Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 20 de 20
Filter
1.
PLoS One ; 16(9): e0256979, 2021.
Article in English | MEDLINE | ID: mdl-34478472

ABSTRACT

PURPOSES: Although clinical and radiological examinations can be used to diagnose oral cancer, and surgical pathology remains the gold standard, these conventional methods have limitations. We evaluated the feasibility of longitudinal next-generation sequencing-based liquid biopsy for oral squamous cell carcinoma surveillance. MATERIALS AND METHODS: Eleven patients were enrolled, and plasma and saliva were collected before, and 1, 3, and 6 months after surgery. Tumor-specific mutations were selected using paired, whole-exome analyses of tumor tissues and whole blood. Genes frequently mutated in head and neck cancer were identified using the Cancer Genome Atlas (TCGA) and Catalogue of Somatic Mutations in Cancer (COSMIC) databases to design targeted deep sequencing panels. RESULTS: In five of the six patients with recurrent cancer, circulating tumor DNA (ctDNA) was detected earlier with liquid biopsy than with conventional monitoring techniques. Moreover, patients without recurrence exhibited decreased ctDNA allele frequency post-treatment. CONCLUSIONS: Longitudinal liquid biopsy of plasma and saliva may be feasible for detecting somatic mutations associated with oral squamous cell carcinomas. It might be attributable to determine early tumor recurrence through genetic analysis of ctDNA.


Subject(s)
Carcinoma, Squamous Cell , Circulating Tumor DNA/metabolism , Liquid Biopsy/methods , Mouth Neoplasms , Aged , Biomarkers, Tumor/metabolism , Carcinoma, Squamous Cell/diagnosis , Carcinoma, Squamous Cell/metabolism , Female , Humans , Longitudinal Studies , Male , Middle Aged , Mouth Neoplasms/diagnosis , Mouth Neoplasms/metabolism , Neoplasm Recurrence, Local , Saliva/metabolism
2.
Life Sci Alliance ; 3(7)2020 07.
Article in English | MEDLINE | ID: mdl-32487689

ABSTRACT

Despite the importance of mitochondrial fatty acid oxidation (FAO) in cancer metabolism, the biological mechanisms responsible for the FAO in cancer and therapeutic intervention based on catabolic metabolism are not well defined. In this study, we observe that Snail (SNAI1), a key transcriptional repressor of epithelial-mesenchymal transition, enhances catabolic FAO, allowing pro-survival of breast cancer cells in a starved environment. Mechanistically, Snail suppresses mitochondrial ACC2 (ACACB) by binding to a series of E-boxes located in its proximal promoter, resulting in decreased malonyl-CoA level. Malonyl-CoA being a well-known endogenous inhibitor of fatty acid transporter carnitine palmitoyltransferase 1 (CPT1), the suppression of ACC2 by Snail activates CPT1-dependent FAO, generating ATP and decreasing NADPH consumption. Importantly, combinatorial pharmacologic inhibition of pentose phosphate pathway and FAO with clinically available drugs efficiently reverts Snail-mediated metabolic reprogramming and suppresses in vivo metastatic progression of breast cancer cells. Our observations provide not only a mechanistic link between epithelial-mesenchymal transition and catabolic rewiring but also a novel catabolism-based therapeutic approach for inhibition of cancer progression.


Subject(s)
Acetyl-CoA Carboxylase/genetics , Fatty Acids/metabolism , Genes, Mitochondrial/genetics , Neoplasms/genetics , Neoplasms/metabolism , Oxidation-Reduction , Snail Family Transcription Factors/metabolism , Acetyl-CoA Carboxylase/metabolism , Adenosine Triphosphate/metabolism , Animals , Cell Line, Tumor , Cell Survival/genetics , Energy Metabolism , Gene Expression Regulation, Neoplastic , Glucose/metabolism , Humans , Lipid Metabolism , Metabolic Networks and Pathways , Metabolome , Metabolomics , Mice , Neoplasms/pathology
3.
Nat Commun ; 9(1): 2301, 2018 06 12.
Article in English | MEDLINE | ID: mdl-29895829

ABSTRACT

Phosphorylation-dependent YAP translocation is a well-known intracellular mechanism of the Hippo pathway; however, the molecular effectors governing YAP cytoplasmic translocation remains undefined. Recent findings indicate that oncogenic YAP paradoxically suppresses Wnt activity. Here, we show that Wnt scaffolding protein Dishevelled (DVL) is responsible for cytosolic translocation of phosphorylated YAP. Mutational inactivation of the nuclear export signal embedded in DVL leads to nuclear YAP retention, with an increase in TEAD transcriptional activity. DVL is also required for YAP subcellular localization induced by E-cadherin, α-catenin, or AMPK activation. Importantly, the nuclear-cytoplasmic trafficking is dependent on the p53-Lats2 or LKB1-AMPK tumor suppressor axes, which determine YAP phosphorylation status. In vivo and clinical data support that the loss of p53 or LKB1 relieves DVL-linked reciprocal inhibition between the Wnt and nuclear YAP activity. Our observations provide mechanistic insights into controlled proliferation coupled with epithelial polarity during development and human cancer.


