Adenosine kinase (ADK, EC: 2.7.1.20) is an evolutionarily ancient ribokinase, which acts as a metabolic regulator by transferring a phosphoryl group to adenosine to form AMP. The enzyme is of interest as a therapeutic target because its inhibition is one of the most effective means to raise the levels of adenosine and hence adenosine receptor activation. For these reasons, ADK has received significant attention in drug discovery efforts in the early 2000s for indications such as epilepsy, chronic pain, and inflammation; however, the report of adverse events regarding cardiovascular and hepatic function as well as instances of microhemorrhage in the brain of preclinical models prevented further development efforts. Recent findings emphasize the importance of compartmentalization of the adenosine system reflected by two distinct isoforms of the enzyme, ADK-S and ADK-L, expressed in the cytoplasm and the cell nucleus, respectively. Newly identified adenosine receptor independent functions of adenosine as a regulator of biochemical transmethylation reactions, which include DNA and histone methylation, identify ADK-L as a distinct therapeutic target for the regulation of the nuclear methylome. This newly recognized role of ADK-L as an epigenetic regulator points toward the potential disease-modifying properties of the next generation of ADK inhibitors. Continued efforts to develop therapeutic strategies to separate nuclear from extracellular functions of adenosine would enable the development of targeted therapeutics with reduced adverse event potential. This review will summarize recent advances in the discovery of novel ADK inhibitors and discuss their potential therapeutic use in conditions ranging from epilepsy to cancer.

Cancer stem-like cells (CSCs), a small subset of cells within the tumor microenvironment, favor tumor relapse and remain a major obstacle in cancer therapy. Hence, hyaluronan-based mesoporous silica nano-formulation with phloroglucinol (MSN-PG-HA) was employed in our study to address the cancer relapse by targeting CD44. Various in vitro cellular assays were conducted to assess the nano-formulation's effect on suppressing CSC characteristics in AGS, HCT-116 and SW-620. The CUA assay indicated increased uptake of FITC-labeled MSN-PG-HA by all GI cancer cell lines and no uptake by CD44- (3T3) cell line confirming the receptor-mediated endocytosis. Moreover, MSN-PG-HA nano-formulation has shown a significantly higher efficacy than free PG in other cellular assays by controlling cell migration, colony and spheroid formation, inducing apoptosis and suppressing the elevated MMP levels, for all the three GI cell lines with p ≤ 0.05, highlighting the nano-formulation's ability to target CSC. Further, the gene and protein expression analyses of hedgehog signaling pathway components, such as GLI1 and SMO, showed a 0.7 to 0.8-fold decrease in expression with MSN-PG-HA treated groups, proving that our nano-formulation could significantly target CSC-related proteins. Furthermore, the expression analysis utilizing inhibitors such as GANT-61 (GLI1 inhibitor) and Sonidegib (SMO inhibitor) have demonstrated that the MSN-PG-HA has shown a significant level of inhibition as that of GANT-61, indicating a similarity in their mechanism of action in inhibiting the cancer cell proliferation. Thus, our findings conclude that MSN-PG-HA could serve as a potential drug for targeting GI cancer stem cells.

The potential regulatory mechanisms of 100 μM melatonin (MT) treatment on proline accumulation in cantaloupe fruit under cold stress were investigated. The results showed that MT treatment increased the content of free proline in cold-stored cantaloupe fruit by upregulating the expressions of the Δ1-pyrroline-5-carboxylate synthase (P5CS) and ornithine δ-aminotransferase (OAT) genes and decreasing the transcripts and enzyme activity of proline dehydrogenase (ProDH). Furthermore, the yeast one-hybrid assay screened for CmDREB1A and CmDREB1E that bind CmP5CS, and electrophoretic mobility shift analysis and dual luciferase reporter gene verified that these two transcription factors could bind the CmP5CS promoter. Further experiments showed that CmDREB1A and CmDREB1E were involved in improving cold tolerance in cantaloupe by MT and regulating the expression of CmP5CS. Our study suggests that MT treatment promotes CmP5CS expression and positively regulates proline accumulation by activating CmDREB1A and CmDREB1E in cantaloupe, thereby improving fruit cold stress tolerance.

