Self-assembling insect ferritin nanoparticles
Inventors
Kwong, Peter • Georgiev, Ivelin • Joyce, Michael Gordon • Kanekiyo, Masaru • Druz, Aliaksandr • Baxa, Ulrich • Van Galen, Joseph • Cheng, Cheng • Mascola, John • Tsybovsky, Yaroslav • Yang, YongPing • Graham, Barney • Moin, Syed Mohammad • Boyington, Jeffrey
Assignees
US Department of Health and Human Services
Publication Number
US-11939356-B2
Publication Date
2024-03-26
Expiration Date
2037-06-27
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Abstract
Disclosed are recombinant insect ferritin nanoparticles that can be used to display two different trimeric antigens at an equal ratio. Also disclosed are nucleic acids encoding the recombinant insect ferritin nanoparticles and methods of producing the recombinant insect ferritin nanoparticles. Methods for eliciting an immune response in a subject are also provided.
Core Innovation
This disclosure provides novel recombinant insect ferritin nanoparticles engineered to display two different trimeric antigens at an equal ratio. Unlike bacterial ferritin nanoparticles, which comprise 24 identical subunits, insect ferritin nanoparticles include twelve copies each of two different subunits termed heavy and light chains, which assemble into a globular nanoparticle presenting four trimers of heavy chains and four trimers of light chains. By fusing different viral protein antigens to the heavy and light chains, the resulting nanoparticle displays two distinct trimeric antigens simultaneously.
The problem addressed is that prior bacterial ferritin nanoparticle technologies permit only random co-assembly of diverse trimeric antigens, lacking control over the pattern and ratio of antigen presentation on a single nanoparticle. This inhibits the ability to ensure a regular geometric pattern and equal ratio display of two different trimeric antigens, which can be crucial for eliciting broadly neutralizing antibody responses. The disclosed recombinant insect ferritin nanoparticles overcome this limitation by self-assembling with defined incorporation of heavy and light chains fused to different antigens, achieving a tetrahedral symmetry with eight heterologous trimeric antigens displayed in a 1:1 ratio.
The recombinant insect ferritin nanoparticles comprise heavy and light chain polypeptides from Lepidoptera insects such as Trichoplusia ni and Manduca, with specific N-terminal truncations to optimally orient attached trimeric antigens like viral envelope ectodomains on the nanoparticle surface. The technology encompasses nucleic acids encoding these fusion polypeptides, methods of producing the nanoparticles, compositions including them, and methods of eliciting immune responses by administering these nanoparticles to subjects. Importantly, the nanoparticles can display trimeric antigens derived from viruses such as HIV-1, influenza, respiratory syncytial virus (RSV), and metapneumovirus (MPV), including heterologous combinations such as different strains or different viruses on the same particle, thereby enhancing breadth of immune recognition.
Claims Coverage
The patent discloses a set of inventive features mainly encompassed in one independent claim directed to a recombinant insect ferritin nanoparticle and related products and methods. Two independent nucleic acid and method claims are also included. The features focus on the composition of the nanoparticle using specific truncated ferritin heavy and light chains fused to viral proteins, and its use to induce immune responses.
Recombinant insect ferritin nanoparticle comprising truncated ferritin subunits fused to viral proteins
A recombinant insect ferritin nanoparticle consisting of twelve recombinant ferritin heavy chains and twelve recombinant ferritin light chains self-assembled into a globular nanoparticle. The heavy chains consist of 172-174 amino acids from the C-terminus of insect ferritin heavy chain, and light chains consist of 182-184 or 177 amino acids from the C-terminus of insect ferritin light chain. Each ferritin chain is N-terminally fused to a viral protein. The ferritin heavy and light chains each have at least 90% sequence identity to either SEQ ID NO: 2 and 6 respectively, or SEQ ID NO: 4 and 8 respectively.
Specific sequence identity levels of ferritin heavy and light chains
Ferritin heavy and light chains can have at least 90% or 95% sequence identity to SEQ ID NO: 2 and 6 (Trichoplusia ni sequences) or SEQ ID NO: 4 and 8 (Manduca sequences), or can include the exact sequences of SEQ ID NO: 2 and 6 or SEQ ID NO: 4 and 8.
Viral protein type fused to ferritin subunits
The viral protein fused to the ferritin heavy and light chains is a viral envelope protein, which can be from viruses such as HIV-1, influenza, RSV, MPV, HPIV, Ebola, Marburg, MERS coronavirus, or SARS coronavirus. The viral protein can be an influenza HA stem protein.
Nucleic acid molecules encoding the recombinant insect ferritin polypeptides
Isolated nucleic acid molecules encoding the recombinant insect ferritin heavy chain and/or light chain fusion proteins, which can be DNA or mRNA, operably linked to promoters, are covered. Vectors containing such nucleic acids and host cells transformed with these vectors are claimed.
Method of producing recombinant insect ferritin nanoparticle
A method including transfecting a permissive cell culture with the expression vector encoding the recombinant ferritin fusion proteins, incubating to allow expression, and purifying the assembled insect ferritin nanoparticles.
Composition comprising the recombinant insect ferritin nanoparticle
A pharmaceutical composition including the recombinant insect ferritin nanoparticle and a pharmaceutically acceptable carrier.
Method of inducing immune response to viral envelope proteins
Administering an effective amount of the recombinant insect ferritin nanoparticle or nucleic acid encoding its components to a subject to induce an immune response to the viral envelope proteins displayed on the nanoparticle.
The claims cover recombinant insect ferritin nanoparticles composed of specific truncated heavy and light chain subunits fused N-terminally to viral proteins, notably viral envelope proteins from diverse viruses, nucleic acid sequences encoding these polypeptides, vectors and host cells for their expression, methods for producing the nanoparticles, their pharmaceutical compositions, and methods of inducing immune responses by administering these nanoparticles. The claims emphasize the use of insect ferritin sequences with defined truncations and fusions to display two different trimeric antigens at an equal ratio on the nanoparticles.
Stated Advantages
Improved breadth of immune recognition by simultaneous display of two diverse trimeric antigens on a single nanoparticle.
Regular geometric pattern and equal ratio presentation of multiple antigen trimers on the nanoparticle surface enhance B cell activation and antibody responses.
Ability to present two distinct antigens in controlled composition and geometry, unlike prior bacterial ferritin nanoparticles which allow only random co-assembly.
Improved expression and stable assembly of the modified insect ferritin nanoparticles with truncated heavy and light chains.
Broad applicability to various viral antigens including HIV-1, influenza, RSV and MPV, demonstrated with recombinant nanoparticles eliciting neutralizing antibodies in animal models.
Documented Applications
Vaccination and immunization to elicit immune responses against viral infections, including HIV-1, influenza, respiratory syncytial virus (RSV), and metapneumovirus (MPV).
Use of recombinant insect ferritin nanoparticles displaying two different trimeric viral envelope protein ectodomains to induce neutralizing antibody responses with improved breadth and potency in subjects.
Production of nanoparticle-based immunogenic compositions for preventing or treating viral infections by vaccination.
Methods of administering recombinant insect ferritin nanoparticles or nucleic acid molecules encoding them to subjects at risk of or infected with viral diseases to generate protective or therapeutic immune responses.
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