A Cell Surface-Binding Antibody Atlas Nominates a MUC18-Directed Antibody-Drug Conjugate for Targeting Melanoma.

Recent advances in targeted therapy and immunotherapy have substantially improved the treatment of melanoma. However, therapeutic strategies are still needed for unresponsive or treatment-relapsed patients with melanoma. To discover antibody-drug conjugate (ADC)-tractable cell surface targets for melanoma, we developed an atlas of melanoma cell surface-binding antibodies (pAb) using a proteome-scale antibody array platform. Target identification of pAbs led to development of melanoma cell killing ADCs against LGR6, TRPM1, ASAP1, and MUC18, among others. MUC18 was overexpressed in both tumor cells and tumor-infiltrating blood vessels across major melanoma subtypes, making it a potential dual-compartment and universal melanoma therapeutic target. AMT-253, an MUC18-directed ADC based on topoisomerase I inhibitor exatecan and a self-immolative T moiety, had a higher therapeutic index compared with its microtubule inhibitor-based counterpart and favorable pharmacokinetics and tolerability in monkeys. AMT-253 exhibited MUC18-specific cytotoxicity through DNA damage and apoptosis and a strong bystander killing effect, leading to potent antitumor activities against melanoma cell line and patient-derived xenograft models. Tumor vasculature targeting by a mouse MUC18-specific antibody-T1000-exatecan conjugate inhibited tumor growth in human melanoma xenografts. Combination therapy of AMT-253 with an antiangiogenic agent generated higher efficacy than single agent in a mucosal melanoma model. Beyond melanoma, AMT-253 was also efficacious in a wide range of MUC18-expressing solid tumors. Efficient target/antibody discovery in combination with the T moiety-exatecan linker-payload exemplified here may facilitate discovery of new ADC to improve cancer treatment. SIGNIFICANCE: Discovery of melanoma-targeting antibodies using a proteome-scale array and use of a cutting-edge linker-payload system led to development of a MUC18-targeting antibody-exatecan conjugate with clinical potential for treating major melanoma subtypes.

Identification of quiescent FOXC2+ spermatogonial stem cells in adult mammals.

In adult mammals, spermatogenesis embodies the complex developmental process from spermatogonial stem cells (SSCs) to spermatozoa. At the top of this developmental hierarchy lie a series of SSC subpopulations. Their individual identities as well as the relationships with each other, however, remain largely elusive. Using single-cell analysis and lineage tracing, we discovered both in mice and humans the quiescent adult SSC subpopulation marked specifically by forkhead box protein C2 (FOXC2). All spermatogenic progenies can be derived from FOXC2+ SSCs and the ablation of FOXC2+ SSCs led to the depletion of the undifferentiated spermatogonia pool. During germline regeneration, FOXC2+ SSCs were activated and able to completely restore the process. Germ cell-specific Foxc2 knockout resulted in an accelerated exhaustion of SSCs and eventually led to male infertility. Furthermore, FOXC2 prompts the expressions of negative regulators of cell cycle thereby ensures the SSCs reside in quiescence. Thus, this work proposes that the quiescent FOXC2+ SSCs are essential for maintaining the homeostasis and regeneration of spermatogenesis in adult mammals.

Clinicopathological features, treatment modalities, and prognosis of esophageal neuroendocrine carcinoma: A single-center retrospective study.