Subject(s)
Active Transport, Cell Nucleus , Adaptor Proteins, Signal Transducing/metabolism , Dishevelled Proteins/metabolism , Genes, Tumor Suppressor , Phosphoproteins/metabolism , A549 Cells , AMP-Activated Protein Kinase Kinases , AMP-Activated Protein Kinases/metabolism , Animals , Cadherins/metabolism , Cell Cycle Proteins , Cell Nucleus/metabolism , Cytoplasm/metabolism , DNA Mutational Analysis , Female , HCT116 Cells , HEK293 Cells , Hippo Signaling Pathway , Humans , MCF-7 Cells , Mice , Mice, Nude , Mutation , Phosphorylation , Protein Serine-Threonine Kinases/metabolism , Protein Transport , Transcription Factors , Tumor Suppressor Protein p53/metabolism , Wnt Proteins/metabolism , Wnt1 Protein/metabolism , YAP-Signaling Proteins , alpha Catenin/metabolism
4.
Genome Res ; 2018 Jan 11.
Article in English | MEDLINE | ID: mdl-29326299

ABSTRACT

KRAS is the most frequently mutated oncogene in human tumors, and its activating mutations represent important therapeutic targets. The combination of Cas9 and guide RNA from the CRISPR-Cas system recognizes a specific DNA sequence and makes a double-strand break, which enables editing of the relevant genes. Here, we harnessed CRISPR to specifically target mutant KRAS alleles in cancer cells. We screened guide RNAs using a reporter system and validated them in cancer cells after lentiviral delivery of Cas9 and guide RNA. The survival, proliferation, and tumorigenicity of cancer cells in vitro and the growth of tumors in vivo were determined after delivery of Cas9 and guide RNA. We identified guide RNAs that efficiently target mutant KRAS without significant alterations of the wild-type allele. Doxycycline-inducible expression of this guide RNA in KRAS-mutant cancer cells transduced with a lentiviral vector encoding Cas9 disrupted the mutant KRAS gene, leading to inhibition of cancer cell proliferation both in vitro and in vivo. Intra-tumoral injection of lentivirus and adeno-associated virus expressing Cas9 and sgRNA suppressed tumor growth in vivo, albeit incompletely, in immunodeficient mice. Expression of Cas9 and the guide RNA in cells containing wild-type KRAS did not alter cell survival or proliferation either in vitro and in vivo. Our study provides a proof-of-concept that CRISPR can be utilized to target driver mutations of cancers in vitro and in vivo.

5.
Biomol Ther (Seoul) ; 26(1): 29-38, 2018 Jan 01.
Article in English | MEDLINE | ID: mdl-29212304

ABSTRACT

During cancer progression, cancer cells are repeatedly exposed to metabolic stress conditions in a resource-limited environment which they must escape. Increasing evidence indicates the importance of nicotinamide adenine dinucleotide phosphate (NADPH) homeostasis in the survival of cancer cells under metabolic stress conditions, such as metabolic resource limitation and therapeutic intervention. NADPH is essential for scavenging of reactive oxygen species (ROS) mainly derived from oxidative phosphorylation required for ATP generation. Thus, metabolic reprogramming of NADPH homeostasis is an important step in cancer progression as well as in combinational therapeutic approaches. In mammalian, the pentose phosphate pathway (PPP) and one-carbon metabolism are major sources of NADPH production. In this review, we focus on the importance of glucose flux control towards PPP regulated by oncogenic pathways and the potential therein for metabolic targeting as a cancer therapy. We also summarize the role of Snail (Snai1), an important regulator of the epithelial mesenchymal transition (EMT), in controlling glucose flux towards PPP and thus potentiating cancer cell survival under oxidative and metabolic stress.

