Biology subjects

McGee, J. E.

Publications and source records attributed to McGee, J. E..

3 recordsLinked to original sources

bioRxiv2026

Single injection of modified self-amplifying RNA encoding a CD19 bispecific T cell engager mediates long-term malignant B cell clearance

Bispecific T cell engagers (BiTEs) are a novel cancer immunotherapy modality that achieves significant clinical success. However, conventional single-chain variable fragment (scFv)-based BiTE therapy requires continuous intravenous infusion due to BiTEs short half-life, limiting patient access and increasing healthcare cost. Self-amplifying RNA (saRNA), an emerging RNA technology, enables durable protein production in situ. Here, we report a 5-methylcytidine (m5C)-modified saRNA encoded BiTE system (saRNA-BiTE) targeting CD19. saRNA-BiTE induces prolonged, antigen-specific target cell lysis in vitro. In an acute lymphoblastic leukemia rechallenge model, a single intravenous injection of saRNA-BiTE formulated in lipid nanoparticles eradicates malignant cells and prevents disease recurrence for 3 months. saRNA-BiTE maintains more stable systemic level than protein BiTE or mRNA-BiTE without generating an initial burst BiTE exposure and affords functional BiTE expression for 6 weeks post administration. This work establishes saRNA-BiTE as a robust platform for extended in situ BiTE expression with enhanced long-term efficacy and facile production.

bioengineering↗

bioRxiv2025

Modified self-amplifying RNAs mediate robust and prolonged gene expression in the mammalian brain

Facile, non-genomic integrating gene delivery technologies are lacking for rapid onset and prolonged protein expression in the brain. Here we report the protein expression and cell type tropism for an advanced messenger ribonucleic acid (mRNA) technology, modified 5-hydroxymethylcytidine (hm5C) self-amplifying ribonucleic acid (saRNA), when injected into the mouse brain or applied to ex vivo human cortical brain slices. saRNA, encoding fluorescent proteins, encapsulated in an LNP formulation comprising ALC-0315 (present in Comirnaty(R)) efficiently mediates long-lasting protein expression in mouse brain cells beyond five weeks, with detectable expression in some neurons at three months. hm5C saRNA substantially outperforms N1m{Psi} mRNA. In addition to transfecting astrocytes and neurons at the injection site, hm5C saRNA-LNPs label neurons retrogradely. Excitingly, hm5C saRNA-LNPs afford protein expression in human cortical brain slices, with expression emerging within 24 hours and lasting beyond 76 days. Modified saRNA provides new opportunities for mechanistic neuroscience research and therapeutic development.

neuroscience↗

bioRxiv2023

Complete substitution with modified nucleotides suppresses the early interferon response and increases the potency of self-amplifying RNA

Self-amplifying RNA (saRNA) will revolutionize vaccines and in situ therapeutics by enabling protein expression for longer duration at lower doses. However, a major barrier to saRNA efficacy is the potent early interferon response triggered upon cellular entry, resulting in saRNA degradation and translational inhibition. Substitution of mRNA with modified nucleotides (modNTPs), such as N1-methylpseudouridine (N1m{Psi}), reduce the interferon response and enhance expression levels. Multiple attempts to use modNTPs in saRNA have been unsuccessful, leading to the conclusion that modNTPs are incompatible with saRNA, thus hindering further development. Here, contrary to the common dogma in the field, we identify multiple modNTPs that when incorporated into saRNA at 100% substitution confer immune evasion and enhance expression potency. Transfection efficiency enhances by roughly an order of magnitude in difficult to transfect cell types compared to unmodified saRNA, and interferon production reduces by >8 fold compared to unmodified saRNA in human peripheral blood mononuclear cells (PBMCs). Furthermore, we demonstrate expression of viral antigens in vitro and observe significant protection against lethal challenge with a mouse-adapted SARS-CoV-2 strain in vivo. A modified saRNA vaccine, at 100-fold lower dose than a modified mRNA vaccine, results in a statistically improved performance to unmodified saRNA and statistically equivalent performance to modified mRNA. This discovery considerably broadens the potential scope of self-amplifying RNA, enabling entry into previously impossible cell types, as well as the potential to apply saRNA technology to non-vaccine modalities such as cell therapy and protein replacement.

bioengineering↗