Endostar is a human recombinant endostatin which is an attractive anti-angiogenesis protein. Because inefficient antigen presenting MHC class I expression (which can be downregulated by HIF-1) is an important strategy for cancer immune evasion, besides its anti-angiogenesis effect, it remains unclear whether Endostar has an inhibitory effect on HIF-1 expression by upregulating MHC class I expression in cancer cells to facilitate immunotherapies, including PD-1/PD-L1 inhibitors. In this study, A549 and NCI-H1299 lung cancer cells were treated with Endostar (6.25 μg/ml, 12.5 μg/ml, and 25 μg/ml, respectively). HIF-1 expression was detected by Immunocytochemistry and Western blot. Proteins of the MHC class I α-heavy chain and β2 m light chain, STAT3 and pSTAT3 were detected by Western blot. The mRNAs of MHC class I α-heavy chain and β2 m light chain were detected by RT-qPCR. It was shown that decreased expression of HIF-1 and promotion of β2-microglobulin were observed after Endostar treatment. In addition, elevated levels of MHC class I α-heavy chain mRNA and protein, as well as downregulation of STAT3 and pSTAT3, were also observed following Endostar treatment. Endostar inhibited HIF-1 expression in A549 and NCI-H1299 lung cancer cells, upregulated expression of MHC class I α-heavy chain and β2 m light chain, with the upregulation of STAT3 and pSTAT3, suggesting involvement of STAT3 pathway. It is important because only in combination with MHC class I on target cells can tumor antigenic peptides be recognized by CD8+ CTLs which destroy target cells. However, MHC class I is frequently deficient in cancer cells.

Benign prostatic hyperplasia (BPH) poses a significant public health challenge, affecting a substantial portion of aging men worldwide. Current therapeutic options offer limited efficacy. The pathogenesis of BPH is multifactorial, involving ferroptosis, oxidative stress, and chronic inflammation. Hydroxycitric acid (HCA) is a natural compound with diverse pharmacological activities, including the inhibition of ferroptosis, anti-inflammatory, anti-oxidative stress, and anti-tumor effects. However, its role in BPH remains unexplored. This study aimed to investigate the effects of HCA on BPH and elucidate the underlying mechanisms, with the goal of providing novel therapeutic insights for BPH treatment.

Heteroresistance and persistence are examples of mechanisms that can allow otherwise drug-susceptible bacteria to survive and resume growth after antibiotic exposure. These temporary forms of antibiotic tolerance can be caused by the upregulation of stress response genes or a decrease in cell growth rate. However, it is not clear how the expression of multiple genes contributes to tolerance phenotypes. Using fluorescent reporters for stress-related genes, we conducted real-time measurements of expression prior to, during, and after antibiotic exposure. We first identified relationships between growth rate and reporter levels based on auto- and cross-correlation analysis, revealing consistent patterns where changes in growth rate were anticorrelated with fluorescence following a delay. We then used pairs of stress gene reporters and time-lapse fluorescence microscopy to measure the growth rate and reporter levels in cells that survived or died following antibiotic exposure. Using these data, we asked whether combined information about reporter expression and growth rate could improve our ability to predict whether a cell would survive or die following antibiotic exposure. We developed a Bayesian inference model to predict how the combination of dual reporter expression levels and growth rate impacts ciprofloxacin survival in Escherichia coli. We found clear evidence of the impact of growth rate and gadX promoter activity on survival. Unexpectedly, our results also revealed examples where additional information from multiple genes decreased prediction accuracy, highlighting an important and underappreciated effect that can occur when integrating data from multiple simultaneous measurements.IMPORTANCETransient increases in bacterial antibiotic tolerance can result in treatment failure despite an infection initially presenting as susceptible, presenting a significant challenge in antibiotic therapy. This phenomenon can also provide a window of opportunity for bacteria to acquire permanent genetic resistance mutations. Although understanding the underlying mechanisms of these antibiotic tolerance phenotypes is crucial for developing effective approaches to treatment, current approaches for studying these transient phenotypes have limitations. Here, we use fluorescent reporters to monitor the expression of genes involved in stress response over time, aiming to link expression with antibiotic survival outcomes. Our results reveal a counterintuitive finding: monitoring multiple gene reporters does not necessarily improve our ability to predict antibiotic survival outcomes compared to single gene reporters. This result emphasizes the need for a deeper mechanistic understanding of the relationship between stress response gene expression and antibiotic tolerance.

