Human ST3Gal II and ST6GalNAc IV genes increase human serum-mediated cytotoxicity to xenogeneic cells.

Carbohydrate-antigens widely existed on glycoproteins and glycosphingolipids of all mammalian cells play a crucial role in self-defense and immunity. Xeno-reactive antibodies included in natural human sera play a protecting role in an acute phase-rejection of xenotransplantation. In this study, we investigated the effect of an alteration of glycosylation-pattern, caused by human sialyltransferases such as hST3Gal II or hST6GalNAc IV, on human serum mediated cytotoxicity in pig kidney PK15 cells. From LDH cytotoxicity assay, cytotoxicity to human serum was significantly increased in hST3Gal II and hST6GalNAc IV-transfected PK15 cells, as compared to the control. In the hST6Gal I-carrying cells, the cytotoxicity to human serum was rather decreased. Moreover, flow cytometry analysis revealed that an alteration of pig glycosylation-pattern by hST3Gal II or hST6GalNAc IV influences on a binding of human IgM or IgG, respectively, in pig kidney cells, regardless of Gal antigen alteration. Finally, we found that hST6GalNAc IV contributed to increase of terminal disialylated tetrasaccharide structure, disialyl T antigen, as evidenced by increase of the MAL II lectin binding capacity in the hST6GalNAc IV-transfected PK15 cells, compared with control. Therefore, our results suggest that carbohydrate antigens, such as disialyl T antigen, newly synthesized by the ST3Gal II- and ST6GalNAc IV are potentially believed to be new xeno-reactive elements.

4-O-Methylascochlorin Synergistically Enhances 5-Fluorouracil-Induced Apoptosis by Inhibiting the Wnt/ß-Catenin Signaling Pathway in Colorectal Cancer Cells.

4-O-Methyl-ascochlorin (MAC), a derivative of the prenyl-phenol antibiotic ascochlorin extracted from the fungus Ascochyta viciae, shows anticarcinogenic effects on various cancer cells. 5-Fluorouracil (5-FU) is used to treat colorectal cancer (CRC); however, its efficacy must be enhanced. In this study, we investigated the molecular mechanisms by which MAC acts synergistically with 5-FU to inhibit cell proliferation and induce apoptosis in CRC cells. MAC enhanced the cytotoxic effects of 5-FU by suppressing the Akt/mTOR/p70S6K and Wnt/ß-catenin signaling pathways. It also reduced the viability of 5-FU-resistant (5-FU-R) cells. Furthermore, expression of anti-apoptosis-related proteins and cancer stem-like cell (CSC) markers by 5-FU-R cells decreased in response to MAC. Similar to MAC, the knockdown of CTNNB1 induced apoptosis and reduced expression of mRNA encoding CRC markers in 5-FU-R cells. In summary, these results suggest that MAC and other ß-catenin modulators may be useful in overcoming the 5-FU resistance of CRC cells.

4-O-Methylascochlorin-Mediated BNIP-3 Expression Controls the Balance of Apoptosis and Autophagy in Cervical Carcinoma Cells.

4-O-methylascochlorin (MAC) is a 4-fourth carbon-substituted derivative of ascochlorin, a compound extracted from a phytopathogenic fungus Ascochyta viciae. MAC induces apoptosis and autophagy in various cancer cells, but the effects of MAC on apoptosis and autophagy in cervical cancer cells, as well as how the interaction between apoptosis and autophagy mediates the cellular anticancer effects are not known. Here, we investigated that MAC induced apoptotic cell death of cervical cancer cells without regulating the cell cycle and promoted autophagy by inhibiting the phosphorylation of serine-threonine kinase B (Akt), mammalian target of rapamycin (mTOR), and 70-kDa ribosomal protein S6 kinase (p70S6K). Additional investigations suggested that Bcl-2/adenovirus E1B 19 kDa protein-interacting protein 3 (BNIP-3), but not Hypoxia-inducible factor 1 alpha (HIF-1α), is a key regulator of MAC-induced apoptosis and autophagy. BNIP-3 siRNA suppressed MAC-induced increases in cleaved- poly (ADP-ribose) polymerase (PARP) and LC3II expression. The pan-caspase inhibitor Z-VAD-FMK suppressed MAC-induced cell death and enhanced MAC-induced autophagy. The autophagy inhibitor chloroquine (CQ) enhanced MAC-mediated cell death by increasing BNIP-3 expression. These results indicate that MAC induces apoptosis to promote cell death and stimulates autophagy to promote cell survival by increasing BNIP-3 expression. This study also showed that co-treatment of cells with MAC and CQ further enhanced the death of cervical cancer cells.

