Social Stigma and Discrimination Toward People With Drug Addiction: A National Survey in Korea.

OBJECTIVE: This study aims to understand the attitudes, stigma, and discrimination of the general adult population toward drug addiction. METHODS: We conducted a cross-sectional nationwide survey with 1,020 Korean adults using an Internet web-based panel. Self-reported data were collected on demographics, experience with substance abuse, perceptions of narcotic analgesic use, beliefs about the legalization of cannabis use, coping with substance abuse and addiction, and perceptions of drug risks. All statistical analysis in this study utilized the IBM SPSS Statistics 26 program. RESULTS: In this study, 1.6% of the participants reported abuse of opioid analgesics, 88.0% reported negative perceptions of drug addiction, and 76.9% reported agreeing to unfair treatment of drug addicts. Logistic regression analysis found that perceived stigma was more prevalent among women (odd ratio [OR]=2.087, p<0.01), old adults (OR=1.939, p<0.01), those with no personal experience of opioid misuse (OR=8.172, p<0.05), and those who were non-smoking (OR=2.011, p<0.01). In addition, the discriminatory attitude was more prevalent among participants with higher income (OR=1.989, p<0.001) and those who are non-smoking (OR=1.608, p<0.05). CONCLUSION: This study provides information and guidelines for public intervention in drug addiction by identifying factors influencing social stigma and discriminatory behaviors toward drug addiction. The findings suggest that education on drug addiction prevention for the general adult population is necessary, and this education should include knowledge on coping with drug addiction and reducing stigma and discrimination toward drug addicts.

The eleven-item Alcohol, Smoking and Substance Involvement Screening Test (ASSIST-11): Cross-cultural psychometric evaluation across 42 countries.

The Alcohol, Smoking and Substance Involvement Screening Test (ASSIST) is an instrument to screen substance-use-related health risks. However, little is known whether the ASSIST could be further shortened while remaining psychometrically sound across different countries, languages, gender identities, and sexual-orientation-based groups. The study aimed to validate a shortened 11-item ASSIST (ASSIST-11). Using the International Sex Survey data, 82,243 participants (M age = 32.39 years) across 42 countries and 26 languages completed questions from the ASSIST-11 regarding gender identity, sexual orientation, and other information. Confirmatory factor analysis (CFA) and multigroup CFA (MGCFA) evaluated the ASSIST-11's structure and tested measurement invariance across groups. Cronbach's α and McDonald's ω were used to examine the internal consistency. Cohen's d and independent t-tests were used to examine known-group validity. The ASSIST-11 was unidimensional across countries, languages, age groups, gender identities (i.e., men, women, and gender-diverse individuals), and sexual orientations (i.e., heterosexual and sexual minority individuals). Cronbach's α was 0.63 and McDonald's ω was 0.68 for the ASSIST-11. Known-group validity was supported by Cohen's d (range between 0.23 and 0.40) with significant differences (p-values<0.001). The ASSIST-11 is a modified instrument with a unidimensional factor structure across different languages, age groups, countries, gender identities, and sexual orientations. The low internal consistency of the ASSIST-11 might be acceptable as it assesses a broad concept (i.e., use of several different substances). Healthcare providers and researchers may use the ASSIST-11 to quickly assess substance-use information from general populations and evaluate the need to follow up with more detailed questions about substance use.

Molecular Detection of Equine Adenovirus 1 in Nasal Swabs from Horses in the Republic of Korea.

Equine adenovirus 1 (EAdV-1) can cause upper respiratory disease in horses and has been reported worldwide. In this study, and for the first time in Korea, the prevalence of EAdV-1 in equine nasal swabs was investigated using a PCR to identify potential risk factors and examine the genetic diversity of its DNA sequences by a comparison with foreign strains. Nasal swabs collected from 359 horses reared at Korea Racing Authority facilities were tested using an EAdV-1 hexon-specific PCR and the associations between EAdV-1 infection and sex, age, region, breed, and activity were analyzed. Five samples (1.4%, 5/359) tested positive for EAdV-1; however, no statistically significant differences were observed with respect to any variable. Among the five EAdV-1-positive horses, a co-infection with equine influenza, equine herpesvirus 1 and 4, or Streptococcus equi was not detected; however, clinical respiratory signs were observed in one. Phylogenetic analyses based on partial EAdV-1 hexon gene sequences revealed that the Korean EAdV-1 isolates shared approximately 98.8-100% similarity among each other and with foreign strains. Three Korean isolates shared high similarity with strains from Australia and India and the remaining two isolates were separate in phylogenetic analyses. These findings highlight the molecular prevalence and genetic diversity of EAdV-1 in horses in Korea.

