Discovery of Novel 1-Phenylpiperidine Urea-Containing Derivatives Inhibiting ß-Catenin/BCL9 Interaction and Exerting Antitumor Efficacy through the Activation of Antigen Presentation of cDC1 Cells.

Aberrant activation of the Wnt/ß-catenin signaling is associated with tumor development, and blocking ß-catenin/BCL9 is a novel strategy for oncogenic Wnt/ß-catenin signaling. Herein, we presented two novel ß-catenin variations and exposed conformational dynamics in several ß-catenin crystal structures at the BCL9 binding site. Furthermore, we identified a class of novel urea-containing compounds targeting ß-catenin/BCL9 interaction. Notably, the binding modalities of inhibitors were greatly affected by the conformational dynamics of ß-catenin. Among them, 28 had a strong affinity for ß-catenin (Kd = 82 nM), the most potent inhibitor reported. In addition, 13 and 35 not only activate T cells but also promote the antigen presentation of cDC1, showing robust antitumor efficacy in the CT26 model. Collectively, our study demonstrated a series of potent small-molecule inhibitors targeting ß-catenin/BCL9, which can enhance antigen presentation and activate cDC1 cells, delivering a potential strategy for boosting innate and adaptive immunity to overcome immunotherapy resistance.

Synthesis and biological evaluation of 4-imidazolidinone-containing compounds as potent inhibitors of the MDM2/p53 interaction.

Inhibition of MDM2/p53 interaction with small-molecule inhibitors stabilizes p53 from MDM2 mediated degradation, which is a promising strategy for the treatment of cancer. In this report, a novel series of 4-imidazolidinone-containing compounds have been synthesized and tested in MDM2/p53 and MDM4/p53 FP binding assays. Upon SAR studies, compounds 2 (TB114) and 22 were identified as the most potent inhibitors of MDM2/p53 but not MDM4/p53 interactions. Both 2 and 22 exhibited strong antiproliferative activities in HCT-116 and MOLM-13 cell lines harboring wild type p53. Mechanistic studies show that 2 and 22 dose-dependently activated p53 and its target genes and induced apoptosis in cells based on the Western blot, qPCR, and flow cytometry assays. In addition, the antiproliferative activities of 2 and 22 were dependent on wild type p53, while they were not toxic to HEK-293 kidney cells. Furthermore, the on-target activities of 2 were general and applicable to other cancer cell lines with wild type p53. These attributes make 2 a good candidate for future optimization to discover a potential treatment of wild-type p53 cancer.

Clinical efficacy and biomarker analysis of neoadjuvant camrelizumab plus chemotherapy for early-stage triple-negative breast cancer: a experimental single-arm phase II clinical trial pilot study.

BACKGROUND: Triple-negative breast cancer (TNBC) is associated with a dismal prognosis. Immune checkpoint inhibitors have shown promising antitumor activity in neoadjuvant settings. This single-arm, phase II trial aimed to evaluate the efficacy and safety of camrelizumab plus chemotherapy as the neoadjuvant therapy (NAT) in early TNBC. METHODS: Patients received eight cycles of camrelizumab plus nonplatinum-based chemotherapy. The primary endpoint was total pathological complete response (pCR). Secondary endpoints included the breast pathological complete response (bpCR), adverse events (AEs). Multiomics biomarkers were assessed as exploratory objective. RESULTS: Twenty of 23 TNBC patients receiving NAT underwent surgery, with the total pCR rate of 65% (13/20) and bpCR rate of 70% (14/20). Grade ≥3 treatment-related AEs were observed in 14 (60.9%) patients, with the most common AE being neutropenia (65.2%). Tumor immune microenvironment was analyzed between pCR and non-pCR samples before and after the NAT. Gene expression profiling showed a higher immune infiltration in pCR patients than non-pCR patients in pre-NAT samples. Through establishment of a predictive model for the NAT efficacy, TAP1 and IRF4 were identified as the potential predictive biomarkers for response to the NAT. Gene set enrichment analysis revealed the glycolysis and hypoxia pathways were significantly activated in non-pCR patients before the NAT, and this hypoxia was aggravated after the NAT. CONCLUSION: Camrelizumab plus nonplatinum-based chemotherapy shows a promising pCR rate in early-stage TNBC, with an acceptable safety profile. TAP1 and IRF4 may serve as potential predictive biomarkers for response to the NAT. Aggravated hypoxia and activated glycolysis after the NAT may be associated with the treatment resistance.

