ATP-binding Cassette Subfamily Research Articles

Identification of Hyperuricemia Alleviating Peptides from Yellow Tuna Thunnus albacares.

The development of food-derived antihyperuricemic substances is important for alleviating hyperuricemia (HUA) and associated inflammation. Here, novel peptides fromThunnus albacares (TAP) with strong antihyperuricemic activity were prepared. TAP was prepared by alkaline protease (molecular weight <1000 Da), with an IC50 value of xanthine oxidase inhibitory activity of 2.498 mg/mL, and 5 mg/mL TAP could reduce uric acid (UA) by 33.62% in human kidney-2 (HK-2) cells (P < 0.01). Mice were fed a high-purine diet and injected with potassium oxonate to induce HUA. Oral administration of TAP (600 mg/kg/d) reduced serum UA significantly by 42.22% and increased urine UA by 79.02% (P < 0.01) via regulating urate transporters GLUT9, organic anion transporter 1, and ATP-binding cassette subfamily G2. Meantime, TAP exhibited hepatoprotective and nephroprotective effects, according to histological analysis. Besides, HUA mice treated with TAP showed anti-inflammatory activity by decreasing the levels of toll-like receptor 4, nuclear factors-κB p65, NLRP3, ASC, and Caspase-1 in the kidneys (P < 0.01). According to serum non-targeted metabolomics, 91 differential metabolites between the MC and TAP groups were identified, and purine metabolism was considered to be the main pathway for TAP alleviating HUA. In a word, TAP exhibited strong antihyperuricemic activity both in vitro and in vivo.

Retinoic acid related orphan receptor α is a genetic modifier that rescues retinal degeneration in a mouse model of Stargardt disease and Dry AMD.

Degeneration of the macula is associated with several overlapping diseases including age-related macular degeneration (AMD) and Stargardt Disease (STGD). Mutations in ATP Binding Cassette Subfamily A Member 4 (ABCA4) are associated with late-onset dry AMD and early-onset STGD. Additionally, both forms of macular degeneration exhibit deposition of subretinal material and photoreceptor degeneration. Retinoic acid related orphan receptor α (RORA) regulates the AMD inflammation pathway that includes ABCA4, CD59, C3 and C5. In this translational study, we examined the efficacy of RORA at attenuating retinal degeneration and improving the inflammatory response in Abca4 knockout (Abca4-/-) mice. AAV5-hRORA-treated mice showed reduced deposits, restored CD59 expression and attenuated amyloid precursor protein (APP) expression compared with untreated eyes. This molecular rescue correlated with statistically significant improvement in photoreceptor function. This is the first study evaluating the impact of RORA modifier gene therapy on rescuing retinal degeneration. Our studies demonstrate efficacy of RORA in improving STGD and dry AMD-like disease.

Alkaliptosis induction counteracts paclitaxel-resistant ovarian cancer cells via ATP6V0D1-mediated ABCB1 inhibition.

Paclitaxel serves as the cornerstone chemotherapy for ovarian cancer, yet its prolonged administration frequently culminates in drug resistance, presenting a substantial challenge. Here we reported that inducing alkaliptosis, rather than apoptosis or ferroptosis, effectively overcomes paclitaxel resistance. Mechanistically, ATPase H+ transporting V0 subunit D1 (ATP6V0D1), a key regulator of alkaliptosis, plays a pivotal role by mediating the downregulation of ATP-binding cassette subfamily B member 1 (ABCB1), a multidrug resistance protein. Both ATP6V0D1 overexpression through gene transfection and pharmacological enhancement of ATP6V0D1 protein stability using JTC801 effectively inhibit ABCB1 upregulation, resulting in growth inhibition in drug-resistant cells. Additionally, increasing intracellular pH to alkaline (pH 8.5) via sodium hydroxide application suppresses ABCB1 expression, whereas reducing the pH to acidic conditions (pH 6.5) with hydrochloric acid amplifies ABCB1 expression in drug-resistant cells. Collectively, these results indicate a potentially effective therapeutic strategy for targeting paclitaxel-resistant ovarian cancer by inducing ATP6V0D1-dependent alkaliptosis.

ABCB1-mediated docetaxel resistance reversed by erastin in prostate cancer.