Rice ß-glucosidase Os12BGlu38 is required for synthesis of intine cell wall and pollen fertility.

Glycoside hydrolase family1 ß-glucosidases play a variety of roles in plants, but their in planta functions are largely unknown in rice (Oryza sativa). In this study, the biological function of Os12BGlu38, a rice ß-glucosidase, expressed in bicellular to mature pollen, was examined. Genotype analysis of progeny of the self-fertilized heterozygous Os12BGlu38 T-DNA mutant, os12bglu38-1, found no homozygotes and a 1:1 ratio of wild type to heterozygotes. Reciprocal cross analysis demonstrated that Os12BGlu38 deficiency cannot be inherited through the male gamete. In cytological analysis, the mature mutant pollen appeared shrunken and empty. Histochemical staining and TEM showed that mutant pollen lacked intine cell wall, which was rescued by introduction of wild-type Os12BGlu38 genomic DNA. Metabolite profiling analysis revealed that cutin monomers and waxes, the components of the pollen exine layer, were increased in anthers carrying pollen of os12bglu38-1 compared with wild type and complemented lines. Os12BGlu38 fused with green fluorescent protein was localized to the plasma membrane in rice and tobacco. Recombinant Os12BGlu38 exhibited ß-glucosidase activity on the universal substrate p-nitrophenyl ß-d-glucoside and some oligosaccharides and glycosides. These findings provide evidence that function of a plasma membrane-associated ß-glucosidase is necessary for proper intine development.

Action of Multiple Rice ß-Glucosidases on Abscisic Acid Glucose Ester.

Conjugation of phytohormones with glucose is a means of modulating their activities, which can be rapidly reversed by the action of ß-glucosidases. Evaluation of previously characterized recombinant rice ß-glucosidases found that nearly all could hydrolyze abscisic acid glucose ester (ABA-GE). Os4BGlu12 and Os4BGlu13, which are known to act on other phytohormones, had the highest activity. We expressed Os4BGlu12, Os4BGlu13 and other members of a highly similar rice chromosome 4 gene cluster (Os4BGlu9, Os4BGlu10 and Os4BGlu11) in transgenic Arabidopsis. Extracts of transgenic lines expressing each of the five genes had higher ß-glucosidase activities on ABA-GE and gibberellin A4 glucose ester (GA4-GE). The ß-glucosidase expression lines exhibited longer root and shoot lengths than control plants in response to salt and drought stress. Fusions of each of these proteins with green fluorescent protein localized near the plasma membrane and in the apoplast in tobacco leaf epithelial cells. The action of these extracellular ß-glucosidases on multiple phytohormones suggests they may modulate the interactions between these phytohormones.

Accuracy of surface-guided patient setup for conventional radiotherapy of brain and nasopharynx cancer.