Fermented cordyceps powder alleviates silica-induced pulmonary inflammation and fibrosis in rats by regulating the Th immune response.

BACKGROUND: Silicosis is an important occupational disease caused by inhalation of free silica and is characterized by persistent pulmonary inflammation, subsequent fibrosis and lung dysfunction. Until now, there has been no effective treatment for the disease due to the complexity of pathogenesis. Fermented cordyceps powder (FCP) has a similar effect to natural cordyceps in tonifying the lung and kidney. It has started to be used in the adjuvant treatment of silicosis. This work aimed to verify the protective effects of FCP against silicosis, and to explore the related mechanism. METHODS: Wistar rats were randomly divided into four groups including the saline-instilled group, the silica-exposed group, the silica + FCP (300 mg/kg) group and the silica + FCP (600 mg/kg) group. Silicosis rat models were constructed by intratracheal instillation of silica (50 mg). Rats in the FCP intervention groups received the corresponding dose of FCP daily by intragastric gavage. Rats were sacrificed on days 7, 28 and 56 after treatment, then samples were collected for further analysis. RESULTS: FCP intervention reduced the infiltration of inflammatory cells and the concentration of interleukin-1ß (IL-1ß), interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α) and transforming growth factor-ß1 (TGF-ß1) at days 7, 28, 56, and decreased the expression of collagen, α-smooth muscle actin (α-SMA) and fibronectin (FN) at days 28 and 56 in the lung of silicosis rats. FCP also decreased the immune response of Th1 and Th17 at days 7, 28, 56 and inhibited the enhancement of the Th2 response at day 56. CONCLUSIONS: FCP intervention could alleviate silica-induced pulmonary inflammation and fibrosis, the protective effect may be achieved by reducing Th1 and Th17 immune responses and inhibiting the enhancement of the Th2 response.

HMGB1 inhibition reduces TDI-induced occupational asthma through ROS/AMPK/autophagy pathway.

Exposure to toluene diisocyanate (TDI) can cause pulmonary diseases such as asthma. Inhibition of high mobility group box 1 protein (HMGB1) has been found to be protective against the toxic effects of TDI on human bronchial epithelial (HBE) cells. Here, we evaluated the in vivo positive roles of HMGB1 in the TDI-caused asthma mice and explored its underlying mechanisms in HBE cells. We found that suppression of HMGB1 obviously alleviated airway inflammation, airway hyperresponsiveness, and airway remodeling in the lung tissue of the asthma mice. The in vitro results showed that inhibition of HMGB1 ameliorated TDI-induced reactive oxygen species (ROS) release, inflammatory response, and activation of autophagy in HBE cells. At the molecular level, inhibition of HMGB1 decreased the expressions of HMGB1, Toll-like receptor 4, Vimentin and matrix metalloproteinase-9 proteins, activated NF-κB and NOD-like receptor protein 3 (NLRP3) inflammasome, and increased E-cadherin expression. Importantly, activation of autophagy could lead to the overactivation of NLRP3 inflammasome in TDI-induced asthma. These results suggest that inhibition of HMGB1 can alleviate TDI-induced asthma through ROS/AMPK/autophagy pathways, which may provide valuable evidence for the pathogenesis and therapeutic targets of TDI-induced asthma.

Abnormality of mussel in the early developmental stages induced by graphene and triphenyl phosphate: In silico toxicogenomic data-mining, in vivo, and toxicity pathway-oriented approach.

Increasing number of complex mixtures of organic pollutants in coastal area (especially for nanomaterials and micro/nanoplastics associated chemicals) threaten aquatic ecosystems and their joint hazards are complex and demanding tasks. Mussels are the most sensitive marine faunal groups in the world, and their early developmental stages (embryo and larvae) are particularly susceptible to environmental contaminants, which can distinguish the probable mechanisms of mixture-induced growth toxicity. In this study, the potential critical target and biological processes affected by graphene and triphenyl phosphate (TPP) were developed by mining public toxicogenomic data. And their combined toxic effects were verified by toxicological assay at early developmental stages in filter-feeding mussels (embryo and larvae). It showed that interactions among graphene/TPP with 111 genes (ABCB1, TP53, SOD, CAT, HSP, etc.) affected phenotypes along conceptual framework linking these chemicals to developmental abnormality endpoints. The PPAR signaling pathway, monocarboxylic acid metabolic process, regulation of lipid metabolic process, response to oxidative stress, and gonad development were noted as the key molecular pathways that contributed to the developmental abnormality. Enriched phenotype analysis revealed biological processes (cell proliferation, cell apoptosis, inflammatory response, response to oxidative stress, and lipid metabolism) affected by the investigated mixture. Combined, our results supported that adverse effects induced by contaminants/ mixture could not only be mediated by single receptor signaling or be predicted by the simple additive effect of contaminants. The results offer a framework for better comprehending the developmental toxicity of environmental contaminants in mussels and other invertebrate species, which have considerable potential for hazard assessment of coastal mixture.

