Discovery and preclinical evaluations of TQB3616, a novel CDK4-biased inhibitor.

Among small-molecule CDK4/6 inhibitors (palbociclib, ribociclib, and abemaciclib) approved for metastatic breast cancers, abemaciclib has a more tolerable adverse effects in clinic. This is attributable to preferential inhibition of CDK4 over CDK6. In our search for a biased CDK4 inhibitor, we discovered a series of pyrimidine-indazole inhibitors. SAR studies led us to TQB3616 as a preferential CDK4 inhibitor. TQB3616 exhibited improvements in both enzymatic and cellular proliferation inhibitory potency when tested side-by-side with the FDA approved palbociclib and abemaciclib. TQB3616 also possessed favorable PK profile in multiple species. These differentiated properties, together with excellent GLP safety profile warranted TQB3616 moving to clinic. TQB3616 entered into clinical development in 2019 and currently in phase III clinical trials (NCT05375461, NCT05365178).

The interplay of soil physicochemical properties, methanogenic diversity, and abundance governs methane production potential in paddy soil subjected to multi-decadal straw incorporation.

Straw incorporation holds significant promise for enhancing soil fertility and mitigating air pollution stemming from straw burning. However, this practice concurrently elevates the production and emission of methane (CH4) from paddy ecosystems. Despite its environmental impact, the precise mechanisms behind the heightened CH4 production resulting from long-term straw incorporation remain elusive. In a 32-year field experiment featuring three fertilization treatments (CFS-chemical fertilizer with wheat straw, CF-chemical fertilizer, and CK-unamended), we investigated the impact of abiotic (soil physicochemical properties) and biotic (methanogenic abundance, diversity, and community composition) factors on CH4 production in paddy fields. Results revealed a significantly higher CH4 production potential under CFS treatment compared to CF and CK treatments. The partial least squares path model revealed that soil physicochemical properties (path coefficient = 0.61), methanogenic diversity (path coefficient = -0.43), and methanogenic abundance (path coefficient = 0.29) collectively determined CH4 production potential, explaining 77% of the variance. Enhanced soil organic carbon content and water content, resulting from straw incorporation, emerged as pivotal factors positively correlated with CH4 production potential. Under CFS treatment, lower Shannon index of methanogens, compared to CF and CK treatments, was attributed to increased Methanosarcina. Notably, the Shannon index and relative abundance of Methanosarcina exhibited negative and positive correlations with CH4 production potential, respectively. Methanogenic abundance, bolstered by straw incorporation, significantly amplified overall potential. This comprehensive analysis underscores the joint influence of abiotic and biotic factors in regulating CH4 production potential during multi-decadal straw incorporation.

Vertical distribution of Candidatus Methylomirabilis and Methanoperedens in agricultural soils.

Candidatus Methylomirabilis-related bacteria conduct anaerobic oxidation of methane (AOM) coupling with NO2- reduction, and Candidatus Methanoperedens-related archaea perform AOM coupling with reduction of diverse electron acceptors, including NO3-, Fe (III), Mn (IV) and SO42-. Application of nitrogen fertilization favors the growth of these methanotrophs in agricultural fields. Here, we explored the vertical variations in community structure and abundance of the two groups of methanotrophs in a nitrogen-rich vegetable field via using illumina MiSeq sequencing and quantitative PCR. The retrieved Methylomirabilis-related sequences had 91.12%-97.32% identity to the genomes of known Methylomirabilis species, and Methanoperedens-related sequences showed 85.49%-97.48% identity to the genomes of known Methanoperedens species which are capable of conducting AOM coupling with reduction of NO3- or Fe (III). The Methanoperedens-related archaeal diversity was significantly higher than Methylomirabilis-related bacteria, with totally 74 and 16 operational taxonomic units, respectively. In contrast, no significant difference in abundance between the bacteria (9.19 × 103-3.83 × 105 copies g-1 dry soil) and the archaea (1.55 × 104-3.24 × 105 copies g-1 dry soil) was observed. Furthermore, the abundance of both groups of methanotrophs exhibited a strong vertical variation, which peaked at 30-40 and 20-30 cm layers, respectively. Soil water content and pH were the key factors influencing Methylomirabilis-related bacterial diversity and abundance, respectively. For the Methanoperedens-related archaea, both soil pH and ammonium content contributed significantly to the changes of these archaeal diversity and abundance. Overall, we provide the first insights into the vertical distribution and regulation of Methylomirabilis-related bacteria and Methanoperedens-related archaea in vegetable soils. KEY POINTS: • The archaeal diversity was significantly higher than bacterial. • There was no significant difference in the abundance between bacteria and archaea. • The abundance of bacteria and archaea peaked at 30-40 and 20-30 cm, respectively.