OBJECTIVES: Esophageal neuroendocrine carcinoma (ENEC) is a rare cancer that is highly malignant and related to a poor prognosis. In this retrospective study we aimed to elucidate the clinical characteristics, diagnosis and management of patients with ENEC and to evaluate the potential prognostic factors. METHODS: Altogether 82 patients diagnosed with ENEC between January 2009 and December 2020 at the Fudan University Shanghai Cancer Center were retrospectively enrolled. Patients' survival was analyzed using the Kaplan-Meier and log-rank methods. Univariate and multivariate analyses and a Cox regression model were used to identify the prognostic factors. RESULTS: The median overall survival (mOS) was 13 months in all patients. Multivariate analysis revealed that advanced tumor stage (hazard ratio [HR] 2.67, 95% confidence interval [CI] 1.07-6.66, P = 0.0353), liver (HR 3.36, 95% CI 1.53-7.41, P = 0.0026) and lung metastasis (HR 3.37, 95% CI 1.20-9.51, P = 0.0214) were associated with a poor prognosis. While positive chromogranin A (CgA) expression was related to a favorable outcome (HR 0.21, 95% CI 0.09-0.49, P < 0.001). Also, patients had adjustment of chemotherapy (dose reduction or less than three cycles) were prone to a worse prognosis compared with those did not (HR 4.36, 95% CI 2.10-9.08, P < 0.001). CONCLUSION: In patients with ENEC, advanced cancer stage, adjustment of chemotherapy, liver and lung metastasis were associated with a poor survival, while CgA expression was related to a favorable prognosis.

Decoding the spermatogonial stem cell niche under physiological and recovery conditions in adult mice and humans.

The intricate interaction between spermatogonial stem cell (SSC) and testicular niche is essential for maintaining SSC homeostasis; however, this interaction remains largely uncharacterized. In this study, to characterize the underlying signaling pathways and related paracrine factors, we delineated the intercellular interactions between SSC and niche cell in both adult mice and humans under physiological conditions and dissected the niche-derived regulation of SSC maintenance under recovery conditions, thus uncovering the essential role of C-C motif chemokine ligand 24 and insulin-like growth factor binding protein 7 in SSC maintenance. We also established the clinical relevance of specific paracrine factors in human fertility. Collectively, our work on decoding the adult SSC niche serves as a valuable reference for future studies on the aetiology, diagnosis, and treatment of male infertility.

Reactive oxygen species may be involved in the distinctive biological effects of different doses of 12C6+ ion beams on Arabidopsis.

Introduction: Heavy ion beam is a novel approach for crop mutagenesis with the advantage of high energy transfer line density and low repair effect after injury, however, little investigation on the biological effect on plant was performed. 50 Gy irradiation significantly stimulated the growth of Arabidopsis seedlings, as indicated by an increase in root and biomass, while 200 Gy irradiation significantly inhibited the growth of seedlings, causing a visible decrease in plant growth. Methods: The Arabidopsis seeds were irradiated by 12C6+. Monte Carlo simulations were used to calculate the damage to seeds and particle trajectories by ion implantation. The seed epidermis received SEM detection and changes in its organic composition were detected using FTIR. Evidence of ROS and antioxidant systems were analyzed. RNA-seq and qPCR were used to detect changes in seedling transcript levels. Results and discussion: Monte Carlo simulations revealed that high-dose irradiation causes various damage. Evidence of ROS and antioxidant systems implies that the emergence of phenotypes in plant cells may be associated with oxidative stress. Transcriptomic analysis of the seedlings demonstrated that 170 DEGs were present in the 50 Gy and 200 Gy groups and GO enrichment indicated that they were mainly associated with stress resistance and cell wall homeostasis. Further GO enrichment of DEGs unique to 50 Gy and 200 Gy revealed 58 50Gy-exclusive DEGs were enriched in response to oxidative stress and jasmonic acid entries, while 435 200 Gy-exclusive DEGs were enriched in relation to oxidative stress, organic cyclic compounds, and salicylic acid. This investigation advances our insight into the biological effects of heavy ion irradiation and the underlying mechanisms.

RING finger 138 deregulation distorts NF-кB signaling and facilities colitis switch to aggressive malignancy.

