Mechanistic Insights from the Crystal Structure and Computational Analysis of the Radical SAM Deaminase DesII.

Radical S-adenosyl-L-methionine (SAM) enzymes couple the reductive cleavage of SAM to radical-mediated transformations that have proven to be quite broad in scope. DesII is one such enzyme from the biosynthetic pathway of TDP-desosamine where it catalyzes a radical-mediated deamination. Previous studies have suggested that this reaction proceeds via direct elimination of ammonia from an α-hydroxyalkyl radical or its conjugate base (i.e., a ketyl radical) rather than 1,2-migration of the amino group to form a carbinolamine radical intermediate. However, without a crystal structure, the active site features responsible for this chemistry have remained largely unknown. The crystallographic studies described herein help to fill this gap by providing a structural description of the DesII active site. Computational analyses based on the solved crystal structure are consistent with direct elimination and indicate that an active site glutamate residue likely serves as a general base to promote deprotonation of the α-hydroxyalkyl radical intermediate and elimination of the ammonia group.

Multimechanism biological profiling of tetrahydro-ß-carboline analogues as selective HDAC6 inhibitors for the treatment of Alzheimer's disease.

With the intensive research on the pathogenesis of Alzheimer's disease (AD), inhibition of HDAC6 appears to be a potential therapeutic approach for AD. In this paper, a series of tetrahydro-ß-carboline derivatives with hydroxamic acid group were fast synthesized. Among all, the most potent 15 selectively inhibited HDAC6 with IC50 of 15.2 nM and markedly increased acetylated alpha-tubulin levels. In cellular assay, 15 showed excellent neurotrophic effect by increasing the expression of GAP43 and Beta-3 tubulin markers. Besides, 15 showed neuroprotective effects in PC12 or SH-SY5Y cells against H2O2 and 6-OHDA injury through activation of Nrf2, catalase and Prx II, and significantly reduced H2O2-induced reactive oxygen species (ROS) production. In vivo, 15 significantly attenuated zebrafish anxiety-like behaviour and memory deficits in a SCOP-induced zebrafish model of AD. To sum up, multifunctional 15 might be a good lead to develop novel tetrahydrocarboline-based agents for the treatment of AD.

Stepwise Ultra-pH-Sensitive Micelles Overcome a pKa Barrier for Systemic Lymph Node Delivery.

While local nanoparticle delivery to lymph nodes is well studied, there are few design criteria for intravenous delivery to the entire lymph node repertoire. In this study, we investigated the effect of NP pH transition on lymph node targeting by employing a series of ultra-pH-sensitive (UPS) polymeric micelles. The UPS library responds to pH thresholds (pKa 6.9, 6.2, and 5.3) over a range of physiological pH. We observed a dependence of intravenous lymph node targeting on micelle pH transition. UPS6.9 (subscript indicates pKa) shows poor lymph node delivery, while UPS5.3 delivers efficiently to lymph node sets. We investigated targeting mechanisms of UPS5.3, observing an accumulation among lymph node lymphatics and a dependence on lymph node-resident macrophages. To overcome the pH-threshold barrier, which limits UPS6.9, we rationally designed a nanoparticle coassembly of UPS6.9 with UPS5.3, called HyUPS. The HyUPS micelle retains the constitutive pH transitions of each polymer, showing stepwise responses to discrete pH thresholds. We demonstrate that HyUPS improves UPS6.9 delivery to lymph nodes, extending this platform for disease detection of lymph node metastasis.

Chemical Structures, Biological Activities, and Biosynthetic Analysis of Secondary Metabolites from Agaricus Mushrooms: A Review.

Agaricus mushrooms are an important genus in the Agaricaceae family, belonging to the order Agaricales of the class Basidiomycota. Among them, Agaricus bisporus is a common mushroom for mass consumption, which is not only nutritious but also possesses significant medicinal properties such as anticancer, antibacterial, antioxidant, and immunomodulatory properties. The rare edible mushroom, Agaricus blazei, contains unique agaricol compounds with significant anticancer activity against liver cancer. Agaricus blazei is believed to expel wind and cold, i.e., the pathogenic factors of wind and cold from the body, and is an important formula in traditional Chinese medicine. Despite its nutritional richness and outstanding medicinal value, Agaricus mushrooms have not been systematically compiled and summarized. Therefore, the present review compiles and classifies 70 natural products extracted from Agaricus mushrooms over the past six decades. These compounds exhibit diverse biological and pharmacological activities, with particular emphasis on antitumor and antioxidant properties. While A. blazei and A. bisporus are the primary producers of these compounds, studies on secondary metabolites from other Agaricus species remain limited. Further research is needed to explore and understand the anticancer and nutritional properties of Agaricus mushrooms. This review contributes to the understanding of the structure, bioactivity, and biosynthetic pathways of Agaricus compounds and provides insights for the development of functional foods using these mushrooms.