Detection and Quantification of Candidatus Methanoperedens-Like Archaea in Freshwater Wetland Soils.

Candidatus Methanoperedens-like archaea, which can use multiple electron acceptors (nitrate, iron, manganese, and sulfate) for anaerobic methane oxidation, could play an important role in reducing methane emissions from freshwater wetlands. Currently, very little is known about the distribution and community composition of Methanoperedens-like archaea in freshwater wetlands, particularly based on their alpha subunit of methyl-coenzyme M reductase (mcrA) genes. Here, the community composition, diversity, and abundance of Methanoperedens-like archaea were investigated in a freshwater wetland through high-throughput sequencing and quantitative PCR on their mcrA genes. A large number of Methanoperedens-like mcrA gene sequences (119,250) were recovered, and a total of 31 operational taxonomic units (OTUs) were generated based on 95% sequence similarity cut-off. The majority of Methanoperedens-like sequences can be grouped into three distinct clusters that were closely associated with the known Methanoperedens species which can couple anaerobic methane oxidation to nitrate or iron reduction. The community composition of Methanoperedens-like archaea differed significantly among different sampling sites, and their mcrA gene abundance was 1.49 × 106 ~ 4.62 × 106 copies g-1 dry soil in the examined wetland. In addition, the community composition of Methanoperedens-like archaea was significantly affected by the soil water content, and the archaeal abundance was significantly positively correlated with the water content. Our results suggest that the mcrA gene is a good biomarker for detection and quantification of Methanoperedens-like archaea, and provide new insights into the distribution and environmental regulation of these archaea in freshwater wetlands.

Effects of abrupt and gradual increase of atmospheric CO2 concentration on methanotrophs in paddy fields.

The simulation of abrupt atmospheric CO2 increase is a common way to examine the response of soil methanotrophs to future climate change. However, atmosphere is undergoing a gradual CO2 increase, and it is unknown whether the previously reported response of methanotrophs to abrupt CO2 increase can well represent their response to the gradual increase. To improve the understanding of the effect of elevated CO2 (eCO2) on methanotrophs in paddy ecosystems, the methane oxidation potential and communities of methanotrophs were examined via open top chambers under the three following CO2 treatments: an ambient CO2 concentration (AC); an abrupt CO2 increase by 200 ppm above AC (AI); a gradual CO2 increase by 40 ppm each year until 200 ppm above AC (GI). Relative to AC treatment, AI and GI treatments significantly (p < 0.05) increased the methane oxidation rate by 43.8% and 36.7%, respectively, during rice growth period. Furthermore, the abundance of pmoA genes was significantly (p < 0.05) increased by 62.4% and 32.5%, respectively, under AI and GI treatments. However, there were no significant variations in oxidation rate or gene abundance between the two eCO2 treatments. In addition, no obvious change of overall community composition of methanotrophs was observed among treatments, while the proportions of Methylosarcina and Methylocystis significantly (p < 0.05) changed. Taken together, our results indicate similar response of methanotrophs to abrupt and gradual CO2 increase, although the magnitude of response under gradual increase was smaller and the abrupt increase may somewhat overestimate the response.

Effect of gradual increase of atmospheric CO2 concentration on nitrification potential and communities of ammonia oxidizers in paddy fields.