Prolonged activation of nuclear factor (NF)-кB signaling significantly contributes to the development of colorectal cancer (CRC). New therapeutic opportunities are emerging from targeting this distorted cell signaling transduction. Here, we discovered the critical role of RING finger 138 (RNF138) in CRC tumorigenesis through regulating the NF-кB signaling, which is independent of its Ubiquitin-E3 ligase activity involved in DNA damage response. RNF138-/- mice were hyper-susceptible to the switch from colitis to aggressive malignancy, which coincided with sustained aberrant NF-кB signaling in the colonic cells. Furthermore, RNF138 suppresses the activation of NF-кB signaling pathway through preventing the translocation of NIK and IKK-Beta Binding Protein (NIBP) to the cytoplasm, which requires the ubiquitin interaction motif (UIM) domain. More importantly, we uncovered a significant correlation between poor prognosis and the downregulation of RNF138 associated with reinforced NF-кB signaling in clinical settings, raising the possibility of RNF138 dysregulation as an indicator for the therapeutic intervention targeting NF-кB signaling. Using the xenograft models built upon either RNF138-dificient CRC cells or the cells derived from the RNF138-dysregulated CRC patients, we demonstrated that the inhibition of NF-кB signaling effectively hampered tumor growth. Overall, our work defined the pathogenic role of aberrant NF-кB signaling due to RNF138 downregulation in the cascade events from the colitis switch to colonic neoplastic transformation and progression, and also highlights the possibility of targeting the NF-кB signaling in treating specific subtypes of CRC indicated by RNF138-ablation.

FMRP ligand circZNF609 destabilizes RAC1 mRNA to reduce metastasis in acral melanoma and cutaneous melanoma.

BACKGROUND: Melanoma is a type of malignant tumor with high aggressiveness and poor prognosis. At present, metastasis of melanoma is still an important cause of death in melanoma patients. However, the potential functions and molecular mechanisms of most circular RNAs (circRNAs) in melanoma metastasis remain unknown. METHODS: circRNAs dysregulated in melanoma cell subgroups with different metastatic abilities according to a screening model based on repeated Transwell assays were identified with a circRNA array. The expression and prognostic significance of circZNF609 in skin cutaneous melanoma and acral melanoma cells and tissues were determined by qRT-PCR, nucleoplasmic separation assays and fluorescence in situ hybridization. In vitro wound healing, Transwell and 3D invasion assays were used to analyse melanoma cell metastasis ability. Tail vein injection and intrasplenic injection were used to study in vivo lung metastasis and liver metastasis, respectively. The mechanism of circZNF609 was further evaluated via RNA immunoprecipitation, RNA pull-down, silver staining, and immunofluorescence colocalization assays. RESULTS: circZNF609 was stably expressed at low levels in melanoma tissues and cells and was negatively correlated with Breslow depth, clinical stage and prognosis of melanoma patients. circZNF609 inhibited metastasis of acral and cutaneous melanoma in vivo and in vitro. Mechanistically, circZNF609 promoted the binding of FMRP protein and RAC1 mRNA, thereby enhancing the inhibitory effect of FMRP protein on the stability of RAC1 mRNA and ultimately inhibiting melanoma metastasis. CONCLUSIONS: Our findings revealed that circZNF609 plays a vital role in the metastasis of acral and cutaneous melanoma through the circRNF609-FMRP-RAC1 axis and indicated that circZNF609 regulates the stability of RAC1 mRNA by combining with FMRP, which might provide insight into melanoma pathogenesis and a new potential target for treatment of melanoma.

Sensing of cytoplasmic chromatin by cGAS activates innate immune response in SARS-CoV-2 infection.

