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Metabolic dysfunction-associated steatohepatitis (MASH) is a form of chronic liver inflammation associated with metabolic syndrome, such as obesity and a major cause of hepatocellular carcinoma (HCC). Multi-biotics, a soymilk fermented with lactic acid bacteria, are known to alleviate obesity by lowering lipid profile. This study aimed to establish and characterize mouse organoids derived from MASH-related HCC models to evaluate drug responses, particularly focusing on Lenvatinib resistance. Organoids were developed using mouse liver tissues subjected to a choline-deficient L-amino acid-defined high-fat diet (CDAHFD) to mimic MASH-related HCC. The study evaluated the effect of multi-biotics, a fermented product, on tumor regression and drug sensitivity. While multi-biotics did not reduce tumor burden, they enhanced the response to Lenvatinib. Additionally, repeated treatment with Lenvatinib led to the development of drug-resistant organoids. Transcriptomic analysis of these resistant organoids identified key pathways related to KRAS signaling, inflammation, and epithelial-mesenchymal transition (EMT), revealing potential targets for overcoming Lenvatinib resistance. This study provides valuable insights into MASH-related HCC progression and drug resistance, offering a model for further therapeutic research., Competing Interests: Declarations Competing interests The authors declare no competing interests., (© 2024. The Author(s).)

Cellular and molecular dynamics of human cells are constantly affected by gravity. Alteration of the gravitational force disturbs the cellular equilibrium, which might modify physiological and molecular characteristics. Nevertheless, biological responses of cancer cells to reduced gravitational force remains obscure. Here, we aimed to comprehend not only transcriptomic patterns but drug responses of colorectal cancer (CRC) under simulated microgravity. We established four organoids directly from CRC patients, and organoids cultured in 3D clinostat were subjected to genome wide expression profiling and drug library screening. Our observations revealed changes in cell morphology and an increase in cell viability under simulated microgravity compared to their static controls. Transcriptomic analysis highlighted a significant dysregulation in the TBC1D3 family of genes. The upregulation of cell proliferation observed under simulated microgravity conditions was further supported by enriched cell cycle processes, as evidenced by the functional clustering of mRNA expressions using cancer hallmark and gene ontology terms. Our drug screening results indicated an enhanced response rate to 5-FU under conditions of simulated microgravity, suggesting potential implications for cancer treatment strategies in simulated microgravity., (© 2024. The Author(s).)

Purpose: Histologic transformation from EGFR-mutant non-small cell lung cancer (NSCLC) to small-cell lung cancer (SCLC) is a key mechanism of resistance to EGFR tyrosine kinase inhibitors (TKI). However, transcriptomic changes between NSCLC and transformed SCLC (t-SCLC) remain unexplored., Experimental Design: We conducted whole-transcriptome analysis of 59 regions of interest through the spatial profiling of formalin-fixed, paraffin-embedded tissues obtained from 10 patients (lung adenocarcinoma, 22; combined SCLC/NSCLC, 7; and t-SCLC, 30 regions of interests). Transcriptomic profiles and differentially expressed genes were compared between pre- and post-transformed tumors., Results: Following EGFR-TKI treatment, 93.7% (15/16) of t-SCLC components evolved into neuroendocrine-high subtypes (SCLC-A or SCLC-N). The transition to t-SCLC occurred regardless of EGFR-TKI treatment and EGFR mutational status, with a notable decrease in EGFR expression (P < 0.001) at both mRNA and protein levels. Pathway analysis revealed that gene overexpression was related to epigenetic alterations in t-SCLC. Interestingly, histone deacetylase inhibitors restored EGFR expression in SNU-2962A cells and their organoid model. The synergistic effects of third-generation EGFR-TKI osimertinib and the histone deacetylase inhibitor fimepinostat were validated in both in vitro and in vivo models., Conclusions: Our study demonstrated that most t-SCLC cases showed neuronal subtypes with low EGFR expression. Differentially expressed gene analysis and t-SCLC preclinical models identified an epigenetic modifier as a promising treatment strategy for t-SCLC., (©2024 American Association for Cancer Research.)

Sarcomas, malignant tumors from mesenchymal tissues, exhibit poor prognosis despite advancements in treatment modalities such as surgery, radiotherapy, and chemotherapy, with doxorubicin being a cornerstone treatment. Resistance to doxorubicin remains a significant hurdle in therapy optimization. This study aims to dissect the molecular bases of doxorubicin resistance in sarcoma cell lines, which could guide the development of tailored therapeutic strategies. Eighteen sarcoma cell lines from 14 patients were established under ethical approvals and classified into seven subtypes. Molecular, genomic, and transcriptomic analyses included whole-exome sequencing, RNA sequencing, drug sensitivity assays, and pathway enrichment studies to elucidate the resistance mechanisms. Variability in doxorubicin sensitivity was linked to specific genetic alterations, including mutations in TP53 and variations in the copy number of genomic loci like 11q24.2. Transcriptomic profiling divided cell lines into clusters by karyotype complexity, influencing drug responses. Additionally, pathway analyses highlighted the role of signaling pathways like WNT/BETA-CATENIN and HEDGEHOG in doxorubicin-resistant lines. Comprehensive molecular profiling of sarcoma cell lines has revealed complex interplays of genetic and transcriptomic factors dictating doxorubicin resistance, underscoring the need for personalized medicine approaches in sarcoma treatment. Further investigations into these resistance mechanisms could facilitate the development of more effective, customized therapy regimens., (© 2024. The Author(s).)

Purpose: Early dissemination of primary pancreatic ductal adenocarcinoma (PDAC) is the main cause of dismal prognosis as it highly limits possible treatment options. A number of PDAC patients experience distant metastasis even after treatment due to the metastatic clones. We aimed to demonstrate the molecular architecture of borderline resectable PDAC manifests cancer dissemination of PDAC., Methods: Here, 36 organoids isolated from primary tumor masses of PDAC patients with diverse metastatic statues are presented. Whole-exome sequencing and RNA sequencing were performed and drug responses to clinically relevant 18 compounds were assessed., Results: Our results revealed that borderline resectable PDAC organoids exhibited distinct patterns according to their metastatic potency highlighted by multiple genetic and transcriptional factors and strong variances in drug responses., Conclusions: These data suggest that the presence of metastatic PDAC can be identified by integrating molecular compositions and drug responses of borderline resectable PDAC., (© 2024. The Author(s).)

Metabolic dysfunction-associated steatotic liver disease (MASLD) is a prevalent metabolic disorder characterized by hepatic steatosis associated with metabolic abnormalities. Recent research has shed light on the intricate interplay among interleukin-1 receptor 1 (IL1R1), gut microbiota, and bile acids in the pathogenesis of MASLD. This review aims to provide a comprehensive overview of the current understanding of the role of IL1R1, gut microbiota, and bile acids in MASLD, exploring their interrelationships and potential mechanisms. We summarize the evidence supporting the involvement of IL1R1 in inflammation, discuss the influence of gut microbiota on bile acid metabolism and its influence on liver health, and elucidate the bidirectional interactions among IL1R1 signaling, gut microbiota composition, and bile acid homeostasis in MASLD. Furthermore, we highlight emerging therapeutic strategies targeting these interrelated pathways for the management of MASLD., (© 2024 The Author(s). Journal of Gastroenterology and Hepatology published by Journal of Gastroenterology and Hepatology Foundation and John Wiley & Sons Australia, Ltd.)

