Genetic Risk Factors for Bortezomib-induced Neuropathic Pain in an Asian Population: A Genome-wide Association Study in South Korea.

Bortezomib-induced neuropathic pain (BINP) poses a challenge in multiple myeloma (MM) treatment. Genetic factors play a key role in BINP susceptibility, but research has predominantly focused on Caucasian populations. This research explored novel genetic risk loci and pathways associated with BINP development in Korean MM patients while evaluating the reproducibility of variants from Caucasians. Clinical data and buffy coat samples from 185 MM patients on bortezomib were collected. The cohort was split into discovery and validation cohorts through random stratification of clinical risk factors for BINP. Genome-wide association study was performed on the discovery cohort (n = 74) with Infinium Global Screening Array-24 v3.0 BeadChip (654,027 single nucleotide polymorphism [SNPs]). Relevant biological pathways were identified using the pathway scoring algorithm. The top 20 SNPs were validated in the validation cohort (n = 111). Previously reported SNPs were validated in the entire cohort (n = 185). Pathway analysis of the genome-wide association study results identified 31 relevant pathways, including immune systems and endosomal vacuolar pathways. Among the top 20 SNPs from the discovery cohort, 16 were replicated, which included intronic variants in ASIC2 and SMOC2, recently implicated in nociception, as well as intergenic variants or long noncoding RNAs. None of the 17 previously reported SNPs remained significant in our cohort (rs2274578, P = .085). This study represents the first investigation of novel genetic loci and biological pathways associated with BINP occurrence. Our findings, in conjunction with existing Caucasian studies, expand the understanding of personalized risk prediction and disease mechanisms. PERSPECTIVE: This article is the first to explore novel genetic loci and pathways linked to BINP in Korean MM patients, offering novel insights beyond the existing research focused on Caucasian populations into personalized risk assessment and therapeutic strategies of BINP.

CD19 chimeric antigen receptor T cell therapy in leukemia xenograft mouse: Anti-leukemic efficacy, kinetics, and 4-week single-dose toxicity.

CD19 Chimeric antigen receptor T (CAR-T) cell therapy has shown a promising response rate for relapsed/refractory B-cell malignancies. However, serious side effects such as cytokine release syndrome and immune effector cell-associated neurotoxicity syndrome arose in early case reports. Though several preclinical and clinical studies of CAR-T cell therapy have been reported, there is a lack of toxicological assessments. This study was carried out as a preclinical assessment of CD19 CAR-T cell therapy, including the anti-leukemic efficacy, kinetics in peripheral blood, and 4-week single-dose toxicity evaluation in leukemia xenograft mice. Leukemia xenograft mice model was established by injecting 1.0 × 105 cells/mouse of luciferase-labeled human B cell acute lymphoblastic leukemia (B-ALL) cell line via the tail vein, and after 3 days, 2.0 or 4.0 × 106 cells/mouse of CD19 CAR-T cells were injected intravenously. CD19 CAR-T cells showed significant anti-leukemic efficacy, showing inhibition of tumor progression in the bioluminescence-based in-vivo imaging system. In the kinetics study using qPCR, CAR-T cells peaked in peripheral blood on day 60 in males and day 30 in females. In a 4-week single-dose toxicity study, CD19 CAR-T cell injected groups showed no mortality and toxicological signs, or changes in body weight, food/water consumption, hematology, clinical chemistry, organ weights, and histopathology compared to control groups. These results suggested that 4.0 × 106 cells/mouse of CD19 CAR-T cells were effective in B-ALL xenograft mice without serious side effects, so the no-observed adverse effect level (NOAEL) was estimated to be higher than 4.0 × 106 cells/mouse, under the condition examined in the current study.

Tagmentation-based analysis reveals the clonal behavior of CAR-T cells in association with lentivector integration sites.