Interaction between SCP3 and JAB1 Confers Cancer Therapeutic Resistance and Stem-like Properties through EGF Expression.

Synaptonemal complex protein 3 (SCP3), a member of the Cor1 family, has been implicated in cancer progression, and therapeutic resistance, as well as cancer stem cell (CSC)-like properties. Previously, we demonstrated that SCP3 promotes these aggressive phenotypes via hyperactivation of the AKT signaling pathway; however, the underlying mechanisms responsible for SCP3-induced AKT activation remain to be elucidated. In this study, we demonstrated that the EGF-EGFR axis is the primary route through which SCP3 acts to activate AKT signaling. SCP3 triggers the EGFR-AKT pathway through transcriptional activation of EGF. Notably, neutralization of secreted EGF by its specific monoclonal antibody reversed SCP3-mediated aggressive phenotypes with a concomitant reversal of EGFR-AKT activation. In an effort to elucidate the molecular mechanisms underlying SCP3-induced transcriptional activation of EGF, we identified Jun activation domain-binding protein 1 (JAB1) as a binding partner of SCP3 using a yeast two-hybrid (Y2H) assay system, and we demonstrated that SCP3 induces EGF transcription through physical interaction with JAB1. Thus, our findings establish a firm molecular link among SCP3, EGFR, and AKT by identifying the novel roles of SCP3 in transcriptional regulation. We believe that these findings hold important implications for controlling SCP3high therapeutic-refractory cancer.

Gram-Negative Bacterial Endotoxin LPS Induces NeuGc Loss through Ets1-Dependent Downregulation of Intestine-Specific pcmah Transcript in Porcine Intestinal Cells.

N-glycolylneuraminic acid (NeuGc), a non-human sialic acid derivative synthesized by cytidine-5'-monophospho-N-acetylneuraminic acid hydroxylase (CMAH), plays a crucial role in mediating infections by certain pathogens. Although it has been postulated that NeuGc biosynthesis and CMAH expression are downregulated during microbial infection, the underlying mechanisms remain unclear. The present study showed that exposure to lipopolysaccharide (LPS), a Gram-negative bacterial endotoxin, leads to loss of NeuGc biosynthesis in pig small intestinal I2I-2I cells. This LPS-induced NeuGc loss was accompanied by decreased CMAH transcript levels, especially intestine-specific 5'pcmah-1. Furthermore, LPS suppressed the activity of the Pi promoter responsible for 5'pcmah-1 by inhibiting DNA binding of Est1. These findings provide insight into the regulatory mechanisms of Neu5Gc biosynthesis during pathogenic infectious events, which may represent a host defense mechanism that protects the self against pathogenic bacterial infections even in non-sanitary environments.

Stabilization of HDAC1 via TCL1-pAKT-CHFR axis is a key element for NANOG-mediated multi-resistance and stem-like phenotype in immune-edited tumor cells.

Cancer immunoediting enriches NANOG expression in tumor cells, resulting in multi-drug resistance and stem-like phenotypes. We previously demonstrated that these NANOG-associated phenotypes are promoted through HDAC1 transcriptional upregulation. In this study, we identified that NANOG also contributes to the stabilization of HDAC1 protein through the AKT signaling pathway. NANOG-AKT axis leads to phosphor-dependent inactivation of CHFR, an E3 ligase for HDAC1 protein, and thereby inhibiting the ubiquitin-mediated degradation of HDAC1. Furthermore, AKT inhibition disrupts HDAC1 WT-mediated phenotypes but had no effect on the phenotypes mediated by HDAC1 FM, a mutant that is unable to interact with CHFR. Critically, we applied a catalytic dead mutant, HDAC1-H141A, to uncover that HDAC1 confers immune-resistance, drug-resistance and stem-like phenotype in tumor cells through its catalytic activity. Collectively, our results establish a firm molecular link in immune-edited tumor cells among NANOG, AKT, CHFR, and HDAC1, identifying HDAC1 as a molecular target in controlling NANOGHIGH immune-refractory cancer.