Four New Polyprenylated Acylphloroglucinols from Hypericum perforatum L.

Hyperforatums A-D (1-4), four new polyprenylated acylphloroglucinols, together with 13 known compounds were isolated and identified from the aerial parts of Hypericum perforatum L. (St. John's wort). Their structures were confirmed with a comprehensive analysis comprising spectroscopic methods, including 1D and 2D NMR, HRESIMS, and electronic circular dichroism (ECD) calculations. Hyperforatum A featured an unusual chromene-1,4-dione bicyclic system, and hyperforatums B and C were two rare monocyclic PPAPs with five-membered furanone cores. Compound 1 exhibited a moderate inhibition effect on NO production in BV-2 microglial cells stimulated by LPS.

Severely polarized extracellular acidity around tumour cells.

Extracellular pH impacts many molecular, cellular and physiological processes, and hence is tightly regulated. Yet, in tumours, dysregulated cancer cell metabolism and poor vascular perfusion cause the tumour microenvironment to become acidic. Here by leveraging fluorescent pH nanoprobes with a transistor-like activation profile at a pH of 5.3, we show that, in cancer cells, hydronium ions are excreted into a small extracellular region. Such severely polarized acidity (pH <5.3) is primarily caused by the directional co-export of protons and lactate, as we show for a diverse panel of cancer cell types via the genetic knockout or inhibition of monocarboxylate transporters, and also via nanoprobe activation in multiple tumour models in mice. We also observed that such spot acidification in ex vivo stained snap-frozen human squamous cell carcinoma tissue correlated with the expression of monocarboxylate transporters and with the exclusion of cytotoxic T cells. Severely spatially polarized tumour acidity could be leveraged for cancer diagnosis and therapy.

Inubritantrimers A-D: Trimerized Sesquiterpenoid [4 + 2] Adducts Featuring a Distinctive Spiro-Polycyclic Scaffold from Inula britannica.

Inubritantrimer A (1), a trace trimerized sesquiterpenoid [4 + 2] adduct featuring an unusual exo-exo type spiro-polycyclic scaffold, together with three new endo-exo [4 + 2] adducts, inubritantrimers B-D (2-4), were discovered from the flowers of Inula britannica. Their structures were elucidated using 1D/2D NMR, X-ray diffraction, and ECD approaches. 1 is characterized as a novel exo-exo trimer, synthesized biogenetically from three sesquiterpenoid monomers, featuring a unique linkage of C-11/C-1', C-13/C-3' and C-13'/C-3″, C-11'/C-1″ through a two-step exo [4 + 2] cycloaddition process. Compounds 1-4 exhibited modest cytotoxicity against breast cancer cells with IC50 values in the range of 5.84-12.01 µM.

STING licensing of type I dendritic cells potentiates antitumor immunity.

Stimulator of interferon genes (STING) is an immune adaptor protein that senses cyclic GMP-AMP in response to self or microbial cytosolic DNA as a danger signal. STING is ubiquitously expressed in diverse cell populations, including cancer cells, with distinct cellular functions, such as activation of type I interferons, autophagy induction, or triggering apoptosis. It is not well understood whether and which subsets of immune cells, stromal cells, or cancer cells are particularly important for STING-mediated antitumor immunity. Here, using a polymeric STING-activating nanoparticle (PolySTING) with a shock-and-lock dual activation mechanism, we show that conventional type 1 dendritic cells (cDC1s) are essential for STING-mediated rejection of multiple established and metastatic murine tumors. STING status in the host but not in the cancer cells (Tmem173-/-) is important for antitumor efficacy. Specific depletion of cDC1 (Batf3-/-) or STING deficiency in cDC1 (XCR1creSTINGfl/fl) abolished PolySTING efficacy, whereas depletion of other myeloid cells had little effect. Adoptive transfer of wild-type cDC1 in Batf3-/- mice restored antitumor efficacy, whereas transfer of cDC1 with STING or IRF3 deficiency failed to rescue. PolySTING induced a specific chemokine signature in wild-type but not Batf3-/- mice. Multiplexed immunohistochemistry analysis of STING-activating cDC1s in resected tumors correlates with patient survival. Furthermore, STING-cDC1 signature was increased after neoadjuvant pembrolizumab therapy in patients with non-small cell lung cancer. Therefore, we have defined that a subset of myeloid cells is essential for STING-mediated antitumor immunity with associated biomarkers for prognosis.