Docetaxel (Doc) currently serves as the primary first-line treatment for patients with castrate-resistant prostate cancer (CRPC). Erastin, a small molecule compound, can trigger inhibition of the cystine-glutamate reverse transport system and other pathways, leading to iron-dependent cell death (ferroptosis). Beyond its role in inducing cancer cell death, erastin demonstrates potential when combined with chemotherapy drugs to heighten cancer cell drug susceptibility. However, the augmentation by erastin of the effects of Doc treatment on prostate cancer, and the underlying mechanisms involved, remain unclear. In the present study, we determined the role and the underlying molecular mechanism of erastin against CRPC. The results showed that CRPC cell lines were resistant to Doc, and the expression of ferroptosis-related factors in drug-resistant cell lines was downregulated. Erastin, in synergy with Doc, exerts a pro-apoptotic effect. Erastin significantly inhibited the activity of ATP-binding cassette subfamily B member 1 (ABCB1) but did not change its protein expression and localization. Finally, in mice, erastin treatment dramatically reduced tumor growth in vivo. Taken together, our findings demonstrate that erastin enhances Doc-induced apoptosis to a certain extent and reverses Doc resistance in prostate cancer by inhibiting the activity of multidrug-resistant protein ABCB1.

Suppression of lysosome metabolism-meditated GARP/TGF-β1 complexes specifically depletes regulatory T cells to inhibit breast cancer metastasis.

Regulatory T cells (Tregs) prevent autoimmunity and contribute to cancer progression. They exert contact-dependent inhibition of immune cells through the production of active transforming growth factor-β1 (TGF-β1). However, the absence of a specific surface marker makes inhibiting the production of active TGF-β1 to specifically deplete human Tregs but not other cell types a challenge. TGF-β1 in an inactive form binds to Tregs membrane protein Glycoprotein A Repetitions Predominant (GARP) and then activates it via an unknown mechanism. Here, we demonstrated that tumour necrosis factor receptor-associated factor 3 interacting protein 3 (TRAF3IP3) in the Treg lysosome is involved in this activation mechanism. Using a novel naphthalenelactam-platinum-based anticancer drug (NPt), we developed a new synergistic effect by suppressing ATP-binding cassette subfamily B member 9 (ABCB9) and TRAF3IP3-mediated divergent lysosomal metabolic programs in tumors and human Tregs to block the production of active GARP/TGF-β1 for remodeling the tumor microenvironment. Mechanistically, NPt is stored in Treg lysosome to inhibit TRAF3IP3-meditated GARP/TGF-β1 complex activation to specifically deplete Tregs. In addition, by promoting the expression of ABCB9 in lysosome membrane, NPt inhibits SARA/p-SMAD2/3 through CHRD-induced TGF-β1 signaling pathway. In addition to expose a previously undefined divergent lysosomal metabolic program-meditated GARP/TGF-β1 complex blockade by exploring the inherent metabolic plasticity, NPt may serve as a therapeutic tool to boost unrecognized Treg-based immune responses to infection or cancer via a mechanism distinct from traditional platinum drugs and currently available immune-modulatory antibodies.

Utilizing Pharmacogenomic Data for a Safer Use of Statins among the Emirati Population.

Statins are the most prescribed lipid-lowering drugs worldwide. The associated adverse events, especially muscle symptoms, have been frequently reported despite their perceived safety. Three pharmacogenes, the solute carrier organic anion transporter family member 1B1 (SLCO1B1), ATP-binding cassette subfamily G member 2 (ABCG2), and cytochrome P450 2C9 (CYP2C9) are suggested as safety biomarkers for statins. The Clinical Pharmacogenomic Implementation Consortium (CPIC) issued clinical guidelines for statin use based on these three genes. The present study aimed to examine variants in these pharmacogenes to predict the safety of statin use among the Emirati population. Analyzing 242 whole exome sequencing data at the three genes enabled the determination of the frequencies of the single nucleotide polymorphisms (SNPs), annotating the haplotypes and the predicted functions of their proteins. In our cohort, 29.8% and 5.4% had SLCO1B1 decreased and poor function, respectively. The high frequency warns of the possibility of significant side effects of some statins and the importance of pharmacogenomic testing. We found a low frequency (6%) of the ABCG2:rs2231142 variant, which indicates the low probability of Emirati patients being recommended against higher rosuvastatin doses compared with other populations with higher frequencies of this variant. In contrast, we found high frequencies of the functionally impaired CYP2C9 alleles, which makes fluvastatin a less favorable choice. Among the sparse studies available, the present one demonstrates all SLCO1B1 and CYP2C9 function-impairing alleles among Emiratis. We highlighted how population-specific pharmacogenomic data can predict safer choices of statins, especially in understudied populations.