Integration site (IS) analysis is essential in ensuring safety and efficacy of gene therapies when integrating vectors are used. Although clinical trials of gene therapy are rapidly increasing, current methods have limited use in clinical settings because of their lengthy protocols. Here, we describe a novel genome-wide IS analysis method, "detection of the integration sites in a time-efficient manner, quantifying clonal size using tagmentation sequencing" (DIStinct-seq). In DIStinct-seq, a bead-linked Tn5 transposome is used, allowing the sequencing library to be prepared within a single day. We validated the quantification performance of DIStinct-seq for measuring clonal size with clones of known IS. Using ex vivo chimeric antigen receptor (CAR)-T cells, we revealed the characteristics of lentiviral IS. We then applied it to CAR-T cells collected at various times from tumor-engrafted mice, detecting 1,034-6,233 IS. Notably, we observed that the highly expanded clones had a higher integration frequency in the transcription units and vice versa in genomic safe harbors (GSH). Also, in GSH, persistent clones had more frequent IS. Together with these findings, the new IS analysis method will help to improve the safety and efficacy of gene therapies.

Stratification Based on Risk Factors at Diagnosis Could Predict Progression in Patients with Pancreatic Cysts.

BACKGROUND: Predicting the risk of malignant transformation in pancreatic cyst patients is challenging. AIM: We retrospectively investigated the risk factors for malignant transformation in pancreatic cyst patients. METHODS: Patients with pancreatic cysts diagnosed using imaging tests were followed from November 2008 to December 2021. A significant change was defined as the additional development of high-risk stigmata (HRS), worrisome features (WFs), or pancreatic cancer during monitoring. RESULTS: In total, 479 patients were analyzed, with a median observation period of 50 months. Forty-four patients (9.2%) showed significant changes, and eight (1.7%) developed pancreatic cancer. The univariate analysis showed that the cyst diameter at diagnosis (≥ 14 mm), main pancreatic duct (MPD) diameter at diagnosis (≥ 3 mm), presence of multilocular cysts, and an inconsistent MPD caliber were significant predictive factors for a significant change. One point was assigned for each significant factor. We grouped the patients into three groups: the low-risk group (total score 0), medium-risk group (score 1-2), and high-risk group (score 3-4). The high-risk group had a higher risk of a significant change than the medium- and low-risk groups (age-adjusted HRs for the medium-risk and high-risk groups were 3.0 and 5.2 compared with the low-risk group). CONCLUSION: Stratification based on risk factors may help predict the development of significant changes in pancreatic cyst patients.

Changes in physical activity and energy intake according to abdominal obesity in Korean adult men before and after COVID-19: Data from 2019 and 2020 Korea National Health and Nutrition Examination Survey (KNHANES).

PURPOSE: This study aimed to investigate changes in physical activity and energy intake according to abdominal obesity in Korean adult men before and after COVID-19. METHODS: Using data from the 2019 and 2020 KNHANES, the physical activity level measured by the Global Physical Activity Questionnaire (GPAQ) the physical activity level by GPAQ, number of days of walking and strength training, aerobic exercise, and total energy, protein, fat, carbohydrate, dietary fiber, and sugar intake for a total of 2,799 participants were analyzed. RESULTS: There were no changes in energy intake during the pandemic. The number of days of weekly walking was higher (2019, p = 0.006; 2020, p = 0.012) and strength training was significantly higher (2019, p < 0.0001; 2020 p < 0.0001) in the non-abdominal obesity group than in the abdominal obesity group before and after COVID-19. Strength training at least once a week suppressed abdominal obesity (0.628 times in 2019, p < 0.0001; 0.605 times in 2020, p < 0.0001). In addition, even when the total energy intake and age were adjusted for, strength training influenced the suppression of abdominal obesity (0.634 times in 2019, p < 0.0001; 0.614 times in 2020, p < 0.0001). CONCLUSION: Even with the change in the physical activity level, such as walking and aerobic exercise, due to the influence of social distancing measures, strength training influenced the suppression of abdominal obesity, regardless of the COVID-19 pandemic.

Efficacy and Safety of a Strategy for Reviewing Intravenous Antibiotics for Hospitalized Japanese Patients with Uncomplicated Diverticulitis: A Single-center Observational Study.