The global coronavirus disease 2019 (COVID-19) pandemic is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a positive-sense RNA virus. How the host immune system senses and responds to SARS-CoV-2 infection remain largely unresolved. Here, we report that SARS-CoV-2 infection activates the innate immune response through the cytosolic DNA sensing cGAS-STING pathway. SARS-CoV-2 infection induces the cellular level of 2'3'-cGAMP associated with STING activation. cGAS recognizes chromatin DNA shuttled from the nucleus as a result of cell-to-cell fusion upon SARS-CoV-2 infection. We further demonstrate that the expression of spike protein from SARS-CoV-2 and ACE2 from host cells is sufficient to trigger cytoplasmic chromatin upon cell fusion. Furthermore, cytoplasmic chromatin-cGAS-STING pathway, but not MAVS-mediated viral RNA sensing pathway, contributes to interferon and pro-inflammatory gene expression upon cell fusion. Finally, we show that cGAS is required for host antiviral responses against SARS-CoV-2, and a STING-activating compound potently inhibits viral replication. Together, our study reported a previously unappreciated mechanism by which the host innate immune system responds to SARS-CoV-2 infection, mediated by cytoplasmic chromatin from the infected cells. Targeting the cytoplasmic chromatin-cGAS-STING pathway may offer novel therapeutic opportunities in treating COVID-19. In addition, these findings extend our knowledge in host defense against viral infection by showing that host cells' self-nucleic acids can be employed as a "danger signal" to alarm the immune system.

Structure-based design and characterization of novel fusion-inhibitory lipopeptides against SARS-CoV-2 and emerging variants.

The ongoing pandemic of COVID-19, caused by SARS-CoV-2, has severely impacted the global public health and socio-economic stability, calling for effective vaccines and therapeutics. In this study, we continued our efforts to develop more efficient SARS-CoV-2 fusion inhibitors and achieved significant findings. First, we found that the membrane-proximal external region (MPER) sequence of SARS-CoV-2 spike fusion protein plays a critical role in viral infectivity and can serve as an ideal template for design of fusion-inhibitory peptides. Second, a panel of novel lipopeptides was generated with greatly improved activity in inhibiting SARS-CoV-2 fusion and infection. Third, we showed that the new inhibitors maintained the potent inhibitory activity against emerging SARS-CoV-2 variants, including those with the major mutations of the B.1.1.7 and B.1.351 strains circulating in the United Kingdom and South Africa, respectively. Fourth, the new inhibitors also cross-inhibited other human CoVs, including SARS-CoV, MERS-CoV, HCoV-229E, and HCoV-NL63. Fifth, the structural properties of the new inhibitors were characterized by circular dichroism (CD) spectroscopy and crystallographic approach, which revealed the mechanisms underlying the high binding and inhibition. Combined, our studies provide important information for understanding the mechanism of SARS-CoV-2 fusion and a framework for the development of peptide therapeutics for the treatment of SARS-CoV-2 and other CoVs.

Corilagin inhibits SARS-CoV-2 replication by targeting viral RNA-dependent RNA polymerase.

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has become one major threat to human population health. The RNA-dependent RNA polymerase (RdRp) presents an ideal target of antivirals, whereas nucleoside analogs inhibitor is hindered by the proofreading activity of coronavirus. Herein, we report that corilagin (RAI-S-37) as a non-nucleoside inhibitor of SARS-CoV-2 RdRp, binds directly to RdRp, effectively inhibits the polymerase activity in both cell-free and cell-based assays, fully resists the proofreading activity and potently inhibits SARS-CoV-2 infection with a low 50% effective concentration (EC50) value of 0.13 µmol/L. Computation modeling predicts that RAI-S-37 lands at the palm domain of RdRp and prevents conformational changes required for nucleotide incorporation by RdRp. In addition, combination of RAI-S-37 with remdesivir exhibits additive activity against anti-SARS-CoV-2 RdRp. Together with the current data available on the safety and pharmacokinetics of corilagin as a medicinal herbal agent, these results demonstrate the potential of being developed into one of the much-needed SARS-CoV-2 therapeutics.

Presence of Anti-MDA5 Antibody and Its Value for the Clinical Assessment in Patients With COVID-19: A Retrospective Cohort Study.