We previously showed that proliferating cell nuclear antigen (PCNA) interacts with androgen receptor (AR) through a PIP-box (PIP-box4) at the N-terminus of AR and regulates AR activity. In this study, we further investigated PCNA/AR interaction. We identified a second PIP-box (PIP-box592) in the DNA binding domain of AR and found that dihydrotestosterone enhances the binding of full-length AR (AR-FL) but not a constitutively active variant (AR-V7) to PCNA. Treatment with R9-AR-PIP, a PIP-box4-mimicking small peptide, inhibits the PCNA/AR interaction, AR occupancy at the androgen response element (ARE) in PSA and p21 genes, and expression of AR target genes, and induces cytotoxicity in AR-positive castration-resistant prostate cancer (CRPC) cells. R9-AR-PIP also significantly inhibits transcriptional activity of AR-FL upon dihydrotestosterone stimulation and the constitutive activity of AR-V7. Moreover, R9-AR-PIP and PCNA-I1S, a small molecule PCNA inhibitor, inhibit the ARE occupancy by AR-FL and AR-Vs in CCNA2 gene that encodes cyclin A2 and cyclin A2 expression. Finally, we found that cyclin A2 is overexpressed in all CRPC cells examined, suggesting that it may contribute to the development of CRPC. These data indicate that targeting PCNA/AR interaction inhibits both AR-FL- and AR-Vs-mediated signaling and implicates it could be a novel therapeutic strategy against CRPC.

Sulforaphane (SFN), a lipophilic small-molecule compound, can be rapidly and completely absorbed upon entering the body. It has garnered extensive research attention for its potential as an antiaging, anticancer, antidiabetic, and antibacterial agent. However, its role and mechanisms of SFN on skeletal muscle postinjury regeneration have not been reported. This research demonstrated that SFN enhanced the regeneration after skeletal muscle injury and up-regulated the proliferation of mouse C2C12 myoblasts. RNA-transcriptome sequencing data revealed that SFN increased Prl2C2 transcription and JAK/STAT signaling pathway activity. CHIP and dual-luciferase reporter gene assays verified that STAT3 binds to the Prl2C2 promoter and regulates its transcription. Consequently, SFN influenced the JAK2/STAT3 signaling activity. Finally, the transcription of Prl2C2 and the proliferation of mouse C2C12 myoblasts were detected by adding JAK2 inhibitor and SFN. The results showed that the JAK2 inhibitor blocked the up-regulation of SFN on the transcription of Prl2C2 and the proliferation of mouse C2C12 myoblasts. The discovery of this phenomenon and its mechanism offer guidance for treating skeletal muscle injuries and supporting animal nutrition research. SFN shows great potential in muscle repair, and future clinical trials could confirm its safety and efficacy, paving the way for new SFN-based treatments and providing new options for patients.

Stomach cancer has become one of the most common types of cancer, with its mortality rate ranking third in the world. Currently, the main treatments for gastric cancer are surgery, radiation therapy, and chemotherapy. Although current treatments can effectively prevent postoperative metastasis and recurrence of gastric cancer, they may also bring various adverse reactions in the gastrointestinal tract and side effects such as bone marrow suppression. Years of research have confirmed that traditional Chinese medicine treatment for gastric and other cancers has distinct characteristics and advantages. Combined treatment can increase the tumor inhibition rate, reduce the side effects of radiation and chemotherapy, improve patients' quality of life, and prolong the survival prognosis.

Gemcitabine (GEM) is used as a first-line therapy for patients diagnosed with any stage of pancreatic cancer (PC); however, patient survival is poor because of GEM resistance. Thus, new approaches to overcome GEM resistance in PC are urgently needed. Here, we aimed to establish an in vivo drug-resistant PC model and identify genes involved in GEM resistance. We focused on one of these factors, CITED4, and elucidated its mechanisms of action in GEM resistance in PC.