6.
Oral Oncol ; 74: 62-67, 2017 11.
Article in English | MEDLINE | ID: mdl-29103753

ABSTRACT

OBJECTIVES: Odontogenic keratocyst (OKC), also known as keratocystic odontogenic tumor (KCOT), has clinical significance due to its high incidence as well as high recurrence rate after surgical enucleation. Current clinical management for OCK is entirely dependent on surgical approach. While various genetic alterations, such as PTCH1 mutation and loss of heterozygosity in tumor suppressor genes, have been reported, the molecular background of OKC is not well-understood. Although recent identification of BRAF V600E mutation and subsequent activation of mitogen-activated protein kinase (MAPK) pathway in ameloblastoma and odontogenic tumors provide additional options with targeted therapeutics, the molecular background of OKC is not well understood. MATERIALS AND METHODS: In this study, we examined BRAF V600E mutation from paraffin embedded OKC samples by tumor cell enriched microdissection and TA cloning of amplified DNA. We further examined the relationship between BRAF V600E mutation and clinical parameters. RESULTS: We found frequent BRAF V600E mutation in OKC (24 of 38 samples, 63.2%). However, BRAF V600E mutational status is not related with clinical indexes such as size, location, and recurrence. In orthokeratinized odontogenic cyst, there is one case of BRAF 600E mutation from 11 samples (9.1%). CONCLUSION: These results indicate that BRAF V600E mutation occurs in OKCs at a high rate and plays an important role in the pathogenesis of OKCs.


Subject(s)
Odontogenic Cysts/genetics , Odontogenic Tumors/genetics , Point Mutation , Proto-Oncogene Proteins B-raf/genetics , Adult , Female , Humans , Male , Middle Aged , Neoplasm Recurrence, Local , Odontogenic Cysts/pathology , Odontogenic Tumors/pathology , Paraffin Embedding , Polymerase Chain Reaction , Young Adult
7.
Sci Rep ; 7(1): 14952, 2017 11 02.
Article in English | MEDLINE | ID: mdl-29097790

ABSTRACT

Injury to the external branch of the superior laryngeal nerve (eSLN) can cause a hoarse or weak voice with dysergia of the cricothyroid. The present study provided the topographic information of the eSLN in the Asian and verified anatomical validity of the landmarks previously recruited to localize the eSLN. Thirty specimens were dissected from 16 human embalmed cadavers (12 men and four women; mean age: 80.5 years). The vertical distance between the eSLN and the apical pole of the thyroid gland (AP) was 8.2 ± 4.2 mm. It descended over the AP with <1 cm distance in 51.7%, >1 cm distance in 27.6% and under the AP in 20.7%. The piercing point (PP) of the eSLN to the muscles located 26.0 ± 5.5 mm posterior and 14.7 ± 5.0 mm inferior to the laryngeal prominence. Generally, the PP located superoposterior to the midpoint of the joint between the joint of inferior constrictor and cricothyroid (ICJ). The distance between the PP and the midpoint was 8.7 ± 5.1 mm. We found that 1) the Asian had the eSLN located over the AP with <1 cm distance about half cases, 2) the PP can be a consistent reference for the eSLN identification, 3) the ICJ can be a useful landmark to preserve the eSLN at the PP.


Subject(s)
Laryngeal Nerves/anatomy & histology , Aged, 80 and over , Asian People , Cadaver , Dissection , Female , Humans , Male
8.
Br J Cancer ; 117(12): 1798-1809, 2017 Dec 05.
Article in English | MEDLINE | ID: mdl-29024940

ABSTRACT

BACKGROUND: Anoctamin (ANO)/transmembrane member 16 (TMEM16) proteins mediate diverse physiological and pathophysiological functions including cancer cell proliferation. The present study aimed to identify the role of ANOs in pancreatic cancer. METHODS: In an initial screen of ANOs, ANO9/TMEM16J was overexpressed in pancreatic cancer cells, and its role in the pathogenesis of pancreatic cancer was evaluated using an integrated in vitro and in vivo approach. To determine clinical relevance of the experimental findings, the prognostic value of ANO9 was evaluated in patients with pancreatic cancer. RESULTS: The ANO9 mRNA and protein levels were increased in pancreatic cancer-derived cells. Exogenous expression of ANO9 in PANC-1 cells significantly increased cell proliferation in cell cultures and in mice. In contrast, knockdown of ANO9 in AsPC-1, BxPC-3, and Capan-2 cells strongly inhibited cell proliferation. Mechanistic analysis suggested that physical association of ANO9 with epidermal growth factor receptor (EGFR) underlies ANO9-induced cell proliferation. Knockdown of ANO9 augmented the effects of the EGFR inhibitor and the cytotoxic agent on pancreatic cancer cell proliferation. In addition, high ANO9 expression is a poor prognostic factor in patients with pancreatic cancer. CONCLUSIONS: The ANO9/TMEM16J appears to be a clinically useful prognostic marker for pancreatic cancer and a potential therapeutic target.


