Development of Ultrapure and Potent Tannic Acids as a Pan-coronal Antiviral Therapeutic.

The rampageous transmission of SARS-CoV-2 has been devastatingly impacting human life and public health since late 2019. The waves of pandemic events caused by distinct coronaviruses at present and over the past decades have prompted the need to develop broad-spectrum antiviral drugs against them. In this study, our Pentarlandir ultrapure and potent tannic acids (UPPTA) showed activities against two coronaviral strains, SARS-CoV-2 and HCoV-OC43, the earliest-known coronaviruses. The mode of inhibition of Pentarlandir UPPTA is likely to act on 3-chymotrypsin-like protease (3CLpro) to prevent viral replication, as supported by results of biochemical analysis, a 3CLpro assay, and a "gain-of-function" 3CLpro overexpressed cell-based method. Even in the 3CLpro overexpressed environment, Pentarlandir UPPTA remained its antiviral characteristic. Utilizing cell-based virucidal and cytotoxicity assays, the 50% effective concentrations (EC50) and 50% cytotoxicity concentration (CC50) of Pentarlandir UPPTA were determined to be ∼0.5 and 52.5 µM against SARS-CoV-2, while they were 1.3 and 205.9 µM against HCoV-OC43, respectively. In the pharmacokinetic studies, Pentarlandir UPPTA was distributable at a high level to the lung tissue with no accumulation in the body, although the distribution was affected by the food effect. With further investigation in toxicology, Pentarlandir UPPTA demonstrated an overall safe toxicology profile. Taking these findings together, Pentarlandir UPPTA is considered to be a safe and efficacious pancoronal antiviral drug candidate that has been advanced to clinical development.

Equilibrative nucleoside transporter 1 inhibition rescues energy dysfunction and pathology in a model of tauopathy.

Tau pathology is instrumental in the gradual loss of neuronal functions and cognitive decline in tauopathies, including Alzheimer's disease (AD). Earlier reports showed that adenosine metabolism is abnormal in the brain of AD patients while consequences remained ill-defined. Herein, we aimed at investigating whether manipulation of adenosine tone would impact Tau pathology, associated molecular alterations and subsequent neurodegeneration. We demonstrated that treatment with an inhibitor (J4) of equilibrative nucleoside transporter 1 (ENT1) exerted beneficial effects in a mouse model of Tauopathy. Treatment with J4 not only reduced Tau hyperphosphorylation but also rescued memory deficits, mitochondrial dysfunction, synaptic loss, and abnormal expression of immune-related gene signatures. These beneficial effects were particularly ascribed to the ability of J4 to suppress the overactivation of AMPK (an energy reduction sensor), suggesting that normalization of energy dysfunction mitigates neuronal dysfunctions in Tauopathy. Collectively, these data highlight that targeting adenosine metabolism is a novel strategy for tauopathies.

Lysophosphatidic acid receptor LPA3 prevents oxidative stress and cellular senescence in Hutchinson-Gilford progeria syndrome.

Hutchinson-Gilford progeria syndrome (HGPS) is a rare laminopathy that produces a mutant form of prelamin A, known as Progerin, resulting in premature aging. HGPS cells show morphological abnormalities of the nuclear membrane, reduced cell proliferation rates, accumulation of reactive oxygen species (ROS), and expression of senescence markers. Lysophosphatidic acid (LPA) is a growth factor-like lipid mediator that regulates various physiological functions via activating multiple LPA G protein-coupled receptors. Here, we study the roles of LPA and LPA receptors in premature aging. We report that the protein level of LPA3 was highly downregulated through internalization and the lysosomal degradation pathway in Progerin-transfected HEK293 cells. By treating Progerin HEK293 cells with an LPA3 agonist (OMPT, 1-Oleoyl-2-O-methyl-rac-glycerophosphothionate) and performing shRNA knockdown of the Lpa3r transcript in these cells, we showed that LPA3 activation increased expression levels of antioxidant enzymes, consequently inhibiting ROS accumulation and ameliorating cell senescence. LPA3 was shown to be downregulated in HGPS patient fibroblasts through the lysosomal pathway, and it was shown to be crucial for ameliorating ROS accumulation and cell senescence in fibroblasts. Moreover, in a zebrafish model, LPA3 deficiency was sufficient to cause premature aging phenotypes in multiple organs, as well as a shorter lifespan. Taken together, these findings identify the decline of LPA3 as a key contributor to the premature aging phenotypes of HGPS cells and zebrafish.

Novel Endogenous Ligands of Aryl Hydrocarbon Receptor Mediate Neural Development and Differentiation of Neuroblastoma.

