Materials and methods for producing improved lentiviral vector particles
Inventors
Nicolai, Christopher James • Tareen, Semih U.
Assignees
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Publication Number
US-10391157-B2
Publication Date
2019-08-27
Expiration Date
Abstract
Materials and methods useful for generating highly mannosylated pseudotyped lentiviral vector particles comprising a Vpx protein are provided.
Core Innovation
The invention relates to improved pseudotyped lentiviral vector particle compositions and generation methods that preferentially bind dendritic cells expressing DC-SIGN. A lentiviral vector genome comprises an exogenous polynucleotide of interest, and the particles are pseudotyped with a polynucleotide encoding a Sindbis E2 glycoprotein that preferentially binds dendritic cells expressing DC-SIGN.
The production is performed in a culture medium comprising kifunensine and uses a virus packaging cell that includes a polynucleotide encoding a Vpx protein or a Vpr protein that retains SAMHD1-inhibiting activity. The approach uses highly mannosylated glycans rich in Man5–Man9, and the document links DC-SIGN-associated targeting performance with kifunensine and mannosidase inhibitors.
The compositions and kits are described for use in vivo, including adjuvants and formulation elements, and the document also reiterates optional integration-deficient lentiviral vectors. The system is presented as a DC-targeted pseudotyped lentiviral vector approach combined with adjuvant-based pharmaceutical compositions.
Claims Coverage
The document provides independent claim coverage focused on generating a pseudotyped lentiviral vector particle that preferentially binds DC-SIGN+ dendritic cells, with three core inventive features.
Kifunensine-containing production for DC-targeting pseudotyped particles
Generating a pseudotyped lentiviral vector particle by culturing a lentiviral vector packaging cell in a culture medium comprising kifunensine, then isolating a pseudotyped lentiviral vector particle that preferentially binds dendritic cells expressing DC-SIGN.
Sindbis E2 glycoprotein preferential for DC-SIGN+ dendritic cells
Including, in the lentiviral vector packaging cell, a polynucleotide encoding a Sindbis E2 glycoprotein that preferentially binds dendritic cells expressing DC-SIGN as part of the generated pseudotyped lentiviral vector particle.
Vpx/Vpr retaining SAMHD1-inhibiting activity
Including, in the lentiviral vector packaging cell, a polynucleotide encoding a Vpx protein or a Vpr protein that retains SAMHD1-inhibiting activity in generating the pseudotyped lentiviral vector particle, followed by isolating the particle that preferentially binds DC-SIGN+ dendritic cells.
Overall, the claim coverage centers on producing kifunensine-containing DC-targeted pseudotyped lentiviral particles by combining a DC-SIGN-preferring Sindbis E2 glycoprotein with a Vpx or Vpr protein retaining SAMHD1-inhibiting activity, and isolating particles that preferentially bind DC-SIGN+ dendritic cells.
Stated Advantages
Preferentially binds dendritic cells expressing DC-SIGN.
Unexpectedly enhanced targeting is described through combining highly mannosylated Sindbis envelope glycoproteins with mannosidase inhibitors to yield glycoforms rich in Man5–Man9.
Documented Applications
DC-based immunization delivery/application with tumor-specific antigens and viral-specific antigens.
Use of tumor-specific antigens including NY-ESO-1, MAGE-A3, MAGE-A1, and MART-1/Melan-A, and viral-specific antigens including HIV antigens.
Vaccines and therapeutics using pseudotyped lentiviral vector particles designed to preferentially bind DC-SIGN+ dendritic cells in vivo.
In vivo monitoring associated with the described pseudotyped lentiviral vector approach.
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