Glioblastoma (GBM), the most lethal primary brain cancer, exhibits intratumoral heterogeneity and molecular plasticity, posing challenges for effective treatment. Despite this, the regulatory mechanisms underlying such plasticity, particularly mesenchymal (MES) transition, remain poorly understood. In this study, we elucidate the role of the RNA-binding protein ELAVL2 in regulating aggressive MES transformation in GBM. We found that ELAVL2 is most frequently deleted in GBM compared to other cancers and associated with distinct clinical and molecular features. Transcriptomic analysis revealed that ELAVL2-mediated alterations correspond to specific GBM subtype signatures. Notably, ELAVL2 expression negatively correlated with epithelial-to-mesenchymal transition (EMT)-related genes, and its loss promoted MES process and chemo-resistance in GBM cells, whereas ELAVL2 overexpression exerted the opposite effect. Further investigation via tissue microarray analysis demonstrated that high ELAVL2 protein expression confers a favorable survival outcome in GBM patients. Mechanistically, ELAVL2 was shown to directly bind to the transcripts of EMT-inhibitory molecules, SH3GL3 and DNM3, modulating their mRNA stability, potentially through an m6A-dependent mechanism. In summary, our findings identify ELAVL2 as a critical tumor suppressor and mRNA stabilizer that regulates MES transition in GBM, underscoring its role in transcriptomic plasticity and glioma progression., (© 2024. The Author(s).)

In pancreatic ductal adenocarcinoma (PDAC), no recurrent metastasis-specific mutation has been found, suggesting that epigenetic mechanisms, such as DNA methylation, are the major contributors of late-stage disease progression. Here, we performed the first whole-genome bisulfite sequencing (WGBS) on mouse and human PDAC organoid models to identify stage-specific and molecular subtype-specific DNA methylation signatures. With this approach, we identified thousands of differentially methylated regions (DMRs) that can distinguish between the stages and molecular subtypes of PDAC. Stage-specific DMRs are associated with genes related to nervous system development and cell-cell adhesions, and are enriched in promoters and bivalent enhancers. Subtype-specific DMRs showed hypermethylation of GATA6 foregut endoderm transcriptional networks in the squamous subtype and hypermethylation of EMT transcriptional networks in the progenitor subtype. These results indicate that aberrant DNA methylation contributes to both PDAC progression and subtype differentiation, resulting in significant and reoccurring DNA methylation patterns with diagnostic and prognostic potential., Competing Interests: The authors declare no competing interests., (© 2024 The Author(s).)

Although gemcitabine-based regimens are widely used as an effective treatment for pancreatic cancer, acquired resistance to gemcitabine has become an increasingly common problem. Therefore, a novel therapeutic strategy to treat gemcitabine-resistant pancreatic cancer is urgently required. Piceamycin has been reported to exhibit antiproliferative activity against various cancer cells; however, its underlying molecular mechanism for anticancer activity in pancreatic cancer cells remains unexplored. Therefore, the present study evaluated the antiproliferation activity of piceamycin in a gemcitabine-resistant pancreatic cancer cell line and patient-derived pancreatic cancer organoids. Piceamycin effectively inhibited the proliferation and suppressed the expression of alpha-actinin-4 , a gene that plays a pivotal role in tumorigenesis and metastasis of various cancers, in gemcitabine-resistant cells. Long-term exposure to piceamycin induced cell cycle arrest at the G 0 /G 1 phase and caused apoptosis. Piceamycin also inhibited the invasion and migration of gemcitabine-resistant cells by modulating focal adhesion and epithelial-mesenchymal transition biomarkers. Moreover, the combination of piceamycin and gemcitabine exhibited a synergistic antiproliferative activity in gemcitabine-resistant cells. Piceamycin also effectively inhibited patient-derived pancreatic cancer organoid growth and induced apoptosis in the organoids. Taken together, these findings demonstrate that piceamycin may be an effective agent for overcoming gemcitabine resistance in pancreatic cancer.

Glioblastoma (GBM) is the most lethal intracranial tumor. Sequencing technologies have supported personalized therapy for precise diagnosis and optimal treatment of GBM by revealing clinically actionable molecular characteristics. Although accumulating sequence data from brain tumors and matched normal tissues have facilitated a comprehensive understanding of genomic features of GBM, these in silico evaluations could gain more biological credibility when they are verified with in vitro and in vivo models. From this perspective, GBM cell lines with whole exome sequencing (WES) datasets of matched tumor tissues and normal blood are suitable biological platforms to not only investigate molecular markers of GBM but also validate the applicability of druggable targets. Here, we provide a complete WES dataset of 26 GBM patient-derived cell lines along with their matched tumor tissues and blood to demonstrate that cell lines can mostly recapitulate genomic profiles of original tumors such as mutational signatures and copy number alterations., (© 2023. The Author(s).)

Krukovine (KV) is an alkaloid isolated from the bark of Abuta grandifolia (Mart.) Sandw. (Menispermaceae) with anticancer potential in some cancers with KRAS mutations. In this study, we explored the anticancer efficacy and mechanism of KV in oxaliplatin-resistant pancreatic cancer cells and patient-derived pancreatic cancer organoids (PDPCOs) with KRAS mutation. After treatment with KV, mRNA and protein levels were determined by RNA-seq and Western blotting, respectively. Cell proliferation, migration, and invasion were measured by MTT, scratch wound healing assay, and transwell analysis, respectively. Patient-derived pancreatic cancer organoids (PDPCOs) with KRAS mutations were treated with KV, oxaliplatin (OXA), and a combination of KV and OXA. KV suppresses tumor progression via the downregulation of the Erk-RPS6K-TMEM139 and PI3K-Akt-mTOR pathways in oxaliplatin-resistant AsPC-1 cells. Furthermore, KV showed an antiproliferative effect in PDPCOs, and the combination of OXA and KV inhibited PDPCO growth more effectively than either drug alone.

Temozolomide (TMZ) has been used as standard-of-care for glioblastoma multiforme (GBM), but the resistance to TMZ develops quickly and frequently. Thus, more studies are needed to elucidate the resistance mechanisms. In the current study, we investigated the relationship among the three important phenotypes, namely TMZ-resistance, cell shape and lipid metabolism, in GBM cells. We first observed the distinct difference in cell shapes between TMZ-sensitive (U87) and resistant (U87R) GBM cells. We then conducted NMR-based lipid metabolomics, which revealed a significant increase in cholesterol and fatty acid synthesis as well as lower lipid unsaturation in U87R cells. Consistent with the lipid changes, U87R cells exhibited significantly lower membrane fluidity. The transcriptomic analysis demonstrated that lipid synthesis pathways through SREBP were upregulated in U87R cells, which was confirmed at the protein level. Fatostatin, an SREBP inhibitor, and other lipid pathway inhibitors (C75, TOFA) exhibited similar or more potent inhibition on U87R cells compared to sensitive U87 cells. The lower lipid unsaturation ratio, membrane fluidity and higher fatostatin sensitivity were all recapitulated in patient-derived TMZ-resistant primary cells. The observed ternary relationship among cell shape, lipid composition, and TMZ-resistance may be applicable to other drug-resistance cases. SREBP and fatostatin are suggested as a promising target-therapeutic agent pair for drug-resistant glioblastoma., (© 2022. The Author(s), under exclusive licence to Shanghai Institute of Materia Medica, Chinese Academy of Sciences and Chinese Pharmacological Society.)