Structure basis of two nanobodies neutralizing SARS-CoV-2 Omicron variant by targeting ultra-conservative epitopes.

The evolving SARS-CoV-2 Omicron strain has repeatedly caused widespread disease epidemics, and effective antibody drugs continue to be in short supply. Here, we identified a batch of nanobodies with high affinity for receptor binding domain (RBD) of SARS-CoV-2 spike protein, separated them into three classes using high performance liquid chromatography (HPLC), and then resolved the crystal structure of the ternary complexes of two non-competing nanobodies (NB1C6 and NB1B5) with RBD using X-ray crystallography. The structures showed that NB1B5 and NB1C6 bind to the left and right flank of the RBD, respectively, and that the binding epitopes are highly conserved cryptic sites in all SARS-CoV-2 mutant strains, as well as that NB1B5 can effectively block the ACE2. These two nanobodies were covalently linked into multivalent and bi-paratopic formats, and have a high affinity and neutralization potency for omicron, potentially inhibiting viral escape. The binding sites of these two nanobodies are relatively conserved, which help guide the structural design of antibodies targeting future variants of SARS-CoV-2 to combat COVID-19 epidemics and pandemics.

Preparation and characterization of nanobodies targeting SARS-CoV-2 papain-like protease.

Coronavirus Papain-like protease (PLpro) mediates the cleavage of viral polyproteins and assists the virus escaping from innate immune response. Thus, PLpro is an attractive target for the development of broad-spectrum drugs as it has a conserved structure across different coronaviruses. In this study, we purified SARS-CoV-2 PLpro as an immune antigen, constructed a nanobody phage display library, and identified a set of nanobodies with high affinity for SARS-CoV-2. In addition, enzyme activity experiments demonstrated that two nanobodies had a significant inhibitory effect on the PLpro. These nanobodies should therefore be investigated as candidates for the treatment of coronaviruses.

A procedure for producing an anti-AXL nanobody in E. coli.

As one of the receptors of the TAM family, AXL plays a vital role in stem cell maintenance, angiogenesis, immune escape of viruses and drug resistance against tumors. In this study, the truncated extracellular segment containing two immunoglobulin-like domains of human AXL (AXL-IG), which has been confirmed to bind growth arrest specific 6 (GAS6) by structural studies [1], was expressed in a prokaryotic expression system and then purified. Immunizing camelid with the purified AXL-IG as antigen could lead to the production of unique nanobodies composed of only variable domain of heavy chain of heavy-chain antibody (VHH), which are around 15 kD and stable. We screened out a nanobody A-LY01 specific binding to AXL-IG. We further determined the affinity of A-LY01 to AXL-IG and revealed that A-LY01 could specifically recognize full-length AXL on the surface of HEK 293T/17 cells. Our study provides appropriate support for the development of diagnostic reagents and antibody therapeutics targeting AXL.

Synthesis and Biological Evaluation of Sclareolide-Indole Conjugates and Their Derivatives.

Sclareolide is a sesquiterpene lactone isolated from various plant sources in tons every year and is commercially used as a flavor ingredient in the cosmetic and food industries. Antitumor and antiviral activities of sclareolide have been previously reported. However, biological studies of sclareolide synthetic analogous are few. In view of these, we developed a robust synthetic method that allows the assembly of 36 novel sclareolide-indole conjugates and their derivatives. The synthetic method was based on TiCl4-promoted nucleophilic substitution of sclareolide-derived hemiacetal 4, while electron-rich aryles including indoles, polyphenol ethers, and pyrazolo [1,5-a]pyridine were good substrates. The stereochemistry of the final products was confirmed by single-crystal X-ray diffraction analysis, while the antiproliferative activities of selected final products were tested in K562 and MV4-11 cancer cell lines. Cytometric flow analysis shows that lead compounds 8k- and 10-induced robust apoptosis in MV4-11 cancer cells, while they exhibited weak impact on cell cycle progression. Taken together, our study suggests that sclareolide could be a good template and substrate for the synthesis of novel antiproliferative compounds.

