E3 ubiquitin ligase UBR5 modulates circadian rhythm by facilitating the ubiquitination and degradation of the key clock transcription factor BMAL1.

The circadian clock is the inner rhythm of life activities and is controlled by a self-sustained and endogenous molecular clock, which maintains a ~ 24 h internal oscillation. As the core element of the circadian clock, BMAL1 is susceptible to degradation through the ubiquitin-proteasome system (UPS). Nevertheless, scant information is available regarding the UPS enzymes that intricately modulate both the stability and transcriptional activity of BMAL1, affecting the cellular circadian rhythm. In this work, we identify and validate UBR5 as a new E3 ubiquitin ligase that interacts with BMAL1 by using affinity purification, mass spectrometry, and biochemical experiments. UBR5 overexpression induced BMAL1 ubiquitination, leading to diminished stability and reduced protein level of BMAL1, thereby attenuating its transcriptional activity. Consistent with this, UBR5 knockdown increases the BMAL1 protein. Domain mapping discloses that the C-terminus of BMAL1 interacts with the N-terminal domains of UBR5. Similarly, cell-line-based experiments discover that HYD, the UBR5 homolog in Drosophila, could interact with and downregulate CYCLE, the BMAL1 homolog in Drosophila. PER2-luciferase bioluminescence real-time reporting assay in a mammalian cell line and behavioral experiments in Drosophila reveal that UBR5 or hyd knockdown significantly reduces the period of the circadian clock. Therefore, our work discovers a new ubiquitin ligase UBR5 that regulates BMAL1 stability and circadian rhythm and elucidates the underlying molecular mechanism. This work provides an additional layer of complexity to the regulatory network of the circadian clock at the post-translational modification level, offering potential insights into the modulation of the dysregulated circadian rhythm.

SENP1 regulates the transformation of lung resident mesenchymal stem cells and is associated with idiopathic pulmonary fibrosis progression.

BACKGROUND: Lung resident mesenchymal stem cells (LR-MSCs) play an important role in idiopathic pulmonary fibrosis (IPF) by transforming into myofibroblasts, thereby losing their repair ability. Evidence suggests that key proteins of multiple signaling pathways are involved in myofibroblast differentiation of LR-MSCs, such as ß-Catenin and GLI family zinc finger 1 (GLI1). These proteins are regulated by SUMO (small ubiquitin-like modifier) modification, which is a post-translational modification that promotes protein degradation, while Sumo specific protein 1 (SENP1)-mediated deSUMOylation produces the opposite biological effects. Therefore, we speculated that SENP1 might be a potential target for treating pulmonary fibrosis by preventing the myofibroblast differentiation of LR-MSCs. METHODS: LR-MSCs were isolated from mice by using immunomagnetic beads. The extracted LR-MSCs were identified by flow cytometric analysis and multilineage differentiation assays. Lentivirus packaged shRNA silenced the expression of SENP1 in vitro and vivo. The silencing efficacy of SENP1 was verified by real-time quantitative PCR. The effect of down-regulated SENP1 on the myofibroblast differentiation of LR-MSCs was assessed by Immunofluorescence and Western blot. Immunoprecipitation was used to clarify that SENP1 was a key target for regulating the activity of multiple signaling pathways in the direction of LR-MSCs differentiation. LR-MSCs resident in the lung was analyzed with in vivo imaging system. HE and Masson staining was used to evaluate the therapeutic effect of LR-MSCs with SENP1 down-regulation on the lung of BLM mice. RESULTS: In this study, we found that the myofibroblast differentiation of LR-MSCs in IPF lung tissue was accompanied by enhanced SENP1-mediated deSUMOylation. The expression of SENP1 increased in LR-MSCs transition of bleomycin (BLM)-induced lung fibrosis. Interfering with expression of SENP1 inhibited the transformation of LR-MSCs into myofibroblasts in vitro and in vivo and restored their therapeutic effect in BLM lung fibrosis. In addition, activation of the WNT/ß-Catenin and Hedgehog/GLI signaling pathways depends on SENP1-mediated deSUMOylation. CONCLUSIONS: SENP1 might be a potential target to restore the repair function of LR-MSCs and treat pulmonary fibrosis. Video Abstract.