We report a proteogenomic analysis of pancreatic ductal adenocarcinoma (PDAC). Mutation-phosphorylation correlations identified signaling pathways associated with somatic mutations in significantly mutated genes. Messenger RNA-protein abundance correlations revealed potential prognostic biomarkers correlated with patient survival. Integrated clustering of mRNA, protein and phosphorylation data identified six PDAC subtypes. Cellular pathways represented by mRNA and protein signatures, defining the subtypes and compositions of cell types in the subtypes, characterized them as classical progenitor (TS1), squamous (TS2-4), immunogenic progenitor (IS1) and exocrine-like (IS2) subtypes. Compared with the mRNA data, protein and phosphorylation data further classified the squamous subtypes into activated stroma-enriched (TS2), invasive (TS3) and invasive-proliferative (TS4) squamous subtypes. Orthotopic mouse PDAC models revealed a higher number of pro-tumorigenic immune cells in TS4, inhibiting T cell proliferation. Our proteogenomic analysis provides significantly mutated genes/biomarkers, cellular pathways and cell types as potential therapeutic targets to improve stratification of patients with PDAC., (© 2022. The Author(s), under exclusive licence to Springer Nature America, Inc.)

The acquisition of drug resistance is a major hurdle for effective cancer treatment. Although several efforts have been made to overcome drug resistance, the underlying mechanisms have not been fully elucidated. This study investigated the role of long non-coding RNA (lncRNA) growth arrest-specific 5 (GAS5) in drug resistance. GAS5 was found to be downregulated in colon cancer cell lines that are resistant to 5-fluorouracil (5-FU). Downregulation of GAS5 decreased the viability of HCT116 cells and the level of the pro-apoptotic BAX protein, while GAS5 overexpression promoted cell death in response to 5-FU. The interaction between GAS5 and BAX mRNA was investigated using MS2-tagged RNA affinity purification (MS2-trap) followed by RT-qPCR, and the results showed that GAS5 bound to the 3'-untranslated region of BAX mRNA and enhanced its expression by interfering with the inhibitory effect of microRNA-128-3p, a negative regulator of BAX. In addition, ectopic expression of GAS5 increased the sensitivity of resistant cells in response to anti-cancer drugs. These results suggest that GAS5 promoted cell death by interfering with miR-128-3p-mediated BAX downregulation. Therefore, GAS5 overexpression in chemo-resistant cancer cells may be a potential strategy to improve the anti-cancer efficacy of drugs.

Background: A significant proportion of colorectal cancer (CRC) patients suffer from early recurrence and progression after surgical treatment. Although the gut microbiota is considered as a key player in the initiation and progression of CRC, most prospective studies have been focused on a particular pathobionts such as Fusobacterium nucleatum. Here, we aimed to identify novel prognostic bacteria for CRC by examining the preoperative gut microbiota through 16S ribosomal RNA gene sequencing., Results: We collected stool samples from 333 patients with primary CRC within 2 weeks before surgery and followed up the patients for a median of 27.6 months for progression and 43.6 months for survival. The sequence and prognosis data were assessed using the log-rank test and multivariate Cox proportional hazard analysis. The gut microbiota was associated with the clinical outcomes of CRC patients (P progress = 0.011, P decease = 0.007). In particular, the high abundance of Prevotella, a representative genus of human enterotypes, indicated lower risks of CRC progression (P = 0.026) and decease (P = 0.0056), while the occurrence of Alistipes assigned to Bacteroides sp., Pyramidobacter piscolens, Dialister invisus, and Fusobacterium nucleatum indicated a high risk of progression. A microbiota-derived hazard score considering the five prognostic bacteria accurately predicted CRC progression in 1000 random subsamples; it outperformed widely accepted clinical biomarkers such as carcinoembryonic antigen and lymphatic invasion, after adjustment for the clinicopathological stage (adjusted HR 2.07 [95% CI, 1.61-2.64], P = 7.8e-9, C-index = 0.78). PICRUSt2 suggested that microbial pathways pertaining to thiamine salvage and L-histidine degradation underlie the different prognoses., Conclusions: The enterotypical genus Prevotella was demonstrated to be useful in improving CRC prognosis, and combined with the four pathobionts, our hazard score based on the gut microbiota should provide an important asset in predicting medical outcomes for CRC patients. Video Abstract., (© 2022. The Author(s).)

Background: The activation of the telomere maintenance mechanism (TMM) is one of the critical drivers of cancer cell immortality. In gliomas, TERT expression and TERT promoter mutation are considered to reliably indicate telomerase activation, while ATRX mutation and/or loss indicates an alternative lengthening of telomeres (ALT). However, these relationships have not been extensively validated in tumor tissues., Methods: Telomerase repeated amplification protocol (TRAP) and C-circle assays were used to profile and characterize the TMM cross-sectionally (n = 412) and temporally (n = 133) across glioma samples. WES, RNA-seq, and NanoString analyses were performed to identify and validate the genetic characteristics of the TMM groups., Results: We show through the direct measurement of telomerase activity and ALT in a large set of glioma samples that the TMM in glioma cannot be defined solely by the combination of telomerase activity and ALT, regardless of TERT expression, TERT promoter mutation, and ATRX loss. Moreover, we observed that a considerable proportion of gliomas lacked both telomerase activity and ALT. This telomerase activation-negative and ALT negative group exhibited evidence of slow growth potential. By analyzing a set of longitudinal samples from a separate cohort of glioma patients, we discovered that the TMM is not fixed and can change with glioma progression., Conclusions: This study suggests that the TMM is dynamic and reflects the plasticity and oncogenicity of tumor cells. Direct measurement of telomerase enzyme activity and evidence of ALT should be considered when defining TMM. An accurate understanding of the TMM in glioma is expected to provide important information for establishing cancer management strategies., (© 2022. The Author(s).)

Organoid culture technique has been taking center stage as a next-generation ex-vivo model due to advancement of stem cell research techniques. The importance of the laboratory-based ex vivo model has increasingly been recognized for recapitulating histological, and physioglocal conditions of in vivo microenviorment. Accordingly, the use of this technique has also broadened the understanding of intratumoral heterogeneity which is closely associated with varied drug responses observed in patients. Likewise, studies on heterogeneity within a single tumor tissue have drawn much attention. Here, we isolated 15 single clones from 4 tumor organoid lines from 1 patient at a primary passage from one patient. Each organoid line showed variable alterations in both genotype and phenotype. Furthermore, our methodological approach on drug test employing a high-throughput screening system enabled us to pinpoint the optimal time frame for anti-cancer drugs within a single tumor. We propose that our method can effectively reveal the heterogeneity of time-point in drug response, and the most optimal therapeutic strategies for individual patient., (Copyright © 2022 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Multifocal colorectal cancer (CRC) comprises both clonally independent primary tumors caused by inherited predisposition and clonally related tumors mainly due to intraluminal spreading along an intact basement membrane. The distinction between these multifocal CRCs is essential because therapeutic strategies vary according to the clonal association of multiple tumor masses. Here, we report one unique case of synchronous intestinal cancer (SIC) with tumors occurring along the entire bowel tract, including the small intestine. We established six patient-derived organoids (PDOs), and patient-derived cell lines (PDCs) from each site of the SIC, which were subjected to extensive genomic, transcriptomic, and epigenomic sequencing. We also estimated the drug responses of each multifocal SIC to 25 clinically relevant therapeutic compounds to validate how the clinically actionable alternations between SICs were associated with drug sensitivity. Our data demonstrated distinct clonal associations across different organs, which were consistently supported by multi-omics analysis, as well as the accordant responses to various therapeutic compounds. Our results indicated the imminent drawback of a single tumor-based diagnosis of multifocal CRC and suggested the necessity of an in-depth molecular analysis of all tumor regions to avoid unexpected resistance to the currently available targeted therapies., (© 2022. The Author(s).)