PURPOSE: To evaluate the accuracy of surface-guided radiotherapy (SGRT) in cranial patient setup by direct comparison between optical surface imaging (OSI) and cone-beam computed tomography (CBCT), before applying SGRT-only setup for conventional radiotherapy of brain and nasopharynx cancer. METHODS AND MATERIALS: Using CBCT as reference, SGRT setup accuracy was examined based on 269 patients (415 treatments) treated with frameless cranial stereotactic radiosurgery (SRS) during 2018-2019. Patients were immobilized in customized head molds and open-face masks and monitored using OSI during treatment. The facial skin area in planning CT was used as OSI region of interest (ROI) for automatic surface alignment and the skull was used as the landmark for automatic CBCT/CT registration. A 6 degrees of freedom (6DOF) couch was used. Immediately after CBCT setup, an OSI verification image was captured, recording the SGRT setup differences. These differences were analyzed in 6DOFs and as a function of isocenter positions away from the anterior surface to assess OSI-ROI bias. The SGRT in-room setup time was estimated and compared with CBCT and orthogonal 2D kilovoltage (2DkV) setups. RESULTS: The SGRT setup difference (magnitude) is found to be 1.0 ± 2.5 mm and 0.1˚±1.4˚ on average among 415 treatments and within 5 mm/3˚ with greater than 95% confidence level (P < 0.001). Outliers were observed for very-posterior isocenters: 15 differences (3.6%) are >5.0mm and 9 (2.2%) are >3.0˚. The setup differences show minor correlations (|r| < 0.45) between translational and rotational DOFs and a minor increasing trend (<1.0 mm) in the anterior-to-posterior direction. The SGRT setup time is 0.8 ± 0.3 min, much shorter than CBCT (5 ± 2 min) and 2DkV (2 ± 1 min) setups. CONCLUSION: This study demonstrates that SGRT has sufficient accuracy for fast in-room patient setup and allows real-time motion monitoring for beam holding during treatment, potentially useful to guide radiotherapy of brain and nasopharynx cancer with standard fractionation.

Identification of Novel Biomarker for Early Detection of Diabetic Nephropathy.

Diabetic nephropathy (DN) is one of the most common complications of diabetes mellitus. After development of DN, patients will progress to end-stage renal disease, which is associated with high morbidity and mortality. Here, we developed early-stage diagnostic biomarkers to detect DN as a strategy for DN intervention. For the DN model, Zucker diabetic fatty rats were used for DN phenotyping. The results revealed that DN rats showed significantly increased blood glucose, blood urea nitrogen (BUN), and serum creatinine levels, accompanied by severe kidney injury, fibrosis and microstructural changes. In addition, DN rats showed significantly increased urinary excretion of kidney injury molecule-1 (KIM-1) and neutrophil gelatinase-associated lipocalin (NGAL). Transcriptome analysis revealed that new DN biomarkers, such as complementary component 4b (C4b), complementary factor D (CFD), C-X-C motif chemokine receptor 6 (CXCR6), and leukemia inhibitory factor (LIF) were identified. Furthermore, they were found in the urine of patients with DN. Since these biomarkers were detected in the urine and kidney of DN rats and urine of diabetic patients, the selected markers could be used as early diagnosis biomarkers for chronic diabetic nephropathy.

Associations between metabolic syndrome and type of dementia: analysis based on the National Health Insurance Service database of Gangwon province in South Korea.

BACKGROUND: Metabolic syndrome is a cluster of conditions that occur together, increasing the risk of cardiovascular disease. However, the relationship between metabolic syndrome and dementia has remained controversial. Using nationwide population cohort data, we investigated the association between metabolic syndrome and dementia, according to the dementia type. METHODS: We analyzed data of 84,144 individuals, in the aged group of more than 60 years, between January 1, 2009, to December 31, 2009, at Gangwon province by using the information of the (Korean) National Health Insurance Service. After eight years of gap, in 2017, we investigated the relationship between metabolic syndrome and dementia. We classified Dementia either as dementia of the Alzheimer type (AD) or vascular dementia (VD). AD and VD were defined as per the criteria of International Classification of Disease, Tenth Revision, Clinical Modification codes. Multiple logistic regression analyses examined the associations between metabolic syndrome or five metabolic syndrome components and dementia. Analyses included factors like age, sex, smoking, alcohol, physical inactivity, previous stroke, and previous cardiac disease. RESULTS: Metabolic syndrome was associated with AD (OR = 11.48, 95% CI 9.03-14.59), not with VD. Each of five components of metabolic syndrome were also associated with AD. (high serum triglycerides: OR = 1.87, 95% CI 1.60-2.19; high blood pressure: OR = 1.85, 95% CI 1.55-2.21; high glucose: OR = 1.77, 95% CI 1.52-2.06; abdominal obesity: OR = 1.88, 95% CI 1.57-2.25; low serum high-density lipoprotein cholesterol: OR = 1.91, 95% CI 1.63-2.24) However, among components of metabolic syndrome, only the high glucose level was associated with VD. (OR = 1.26, 95% CI 1.01-1.56) body mass index (BMI), fasting glucose, and smoking were also associated with AD. (BMI: OR = 0.951, 95% CI 0.927-0.975; fasting glucose: OR = 1.003, 95% CI 1.001-1.005; smoking: OR = 1.020, 95% CI 1.003-1.039) A history of the previous stroke was associated with both AD and VD. (AD: OR = 1.827, 95% CI 1.263-2.644; VD: OR 2.775, 95% CI 1.747-4.406) CONCLUSIONS: Metabolic syndrome was associated with AD but not with VD. Patients with metabolic syndrome had an 11.48 times more likeliness to develop AD compared to those without metabolic syndrome. VD was associated only with several risk factors that could affect the vascular state rather than a metabolic syndrome. We suggested that the associations between metabolic syndrome and dementia would vary depending on the type of dementia.