Subject(s)
Anoctamins/genetics , Anoctamins/metabolism , Carcinoma, Pancreatic Ductal/genetics , Carcinoma, Pancreatic Ductal/metabolism , ErbB Receptors/metabolism , Membrane Proteins/metabolism , Pancreatic Neoplasms/genetics , Pancreatic Neoplasms/metabolism , Phospholipid Transfer Proteins/genetics , Phospholipid Transfer Proteins/metabolism , Adult , Aged , Aged, 80 and over , Animals , Anti-Bacterial Agents/therapeutic use , Antineoplastic Agents/pharmacology , Biomarkers, Tumor/genetics , Biomarkers, Tumor/metabolism , Carcinogenesis , Carcinoma, Pancreatic Ductal/drug therapy , Cell Line, Tumor , Cell Proliferation/genetics , Deoxycytidine/analogs & derivatives , Deoxycytidine/pharmacology , Disease-Free Survival , Doxycycline/therapeutic use , ErbB Receptors/antagonists & inhibitors , Erlotinib Hydrochloride/pharmacology , Female , Gene Knockdown Techniques , HEK293 Cells , Humans , Male , Membrane Proteins/genetics , Mice , Middle Aged , Neoplasm Transplantation , Pancreatic Neoplasms/drug therapy , Prognosis , RNA, Messenger/metabolism , Survival Rate , Tumor Stem Cell Assay , Gemcitabine
9.
Maxillofac Plast Reconstr Surg ; 39(1): 24, 2017 Dec.
Article in English | MEDLINE | ID: mdl-28894727

ABSTRACT

BACKGROUND: Tunneled transposition of the facial artery myomucosal (FAMM) island flap on the lingual side of the mandible has been reported for intraoral as well as oropharyngeal reconstruction. This modified technique overcomes the limitations of short range and dentition and further confirms the flexibility of the flap. This paper presents a case of reconstructing secondary soft palatal defect due to flap necrosis following two-flap palatoplasty in irradiated patient with lingually transposed facial artery myomucosal island flap. CASE PRESENTATION: The authors successfully reconstructed secondary soft palatal defect due to flap necrosis following two-flap palatoplasty in an irradiated 59-year-old female patient with tunnelized-facial artery myomucosal island flap (t-FAMMIF). CONCLUSIONS: Islanding and tunneling modification extends the versatility of the FAMM flap in the reconstruction of soft palatal defects post tumor excision and even after radiation, giving a great range of rotation and eliminating the need for revision in a second stage procedure. The authors thus highly recommend this versatile flap for the reconstruction of small and medium-sized oral defects.

10.
Maxillofac Plast Reconstr Surg ; 39(1): 25, 2017 Dec.
Article in English | MEDLINE | ID: mdl-28944219

ABSTRACT

BACKGROUND: The conventional transcervical resection for submandibular gland disease has some risks and an unsatisfactory cosmetic result. Recently, robot-assisted surgery has been developed as a plausible substitute for conventional surgery which provides an excellent cosmetic outcome. CASE PRESENTATION: The authors performed robot-assisted sialadenectomy via modified facelift incision using the da Vinci Xi surgical system (Intuitive Surgical Inc., CA, USA) with two endowrist arms (monopolar curved scissors and Maryland bipolar forceps) successfully in a 44-year-old female patient who suffered from sialolith and severe atrophic submandibular gland. CONCLUSIONS: If similar studies are done in the future, this robot-assisted sialadenectomy may become established as an alternative to existing disadvantageous surgical methods.

11.
Maxillofac Plast Reconstr Surg ; 39(1): 14, 2017 Dec.
Article in English | MEDLINE | ID: mdl-28603708

ABSTRACT

Tissue defect reconstruction using radial forearm free flap (RFFF) is a common surgical technique whose success or failure is mainly dependent on venous drainage. RFFF has two major venous outflow systems, superficial and deep vein. Drainage methods include combining both systems or using one alone. This review aims to recapitulate the vascular anatomy and network of RFFF as well as shed light on deep vein as a reliable venous drainage system. We also discuss basic evidence for and advantages of single microanastomosis with coalesced vein to overcome technical difficulties associated with the deep vein system.