Background/aims: Three-dimensional cultures of human pancreatic cancer tissue also known as "organoids" have largely been developed from surgical specimens. Given that most patients present with locally advanced and/or metastatic disease, such organoids are not representative of the majority of patients. Therefore, we used endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) to collect pancreatic cancer tissues from patients with advanced pancreatic cancer to create organoids, and evaluated their utility in pancreatic cancer research., Methods: Single-pass EUS-FNA samplings were employed to obtain the tissue for organoid generation. After establishment of the organoid, we compared the core biopsy tissues with organoids using hematoxylin and eosin staining, and performed whole exome sequencing (WES) to detect mutational variants. Furthermore, we compared patient outcome with the organoid drug response to determine the potential utility of the clinical application of such organoid-based assays., Results: Organoids were successfully generated in 14 of 20 tumors (70%) and were able to be passaged greater than 5 times in 12 of 20 tumors (60%). Among them, we selected eight pairs of organoid and core biopsy tissues for detailed analyses. They showed similar patterns in hematoxylin and eosin staining. WES revealed mutations in KRAS, TP53, CDKN2A, SMAD4, BRCA1 , and BRCA2 which were 93% homologous, and the mean nonreference discordance rate was 5.47%. We observed moderate drug response correlations between the organoids and clinical outcomes in patients who underwent FOLFIRINOX chemotherapy., Conclusions: The established organoids from EUS-FNA core biopsies can be used for a suitable model system for pancreatic cancer research.

The poor prognosis of ovarian cancer patients mainly results from a lack of early diagnosis approaches and a high rate of relapse. Only a very modest improvement has been made in ovarian cancer patient survival with traditional treatments. More targeted therapies precisely matching each patient are strongly needed. The aberrant activation of Wnt/β-catenin signaling pathway plays a fundamental role in cancer development and progression in various types of cancer including ovarian cancer. Recent insight into this pathway has revealed the potential of targeting Wnt/β-catenin in ovarian cancer treatment. This study aims to investigate the effect of CWP232291, a small molecular Wnt/β-catenin inhibitor on ovarian cancer progression. Various in vitro , in vivo and ex vivo models are established for CWP232291 testing. Results show that CWP232291 could significantly attenuate ovarian cancer growth through inhibition of β-catenin. Noticeably, CWP232291 could also s suppress the growth of cisplatin-resistant cell lines and ovarian cancer patient-derived organoids. Overall, this study has firstly demonstrated the anti-tumor effect of CWP232291 in ovarian cancer and proposed Wnt/β-catenin pathway inhibition as a novel therapeutic strategy against ovarian cancer., Competing Interests: Authors AY and C-LJ were employed by JW Pharmaceutical Corporation. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The authors declare that this study received funding from JW Pharmaceutical Corporation. The funder had the following involvement with the study: design and performance of the mouse experiment., (Copyright © 2022 Wang, Cho, Yoo, Jung, Kim, Kim, Lee, Jo, Han, Song, Lee, Kim, Lee, Ku, Lee and Song.)

Malignant pleural effusion (MPE) is an independent determinant of poor prognostic factor of non-small cell lung cancer (NSCLC). The course of anchorage independent growth within the pleural cavity likely reforms the innate molecular characteristics of malignant cells, which largely accounts for resistance to chemotherapy and poor prognosis after the surgical resection. Nevertheless, the genetic and transcriptomic features with respect to various drug responses of MPE-complicated NSCLC remain poorly understood. To obtain a clearer overview of the MPE-complicated NSCLC, we established 28 MPE-derived lung cancer cell lines which were subjected to genomic, transcriptomic and pharmacological analysis. Our results demonstrated MPE-derived NSCLC cell lines recapitulated representative driver mutations generally found in the primary NSCLC. It also exhibited the presence of distinct translational subtypes in accordance with the mutational profiles. The drug responses of several targeted chemotherapies accords with both genomic and transcriptomic characteristics of MPE-derived NSCLC cell lines. Our data also suggest that the impending drawback of mutation-based clinical diagnosis in evaluating MPE-complicated NSCLS patient responses. As a potential solution, our work showed the importance of comprehending transcriptomic characteristics in order to defy potential drug resistance caused by MPE., (© 2022. The Author(s).)

Glioblastomas (GBM) exhibit intratumoral heterogeneity of various oncogenic evolutional processes. We have successfully isolated and established two distinct cancer cell lines with different morphological and biological characteristics that were derived from the same tissue sample of a GBM. When we compared their genomic and transcriptomic characteristics, each cell line harbored distinct mutation clusters while sharing core driver mutations. Transcriptomic analysis revealed that one cell line was undergoing a mesenchymal transition process, unlike the other cell line. Furthermore, we could identify four tumor samples containing our cell line-like clusters from the publicly available single-cell RNA-seq data, and in a set of paired longitudinal GBM samples, we could confirm three pairs where the recurrent sample was enriched in the genes specific to our cell line undergoing mesenchymal transition. The present study provides direct evidence and a valuable source for investigating the ongoing process of subcellular mesenchymal transition in GBM, which has prognostic and therapeutic implications., (© 2022. The Author(s).)

Intratumor heterogeneity (ITH) stands as one of the main difficulties in the treatment of colorectal cancer (CRC) as it causes the development of resistant clones and leads to heterogeneous drug responses. Here, 12 sets of patient-derived organoids (PDOs) and cell lines (PDCs) isolated from multiple regions of single tumors from 12 patients, capturing ITH by multiregion sampling of individual tumors, are presented. Whole-exome sequencing and RNA sequencing of the 12 sets are performed. The PDOs and PDCs of the 12 sets are also analyzed with a clinically relevant 24-compound library to assess their drug responses. The results reveal unexpectedly widespread subregional heterogeneity among PDOs and PDCs isolated from a single tumor, which is manifested by genetic and transcriptional heterogeneity and strong variance in drug responses, while each PDO still recapitulates the major histologic, genomic, and transcriptomic characteristics of the primary tumor. The data suggest an imminent drawback of single biopsy-originated PDO-based clinical diagnosis in evaluating CRC patient responses. Instead, the results indicate the importance of targeting common somatic driver mutations positioned in the trunk of all tumor subregional clones in parallel with a comprehensive understanding of the molecular ITH of each tumor., (© 2021 The Authors. Advanced Science published by Wiley-VCH GmbH.)

Despite the essential roles of natural killer (NK) cells in cancer treatment, the physical barrier and biological cues of the tumor microenvironment (TME) may induce NK cell dysfunction, causing their poor infiltration into tumors. The currently available two-dimensional (2D) cancer-NK co-culture systems hardly represent the characteristics of TME and are not suitable for tracking the infiltration of immune cells and assessing the efficacy of immunotherapy. This study aims to monitor NK-mediated cancer cell killing using a polymer thin film-based, 3D assay platform that contains highly tumorigenic cancer spheroids. A poly(cyclohexyl methacrylate) (pCHMA)-coated surface enables the generation of tumorigenic spheroids from pancreatic cancer patient-derived cancer cells, showing considerable amounts of extracellular matrix (ECM) proteins and cancer stem cell (CSC)-like characteristics. The 3D spheroid-based assay platform allows rapid discovery of a therapeutic agent for synergistic NK-mediated cytotoxicity through imaging-based high-content screening. In detail, the small molecule C19, known as a multi-epithelial-mesenchymal transition pathway inhibitor, is shown to enhance NK activation and infiltration via modulation of the ECM, resulting in synergistic cytotoxicity against cancer spheroids. This 3D biomimetic co-culture assay platform provides promising applications for predicting patient-specific responses to immunotherapy through advanced therapeutic combinations involving a chemical drug and immune cells.