Ubiquitin-conjugating enzyme UBE2O regulates cellular clock function by promoting the degradation of the transcription factor BMAL1.

Dysregulation of the circadian rhythm is associated with many diseases, including diabetes, obesity, and cancer. Aryl hydrocarbon receptor nuclear translocator-like protein 1 (Arntl or Bmal1) is the only clock gene whose loss disrupts circadian locomotor behavior in constant darkness. BMAL1 levels are affected by proteasomal inhibition and by several enzymes in the ubiquitin-proteasome system, but the exact molecular mechanism remains unclear. Here, using immunoprecipitation and MS analyses, we discovered an interaction between BMAL1 and ubiquitin-conjugating enzyme E2 O (UBE2O), an E3-independent E2 ubiquitin-conjugating enzyme (i.e. hybrid E2/E3 enzyme). Biochemical experiments with cell lines and animal tissues validated this specific interaction and uncovered that UBE2O expression reduces BMAL1 levels by promoting its ubiquitination and degradation. Moreover, UBE2O expression/knockdown diminished/increased, respectively, BMAL1-mediated transcriptional activity but did not affect BMAL1 gene expression. Bioluminescence experiments disclosed that UBE2O knockdown elevates the amplitude of the circadian clock in human osteosarcoma U2OS cells. Furthermore, mapping of the BMAL1-interacting domain in UBE2O and analyses of BMAL1 stability and ubiquitination revealed that the conserved region 2 (CR2) in UBE2O significantly enhances BMAL1 ubiquitination and decreases BMAL1 protein levels. A Cys-to-Ser substitution experiment identified the critical Cys residue in the CR2 domain responsible for BMAL1 ubiquitination. This work identifies UBE2O as a critical regulator in the ubiquitin-proteasome system, which modulates BMAL1 transcriptional activity and circadian function by promoting BMAL1 ubiquitination and degradation under normal physiological conditions.

Sophocarpine Inhibits Tumorgenesis of Colorectal Cancer via Downregulation of MEK/ERK/VEGF Pathway.

Colorectal cancer (CRC) is one of the most common malignant tumors and the third leading cause of cancer-related deaths in the world. It was reported that sophocarpine could attenuate the progression of CRC in mice. However, the mechanisms by which sophocarpine regulate the proliferation and migration in CRC remain unclear. Thus, this study aimed to investigate anti-tumor mechanisms of sophocarpine in CRC cells. CCK-8 assay, wound healing assay and transwell migration were used to detect cell proliferation and migration, respectively. In addition, Western blotting and enzyme-linked immunosorbent assay (ELISA) were used to further detect protein expressions and cytokines in vitro. The results revealed that sophocarpine significantly inhibited proliferation in HCT116 and SW620 cells, respectively. Meanwhile, sophocarpine inhibited CRC cells migration via downregulation of the levels of N-cadherin, matrix metalloproteinase (MMP)-9, phosphorylated extracellular signal-regulated kinase (p-ERK), p-mitogen-activated protein kinase kinase (MEK), vascular endothelial growth factor (VEGF)-A, VEGF-C and VEGF-D. Moreover, overexpression of MEK reversed the anti-migration effects of sophocarpine on CRC cells via upregulation of VEGF-A/C/D. Our findings indicated that sophocarpine could inhibit CRC cells migration via downregulation of MEK/ERK/VEGF pathway. Thus, sophocarpine may act as a potential agent for the treatment of CRC.

Ursolic Acid Inhibits Tumor Growth via Epithelial-to-Mesenchymal Transition in Colorectal Cancer Cells.

Ursolic acid (UA), a natural pentacyclic triterpenoid, is a promising compound for cancer prevention and therapy. However, its mechanisms of action have not been well elucidated in colorectal cancer cells. Here, using cultured human colon cancer cell lines SW620 and HCT116, this assay demonstrates that UA reduces cell viability, inhibits cell clone formation, and induces caspase-3 mediated apoptosis. Additional experiments show that UA inhibits cell migration and epithelial-mesenchymal transition (EMT), including E-cadherin, Vimentin, Integrin, Twist, and Zeb1 biomakers. These results suggest that UA inhibits cell proliferation, invasion, and metastasis in colorectal cancer cells by affecting mechanisms that regulate EMT. Taken together, the results suggested that the anti-proliferation and anti-metastasis activities of UA was through EMT inhibition in colorectal cancer.