Background: Striking similarities have been found between coronavirus disease 2019 (COVID-19) and anti-melanoma differentiation-associated gene 5 (MDA5) antibody (Ab)-related dermatomyositis, implying a shared autoinflammatory aberrance. Herein, we aim to investigate whether the anti-MDA5 Ab is present in COVID-19 and correlates with the severity and adverse outcome of COVID-19 patients. Methods and Findings: We retrospectively recruited 274 adult inpatients with COVID-19 in this study, including 48, 164, and 62 cases of deaths, severe, and non-severe patients respectively. The anti-MDA5 Ab was determined by ELISA and verified by Western Blotting, which indicated that the positive rate of anti-MDA5 Ab in COVID-19 patients was 48.2% (132/274). The clinical and laboratory features, as well as outcomes between patients with positive and negative anti-MDA5 Ab were compared and we found that the anti-MDA5 Ab positive patients tended to represent severe disease (88.6% vs 66.9%, P<0.0001). We also demonstrated that the titer of anti-MDA5 Ab was significantly elevated in the non-survivals (5.95 ± 5.16 vs 8.22 ± 6.64, P=0.030) and the positive rate was also higher than that in the survivals (23.5% vs 12.0%, P=0.012). Regarding severe COVID-19 patients, we found that high titer of anti-MDA5 Ab (≥10.0 U/mL) was more prevalent in the non-survivals (31.2% vs 14.0%, P=0.006). Moreover, a dynamic analysis of anti-MDA5 Ab was conducted at different time-points of COVID-19, which revealed that early profiling of anti-MDA5 Ab could distinguish severe patients from those with non-severe ones. Conclusions: Anti-MDA5 Ab was prevalent in the COVID-19 patients and high titer of this antibody is correlated with severe disease and unfavorable outcomes.

Identification of Potent and Safe Antiviral Therapeutic Candidates Against SARS-CoV-2.

COVID-19 pandemic has infected millions of people with mortality exceeding >1 million. There is an urgent need to find therapeutic agents that can help clear the virus to prevent severe disease and death. Identifying effective and safer drugs can provide more options to treat COVID-19 infections either alone or in combination. Here, we performed a high throughput screening of approximately 1,700 US FDA-approved compounds to identify novel therapeutic agents that can effectively inhibit replication of coronaviruses including SARS-CoV-2. Our two-step screen first used a human coronavirus strain OC43 to identify compounds with anti-coronaviral activities. The effective compounds were then screened for their effectiveness in inhibiting SARS-CoV-2. These screens have identified 20 anti-SARS-CoV-2 drugs including previously reported compounds such as hydroxychloroquine, amlodipine besylate, arbidol hydrochloride, tilorone 2HCl, dronedarone hydrochloride, mefloquine, and thioridazine hydrochloride. Five of the newly identified drugs had a safety index (cytotoxic/effective concentration) of >600, indicating a wide therapeutic window compared to hydroxychloroquine which had a safety index of 22 in similar experiments. Mechanistically, five of the effective compounds (fendiline HCl, monensin sodium salt, vortioxetine, sertraline HCl, and salifungin) were found to block SARS-CoV-2 S protein-mediated cell fusion. These FDA-approved compounds can provide much needed therapeutic options that we urgently need during the midst of the pandemic.

P53-R273H mutation enhances colorectal cancer stemness through regulating specific lncRNAs.

