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Nature Communications Aug 2023The delivery of genetic cargo remains one of the largest obstacles to the successful translation of experimental therapies, in large part due to the absence of...
The delivery of genetic cargo remains one of the largest obstacles to the successful translation of experimental therapies, in large part due to the absence of targetable delivery vectors. Enveloped delivery modalities use viral envelope proteins, which determine tropism and induce membrane fusion. Here we develop DIRECTED (Delivery to Intended REcipient Cells Through Envelope Design), a modular platform that consists of separate fusion and targeting components. To achieve high modularity and programmable cell type specificity, we develop multiple strategies to recruit or immobilize antibodies on the viral envelope, including a chimeric antibody binding protein and a SNAP-tag enabling the use of antibodies or other proteins as targeting molecules. Moreover, we show that fusogens from multiple viral families are compatible with DIRECTED and that DIRECTED components can target multiple delivery chassis (e.g., lentivirus and MMLV gag) to specific cell types, including primary human T cells in PBMCs and whole blood.
Topics: Humans; Antibodies; Lentivirus; Membrane Fusion; Tropism; Viral Envelope Proteins
PubMed: 37612276
DOI: 10.1038/s41467-023-40788-8 -
Emerging Microbes & Infections Dec 2023Chronic inflammation and T cell dysregulation persist in individuals infected with human immunodeficiency virus type 1 (HIV-1), even after successful antiretroviral...
Chronic inflammation and T cell dysregulation persist in individuals infected with human immunodeficiency virus type 1 (HIV-1), even after successful antiretroviral treatment. The mechanism involved is not fully understood. Here, we used Olink proteomics to comprehensively analyze the aberrant inflammation-related proteins (IRPs) in chronic HIV-1-infected individuals, including in 24 treatment-naïve individuals, 33 immunological responders, and 38 immunological non-responders. T cell dysfunction was evaluated as T cell exhaustion, activation, and differentiation using flow cytometry. We identified a cluster of IRPs (cluster 7), including CXCL11, CXCL9, TNF, CXCL10, and IL18, which was closely associated with T cell dysregulation during chronic HIV-1 infection. Interestingly, IRPs in cluster 5, including ST1A1, CASP8, SIRT2, AXIN1, STAMBP, CD40, and IL7, were negatively correlated with the HIV-1 reservoir size. We also identified a combination of CDCP1, CXCL11, CST5, SLAMF1, TRANCE, and CD5, which may be useful for distinguishing immunological responders and immunological non-responders. In conclusion, the distinct inflammatory milieu is closely associated with immune restoration of T cells, and our results provide insight into immune dysregulation during chronic HIV-1 infection.
Topics: Humans; T-Lymphocytes; HIV-1; HIV Infections; Inflammation; Antigens, Neoplasm; Cell Adhesion Molecules
PubMed: 36408648
DOI: 10.1080/22221751.2022.2150566 -
Viruses Aug 2023The HIV-1 latent reservoir is considered the major barrier to achieve the eradication, although some evidences indicate that curing HIV-1 is a feasible goal [...].
The HIV-1 latent reservoir is considered the major barrier to achieve the eradication, although some evidences indicate that curing HIV-1 is a feasible goal [...].
Topics: Humans; Receptors, Chimeric Antigen; Immunotherapy, Adoptive; HIV Seropositivity; HIV-1; Cell- and Tissue-Based Therapy
PubMed: 37766199
DOI: 10.3390/v15091793 -
Nature Biomedical Engineering Apr 2024The blood-brain barrier (BBB) restricts the systemic delivery of messenger RNAs (mRNAs) into diseased neurons. Although leucocyte-derived extracellular vesicles (EVs)...
The blood-brain barrier (BBB) restricts the systemic delivery of messenger RNAs (mRNAs) into diseased neurons. Although leucocyte-derived extracellular vesicles (EVs) can cross the BBB at inflammatory sites, it is difficult to efficiently load long mRNAs into the EVs and to enhance their neuronal uptake. Here we show that the packaging of mRNA into leucocyte-derived EVs and the endocytosis of the EVs by neurons can be enhanced by engineering leucocytes to produce EVs that incorporate retrovirus-like mRNA-packaging capsids. We transfected immortalized and primary bone-marrow-derived leucocytes with DNA or RNA encoding the capsid-forming activity-regulated cytoskeleton-associated (Arc) protein as well as capsid-stabilizing Arc 5'-untranslated-region RNA elements. These engineered EVs inherit endothelial adhesion molecules from donor leukocytes, recruit endogenous enveloping proteins to their surface, cross the BBB, and enter the neurons in neuro-inflammatory sites. Produced from self-derived donor leukocytes, the EVs are immunologically inert, and enhanced the neuronal uptake of the packaged mRNA in a mouse model of low-grade chronic neuro-inflammation.
Topics: Animals; Neurons; Extracellular Vesicles; RNA, Messenger; Mice; Blood-Brain Barrier; Retroviridae; Capsid; Leukocytes; Humans; Mice, Inbred C57BL
PubMed: 38374224
DOI: 10.1038/s41551-023-01150-x -
Viruses Jul 2023Despite the availability of effective anti-HIV drug therapy, according to UNAIDS estimates, 1 [...].