Deubiquitinating enzyme USP9X regulates cellular clock function by modulating the ubiquitination and degradation of a core circadian protein BMAL1.

Living organisms on the earth maintain a roughly 24 h circadian rhythm, which is regulated by circadian clock genes and their protein products. Post-translational modifications of core clock proteins could affect the circadian behavior. Although ubiquitination of core clock proteins was studied extensively, the reverse process, deubiquitination, has only begun to unfold and the role of this regulation on circadian function is not completely understood. Here, we use affinity purification and mass spectrometry analysis to identify probable ubiquitin carboxyl-terminal hydrolase FAF-X (USP9X) as an interacting protein of the core clock protein aryl hydrocarbon receptor nuclear translocator-like protein 1 (ARNTL or BMAL1). Through biochemical experiments, we discover that USP9X reduces BMAL1 ubiquitination, enhances its stability, and increases its protein level, leading to the elevated transcriptional activity. Bioluminescence measurement reveals that USP9X knockdown decreases the amplitude of the cellular circadian rhythm but the period and phase are not affected. Our experiments find a new regulator for circadian clock at the post-translational level and demonstrate a different regulatory function for the circadian clock through the deubiquitination and the up-regulation of the core clock protein BMAL1 in the positive limb of the transcription-translation feedback loop.

Compound C induces protective autophagy in human cholangiocarcinoma cells via Akt/mTOR-independent pathway.

Compound C, a well-known inhibitor of AMP-activated protein kinase (AMPK), has been reported to exert antitumor activities in some types of cells. Whether compound C can exert antitumor effects in human cholangiocarcinoma (CCA) remains unknown. Here, we demonstrated that compound C is a potent inducer of cell death and autophagy in human CCA cells. Autophagy inhibitors increased the cytotoxicity of compound C towards human CCA cells, as confirmed by increased LDH release, and PARP cleavage. It is notable that compound C treatment increased phosphorylated Akt, sustained high levels of phosphorylated p70S6K, and decreased mTOR regulated p-ULK1 (ser757). Based on the data that blocking PI3K/Akt or mTOR had no apparent influence on autophagic response, we suggest that compound C induces autophagy independent of Akt/mTOR signaling in human CCA cells. Further study demonstrated that compound C inhibited the phosphorylation of JNK and its target c-Jun. Blocking JNK by SP600125 or siRNA suppressed autophagy induction upon compound C treatment. Moreover, compound C induced p38 MAPK activation, and its inhibition promoted autophagy induction via JNK activation. In addition, compound C induced p53 expression, and its inhibition attenuated compound C-induced autophagic response. Thus, compound C triggers autophagy, at least in part, via the JNK and p53 pathways in human CCA cells. In conclusion, suppresses autophagy could increase compound C sensitivity in human CCA.

Puerarin prevents LPS-induced acute lung injury via inhibiting inflammatory response.

Acute lung injury (ALI) is a critical illness syndrome with high morbidity and mortality in patients. Inflammation has been known to be involved in the development of ALI. The purpose of this study was to investigate the effect of puerarin on lipopolysaccharide (LPS)-induced ALI in mice. The pro-inflammatory cytokines TNF-α, IL-6 and IL-1ß were determined by ELISA. Western blot analysis was used for detecting the expression of NF-κB, IκBα, and LXRα. And myeloperoxidase (MPO) activity, lung wet/dry (W/D) ratio, and histopathological examination were also detected in lung tissues. The results showed that puerarin significantly inhibited LPS-stimulated MPO activity in lung tissues. Meanwhile, puerarin attenuated lung histopathological changes and lung wet/dry (W/D) ratio. We also found that the expression of pro-inflammatory cytokines, TNF-α, IL-6 and IL-1ß were inhibited by puerarin. Puerarin also inhibited LPS-induced TNF-α in RAW264.7 cells and IL-8 in A549 cells. From the results of western blotting, puerarin significantly suppressed LPS-stimulated NF-κB activation. And the expression of LXRα was dose-dependently increased by treatment of puerarin. The inhibition of puerarin on TNF-α production in RAW264.7 cells and IL-8 production in A549 cells were blocked by LXRα inhibitor geranylgeranyl pyrophosphate (GGPP). These results suggested that puerarin attenuated ALI by activating LXRα, which subsequently inhibited LPS-induced inflammatory response.