BACKGROUND: TP53 is one of the most frequently mutated genes among all cancer types, and TP53 mutants occur more than 60% in colorectal cancer (CRC). Among all mutants, there are three hot spots, including p53-R175H, p53-R248W and p53-R273H. Emerging evidence attributes cancer carcinogenesis to cancer stem cells (CSCs). Long noncoding RNAs (lncRNAs) play crucial roles in maintaining the stemness of CSCs. However, it is unknown if mutant p53-regulated lncRNAs are implicated in the maintenance of CSC stemness. METHODS: RNA-sequencing (RNA-seq) and ChIP-sequencing (ChIP-seq) were used to trace the lncRNA network regulated by p53-R273H in HCT116 endogenous p53 point mutant spheroid cells generated by the somatic cell knock-in method. RT-qPCR was used to detect lncRNA expression patterns, verifying the bioinformatics analysis. Transwell, spheroid formation, fluorescence activated cell sorter (FACS), xenograft nude mouse model, tumor frequency assessed by extreme limiting dilution analysis (ELDA), Western blot assays and chemoresistance analysis were performed to elucidate the functions and possible mechanism of lnc273-31 and lnc273-34 in cancer stem cells. RESULTS: p53-R273H exhibited more characteristics of CSC than p53-R175H and p53-R248W. RNA-seq profiling identified 37 up regulated and 4 down regulated differentially expressed lncRNAs regulated by p53-R273H. Combined with ChIP-seq profiling, we further verified two lncRNAs, named as lnc273-31 and lnc273-34, were essential in the maintenance of CSC stemness. Further investigation illustrated that lnc273-31 or lnc273-34 depletion dramatically diminished colorectal cancer migration, invasion, cancer stem cell self-renewal and chemoresistance in vitro. Moreover, the absence of lnc273-31 or lnc273-34 dramatically delayed cancer initiation and tumorigenic cell frequency in vivo. Also, lnc273-31 and lnc273-34 have an impact on epithelial-to mesenchymal transition (EMT). Finally, lnc273-31 and lnc273-34 were significantly highly expressed in CRC tissues with p53-R273H mutation compared to those with wildtype p53. CONCLUSIONS: The present study unveiled a high-confidence set of lncRNAs regulated by p53-R273H specific in colorectal CSCs. Furthermore, we demonstrated that two of them, lnc273-31 and lnc273-34, were required for colorectal CSC self-renewal, tumor propagation and chemoresistance. Also, the expression of these two lncRNAs augmented in colorectal cancer patient samples with p53-R273H mutation. These two lncRNAs may serve as promising predictors for patients with p53-R273H mutation and are vital for chemotherapy.

Correction to: RHBDD1 promotes colorectal cancer metastasis through the Wnt signaling pathway and its downstream target ZEB1.

In the publication of this article [1], there is an error in the Methods section at paragraph Cell culture and reagents and the Additional file2: Fig. S1 was erroneous linked.

RHBDD1 promotes colorectal cancer metastasis through the Wnt signaling pathway and its downstream target ZEB1.

BACKGROUND: 40-50% of colorectal cancer (CRC) patients develop metastatic disease; the presence of metastasis hinders the effective treatment of cancer through surgery, chemotherapy and radiotherapy, which makes 5-year survival rate extremely low; therefore, studying CRC metastasis is crucial for disease therapy. In the present study, we investigated the role of rhomboid domain containing 1 (RHBDD1) in tumor metastasis of CRC. METHODS: The expression of RHBDD1 was analyzed in 539 colorectal tumor tissues for its correlation with lymphatic metastasis and distal metastasis. Transwell assay in vitro and pleural metastasis analysis in vivo were performed to determine the functions of RHBDD1 during CRC cells metastasis. RNA-seq analysis, TOP/FOP flash reporter assay, western blot and transwell assay were performed to investigate the underlying mechanism for the function of RHBDD1 on Wnt signaling pathway. Bioinformatics analysis was conducted to investigate epithelial-mesenchymal transition (EMT) and stemness in HCT-116 cells. Tissue microarray analysis, Q-PCR and western blot were performed to determine the correlation of RHBDD1 and Zinc Finger E-Box Binding Homeobox 1 (ZEB1). RESULTS: In this study, we found that RHBDD1 expression was positively correlated with lymphatic metastasis and distal metastasis in 539 colorectal tumor tissues. RHBDD1 expression can promote CRC cells metastasis in vitro and in vivo. RNA-Seq analysis showed that the Wnt signaling pathway played a key role in this metastatic regulation. RHBDD1 mainly regulated ser552 and ser675 phosphorylation of ß-catenin to activate the Wnt signaling pathway. Rescuing ser552 and ser675 phosphorylation of ß-catenin resulted in the recovery of signaling pathway activity, migration, and invasion in CRC cells. RHBDD1 promoted EMT and a stem-like phenotype of CRC cells. RHBDD1 regulated the Wnt/ß-catenin target gene ZEB1, a potent EMT activator, at the RNA and protein levels. Clinically, RHBDD1 expression was positively correlated with ZEB1 at the protein level in 71 colon tumor tissues. CONCLUSIONS: Our findings therefore indicated that RHBDD1 can promote CRC metastasis through the Wnt signaling pathway and ZEB1. RHBDD1 may become a new therapeutic target or clinical biomarker for metastatic CRC.