Objective Treatment for uncomplicated diverticulitis (UD) is not well established. We evaluated the strategy of reviewing intravenous antibiotics for hospitalized Japanese patients with UD. Methods Treatment was based on the physician's choice until August 2018; the indications for hospitalization and treatment have been standardized since September 2018. In this study, we monitored the use of intravenous antibiotics administered to patients hospitalized for UD and then reviewed the need for them on hospital day 3. We compared patients' length of antibiotic use, hospital stay, health care cost, and complications via the review strategy from September 2018 to December 2020 and via the previous physicians' choice strategy from January 2016 to August 2018. Results Two hundred and forty-seven patients were admitted to our hospital because of acute colonic diverticulitis from January 2016 to December 2020. After excluding complicated cases, 106 individuals were enrolled during the period of physician's choice; 87 were enrolled when treatment review was employed. There were no significant differences in age, sex, inflammation site, or severity during the first hospital visit. The median duration of antibiotic use was significantly reduced from 5 to 4 days (p=0.0075), with no marked increase in rates of transfer to surgery, mortality, or readmission due to recurrence. A more significant proportion of patients completed 3-day antibiotic treatment with the review strategy than with the physician's choice strategy (6.6% vs. 25.3%, p=0.0004). However, the length of hospital stay and total medical costs did not decrease. Conclusion The strategy of reviewing treatment on day 3 after hospitalization for UD safety reduced the duration of antibiotic use, but the hospital stay and health care costs did not decrease.

Factors Influencing the Eating Behaviors of Young Adults Using the 2013-2018 Korea National Health and Nutrition Examination Survey.

Background: There is a substantial gap in the eating behaviors between the sexes. We aimed to analyze the predictors of eating behaviors by sex in the young adult population. Methods: We used the data of the sixth and seventh Korea National Health and Nutrition Examination Survey conducted by the Korea Disease Control and Prevention Agency last 2013-2018. Data from 2,502 women and 2,101 men aged 19-29 yr were included in the analysis using frequency, percentage, x 2-test, and multiple logistic regression. Results: Education (0.612, CI=0.465∼0.805), economic status (2.104, CI=1.435-3.086), marital status (3.162, CI=2.356∼4.243), and household structure (0.403, CI=0.208∼0.782) were identified to predict the frequency of dining-out in women, while marital status (0.302, CI=0.121∼0.749), economic activity (1.969, CI=1.483∼2.613), and household structure (0.243, CI=0.137∼0.432) predicted dining-out frequency in men. The current smoking status predicted most eating behaviors, including breakfast skipping frequency (1.864, CI=1.318∼2.637), use of supplements (2.062, CI=1.439∼2.953), and use of nutrition labels (1.545, 1.084∼2.204) for men. Meanwhile, nutrition labeling was used less in both men (0.550, CI=0.343∼0.882) and women (0.646, CI=0.473∼0.882) who subjective body recognition as obesity. Conclusion: The factors that primarily predicted the frequency of dining out in young women and the behavior of breakfast skipping, use of nutrition labels, and frequency of dining-out in men can be used as foundational data for developing sex-specific intervention programs to improve eating behaviors.

A TRPV1 antagonist, PAC-14028 does not increase the risk of tumorigenesis in chemically induced mouse skin carcinogenesis.

PAC-14028 (Asivatrep: C21H22F5N3O3S) cream is a novel, topical nonsteroidal, anti-inflammatory, and TRPV1 (transient receptor potential vanilloid subfamily, member 1) antagonist for the treatment of mild to moderate atopic dermatitis. Concerns about the risk of tumor development by TRPV1 blockade in the skin have been prompted, but these findings were proved to be indirect or are still controversial. This study was tested to determine whether TRPV1 selective antagonist, PAC-14028 cream is safe from the promotion of skin tumorigenesis in the two-stage carcinogenesis model. PAC-14028 cream, 0.25%, 0.5%, or 1.0% was applied once daily topically to mouse skin for up to 24 weeks in two-stage chemical carcinogenesis testing using 7, 12-dimethylbenz[a]anthracene (DMBA) and 12-O-tetradecanoylphorbol-13-acetate (TPA). Morbidity/death, clinical signs, tumor formation, activity of EGFR/Akt/mTOR signaling, and systemic exposure to PAC-14028 were investigated. Daily dermal administration of PAC-14028, was not skin carcinogenic. There was also no evidence on the activation of EGFR/Akt/mTOR signaling pathway by the topical treatment of PAC-14028. On Day 169, 1.0% (20 mg/kg/day) of PAC-14028 in female mice resulted in a Cmax and AUC0-τ of 12916.0 ng/mL and 78962.9 ng‧hr/mL, respectively. PAC-14028 cream was well tolerated and did not increase the risk of skin tumorigenesis in two-stage carcinogenesis study.

