Michael D. Swanson

A Lectin Isolated from Bananas Is a Potent Inhibitor of HIV Replication

BanLec is a jacalin-related lectin isolated from the fruit of bananas, Musa acuminata. This lectin binds to high mannose carbohydrate structures, including those found on viruses containing glycosylated envelope proteins such as human immunodeficiency virus type-1 (HIV-1). Therefore, we hypothesized that BanLec might inhibit HIV-1 through binding of the glycosylated HIV-1 envelope protein, gp120. We determined that BanLec inhibits primary and laboratory-adapted HIV-1 isolates of different tropisms and subtypes. BanLec possesses potent anti-HIV activity, with IC50 values in the low nanomolar to picomolar range. The mechanism for BanLec-mediated antiviral activity was investigated by determining if this lectin can directly bind the HIV-1 envelope protein and block entry of the virus into the cell. An enzyme-linked immunosorbent assay confirmed direct binding of BanLec to gp120 and indicated that BanLec can recognize the high mannose structures that are recognized by the monoclonal antibody 2G12. Furthermore, BanLec is able to block HIV-1 cellular entry as indicated by temperature-sensitive viral entry studies and by the decreased levels of the strong-stop product of early reverse transcription seen in the presence of BanLec. Thus, our data indicate that BanLec inhibits HIV-1 infection by binding to the glycosylated viral envelope and blocking cellular entry. The relative anti-HIV activity of BanLec compared favorably to other anti-HIV lectins, such as snowdrop lectin and Griffithsin, and to T-20 and maraviroc, two anti-HIV drugs currently in clinical use. Based on these results, BanLec is a potential component for an anti-viral microbicide that could be used to prevent the sexual transmission of HIV-1.

Discovery of selective bioactive small molecules by targeting an RNA dynamic ensemble

Current approaches used to identify protein-binding small molecules are not suited for identifying small molecules that can bind emerging RNA drug targets. By docking small molecules onto an RNA dynamic ensemble constructed by combining NMR spectroscopy and computational molecular dynamics, we virtually screened small molecules that target the entire structure landscape of the transactivation response element (TAR) from HIV type 1 (HIV-1). We quantitatively predict binding energies for small molecules that bind different RNA conformations and report the de novo discovery of six compounds that bind TAR with high affinity and inhibit its interaction with a Tat peptide in vitro (K(i) values of 710 nM-169 μM). One compound binds HIV-1 TAR with marked selectivity and inhibits Tat-mediated activation of the HIV-1 long terminal repeat by 81% in T-cell lines and HIV replication in an HIV-1 indicator cell line (IC(50) ∼23.1 μM).

Expression of Human Endogenous Retrovirus Type K (HML-2) Is Activated by the Tat Protein of HIV-1

ABSTRACT Human endogenous retroviruses (HERVs) make up 8% of the human genome. The expression of HERV-K (HML-2), the family of HERVs that most recently entered the genome, is tightly regulated but becomes markedly increased after infection with HIV-1. To better understand the mechanisms involved in this activation, we explored the role of the HIV-1 Tat protein in inducing the expression of these endogenous retroviral genes. Administration of recombinant HIV-1 Tat protein caused a 13-fold increase in HERV-K (HML-2) gag RNA transcripts in Jurkat T cells and a 10-fold increase in primary lymphocytes, and the expression of the HERV-K (HML-2) rec and np9 oncogenes was also markedly increased. This activation was seen especially in lymphocytes and monocytic cells, the natural hosts for HIV-1 infection. Luciferase reporter gene assays demonstrated that the effect of Tat on HERV-K (HML-2) expression occurred at the level of the transcriptional promoter. The transcription factors NF-κB and NF-AT contribute to the Tat-induced activation of the promoter, as shown by chromatin immunoprecipitation assays, mutational analysis of the HERV-K (HML-2) long terminal repeat, and treatments with agents that inhibit NF-κB or NF-AT activation. These studies demonstrate that HIV-1 Tat plays an important role in activating expression of HERV-K (HML-2) in the setting of HIV-1 infection.