Recent research indicates that programmed death 1 (PD-1) and programmed death-ligand 1 (PD-L1) inhibitors show promise in treating triple-negative breast cancer (TNBC), but their efficacy is lower than anticipated, especially when used alone. Therefore, enhancing the anti-tumor immune response strategy for TNBC is crucial. Ubiquitin carboxy-terminal hydrolase L1 (UCH-L1), involved in tumor cell regulation and a potential therapeutic target, has an undefined role in TNBC immunotherapy. In this study, we explored the inverse correlation between UCH-L1 and PD-L1 in TNBC patient tissues. Through in vitro experiments, we found that UCH-L1 negatively regulates PD-L1 by stabilizing the E3 ubiquitin ligase ariadne-1 homolog (ARIH1), which promotes PD-L1 ubiquitination and degradation. Further analysis in Balb/c mice xenograft tumors showed that UCH-L1 correlates with GZMB+/CD8+ T cell infiltration in TNBC, suggesting potential synergistic effects when combining UCH-L1 inhibitors with PD-L1 antibodies. Overall, in TNBC, UCH-L1 stabilizes ARIH1, leading to low PD-L1 expression, which may explain the limited effectiveness of immunotherapy in TNBC patients. Our mouse experiments showed improved therapeutic effects when combining UCH-L1 inhibitors with PD-L1 antibodies. These findings offer a new avenue for immunotherapy in TNBC patients.

(R)-β-homoserine (RBH) is a structural analogue of β-aminobutyric acid (BABA) that can enhance plant resistance to a wide range of pathogens. Here, we investigated the regulatory role of VvTCP7 on the RBH-induced priming response against Botrytis cinerea in grapes. The results showed that RBH primed a defense mechanism in grape berries and enhanced their response to fungal infection. RBH upregulated the expression of a group of genes involved in SA synthesis, thus inducing SA accumulation in grapes. VvTCP7 has high homology to AtCHE in Arabidopsis thaliana and is recognized to be a nucleus-localized protein that promotes SA synthesis. Notably, RBH elevated VvTCP7 expression in harvested grape berries, which was accompanied by enhanced expression of VvNPR1, a master regulator of SAR, as well as the SA-responsive PR genes. Additionally, Y1H, EMSA and DLR assays confirmed that VvTCP7 has the ability to bind directly to the GGNCCC motif within the VvICS promoter to induce VvICS transcription and SA synthesis. Overexpression of VvTCP7 in Arabidopsis led to a marked increase in the transcription of PR genes, enhancing defensive response to B. cinerea. However, the VvTCP7 knockout led to a decrease in PR gene expression and increased susceptibility to the fungus. Collectively, the data suggest that VvTCP7 contributes to RBH-induced SAR priming by activating the SA synthesis and resultant enhances SAR defenses to combat fungal invasion.

We demonstrate that exposure to the AB5 subtilase cytotoxin (SubAB) induces the unfolded protein response (UPR) in human peripheral blood mononuclear cells, concomitant with a proinflammatory response across distinct cell subsets. Notably, SubAB selectively induces type-I interferon (IFN) expression in plasmacytoid dendritic cells, acting synergistically with Toll-like receptor 7 stimulation. The induction of type-I IFN in response to SubAB relies on stimulator of interferon genes (STING) activation, coupled with protein synthesis inhibition mediated by protein kinase R-like endoplasmic reticulum kinase (PERK) and phosphorylation of the eukaryotic translation initiation factor 2 subunit-alpha. By impeding mRNA translation through the integrated stress response, SubAB precipitates the downregulation of the negative innate signaling feedback regulator Tax1-binding protein 1. This downregulation is necessary to unleash TANK-binding kinase 1 signaling associated with STING activation. These findings shed light on how UPR-inducing conditions may regulate the immune system during infection or pathogenesis.