The inhibition role of miR-22 in hepatocellular carcinoma cell migration and invasion via targeting CD147.

BACKGROUND: Recently, miR-22 is identified as a tumor-suppressing microRNA in many human cancers. CD147 is a novel cancer-associated biomarker that plays an important role in the invasion and metastasis of malignant tumor. However, the involvement of miR-22 in CD147 regulation and hepatocellular carcinoma (HCC) progression and metastasis has not been investigated. METHODS: We measured miR-22 expression level in 34 paired of HCC and matched normal tissues, HCC cell lines by real-time quantitative RT-PCR. Invasion assay, MTT proliferation assay and wound-healing assay were performed to test the invasion and proliferation of HCC cell after overexpression of miR-22. The effect of miR-22 on HCC in vivo was validated by murine xenograft model. The relationship of miR-22 and its target gene CD147 was also investigated. RESULTS: We found that the expression of miR-22 in HCC tissues and cell lines were much lower than that in normal control, respectively. The expression of miR-22 was inversely correlated with HCC metastatic ability. Moreover, overexpression of miR-22 could significantly inhibit the HCC cell proliferation, migration and invasion in vitro and decrease HCC tumor growth in vivo. Finally, we found that miR-22 interacted with CD147 and decreased its expression, via a specific target site within the CD147 3'UTR by luciferase reporter assay. The expression of CD147 was inversely correlated with miR-22 expression in HCC tissues. CONCLUSION: Our results suggested that miR-22 was downexpressed in HCC and inhibited HCC cell proliferation, migration and invasion through downregulating cancer-associated gene CD147 which may provide a new bio-target for HCC therapy.

Expression of activated signal transducer and activator of transcription-3 as a predictive and prognostic marker in advanced esophageal squamous cell carcinoma.

BACKGROUND: Signal transducer and activator of transcription-3 (STAT3) is an oncogenic transcription factor constitutively active and aberrantly expressed in various types of malignancies, and the expression of p-STAT3 has been recognized as a predictor of poor survival. It remains unclear how variations in p-STAT3 expression influence clinical outcomes in esophageal squamous cell carcinoma (ESCC). METHODS: Between 1 January 2008 and 1 November 2013, 153 advanced esophageal squamous cell carcinoma patients (stage IV) from two cancer centers in West China were treated with paclitaxel and cisplatin. We retrospectively analyzed the clinical outcomes of patients with ESCC and examined the correlation between p-STAT3 levels and clinical outcomes in esophageal cancer patients. RESULTS: Among the 153 patients, positive p-STAT3 expression was observed in 73 of 153 (47.7 %) cases. The median PFS for patients with positive expression of p-STAT3 and negative expression of p-STAT3 was 5.0 months and 6.9 months, respectively (P < 0.001). The median overall survival was significantly higher in patients with p-STAT3 negative tumors than in those with p-STAT3 positive tumors (9.9 vs 8.9 months, P = 0.026). Kaplan-Meier survival analysis showed that p-STAT3 expression was statistically indicative of a poor prognosis for progression-free survival. CONCLUSIONS: These data showed that p-STAT3 expression was significantly associated with poor prognosis in patients with esophageal cancer and could be used as a predictive and prognostic marker in esophageal cancer.

The suppressive role of p38 MAPK in cellular vacuole formation.

Vacuolization of the cytoplasm is one of the dramatic and frequently observed phenomena in various cell types. Cellular vacuoles occur spontaneously or via a wide range of inductive stimuli, but the molecular mechanism involved in this process remains largely unknown. In this study, we investigated the role of the p38 and JNK pathways in the formation of cytoplasmic vacuoles. We found that p38 and JNK agonist anisomycin abolishes spontaneous cytoplasmic vacuolization of HepG2 cells through p38 activation, but not through JNK activation. Importantly, blocking the activity of p38 or suppression the expression of p38 elicits cytoplasmic vacuoles formation in various cancer cells. Furthermore, cytoplasmic vacuoles induced by p38 blocking are derived from the perinuclear region. These observations provide direct evidence for a role of p38 signaling in regulating the formation of cytoplasmic vacuoles.

