Biology subjects

Tietjen, I.

Publications and source records attributed to Tietjen, I..

6 recordsLinked to original sources

bioRxiv2024

Ex vivo and in vivo HIV-1 latency reversal by Mukungulu, a protein kinase C-activating African medicinal plant extract

Current HIV latency reversing agents (LRAs) have had limited success in clinic, indicating the need for new strategies that can reactivate and/or eliminate HIV reservoirs. "Mukungulu," prepared from the bark of Croton megalobotrys Mull. Arg., is traditionally used for HIV/AIDS management in Northern Botswana despite containing an abundance of protein kinase C (PKC)-activating phorbol esters ("namushens"). Here we show that Mukungulu is tolerated in mice at up to 12.5 mg/kg while potently reversing latency in antiretroviral therapy (ART)-suppressed HIV-infected humanized mice at 5 mg/kg. In peripheral blood mononuclear cells (PBMC) and isolated CD4+ T-cells from ART-suppressed people living with HIV-1, 1 {micro}g/mL Mukungulu reverses latency on par with or superior to anti-CD3/CD28 positive control, as measured by HIV gag-p24 protein expression, where the magnitude of HIV reactivation in PBMC corresponds to intact proviral burden levels in CD4+ T-cells. Bioassay-guided fractionation identifies 5 namushen phorbol ester compounds that reactivate HIV expression, yet namushens alone do not match Mukungulus activity, suggesting additional enhancing factors. Together, these results identify Mukungulu as a robust natural LRA which may warrant inclusion in future LRA-based HIV cure and ART-free remission efforts.

microbiology↗

bioRxiv2024

A high-throughput, microplate reader-based method to monitor in vitro HIV latency reversal in the absence of flow cytometry

J-Lat cells are derivatives of the Jurkat CD4+ T cell line that contain a non-infectious, inducible HIV provirus with a GFP tag. While these cells have substantially advanced our understanding of HIV latency, their use by many laboratories in low and middle-income countries is restricted by limited access to flow cytometry. To overcome this barrier, we describe a modified J-Lat assay using a standard microplate reader that detects HIV-GFP expression following treatment with latency-reversing agents (LRAs). We show that HIV reactivation by control LRAs like prostratin and romidepsin is readily detected with dose dependence and with significant correlation and sensitivity to standard flow cytometry. For example, 10 {micro}M prostratin induced a 20.1 {+/-} 3.3-fold increase in GFP fluorescence in the microplate reader assay, which corresponded to 64.2 {+/-} 5.0% GFP-positive cells detected by flow cytometery. Similarly, 0.3 {micro}M prostratin induced a 1.7 {+/-} 1.2-fold increase compared to 8.7 {+/-} 5.7% GFP-positive cells detected. Using this method, we screen 79 epigenetic modifiers and identify molibresib, quisinostat, and CUDC-101 as novel LRAs. This microplate reader-based method offers accessibility to researchers in resource-limited regions to work with J-Lat cells and more actively participate in global HIV cure research efforts. HighlightsO_LIJ-Lat T-cell lines are important to HIV cure research but require flow cytometry C_LIO_LIWe describe a method to work with J-Lat cells using a standard microplate reader C_LIO_LIThis assay can detect control LRAs similar to flow cytometry and discover new LRAs C_LIO_LIThis assay allows low-resourced laboratories to contribute to HIV cure research C_LI

microbiology↗

bioRxiv2024

Antiviral Mechanisms and Preclinical Evaluation of Amantadine Analogs that Continue to Inhibit Influenza A Viruses with M2 S31N-Based Drug Resistance

To better manage seasonal and pandemic influenza infections, new drugs are needed with enhanced activity against contemporary amantadine- and rimantadine-resistant influenza A virus (IAV) strains containing the S31N variant of the viral M2 ion channel (M2S31N). Here we tested 36 amantadine analogs against a panel of viruses containing either M2S31N or the parental, M2 S31 wild-type variant (M2WT). We found that several analogs, primarily those with sizeable lipophilic adducts, inhibited up to three M2S31N-containing viruses with activities at least 5-fold lower than rimantadine, without inhibiting M2S31N proton currents or modulating endosomal pH. While M2WT viruses in passaging studies rapidly gained resistance to these analogs through the established M2 mutations V27A and/or A30T, resistance development was markedly slower for M2S31N viruses and did not associate with additional M2 mutations. Instead, a subset of analogs, exemplified by 2-propyl-2-adamantanamine (38), but not 2-(1-adamantyl)piperidine (26), spiro[adamantane-2,2-pyrrolidine] (49), or spiro[adamantane-2,2-piperidine] (60), inhibited cellular entry of infectious IAV following pre-treatment and/or H1N1 pseudovirus entry. Conversely, an overlapping subset of the most lipophilic analogs including compounds 26, 49, 60, and others, disrupted viral M2-M1 protein colocalization required for intracellular viral assembly and budding. Finally, a pilot toxicity study in mice demonstrated that 38 and 49 were tolerated at doses approaching those of amantadine. Together, these results indicate that amantadine analogs act on multiple, complementary mechanisms to inhibit replication of M2S31N viruses. Highlights- Current IAVs have M2 mutations that confer resistance to amantadine and rimantadine - Several amantadine analogs inhibit these viruses without acting on M2 proton currents - Alternative antiviral targets include IAV entry and M2-M1 protein colocalization - Amantadine analogs are also tolerated in mice - Future amantadine antivirals could simultaneously act on multiple IAV mechanisms

microbiology↗

bioRxiv2024

Traditional uses and pharmacological activities of Tetracera alnifolia (Wild) Drake