To investigate if FVIII-Fc Fusion protein (FcFVIII) may modulate inflammation and immune stimulation in hemophilic synovium via the Fc-portion of immunoglobulin used for half-life extension we performed gene expression profiling in FVIII-deficient mice. Hemarthrosis was induced by sub-patellar puncture in FVIII-KO mice, + /- periprocedural recombinant human (rh)FVIII,murine (m)FcFVIII, or mIgG2a. Synovium was harvested at baseline and on days (D) 3 and 14, followed by RNA extraction and sequencing, and histological analysis. RNASeq data were processed using standard protocols followed by differential gene expression (DGE) analysis. Functional enrichment analysis generated molecular pathways (KEGG and Reactome). To distinguish between on-target and off-target (related and unrelated to injury/bleed) effects the following groups were compared: i) Baseline vs. injured-saline, ii) injured-saline vs. injured-rhFVIII, iii) injured-saline vs. injured-mFcFVIII. Knee injury in FVIII-KO mice resulted in hemarthrosis, which was prevented by peri-procedural rhFVIII and mFcFVIII treatments. Only a small proportion of genes was affected by FVIII treatment, exhibiting overlap but also distinct differences between both FVIII-preparations. Acutely (D3), mFcFVIII had unique on-target effects related to immune and inflammatory regulation, whereas rhFVIII mostly affected mRNA and protein processing. On day 14, macrophage profiling indicated a transition from M1 to M2, and only mFcFVIII uniquely influenced pathways and genes associated with tissue remodeling and repair. Some mFcFVIII DGE patterns resembled mIgG2a patterns. Synovial vascular remodeling and cartilage health were better with mFcFVIII than rhFVIII. Interestingly, both FVIII-preparations exerted off-target effects on immune system pathways, albeit with temporal differences. These observations provide proof-of-principle that the type of FVIII preparation can influence synovial processes beyond acute hemostasis control, deserving exploration in the setting of joint bleed control in hemophilia.

Acute lung injury (ALI) is a severe inflammatory lung disorder that requires effective therapeutic strategies. Ephedrine (EPH) is the main active component found in medicinal plants of the Ephedra genus and is commonly used to modulate inflammatory responses in various diseases. Hypoxia-inducible factor 1-alpha (HIF1α) is a subunit of hypoxia-inducible factor 1 (HIF1), which plays a critical regulatory role in cellular responses under hypoxic conditions. Moreover, the degradation pathway of HIF1α is regulated by the deubiquitinase Ovarian Tumour Domain-containing Ubiquitin Aldehyde Binding Protein 1 (OTUB1). The aim of this study is to investigate the therapeutic effects of EPH on ALI and its potential therapeutic mechanism. We utilised a lipopolysaccharide (LPS)-induced ALI mouse model and employed various methods for evaluation. Ultimately, our research findings demonstrate that EPH exhibits anti-ALI effects, with the involvement of HIF1α and OTUB1 in the pharmacological actions of EPH. In conclusion, our study results demonstrate that EPH exhibits anti-ALI effects and exerts its protective effects through modulation of the OTUB1 and HIF1α pathways. Our research findings not only lay the foundation for expanding the medicinal applications of EPH but also provide direction for seeking improved treatment strategies for ALI.

Ovarian cancer (OC) remains a leading cause of gynaecological cancer deaths due to late diagnosis and the emergence of resistance to platinum-based chemotherapy, like cisplatin (Cis). Here, we investigated the potential of metformin (Met), a drug commonly used for type 2 diabetes, to overcome Cis resistance in OC. Our findings revealed a synergistic effect of Met with Cis in inhibiting cell viability, proliferation and colony/sphere formation capacity in both cisplatin-sensitive (A2780) and -resistant (A2780/CDDP) ovarian cancer cell lines. This synergistic action triggered apoptosis through DNA damage, S-phase cell cycle arrest and modulation of autophagy. Met also significantly decreased the expression of pluripotency transcription factors (Oct-4, Sox2 and Nanog), indicating its potential to target cancer stem cells (CSCs). Furthermore, the combination therapy downregulated multidrug resistance protein 1 (MDR1) and excision repair cross-complementation group 1 (ERCC1) expression, thereby sensitising resistant cells to Cis-induced cytotoxicity. Additionally, the combination treatment suppressed the Hedgehog (Hh) signalling pathway, which is an important factor in inhibiting CSCs. Our study highlights the potential of the Met signalling pathway to synergise with Cis, overcoming therapeutic resistance in OC by targeting diverse cellular processes, including CSCs, and warrants further investigation in preclinical models.