ABCG2-Expressing Clonal Repopulating Endothelial Cells Serve to Form and Maintain Blood Vessels.

Most organs are maintained lifelong by resident stem/progenitor cells. During development and regeneration, lineage-specific stem/progenitor cells can contribute to the growth or maintenance of different organs, whereas fully differentiated mature cells have less regenerative potential. However, it is unclear whether vascular endothelial cells (ECs) are also replenished by stem/progenitor cells with EC-repopulating potential residing in blood vessels. It has been reported recently that some EC populations possess higher clonal proliferative potential and vessel-forming capacity compared with mature ECs. Nevertheless, a marker to identify vascular clonal repopulating ECs (CRECs) in murine and human individuals is lacking, and, hence, the mechanism for the proliferative, self-renewal, and vessel-forming potential of CRECs is elusive. We analyzed colony-forming, self-renewal, and vessel-forming potential of ABCG2 (ATP binding cassette subfamily G member 2)-expressing ECs in human umbilical vessels. To study the contribution of Abcg2-expressing ECs to vessel development and regeneration, we developed Abcg2CreErt2;ROSA TdTomato mice and performed lineage tracing during mouse development and during tissue regeneration after myocardial infarction injury. RNA sequencing and chromatin methylation chromatin immunoprecipitation followed by sequencing were conducted to study the gene regulation in Abcg2-expressing ECs. In human and mouse vessels, ECs with higher ABCG2 expression (ABCECs) possess higher clonal proliferative potential and in vivo vessel-forming potential compared with mature ECs. These cells could clonally contribute to vessel formation in primary and secondary recipients after transplantation. These features of ABCECs meet the criteria of CRECs. Results from lineage tracing experiments confirm that Abcg2-expressing CRECs (AbcCRECs) contribute to arteries, veins, and capillaries in cardiac tissue development and vascular tissue regeneration after myocardial infarction. Transcriptome and epigenetic analyses reveal that a gene expression signature involved in angiogenesis and vessel development is enriched in AbcCRECs. In addition, various angiogenic genes, such as Notch2 and Hey2, are bivalently modified by trimethylation at the 4th and 27th lysine residue of histone H3 (H3K4me3 and H3K27me3) in AbcCRECs. These results are the first to establish that a single prospective marker identifies CRECs in mice and human individuals, which holds promise to provide new cell therapies for repair of damaged vessels in patients with endothelial dysfunction.

Histological Findings in the Eyes of Abcc6 Knockout Rat Model of Pseudoxanthoma Elasticum.

To describe the ocular findings of murine pseudoxanthoma elasticum (PXE) models with ATP-binding cassette subfamily C member 6 (Abcc6) gene knockout. This experiment was conducted in four Abcc6-/- rats and compared with six wild-type Abcc6+/+ control rats. The animals underwent necropsy at 6 months of age. Histological examination of the eyes was performed. Histological examination of eight eyes from four Abcc6-/- rats revealed multiple nodular foci of calcification in the uvea, sclera, and conjunctiva, focally in perivascular distribution, as well as linear and nodular calcification of Bruch's membrane. Calcific foci were not associated with inflammation in the knockout rats. There was no evidence of calcification in control eyes. The Abcc6-/- rat model shows that PXE can affect multiple ocular tissues beyond the calcification in Bruch's membrane noted in human eyes. Nodular calcific foci probably correspond to comet lesions seen in patients with PXE. The presence of ectopic calcium without inflammation distinguishes it from inflammatory calcium deposition in atherosclerosis. Further studies are needed to determine why PXE does not cause inflammatory infiltration. The Abcc6-/- murine model may be suitable for studying ocular PXE pathophysiology and ectopic calcification and developing effective therapies.