Dietary fat intake and age modulate the composition of the gut microbiota and colonic inflammation in C57BL/6J mice.

BACKGROUND: More than half of the adult population worldwide is overweight or obese, while excess adiposity has been linked to chronic low-grade inflammation, contributing to the development of chronic diseases. Recent studies have showed that diet-induced alterations to the gut microbiota composition play a pivotal role in the development of obesity. However, the cause-effect relationship between obesity and gut microbiota composition is not yet fully understood. In this study, we investigated the short-term responses of gut microbiota composition to diets with different fat contents and their associations with inflammatory biomarkers. RESULTS: Sixty male C57BL/6 J mice were fed a normal diet (ND; 15% fat) or a high-fat diet (HFD; 45% fat) for 10 or 20 weeks. The relative proportion of the phylum Actinobacteria was elevated by the HFD and was positively associated with body weight and proinflammatory cytokines including TNF-α, IL-1ß, and IL-6. The proportion of the phylum Firmicutes increased with aging and was also positively correlated with proinflammatory cytokines. The proportions of Actinobacteria and Firmicutes were inversely associated with tight junction proteins claudin-1 and E-cadherin, respectively. The proportions of the class Clostridia and the family Ruminococcaceae within the phylum Firmicutes were affected by both diet and age. In addition, the proportions of the phylum Bacteroidetes, the family Bacteroidaceae, and the genus Bacteroides decreased with aging and were inversely correlated with colonic proinflammatory cytokines representing a positive association with tight junction proteins. CONCLUSIONS: Host age and dietary fat intake are important elements that induce proportional changes in gut microbiota, and these changes are also associated with systemic inflammation. This study provides evidence that diet affects the gut microbiota composition within a short period of time.

GM-CSF Promotes the Expansion and Differentiation of Cord Blood Myeloid-Derived Suppressor Cells, Which Attenuate Xenogeneic Graft-vs.-Host Disease.

Myeloid-derived suppressor cells (MDSCs) are increased in tumor patients. Studies have shown generation of MDSCs from human peripheral blood mononuclear cells (PBMCs) by various cytokine combinations. However, large scale expansion of human MDSCs has not been demonstrated or applied in clinic settings. We investigated which cytokine combinations among GM-CSF/SCF, G-CSF/SCF, or M-CSF/SCF efficiently expand and differentiate human MDSCs following culture CD34+ cells of umbilical cord blood (CB). GM-CSF/SCF showed the greatest expansion of MDSCs. Up to 108 MDSCs (HLA-DRlowCD11b+CD33+) could be produced from 1 unit of CB following 6 weeks of continuous culture. MDSCs produced from culture of CD34+ cells with GM-CSF/SCF for 6 weeks had the greatest suppressive function of T cell proliferation and had the highest expression of immunosuppressive molecules including iNOS, arginase 1 and IDO compared to those differentiated with G-CSF/SCF or M-CSF/SCF. MDSCs secreted IL-10, TGB-ß, and VEGF. The infusion of expanded MDSCs significantly prolonged the survival and decreased the GVHD score in a NSG xenogeneic model of GVHD. Injected MDSCs increased IL-10 and TGF-ß but decreased the level of TNF-α and IL-6 in the serum of treated mice. Notably, FoxP3 expressing regulatory T (Treg) cells were increased while IFN-γ (Th1) and IL-17 (Th17) producing T cells were decreased in the spleen of MDSC treated mice compared to untreated GVHD mice. Our results demonstrate that human MDSCs are generated from CB CD34+ cells using GM-CSF/SCF. These MDSCs exhibited potent immunosuppressive function, suggesting that they are useable as a treatment for inflammatory diseases such as GVHD.