12.
Oncotarget ; 8(19): 31842-31855, 2017 May 09.
Article in English | MEDLINE | ID: mdl-28418862

ABSTRACT

The epithelial-mesenchymal transition (EMT) is implicated in tumorigenesis and cancer progression, and canonical Wnt signaling tightly controls Snail, a key transcriptional repressor of EMT. While the suppression of canonical Wnt signaling and EMT comprises an attractive therapeutic strategy, molecular targets for small molecules reverting Wnt and EMT have not been widely studied. Meanwhile, the anti-helminthic niclosamide has been identified as a potent inhibitor of many oncogenic signaling pathways although its molecular targets have not yet been clearly identified. In this study, we show that niclosamide directly targets Axin-GSK3 interaction, at least in part, resulting in suppression of Wnt/Snail-mediated EMT. In vitro and in vivo, disruption of Axin-GSK3 complex by niclosamide induces mesenchymal to epithelial reversion at nM concentrations, accompanied with suppression of the tumorigenic potential of colon cancer. Niclosamide treatment successfully attenuates Snail abundance while increasing E-cadherin abundance in xenograft tumor. Notably, oral administration of niclosamide significantly suppressed adenoma formation in an APC-MIN mice model, indicating that niclosamide is an effective therapeutic for familial adenomatosis polyposis (FAP) patients. In this study, we identified a novel target to control the canonical Wnt pathway and Snail-mediated EMT program, and discovered a repositioned therapeutics for FAP patients.


Subject(s)
Adenomatous Polyposis Coli/metabolism , Axin Protein/metabolism , Glycogen Synthase Kinase 3/metabolism , Niclosamide/pharmacology , Adenomatous Polyposis Coli/drug therapy , Adenomatous Polyposis Coli/genetics , Animals , Axin Protein/chemistry , Cell Line, Tumor , Cell Movement/drug effects , Cell Survival/drug effects , Disease Models, Animal , Epithelial-Mesenchymal Transition/drug effects , Glycogen Synthase Kinase 3/chemistry , Heterografts , Mice , Models, Molecular , Molecular Conformation , Niclosamide/chemistry , Protein Binding/drug effects , Wnt Signaling Pathway/drug effects
13.
Oncotarget ; 8(19): 31856-31863, 2017 May 09.
Article in English | MEDLINE | ID: mdl-28418865

ABSTRACT

Despite the importance of Ras oncogenes as a therapeutic target in human cancer, their 'undruggable' tertiary structures limit the effectiveness of anti-Ras drugs. Canonical Wnt signaling contributes to Ras activity by glycogen synthase kinase 3 (GSK-3)-dependent phosphorylation at the C-terminus and subsequent degradation. In the accompanying report, we show that the anti-helminthic niclosamide directly binds to GSK-3 and inhibits Axin functions in colon cancer cells, with reversion of Snail-mediated epithelial-mesenchymal transition. In this study, we report that niclosamide effectively suppresses Ras and nuclear NFAT activities regardless of the mutational status of Ras at nM levels. Mechanistically, niclosamide increased endogenous GSK-3 activity, shortening the half-life of mutant Ras. Further, niclosamide activates Raf-1 kinase inhibitory protein, a downstream target of Snail repressor. Niclosamide treatment attenuates Ras-induced oncogenic potential in vitro and in vivo. These findings provide a clinically available repositioned Ras inhibitor as well as a novel strategy for inhibiting the Ras via GSK-3.


Subject(s)
Cell Transformation, Neoplastic/drug effects , Cell Transformation, Neoplastic/genetics , Cell Transformation, Neoplastic/metabolism , Genes, ras , Glycogen Synthase Kinase 3/metabolism , Niclosamide/pharmacology , Animals , Cell Line, Tumor , Disease Models, Animal , Enzyme Activation/drug effects , Humans , Models, Biological , Mutation , Xenograft Model Antitumor Assays
14.
Nat Commun ; 8: 14374, 2017 02 08.
Article in English | MEDLINE | ID: mdl-28176759

ABSTRACT

Dynamic regulation of glucose flux between aerobic glycolysis and the pentose phosphate pathway (PPP) during epithelial-mesenchymal transition (EMT) is not well-understood. Here we show that Snail (SNAI1), a key transcriptional repressor of EMT, regulates glucose flux toward PPP, allowing cancer cell survival under metabolic stress. Mechanistically, Snail regulates glycolytic activity via repression of phosphofructokinase, platelet (PFKP), a major isoform of cancer-specific phosphofructokinase-1 (PFK-1), an enzyme involving the first rate-limiting step of glycolysis. The suppression of PFKP switches the glucose flux towards PPP, generating NADPH with increased metabolites of oxidative PPP. Functionally, dynamic regulation of PFKP significantly potentiates cancer cell survival under metabolic stress and increases metastatic capacities in vivo. Further, knockdown of PFKP rescues metabolic reprogramming and cell death induced by loss of Snail. Thus, the Snail-PFKP axis plays an important role in cancer cell survival via regulation of glucose flux between glycolysis and PPP.


