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In response to stress stimuli, eukaryotic cells typically suppress protein synthesis. This leads to the release of mRNAs from polysomes, their condensation with RNA-binding proteins, and the formation of non-membrane-bound cytoplasmic compartments called stress granules (SGs). SGs contain 40S but generally lack 60S ribosomal subunits. It is known that cycloheximide, emetine, and anisomycin, the ribosome inhibitors that block the progression of 80S ribosomes along mRNA and stabilize polysomes, prevent SG assembly. Conversely, puromycin, which induces premature termination, releases mRNA from polysomes and stimulates the formation of SGs. The same effect is caused by some translation initiation inhibitors, which lead to polysome disassembly and the accumulation of mRNAs in the form of stalled 48S preinitiation complexes. Based on these and other data, it is believed that the trigger for SG formation is the presence of mRNA with extended ribosome-free segments, which tend to form condensates in the cell. In this study, we evaluated the ability of various small-molecule translation inhibitors to block or stimulate the assembly of SGs under conditions of severe oxidative stress induced by sodium arsenite. Contrary to expectations, we found that ribosome-targeting elongation inhibitors of a specific type, which arrest solitary 80S ribosomes at the beginning of the mRNA coding regions but do not interfere with all subsequent ribosomes in completing translation and leaving the transcripts (such as harringtonine, lactimidomycin, or T-2 toxin), completely prevent the formation of arsenite-induced SGs. These observations suggest that the presence of even a single 80S ribosome on mRNA is sufficient to prevent its recruitment into SGs, and the presence of extended ribosome-free regions of mRNA is not sufficient for SG formation. We propose that mRNA entry into SGs may be mediated by specific contacts between RNA-binding proteins and those regions on 40S subunits that remain inaccessible when ribosomes are associated. [ABSTRACT FROM AUTHOR]

Eukaryotic ribosome and cap-dependent translation are attractive targets in the antitumor, antiviral, anti-inflammatory, and antiparasitic therapies. Currently, a broad array of small-molecule drugs is known that specifically inhibit protein synthesis in eukaryotic cells. Many of them are well-studied ribosome-targeting antibiotics that block translocation, the peptidyl transferase center or the polypeptide exit tunnel, modulate the binding of translation machinery components to the ribosome, and induce miscoding, premature termination or stop codon readthrough. Such inhibitors are widely used as anticancer, anthelmintic and antifungal agents in medicine, as well as fungicides in agriculture. Chemicals that affect the accuracy of stop codon recognition are promising drugs for the nonsense suppression therapy of hereditary diseases and restoration of tumor suppressor function in cancer cells. Other compounds inhibit aminoacyl-tRNA synthetases, translation factors, and components of translation-associated signaling pathways, including mTOR kinase. Some of them have antidepressant, immunosuppressive and geroprotective properties. Translation inhibitors are also used in research for gene expression analysis by ribosome profiling, as well as in cell culture techniques. In this article, we review well-studied and less known inhibitors of eukaryotic protein synthesis (with the exception of mitochondrial and plastid translation) classified by their targets and briefly describe the action mechanisms of these compounds. We also present a continuously updated database (http://eupsic.belozersky.msu.ru/) that currently contains information on 370 inhibitors of eukaryotic protein synthesis. [ABSTRACT FROM AUTHOR]

