Madurastatins with Imidazolidinone Rings: Natural Products or Side-Reaction Products from Extraction Solvents?

Madurastatins are a group of pentapeptides containing an oxazoline moiety, and, in a few cases, an imidazolidinone ring as an additional structural feature. In our search for new potential antiparasitic metabolites from natural sources, we studied the acetone extracts from a culture of Actinomadura sp. CA-135719. The LC/HRMS analysis of this extract identified the presence of the known madurastatins C1 (1), D1 (4), and D2 (5) together with additional members of the family that were identified as the new madurastatins H2 (2) and 33-epi-D1 (3) after isolation and spectroscopic analysis. The planar structures of the new compounds were established by HRMS, ESI-qTOF-MS/MS, and 1D and 2D NMR data, and their absolute configuration was proposed using Marfey's and bioinformatic analyses of the biosynthetic gene cluster (BGC). A revision of the absolute configuration of madurastatins D1 and D2 is proposed. Additionally, madurastatins containing imidazolidinone rings are proved to be artifacts originating during acetone extraction of the bacterial cultures.

Screening of FDA-Approved Drugs Using a MERS-CoV Clinical Isolate from South Korea Identifies Potential Therapeutic Options for COVID-19.

Therapeutic options for coronaviruses remain limited. To address this unmet medical need, we screened 5406 compounds, including United States Food and Drug Administration (FDA)-approved drugs and bioactives, for activity against a South Korean Middle East respiratory syndrome coronavirus (MERS-CoV) clinical isolate. Among 221 identified hits, 54 had therapeutic indexes (TI) greater than 6, representing effective drugs. The time-of-addition studies with selected drugs demonstrated eight and four FDA-approved drugs which acted on the early and late stages of the viral life cycle, respectively. Confirmed hits included several cardiotonic agents (TI > 100), atovaquone, an anti-malarial (TI > 34), and ciclesonide, an inhalable corticosteroid (TI > 6). Furthermore, utilizing the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), we tested combinations of remdesivir with selected drugs in Vero-E6 and Calu-3 cells, in lung organoids, and identified ciclesonide, nelfinavir, and camostat to be at least additive in vitro. Our results identify potential therapeutic options for MERS-CoV infections, and provide a basis to treat coronavirus disease 2019 (COVID-19) and other coronavirus-related illnesses.

Noncanonical immune response to the inhibition of DNA methylation by Staufen1 via stabilization of endogenous retrovirus RNAs.

DNA-methyltransferase inhibitors (DNMTis), such as azacitidine and decitabine, are used clinically to treat myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). Decitabine activates the transcription of endogenous retroviruses (ERVs), which can induce immune response by acting as cellular double-stranded RNAs (dsRNAs). Yet, the posttranscriptional regulation of ERV dsRNAs remains uninvestigated. Here, we find that the viral mimicry and subsequent cell death in response to decitabine require the dsRNA-binding protein Staufen1 (Stau1). We show that Stau1 directly binds to ERV RNAs and stabilizes them in a genome-wide manner. Furthermore, Stau1-mediated stabilization requires a long noncoding RNA TINCR, which enhances the interaction between Stau1 and ERV RNAs. Analysis of a clinical patient cohort reveals that MDS and AML patients with lower Stau1 and TINCR expressions exhibit inferior treatment outcomes to DNMTi therapy. Overall, our study reveals the posttranscriptional regulatory mechanism of ERVs and identifies the Stau1-TINCR complex as a potential target for predicting the efficacy of DNMTis and other drugs that rely on dsRNAs.

Identification of Antiviral Drug Candidates against SARS-CoV-2 from FDA-Approved Drugs.

Drug repositioning is the only feasible option to immediately address the COVID-19 global challenge. We screened a panel of 48 FDA-approved drugs against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) which were preselected by an assay of SARS-CoV. We identified 24 potential antiviral drug candidates against SARS-CoV-2 infection. Some drug candidates showed very low 50% inhibitory concentrations (IC50s), and in particular, two FDA-approved drugs-niclosamide and ciclesonide-were notable in some respects.

In Vitro and in Vivo Activity of mTOR Kinase and PI3K Inhibitors Against Leishmania donovani and Trypanosoma brucei.

Kinetoplastid parasites, including Leishmania and Trypanosoma spp., are life threatening pathogens with a worldwide distribution. Next-generation therapeutics for treatment are needed as current treatments have limitations, such as toxicity and drug resistance. In this study, we examined the activities of established mammalian target of rapamycin (mTOR)/phosphoinositide 3-kinase (PI3K) inhibitors against these tropical diseases. High-throughput screening of a library of 1742 bioactive compounds against intracellular L. donovani was performed, and seven mTOR/PI3K inhibitors were identified. Dose-dilution assays revealed that these inhibitors had half maximal effective concentration (EC50) values ranging from 0.14 to 13.44 µM for L. donovani amastigotes and from 0.00005 to 8.16 µM for T. brucei. The results of a visceral leishmaniasis mouse model indicated that treatment with Torin2, dactolisib, or NVP-BGT226 resulted in reductions of 35%, 53%, and 54%, respectively, in the numbers of liver parasites. In an acute T. brucei mouse model using NVP-BGT226 parasite numbers were reduced to under the limits of detection by five consecutive days of treatment. Multiple sequence and structural alignment results indicated high similarities between mTOR and kinetoplastid TORs; the inhibitors are predicted to bind in a similar manner. Taken together, these results indicated that the TOR pathways of parasites have potential for the discovery of novel targets and new potent inhibitors.