Subject(s)
Glucose/metabolism , Neoplasms/pathology , Oxidative Stress/genetics , Phosphofructokinase-1, Type C/genetics , Phosphofructokinase-1/genetics , Snail Family Transcription Factors/metabolism , Cell Survival/genetics , Epithelial-Mesenchymal Transition/genetics , Gene Expression Regulation, Neoplastic , Gene Knockdown Techniques , Glycolysis , Humans , NADP/metabolism , Neoplasms/genetics , Neoplasms/metabolism , Pentose Phosphate Pathway/genetics , Phosphofructokinase-1/metabolism , Phosphofructokinase-1, Type C/metabolism , RNA, Small Interfering/metabolism , Snail Family Transcription Factors/genetics
15.
Arch Pharm Res ; 38(3): 313-20, 2015 Mar.
Article in English | MEDLINE | ID: mdl-25634102

ABSTRACT

Aerobic glycolysis is widely accepted as the glucose metabolism for production of biomass such as nucleotides, amino acids, and fatty acids which underlie the anabolic process of cancer cell proliferation. The epithelial-mesenchymal transition (EMT) is a complex cellular mechanism for invasion and metastatic progression in cancer cells. While Snail-mediated EMT regulated by major oncogenic signaling has been well-studied over the last decade, metabolic reprogramming during the EMT has not. In this work, we emphasize the importance of catabolic metabolism for cancer cell survival during cancer cell EMT. Because specific catabolic processes such as autophage and fatty acid oxidation have been well explained, we mainly focus on the general aspects of energy metabolism promoting cancer cell survival under metabolic stress. We also revisit the role of mitochondria in catabolism as oxidative phosphorylation in cancer has long been underestimated. Considering the highly inefficient process of metastatic progression and profound metabolic stress following matrix detachment of solid cancer, catabolic reprogramming during the EMT may play an important role in overcoming metastatic inefficiency of cancer cells.


Subject(s)
Energy Metabolism/physiology , Epithelial-Mesenchymal Transition/physiology , Neoplasms/metabolism , Cell Survival/physiology , Glucose/metabolism , Glycolysis/physiology , Humans , Mitochondria/metabolism , Neoplasm Metastasis , Neoplasms/pathology , Oxidative Phosphorylation , Stress, Physiological/physiology
16.
J Control Release ; 194: 316-22, 2014 Nov 28.
Article in English | MEDLINE | ID: mdl-25245775

ABSTRACT

Diabetes is the fastest growing metabolic disease that fails to utilize glucose properly due to insulin deficiency or insulin resistance. Although several limited studies demonstrated non-invasive means of protein delivery, major hurdles for commercial success such as short half-life, enzymatic degradation and low bioavailability still remain to overcome. Methylcellulose (MC), a hydrophobically-modified cellulose derivative, forms temperature reversible gel in aqueous solution. However, as the gelling temperature of MC is higher than body temperature, it should be lowered to below body temperature for practical clinical application. In order to decrease gelling temperature and increase bio-compatibility and bio-elimination of MC, the molecular weight of MC was decreased using enzymatic degradation method and confirmed by gel permeation chromatography. Bio-elimination of low molecular weight (LMw) MC was confirmed with non-invasive live image and ex vivo experiment. The exenatide and FGF 21 were physically loaded 100% into LMwMC-based thermo-reversible gel and slowly released from gel with no initial bursts. Exenatide-loaded LMwMC gel showed reduction of blood glucose level for a week in type 1 diabetic animal model. FGF 21-loaded LMwMC gel reduced glucose level to normal condition and maintained over 10 days in type 2 diabetic animal model. LMwMC-based thermo-reversible and injectable hydrogel provides a strong potential to be efficient protein drug delivery system for the treatment of type 1 and type 2 diabetes.