Harringtonine (HT) is an anticancer alkaloid early extracted and isolated from cephalotaxus fortunei Hook. f., also has various pharmacological activities such as antiviral, antibacterial, antimalarial, anti-inflammatory, antioxidant, herbicidal and insecticidal. However, the factors affecting the stability of HT, the main degradation sites and mechanisms involved in its disposal process in vivo have not yet been elucidated. This study utilized HPLC-fluorescence detection method to establish a simple quantitative detection method for HT with good accuracy, precision, and high sensitivity. Temperature and pH were the main factors affecting the stability of HT, which underwent significant degradation in high temperature and alkaline environments because of the occurrence of hydrolysis reactions. In isolated biological homogenates of SD rats, except gastrointestinal tract, HT was degraded in other sites, especially respiratory, mainly in airway and lungs, and systemic metabolism, mainly in livers, spleens, and kidneys. Through UPLC-Q-TOF-MS, three forced degradation products were identified as 4′-demethyl HT, cephalotaxine, and dehydrated HT, respectively. However, the degradation product in isolated biological homogenates of SD rats was only 4′-demethyl HT due to the relatively mild environment. Our findings contributed to a necessary study basis for HT in terms of structural optimization, dosage form selection, storage and transportation. • HPLC-fluorescence method can quantify HT with high selectivity and sensitivity. • HT is unstable in high temperature and alkaline environment in PBS. • HT is degraded into 4′-demethyl HT in isolated biological homogenates of SD rats. • Forced degradation products of HT are 4′-demethyl HT, cephalotaxine, dehydrated HT. [ABSTRACT FROM AUTHOR]

[Display omitted] Fusion with host cell membrane is the main mechanism of infection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Here, we propose that a new strategy to screen small-molecule antagonists blocking SARS-CoV-2 membrane fusion. Using cell membrane chromatography (CMC), we found that harringtonine (HT) simultaneously targeted SARS-CoV-2 S protein and host cell surface TMPRSS2 expressed by the host cell, and subsequently confirmed that HT can inhibit membrane fusion. HT effectively blocked SARS-CoV-2 original strain entry with the IC 50 of 0.217 μM, while the IC 50 in delta variant decreased to 0.101 μM, the IC 50 in Omicron BA.1 variant was 0.042 μM. Due to high transmissibility and immune escape, Omicron subvariant BA.5 has become the dominant strain of the SARS-CoV-2 virus and led to escalating COVID-19 cases, however, against BA.5, HT showed a surprising effectiveness. The IC 50 in Omicron BA.5 was even lower than 0.0019 μM. The above results revealed the effect of HT on Omicron is very significant. In summary, we characterize HT as a small-molecule antagonist by direct targeting on the Spike protein and TMPRSS2. [ABSTRACT FROM AUTHOR]

Harringtonine (HT) is a natural compound, which is mainly produced by the genus Cephalotaxus , and has been clinically utilized in China for the treatment of acute leukemia and lymphoma. However, the amounts of HT in the Cephalotaxus species are very small; therefore, plant tissue cultures have been focused upon to enhance HT production. Qualitative/quantitative methods for HT detection are required to screen superior cell lines. We developed a one-step indirect competitive immunochromatographic assay (ICA) using colloidal gold nanoparticles conjugated with highly specific monoclonal antibodies against HT (MAb 1D2) for simple, rapid, and sensitive detection of HT in plant samples. This ICA can be completed in 15 min after dipping the strip into analytes with a limit of detection of ∼313 ng/mL. In developed ICA, fiber pad which is usually used for conventional ICA, was not used to shorten the time for preparing chromatographic strip, resulting in a decrease in the volume of valuable analytes (20 μL). Considering simplicity, rapidity, and sensitivity of the developed ICA, this study could be applied to a fieldwork study for finding new natural resources containing HT. [ABSTRACT FROM AUTHOR]

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has recently posed a serious threat to global public health. Harringtonine (HT), as a small-molecule antagonist, has antiviral activity against a variety of viruses. There is evidence that HT can inhibit the SARS-CoV-2 entry into host cells by blocking the Spike protein and transmembrane protease serine 2 (TMPRSS2). However, the molecular mechanism underlying the inhibition effect of HT is largely elusive. Here, docking and all-atom molecular dynamics simulations were used to investigate the mechanism of HT against the receptor binding domain (RBD) of Spike, TMPRSS2, as well as the complex of RBD and angiotensin-converting enzyme 2 complex (RBD-ACE2). The results reveal that HT binds to all proteins primarily through hydrogen bond and hydrophobic interactions. Binding with HT influences the structural stability and dynamic motility processes of each protein. The interactions of HT with residues N33, H34 and K353 of ACE2, and residue K417 and Y453 of RBD contribute to disrupting the binding affinity between RBD and ACE2, which may hinder the virus entry into host cells. Our research provides molecular insights into the inhibition mechanism of HT against SARS-CoV-2 associated proteins, which will help for the novel antiviral drugs development. [ABSTRACT FROM AUTHOR]