Particulate Matter and Its Components Induce Alteration on the T-Cell Response: A Population Biomarker Study.

Compared with the T-cell potential of particulate matter (PM) in animal studies, comprehensive evaluation on the impairments of T-cell response and exposure-response from PM and its components in human population is limited. There were 768 participants in this study. We measured environmental PM and its polycyclic aromatic hydrocarbons (PAHs) and metals and urinary metabolite levels of PAHs and metals among population. T lymphocyte and its subpopulation (CD4+ T cells and CD8+ T cells) and the expressions of T-bet, GATA3, RORγt, and FoxP3 were measured. We explored the exposure-response of PM compositions by principal component analysis and mode of action by mediation analysis. There was a significant decreasing trend for T lymphocytes and the levels of T-bet and GATA3 with increased PM levels. Generally, there was a negative correlation between PM, urinary 1-hydroxypyrene, urinary metals, and the levels of T-bet and GATA3 expression. Additionally, CD4+ T lymphocytes were found to mediate the associations of PM2.5 with T-bet expression. PM and its bound PAHs and metals could induce immune impairments by altering the T lymphocytes and genes of T-bet and GATA3.

Bile acids-gut microbiota crosstalk contributes to the improvement of type 2 diabetes mellitus.

Type 2 diabetes mellitus (T2DM) occurs that cannot effectively use the insulin. Insulin Resistance (IR) is a significant characteristic of T2DM which is also an essential treatment target in blood glucose regulation to prevent T2DM and its complications. Bile acids (BAs) are one group of bioactive metabolites synthesized from cholesterol in liver. BAs play an important role in mutualistic symbiosis between host and gut microbiota. It is shown that T2DM is associated with altered bile acid metabolism which can be regulated by gut microbiota. Simultaneously, BAs also reshape gut microbiota and improve IR and T2DM in the bidirectional communications of the gut-liver axis. This article reviewed the findings on the interaction between BAs and gut microbiota in improving T2DM, which focused on gut microbiota and its debinding function and BAs regulated gut microbiota through FXR/TGR5. Meanwhile, BAs and their derivatives that are effective for improving T2DM and other treatments based on bile acid metabolism were also summarized. This review highlighted that BAs play a critical role in the glucose metabolism and may serve as therapeutic targets in T2DM, providing a reference for discovering and screening novel therapeutic drugs.

The mitochondrial complex I proteins of Candida albicans moderate phagocytosis and the production of pro-inflammatory cytokines in murine macrophages and dendritic cells.

Loss of respiratory functions impairs Candida albicans colonization of host tissues and virulence in a murine model of candidiasis. Furthermore, it is known that respiratory inhibitors decrease mannan synthesis and glucan exposure and thereby promotes phagocytosis. To understand the impact of respiratory proteins of C. albicans on host innate immunity, we characterized cell wall defects in three mitochondrial complex I (CI) null mutants (nuo1Δ, nuo2Δ and ndh51Δ) and in one CI regulator mutant (goa1Δ), and we studied the corresponding effects of these mutants on phagocytosis, neutrophil killing and cytokine production by dendritic cells (DCs). We find that reductions of phosphopeptidomannan (PPM) in goa1Δ, nuo1Δ and phospholipomannan (PLM) in nuo2Δ lead to reductions of IL-2, IL-4, and IL-10 but increase of TNF-α in infected DCs. While PPM loss is a consequence of a reduced phospho-Cek1/2 MAPK that failed to promote phagocytosis and IL-22 production in goa1Δ and nuo1Δ, a 30% glucan reduction and a defective Mek1 MAPK response in ndh51Δ lead to only minor changes in phagocytosis and cytokine production. Glucan exposure and PLM abundance seem to remain sufficient to opsonize neutrophil killing perhaps via humoral immunity. The diversity of immune phenotypes in these mutants possessing divergent cell wall defects is further supported by their transcriptional profiles in each infected murine macrophage scenario. Since metabolic processes, oxidative stress-induced senescence, and apoptosis are differently affected in these scenarios, we speculate that during the early stages of infection, host immune cells coordinate their bioactivities based upon a mixture of signals generated during host-fungi interactions.