Discovery from Hypericum elatoides and synthesis of hyperelanitriles as α-aminopropionitrile-containing polycyclic polyprenylated acylphloroglucinols.

The search for lead compounds with anti-neuroinflammatory activity from structurally 'optimized' natural products is a crucial and promising strategy in the quest to discover safe and efficacious agents for treating neurodegenerative diseases. A phytochemical investigation on the aerial portions of Hypericum elatoides led to the isolation of five nitrogenous polycyclic polyprenylated acylphloroglucinols (PPAPs), hyperelanitriles A-D (1-4) and hyperelamine A (5). Their structures were determined by spectroscopic analysis, ECD and NMR calculations, and X-ray crystallography. To the best of our knowledge, compounds 1-4 represent the first examples of acylphloroglucinols featuring an α-aminonitrile moiety, while 5 is a rare enamine-containing PPAP. Further, the synthesis of these naturally occurring PPAP-based nitriles or amines was accomplished. Compound 5 exhibited inhibitory activity against LPS-activated NO production in BV-2 cells, potentially through the suppression of TLR-4/NF-κB signaling. Here we show the isolation, structural elucidation, synthesis, and bioactive evaluation of compounds 1-5.

STING Licensing of Type I Dendritic Cells Potentiates Antitumor Immunity.

Stimulator of interferon genes (STING) is an immune adaptor protein that senses cyclic GMP-AMP (cGAMP) in response to self or microbial cytosolic DNA as a danger signal. STING is ubiquitously expressed in diverse cell populations including cancer cells with distinct cellular functions such as activation of type I interferons, autophagy induction, or triggering apoptosis. It is not well understood whether and which subsets of immune cells, stromal cells, or cancer cells are particularly important for STING-mediated antitumor immunity. Here using a polymeric STING-activating nanoparticle (PolySTING) with a "shock-and-lock" dual activation mechanism, we show type 1 conventional dendritic cell (cDC1) is essential for STING-mediated rejection of multiple established and metastatic murine tumors. STING status in the host but not in the cancer cells ( Tmem173 -/- ) is important for antitumor efficacy. Specific depletion of cDC1 ( Batf3 -/- ) or STING deficiency in cDC1 ( XCR1 cre STING fl/fl ) abolished PolySTING efficacy, whereas depletion of other myeloid cells had little effect. Adoptive transfer of wildtype cDC1 in Batf3 -/- mice restored antitumor efficacy while transfer of cDC1 with STING or IRF3 deficiency failed to rescue. PolySTING induced a specific chemokine signature in wildtype but not Batf3 -/- mice. Multiplexed immunohistochemistry analysis of STING-activating cDC1s in resected tumors correlates with patient survival while also showing increased expressions after neoadjuvant pembrolizumab therapy in non-small cell lung cancer patients. Therefore, we have defined that a subset of myeloid cells is essential for STING-mediated antitumor immunity with associated biomarkers for prognosis. One Sentence Summary: A "shock-and-lock" nanoparticle agonist induces direct STING signaling in type 1 conventional dendritic cells to drive antitumor immunity with defined biomarkers.

Dimerized sesquiterpenoid [4 + 2] adducts with ferroptosis-promoting activity from Inula britannica Linn.

Inubritanolides C and D (1 and 2), two exo sesquiterpenoid [4 + 2] adducts with unprecedented interconverting conformations of twist-chair and chair, together with two previously undescribed endo [4 + 2] dimers (3 and 4) were discovered from Inula britannica flowers. Dimers 1 and 2 have an undescribed carbon skeleton comprising of eudesmanolide and guaianolide units with the linkage mode of C-11/C-1' and C-13/C-3' via a Diels-Alder cycloaddition reaction. Their structures were elucidated using 1D/2D NMR, X-ray diffraction, ECD, and variable-temperature NMR experiments. Dimer 2 displayed a strong inhibitory effect on breast cancer cells by promoting lipid ROS production, showing its potential as ferroptosis inducer.

The alkynyl-containing compounds from mushrooms and their biological activities.