A versatile intermediate γ-butyrolactone 7 was prepared from L-alanine with 96% ee value, which could be used as a common intermediate for the preparation of the side-chain acid of homoharringtonine and harringtonine in 26.6% and 27% overall yield respectively. The key steps involved [2, 3]-Meisenheimer rearrangement, allylic bromination, and Barbier reaction. Some interesting experimental phenomena and effective solutions to these issues are reported herein. [ABSTRACT FROM AUTHOR]

Babesia gibsoni is a tick-transmitted intraerythrocytic apicomplexan parasite that causes babesiosis in dogs. Due to the strong side effects and lack of efficacy of current drugs, novel drugs against B. gibsoni are urgently needed. Natural products as a source for new drugs is a good choice for screening drugs against B. gibsoni. The current study focuses on identifying novel potential drugs from natural products against B. gibsoni in vitro. Parasite inhibition was verified using a SYBR green I-based fluorescence assay. A total of 502 natural product compounds were screened for anti- B. gibsoni activity in vitro. Twenty-four compounds showed high growth inhibition (>80%) on B. gibsoni and 5 plant-derived compounds were selected for further study. The half-maximal inhibitory concentration (IC 50) values of lycorine (LY), vincristine sulfate (VS), emetine·2HCl (EME), harringtonine (HT) and cephaeline·HBr (CEP) were 784.4 ± 3.3, 643.0 ± 2.8, 253.1 ± 1.4, 23.4 ± 1.2, and 108.1 ± 4.3 nM, respectively. The Madin-Darby canine kidney (MDCK) cell line was used to assess cytotoxicity of hit compounds. All compounds showed minimal toxicity to the MDCK cells. The effects of hit compounds combined with diminazene aceturate (DA) on B. gibsoni were further evaluated in vitro. VS, EME, HT or CEP combined with DA showed synergistic effects against B. gibsoni , whereas LY combined with DA showed an antagonistic effect against B. gibsoni. The results obtained in this study indicate that LY, VS, EME, HT and CEP are promising compounds for B. gibsoni treatment. [Display omitted] • Natural product compounds were screened for anti- Babesia gibsoni activity in vitro. • Twenty-four compounds showed significant inhibition on B. gibsoni growth. • Five plant-derived compounds were further evaluated for their IC 50 , CC 50 and combination indices. [ABSTRACT FROM AUTHOR]

Mosquito-borne Zika virus (ZIKV) is a Flavivirus that came under intense study from 2014 to 2016 for its well-known ability to cause congenital microcephaly in fetuses and neurological Guillain–Barré disease in adults. Substantial research on screening antiviral agents against ZIKV and preventing ZIKV infection are globally underway, but Food and Drug Administration (FDA)-approved treatments are not available yet. Compounds from Chinese medicinal herbs may offer an opportunity for potential therapies for anti-ZIKV infection. In this study, we evaluated the antiviral efficacy of harringtonine against ZIKV. Harringtonine possessed anti-ZIKV properties against the binding, entry, replication, and release stage through the virus life cycle. In addition, harringtonine have strong virucidal effects in ZIKV and exhibited prophylaxis antiviral ability prior ZIKV infection. The antiviral activity also observed in the treatment against Japanese encephalitis reporter virus (RP9-GFP strain). Overall, this study demonstrated that harringtonine would be a favorable potential candidate for the development of anti-ZIKV infection therapies. [ABSTRACT FROM AUTHOR]