Pancreatic Cancer Progression Relies upon Mutant p53-Induced Oncogenic Signaling Mediated by NOP14.

Mutant p53 (mutp53) proteins promote tumor invasion and metastasis in pancreatic ductal adenocarcinoma (PDAC). However, the mechanism underlying sustained activation of mutp53 oncogenic signaling is currently unclear. In this study, we report that NOP14 nucleolar protein (NOP14) expression is upregulated in PDAC tumors and metastatic tissue specimens. NOP14 overexpression promoted cell motility, whereas NOP14 inhibition decreased invasive capacity of PDAC cells. In vivo invasion assays conducted on established subcutaneously, orthotopically, and intravenously injected tumor mouse models also indicated NOP14 as a promoter of PDAC metastasis. Mechanistically, mutp53 was validated as a functional target of NOP14; NOP14 primed tumor invasion and metastasis by increasing the stability of mutp53 mRNA. The NOP14/mutp53 axis suppressed p21 expression at both the transcriptional and posttranscriptional levels via induction of miR-17-5p in PDAC cells. In vivo, high NOP14 expression in PDAC patient tumors correlated with local metastasis and lymph invasion. Overall, our findings define a novel mechanism for understanding the function of NOP14 in the metastatic cascade of PDAC. Targeting NOP14 allows for effective suppression of tumor invasion in a mutp53-dependent manner, implicating NOP14 inhibition as a potential approach for attenuating metastasis in p53-mutant tumors. Cancer Res; 77(10); 2661-73. ©2017 AACR.

The hepatic FOXQ1 transcription factor regulates glucose metabolism in mice.

AIM/HYPOTHESIS: Hepatic forkhead box q1 (FOXQ1) expression levels are regulated by nutritional and pathophysiological status. In this study we investigated the role of FOXQ1 in the regulation of hepatic gluconeogenesis. METHODS: We used multiple mouse and cell models to study the role of FOXQ1 in regulating expression of gluconeogenic genes, and cellular and hepatic glucose production. RESULTS: Expression of hepatic FOXQ1 was regulated by fasting in normal mice and was dysregulated in diabetic mice. Overexpression of FOXQ1 in primary hepatocytes inhibited expression of gluconeogenic genes and decreased cellular glucose output. Hepatic FOXQ1 rescue in db/db and high-fat diet-induced obese mice markedly decreased blood glucose level and improved glucose intolerance. In contrast, wild-type C57 mice with hepatic FOXQ1 deficiency displayed increased blood glucose levels and impaired glucose tolerance. Interestingly, studies into molecular mechanisms indicated that FOXQ1 interacts with FOXO1, thereby blocking FOXO1 activity on hepatic gluconeogenesis, preventing it from directly binding to insulin response elements mapped in the promoter region of gluconeogenic genes. CONCLUSIONS/INTERPRETATION: FOXQ1 is a novel factor involved in regulating hepatic gluconeogenesis, and the decreased FOXQ1 expression in liver may contribute to the development of type 2 diabetes.

Forkhead Box P1 (FOXP1) Transcription Factor Regulates Hepatic Glucose Homeostasis.