Subject(s)
Diabetes Mellitus, Type 1/drug therapy , Diabetes Mellitus, Type 2/drug therapy , Fibroblast Growth Factors/administration & dosage , Fibroblast Growth Factors/therapeutic use , Gels/chemistry , Hypoglycemic Agents/administration & dosage , Hypoglycemic Agents/therapeutic use , Methylcellulose/chemistry , Peptides/administration & dosage , Peptides/therapeutic use , Pharmaceutic Aids/chemistry , Venoms/administration & dosage , Venoms/therapeutic use , Animals , Blood Glucose/metabolism , Diabetes Mellitus, Experimental/drug therapy , Exenatide , Fibroblast Growth Factors/pharmacokinetics , Hypoglycemic Agents/pharmacokinetics , Male , Mice , Mice, Inbred BALB C , Molecular Weight , Peptides/pharmacokinetics , Thermodynamics , Tissue Distribution , Venoms/pharmacokinetics
17.
Biomaterials ; 35(37): 9888-9896, 2014 Dec.
Article in English | MEDLINE | ID: mdl-25218859

ABSTRACT

Though growth factors allow tissue regeneration, the trade-off between their effectiveness and adverse effects limits clinical application. The key issues in current growth factor therapy largely derive from initial burst pharmacokinetics, rapid clearance, and proteolytic cleavage resulting in clinical ineffectiveness and diverse complications. While a number of studies have focused on the development of carriers, issues arising from soluble growth factor remain. In this study, we report a prodrug of growth factors constituting a novel mode of action (MoA). To mimic endogenous protein processing in cells, we developed a recombinant BMP-2 polypeptide based on a protein transduction domain (PTD) to transduce the protein into cells followed by furin-mediated protein cleavage and secretion of active growth factor. As proof of concept, a few micrograms scale of PTD-BMP-2 polypeptide sufficed to induce bone regeneration in vivo. As a simple platform, our technique can easily be extended to delivery of BMP-7 and DKK-1 as therapeutics for TGF-ß and canonical Wnt signaling, respectively, to suppress the epithelial-mesenchymal transition (EMT), which constitutes a fundamental biological mechanism of many diseases. This technique largely overcomes the limitations of current soluble growth factors and opens the door to next generation growth factor therapeutics.


Subject(s)
Bone Morphogenetic Protein 2/genetics , Peptides/genetics , Transduction, Genetic/methods , Transforming Growth Factor beta/genetics , Animals , Bone Morphogenetic Protein 2/metabolism , Bone Morphogenetic Protein 2/pharmacology , Bone Regeneration/drug effects , Cell Line , Dogs , Female , Furin/metabolism , Humans , Mice , Peptides/metabolism , Peptides/pharmacology , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolism , Recombinant Fusion Proteins/pharmacology , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Recombinant Proteins/pharmacology , Regenerative Medicine , Transforming Growth Factor beta/metabolism , Transforming Growth Factor beta/pharmacology , Wnt Signaling Pathway/drug effects
18.
Nat Commun ; 5: 4423, 2014 Jul 23.
Article in English | MEDLINE | ID: mdl-25055241

ABSTRACT

Cytotoxin-associated gene A (CagA) is an oncoprotein and a major virulence factor of H. pylori. CagA is delivered into gastric epithelial cells via a type IV secretion system and causes cellular transformation. The loss of epithelial adhesion that accompanies the epithelial-mesenchymal transition (EMT) is a hallmark of gastric cancer. Although CagA is a causal factor in gastric cancer, the link between CagA and the associated EMT has not been elucidated. Here, we show that CagA induces the EMT by stabilizing Snail, a transcriptional repressor of E-cadherin expression. Mechanistically we show that CagA binds GSK-3 in a manner similar to Axin and causes it to shift to an insoluble fraction, resulting in reduced GSK-3 activity. We also find that the level of Snail protein is increased in H. pylori infected epithelium in clinical samples. These results suggest that H. pylori CagA acts as a pathogenic scaffold protein that induces a Snail-mediated EMT via the depletion of GSK-3.


Subject(s)
Antigens, Bacterial/physiology , Bacterial Proteins/physiology , Down-Regulation/physiology , Epithelial-Mesenchymal Transition/physiology , Glycogen Synthase Kinase 3/metabolism , Helicobacter pylori/physiology , Transcription Factors/physiology , Biopsy , Carcinogenesis/metabolism , Carcinogenesis/pathology , Cells, Cultured , Epithelial Cells/metabolism , Epithelial Cells/pathology , Gastric Mucosa/metabolism , Gastritis/metabolism , Gastritis/pathology , Humans , Signal Transduction/physiology , Snail Family Transcription Factors , Stomach/pathology , Stomach Neoplasms/metabolism , Stomach Neoplasms/pathology
19.
Cell Cycle ; 12(10): 1578-87, 2013 May 15.
Article in English | MEDLINE | ID: mdl-23624843