The novel cereblon modulator CC-885 inhibits mitophagy via selective degradation of BNIP3L.

Mitophagy is a degradative pathway that mediates the degradation of the entire mitochondria, and defects in this process are implicated in many diseases including cancer. In mammals, mitophagy is mediated by BNIP3L (also known as NIX) that is a dual regulator of mitochondrial turnover and programmed cell death pathways. Acute myeloid leukemia (AML) cells with deficiency of BNIP3L are more sensitive to mitochondria-targeting drugs. But small molecular inhibitors for BNIP3L are currently not available. Some immunomodulatory drugs (IMiDs) have been proved by FDA for hematologic malignancies, however, the underlining molecular mechanisms are still elusive, which hindered the applications of BNIP3L inhibition for AML treatment. In this study we carried out MS-based quantitative proteomics analysis to identify the potential neosubstrates of a novel thalidomide derivative CC-885 in A549 cells. In total, we quantified 5029 proteins with 36 downregulated in CRBN+/+ cell after CC-885 administration. Bioinformatic analysis showed that macromitophagy pathway was enriched in the negative pathway after CC-885 treatment. We further found that CC-885 caused both dose- and time-dependent degradation of BNIP3L in CRBN+/+, but not CRBN-/- cell. Thus, our data uncover a novel role of CC-885 in the regulation of mitophagy by targeting BNIP3L for CRL4CRBN E3 ligase-dependent ubiquitination and degradation, suggesting that CC-885 could be used as a selective BNIP3L degradator for the further investigation. Furthermore, we demonstrated that CC-885 could enhance AML cell sensitivity to the mitochondria-targeting drug rotenone, suggesting that combining CC-885 and mitochondria-targeting drugs may be a therapeutic strategy for AML patients.

LIKE SEX4 1 Acts as a ß-Amylase-Binding Scaffold on Starch Granules during Starch Degradation.

In Arabidopsis (Arabidopsis thaliana) leaves, starch is synthesized during the day and degraded at night to fuel growth and metabolism. Starch is degraded primarily by ß-amylases, liberating maltose, but this activity is preceded by glucan phosphorylation and is accompanied by dephosphorylation. A glucan phosphatase family member, LIKE SEX4 1 (LSF1), binds starch and is required for normal starch degradation, but its exact role is unclear. Here, we show that LSF1 does not dephosphorylate glucans. The recombinant dual specificity phosphatase (DSP) domain of LSF1 had no detectable phosphatase activity. Furthermore, a variant of LSF1 mutated in the catalytic cysteine of the DSP domain complemented the starch-excess phenotype of the lsf1 mutant. By contrast, a variant of LSF1 with mutations in the carbohydrate binding module did not complement lsf1 Thus, glucan binding, but not phosphatase activity, is required for the function of LSF1 in starch degradation. LSF1 interacts with the ß-amylases BAM1 and BAM3, and the BAM1-LSF1 complex shows amylolytic but not glucan phosphatase activity. Nighttime maltose levels are reduced in lsf1, and genetic analysis indicated that the starch-excess phenotype of lsf1 is dependent on bam1 and bam3 We propose that LSF1 binds ß-amylases at the starch granule surface, thereby promoting starch degradation.

3-dimensional analysis about the effects of aging and risk factors on changes in oral environment.