Purpose: Epidermal growth factor receptor kinase domain duplication (EGFR-KDD) is a rare and poorly understood oncogenic mutation in non-small cell lung cancer (NSCLC). We aimed to investigate the acquired resistance mechanism of EGFR-KDD against EGFR-TKIs., Materials and Methods: We identified EGFR-KDD in tumor tissue obtained from a patient with stage IV lung adenocarcinoma and established the patient-derived cell line SNU-4784. We also established several EGFR-KDD Ba/F3 cell lines: EGFR-KDD wild type (EGFR-KDDWT), EGFR-KDD domain 1 T790M (EGFR-KDDD1T), EGFR-KDD domain 2 T790M (EGFR-KDDD2T), and EGFR-KDD both domain T790M (EGFR-KDDBDT). We treated the cells with EGFR tyrosine kinase inhibitors (TKIs) and performed cell viability assays, immunoblot assays, and ENU (N-ethyl-N-nitrosourea) mutagenesis screening., Results: In cell viability assays, SNU-4784 cells and EGFR-KDDWT Ba/F3 cells were sensitive to 2nd generation and 3rd generation EGFR TKIs. In contrast, the T790M-positive EGFR-KDD Ba/F3 cell lines (EGFR-KDDT790M) were only sensitive to 3rd generation EGFR TKIs. In ENU mutagenesis screening, we identified the C797S mutation in kinase domain 2 of EGFR-KDDBDT Ba/F3 cells. Based on this finding, we established an EGFR-KDD domain 1 T790M/domain 2 cis-T790M+C797S (EGFR-KDDT/T+C) Ba/F3 model, which was resistant to EGFR TKIs and anti-EGFR monoclonal antibody combined with EGFR TKIs., Conclusion: Our study reveals that the T790M mutation in EGFR-KDD confers resistance to 1st and 2nd generation EGFR TKIs, but is sensitive to 3rd generation EGFR TKIs. In addition, we identified that the C797S mutation in kinase domain 2 of EGFR-KDDT790M mediates a resistance mechanism against 3rd generation EGFR TKIs.

Recently, several panels using two representative targeting methods have been developed but they do not reflect racial specificity, especially for Asians. We have developed and analytically validated the Korean Pan-cancer Companion Diagnostic (CDX) Panel to apply targeted anticancer drugs to Korean patients based on the molecular characteristics of tumors using tumor samples without matched patient normal samples. The panel included 31 genes with reported single nucleotide variants, 9 genes with reported copy number variations, and 15 genes with predictive responses to targeted drugs under clinical testing, enabling the panel to be analyzed for the targets of 30 targeted anticancer drugs. It is cost-effective and optimized for cancer type-specific therapy in Korean cancer patients across solid cancer types while minimizing the limitations of existing approaches. In addition, the optimized filtering protocol for somatic variants from tumor-only samples enables researchers to use this panel without matched normal samples. To verify the panel, 241 frozen tumor tissues and 71 formalin-fixed paraffin-embedded (FFPE) samples from several institutes were registered. This gene screening method is expected to reduce test turnaround time and cost, making it a balanced approach to investigate solid cancer-related gene regions.

Abnormalities of the tumor vasculature result in insufficient blood supply and development of a tumor microenvironment that is characterized by low glucose concentrations, low extracellular pH, and low oxygen tensions. We previously reported that glucose-deprived conditions induce metabolic stress and promote tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced cytotoxicity. In this study, we examined whether the metabolic stress-associated endoplasmic reticulum (ER) stress response pathway plays a pivotal role in the enhancement of TRAIL cytotoxicity. We observed no significant cytotoxicity when human colorectal cancer SW48 cells were treated with various doses of TRAIL (2-100 ng/ml) for 4 h or glucose (0-25 mM) for 24 h. However, a combination of TRAIL and low glucose-induced dose-dependent apoptosis through activation of caspases (-8, -9, and -3). Studies with activating transcription factor 4 (ATF4), C/EBP-homologous protein (CHOP), p53 upregulated modulator of apoptosis (PUMA), or death receptor 5 (DR5)-deficient mouse embryonic fibroblasts or HCT116 cells suggest that the ATF4-CHOP-PUMA axis and the ATF4-CHOP-DR5 axis are involved in the combined treatment-induced apoptosis. Moreover, the combined treatment-induced apoptosis was completely suppressed in BH3 interacting-domain death agonist (Bid)- or Bcl-2-associated X protein (Bax)-deficient HCT116 cells, but not Bak-deficient HCT116 cells. Interestingly, the combined treatment-induced Bax oligomerization was suppressed in PUMA-deficient HCT116 cells. These results suggest that glucose deprivation enhances TRAIL-induced apoptosis by integrating the ATF4-CHOP-PUMA axis and the ATF4-CHOP-DR5 axis, consequently amplifying the Bid-Bax-associated mitochondria-dependent pathway., (© 2021 Wiley Periodicals LLC.)

In epithelial ovarian cancer (EOC), carboplatin/cisplatin-induced chemoresistance is a major hurdle to successful treatment. Aerobic glycolysis is a common characteristic of cancer. However, the role of glycolytic metabolism in chemoresistance and its impact on clinical outcomes in EOC are not clear. Here, we show a functional interaction between the key glycolytic enzyme hexokinase II (HKII) and activated P-p53 (Ser15) in the regulation of bioenergetics and chemosensitivity. Using translational approaches with proximity ligation assessment in cancer cells and human EOC tumor sections, we showed that nuclear HKII-P-p53 (Ser15) interaction is increased after chemotherapy, and functions as a determinant of chemoresponsiveness as a prognostic biomarker. We also demonstrated that p53 is required for the intracellular nuclear HKII trafficking in the control of glycolysis in EOC, associated with chemosensitivity. Mechanistically, cisplatin-induced P-p53 (Ser15) recruits HKII and apoptosis-inducing factor (AIF) in chemosensitive EOC cells, enabling their translocation from the mitochondria to the nucleus, eliciting AIF-induced apoptosis. Conversely, in p53-defective chemoresistant EOC cells, HKII and AIF are strongly bound in the mitochondria and, therefore, apoptosis is suppressed. Collectively, our findings implicate nuclear HKII-P-p53(Ser15) interaction in chemosensitivity and could provide an effective clinical strategy as a promising biomarker during platinum-based therapy.