Housekeeping promoter 5'pcmah-2 of pig CMP-N-acetylneuraminic acid hydroxylase gene for NeuGc expression.

In the present study, we isolated pCMAH house-keeping promoter regions (Ph), which are responsible for transcriptional regulation and which are located upstream of the alternative transcript pcmah-2. Luciferase reporter assays using serial construction of each deleted promoter demonstrated that the Ph promoter was highly active in pig-derived kidney PK15. Ph promoter of pcmah lacked a TATA box, but contained three putative Sp1 binding sites. Mutations of these Sp1 binding sites always resulted in the reduction of luciferase activities in Ph-334. In addition, treatment with mithramycin A (25-100 nM) decreased the luciferase activities of the Ph promoters and NeuGc expression in a dose-dependent manner. Electrophoretic mobility shift assay analysis revealed that the probes containing each Sp1 binding site bound to Sp1. Taken together, the results indicate that Sp1 bind to their putative binding sites on the Ph promoter regions of pcmah and positively regulate the promoter activity in pig kidney cells. Interspecies comparison of 5'UTRs and 5'flanking regions shows high homology between pig and cattle, and Sp1 binding sites existing in genomic regions corresponding Ph region are evolutionally conserved.

Hepatitis B virus X protein specially regulates the sialyl lewis a synthesis among glycosylation events for metastasis.

BACKGROUND: The metastasis of hematogenous cancer cells is associated with abnormal glycosylation such as sialyl lewis antigens. Although the hepatitis B virus X protein (HBx) plays important role in liver disease, the precise function of HBx on aberrant glycosylation for metastasis remains unclear. METHODS: The human hepatocellular carcinoma tissues, HBx transgenic mice and HBx-transfected cells were used to check the correlation of expressions between HBx and Sialyl lewis antigen for cancer metastasis. To investigate whether expression levels of glycosyltransferases induced in HBx-transfected cells are specifically associated with sialyl lewis A (SLA) synthesis, which enhances metastasis by interaction of liver cancer cells with endothelial cells, ShRNA and siRNAs targeting specific glycosyltransferases were used. RESULTS: HBx expression in liver cancer region of HCC is associated with the specific synthesis of SLA. Furthermore, the SLA was specifically induced both in liver tissues from HBx-transgenic mice and in in vitro HBx-transfected cells. HBx increased transcription levels and activities of α2-3 sialyltransferases (ST3Gal III), α1-3/4 fucosyltransferases III and VII (FUT III and VII) genes, which were specific for SLA synthesis, allowing dramatic cell-cell adhesion for metastatic potential. Interestingly, HBx specifically induced expression of N-acetylglucosamine-ß1-3 galactosyltransferase V (ß1-3GalT 5) gene associated with the initial synthesis of sialyl lewis A, but not ß1-4GalT I. The ß1-3GalT 5 shRNA suppressed SLA expression by HBx, blocking the adhesion of HBx-transfected cells to the endothelial cells. Moreover, ß1-3GalT 5 silencing suppressed lung metastasis of HBx-transfected cells in in vivo lung metastasis system. CONCLUSION: HBx targets the specific glycosyltransferases for the SLA synthesis and this process regulates hematogenous cancer cell adhesion to endothelial cells for cancer metastasis.

GM-CSF-loaded chitosan hydrogel as an immunoadjuvant enhances antigen-specific immune responses with reduced toxicity.