PURPOSE: The purpose of this study is to investigate the effects of aging and various risk factors on the oral environment and to analyze them in 3-dimensions. MATERIALS AND METHODS: A total of 800 patients were enrolled in this study, and subjects were divided into 4 groups by age-under 55, 56 - 65, 66 - 75, and over 76. Based on their most recent visit, the number of crowns, bridges, implants, and the remaining natural teeth were recorded. Smoking habits, along with presence of diabetes and hypertension, were surveyed, as risk factors were also set as a variable. Comparisons among the groups or within the groups were performed by independent t-test, and one-way and two-way ANOVA. Kruskal-Wallis test and Mann-Whitney U test were used for analysis. It was assumed to be statistically significant when P value is below .05. RESULTS: Changes in the number of crowns, bridges, implants, and the remaining natural teeth by age were statistically significant. When we examined the effect of risk factors on the change of variables with age, hypertension was found to affect the number of bridges. Diabetes and smoking were found to affect the number of the remaining natural teeth. The other variables were not statistically significant. CONCLUSION: Aging is considered to be an important variable affecting the change of oral environment. Among the risk factors, the presence of smoking habit and diabetes is thought to have a great influence on the change of the number of the remaining natural teeth.

A Therapeutic Strategy for Chemotherapy-Resistant Gastric Cancer via Destabilization of Both ß-Catenin and RAS.

: Treatment of advanced gastric cancer patients with current standard chemotherapeutic agents frequently results in resistance, leading to poor overall survival. However, there has been no success in developing strategies to overcome it. We showed the expression levels of both ß-catenin and RAS were significantly increased and correlated in tissues of 756 gastric cancer (GC) patients and tissues of primary- and acquired-resistance patient-derived xenograft tumors treated with 5-fluorouracil and oxaliplatin modulated with leucovorin (FOLFOX). On the basis of our previous studies, where small molecules to suppress colorectal cancer (CRC) via degrading both ß-catenin and RAS were developed, we tested the effectiveness of KYA1797K, a representative compound functioning by binding axin, in the growth of GC cells. The efficacy test of the drugs using gastric tumor organoids of Apc1638N mice showed that the CD44 and ALDH1A3 cancer stem cell markers were induced by FOLFOX, but not by KYA1797K. KYA1797K also efficiently suppressed tumors generated by re-engrafting the FOLFOX-resistant patient-derived xenograft (PDX) tumors, which also showed resistance to paclitaxel. Overall, the small-molecule approach degrading both ß-catenin and RAS has potential as a therapeutic strategy for treating GC patients resistant to current standard chemotherapies.

Molecular Detection of Equus caballus Papillomavirus Type 2 in Genital Swabs From Healthy Horses in the Republic of Korea.

Equus caballus papillomavirus type 2 (EcPV-2) is implicated in genital neoplasms in horses, including equine penile papillomas, penile intraepithelial neoplasia, and squamous cell carcinoma (SCC). This virus seldom regresses spontaneously and can result in the development of SCC, which may result in significant clinical damage and economic cost. However, the prevalence of this virus is unknown in clinically unaffected horses. The aim of this study was to determine the prevalence of EcPV-2 DNA in genital swabs from healthy horses in the Republic of Korea and to investigate genetic variability within EcPV-2. A total of 797 genital swab samples obtained from Thoroughbred horses (35 stallions and 762 mares) were tested for the presence of the EcPV-2 gene E1 using a polymerase chain reaction (PCR) assay. Positive results were confirmed by repeat PCR to detect the presence of E6 and amplicon sequencing. Equus caballus papillomavirus type 2 DNA was detected in 1% (8/762) of mare swabs but in none of stallion swabs (0/35). All EcPV-2-positive mares had been imported from the United States. Equus caballus papillomavirus type 2 E6 gene sequences were used for phylogenetic analysis and revealed three subdivisions related to the European strains, and three isolates were located on two separate branches. This study documents the molecular prevalence and phylogenetic analysis of EcPV-2 DNA in the Republic of Korea.

A Ras destabilizer KYA1797K overcomes the resistance of EGFR tyrosine kinase inhibitor in KRAS-mutated non-small cell lung cancer.