Currently, the influence of elevated atmospheric CO2 concentration (eCO2) on ammonia oxidation to nitrite, the rate-limiting step of nitrification in paddy soil, is poorly known. Previous studies that simulate the effect of eCO2 on nitrification are primarily based on an abrupt increase of atmospheric CO2 concentration. However, paddy ecosystems are experiencing a gradual increase of CO2 concentration. To better understand how the nitrification potential, abundance and communities of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) respond to eCO2 in paddy ecosystems, a field experiment was conducted using the following two treatments: a gradual increase of CO2 (EC, increase of 40 ppm per year until 200 ppm above ambient) and ambient CO2 (CK). The results demonstrated that the EC treatment significantly (P < 0.05) stimulated the soil potential nitrification rate (PNR) at the jointing and milky stages, which increased by 127.83% and 27.35%, respectively, compared with CK. Furthermore, the EC treatment significantly (P < 0.05) stimulated the AOA and AOB abundance by 56.60% and 133.84%, respectively, at the jointing stage. Correlation analysis showed that the PNR correlated well with the abundance of AOB (R2 = 0.7389, P < 0.001). In addition, the EC treatment significantly (P < 0.05) altered the community structure of AOB, while it had little effect on that of AOA. A significant difference in the proportion of Nitrosospira was observed between CO2 treatments. In conclusion, the gradual increase of CO2 positively influenced the PNR and abundance of ammonia oxidizers, and AOB could be more important than AOA in nitrification under eCO2.

Effect of microwave ablation treatment of hepatic malignancies on serum cytokine levels.

BACKGROUND: Microwave ablation (MWA) is widely used to treat unresectable primary and secondary malignancies of the liver, and a limited number of studies indicate that ablation can cause not only necrosis at the in situ site but also an immunoreaction of the whole body. This study aimed to investigate the effects of MWA on cytokines in patients who underwent MWA for a hepatic malignancy. METHODS: Patients admitted to the Oncology Department in the First Affiliated Hospital of Soochow University between June 2015 and February 2019 were selected. Peripheral blood was collected from patients with a hepatic malignancy treated with MWA. The levels of cytokines (IL-2, IFN-γ, TNF-α, IL-12 p40, IL-12 p70, IL-4, IL-6, IL-8, IL-10, and vascular endothelial growth factor (VEGF)) were detected with a Milliplex® MAP Kit. The comparison times were as follows: before ablation, 24 h after ablation, 15 days after ablation, and 30 days after ablation. Data were analyzed using a paired sample t-tests and Spearman's correlation analysis. RESULTS: A total of 43 patients with hepatic malignancies were assessed. There were significant differences in IL-2, IL-12 p40, IL-12 p70, IL-1ß, IL-8, and TNF-α at 24 h after MWA. Significant increases (> 2-fold vs. before ablation) were observed in IL-2, IL-1ß, IL-6, IL-8, IL-10, and TNF-α after MWA. Elevated IL-2 and IL-6 levels after ablation were positively correlated with energy output during the MWA procedure. CONCLUSIONS: WA treatment for hepatic malignancies can alter the serum levels of several cytokines such as IL-2 and IL-6.

Ethanolic extract of Arctium lappa leaves alleviates cerebral ischemia reperfusion-induced inflammatory injury via HDAC9-mediated NF-κB pathway.

BACKGROUND: Ischemic stroke (IS) is a major cause of mortality and disability worldwide. Inflammatory response is crucial in the pathogenesis of tissue injury in cerebral infarction. Arctium lappa leaves are traditionally used to treat IS. PURPOSES: To investigate the neuroprotective effects and molecular mechanisms of the ethanolic extract of A. lappa leaves (ALLEE) on cerebral ischemia-reperfusion (CIR). METHODS: Middle cerebral artery obstruction reperfusion (MCAO/R) rats and an oxygen-glucose deprivation/reoxygenation (OGD/R) cell model were used to evaluate ALLEE pharmacodynamics. Various methods, including neurological function, 2,3,5-triphenyltetrazolium chloride, hematoxylin and eosin, and Nissl, enzyme-linked immunosorbent, and TdT-mediated dUTP nick-end labeling assays, were used to analyze the neuroprotective effects of ALLEE in vitro and in vivo. The major chemical components and potential target genes of ALLEE were screened using network pharmacology. Molecular docking, western blotting, and immunofluorescence analyses were performed to confirm the effectiveness of the targets in related pathways. RESULTS: ALLEE exerted potent effects on the MCAO/R model by decreasing the neurological scores, infarct volumes, and pathological features (p < 0.01). Furthermore, network pharmacology results revealed that the treatment of IS with ALLEE involved the regulation of various inflammatory pathways, such as the tumor necrosis factor (TNF) and chemokine signaling pathways. ALLEE also played key roles in targeting key molecules, including nuclear factor (NF)-κBIA, NF-κB1, interleukin (IL)-6, TNF-α and IL1ß, and regulating the histone deacetylase (HDAC)-9-mediated signaling pathway. In vivo and in vitro analyses revealed that ALLEE significantly regulated the NF-κB pathway, promoted the phosphorylation activation of NF-κB P65, IκB and IKK (p < 0.01 or p < 0.05), and decreased the expression levels of the inflammatory factors, IL-1ß, IL-6 and TNF-α (p < 0.01). Moreover, ALLEE significantly decreased the expression of HDAC9 (p < 0.01) that is associated with inflammatory responses. However, HDAC9 overexpression partially reversed the neuroprotective effects of ALLEE and its suppressive effects on inflammation and phosphorylation of NF-κB (p < 0.01). CONCLUSIONS: In conclusion, our results revealed that ALLEE ameliorates MCAO/R-induced experimental CIR by modulating inflammatory responses via the inhibition of HDAC9-mediated NF-κB pathway.