BACKGROUND: The application of vaccine adjuvants has been vigorously studied for a diverse range of diseases in order to improve immune responses and reduce toxicity. However, most adjuvants have limited uses in clinical practice due to their toxicity. METHODS: Therefore, to reduce health risks associated with the use of such adjuvants, we developed an advanced non-toxic adjuvant utilizing biodegradable chitosan hydrogel (CH-HG) containing ovalbumin (OVA) and granulocyte-macrophage colony-stimulating factor (GM-CSF) as a local antigen delivery system. RESULTS: After subcutaneous injection into mice, OVA/GM-CSF-loaded CH-HG demonstrated improved safety and enhanced OVA-specific antibody production compared to oil-based adjuvants such as Complete Freund's adjuvant (CFA) or Incomplete Freund's adjuvant (IFA). Moreover, CH-HG system-mediated immune responses was characterized by increased number of OVA-specific CD4(+) and CD8(+) INF-γ(+) T cells, leading to enhanced humoral and cellular immunity. CONCLUSIONS: In this study, the improved safety and enhanced immune response characteristics of our novel adjuvant system suggest the possibility of the extended use of adjuvants in clinical practice with reduced apprehension about toxic side effects.

Ganglioside GM3 is required for caffeic acid phenethyl ester-induced megakaryocytic differentiation of human chronic myelogenous leukemia K562 cells.

The human chronic myelogenous cell line K562 has been used extensively as a model for the study of leukemia differentiation. We show here that treatment of K562 cells with caffeic acid phenethyl ester (CAPE) induced a majority of cells to differentiate towards the megakaryocytic lineage. Microscopy analysis showed that K562 cells treated with CAPE exhibited characteristic features of physiological megakaryocytic differentiation, including the presence of vacuoles and demarcation membranes. Differentiation of K562 cells treated with CAPE was also accompanied by a net increase in megakaryocytic markers. The transcriptional activity of lactosylceramide α-2,3-sialyltransferase (GM3 synthase) and synthesis of ganglioside GM3 were increased by CAPE treatment. The promoter analysis of GM3 synthase demonstrated that CAPE induced the expression of GM3 synthase mRNA via activation of the cAMP response element-binding protein (CREB), transcription factor in nucleus. Interestingly, the inhibition of ganglioside GM3 synthesis by D-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propranol (D-PDMP) and GM3 synthase-siRNA blocked the CAPE-induced expression of the megakaryocytic markers and differentiation of K562 cells. Taken together, these results suggest that CAPE induces ganglioside GM3-mediated megakaryocytic differentiation of human chronic myelogenous cells.

Apoptosis inhibitor-5 overexpression is associated with tumor progression and poor prognosis in patients with cervical cancer.

BACKGROUND: The apoptosis inhibitor-5 (API5), anti-apoptosis protein, is considered a key molecule in the tumor progression and malignant phenotype of tumor cells. Here, we investigated API5 expression in cervical cancer, its clinical significance, and its relationship with phosphorylated extracellular signal-regulated kinase 1 and 2 (pERK1/2) in development and progression of cervical cancer. METHODS: API5 effects on cell growth were assessed in cervical cancer cell lines. API5 and pERK1/2 immunohistochemical staining were performed on a cervical cancer tissue microarray consisting of 173 primary cervical cancers, 306 cervical intraepithelial neoplasias (CINs), and 429 matched normal tissues. RESULTS: API5 overexpression promoted cell proliferation and colony formation in CaSki cells, whereas API5 knockdown inhibited the both properties in HeLa cells. Immunohistochemical staining showed that API5 expression increased during the normal to tumor transition of cervical carcinoma (P < 0.001), and this increased expression was significantly associated with tumor stage (P = 0.004), tumor grade (P < 0.001), and chemo-radiation response (P = 0.004). API5 expression levels were positively associated with pERK1/2 in cervical cancer (P < 0.001) and high grade CIN (P = 0.031). In multivariate analysis, API5+ (P = 0.039) and combined API5+/pERK1/2+ (P = 0.032) were independent prognostic factors for overall survival. CONCLUSIONS: API5 expression is associated with pERK1/2 in a subset of cervical cancer patients and its expression predicts poor overall survival, supporting that API5 may be a promising novel target for therapeutic interventions.

Ganglioside GM3 participates in the TGF-ß1-induced epithelial-mesenchymal transition of human lens epithelial cells.