The epidermal growth factor receptor (EGFR) inhibitors such as erlotinib and gefitinib are widely used for treatment of non-small cell lung cancer (NSCLC), but they have shown limited efficacy in an unselected population of patients. The KRAS mutations, which are identified in approximately 20% of NSCLC patients, have shown to be associated with the resistance to the EGFR tyrosine kinase inhibitors (TKIs). Currently, there is no clinically available targeted therapy which can effectively inhibit NSCLC tumors harboring KRAS mutations. This study aims to show the effectiveness of KYA1797K, a small molecule which revealed anti-cancer effect in colorectal cancer by destabilizing Ras via inhibiting the Wnt/ß-catenin pathway, for the treatment of KRAS-mutated NSCLC. While erlotinib fail to have anti-transforming effect in NSCLC cell lines harboring KRAS mutations, KYA1797K effectively inhibited the Ras-ERK pathway in KRAS-mutant NSCLC cell lines. As a result, KYA1797K treatment suppressed the growth and transformation of KRAS mutant NSCLC cells and also induced apoptosis. Furthermore, KYA1797K effectively inhibited Kras-driven tumorigenesis in the KrasLA2 mouse model by suppressing the Ras-ERK pathway. The destabilization of Ras via inhibition of the Wnt/ß-catenin pathway is a potential therapeutic strategy for KRAS-mutated NSCLC that is resistant to EGFR TKI.

WDR76 is a RAS binding protein that functions as a tumor suppressor via RAS degradation.

Stability regulation of RAS that can affect its activity, in addition to the oncogenic mutations, occurs in human cancer. However, the mechanisms for stability regulation of RAS involved in their activity and its roles in tumorigenesis are poorly explored. Here, we identify WD40-repeat protein 76 (WDR76) as one of the HRAS binding proteins using proteomic analyses of hepatocellular carcinomas (HCC) tissue. WDR76 plays a role as an E3 linker protein and mediates the polyubiquitination-dependent degradation of RAS. WDR76-mediated RAS destabilization results in the inhibition of proliferation, transformation, and invasion of liver cancer cells. WDR76-/- mice are more susceptible to diethylnitrosamine-induced liver carcinogenesis. Liver-specific WDR76 induction destabilizes Ras and markedly reduces tumorigenesis in HRasG12V mouse livers. The clinical relevance of RAS regulation by WDR76 is indicated by the inverse correlation of their expressions in HCC tissues. Our study demonstrates that WDR76 functions as a tumor suppressor via RAS degradation.

A small molecule approach to degrade RAS with EGFR repression is a potential therapy for KRAS mutation-driven colorectal cancer resistance to cetuximab.

Drugs targeting the epidermal growth factor receptor (EGFR), such as cetuximab and panitumumab, have been prescribed for metastatic colorectal cancer (CRC), but patients harboring KRAS mutations are insensitive to them and do not have an alternative drug to overcome the problem. The levels of ß-catenin, EGFR, and RAS, especially mutant KRAS, are increased in CRC patient tissues due to mutations of adenomatous polyposis coli (APC), which occur in 90% of human CRCs. The increases in these proteins by APC loss synergistically promote tumorigenesis. Therefore, we tested KYA1797K, a recently identified small molecule that degrades both ß-catenin and Ras via GSK3ß activation, and its capability to suppress the cetuximab resistance of KRAS-mutated CRC cells. KYA1797K suppressed the growth of tumor xenografts induced by CRC cells as well as tumor organoids derived from CRC patients having both APC and KRAS mutations. Lowering the levels of both ß-catenin and RAS as well as EGFR via targeting the Wnt/ß-catenin pathway is a therapeutic strategy for controlling CRC and other types of cancer with aberrantly activated the Wnt/ß-catenin and EGFR-RAS pathways, including those with resistance to EGFR-targeting drugs attributed to KRAS mutations.

ß-Catenin-RAS interaction serves as a molecular switch for RAS degradation via GSK3ß.