Anti-psoriatic effect of myeloid-derived suppressor cells on imiquimod-induced skin inflammation in mice.

Myeloid-derived suppressor cells (MDSCs) play an important role in controlling the immune response against cancer and in suppression of autoimmunity and allergic inflammation. However, the beneficial effects of MDSCs on the experimental mouse model of psoriasis have not been reported. Therefore, we investigated the anti-psoriatic effect of MDSCs on IMQ-induced skin inflammation in mice and explored the mechanisms involved. Our results showed that administration of MDSCs (1 × 106 or 2 × 106  cells) suppressed the development of IMQ-induced skin inflammation in mice as exemplified by a significant reduction in clinical severity scores and was associated with a reduction of histopathological changes, including inflammatory infiltration, epidermal hyperplasia and hyperkeratosis. The immunosuppressive effect of MDSCs (1 × 106 or 2 × 106  cells) corresponded to the production of Th1 cytokines (TNF-α, IFN-γ) and Th17 cytokines (IL-17A and IL-23) in the serum and dorsal skin. Administration of MDSCs (1 × 106 or 2 × 106  cells) also inhibited splenomegaly. Moreover, an increased percentage of CD4+ CD25+ FoxP3+ regulatory T (Treg) cells and decreased percentage of Th1 and Th17 cells were found in mice treated with MDSCs. Taken together, these results imply that MDSCs have immunomodulatory and immunosuppressive effects on disease progression in a murine model of psoriasis and that MDSCs could be used in preventive or therapeutic strategies for the management of autoimmune inflammatory skin disorders, such as psoriasis.

Ex Vivo Generated Human Cord Blood Myeloid-Derived Suppressor Cells Attenuate Murine Chronic Graft-versus-Host Diseases.

Myeloid-derived suppressor cells (MDSCs) are a heterogeneous population of immature myeloid cells with anti-inflammatory activity, and expanded murine MDSCs are capable of attenuating preclinical acute graft-versus-host disease (aGVHD) severity. Two murine cGVHD models were used to evaluate the effectiveness of ex vivo cultured human cord blood (hCB) MDSCs in chronic GVHD (cGVHD). First, GVHD recipients surviving in a classic C57BL/6 into MHC-mismatched BALB/c aGVHD model developed cGVHD. Second, donor pretreatment with granulocyte colony-stimulating factor (G-CSF) induced cGVHD. hCB-MDSCs (1 × 106) were intravenously injected to determine their preventive effects (on days 5, 7, 10, and 21) or therapeutic effects (on days 21, 28, and 35). In the first model the onset of clinical cutaneous cGVHD was significantly delayed in preventive hCB-MDSCs-treated allogeneic recipients. Pathologic scoring of target organs confirmed these clinical results. Importantly, thymic tissues of GVHD mice treated with hCB-MDSCs were less severely damaged, showing higher numbers of double (CD4 and CD8) positive T cells with reduced expansion of donor-type CD4 and CD8 T cells. Moreover, late infusion of hCB-MDSCs controlled the severity of established cGVHD that had occurred in control recipients. In the second model, cGVHD induced by G-CSF-mobilized stem cell graft was associated with promotion of Th 17 and Th 2 differentiation. hCB-MDSCs attenuated clinical and pathologic cGVHD severity. Increased production of IL-17 and more infiltration of T cells and macrophages in cGVHD mice were markedly reduced after hCB-MDSCs treatment. Importantly, Foxp3+ regulatory T cells and IFN-γ-producing T cells were expanded, whereas IL-17- and IL-4-producing T cells were decreased in allogeneic recipients of hCB-MDSCs. Taken together, these results showed that hCB-MDSCs have preclinical capability of attenuating cGVHD by preserving thymus function and regulating Th 17 signaling, suggesting a possible therapeutic strategy for clinical application.

