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Chimeric antigen receptor (CAR) T cell therapy is a relatively new and powerful way of transforming T cells with receptors needed to recognize and kill diseased cells. Traditionally, it involves extraction of T cells from a patient, ex vivo transformation of them with CARs, expansion, and subsequent re-infusion into the patient. Recent developments aim to avoid this lengthy, costly patient-specific procedure by using var- ious viral and non-viral vector particles for direct in vivo delivery of CAR-encoding genes. In this paper we highlight several fundamental connections between in vitro and in vivo aspects of this process. We dis- cuss the proposed use of in vitro-reconstituted virus-like particles (VLPs), prepared from purified CAR- encoding mRNA and viral capsid protein, and functionalized with a T cell-targeting antibody. We compare and contrast these particles – and their use as gene vectors – with the several modalities currently employed that involve in cellulo generation of lentiviral or AAV vectors or in vitro complexation of nucleic acids with cationic polymers or lipid vesicles. We report the unique stoichiometric preciseness and ther- modynamic stability of VLPs formed from anti-HIV-glycoprotein CAR-encoding mRNA and the capsid pro- tein from a plant virus, and quantify the extent to which these monodisperse spherical VLPs are RNase resistant and lead to strong CAR expression in T cells. Further, in vitro cell-killing experiments are pro- posed, in which these CAR VLP-transformed T cells are mixed with HIV-infected cells, to be followed by in vivo experiments involving injection of the particles into HIV-infected humanized mice.
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PSGL-1 restricts HIV-1 infectivity by blocking virus particle attachment to target cells
P-selectin glycoprotein ligand-1 (PSGL-1) is a dimeric, mucin-like, 120-kDa glycoprotein that binds to P-, E-, and L-selectins. PSGL-1 is expressed primarily on the surface of lymphoid and myeloid cells and is up-regulated during inflammation to mediate leukocyte tethering and rolling on the surface of endothelium for migration into inflamed tissues. Although it has been reported that PSGL-1 expression inhibits HIV-1 replication, the mechanism of PSGL-1–mediated anti-HIV activity remains to be elucidated. Here we report that PSGL-1 in virions blocks the infectivity of HIV-1 particles by preventing the binding of particles to target cells. This inhibitory activity is independent of the viral glycoprotein present on the virus particle; the binding of particles bearing the HIV-1 envelope glycoprotein or vesicular stomatitis virus G glycoprotein or even lacking a viral glycoprotein is impaired by PSGL-1. Mapping studies show that the extracellular N-terminal domain of PSGL-1 is necessary for its anti–HIV-1 activity, and that the PSGL-1 cytoplasmic tail contributes to inhibition. In addition, we demonstrate that the PSGL-1–related monomeric E-selectin–binding glycoprotein CD43 also effectively blocks HIV-1 infectivity. HIV-1 infection, or expression of either Vpu or Nef, down-regulates PSGL-1 from the cell surface; expression of Vpu appears to be primarily responsible for enabling the virus to partially escape PSGL-1–mediated restriction. Finally, we show that PSGL-1 inhibits the infectivity of other viruses, such as murine leukemia virus and influenza A virus. These findings demonstrate that PSGL-1 is a broad-spectrum antiviral host factor with a unique mechanism of action.
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- Award ID(s):
- 1662096
- PAR ID:
- 10192122
- Date Published:
- Journal Name:
- Proceedings of the National Academy of Sciences
- Volume:
- 117
- Issue:
- 17
- ISSN:
- 0027-8424
- Page Range / eLocation ID:
- 9537 to 9545
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.1073/pnas.1916054117