Dan-shen Yin promotes bile acid metabolism and excretion to prevent atherosclerosis via activating FXR/BSEP signaling pathway

Ethnopharmacological relevanceDan-shen Yin (DSY), a traditional prescription, has been demonstrated to be effective in decreasing hyperlipidemia and preventing atherosclerosis (AS), but its mechanism remains unknown. We hypothesized that DSY activates farnesoid X receptor (FXR) to promote bile acid metabolism and excretion, thereby alleviating AS. Aim of the studyThis study was designed to explore whether DSY reduces liver lipid accumulation and prevents AS by activating FXR and increasing cholesterol metabolism and bile acid excretion. Materials and methodsThe comprehensive chemical characterization of DSY was analyzed by UHPLC-MS/MS. The AS models of ApoE−/− mice and SD rats was established by high-fat diet and high-fat diet combined with intraperitoneal injection of vitamin D3, respectively. The aortic plaque and pathological changes were used to evaluate AS. Lipid levels, H&E staining and oil red O staining were used to evaluate liver lipid accumulation. The cholesterol metabolism and bile acid excretion were evaluated by enzyme-linked immunosorbent assay, UPLC-QQQ/MS. In vitro, the lipid and FXR/bile salt export pump (BSEP) levels were evaluated by oil red O staining, real-time quantitative polymerase chain reaction (RT-qPCR) and western blotting. ResultsA total of 36 ingredients in DSY were identified by UPLC-MS/MS analysis. In vivo, high-dose DSY significantly inhibited aortic intimal thickening, improved arrangement disorder, tortuosity, and rupture of elastic fibers, decreased lipid levels, and reduced the number of fat vacuoles and lipid droplets in liver tissue in SD rats and ApoE−/− mice. Further studies found that high-dose DSY significantly reduced liver lipid and total bile acids levels, increased liver ursodeoxycholic acid (UDCA) and other non-conjugated bile acids levels, increased fecal total cholesterol (TC) levels, and augmented FXR, BSEP, cholesterol 7-alpha hydroxylase (CYP7A1), ATP binding cassette subfamily G5/G8 (ABCG5/8) expression levels, while decreasing ASBT expression levels. In vitro studies showed that DSY significantly reduced TC and TG levels, as well as lipid droplets, while also increasing the expression of ABCG5/8, FXR, and BSEP in both HepG2 and Nr1h4 knockdown HepG2 cells. ConclusionThis study demonstrated that DSY promotes bile acid metabolism and excretion to prevent AS by activating FXR. For the prevent of AS and drug discovery provided experimental basis.

SMARCA4 (BRG1) activates ABCC3 transcription to promote hepatocellular carcinogenesis

AimsHepatocellular carcinoma (HCC) is a lead cause of cancer-related deaths. In the present study we investigated the role of Brahma-related gene 1 (BRG1), a chromatin remodeling protein, in HCC the pathogenesis focusing on identifying novel transcription targets. Methods and materialsHepatocellular carcinogenesis was modeled in mice by diethylnitrosamine (DEN). Cellular transcriptome was evaluated by RNA-seq. ResultsHepatocellular carcinoma was appreciably retarded in BRG1 knockout mice compared to wild type littermates. Transcriptomic analysis identified ATP Binding Cassette Subfamily C Member 3 (ABCC3) as a novel target of BRG1. BRG1 over-expression in BRG1low HCC cells (HEP1) up-regulated whereas BRG1 depletion in BRG1high HCC cells (SNU387) down-regulated ABCC3 expression. Importantly, BRG1 was detected to directly bind to the ABCC3 promoter to activate ABCC3 transcription. BRG1 over-expression in HEP1 cells promoted proliferation and migration, both of which were abrogated by ABCC3 silencing. On the contrary, BRG1 depletion in SNU387 cells decelerated proliferation and migration, both of which were rescued by ABCC3 over-expression. Importantly, high BRG1/ABCC3 expression predicted poor prognosis in HCC patients. Mechanistically, ABCC3 regulated hepatocellular carcinogenesis possibly by influencing lysosomal homeostasis. SignificanceIn conclusion, our data suggest that targeting BRG1 and its downstream target ABCC3 can be considered as a reasonable approach for the intervention of hepatocellular carcinoma.

The use of human iPSC-derived alveolar organoids to explore SARS-CoV-2 variant infections and host responses.