Murine Colitis is Mediated by Vimentin

Vimentin, an abundant intermediate filament protein, presumably has an important role in stabilizing intracellular architecture, but its function is otherwise poorly understood. In a vimentin knockout (Vim KO) mouse model, we note that Vim KO mice challenged with intraperitoneal Escherichia coli control bacterial infection better than do wild-type (WT) mice. In vitro, Vim KO phagocytes show significantly increased capacity to mediate bacterial killing by abundant production of reactive oxygen species (ROS) and nitric oxides, likely due to interactions with the p47phox active subunit of NADPH oxidase. In acute colitis induced by dextran sodium sulfate (DSS), Vim KO mice develop significantly less gut inflammation than do WT mice. Further, Vim KO mice have markedly decreased bacterial extravasation in the setting of DSS-induced acute colitis, consistent with decreased intestinal disease. Our results suggest that vimentin impedes bacterial killing and production of ROS, thereby contributing to the pathogenesis of acute colitis.

Synergistic in vitro anti-HIV type 1 activity of tenofovir with carbohydrate-binding agents (CBAs)

Tenofovir, a well-known and highly prescribed anti-HIV-1 drug for the treatment of HIV/AIDS infections, has recently also shown its effectiveness as a potential microbicide drug in the prevention of HIV transmission. Here, we evaluated the combination of tenofovir with various members of the class of carbohydrate-binding agents (CBAs) targeting the glycans on the viral envelope gp120 for their anti-HIV efficacy. The tenofovir/CBA combinations predominantly showed synergistic antiviral activity using the median effect principle. These findings illustrate that combination of tenofovir with CBAs may increase the antiviral potency of the individual drugs and reducing the risk on potential side-effects.

T cells establish and maintain CNS viral infection in HIV-infected humanized mice

&NA; The human brain is an important site of HIV replication and persistence during antiretroviral therapy (ART). Direct evaluation of HIV infection in the brains of otherwise healthy individuals is not feasible; therefore, we performed a large‐scale study of bone marrow/liver/thymus (BLT) humanized mice as an in vivo model to study HIV infection in the brain. Human immune cells, including CD4+ T cells and macrophages, were present throughout the BLT mouse brain. HIV DNA, HIV RNA, and/or p24+ cells were observed in the brains of HIV‐infected animals, regardless of the HIV isolate used. HIV infection resulted in decreased numbers of CD4+ T cells, increased numbers of CD8+ T cells, and a decreased CD4+/CD8+ T cell ratio in the brain. Using humanized T cell‐only mice (ToM), we demonstrated that T cells establish and maintain HIV infection of the brain in the complete absence of human myeloid cells. HIV infection of ToM resulted in CD4+ T cell depletion and a reduced CD4+/CD8+ T cell ratio. ART significantly reduced HIV levels in the BLT mouse brain, and the immune cell populations present were indistinguishable from those of uninfected controls, which demonstrated the effectiveness of ART in controlling HIV replication in the CNS and returning cellular homeostasis to a pre‐HIV state.

BanLec, a banana lectin, is a potent inhibitor of Middle East respiratory syndrome coronavirus in in vitro assays

in in vitro assays Jasper Fuk-Woo Chan1,2,*, Kwok-Hung Chan1,2, Dan M. Boudreaux3, Michael D. Swanson4, David M. Markovitz3, Kwok-Yung Yuen1,2 1State Key Laboratory of Emerging Infectious Diseases, 2Department of Microbiology, The University of Hong Kong, Queen Mary Hospital, HKSAR, China; 3Department of Internal Medicine, Division of Infectious Diseases, University of Michigan, Ann Arbor, MI, USA; and 4Division of Infectious Diseases, University of North Carolina, Chapel Hill, NC, USA. (*Correspondence: Jasper F. W. Chan. Email: jfwchan@hku.hk. Tel: (852) 22554892. Fax: (852) 28551241.)