Purpose: Long-term oncologic differences in outcome between groups of patients with Lynch syndrome (LS) colorectal cancer (CRC) and sporadic CRC with microsatellite instability-high (MSI-H) are the focus of investigation in the current study., Methods: Patients registered in the Korean Hereditary Tumor Registry and 2 tertiary referral hospitals treated for stage I-III CRC between 2005 and 2015 were retrospectively analyzed. Detection for both groups was performed using pedigree, microsatellite instability, and mismatch repair (MMR) gene testing. Multivariate analyses for overall survival (OS) and disease-free survival (DFS) were conducted., Results: Cases of LS (n = 77) and sporadic CRC with MSI-H (n = 96) were identified. LS CRC patients were younger in age and displayed tumor sidedness, typically involving left-sided colon and rectum, compared to patients with sporadic CRC with MSI-H. OS and DFS were lower for LS CRC relative to CRC with MSI-H (OS, 72.7% vs. 93.8%, P = 0.001; DFS, 71.4% vs. 88.5%, P = 0.001). In multivariate analyses, tumor sidedness, stage, and chemotherapy were independent factors for OS and DFS. LS CRC was a prognostic factor for poorer OS (hazard ratio, 2.740; 95% confidence interval, 1.003-7.487; P = 0.049), but not DFS., Conclusion: Our findings indicate that LS CRC is associated with poorer outcomes compared to sporadic CRC with MSI-H, presenting distinct clinical features. In view of the current lack of knowledge on genetic and molecular mechanisms, appropriate management taking into consideration the difficulty of identification of CRC with hypermutable tumors harboring heterogeneity is essential., Competing Interests: Conflicts of Interest: No potential conflict of interest relevant to this article was reported., (Copyright © 2021, the Korean Surgical Society.)

A metastasis of colorectal cancer is difficult to diagnose, and has a poor prognosis. Therefore, we tried to elucidate the possibility of a diagnostic and prognostic marker. Exosomal miR-193a and let-7g were sorted by miRNA microarray. The expression of miR-193a in the PTM group was lower than that of the primary CRC group, and the expression of let-7g was higher than that of the primary CRC. MMP16 and CDKN1A expression was confirmed respectively for target genes of two miRNAs. When the mimics of these miRNAs were treated with cell lines, both MMP16 and CDKN1A decreased intracellular expression. Cell invasiveness and proliferation were decreased by miR-193a and increased by let-7g. The differences in expression of exosomal miR-193a and let-7g extracted from the plasma of patients were classified as cancer progression indicators. Furthermore, the survival rate decreased in the group with low miR-193a expression and high let-7g expression. Our study confirmed the possibility of using this as a diagnostic and prognostic marker for colorectal cancer by measuring the expression levels of exosomal miR-193a and let-7g in blood., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

The emergence of T-cell engineering with chimeric antigen receptors (CARs) has led to attractive therapeutics; however, autologous CAR-T cells are associated with poor clinical outcomes in solid tumors because of low safety and efficacy. Therefore, the aim of our study was to develop a CAR therapy with enhanced cytotoxicity against solid cancer using allogeneic NK cells. In this study, we engineered "off-the-shelf" NK cells to redirect them towards pancreatic ductal adenocarcinoma (PDAC) by improving their target-specific cytotoxic potential. By integrated bioinformatic and clinicopathological analyses, folate receptor alpha (FRα) and death receptor 4 (DR4) were significantly highly expressed in patient-derived tumor cells. The combined expression of FRα and DR4/5 was associated with inferior clinical outcomes, therefore indicating their use as potential targets for biomolecular treatment. Thus, FRα and DR4 expression pattern can be a strong prognostic factor as promising therapeutic targets for the treatment of PDAC. For effective PDAC treatment, allogeneic CAR-NK cells were reprogrammed to carry an apoptosis-inducing ligand and to redirect them towards FRα and initiate DR4/5-mediated cancer-selective cell death in FRα- and DR4/5-positive tumors. As a result, the redirected cytotoxic ligand-loaded NK cells led to a significantly enhanced tumor-selective apoptosis. Accordingly, use of allogeneic CAR-NK cells that respond to FRα and DR4/5 double-positive cancers might improve clinical outcomes based on personal genome profiles. Thus, therapeutic modalities based on allogeneic NK cells can potentially be used to treat large numbers of patients with optimally selective cytotoxicity., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Gastrointestinal stromal tumor (GIST) is the most common sarcoma in the gastrointestinal (GI) tract. Approximately 85% of the GIST is associated with a c-KIT mutation. A few GISTs show mutations in the gene encoding platelet-derived growth factor receptor alpha (PDGFR α or PDGFRA) without c-KIT gene mutation. GIST without c-KIT or PDGFRA mutations, which called wild type GIST, is about 5-10% of the total GIST. Fusion genes were also reported as one of the factors associated with carcinogenesis and drug resistance. With five cell lines derived from imatinib-resistant patients, novel fusion genes were identified from RNA sequencing and both physiological role and therapeutic potential were elucidated. Next-generation sequencing (NGS) analysis and lentiviral transduction were used to effect of fusion gene on GISTs. All the GIST cell lines carried c-KIT-positivity. Three different fusion gene analysis methods were used to find candidate fusion genes, including EIF3K-ACTN4, SYNCRIP-SNX14 and EXOC2-AK7. A novel interchromosomal fusion gene of the candidates, especially EXOC2-AK7, was confirmed in both tissue and cell line. The transduction of fusion gene increased the proliferation compared with the control group. Additionally, the fusion gene increased wound coverage capability. The fusion gene-transduced cell lines were more sensitive than the control group in the treatment of imatinib. In conclusion, five different imatinib-resistant GIST cell lines including the EXOC2-AK7 fusion gene derived from GIST-R5 represent important research tools for the investigation of cancer cell mechanisms underlying drug resistance and genetic variation. Furthermore, our study may facilitate pre-clinical studies of new therapeutic strategies., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.)

The clinical phenotype linked with mutations in ABCB1, encoding P-glycoprotein, has never been reported. Here, we describe twin sisters with biallelic mutations in ABCB1 who showed recurrent reversible encephalopathy accompanied by acute febrile or afebrile illness. Whole-exome sequencing was performed on one of the twin and her healthy parents, and revealed compound heterozygous loss-of-function variants in ABCB1. The patient brains displayed substantial loss of xenobiotic clearance ability, as demonstrated by [ 11 C]verapamil positron emission tomography (PET) study, linking this phenotype with ABCB1 function. The endogenous cytokine clearance from the brain was also decreased in LPS-treated ABCB1 knockout mice compared to controls. The results provide insights into the physiological requirement of ABCB1 in maintaining homeostasis of various compounds for normal brain function., (© 2020 The Authors. Annals of Clinical and Translational Neurology published by Wiley Periodicals LLC on behalf of American Neurological Association.)

Due to its regulation of CDK1/2 phosphorylation, WEE1 plays essentially roles in the regulations of G2/M checkpoint and DNA damage response (DDR). WEE1 inhibition can increase genomic instability by inducing replication stress and G2/M checkpoint inactivation, which result in increased cellular sensitivity to DNA damaging agents. We considered an increase in genomic instability induced by WEE1 inhibition might be used to augment the effects of drugs targeting DNA repair protein. Typically, PARP inhibitors are effective in germline BRCA 1/2 mutated breast and ovarian cancer, but their applicabilities in triple-negative breast cancer (TNBC) are limited. This study was conducted to investigate the anti-tumor effects of the WEE1 inhibitor, AZD1775, and the mechanism responsible for its potentiation of sensitivity to olaparib (a PARP inhibitor) via the modulation of DDR in TNBC cells. Our results suggest that AZD1775 could be used to broaden the application range of olaparib in TNBC and provide a rationale for a clinical trial of combined olaparib and AZD1775 therapy.