Severe acute respiratory syndrome-relatedcoronavirus 2 (SARS-CoV-2) primarily targets the respiratory system. Physiologically relevant human lung models are indispensable to investigate virus-induced host response and disease pathogenesis. In this study, we generated human induced pluripotent stem cell (iPSC)-derived alveolar organoids (AOs) using an established protocol that recapitulates the sequential steps of in vivo lung development. AOs express alveolar epithelial type II cell protein markers including pro-surfactant protein C and ATP binding cassette subfamily A member 3. Compared to primary human alveolar type II cells, AOs expressed higher mRNA levels of SARS-CoV-2 entry factors, angiotensin-converting enzyme 2 (ACE2), asialoglycoprotein receptor 1 (ASGR1) and basigin (CD147). Considering the localization of ACE2 on the apical side in AOs, we used three AO models, apical-in, sheared and apical-out for SARS-CoV-2 infection. All three models of AOs were robustly infected with the SARS-CoV-2 irrespective of ACE2 accessibility. Antibody blocking experiment revealed that ASGR1 was the main receptor for SARS-CoV2 entry from the basolateral in apical-in AOs. AOs supported the replication of SARS-CoV-2 variants WA1, Alpha, Beta, Delta, and Zeta and Omicron to a variable degree with WA1 being the highest and Omicron being the least. Transcriptomic profiling of infected AOs revealed the induction of inflammatory and interferon-related pathways with NF-κB signaling being the predominant host response. In summary, iPSC-derived AOs can serve as excellent human lung models to investigate infection of SARS-CoV-2 variants and host responses from both apical and basolateral sides.

Development of an orally bioavailable mSWI/SNF ATPase degrader and acquired mechanisms of resistance in prostate cancer

Mammalian switch/sucrose nonfermentable (mSWI/SNF) ATPase degraders have been shown to be effective in enhancer-driven cancers by functioning to impede oncogenic transcription factor chromatin accessibility. Here, we developed AU-24118, an orally bioavailable proteolysis-targeting chimera (PROTAC) degrader of mSWI/SNF ATPases (SMARCA2 and SMARCA4) and PBRM1. AU-24118 demonstrated tumor regression in a model of castration-resistant prostate cancer (CRPC) which was further enhanced with combination enzalutamide treatment, a standard of care androgen receptor (AR) antagonist used in CRPC patients. Importantly, AU-24118 exhibited favorable pharmacokinetic profiles in preclinical analyses in mice and rats, and further toxicity testing in mice showed a favorable safety profile. As acquired resistance is common with targeted cancer therapeutics, experiments were designed to explore potential mechanisms of resistance that may arise with long-term mSWI/SNF ATPase PROTAC treatment. Prostate cancer cell lines exposed to long-term treatment with high doses of a mSWI/SNF ATPase degrader developed SMARCA4 bromodomain mutations and ABCB1 (ATP binding cassette subfamily B member 1) overexpression as acquired mechanisms of resistance. Intriguingly, while SMARCA4 mutations provided specific resistance to mSWI/SNF degraders, ABCB1 overexpression provided broader resistance to other potent PROTAC degraders targeting bromodomain-containing protein 4 and AR. The ABCB1 inhibitor, zosuquidar, reversed resistance to all three PROTAC degraders tested. Combined, these findings position mSWI/SNF degraders for clinical translation for patients with enhancer-driven cancers and define strategies to overcome resistance mechanisms that may arise.

Development of an efficient liposomal DOX delivery formulation for HCC therapy by targeting CK2α.

Hepatocellular carcinoma (HCC) is a digestive tract cancer with high mortality and poor prognosis, especially in China. Current chemotherapeutic drugs lead to poor prognosis, low efficacy, and high side effects due to weak targeting specificity and rapidly formed multidrug resistance (MDR). Based on the previous studies on the doxorubicin (DOX) formulation for cancer targeting therapy, we developed a novel DOX delivery formulation for the targeting chemotherapy of HCC and DOX resistant HCC. HCSP4 was previously screened and casein kinase 2α (CK2α) was predicted as its specific target on HCC cells in our lab. In the study, miR125a-5p was firstly predicted as an MDR inhibiting miRNA, and then CK2α was validated as the target of HCSP4 and miR125a-5p using CK2α-/-HepG2 cells. Based on the above, an HCC targeting and MDR inhibiting DOX delivery liposomal formulation, HCSP4/Lipo-DOX/miR125a-5p was synthesized and tested for its HCC therapeutic efficacy in vitro. The results showed that the liposomal DOX delivery formulation targeted to HCC cells specifically and sensitively, and presented the satisfied therapeutic efficacy for HCC, particularly for DOX resistant HCC. The potential therapeutic mechanism of the DOX delivery formulation was explored, and the formulation inhibited the expression of MDR-relevant genes including ATP-binding cassette subfamily B member 1 (ABCB1, also known as P-glycoprotein), ATP-binding cassette subfamily C member 5 (ABCC5), enhancer of zeste homolog 2 (EZH2), and ATPase Na+/K+ transporting subunit beta 1 (ATP1B1). Our study presents a novel targeting chemotherapeutic drug formulation for the therapy of HCC, especially for drug resistant HCC, although it is primarily and needs further study in vivo, but provided a new strategy for the development of novel anticancer drugs.