A chemical perspective on the modulation of TEAD transcriptional activities: Recent progress, challenges, and opportunities.

TEADs are transcription factors and core downstream components of the Hippo pathway. Mutations of the Hippo pathway and/or dysregulation of YAP/TAZ culminate in aberrant transcriptional activities of TEADs, which were considered as key contributing factors of mesotheliomas, fibrotic diseases, Alzheimer's diseases, Huntington's diseases, suppressive immune response, and drug resistance, among others. To modulate transcriptional activities of TEADs, several pharmacological approaches have been pursued, including TEAD/YAP protein-protein interaction inhibitors, TEAD PBP inhibitors, and TEAD activators. As summarized in this review, a large number of inhibitors and activators of TEADs have been reported with decent in vitro potencies, a few exerted robust and compelling in vivo efficacies, and three that are undergoing clinical trials for the treatment of human cancers. Despite clinical advancement of the TEAD PBP inhibitors, development of other types TEAD inhibitors and activators generally lags behind. Information showcased herein might benefit discovery of next generation TEAD modulators for treatment of human oncological diseases and beyond.

Individualized CT image-guided free-hand catheter technique: A new and reliable method for minimally invasive evacuation of basal ganglia hematoma.

Objective: To validate the clinical reliability of an individualized CT image-guided' free-hand catheter technique (CTGFC) for basal ganglia hematoma (BGH) evacuation. Methods: From January 2017 to December 2020, 58 cases of patients with BGH who underwent catheter evacuation were enrolled. The surgery was conducted using the CTGFC (n = 31) or stereotactic catheter technique (STC, n = 27). The authors evaluated the baseline characteristics, operation-related indicators, postoperative complications, hospitalization-related indicators, short-term and long-term functional outcomes, and mortality rate 1 year after surgery. Results: All patients underwent BGH evacuation under non-general anesthesia in the CTGFC group. The operative time (p < 0.01) and operation costs (p < 0.05) were significantly shorter in the CTGFC group than that in the STC group (p < 0.01). Comparable results were found in the catheter indwelling duration, residual hematoma volume, hematoma evacuation rate, incidence of postoperative complications, hospital ICU stay, and hospital costs between these two groups (p > 0.05). The duration of hospital stay was remarkably shorter in the CTGFC group than that in the STC group (p < 0.01). There were no differences in terms of the short-time functional outcomes score at discharge, including the Glasgow outcome scale (GOS) score, the activities of daily living (ADL) score, and the Karnofsky performance score (KPS). Moreover, comparable findings were also found in the 1-year postoperative GOS score, ADL score, KPS score, and mortality rate between these two groups. Conclusion: The simple CTGFC-assisted surgery was a safe and reliable option for BGH evacuation, especially in primary medical institutes and emergency situations with limited medical resources.

Identification of key pathways and genes in the progression of silicosis based on WGCNA.

Silicosis, induced by inhaling silica particles in workplaces, is one of the most common occupational diseases. The prognosis of silicosis and its consequent fibrosis is extremely poor due to limited treatment modalities and lack of understanding of the disease mechanisms. In this study, a Wistar rat model for silicosis fibrosis was established by intratracheal instillation of silica (0, 50, 100 and 200 mg/mL, 1 mL) with the evidence of Hematoxylin and Eosin (HE) and Masson staining and the expressions of inflammatory and fibrotic proteins of rats' lung tissues. RNA of lung tissues of rats exposed to 200 mg/mL silica particles and normal saline for 14 d and 28 d was extracted and sequenced to detect differentially expressed genes (DEGs) and to identify silicosis fibrosis-associated modules and hub genes by Weighted gene co-expression network analysis (WGCNA). Predictions of gene functions and signaling pathways were conducted using Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) databases. In this study, it has been demonstrated the promising role of the Hippo signaling pathway in silicosis fibrosis, which will be conducive to elucidating the specific mechanism of pulmonary fibrosis induced by silica and to determining molecular initiating event (MIE) and adverse outcome pathway (AOP) of silicosis fibrosis.

Structure-Based Discovery of MDM2/4 Dual Inhibitors that Exert Antitumor Activities against MDM4-Overexpressing Cancer Cells.