Two of the most critical factors for the survival of glioblastoma (GBM) patients are precision diagnosis and the tracking of treatment progress. At the moment, various sophisticated and specific diagnostic procedures are being used, but there are relatively few simple diagnosis methods. This work introduces a sensing probe based on a turn-on type fluorescence response that can measure the cysteine (Cys) level, which is recognized as a new biomarker of GBM, in human-derived cells and within on-site human clinical biopsy samples. The Cys-initiated chemical reactions of the probe cause a significant fluorescence response with high selectivity, high sensitivity, a fast response time, and a two-photon excitable excitation pathway, which allows the imaging of GBM in both mouse models and human tissue samples. The probe can distinguish the GBM cells and disease sites in clinical samples from individual patients. Besides, the probe has no short or long-term toxicity and immune response. The present findings hold promise for application of the probe to a relatively simple and straightforward following of GBM at clinical sites., (This journal is © The Royal Society of Chemistry 2020.)

Cbl‑associated protein (CAP) is encoded by the sorbin and SH3 domain‑containing 1 (SORBS1) gene. CAP has been reported to be associated with the actin cytoskeleton, receptor tyrosine kinase signaling and cell adhesion through interactions with various proteins. It may be hypothesized that SORBS1 has numerous unknown functions, which may include providing a favorable condition for metastasis. Although CAP has been demonstrated to possess a number of functions, the role of this protein has only been reported in metabolic signaling pathways and its function in cancer remains to be elucidated. In the present study, SORBS1 expression was detected in colorectal cancer cell lines divided into the primary group and the metastatic group by reverse transcription‑quantitative PCR and western blot analysis. In addition, SORBS1 expression was manipulated by vector transfection and lentivirus transduction. The metastatic role of SORBS1, as determined by assessing its effects on cell proliferation and migration, was determined by colony formation assay, cell cycle analysis and Boyden chamber assay. To elucidate the SORBS1‑binding protein, immunoprecipitation was performed. Co‑localization of SORBS1 and AHNAK nucleoprotein (AHNAK) was identified by confocal microscopy. Notably, the protein expression levels of CAP were higher in SNU‑769A and SW480 cells than in SNU‑769B and SW620 cells. In addition, the number of colonies in the SORBS1‑overexpressing group was significantly increased compared with that of the control group, as determined using the colony formation assay; the SORBS1 overexpression group formed >8‑fold more colonies than the control group. The proliferative ability of the SORBS1 overexpression group was also significantly increased compared with the control group over the entire incubation period. Cell migration assays revealed that the number of migrated SORBS1‑knockdown cells was reduced compared with the control in both HCT‑116 and SNU‑C4 cell lines; migration area was decreased to 31 and 26% in HCT‑116 and SNU‑C4 cell lines, respectively. Consequently, it was confirmed that SORBS1 could form a complex with AHNAK, which functions as a tumor suppressor through inhibition of phosphorylated‑ERK and Rho‑associated coiled‑coil containing protein kinase 1. In conclusion, SORBS1 may serve a crucial role in cancer growth and migration via inhibition of AHNAK expression.

Colorectal cancer (CRC) represents the third most frequently diagnosed malignancy worldwide and is the second most common cause of tumor-associated mortalities in Korea. Due to the disease's aggressive behavior, the 5-year survival rate for CRC patients remains unpromising. Well-characterized cell lines have been used as a biological model for studying the biology of cancer and developing novel therapeutics. To assist in vitro studies, 18 CRC cell lines (SNU-1566, SNU-1983, SNU-2172, SNU-2297, SNU-2303, SNU-2353B, SNU-2359, SNU-2373B, SNU-2407, SNU-2423, SNU-2431, SNU-2465, SNU-2493, SNU-2536C, SNU-2621B, SNU-NCC-61, SNU-NCC-376, and SNU-NCC-377) derived from Korean patients were established and characterized in the present study. General characteristics of each cell line including doubling time, in vitro morphology, mutational profiles, and protein expressions of CRC-related genes were described. Whole exome sequencing was performed on each cell line to configure mutational profiles. Single nucleotide variation, frame shift, in-frame deletions and insertions, start codon deletion, and splice stop codon mutation of various genes were found and classified based on their pathogenicity reports. In addition, cell viability was assayed to measure their sensitivities to 24 anti-cancer drugs including anti-metabolites, kinase inhibitors, histone deacetylase inhibitors, alkylating inhibitors, and topoisomerase inhibitors, all widely used for various cancers. On testing, five CRC cell lines showed MSI, of which MLH1 or MSH6 gene was mutated. These newly established CRC cell lines can be used to investigate biological characteristics of CRC, particularly for investigating gene alterations associated with CRC.

Objective: The incidence of pancreatic adenocarcinoma (PA) approximates its prevalence, as the malignancy is almost consistently fatal within a year. Although the currently available adjuvant therapy seems to provide survival benefit, it is only moderate, and the standard regimen has not yet been established. Therefore, more biological resources to investigate the PA are needed., Methods: Here, we established and characterized 10 human pancreatic cancer cell lines derived from primary tumor mass. Whole exome sequencing technique was used to identify driver mutations and aberrant pathways in each cell line., Results: Five anticancer drugs were treated to find half maximal effective concentration (EC50), and the response was analyzed in reference to mutational status. Frame shift mutations in ARID1A gene and HER2 amplification were mutually related to better response to the anticancer drugs. In contrast, frame shift mutation in MSH6 gene was associated with resistance to anticancer drugs., Conclusions: In summary, we established 10 pancreatic cancer cell lines and integrated various molecular aberrations and features of pancreatic cancer cells. Our biological resources are expected to contribute to facilitating research on PA.

Introduction: NSCLC with EGFR exon 20 insertion mutations is the third most common type of EGFR-mutant NSCLC and is resistant to EGFR tyrosine kinase inhibitors (TKIs). This study was conducted to evaluate the efficacies of first- to third-generation EGFR TKIs against NSCLC cells harboring EGFR exon 20 insertion mutations., Methods: We developed seven EGFR exon 20 insertion-mutant Ba/F3 models and one patient-derived NSCLC (SNU-3173) of subtypes A763insFQEA, V769insASV, D770insSVD, D770insNPG, P772insPR, H773insH, H773insNPH, and H773insAH. Cell viability assays, immunoblotting, and N-ethyl-N-nitrosourea mutagenesis screenings were performed. EGFR exon 20 insertion-mutant structures and couplings with osimertinib, a third-generation EGFR TKI, were modeled and compared., Results: EGFR exon 20 insertion-mutant NSCLC cells, excluding EGFR A763insFQEA, were resistant to first-generation EGFR TKIs (concentration that inhibits 50% [IC 50 ], 1.1 ± 0.067 to 5.4 ± 0.115 μM). Mutants were sensitive to second-generation EGFR TKIs (IC 50 , 0.02 ± 0.0002 to 161.8 ± 18.7nM), except EGFR H773insH (IC 50 , 46.3 ± 8.0 to 352.5 ± 22.7nM). The IC 50 ratios for mutant to wild-type cells were higher than those for third-generation EGFR TKIs. Third-generation EGFR TKI osimertinib was highly potent against EGFR exon 20 insertion-mutant cells (IC 50 , 14.7-62.7 nM), including EGFR H773insH, and spared wild-type EGFR cells. N-ethyl-N-nitrosourea mutagenesis screening of EGFR exon 20 insertion-mutant Ba/F3 cells showed various second sites for EGFR mutations, mostly at exons 20 and 21, including E762K, P794S, and G796D. In addition, osimertinib-resistant cells were established by stepwise exposure to osimertinib and harbored EGFR E762K mutation., Conclusions: Osimertinib is active against EGFR exon 20 insertion-mutant NSCLC and flexibly binds within drug-binding pockets in preclinical models., (Copyright © 2019 International Association for the Study of Lung Cancer. Published by Elsevier Inc. All rights reserved.)