Aryl hydrocarbon receptor (AHR) signaling has been suggested to play roles in various physiological functions independent of its xenobiotic activity, including cell cycle regulation, immune response, and embryonic development. Several endogenous ligands were also identified by high-throughput screening techniques. However, the mechanism by which these molecules mediate AHR signaling in certain functions is still elusive. In this study, we investigated the possible pathway through which AHR and its endogenous ligands regulate neural development. We first identified two neuroactive steroids, 3α,5α-tetrahydrocorticosterone and 3α,5ß-tetrahydrocorticosterone (5α- and 5ß-THB), as novel AHR endogenous ligands through the use of an ultrasensitive dioxin-like compound bioassay and liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS). We then treated zebrafish embryos with 5α- and 5ß-THB, which enhance the expression of neurogenesis marker HuC. Furthermore, 5α- and 5ß-THB both enhanced the expression of myelinating glial cell markers, sex determining region Y-box 10 (Sox10), and myelin-associated proteins myelin basic protein (Mbp) and improved the mobility of zebrafish larvae via the Ahr2 pathway. These results indicated that AHR mediates zebrafish neurogenesis and gliogenesis, especially the differentiation of oligodendrocyte or Schwann cells. Additionally, we showed that these molecules may induce neuroblastoma (NB) cell differentiation suggesting therapeutic potential of 5α- and 5ß-THB in NB treatment. In summary, our results reveal that 5α- and 5ß-THB are endogenous ligands of AHR and have therapeutic potential for NB treatment. By the interaction with THB, AHR signaling regulates various aspects of neural development.

A Positive Feedback Loop between Glial Cells Missing 1 and Human Chorionic Gonadotropin (hCG) Regulates Placental hCGß Expression and Cell Differentiation.

Human chorionic gonadotropin (hCG) is composed of a common α subunit and a placenta-specific ß subunit. Importantly, hCG is highly expressed in the differentiated and multinucleated syncytiotrophoblast, which is formed via trophoblast cell fusion and stimulated by cyclic AMP (cAMP). Although the ubiquitous activating protein 2 (AP2) transcription factors TFAP2A and TFAP2C may regulate hCGß expression, it remains unclear how cAMP stimulates placenta-specific hCGß gene expression and trophoblastic differentiation. Here we demonstrated that the placental transcription factor glial cells missing 1 (GCM1) binds to a highly conserved promoter region in all six hCGß paralogues by chromatin immunoprecipitation-on-chip (ChIP-chip) analyses. We further showed that cAMP stimulates GCM1 and the CBP coactivator to activate the hCGß promoter through a GCM1-binding site (GBS1), which also constitutes a previously identified AP2 site. Given that TFAP2C may compete with GCM1 for GBS1, cAMP enhances the association between the hCGß promoter and GCM1 but not TFAP2C. Indeed, the hCG-cAMP-protein kinase A (PKA) signaling pathway also stimulates Ser269 and Ser275 phosphorylation of GCM1, which recruits CBP to mediate GCM1 acetylation and stabilization. Consequently, hCG stimulates the expression of GCM1 target genes, including the fusogenic protein syncytin-1, to promote placental cell fusion. Our study reveals a positive feedback loop between GCM1 and hCG regulating placental hCGß expression and cell differentiation.

Functional antagonism between high temperature requirement protein A (HtrA) family members regulates trophoblast invasion.

Human trophoblast invasion of decidualized endometrium is essential for placentation and is tightly regulated and involves trophoblast-decidual cell interaction. High temperature requirement A4 (HtrA4) is a secreted serine protease highly expressed in the invasive extravillous trophoblasts that invade decidua. In contrast, both HtrA1 and HtrA3 have been shown to inhibit trophoblast invasion. Here we provide evidence that decidua-secreted HtrA1 and HtrA3 antagonize HtrA4-mediated trophoblast invasion. We demonstrated that HtrA1 and HtrA3 interact with and degrade HtrA4 and thereby inhibit trophoblast-like JAR cell invasion. Specifically, HtrA1 and HtrA3 expression is up-regulated under decidualization conditions in endometrial stromal and epithelial cells, T-HESCs and Ishikawa cells, respectively. Conditioned media from these two cell lines after decidualization treatment suppress HtrA4-expressing JAR cell invasion in an HtrA1- or HtrA3-dependent manner. Co-culture of the HtrA4-expressing JAR cells with decidualization stimuli-treated T-HESC or Ishikawa monolayer also impairs JAR cell invasion, which can be reversed by HtrA1 or HtrA3 knockdown, supporting that HtrA1 and HtrA3 are crucial for trophoblast-decidual cell interaction in the control of trophoblast invasion. Our study reveals a novel regulatory mechanism of trophoblast invasion through physical and functional interaction between HtrA family members.