Mushrooms have been utilized by humans for thousands of years due to their medicinal and nutritional properties. They are a crucial natural source of bioactive secondary metabolites, and recent advancements have led to the isolation of several alkynyl-containing compounds with potential medicinal uses. Despite their relatively low abundance, naturally occurring alkynyl compounds have attracted considerable attention due to their high reactivity. Bioactivity studies have shown that alkynyl compounds exhibit significant biological and pharmacological activities, including antitumor, antibacterial, antifungal, insecticidal, phototoxic, HIV-inhibitory, and immunosuppressive properties. This review systematically compiles 213 alkynyl-containing bioactive compounds isolated from mushrooms since 1947 and summarizes their diverse biological activities, focusing mainly on cytotoxicity and anticancer effects. This review serves as a detailed and comprehensive reference for the chemical structures and bioactivity of alkynyl-containing secondary metabolites from mushrooms. Moreover, it provides theoretical support for the development of chemical constituents containing alkynyl compounds in mushrooms based on academic research and theory.

Torreya grandis oil attenuates cognitive impairment in scopolamine-induced mice.

The oil of Torreya grandis (TGO), a common nut in China, is considered to be a bioactive edible oil and has a great value in functional food development. In this study, the neuroprotective effects of TGO were investigated on a scopolamine (SCOP)-induced C57BL/6J mouse model. The mice were pretreated with TGO for 30 days (1000 mg per kg per day and 3000 mg per kg per day, i.g.). Behavioral tests showed that the supplementation of TGO could prevent the cognitive deficits induced by SCOP. TGO rebalanced the disorder of the cholinergic system by upgrading the level of acetylcholine. TGO also alleviated the over-activation of microglia and inhibited neuroinflammation and oxidative stress. Additionally, TGO could regulate the composition of gut microbiota, increase the production of short-chain fatty acids, and decrease the content of lipopolysaccharides in the serum. In conclusion, TGO has the potential to prevent loss of memory and impairment of cognition, which may be related to its regulation of the gut microbiota-metabolite-brain axis.

Tetrahydro-ß-carboline derivatives as potent histone deacetylase 6 inhibitors with broad-spectrum antiproliferative activity.

A series of tetrahydro-ß-carboline (THßC)-based hydroxamic acids were rationally designed and synthesized as novel selective HDAC6 inhibitors (sHDAC6is) by the application of scaffold hopping strategy. Several THßC analogues were highly potent (IC50 < 5 nM) and selective against HDAC6 enzyme and exhibited good antiproliferative activity against human multiple myeloma (MM) cell. Molecular docking interpreted the structure activity relationship (SAR). Target engagement of HDAC6 was confirmed in RPMI-8226 cells using the WB assay. In vitro, (1S, 3R)-1-(4-chlorophenyl)-N-(4-(hydroxycarbamoyl)benzyl)-2,3,4,9-tetrahydro-1H-pyrido[3, 4-b]indole-3-carboxamide (14g) showed potent broad antiproliferative activity against various tumors including leukemia, colon cancer, melanoma, and breast cancer cell lines, better than ACY-1215. Moreover, 14g also showed good pharmacokinetics properties in mice via oral administration.

Loss of p53 and mutational heterogeneity drives immune resistance in an autochthonous mouse lung cancer model with high tumor mutational burden.

The role of tumor mutational burden (TMB) in shaping tumor immunity is a key question that has not been addressable using genetically engineered mouse models (GEMMs) of lung cancer. To induce TMB in lung GEMMs, we expressed an ultra-mutator variant of DNA polymerase-E (POLE)P286R in lung epithelial cells. Introduction of PoleP286R allele into KrasG12D and KrasG12D; p53L/L (KP) models significantly increase their TMB. Immunogenicity and sensitivity to immune checkpoint blockade (ICB) induced by Pole is partially dependent on p53. Corroborating these observations, survival of NSCLC patients whose tumors have TP53truncating mutations is shorter than those with TP53WT with immunotherapy. Immune resistance is in part through reduced antigen presentation and in part due to mutational heterogeneity. Total STING protein levels are elevated in Pole mutated KP tumors creating a vulnerability. A stable polyvalent STING agonist or p53 induction increases sensitivity to immunotherapy offering therapeutic options in these polyclonal tumors.

Elucidation of Protonation Cooperativity of a STING-Activating Polymer.