TGF-ß (transforming growth factor-ß)-induced EMT (epithelial-mesenchymal transition) induces the proliferation and migration of the HLE (human lens epithelial) cells. Ganglioside GM3, simple sialic-acid-containing glycosphingolipids on mammalian cell membranes, regulates various pathological phenomena such as insulin resistance and tumour progression. However, the relationship between ganglioside GM3 and TGF-ß-induced EMT in the HLE B-3 cells is poorly understood. In the present study we demonstrated that ganglioside GM3 was involved in TGF-ß1-induced EMT in HLE B-3 cells. Our results indicated that the expression of ganglioside GM3 and GM3 synthase mRNA were significantly increased in TGF-ß1-induced HLE B-3 cells. Reporter gene analysis also demonstrated that transcriptional activation of the GM3 synthase gene was regulated by Sp1 (specificity protein 1) in HLE B-3 cells upon TGF-ß1 stimulation. Interestingly, the inhibition of ganglioside GM3 expression by d-PDMP [d-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol] and GM3 synthase shRNA (short hairpin RNA) resulted significantly in the suppression of cell migration and EMT-related signalling in HLE B-3 cells stimulated by TGF-ß. Furthermore, exogenous treatment of ganglioside GM3 rescued the expression of EMT molecules and cell migration suppressed by the depletion of ganglioside GM3 in TGF-ß1-induced HLE B-3 cells. We also found that ganglioside GM3 interacted with TGFßRs (TGF-ß receptors) in TGF-ß1-induced HLE B-3 cells. Taken together, these results suggest that ganglioside GM3 induced by TGF-ß1 regulates EMT by potential interaction with TGFßRs.

Rg3-enriched red ginseng extracts enhance apoptosis in CoCl2-stimulated breast cancer cells by suppressing autophagy.

Background: Ginsenoside Rg3, a primary bioactive component of red ginseng, has anti-cancer effects. However, the effects of Rg3-enriched ginseng extract (Rg3RGE) on apoptosis and autophagy in breast cancer have not yet been investigated. In the present study, we explored the anti-tumor effects of Rg3RGE on breast cancer cells stimulated CoCl2, a mimetic of the chronic hypoxic response, and determined the operative mechanisms of action. Methods: The inhibitory mechanisms of Rg3RGE on breast cancer cells, such as apoptosis, autophagy and ROS levels, were detected both in vitro. To determine the anti-cancer effects of Rg3RGE in vivo, the cancer xenograft model was used. Results: Rg3RGE suppressed CoCl2-induced spheroid formation and cell viability in 3D culture of breast cancer cells. Rg3RGE promoted apoptosis by increasing cleaved caspase 3 and cleaved PARP and decreasing Bcl2 under the hypoxia mimetic conditions. Further, we identified that Rg3RGE promoted apoptosis by inhibiting lysosomal degradation of autophagosome contents in CoCl2-induced autophagy. We further identified that Rg3RGE-induced apoptotic cell death and autophagy inhibition was mediated by increased intracellular ROS levels. Similarly, in the in vivo xenograft model, Rg3RGE induced apoptosis and inhibited cell proliferation and autophagy. Conclusion: Rg3RGE-stimulated ROS production promotes apoptosis and inhibits protective autophagy under hypoxic conditions. Autophagosome accumulation is critical to the apoptotic effects of Rg3RGE. The in vivo findings also demonstrate that Rg3RGE inhibits breast cancer cell growth, suggesting that Rg3RGE has potential as potential as a putative breast cancer therapeutic.

PPIB/Cyclophilin B expression associates with tumor progression and unfavorable survival in patients with pulmonary adenocarcinoma.