5α-dihydrotestosterone reduces renal Cyp24a1 expression via suppression of progesterone receptor.

Androgens act in concert with vitamin D to influence reabsorption of calcium. However, it is unclear whether androgens directly regulate vitamin D homeostasis or control other cellular events that are related to vitamin D metabolism. To examine whether the expression of vitamin D-related genes in mouse kidney is driven by androgens or androgen-dependent effects, the androgen receptor and other sex steroid receptors were monitored in orchidectomized mice treated with 5α-dihydrotestosterone (DHT). Our results revealed that exposing orchidectomized mice to DHT inhibited the expression of progesterone receptor (Pgr) with or without estrogen receptor α expression, the latter was confirmed by ER-positive (MCF7 and T47D) or -negative (PCT) cells analysis. The loss of Pgr in turn decreased the expression of renal 24-hydroxylase via transcriptional regulation because Cyp24a1 gene has a progesterone receptor-binding site on promoter. When male kidneys preferentially hydroxylate 25-hydroxyvitamin D3 using 24-hydroxylase rather than 25-hydroxyvitamin D3-1-alpha hydroxylase, DHT suppressed the Pgr-mediated 24-hydroxylase expression, and it is important to note that DHT increased the blood 25-hydroxyvitamin D3 levels. These findings uncover an important link between androgens and vitamin D homeostasis and suggest that therapeutic modulation of Pgr may be used to treat vitamin D deficiency and related disorders.

Do TRPV1 antagonists increase the risk for skin tumourigenesis? A collaborative in vitro and in vivo assessment.

A recent hypothesis suggesting that the pharmacological target TRPV1 (transient receptor potential vanilloid subfamily, member 1) may function as a tumour suppressor, which potentially impacts the development of TRPV1 antagonist therapeutics for a range of conditions. However, little is known about the long-term physiologic effects of TRPV1 blockade in the skin. In vitro and in vivo studies suggested that the potent TRPV1 competitive antagonist AMG-9810 promoted proliferation in N/TERT1 cells (telomerase-immortalised primary human keratinocytes 1) and tumour development in mouse skin that was mediated through EGFR/Akt/mTOR signalling. We attempted to reproduce the reported in vitro and in vivo findings to further explore this hypothesis to understand the underlying mechanism and the risk associated with TRPV1 antagonism in the skin. In vitro proliferation studies using multiple methods and topical application with AMG-9810 and structurally similar TRPV1 antagonists such as SB-705498 and PAC-14028 were performed. Although we confirmed expression of TRPV1 in primary human epidermal keratinocytes (HEKn) and spontaneously immortalised human keratinocytes (HaCaT), we were unable to demonstrate cell proliferation in either cell type or any clear evidence of increased expression of proteins in the EGFR/Akt/mTOR signalling pathway with these molecules. We were also unable to demonstrate skin tumour promotion or underlying molecular mechanisms involved in the EGFR/Akt/mTOR signalling pathway in a single-dose and two-stage carcinogenesis mouse study treated with TRPV1 antagonists. In conclusion, our data suggest that inhibiting the pharmacological function of TRPV1 in skin by specific antagonists has not been considered to be indicative of skin tumour development.

Association of daily physical activity level with health-related factors by gender and age-specific differences among Korean adults based on the sixth (2014-2015) Korea national health and nutrition examination survey