In Vitro and In Vivo Investigation of the Inhibition of Trypanosoma brucei Cell Growth by Lipophilic Bisphosphonates.

We report the results of a screen of a library of 925 potential prenyl synthase inhibitors against Trypanosoma brucei farnesyl diphosphate synthase (TbFPPS) and against T. brucei, the causative agent of human African trypanosomiasis. The most potent compounds were lipophilic analogs of the bone resorption drug zoledronate, some of which had submicromolar to low micromolar activity against bloodstream form T. brucei and selectivity indices of up to ∼ 300. We evaluated the effects of two such inhibitors on survival and parasitemia in a T. brucei mouse model of infection and found that survival increased by up to 16 days. We also investigated the binding of three lipophilic bisphosphonates to an expressed TbFPPS using crystallography and investigated the thermodynamics of binding using isothermal titration calorimetry.

In Vitro and in Vivo Activity of Multitarget Inhibitors against Trypanosoma brucei.

We tested a series of amidine and related compounds against Trypanosoma brucei. The most active compound was a biphenyldiamidine that had an EC 50 of 7.7 nM against bloodstream-form parasites. There was little toxicity against two human cell lines with CC 50 > 100 µM. There was also good in vivo activity in a mouse model of infection with 100% survival at 3 mg/kg i.p. The most potent lead blocked replication of kinetoplast DNA (k-DNA), but not nuclear DNA, in the parasite. Some compounds also inhibited the enzyme farnesyl diphosphate synthase (FPPS), and some were uncouplers of oxidative phosphorylation. We developed a computational model for T. brucei cell growth inhibition (R (2) = 0.76) using DNA ΔT m values for inhibitor binding combined with T. brucei FPPS IC 50 values. Overall, the results suggest that it may be possible to develop multitarget drug leads against T. brucei that act by inhibiting both k-DNA replication and isoprenoid biosynthesis.

A novel highly potent and selective 11ß-hydroxysteroid dehydrogenase type 1 inhibitor, UI-1499.

AIMS: Elevated levels of glucocorticoid hormones cause glucose intolerance, visceral obesity, insulin resistance, hypertension, and dyslipidemia. 11ß-Hydroxysteroid dehydrogenase type 1 (11ß-HSD1) represents as an attractive therapeutic target for treatment of metabolic syndrome and type 2 diabetes. This study investigated whether 11ß-HSD1 inhibition by a novel selective inhibitor, (1S,3R,4S,5S,7S)-4-(1-((2-fluoro-N-methylphenylsulfonamido)-methyl) cyclopropanecarboxamido) adamantane-1-carboxamide (UI-1499) ameliorated metabolic abnormalities in diabetic mice. MAIN METHODS: The in vitro activity of 11ß-HSD1 was measured using the homogeneous time resolved fluorescence (HTRF) assay. Differentiated adipocytes were used to evaluate cellular 11ß-HSD1 activity. To determine the inhibitory effects on 11ß-HSD1 in tissues, we performed ex vivo studies using liver and epididymal fat isolated from C57BL/6J mice. KKAy mice were administered with UI-1499 to evaluate whether this compound ameliorated metabolic abnormalities in vivo in diabetic animals. KEY FINDINGS: UI-1499 had highly potent inhibitory activity in mouse, monkey and human 11ß-HSD1, derived from liver microsomes. Oral administration of 45 mg/kg UI-1499 significantly inhibited 11ß-HSD1 activity in C57BL/6J mouse liver and epididymal fat tissues. In KKAy mice, oral administration of UI-1499 (10 and 30 mg/kg for 3 weeks) lowered fasting blood glucose and HbA1c levels; these effects were comparable to those of pioglitazone. Further, UI-1499 treatment lowered plasma low-density lipoprotein (LDL) level in KKAy mice. SIGNIFICANCE: These results suggest that the 11ß-HSD1 inhibitor, UI-1499, may serve as a novel drug candidate for the treatment of type 2 diabetes and metabolic syndrome.

Secondary metabolites from Vietnamese marine invertebrates with activity against Trypanosoma brucei and T. cruzi.

Marine-derived natural products from invertebrates comprise an extremely diverse and promising source of the compounds from a wide variety of structural classes. This study describes the discovery of five marine natural products with activity against Trypanosoma species by natural product library screening using whole cell in vitro assays. We investigated the anti-trypanosomal activity of the extracts from the soft corals and echinoderms living in Vietnamese seas. Of the samples screened, the methanolic extracts of several marine organisms exhibited potent activities against cultures of Trypanosoma brucei and T. cruzi (EC50 < 5.0 µg/mL). Among the compounds isolated from these extracts, laevigatol B (1) from Lobophytum crassum and L. laevigatum, (24S)-ergost-4-ene-3-one (2) from Sinularia dissecta, astropectenol A (3) from Astropecten polyacanthus, and cholest-8-ene-3ß,5α,6ß,7α-tetraol (4) from Diadema savignyi showed inhibitory activity against T. brucei with EC50 values ranging from 1.57 ± 0.14 to 14.6 ± 1.36 µM, relative to the positive control, pentamidine (EC50 = 0.015 ± 0.003 µM). Laevigatol B (1) and 5α-cholest-8(14)-ene-3ß,7α-diol (5) exhibited also significant inhibitory effects on T. cruzi. The cytotoxic activity of the pure compounds on mammalian cells was also assessed and found to be insignificant in all cases. This is the first report on the inhibitory effects of marine organisms collected in Vietnamese seas against Trypanosoma species responsible for neglected tropical diseases.