ABSTRACT

p53 is a bona fide tumor suppressor gene whose loss of function marks the most common genetic alteration in human malignancy. Although the causal link between loss of p53 function and tumorigenesis has been clearly demonstrated, the mechanistic links by which loss of p53 potentiates oncogenic signaling are not fully understood. Recent evidence indicates that the microRNA-34 (miR-34) family, a transcriptional target of the p53, directly suppresses a set of canonical Wnt genes and Snail, resulting in p53-mediated suppression of Wnt signaling and the EMT process. In this study, we report that p53 regulates GSK-3ß nuclear localization via miR-34-mediated suppression of Axin2 in colorectal cancer. Exogenous miR-34a decreases Axin2 UTR-reporter activity through multiple binding sites within the 5' and 3' UTR of Axin2. Suppression of Axin2 by p53 or miR-34 increases nuclear GSK-3ß abundance and leads to decreased Snail expression in colorectal cancer cells. Conversely, expression of the non-coding UTR of Axin2 causes depletion of endogenous miR-34 via the miR-sponge effect together with increased Axin2 function, supporting that the RNA-RNA interactions with Axin2 transcripts act as an endogenous decoy for miR-34. Further, RNA transcripts of miR-34 target were correlated with Axin2 in clinical data set of colorectal cancer patients. Although the biological relevance of nuclear GSK-3 level has not been fully studied, our results demonstrate that the tumor suppressor p53/miR-34 axis plays a role in regulating nuclear GSK-3 levels and Wnt signaling through the non-coding UTR of Axin2 in colorectal cancer.


Subject(s)
Axin Protein/metabolism , Glycogen Synthase Kinase 3/metabolism , MicroRNAs/metabolism , Tumor Suppressor Protein p53/metabolism , 3' Untranslated Regions , 5' Untranslated Regions , Antibiotics, Antineoplastic/pharmacology , Axin Protein/antagonists & inhibitors , Axin Protein/genetics , Binding Sites , Cell Line, Tumor , Colorectal Neoplasms/metabolism , Colorectal Neoplasms/pathology , Doxorubicin/pharmacology , Gene Expression Regulation, Neoplastic/drug effects , HCT116 Cells , Humans , RNA Interference , RNA, Messenger/metabolism , RNA, Small Interfering/metabolism , Snail Family Transcription Factors , Transcription Factors/metabolism , Transcription, Genetic , Wnt Signaling Pathway
20.
Cell Cycle ; 11(7): 1273-81, 2012 Apr 01.
Article in English | MEDLINE | ID: mdl-22421157

ABSTRACT

Though tumor suppressor p53 and the canonical Wnt cascade have been extensively studied for the last 30 years, due to their important physiological roles, the two signaling pathways have been largely considered independent. Recently, the miR-34 family was found to directly link p53 and Wnt, revealing the tight connection between loss of tumor suppressor function and activation of oncogenic signaling. These observations demonstrate that miR-34, known to be directly downstream of p53, targets a set of highly conserved sites in the UTR of Wnt and EMT genes, specifically WNT1, WNT3, LRP6, AXIN2, ß-catenin, LEF1 and Snail, resulting in suppression of TCF/LEF transcriptional activity and the EMT program. The loss of p53 function increases Wnt activities and promotes the Snail-dependent EMT program at multiple levels in a miR-34/UTR-specific manner. The TCF/LEF transcriptional signature was closely associated with functionality of p53 and miR-34 in clinical samples, suggesting the pervasive impact of miR-34 loss on the oncogenic pathway in human cancer. Here, we review recent findings on ceRNA in light of novel data to elucidate the physiological relevance of the p53-miR-34-Wnt network, which encompasses sets of genes and directions of signaling. As loss of wt-p53 or hyperactivation of Wnt is critical in maintaining cancer stem cell properties and in establishing the metastatic program, these observations indicate a mechanism of miR-mediated quasi-sufficiency which connects tumor suppressor and oncogenic signaling pathways, supporting a continuum model of human cancer.


Subject(s)
MicroRNAs/genetics , Neoplasms/metabolism , Tumor Suppressor Protein p53/metabolism , Wnt Signaling Pathway , Wnt3 Protein/metabolism , Humans , Neoplasms/genetics , Neoplasms/mortality , Neoplasms/pathology , Protein-Tyrosine Kinases/genetics , Protein-Tyrosine Kinases/metabolism , RNA Interference , Snail Family Transcription Factors , Transcription Factors/genetics , Transcription Factors/metabolism , Transcriptional Activation , Tumor Suppressor Protein p53/genetics , Wnt Signaling Pathway/genetics , Wnt3 Protein/genetics
SELECTION OF CITATIONS
SEARCH DETAIL
...