PURPOSE: This study examined the effects of daily physical activity level on health-related factors according to gender and identified age-specific differences among Korean adults. METHODS: Using data from the Korea National Health and Nutrition Examination Survey VI (2014-2015), we selected adults aged 19-64 years who participated in both a health examination and health interview survey. The study included 6,457 participants 19-64 years of age (2,611 men, 3,846 women). RESULTS: Assessment of the differences in health-related factors according to age and physical activity in men and women by repeated two-way analysis of variance (ANOVA) revealed significant interaction effects on total cholesterol (TC) and triglyceride (TG) levels and diastolic blood pressure (DBP) in male participants, but there were no significant interaction effects for any health-related factors in female participants. The group of female participants aged 40-64 years with daily physical activity levels over 200 kcal showed a significantly increased prevalence of 46% for dyslipidemia compared to that in female participants with daily physical activity levels below 200 kcal. Physical activity was positively correlated with weight and high-density lipoprotein cholesterol (HDL-C) levels in men 19-39 years of age, compared to weight, waist circumference (WC), body mass index (BMI), and DBP in men 40-64 years of age, and weight, WC, BMI, glycated hemoglobin (HbA1c) and triglyceride (TG) levels in women 19-39 years of age. In women 40- 64 years of age, physical activity was especially significantly positively correlated with weight, BMI, HDL-C and negatively correlated with fasting glucose and TG levels. CONCLUSION: In male and female participants, the 40-64-year age group showed negative results for health-related factors compared to those in the 19-39-year age group. The higher the weight, WC, BMI, the higher is the physical activity level. Physical activity levels were significantly positively correlated with health-related variables.

Generation of lung cancer cell lines harboring EGFR T790M mutation by CRISPR/Cas9-mediated genome editing.

Tyrosine kinase inhibitors (TKIs) such as gefitinib and erlotinib are effective against lung adenocarcinomas harboring epidermal growth factor receptor (EGFR) mutations. However, cancer cells can develop resistance to these agents with prolonged exposure; in over 50% of cases, this is attributable to the EGFR T790M mutation. Moreover, additional resistance mutations can arise with the use of new drugs. Cancer cell lines with specific mutations can enable the study of resistance mechanisms. In this study, we introduced the EGFR T790M mutation into the PC9 human lung cancer cell line-which has a deletion in exon 19 of the EGFR gene-by clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas)9-mediated genome editing. EGFR pyrosequencing and peptide nucleic acid clamping revealed that PC9 cells with EGFR T790M generated by CRISPR/Cas 9 had a higher T790M mutation rate than those with the same mutation generated by long-term exposure to gefitinib (PC9-G); moreover, resistance to gefitinib in these clones was higher than that in PC9-G cells. The clones were also highly sensitive to the 3rd-generation EGFR TKI AZD9291, which is cytotoxic to lung cancer cells with EGFR T790M. The CRISPR/Cas9 programmable nuclease system can be used to generate various cancer cell lines with specific mutations that can facilitate studies on resistance mechanisms and drug efficacy.

Erratum: High-fat Diet Accelerates Intestinal Tumorigenesis Through Disruption Intestinal Cell Membrane Integrity.

[This corrects the article on p. 95 in vol. 21, PMID: 27390738.].

Protective effects of agonistic anti-4-1BB antibody on the development of imiquimod-induced psoriasis-like dermatitis in mice.

Agonistic anti-4-1BB antibodies (Abs) play a central role in immunomodulatory conditions that control the pathogenesis of immune-mediated autoimmune and allergic diseases. However, the effects of agonistic anti-4-1BB Abs have not been examined in an experimental mouse model of psoriasis. Therefore, we investigated the protective effects of agonistic anti-4-1BB Abs, using imiquimod (IMQ)-induced psoriasis-like dermatitis in mice, a condition histologically and clinically similar to human psoriasis. We found that administration of agonistic anti-4-1BB Abs (10mg/kg) significantly alleviated the severity of IMQ-induced psoriasis-like skin inflammation in mice, with reduced histologic symptoms, including inflammatory infiltration, parakeratosis, and hyperkeratosis. Subsequent analyses revealed that the production of Th17 cytokines (IL-17A and IL-23) in the serum and skin of IMQ-induced mice was significantly inhibited by agonistic anti-4-1BB Abs (10mg/kg), although Th1 cytokines (TNF-α and IFN-γ) were not. Moreover, administration of agonistic anti-4-1BB Abs (10mg/kg) induced a relative increase of CD4(+)FoxP3(+) regulatory T (Treg) cells in the spleen and draining lymph node (DLN). Taken together, our data provide evidence that agonistic anti-4-1BB Abs possesses immunosuppressive properties in IMQ-induced psoriasis-like skin inflammation, providing insight into the immunomodulatory effect of agonistic anti-4-1BB Abs for psoriasis immunotherapy.