Despite recent clinical progress in peptide-based dual inhibitors of MDM2/4, small-molecule ones with robust antitumor activities remain challenging. To tackle this issue, 31 (YL93) was structure-based designed and synthesized, which had MDM2/4 binding Ki values of 1.1 and 642 nM, respectively. In three MDM4-overexpressing cancer cell lines harboring wild-type p53, 31 shows improved cell growth inhibition activities compared to RG7388, an MDM2-selective inhibitor in late-stage clinical trials. Mechanistic studies show that 31 increased cellular protein levels of p53 and p21 and upregulated the expression of p53-targeted genes in RKO cells with MDM4 amplification. In addition, 31 induced cell-cycle arrest and apoptosis in western blot and flow cytometry assays. Taken together, dual inhibition of MDM2/4 by 31 elicited stronger antitumor activities in vitro compared to selective MDM2 inhibitors in wild-type p53 and MDM4-overexpressing cancer cells.

Wnt signaling pathway in cancer immunotherapy.

Wnt/ß-catenin signaling is a highly conserved pathway that regulates cell proliferation, differentiation, apoptosis, stem cell self-renewal, tissue homeostasis, and wound healing. Dysregulation of the Wnt pathway is intricately involved in almost all stages of tumorigenesis in various cancers. Through direct and/or indirect effects on effector T cells, T-regulatory cells, T-helper cells, dendritic cells, and other cytokine-expressing immune cells, abnormal activation of Wnt/ß-catenin signaling benefits immune exclusion and hinders T-cell-mediated antitumor immune responses. Activation of Wnt signaling results in increased resistance to immunotherapies. In this review, we summarize the process by which Wnt signaling affects cancer and immune surveillance, and the potential for targeting the Wnt-signaling pathway via cancer immunotherapy.

Targeting Wnt Signaling in the Tumor Immune Microenvironment to Enhancing EpCAM CAR T-Cell therapy.

Colorectal cancer (CRC) patients are still lacking viable treatments. Chimeric antigen receptor (CAR) T cells have shown promise in hematologic malignancies, but their efficacy in solid tumors has been limited due to the immunosuppressive tumor microenvironment. We found that cancer antigen- EpCAM expression increased in the metastatic stage compared with the primary stage in cancers and the activation of Wnt and TGFß pathways was positively correlated with EpCAM expression in multiple cancers, including colorectal cancer. We constructed CAR T cells targeting EpCAM that successfully showed selective cytotoxicity in highly EpCAM-expressing cancer cell lines. The combination of EpCAM CAR-T with the Wnt inhibitor-hsBCL9CT-24 displayed synergetic effect against EpCAM-positive colon cells in vitro and also in vivo. A mechanistic study showed that hsBCL9CT-24 treatment could modulate the tumor environment and improve infiltration of T cells, while possibly promoting the effector T cells at the early stages and postponing the exhaustion of CAR T cells at advanced stages. Overall, these results demonstrated that the combination of EpCAM CAR T-cell therapy with the Wnt inhibitor can overcome the limitations of CAR T cells in treating solid tumors.

Graphene-triphenyl phosphate (TPP) co-exposure in the marine environment: Interference with metabolism and immune regulation in mussel Mytilus galloprovincialis.

Both immune regulation and endocrine systems are great challenges to marine organisms, and effective protocols for determining these adverse outcome pathways are limited, especially in vivo. The increasing usage of graphene nanomaterials can lead to the frequent exposure to marine organisms. Triphenyl phosphate (TPP), an organophosphate flame retardant, is frequently detected in natural environments. In this study, the combined toxic effects of co-exposure to graphene and TPP was investigated in Mytilus galloprovincialis using computational toxicology and multi-omics technology. Noticeably, graphene could disturb the membrane stability and increase the tissue accumulation of TPP. The adsorption behavior of TPP on graphene could inhibit the surface activity of graphene. In the digestive gland, transcriptomics analysis revealed the down-regulated genes in graphene + TPP treatment, including glyceraldehyde-3-phosphate dehydrogenase (GAPDH), sorbitol dehydrogenase (SORD), glutathione s-transferase mu 3 (GSTM3) and 4-aminobutyrate aminotransferase (ABAT), were mainly associated with oxidative stress and energy metabolism. Moreover, metabolic responses indicated that graphene + TPP could cause disturbances in energy metabolism and osmotic regulation marked by differentially altered ATP, glucose and taurine in mussels. These data underline the need for further knowledge on the potential interactions of nanomaterials with existing contaminants in marine organisms.