c-Met function requires N-linked glycosylation modification of pro-Met.

c-Met, the receptor for hepatocyte growth factor (HGF), is cell surface tyrosine kinase that controls cancer cell growth, survival, invasion, and metastasis. Post-translational modification, such as glycosylation, plays an essential role in regulating the function of cell surface molecules. Whether glycosylation modification regulates the enzymatic properties of c-Met is unknown. In this study, we investigated the effect of glycosylation on the function of c-Met. We found that c-Met is an N-linked glycosylated protein. Both pro-Met and p145Met (the ß subunit of mature c-Met) have N-linked glycosylation. Glycosylation inhibitor studies revealed that the N-glycosylation modification of p145Met is from pro-Met, but not due to the further modification of pro-Met. Importantly, blocking the N-glycosylation targets pro-Met to cytoplasm and initiates its phosphorylation independent of HGF engagement. Nonglycosylated pro-Met activates c-Met downstream pathways to a certain extent to compensate for the degradation of p145Met induced by glycosylation blocking-mediated endoplasmic reticulum (ER) stress.

A Chinese herbal formula, Yi-Qi-Fu-Sheng, inhibits migration/invasion of colorectal cancer by down-regulating MMP-2/9 via inhibiting the activation of ERK/MAPK signaling pathways.

BACKGROUND: A Chinese herbal formula, Yi-Qi-Fu-Sheng (YQFS), has long been employed clinically to treat cancer patients. We aimed to determine its effectiveness as a treatment method for colorectal cancer. We investigated the therapeutic effects of YQFS on colorectal cancer, as well as the underlying mechanisms, which have not previously been explored. METHODS: First, YQFS was extracted and chemically characterized. We then tested the effects of YQFS on proliferation and migration by MTT and transwell migration assays in vitro. Mouse xenograft models of colorectal cancer were established by inoculation with HCT-116 cells, and mice received one of three oral doses (200, 400 and 800 mg/kg/day) to evaluate the effects of YQFS extract. Metalloproteinase-2/9 (MMP-2/9) expression in mice was evaluated by gelatin zymography assay. Apoptosis was evaluated by flow cytometry (FCM) analysis in vitro and by TUNEL assay in vivo. ERK and p-ERK expression were evaluated by western blot analysis at the protein level in vitro, and by quantitative RT-PCR at mRNA level in vivo. RESULTS: Our results show that YQFS significantly inhibits colorectal cancer cell proliferation and induces apoptosis and cell cycle arrest at the G1- and S-phase in HCT-116 cells. Furthermore, YQFS effectively retards tumor cell migration and invasion by inhibiting metalloproteinase-2/9 (MMP-2/9) expression, both in vitro and in vivo. Moreover, YQFS had an inhibitory effect on tumor growth in vivo, and induced apoptosis through the inhibition of the ERK1/2 pathway both in vitro and in vivo. CONCLUSION: These findings demonstrate that YQFS extract has an anti-tumor effect in colorectal cancer, which could be attributed to ERK1/2-dependent inhibition of MMP-2/9 expression.

[Phosphorylation of eukaryotic initiation factor 2-alpha inhibits cisplatin-mediated apoptosis of hepatocellular carcinoma cells].

OBJECTIVE: To investigate whether the phosphorylation (functionally inhibitive) of eukaryotic initiation factor 2-alpha (eIF2-a) affects the molecular mechanism of cisplatin-induced cellular apoptosis in human hepatocellular carcinoma (HCC). METHODS: The human HCC cultured cell lines SMMC-7221 and HepG2 were treated with cisplatin alone (controls; 24 h) or in combination with pre-transfection of a dominant-negative eIF2-a mutant (eIF2aS51A) or pre-exposure to an eIF2-a-specific phosphatase inhibitor (salubrinal) to decrease or increase the phosphorylation level, respectively. Changes in expression of apoptosis markers were quantitatively and qualitatively assessed by flow cytometry and western blot analysis. The significance of differences among groups was assessed by analysis of variance testing and of differences between groups was assessed by t-test. RESULTS: Cisplatin treatment induced the appropriate functional-inhibitive phosphorylation of eIF2-a on serine 51. Cisplatin treatment (10 mg/ml) induced significant apoptosis in the eIF2aS51A pre-transfected SMMC-7721 (control: 21.7 +/- 1.5% vs. 50.7 +/- 2.1%, t = 19.454, P less than 0.05) and HepG2 (21.0 +/- 1.0% vs. 57.3 +/- 2.1%, t = 27.250, P less than 0.05). Salubrinal pre-treatment significantly inhibited the cisplatin (15 mg/ml)-induced apoptosis in SMMC-7721 (control: 50.3 +/- 2.5% vs. 16.3 +/- 2.1%, t = 18.031, P less than 0.05) and HepG2 (42.0 +/- 2.6% vs. 12.0 +/- 2.0%, t = 15.667, P less than 0.05). CONCLUSION: Phosphorylation of eIF2-a may act to inhibit cisplatin-induced apoptosis of HCC.