Stimuli-responsive nanomaterials have the potential to improve the performance and overcome existing barriers of conventional nanotherapeutics. Molecular cooperativity design in stimuli-responsive nanomedicine can amplify physiological signals, enabling a cooperative response for improved diagnostic and therapeutic precision. Previously, this work reported an ultra-pH-sensitive polymer, PEG-b-PC7A, that possesses innate immune activating properties by binding to the stimulator of interferon genes (STING) through polyvalent phase condensation. This interaction enhances STING activation and synergizes with the endogenous STING ligand for robust cancer immunotherapy. Despite its successes in innate immune activation, the fundamental physicochemical and pH-responsive properties of PC7A require further investigation. Here, this study elucidates the protonation cooperativity driven by the phase transition of PC7A copolymer. The highly cooperative system displays an "all-or-nothing" proton distribution between highly charged unimer (all) and neutral micelle (nothing) states without gradually protonated intermediates. The binary protonation behavior is further illustrated in pH-precision-controlled release of a representative anticancer drug, ß-lapachone, by PC7A micelles over a noncooperative PE5A polymer. Furthermore, the bimodal distribution of protons is represented by a high Hill coefficient (nH  > 9), featuring strong positive cooperativity. This study highlights the nanoscale pH cooperativity of an immune activating polymer, providing insights into the physicochemical characterization and design parameters for future nanotherapeutics development.

The Cancer Moonshot Immuno-Oncology Translational Network at 5: accelerating cancer immunotherapies.

The Immuno-Oncology Translational Network (IOTN) was established in 2018 as part of the Cancer Moonshot. In 2022, President Joe Biden set new goals to reduce the cancer death rate by half within 25 years and improve the lives of people with cancer and cancer survivors. The IOTN is focused on accelerating translation of cancer immunology research, from bench to bedside, and improving immunotherapy outcomes across a wide array of cancers in the adult population. The unique structure and team science approach of the IOTN is designed to accelerate discovery and evaluation of novel immune-based therapeutic and prevention strategies. In this article, we describe IOTN progress to date, including new initiatives and the development of a robust set of resources to advance cancer immunology research. We summarize new insights by IOTN researchers, some of which are ripe for translation for several types of cancers. Looking to the future, we identify barriers to the translation of immuno-oncology concepts into clinical trials and key areas for action and improvements that are suitable for high-yield investments. Based on these experiences, we recommend novel National Institutes of Health funding mechanisms and development of new resources to address these barriers.

Iterative-Acting Thioesterase from Polyketide Biosynthesis Accepts Diverse Nucleophilic Alcohols to Yield Oxazole-Containing Esters.

The biosynthesis of antitumor oxazole-containing conglobatin is directed by a multienzyme assembly line of nonribosomal peptide synthetase (NRPS) and polyketide synthase (PKS), in which an uncanonical iterative-acting C-terminal thioesterase domain, Cong-TE, ligated two fully elongated chains/conglobatin monomers on the terminal acylcarrier protein and subsequently cyclized the resulting dimer to a C2-symmetric macrodiolide. Screening of the conglobatin producer for secondary metabolites led to the discovery of two new compounds conglactones A (1) and B (2), possessing inhibitory activities to phytopathogenic microorganisms and cancer cells, respectively. The compounds 1 and 2 feature the ester bond-linked hybrid structures consisting of an aromatic polyketide benwamycin I (3) and one (for 1)/two (for 2) molecules of the conglobatin monomer (5). Genetic mutational analysis revealed that the production of 1 and 2 was correlated with the biosynthetic pathways of 3 and 5. Biochemical analysis indicated that 1 and 2 were produced by Cong-TE from 3 and an N-acetylcysteamine thioester form of 5 (7). Furthermore, the substrate compatibility of Cong-TE was demonstrated by enzymatically generating a bunch of ester products from 7 and 43 exotic alcohols. This property of Cong-TE was further validated by producing 36 hybrid esters in the fermentation of conglobatin producer fed with nonindigenous alcohols. This work shows a prospect of developing Cong-TE for green synthesis of valuable oxazole-containing esters, thus complementing the environmentally unfriendly chemosynthesis strategies.

Trienomycin A-simplified analogs: Synthesis and anti-neuroinflammatory activity.

A series of novel trienomycin A (TA)-mimetic compounds (5a-p) have been designed, synthesized, and evaluated for their in vitro anti-neuroinflammatory and neuroprotective activities. Among them, compounds 5h, 5n, and 5o exhibits relatively strong NO inhibitory activity in LPS-activated BV-2 cells with the EC50 values of 12.4, 17.3, and 8.9 µM, respectively. Moreover, 5h showed evidently neuroprotective effect against H2O2-induced PC-12 cells without cytotoxicity at 20 µM. Overall, these compounds can provide a better understanding of the structure-activity relationship of TA and furnish research ideas for anti-neuroinflammatory and neuroprotective agents.