Cyclophilin B (CypB), encoded by peptidylprolyl isomerase B (PPIB), is involved in cellular transcriptional regulation, immune responses, chemotaxis, and proliferation. Recent studies have shown that PPIB/CypB is associated with tumor progression and chemoresistance in various cancers. However, the clinicopathologic significance and mechanism of action of PPIB/CypB in non-small cell lung cancer (NSCLC) remain unclear. In this study, we used RNA in situ hybridization to examine PPIB expression in 431 NSCLC tissue microarrays consisting of 295 adenocarcinomas (ADCs) and 136 squamous cell carcinomas (SCCs). Additionally, Ki-67 expression was evaluated using immunohistochemistry. The role of PPIB/CypB was assessed in five human NSCLC cell lines. There was a significant correlation between PPIB/CypB expression and Ki-67 expression in ADC (Spearman correlation r=0.374, P<0.001) and a weak correlation in SCC (r=0.229, P=0.007). In ADCs, high PPIB expression (PPIBhigh) was associated with lymph node metastasis (P=0.023), advanced disease stage (P=0.014), disease recurrence (P=0.013), and patient mortality (P=0.015). Meanwhile, high Ki-67 expression (Ki-67high) was correlated with male sex, smoking history, high pT stage, lymph node metastasis, advanced stage, disease recurrence, and patient mortality in ADC (all P<0.001). However, there was no association between either marker or clinicopathological factors, except for old age and PPIBhigh (P=0.038) in SCC. Survival analyses revealed that the combined expression of PPIBhigh/Ki-67high was an independent prognosis factor for poor disease-free survival (HR 1.424, 95% CI 1.177-1.723, P<0.001) and overall survival (HR 1.266, 95% CI 1.036-1.548, P=0.021) in ADC, but not in SCC. Furthermore, PPIB/CypB promoted the proliferation, colony formation, and migration of NSCLC cells. We also observed the oncogenic properties of PPIB/CypB expression in human bronchial epithelial cells. In conclusion, PPIB/CypB contributes to tumor growth in NSCLC, and elevated PPIB/Ki-67 levels are linked to unfavorable survival, especially in ADC.

TRPV1 inhibition overcomes cisplatin resistance by blocking autophagy-mediated hyperactivation of EGFR signaling pathway.

Cisplatin resistance along with chemotherapy-induced neuropathic pain is an important cause of treatment failure for many cancer types and represents an unmet clinical need. Therefore, future studies should provide evidence regarding the mechanisms of potential targets that can overcome the resistance as well as alleviate pain. Here, we show that the emergence of cisplatin resistance is highly associated with EGFR hyperactivation, and that EGFR hyperactivation is arisen by a transcriptional increase in the pain-generating channel, TRPV1, via NANOG. Furthermore, TRPV1 promotes autophagy-mediated EGF secretion via Ca2+ influx, which activates the EGFR-AKT signaling and, consequentially, the acquisition of cisplatin resistance. Importantly, TRPV1 inhibition renders tumors susceptible to cisplatin. Thus, our findings indicate a link among cisplatin resistance, EGFR hyperactivation, and TRPV1-mediated autophagic secretion, and implicate that TRPV1 could be a crucial drug target that could not only overcome cisplatin resistance but also alleviate pain in NANOG+ cisplatin-resistant cancer.

The lipoprotein-associated phospholipase A2 inhibitor Darapladib sensitises cancer cells to ferroptosis by remodelling lipid metabolism.

Arachidonic and adrenic acids in the membrane play key roles in ferroptosis. Here, we reveal that lipoprotein-associated phospholipase A2 (Lp-PLA2) controls intracellular phospholipid metabolism and contributes to ferroptosis resistance. A metabolic drug screen reveals that darapladib, an inhibitor of Lp-PLA2, synergistically induces ferroptosis in the presence of GPX4 inhibitors. We show that darapladib is able to enhance ferroptosis under lipoprotein-deficient or serum-free conditions. Furthermore, we find that Lp-PLA2 is located in the membrane and cytoplasm and suppresses ferroptosis, suggesting a critical role for intracellular Lp-PLA2. Lipidomic analyses show that darapladib treatment or deletion of PLA2G7, which encodes Lp-PLA2, generally enriches phosphatidylethanolamine species and reduces lysophosphatidylethanolamine species. Moreover, combination treatment of darapladib with the GPX4 inhibitor PACMA31 efficiently inhibits tumour growth in a xenograft model. Our study suggests that inhibition of Lp-PLA2 is a potential therapeutic strategy to enhance ferroptosis in cancer treatment.

4-O-methylascochlorin-stimulated HIF-1α expression induces the epithelial mesenchymal transition and cell survival in breast cancer cells.