Equal importance of humic acids and nitrate in driving anaerobic oxidation of methane in paddy soils.

Methane (CH4) is both generated and consumed in paddy soils, where anaerobic oxidation of methane (AOM) serves as a crucial process for mitigating CH4 emissions. Although the participation of humic acids (HA) and nitrate in AOM has been recognized, their relative roles and significance in paddy soils remain insufficiently investigated. In this study, we explored the potential activity of AOM driven by HA and nitrate, as well as the composition of archaeal communities in paddy soils across different rice growth periods and fertilization treatments. AOM activity ranged from 0.81 to 1.33 and 1.26 to 2.38 nmol of 13CO2 g-1 (dry soil) day-1 with HA and nitrate, respectively. No significant differences (p < 0.05) were observed between the AOM activity driven by HA and nitrate across the three fertilization treatments. According to AOM activity, the annual consumption of CH4 was estimated at approximately 0.49 ± 0.06 and 0.83 ± 0.19 Tg for AOM processes driven by HA and nitrate in Chinese paddy soils. Nitrate-driven AOM activity exhibited a positive (p < 0.05) correlation with the abundance of the ANME-2d mcrA gene but a negative (p < 0.05) correlation with the content of dissolved organic carbon. Intriguingly, HA-driven AOM activity was only correlated positively with the nitrate-driven AOM activity. Soil water content, soil organic carbon, nitrate and nitrite contents were significantly correlated with the relative abundance of methanogenic and methanotrophic archaea. These results identified the potential importance of HA and nitrate in driving AOM processes within paddy soils, providing a comprehensive understanding of the complex microbial processes regulating greenhouse gas emissions from paddy soils.

Nitrogen fertilization rate affects communities of ammonia-oxidizing archaea and bacteria in paddy soils across different climatic zones of China.

Nitrogen fertilization has important effects on nitrification. However, how the rate of nitrogen fertilization affects nitrification potential, as well as the communities of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB), remains unclear. We performed a large-scale investigation of nitrification potential and ammonia-oxidizer communities in Chinese paddy fields at different nitrogen fertilization rates across different climatic zones. It was found that the nitrification potential at the high nitrogen fertilization rate (≥150 kg-1 N ha-1) was 23.35 % higher than that at the intermediate rate (100-150 kg-1 N ha-1) and 20.77 % higher than that at the low rate (< 100 kg-1 N ha-1). The nitrification potential showed no significant variation among different nitrogen fertilization rates across climatic zones. Furthermore, the AOA and AOB amoA gene abundance at the high nitrogen fertilization rate was 481.67 % and 292.74 % higher (p < 0.05) than that at the intermediate rate, respectively. Correlation analysis demonstrated a significant positive correlation between AOB abundance and nitrification potential. AOA and AOB community composition differed significantly among nitrogen fertilization rates. Moreover, soil NH4+ content, pH, water content, bulk density, and annual average temperature were regarded as key environmental factors influencing the community structure of ammonia-oxidizers. Taken together, the nitrogen fertilization rate had a significant impact on the communities of AOA and AOB but did not significantly alter the nitrification potential. Our findings provide new insights into the impact of nitrogen fertilization management on nitrification in rice paddy fields.