Submitter Withdrawn 12th Australasian Virology Society Meeting 2024

mRNA delivery of ultra-potent HIV broadly neutralising monoclonal antibodies with engineered Fc-regions for expanded immune function. (#231)

Behnaz Heydarchi 1 , Fahimeh Mousavi 1 , Mehrnaz Bakhti 1 , Linda Earnest 1 , Melissa Barrow 1 , Damian Purcell 1
  1. Microbiology and Immunology, Doherty Institute, University of Melbourne, Melbourne, VIC, Australia

Broadly neutralising monoclonal antibodies (bNAbs) targeting conserved epitopes on Human Immunodeficiency Virus (HIV) envelope (Env) are effective antiviral therapeutics. We made an ultra-potent humanized bNAb (MEL-1872) with distinctive ultra-long CDRH3 regions of 57aa from bovine that was repeatedly vaccinated with a series of different stabilised Env-trimers1.  MEL-1872 binds the highly conserved CD4 binding site on HIV Env (IC50 0.009 mg/mL) without cross-reactivity to human cellular antigens. We explored mRNA delivery for expression of this bNAb antiviral preventing infectious spread and clearing HIV-infected cells. Sequence optimized immunoglobulin heavy (IgH) and light (IgL) chain plasmid templates were used for T7-RNA-pol in vitro transcription of pseudouridine-modified mRNAs incorporating a 5′ Cleancap, optimized 5′ and 3′ UTR, and a poly(A) tail. Stoichiometrically matched mixtures of purified IgH and IgL mRNAs achieved efficient expression of potent HIV-neutralising MEL-1872 after transfection into human and mouse cell-lines. We further engineered the IgG1 heavy chain2 to incorporate: a) the LS mutation binding neonatal Fc-receptor (FcRN) imparting systemic longevity from uptake recirculation; b) GASDALI mutation for FcgRIIA and FcgRIIIA binding for ADCC and NK-cell targeting; c) GASDALI/LS  dual mutants, and d) substituting IgG1H for the IgAH1&2 and J-chain for dimeric dIgA engagement of poly-IgA-receptor imparting mucosal secretion.  The suitability of mRNA delivery of MEL-1872 for viral neutralisation and cell-mediated antiviral activity was assessed in vitro using cell models including HIV productive or latent-infected cells. The mRNA mixtures were formulated into lipid nanoparticles (LNPs) tuned for either vaccination or immune-tolerant expression and delivered to ex vivo HIV-infected human T-cells or mice. Engineered MEL-1872 bNAb expression and potency for HIV neutralisation was confirmed from cell supernatant or mouse serum and mucosal washings. The anti-drug antibody responses against the distinctive bovine CDRH3 was assessed for different formulations to assess suitability of these mRNA-LNP formulations for clinical applications in HIV treatment and prevention.

  1. Heydarchi B, Fong DS, Gao H, Salazar-Quiroz NA, Edwards JM, Gonelli CA, Grimley S, Aktepe TE, Mackenzie C, Wales WJ, van Gils MJ, Cupo A, Rouiller I, Gooley PR, Moore JP, Sanders RW, Montefiori D, Sethi A, Purcell DFJ. Broad and ultra-potent cross-clade neutralization of HIV-1 by a vaccine-induced CD4 binding site bovine antibody. Cell Rep Med. 2022;3(5):100635.
  2. Edwards J, Heydarchi B, Khoury G, Salazar-Quiroz NA, Gonelli CA, Wines B, Hogarth PM, Kristensen AB, Parsons MS, Purcell DFJ. 2021. Enhancement of antibody‐dependent cellular cytotoxicity and phagocytosis in anti‐HIV‐1 bovine chimeric broadly-neutralizing antibodies. J Virol. 2021;95(13):e0021921.