Ethnopharmacological relevanceTetracera alnifolia (Wild) Drake, is well used in traditional Guinean medicine for the treatment of infectious skin diseases. The present aim was to contribute to the valorization of Tetracera alnifolia leaves, focused on ethnomedical, biological and phytochemical investigations. Materials and methodswe conducted an ethnomedical survey across several markets of the city of Conakry to identify 39 healers. Chloroform, methanol, dichloromethane, and aqueous extracts were tested for activities against protozoa, bacteria, fungi, HIV, and SARS-CoV-2. Resultsthe traditional healers indicated that T. alnifolia is used in the treatment of more than 15 pathologies including Fassa (marasmus/malnutrition), Soukhou kouye (white discharge in women), and Temou bankhi (sexual weakness in men). Leaves were the most used part. The modes of preparation included decoction and powder. Data from biological activities identicatied good activities of the methanolic extract against Leishmania infantum (MIC = 8.11 g / ml) and a moderate activity on Trypanosoma brucei (MIC = 28.15 g / ml) and Staphylococcus aureus (MIC = 29.91 g / ml), while dichloromethane extracts acted on live SARS-CoV-2 replication with up to 53.4% inhibition at 50 g/mL. Conclusionthese results explain at least in part the traditional use of T. alnifolia

pharmacology and toxicology↗

bioRxiv2022

Potent, Gut-Restricted Inhibitors of Divalent Metal Transporter 1 (DMT1): Preclinical Efficacy Against Iron Overload and Safety Evaluation

Divalent metal transporter 1 (DMT1) cotransports ferrous iron and protons and is the primary mechanism for uptake of non-heme iron by enterocytes. Inhibitors are potentially useful as therapeutic agents to treat iron overload disorders such as hereditary hemochromatosis or {beta}-thalassemia intermedia, provided that inhibition can be restricted to the duodenum. We used a calcein quench assay to identify human DMT1 (hDMT1) inhibitors. Dimeric constructs were made to generate more potent compounds with low systemic exposure. Direct block of DMT1 was confirmed by voltage clamp measurements. The lead compound, XEN602, strongly inhibits dietary iron uptake in both rats and pigs, yet has negligible systemic exposure. Efficacy is maintained for >2 weeks in a rat subchronic dosing assay. Doses that lowered iron content in the spleen and liver by >50% had no effect on the tissue content of other divalent cations except for cobalt. XEN602 represents a powerful pharmacological tool for understanding the physiological function of DMT1. SIGNIFICANCE STATEMENTThis report introduces methodology to develop potent, gut-restricted inhibitors of DMT1 and identifies XEN602 as a suitable compound for in vivo studies. We also report novel animal models to quantify the inhibition of dietary uptake of iron in both rodents and pigs. This research shows that inhibition of DMT1 is a promising means to treat iron overload disorders.

pharmacology and toxicology↗

bioRxiv2021

The natural stilbenoid (-)-hopeaphenol inhibits cellular entry of SARS-CoV-2 USA-WA1/2020, B.1.1.7 and B.1.351 variants

Antivirals are urgently needed to combat the global SARS-CoV-2/COVID-19 pandemic, supplement existing vaccine efforts, and target emerging SARS-CoV-2 variants of concern. Small molecules that interfere with binding of the viral spike receptor binding domain (RBD) to the host ACE2 receptor may be effective inhibitors of SARS-CoV-2 cell entry. Here we screened 512 pure compounds derived from natural products using a high-throughput RBD/ACE2 binding assay and identified (-)-hopeaphenol, a resveratrol tetramer, in addition to vatalbinoside A and vaticanol B, as potent and selective inhibitors of RBD/ACE2 binding and viral entry. For example, (-)-hopeaphenol disrupted RBD/ACE2 binding with a 50% inhibitory concentration (IC50) of 0.11 M in contrast to an IC50 of 28.3 M against the unrelated host ligand/receptor binding pair PD-1/PD-L1 (selectivity index = 257.3). When assessed against the USA-WA1/2020 variant, (-)-hopeaphenol also inhibited entry of a VSV{Delta}G-GFP reporter pseudovirus expressing SARS-CoV-2 spike into ACE2-expressing Vero-E6 cells and in vitro replication of infectious virus in cytopathic effect assays (IC50 = 10.2 M) without cytotoxicity. Notably, (-)- hopeaphenol also inhibited two emerging variants of concern originating from the United Kingdom (B.1.1.7) and South Africa (B.1.351) in both cytopathic effect and spike-containing pseudovirus assays with similar (B.1.1.7) or improved (B.1.351) efficacies over the USA- WA1/2020 variant. These results identify (-)-hopeaphenol and related stilbenoid analogues as potent and selective inhibitors of viral entry across multiple SARS-CoV-2 variants including those with increased infectivity and/or reduced susceptibility to existing vaccines. ImportanceSARS-CoV-2 antivirals are needed to supplement existing vaccine efforts and target emerging viral variants with increased infectivity or reduced susceptibility to existing vaccines. Here we show that (-)-hopeaphenol, a naturally-occurring stilbenoid compound, in addition to its analogues vatalbinoside A and vaticanol B, inhibit SARS-CoV-2 by blocking the interaction of the viral spike protein with the cellular ACE2 entry receptor. Importantly, in addition to inhibiting the early USA-WA1/2020 SARS-CoV-2 variant, hopeaphenol also inhibits emerging variants of concern including B.1.1.7 ("United Kingdom variant") and B.1.351 ("South Africa variant"), with improved efficacy against B.1.351. (-)-Hopeaphenol therefore represents a new antiviral lead against infection from multiple SARS-CoV-2 variants.

microbiology↗