Analyzing chemical composition of Sargentodoxae caulis water extract and their hypouricemia effect in hyperuricemic mice

Hyperuricemia (HUA) is a metabolic disease characterized by the increase of serum uric acid (UA) level. Sargentodoxae Caulis (SC) is a commonly used herbal medicine for the treatment of gouty arthritis, traumatic swelling, and rheumatic arthritis in clinic. In this study, a total of fifteen compounds were identified in SC water extract using UHPLC-Q-TOF-MS/MS, including three phenolic acids, seven phenolic glycosides, four organic acids, and one lignan. Then, to study the hypouricemia effect of SC, a HUA mouse model was induced using a combination of PO, HX, and 20% yeast feed. After 14 days of treatment with the SC water extract, the levels of serum UA, creatinine (CRE), blood urea nitrogen (BUN) were reduced significantly, and the organ indexes were restored, the xanthine oxidase (XOD) activity were inhibited as well. Meanwhile, SC water extract could ameliorate the pathological status of kidneys and intestine of HUA mice. Additionally, quantitative real-time PCR (qRT-PCR) and western blotting results showed that SC water extract could increase the expression of ATP binding cassette subfamily G member 2 (ABCG2), organic cation transporter 1 (OCT1), organic anion transporter 1 (OAT1) and organic anion transporter 3 (OAT3), whereas decrease the expression of glucose transporter 9 (GLUT9). This study provided a data support for the clinical application of SC in the treatment of HUA.

Proteomic study of left ventricle and cortex in rats after myocardial infarction

Myocardial infarction (MI) induces neuroinflammation indirectly, chronic neuroinflammation may cause neurodegenerative diseases. Changes in the proteomics of heart and brain tissue after MI may shed new light on the mechanisms involved in neuroinflammation. This study explored brain and heart protein changes after MI with a data-independent acquisition (DIA) mode proteomics approach. Permanent ligation of the left anterior descending coronary artery (LAD) was performed in the heart of rats, and the immunofluorescence of microglia in the brain cortex was performed at 1d, 3d, 5d, and 7d after MI to detect the neuroinflammation. Then proteomics was accomplished to obtain the vital proteins in the heart and brain post-MI. The results show that the number of microglia was significantly increased in the Model-1d group, the Model-3d group, the Model-5d group, and the Model-7d group compared to the Sham group. Various proteins were obtained through DIA proteomics. Linking to key targets of brain disease, 14 proteins were obtained in the brain cortex. Among them, elongation of very long chain fatty acids protein 5 (ELOVL5) and ATP-binding cassette subfamily G member 4 (ABCG4) were verified through western blotting (WB). The results of WB were consistent with the proteomics results. Therefore, these proteins may be related to the pathogenesis of neuroinflammation after MI.

Abstract 4232: Exploring chemotherapy resistance in pancreatic cancer: Insights from a 3D organoid model