RAS proteins play critical roles in various cellular processes, including growth and transformation. RAS proteins are subjected to protein stability regulation via the Wnt/ß-catenin pathway, and glycogen synthase kinase 3 beta (GSK3ß) is a key player for the phosphorylation-dependent RAS degradation through proteasomes. GSK3ß-mediated RAS degradation does not occur in cells that express a nondegradable mutant (MT) ß-catenin. Here, we show that ß-catenin directly interacts with RAS at the α-interface region that contains the GSK3ß phosphorylation sites, threonine 144 and threonine 148 residues. Exposure of these sites by prior ß-catenin degradation is required for RAS degradation. The introduction of a peptide that blocks the ß-catenin-RAS interaction by binding to ß-catenin rescues the GSK3ß-mediated RAS degradation in colorectal cancer (CRC) cells that express MT ß-catenin. The coregulation of ß-catenin and RAS stabilities by the modulation of their interaction provides a mechanism for Wnt/ß-catenin and RAS-ERK pathway cross-talk and the synergistic transformation of CRC by both APC and KRAS mutations.

Identification of Ras-degrading small molecules that inhibit the transformation of colorectal cancer cells independent of ß-catenin signaling.

Although the development of drugs that control Ras is an emerging topic in cancer therapy, no clinically applicable drug is currently available. We have previously utilized knowledge of the Wnt/ß-catenin signaling-dependent mechanism of Ras protein stability regulation to identify small molecules that inhibit the proliferation and transformation of various colorectal cancer (CRC) cells via degradation of both ß-catenin and Ras. Due to the absence of Ras degradation in cells expressing a nondegradable mutant form of ß-catenin and the need to determine an alternative mechanism of Ras degradation, we designed a cell-based system to screen compounds that degrade Ras independent of the Wnt/ß-catenin signaling pathway. A cell-based high-content screening (HCS) system that monitors the levels of EGFP-K-RasG12V was established using HCT-116 cells harboring a nondegradable mutant CTNNB1 (ΔS45). Through HCS of a chemical library composed of 10,000 compounds and subsequent characterization of hits, we identified several compounds that degrade Ras without affecting the ß-catenin levels. KY7749, one of the most effective compounds, inhibited the proliferation and transformation of CRC cells, especially KRAS-mutant cells that are resistant to the EGFR monoclonal antibody cetuximab. Small molecules that degrade Ras independent of ß-catenin may able to be used in treatments for cancers caused by aberrant EGFR and Ras.

Interaction of the Wnt/ß-catenin and RAS-ERK pathways involving co-stabilization of both ß-catenin and RAS plays important roles in the colorectal tumorigenesis.

Cancer development is usually driven by multiple genetic and molecular alterations rather than by a single defect. In the human colorectal cancer (CRC), series of mutations of genes are involved in the different stages of tumorigenesis. For example, adenomatous polyposis coli (APC) and KRAS mutations have been known to play roles in the initiation and progression of the tumorigenesis, respectively. However, many studies indicate that mutations of these two genes, which play roles in the Wnt/ß-catenin and RAS-extra-cellular signal regulated kinase (ERK) pathways, respectively, cooperatively interact in the tumorigenesis in several different cancer types including CRC. Both Apc and Kras mutations critically increase number and growth rate of tumors although single mutation of these genes does not significantly enhance the small intestinal tumorigenesis of mice. Both APC and KRAS mutations even result in the liver metastasis with inductions of the cancer stem cells (CSCs) markers in a mice xenograft model. In this review, we are going to describe the history for interaction between the Wnt/ß-catenin and RAS/ERK pathways especially related with CRC, and provide the mechanical basis for the cross-talk between the two pathways. The highlight of the crosstalk involving the stability regulation of RAS protein via the Wnt/ß-catenin signaling which is directly related with the cellular proliferation and transformation will be discussed. Activation status of GSK3ß, a key enzyme involving both ß-catenin and RAS degradations, is regulated by the status of the Wnt/ß-catenin signaling dependent upon extracellular stimuli or intracellular abnormalities of the signaling components. The levels of both ß-catenin and RAS proteins are co-regulated by the Wnt/ß-catenin signaling, and these proteins are overexpressed with a positive correlation in the tumor tissues of CRC patients. These results indicate that the elevation of both ß-catenin and RAS proteins is pathologically significant in CRC. In this review, we also will discuss further involvement of the increments of both ß-catenin and RAS especially mutant KRAS in the activation of CSCs and metastasis. Overall, the increments of ß-catenin and RAS especially mutant KRAS by APC loss play important roles in the cooperative tumorigenesis of CRC.