4-O-Methyl-ascochlorin (MAC), a derivative of the prenyl-phenol antibiotic ascochlorin, promotes accumulation of HIF-1α. In this study, we investigated the molecular mechanisms of the effect of MAC on cell migration and mesenchymal epithelial transition (EMT) processes in breast cancer cells. MAC upregulated cell motility and migration regardless of cell viability, and promoted EMT features by regulating EMT-related proteins and transcription. In addition, the MAC-induced increase in the EMT was closely related to activation of HIF-1α expression. However, the MAC-induced EMT was not associated with AMPK phosphorylation or intracellular ROS, which stimulate HIF-1α expression. Similarly, HIF-1α-mediated autophagy induced by MAC was not related to EMT-related proteins. Inhibition of HIF-1α activity inhibited MAC-stimulated cell migration and increased MAC-induced cell death, indicating that HIF-1α activation is important for MAC-mediated cell migration and survival in breast cancer cells. Together, these results suggest that MAC can be used to investigate the link between HIF-1α activation and other oncogenes or tumor suppressors in breast cancer cells.

Targeting the NANOG/HDAC1 axis reverses resistance to PD-1 blockade by reinvigorating the antitumor immunity cycle.

Immune checkpoint blockade (ICB) therapy has shifted the paradigm for cancer treatment. However, the majority of patients lack effective responses because of the emergence of immune-refractory tumors that disrupt the amplification of antitumor immunity. Therefore, the identification of clinically available targets that restrict antitumor immunity is required to develop potential combination therapies. Here, using transcriptomic data on patients with cancer treated with programmed cell death protein 1 (PD-1) therapy and newly established mouse preclinical anti-PD-1 therapy-refractory models, we identified NANOG as a factor restricting the amplification of the antitumor immunity cycle, thereby contributing to the immune-refractory feature of the tumor microenvironment (TME). Mechanistically, NANOG induced insufficient T cell infiltration and resistance to CTL-mediated killing via the histone deacetylase 1-dependent (HDAC1-dependent) regulation of CXCL10 and MCL1, respectively. Importantly, HDAC1 inhibition using an actionable agent sensitized NANOGhi immune-refractory tumors to PD-1 blockade by reinvigorating the antitumor immunity cycle. Thus, our findings implicate the NANOG/HDAC1 axis as a central molecular target for controlling immune-refractory tumors and provide a rationale for combining HDAC inhibitors to reverse the refractoriness of tumors to ICB therapy.

Rg3-enriched red ginseng extract promotes lung cancer cell apoptosis and mitophagy by ROS production.

BACKGROUND: Red Ginseng has been used for many years to treat diseases. Ginsenoside Rg3 has documented therapeutic effects, including anticancer and anti-inflammatory activities. However, the anticancer effect of Rg3-enriched red ginseng extract (Rg3-RGE) and its underlying mechanisms have not been fully explored. We investigated whether Rg3-RGE plays an anti-tumor role in lung cancer cells. METHODS: To examine the effect of Rg3-RGE on lung cancer cells, we performed cell viability assays, flow cytometry, western blotting analysis, and immunofluorescence to monitor specific markers. RESULTS: Rg3-RGE significantly inhibited cell proliferation and induced mitochondria-dependent apoptosis. Furthermore, Rg3-RGE also increased expression of mitophagy-related proteins such as PINK1 and Parkin. In addition, treatment with Rg3-RGE and mitophagy inhibitors stimulated cell death by inducing mitochondria dysfunction. CONCLUSIONS: Rg3-RGE could be used as a therapeutic agent against lung cancer.

NANOG confers resistance to complement-dependent cytotoxicity in immune-edited tumor cells through up-regulating CD59.

Cancer immunoediting drives the adaptation of tumor cells to host immune surveillance. Previously, we have demonstrated that immunoediting driven by cytotoxic T lymphocytes (CTLs) enriches NANOG+ tumor cells with immune-refractory properties. Here, we found that CTL-mediated immune pressure triggered cross-resistance of tumor cells to the complement system, a part of the innate immune system. In this process, NANOG upregulated the membrane-bound complement regulatory protein (mCRP) CD59 through promoter occupancy, thereby contributing to the resistance of tumor cells against complement-dependent cytotoxicity (CDC). Notably, targeting of NANOG sensitized the immune-refractory tumor cells to trastuzumab-mediated CDC. Collectively, our results revealed a possible mechanism through which selection imposed by T-cell based immunotherapy triggered complement-resistant phenotypes in the tumor microenvironment (TME), by establishing a firm molecular link between NANOG and CD59 in immune-edited tumor cells. We believe these results hold important implications for the clinical application of CDC-mediated therapeutic antibody.