Abstract Pancreatic cancer is marked by a dismal prognosis, with a 5-year survival rate at approximately 10%. A key contributor to its lethality is the development of chemotherapy resistance, followed by early metastasis. However, studies elucidating the mechanisms behind chemotherapy resistance in pancreatic cancer are limited. Previous research has relied on 2D cell line models, which present limitations, such as uniform cell types that fail to capture tumor heterogeneity and an inaccurate representation of clinical realities, wherein resistance often develops gradually rather than through continuous exposure to high drug concentrations. In this study, we developed a more sophisticated organoid model of chemotherapy resistance. Six organoids from three patients were matched to study chemotherapy resistance in a personalized context. Comprehensive whole-exome sequencing and transcriptomic analyses were performed, along with an examination of chemotherapy resistance protein expression and EMT-related markers. Notably, differential expression levels of CK19 and Vimentin were observed between samples. Our analysis also focused on the expression of the ATP-Binding Cassette Subfamily G Member 2 (ABCG2) gene in chemoresistant organoids. Western blot analysis indicated a significant increase in ABCG2 expression in these organoids compared to controls. According to the Moffitt RNA signature, the primary and corresponding chemoresistant organoids retained their molecular subtype classifications both before and after developing chemoresistance. Gene set enrichment analysis uncovered significant upregulation of pathways related to DNA repair and cell cycle regulation, particularly the G2M checkpoint, in chemoresistant organoids. Furthermore, gene ontology functional analysis suggested a notable overrepresentation of activities related to amino acid transmembrane transport and extracellular matrix constituents, underscoring their potential role in the chemoresistance phenotype. Citation Format: Hee Seung Lee. Exploring chemotherapy resistance in pancreatic cancer: Insights from a 3D organoid model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4232.

Exploring the effects of calycosin on anthracycline-induced cardiotoxicity: a network pharmacology, molecular docking, and experimental study.

Chemotherapy with anthracyclines can cause cardiotoxicity, possibly leading to stopping treatment in some cancer patients. In cardio-oncology research, preventing and minimizing anthracycline-induced cardiotoxicity (AIC) is a hot issue. For the treatment of AIC, calycosin (CA), an isoflavone component in astragali radix (AR), has become a research focus. However, the elaborate mechanisms of calycosin treating AIC remain to be unrevealed. To explore the effects of CA on AIC through multiple dimensions concerning network pharmacology, molecular docking, and experimental evaluations. The study evaluated calycosin's potential targets and mechanisms for treating AIC using network pharmacology and molecular docking. The candidate genes/targets of CA and AIC were screened using the online-available database. Protein-protein interactions (PPI) between the common targets were constructed using the STRING platform, and the results were then visualized using Cytoscape. Molecular docking was used to evaluate the strength of the binding force between CA and the common targets. The possible pharmacological mechanisms of CA were explained by pathway enrichment and GSEA. Subsequently, the candidate targets were identified in vitro experiments. Network pharmacology effectively discovered the CA's multitarget intervention in AIC, including TNF, ABCC1, TOP2A, ABCB1, and XDH. CA binds to the ATP-binding cassette subfamily B member 1(ABCB1) had the highest binding energy (-7.5 kcal/mol) according to the molecular docking analysis and was selected and visualized for subsequent analysis. In vitro experiments showed that ABCB1 exhibited significant time-curve changes under different doses of doxorubicin (DOX) compared with DMSO control experiments. The anti-AIC pharmacological mechanism of CA were revealed by highlighting the biological processes of oxidative stress (OR) and inflammation. We employed a practicable bioinformatics method to connect network and molecular docking to determine the calycosin's therapeutic mechanism against AIC and identified some bioinformatics results in in vitro experiments. The results presented show that CA may represent an encouraging treatment for AIC.

Aging and brain free cholesterol concentration on amyloid-β peptide accumulation in guinea pigs.

Alzheimer's disease is a complex multifactorial neurodegenerative disorder wherein age is a major risk factor. The appropriateness of the Hartley guinea pig (GP), which displays high sequence homologies of its amyloid-β (Aβ40 and Aβ42) peptides, Mdr1 and APP(amyloid precursor protein) and similarity in lipid handling to humans, was appraised among 9-40weeks old guinea pigs. Protein expression levels of P-gp (Abcb1) and Cyp46a1 (24(S)-hydroxylase) for Aβ40, and Aβ42 efflux and cholesterol metabolism, respectively, were decreased with age, whereas those for Lrp1 (low-density lipoprotein receptor related protein 1), Rage (receptor for advanced glycation endproducts) for Aβ efflux and influx, respectively, and Abca1 (the ATP binding cassette subfamily A member 1) for cholesterol efflux, were unchanged among the ages examined. There was a strong, negative correlation of the brain Aβ peptide concentrations and Abca1 protein expression levels with free cholesterol. The correlation of Aβ peptide concentrations with Cyp46a1 was, however, not significant, and concentrations of the 24(S)-hydroxycholesterol metabolite revealed a decreasing trend from 20weeks old toward 40weeks old guinea pigs. The composite data suggest a role for free cholesterol on brain Aβ accumulation. The decreases in P-gp and Lrp1 protein levels should further exacerbate the accumulation of Aβ peptides in guinea pig brain.