Despite the availability of effective anti-HIV drug therapy, according to UNAIDS estimates, 1 [...].
Topics: Humans; HIV Infections; Retroviridae; Anti-HIV Agents; Molecular Biology
PubMed: 37515235
DOI: 10.3390/v15071549 -
Current Opinion in HIV and AIDS Mar 2024We explore the current status of research on HIV-1 subtype-specific variations and their impact on HIV-1 latency. We also briefly address the controversy surrounding the... (Review)
Review
PURPOSE OF REVIEW
We explore the current status of research on HIV-1 subtype-specific variations and their impact on HIV-1 latency. We also briefly address the controversy surrounding the decision-making process governing the ON/OFF states of HIV-1 transcription, specifically focusing on the regulatory elements, the long terminal repeat (LTR), and Tat. Understanding the decision-making process is crucial for developing effective intervention strategies, such as the 'shock-and-kill' approach, to reactivate latent HIV-1.
RECENT FINDINGS
Attention has been drawn to subtype-specific transcription factor binding site (TFBS) variations and the possible impact of these variations on viral latency. Further, diverse subtype-specific assays have been developed to quantify the latent viral reservoirs. One interesting observation is the relatively larger latent reservoirs in HIV-1B infection than those of other viral subtypes, which needs rigorous validation. The emergence of LTR-variant viral strains in HIV-1C demonstrating significantly higher levels of latency reversal has been reported.
SUMMARY
Despite persistent and substantial efforts, latent HIV-1 remains a formidable challenge to a functional cure. Determined and continued commitment is needed to understand the ON/OFF decision-making process of HIV-1 latency, develop rigorous assays for accurately quantifying the latent reservoirs, and identify potent latency-reversing agents and cocktails targeting multiple latency stages. The review emphasizes the importance of including diverse viral subtypes in future latency research.
Topics: Humans; Virus Latency; HIV Infections; HIV-1; Virus Activation; CD4-Positive T-Lymphocytes
PubMed: 38169308
DOI: 10.1097/COH.0000000000000835 -
Proceedings of the National Academy of... Jul 2023HIV-1 assembly occurs at the inner leaflet of the plasma membrane (PM) in highly ordered membrane microdomains. The size and stability of membrane microdomains is...
HIV-1 assembly occurs at the inner leaflet of the plasma membrane (PM) in highly ordered membrane microdomains. The size and stability of membrane microdomains is regulated by activity of the sphingomyelin hydrolase neutral sphingomyelinase 2 (nSMase2) that is localized primarily to the inner leaflet of the PM. In this study, we demonstrate that pharmacological inhibition or depletion of nSMase2 in HIV-1-producer cells results in a block in the processing of the major viral structural polyprotein Gag and the production of morphologically aberrant, immature HIV-1 particles with severely impaired infectivity. We find that disruption of nSMase2 also severely inhibits the maturation and infectivity of other primate lentiviruses HIV-2 and simian immunodeficiency virus, has a modest or no effect on nonprimate lentiviruses equine infectious anemia virus and feline immunodeficiency virus, and has no effect on the gammaretrovirus murine leukemia virus. These studies demonstrate a key role for nSMase2 in HIV-1 particle morphogenesis and maturation.
Topics: Animals; Cats; Horses; Mice; HIV-1; Sphingomyelin Phosphodiesterase; Virus Assembly; Lentivirus; Infectious Anemia Virus, Equine
PubMed: 37406093
DOI: 10.1073/pnas.2219475120 -
The Veterinary Record Nov 2023
Topics: Cats; Animals; Leukemia, Feline; Vaccination; Antibodies, Viral; Leukemia Virus, Feline; Cat Diseases; Immunodeficiency Virus, Feline
PubMed: 37921294
DOI: 10.1002/vetr.3641 -
The EMBO Journal Dec 2023Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication....
Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression.
Topics: Animals; Mice; Endogenous Retroviruses; Extracellular Vesicles; Cell Line, Tumor; Cell Differentiation; Dendritic Cells; Neoplasms
PubMed: 38073509
DOI: 10.15252/embj.2023113590 -
Nucleic Acids Research Aug 2023The discovery of reverse transcriptases (RTs) challenged the central dogma by establishing that genetic information can also flow from RNA to DNA. Although they act as...
The discovery of reverse transcriptases (RTs) challenged the central dogma by establishing that genetic information can also flow from RNA to DNA. Although they act as DNA polymerases, RTs are distantly related to replicases that also possess de novo primase activity. Here we identify that CRISPR associated RTs (CARTs) directly prime DNA synthesis on both RNA and DNA. We demonstrate that RT-dependent priming is utilized by some CRISPR-Cas complexes to synthesise new spacers and integrate these into CRISPR arrays. Expanding our analyses, we show that primer synthesis activity is conserved in representatives of other major RT classes, including group II intron RT, telomerase and retroviruses. Together, these findings establish a conserved innate ability of RTs to catalyse de novo DNA primer synthesis, independently of accessory domains or alternative priming mechanisms, which likely plays important roles in a wide variety of biological pathways.
Topics: DNA-Directed DNA Polymerase; HIV Reverse Transcriptase; Introns; Retroviridae; RNA; RNA-Directed DNA Polymerase; DNA Replication
PubMed: 37279911
DOI: 10.1093/nar/gkad478