Polymeric microcarriers for minimally-invasive cell delivery.

Tissue engineering (TE) aims at restoring tissue defects by applying the three-dimensional (3D) biomimetic pre-formed scaffolds to restore, maintain, and enhance tissue growth. Broadly speaking, this approach has created a potential impact in anticipating organ-building, which could reduce the need for organ replacement therapy. However, the implantation of such cell-laden biomimetic constructs based on substantial open surgeries often results in severe inflammatory reactions at the incision site, leading to the generation of a harsh adverse environment where cell survival is low. To overcome such limitations, micro-sized injectable modularized units based on various biofabrication approaches as ideal delivery vehicles for cells and various growth factors have garnered compelling interest owing to their minimally-invasive nature, ease of packing cells, and improved cell retention efficacy. Several advancements have been made in fabricating various 3D biomimetic microscale carriers for cell delivery applications. In this review, we explicitly discuss the progress of the microscale cell carriers that potentially pushed the borders of TE, highlighting their design, ability to deliver cells and substantial tissue growth in situ and in vivo from different viewpoints of materials chemistry and biology. Finally, we summarize the perspectives highlighting current challenges and expanding opportunities of these innovative carriers.

Roles of lipid metabolism and its regulatory mechanism in idiopathic pulmonary fibrosis: A review.

Idiopathic pulmonary fibrosis is a progressive lung disease of unknown etiology characterized by distorted distal lung architecture, inflammation, and fibrosis. Several lung cell types, including alveolar epithelial cells and fibroblasts, have been implicated in the development and progression of fibrosis. However, the pathogenesis of idiopathic pulmonary fibrosis is still incompletely understood. The latest research has found that dysregulation of lipid metabolism plays an important role in idiopathic pulmonary fibrosis. The changes in the synthesis and activity of fatty acids, cholesterol and other lipids seriously affect the regenerative function of alveolar epithelial cells and promote the transformation of fibroblasts into myofibroblasts. Mitochondrial function is the key to regulating the metabolic needs of a variety of cells, including alveolar epithelial cells. Sirtuins located in mitochondria are essential to maintain mitochondrial function and cellular metabolic homeostasis. Sirtuins can maintain normal lipid metabolism by regulating respiratory enzyme activity, resisting oxidative stress, and protecting mitochondrial function. In this review, we aimed to discuss the difference between normal and idiopathic pulmonary fibrosis lungs in terms of lipid metabolism. Additionally, we highlight recent breakthroughs on the effect of abnormal lipid metabolism on idiopathic pulmonary fibrosis, including the effects of sirtuins. Idiopathic pulmonary fibrosis has its high mortality and limited therapeutic options; therefore, we believe that this review will help to develop a new therapeutic direction from the aspect of lipid metabolism in idiopathic pulmonary fibrosis.

Overcoming Cancer Multi-drug Resistance (MDR): Reasons, mechanisms, nanotherapeutic solutions, and challenges.