Cholangiocyte organoids to study drug-induced injury.

Drug induced bile duct injury is a frequently observed clinical problem leading to a wide range of pathological features. During the past decades, several agents have been identified with various postulated mechanisms of bile duct damage, however, mostly still poorly understood. Here, we investigated the mechanisms of chlorpromazine (CPZ) induced bile duct injury using advanced in vitro cholangiocyte cultures. Intrahepatic cholangiocyte organoids (ICOs) were driven into mature cholangiocyte like cells (CLCs), which were exposed to CPZ under cholestatic or non-cholestatic conditions through the addition of a bile acid cocktail. CPZ caused loss of monolayer integrity by reducing expression levels of tight junction protein 1 (TJP1), E-cadherin 1 (CDH1) and lysyl oxidase homolog 2 (LOXL2). Loss of zonula occuludens-1 (ZO-1) and E-cadherin was confirmed by immunostaining after exposure to CPZ and rhodamine-123 leakage further confirmed disruption of the cholangiocyte barrier function. Furthermore, oxidative stress seemed to play a major role in the early damage response by CPZ. The drug also decreased expression of three main basolateral bile acid transporters, ABCC3 (ATP binding cassette subfamily C member 3), SLC51A/B (solute carrier family 51 subunit alpha/beta) and multidrug resistance transporter ABCB1 (ATP binding cassette subfamily B member 1), thereby contributing to bile acid accumulation. CPZ did not induce an inflammatory response by itself, but addition of TNFα revealed a synergistic effect. These results show that ICOs present a model to identify toxic drugs affecting the bile ducts while providing mechanistic insights into hepatotoxicity.

Xanthoceras sorbifolium leaves alleviate hyperuricemic nephropathy by inhibiting the PI3K/AKT signaling pathway to regulate uric acid transport

Ethnopharmacological relevanceIn China, Xanthoceras sorbifolium Bunge was first documented as “Wen Guan Hua” in the “Jiu Huang Ben Cao” in 1406 A.D. According to the “National Compilation of Chinese Herbal Medicine,” X. sorbifolium leaves are sweet and flat in nature and can dispel wind and dampness, suggesting that their extract can be used to treat rheumatoid arthritis. X. sorbifolium Bunge has also been used to treat arteriosclerosis, hyperlipidemia, hypertension, chronic hepatitis, and rheumatism, complications associated with hyperuricemic nephropathy (HN), a condition characterized by kidney damage resulting from high levels of uric acid (UA) in the blood. Aim of the studyThe purpose of this study was to investigate the effects and underlying mechanisms of a 70% ethanol extract from X. sorbifolium leaves (EX) in alleviating HN. Materials and methodsA mouse model of hyperuricemia was established to initially evaluate the hypouricemic effects and determine the effective dose of EX. Phytochemical analyses were conducted using ultra high-performance liquid chromatography and liquid chromatography-mass spectroscopy. The potential key pathways of EX in the alleviation of HN were inferred using network pharmacology and bioinformatics. An HN rat model was then established, and experiments including biomarker detection, western blotting, reverse transcription quantitative polymerase chain reaction, immunohistochemical and Masson's trichrome staining, and transmission electron microscopy were conducted to evaluate the effect of EX on UA transporter expression in vitro. ResultsNetwork pharmacology and bioinformatics analyses revealed that the phosphoinositide 3-kinase (PI3K)/AKT signaling pathway was the key pathway for the alleviation of HN progression by EX. EX treatment reduced serum biomarkers in HN rats, downregulated the expression of p-PI3K, p-AKT, glucose transporter 9 (GLUT9), urate transporter 1 (URAT1), Collagen I, matrix metalloproteinase (MMP)-2, and MMP-9, and upregulated the expression of ATP binding cassette subfamily G member 2 (ABCG2) to improve renal interstitial fibrosis in HN rats. A high content of both quercitrin and cynaroside were identified in EX; their administration inhibited the increased expression of GLUT9 and URAT1 in damaged HK-2 cells. ConclusionOur study provides evidence that EX alleviates HN. The potential mechanism underlying this effect may be the regulation of UA transporters, such as GLUT9 and URAT1, by limiting the activation of the PI3K/AKT signaling pathway to improve renal injury.