higher expression compared to HML-2, predominantly within myeloid cells. Effective combined antiretroviral therapy (cART) was able to significantly reduce HML-6 transcription, regardless of whether the treatment was initiated in the acute or late chronic phases of HIV infection. Notably, chronic infections showed higher HML-6 transcript levels compared to acute infections in both naïve and successfully cART-treated subjects, potentially associated with persistent immune dysregulation observed in chronic HIV infection, although a direct causal role of HML-6 expression remains to be established.
E. Lazzari, Gabriella Rozera, Lucrezia Pierfederici et al.· International Journal of Mol...· 0 citations
Lentiviral vector (LV) production for CAR-T therapy remains challenging due to the limited scalability of adherent HEK293 cell cultures. To address this, we adapted HEK293FT cells directly to serum-free FreeStyle 293 Expression Medium, generating a novel suspension cell line, HEK293FT-DS. We characterized growth kinetics, stability over 35 passages, and transient transfection parameters (PEI:DNA ratio, cell density). The suspension-adapted cells grew with >93% viability and a specific growth rate of 0.54 day−1, and maintained stable viable cell density (1.51 ± 0.03 × 106 cells/mL) over 35 passages. The optimal PEI:DNA ratio was 2.5:1, and increasing the transfection cell density to 16 × 106 cells/mL boosted functional titers to approximately 9 × 106 TU/mL in clarified supernatant. Scaled production (1 L culture supernatant per batch) yielded up to 5 × 109 TU. The resulting vectors efficiently transduced primary human T cells (57% CAR-positive cells) in a CliniMACS Prodigy-based process. The platform based on suspension-adapted HEK293FT-DS cells enables high-density transfection and provides a cost-effective, scalable alternative to commercial LV production systems, particularly suited for academic CAR-T cell manufacturing.
A. Shevtsov, Aitolkyn Kydyrbayeva, Gaziza Nigmatulla et al.· Bioengineering· 0 citations