Multi-drug resistance (MDR) in cancer cells, either intrinsic or acquired through various mechanisms, significantly hinders the therapeutic efficacy of drugs. Typically, the reduced therapeutic performance of various drugs is predominantly due to the inherent over expression of ATP-binding cassette (ABC) transporter proteins on the cell membrane, resulting in the deprived uptake of drugs, augmenting drug detoxification, and DNA repair. In addition to various physiological abnormalities and extensive blood flow, MDR cancer phenotypes exhibit improved apoptotic threshold and drug efflux efficiency. These severe consequences have substantially directed researchers in the fabrication of various advanced therapeutic strategies, such as co-delivery of drugs along with various generations of MDR inhibitors, augmented dosage regimens and frequency of administration, as well as combinatorial treatment options, among others. In this review, we emphasize different reasons and mechanisms responsible for MDR in cancer, including but not limited to the known drug efflux mechanisms mediated by permeability glycoprotein (P-gp) and other pumps, reduced drug uptake, altered DNA repair, and drug targets, among others. Further, an emphasis on specific cancers that share pathogenesis in executing MDR and effluxed drugs in common is provided. Then, the aspects related to various nanomaterials-based supramolecular programmable designs (organic- and inorganic-based materials), as well as physical approaches (light- and ultrasound-based therapies), are discussed, highlighting the unsolved issues and future advancements. Finally, we summarize the review with interesting perspectives and future trends, exploring further opportunities to overcome MDR.

Efficacy and safety of immune checkpoint inhibitors in post-TKI NSCLC patients harboring EGFR mutations.

OBJECTIVE: Immune checkpoint inhibitors (ICIs) have been validated in epidermal growth factor receptor (EGFR) wild-type advanced non-small cell lung cancer (NSCLC) patients. However, there exists no evidence regarding NSCLC patients harboring EGFR mutations, experiencing EGFR-TKI (tyrosine kinase inhibitor) treatment failure. We collected clinical information from real world and conducted a time series-based meta-analysis to determine the efficacy and safety of ICIs in patients harboring EGFR mutations and experienced EGFR-TKIs resistance. METHODS: Twenty-two NSCLC patients with EGFR mutations after TKI resistance were included from two hospitals. PubMed, Embase and Cochrane Library were searched for relevant literature published until December 31, 2021. Endpoint outcomes included mortality and progression-free survival (PFS) at different times of follow-up. RESULTS: In total, 22 patients showed that the median PFS was 5.6 months (range 2.0-9.0 months). According to treatment strategies, the median PFS was 2.4 months (range 2.0-5.3 months) in the ICI monotherapy group and 5.9 months (range 2.8-9.0 months) in the ICI combined Chemotherapy group. Additionally, sixteen studies, including 5 trials, 10 controlled cohorts and 1 real-world study, were assessed, involving a total of ICI-treated NSCLC patients with EGFR mutation after TKI failure. The 6-month survival and PFS rate were 0.82 (95% CI 0.36-0.97) and 0.55 (95% CI 0.34-0.74), respectively. ICI combined chemotherapy showed the best survival outcome among these groups, as demonstrated by the 12-month survival rate and PFS. No new safety signals were identified with the combination therapy. The frequency of treatment-related adverse events was similar to that in previously reported studies of chemotherapy combined with checkpoint inhibitors. CONCLUSIONS: The addition of ICIs plus chemotherapy may significantly improve progression-free survival among patients with locally advanced or metastatic non-squamous NSCLC who EGFR-TKIs resistance.

Activation of PKR/eIF2α signaling cascade is associated with dihydrotestosterone-induced cell cycle arrest and apoptosis in human liver cells.

Androgen receptor (AR) signaling plays an important role in the development and progression of several liver diseases, including hepatocellular carcinoma (HCC) and non-alcoholic fatty liver disease (NAFLD). Dihydrotestosterone (DHT) is the active metabolite of the major circulating androgen, testosterone. In this study, we investigated the effect of DHT on human liver cells. We found that DHT not only induces cell cycle arrest but also initiates apoptosis in androgen-sensitive liver cells, such as SMMC-7721 and L02. Importantly, DHT/AR induces the activation of RNA-dependent protein kinase (PKR)/eukaryotic initiation factor-2 alpha (eIF2α) cascades in androgen-sensitive liver cells. PKR/eIF2α activation-induced growth arrest and DNA damage-inducible gene 153 (GADD153) and heat shock protein 27 (Hsp27) expression contribute to cell cycle arrest in response to DHT. It is notable that DHT administration results in androgen-sensitive liver cells apoptosis, at least in part, through PKR/eIF2α/GADD153 cascades. These results suggest that the androgen/AR pathway plays a pivotal role in liver cell growth and apoptosis regulating, whose deregulation might be involved in the pathogenesis of liver diseases.