High-fat Diet Accelerates Intestinal Tumorigenesis Through Disrupting Intestinal Cell Membrane Integrity.

BACKGROUND: Excess energy supply induces chronic low-grade inflammation in association with oxidative stress in various tissues including intestinal epithelium. The objective of this study was to investigate the effect of high-fat diet (HFD) on intestinal cell membrane integrity and intestinal tumorigenesis in Apc(Min/+) mice. METHODS: Mice were fed with either normal diet (ND) or HFD for 12 weeks. The number of intestinal tumors were counted and biomarkers of endotoxemia, oxidative stress, and inflammation were determined. Changes in intestinal integrity was measured by fluorescein isothiocyanate (FITC)-dextran penetration and membrane gap junction protein expression. RESULTS: HFD group had significantly higher number of tumors compared to ND group (P < 0.05). Blood total antioxidant capacity was lower in HFD group, while colonic 8-hydroxy-2'-deoxyguanosine level, a marker of oxidative damage, was higher in HFD group compared to that of ND group (P < 0.05). The penetration of FITC-dextran was substantially increased in HFD group (P < 0.05) while the expressions of membrane gap junction proteins including zonula occludens-1, claudin-1, and occludin were lower in HFD group (P < 0.05) compared to those in ND group. Serum concentration of lipopolysaccharide (LPS) receptor (CD14) and colonic toll-like receptor 4 (a LPS receptor) mRNA expression were significantly higher in HFD group than in ND group (P < 0.05), suggesting that significant endotoxemia may occur in HFD group due to the increased membrane permeability. Serum interleukin-6 concentration and myeloperoxidase activity were also higher in HFD group compared to those of ND group (P < 0.05). CONCLUSIONS: HFD increases oxidative stress disrupting intestinal gap junction proteins, thereby accelerating membrane permeability endotoxemia, inflammation, and intestinal tumorigenesis.

Genome-wide association study for the interaction between BMR and BMI in obese Korean women including overweight.

BACKGROUND/OBJECTIVES: This is the first study to identify common genetic factors associated with the basal metabolic rate (BMR) and body mass index (BMI) in obese Korean women including overweight. This will be a basic study for future research of obese gene-BMR interaction. SUBJECTS/METHODS: The experimental design was 2 by 2 with variables of BMR and BMI. A genome-wide association study (GWAS) of single nucleotide polymorphisms (SNPs) was conducted in the overweight and obesity (BMI > 23 kg/m(2)) compared to the normality, and in women with low BMR (< 1426.3 kcal/day) compared to high BMR. A total of 140 SNPs reached formal genome-wide statistical significance in this study (P < 1 × 10(-4)). Surveys to estimate energy intake using 24-h recall method for three days and questionnaires for family history, a medical examination, and physical activities were conducted. RESULTS: We found that two NRG3 gene SNPs in the 10q23.1 chromosomal region were highly associated with BMR (rs10786764; P = 8.0 × 10(-7), rs1040675; 2.3 × 10(-6)) and BMI (rs10786764; P = 2.5 × 10(-5), rs10786764; 6.57 × 10(-5)). The other genes related to BMI (HSD52, TMA16, MARCH1, NRG1, NRXN3, and STK4) yielded P <10 × 10(-4). Five new loci associated with BMR and BMI, including NRG3, OR8U8, BCL2L2-PABPN1, PABPN1, and SLC22A17 were identified in obese Korean women (P < 1 × 10(-4)). In the questionnaire investigation, significant differences were found in the number of starvation periods per week, family history of stomach cancer, coffee intake, and trial of weight control in each group. CONCLUSION: We discovered several common BMR- and BMI-related genes using GWAS. Although most of these newly established loci were not previously associated with obesity, they may provide new insights into body weight regulation. Our findings of five common genes associated with BMR and BMI in Koreans will serve as a reference for replication and validation of future studies on the metabolic rate.