The combination of gemcitabine and cisplatin serves as a cornerstone in bladder cancer (BC) treatment, yet chemotherapy resistance continues to pose a significant challenge. This study utilizes a novel BC organoid model integrated with drug sensitivity assays to uncover the mechanisms underlying resistance and identify potential therapeutic targets. Our findings reveal that AP1M2 expression is markedly upregulated in gemcitabine- and cisplatin-resistant BC cells and tissues. Elevated AP1M2 levels contribute to enhanced chemotherapy resistance and tumor cell proliferation by facilitating the DNA damage response and increasing RAD54B expression. Mechanistically, AP1M2 interacts with the RNA-binding protein PUM1 to stabilize RAD54B mRNA, thereby supporting DNA repair and survival under chemotherapeutic stress. Notably, inhibition of AP1M2/PUM1-mediated RAD54B expression sensitized BC xenografts to gemcitabine-cisplatin treatment in vivo. These findings unveil a novel mechanism of chemotherapy resistance in BC and highlight the AP1M2/PUM1/RAD54B pathway as a promising therapeutic target to counter resistance and enhance treatment outcomes.

Pseudomonas aeruginosa is an opportunistic pathogen able to form biofilms, contributing to its virulence. With the increasing use of sweeteners in various foods, understanding their influence on bacterial behavior is critical. This study investigated the virulence of P. aeruginosa PAO1 exposed to sweeteners (erythritol, stevia, fructose, coconut sugar, cane sugar, demerara). Sweeteners didn't affect growth rates. Erythritol stimulated biofilm (100 µg/mL, 159.98% formation), while 10 µg/mL of coconut sugar, cane sugar, and demerara promoted lower levels (∼70% formation). Erythritol stimulated exopolysaccharides production but reduced biofilm eDNA. Stevia, fructose, and coconut sugar increased the expression of lasI, lasR, rhlI, rhlR, pqsA, mvfR, and pvdF. HPLC analysis confirmed sucrose as the major sugar in demerara, coconut and cane sugar. Erythritol stimulated biofilm and some virulence genes expression, while other sweeteners' effects varied. Cane sugar was a biofilm inhibitor with a limited gene expression effect. The sweeteners' impact on microorganisms is diverse and should be further investigated.

We investigated the role of Nuclear Receptor Binding SET Domain Protein 2 (NSD2) and microRNAs (miRNAs) in melanoma de-differentiation following Romidepsin and Interferon-α2b (RI) treatment. Melanoma is the most lethal form of skin cancer, and despite advancements in therapy, treatment resistance remains a major challenge. De-differentiation has been widely recognized as a key factor contributing to therapy resistance.

Saline-alkaline stress is a common problem in Akebia trifoliata cultivation. In this study, the enhancing effects of 5-azacytidine (5-AzaC) on the resistance of A. trifoliata to saline-alkaline stress and the underlying mechanisms were investigated. Plant height, stem diameter, biomass, root length, fresh weight of root, and root/shoot ratio of 6-month-old A. trifoliata seedlings were measured after saline-alkaline stress with or without 5-AzaC treatment. Moreover, the contents of photosynthetic pigments, malondialdehyde (MDA), H2O2, sodium, soluble sugar, and proline; activities of superoxide dismutase, peroxidase (POD), and catalase (CAT); and anatomical structures of root, stem, and leaf were assessed. Furthermore, comparative transcriptome sequencing was performed. The results demonstrated that growth and development of A. trifoliata were severely inhibited under saline-alkaline stress, suggesting that the seedlings were exposed to severe oxidative and osmotic stresses. Treatment with exogenous 5-AzaC could significantly relieve the symptoms of saline-alkaline stress in A. trifoliata. Under saline-alkaline stress, 5-AzaC could increase the stem diameter, biomass, root length, fresh weight of root, and root/shoot ratio and minimize damages to the anatomical structure. Moreover, absorption of Na+ was reduced; ionic balance was maintained; POD and CAT activities were significantly improved; proline and soluble sugar contents increased, and H2O2 and MDA contents decreased. Transcriptome analysis revealed that 5-AzaC functioned via regulating KEGG pathways such as plant hormone signal transduction, phenylpropanoid biosynthesis, photosynthesis, amino sugar and nucleotide sugar metabolism, and glutathione metabolism under saline-alkaline stress. Particularly, enhanced expression of genes from the auxin pathway in plant hormone signal transduction; the lignin synthetic pathway in phenylpropanoid biosynthesis; and photosystem II, photosystem I, photosynthetic electron transport, and F-type ATP pathway in photosynthesis may be related to 5-AzaC-induced saline-alkaline resistance. The results provided theoretical references for A. trifoliata cultivation in saline-alkaline soil and application of 5-AzaC to improve saline-alkaline tolerance in plants.