Dysregulation of hepatic gluconeogenesis contributes to the pathogenesis of diabetes, yet the detailed molecular mechanisms remain to be fully elucidated. Here we show that FOXP1, a transcriptional repressor, plays a key role in the regulation of systemic glucose homeostasis. Hepatic expression levels of FOXP1 are decreased in diabetic mice. Modest hepatic overexpression of FOXP1 in mice inhibited the expression of gluconeogenic genes, such as peroxisome proliferators-activated receptor γ coactivator-1α (PGC-1α), phosphoenolpyruvate carboxykinase (PEPCK), and glucose-6-phosphatase (G6PC), leading to a decrease in hepatic glucose production and fasting blood glucose levels in normal mice and different mouse models of diabetes, including db/db diabetic and high-fat diet-induced obese mice. FOXP1 physically interacted with FOXO1 in vivo and competed with FOXO1 for binding to the insulin response element in the promoter region of gluconeogenic genes, thereby interfering expression of these genes. These results identify a previously unrecognized role for FOXP1 in the transcriptional control of hepatic glucose homeostasis.

Sirt1 mediates the effect of the heme oxygenase inducer, cobalt protoporphyrin, on ameliorating liver metabolic damage caused by a high-fat diet.

BACKGROUND & AIMS: Heme oxygenase 1 (HO-1)-mediated increases in adiponectin, ameliorate the deleterious effects of obesity and metabolic syndrome; however, the effect of HO-1 on hepatic lipid metabolism remains elusive. The aim of this study is to evaluate the role of HO-1 in hepatic lipid metabolism. METHODS: Functional studies were performed using C57BL/6J (WT) mice and Sirt1 liver specific mutant (Sirt1-deficient) mice. The molecular mechanism was explored in primary hepatocytes and mouse liver. RESULTS: Chronic exposure to high-fat diet (HFD) induced hepatic steatosis in WT mice. Treatment of WT mice on HFD with cobalt protoporphyrin (CoPP), an inducer of HO-1 activity, decreased body weight and visceral fat content, reduced intracellular hepatic triglyceride and serum total cholesterol concentrations, and decreased liver lipid droplet formation. Compared with WT mice, the administration of CoPP to Sirt1-deficient mice on HFD increased visceral fat content, and slightly promoted liver lipid droplet formation. CoPP improved glucose tolerance and insulin sensitivity in WT mice on HFD, but compromised insulin sensitivity in Sirt1-deficient mice on HFD. Furthermore, CoPP-induced Sirt1 expression and decreased sterol regulatory element binding protein 1c (SREBP-1c) expression in WT mice on HFD. However, CoPP promoted SREBP-1c expression in Sirt1-deficient hepatocytes, which was reversed by a protein tyrosine phosphatase 1b inhibitor. Additionally, while the administration of CoPP to WT mice on HFD improved antioxidant and anti-inflammatory states, these CoPP-mediated effects were abolished in Sirt1-deficient mice. CONCLUSIONS: Sirt1 mediates the effect of CoPP on ameliorating liver metabolic damage caused by HFD.

Involvement of KLF11 in hepatic glucose metabolism in mice via suppressing of PEPCK-C expression.

BACKGROUND: Abnormal hepatic gluconeogenesis is related to hyperglycemia in mammals with insulin resistance. Despite the strong evidences linking Krüppel-like factor 11 (KLF11) gene mutations to development of Type 2 diabetes, the precise physiological functions of KLF11 in vivo remain largely unknown. RESULTS: In current investigation, we showed that KLF11 is involved in modulating hepatic glucose metabolism in mice. Overexpression of KLF11 in primary mouse hepatocytes could inhibit the expression of gluconeogenic genes, including phosphoenolpyruvate carboxykinase (cytosolic isoform, PEPCK-C) and peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α), subsequently decreasing the cellular glucose output. Diabetic mice with overexpression of KLF11 gene in livers significantly ameliorated hyperglycemia and glucose intolerance; in contrast, the knockdown of KLF11 expression in db/m and C57BL/6J mice livers impaired glucose tolerance. CONCLUSIONS: Our data strongly indicated the involvement of KLF11 in hepatic glucose homeostasis via modulating the expression of PEPCK-C.