Colorectal cancer (CRC) is the third most common cancer diagnosed worldwide and represents the second most common cause of all cancer-related deaths in Korea. Although epidemiological data indicate a sharp increase in the incidence of CRC among individuals older than 50 years, more than 10% of CRCs occur before reaching 50. These are known as early-onset CRCs (EOCRCs) and are likely to be suggestive of hereditary predisposition. However, known familial CRC syndromes account for only 20% of genetic aberrations of EOCRC, and the remaining 80% are still in question. Therefore, we aimed to establish reproducible biological resources and contribute to expand the mutation database of EOCRC. Four cell lines derived from the original tumor mass of CRC patients diagnosed under age 30 years were established, and next-generation sequencing technique was used to identify the genetic features of EOCRC. We have identified one novel fusion gene, FAM174A-WWC1, and analyzed its functional role. The induction of FAM174A-WWC1 to normal fibroblast caused alternations in cellular morphology as well as intercellular expression of E-cadherin and N-cadherin. Moreover, WWC1 carrying the fused FAM174A domain not only abrogated the membrane expression of YAP1 but also significantly increased the levels of nucleic YAP1. As a result, the FAM174A-WWC1 expression increased the oncogenic capacity and invasiveness of normal fibroblasts, which suggests its role as a potential driver mutation of EOCRC., (Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.)

Exposure to persistent organic pollutants (POPs) during pregnancy is associated with a disruption in thyroid hormone balance. The placenta serves as an important environment for fetal development and also regulates thyroid hormone supply to the fetus. However, epigenetic changes of thyroid regulating genes in placenta have rarely been studied. This study was conducted to evaluate the association between several POP concentrations in maternal serum and DNA methylation of thyroid hormone-related genes in the placenta. The placenta samples were collected from 106 Korean mother at delivery, and the promoter methylation of the placental genes was measured by a bisulfite pyrosequencing. The deiodinase type 3 (DIO3), monocarboxylate transporter 8 (MCT8), and transthyretin (TTR) genes were selected as the target genes as they play an important role in the regulation of fetal thyroid balance. Because people are exposed to multiple chemicals at the same time, a multiple-POP model using principal component analysis (PCA) was applied to evaluate the association between the multiple POPs exposure and the epigenetic change in placenta. In addition, a single-POP model which includes one chemical each in the statistical model for association was conducted. Based on the single-POP models, serum concentrations of p,p'-dichlorodiphenyldichloroethylene (p,p'-DDE) and brominated diphenyl ether-47 (BDE-47) were significantly associated with an increase in placental DIO3 methylation, but only among female infants. Among male infants, a positive association between serum p,p'-DDT and MCT8 methylation level was found. According to the multiple-POP models, serum DDTs were positively associated with DIO3 methylation in the placenta of female infants, while a positive association with MCT8 methylation was observed in those of the male infants. Our observation showed that in utero exposure to DDTs may influence the DNA methylation of DIO3 and MCT8 genes in the placenta, in a sexually dimorphic manner. These alterations in placental epigenetic regulation may in part explain the thyroid hormone disruption observed among the newborns or infants followed by in utero exposure to POPs., (Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Precise mechanisms underlying interleukin-7 (IL-7)-mediated tumor invasion remain unclear. Thus, we investigated the role of IL-7 in tumor invasiveness using metastatic prostate cancer PC-3 cell line derivatives, and assessed the potential of IL-7 as a clinical target using a Janus kinase (JAK) inhibitor and an IL-7-blocking antibody. We found that IL-7 stimulated wound-healing migration and invasion of PC-3 cells, increased phosphorylation of signal transducer and activator of transcription 5, Akt, and extracellular signal-regulated kinase. On the other hand, a JAK inhibitor and an IL-7-blocking antibody decreased the invasiveness of PC-3 cells. IL-7 increased tumor sphere formation and expression of epithelial-mesenchymal transition (EMT) markers. Importantly, lentiviral delivery of IL-7Rα to PC-3 cells significantly increased bone metastasis in an experimental murine metastasis model compared to controls. The gene expression profile of human prostate cancer cells from The Cancer Genome Atlas revealed that EMT pathways are strongly associated with prostate cancers that highly express both IL-7 and IL-7Rα. Collectively, these data suggest that IL-7 and/or IL-7Rα are promising targets of inhibiting tumor metastasis.

Peritoneal metastasis is one of the major patterns of unresectability in colorectal cancer (CRC) and a cause of death in advanced CRC. Identification of distinct gene expressions between primary CRC and peritoneal seeding metastasis is to predict the metastatic potential of primary human CRC. Three pairs of primary CRC (SNU-2335A, SNU-2404A, and SNU-2414A) and corresponding peritoneal seeding (SNU-2335D, SNU-2404B, and SNU-2414B) cell lines were established to determine the different gene expressions and resulting aberrated signaling pathways in peritoneal metastasis tumor using whole exome sequencing and microarray. Whole exome sequencing detected that mutation in CYP2A7 was exclusively shared in peritoneal seeding cell lines. Microarray identified that there were five upregulated genes (CNN3, SORBS1, BST2, EPSTI1, and KLHL5) and two downregulated genes (TRY6 and STYL5) in the peritoneal metastatic cell lines. CNN3 expression was highly augmented in both mRNA and protein levels in peritoneal metastasis cells. Knockdown of Calponin 3 resulted in augmented level of E-cadherin in peritoneal metastasis cells, and migration and invasiveness decreased accordingly. We suggest that CNN3 takes part in cell projection and movement, and the detection and distribution of CNN3 may render prognostic information for predicting peritoneal seeding metastasis from primary colorectal cancer., (Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.)

Prenatal exposure to persistent organic pollutants (POPs) has been linked to numerous adverse birth outcomes among newborn infants in many epidemiological studies. Although epigenetic modifications have been suggested as possible explanations for those associations, studies have rarely reported a relationship between POP exposure during pregnancy and DNA methylation in the placenta. In the present study, we investigated the association between prenatal exposure to several POPs, including organochlorine pesticides (OCPs), polybrominated diphenyl ethers (PBDEs), and polychlorinated biphenyls (PCBs), and methylation levels of long interspersed element 1 (LINE-1), as well as imprinted genes in placental DNAs among Korean mother-child pairs (N = 109). We assessed the association of DNA methylation not only with each target POP (single-POP models) but also with multiple POPs applying principal component analysis (multiple-POP models). Potential associations between placental DNA methylation and birth outcomes of newborn infants were also estimated. In single-POP models, significant associations were detected between OCP measurements and placental DNA methylation. Elevated concentrations of β-hexachlorhexane (β-HCH) in maternal serum collected during delivery were significantly associated with a decrease in methylation of LINE-1 in the placenta. Higher levels of p,p'-dichlorodiphenyltrichloroethane (p,p'-DDT) in maternal serum were associated with hypermethylation of insulin-like growth factor 2 (IGF2). In multiple-POP models, a significant and positive association between DDTs and IGF2 methylation was also observed. Placental LINE-1 methylation was inversely associated with birth length. Our observations indicate that prenatal exposure to several POPs including DDTs is associated with the changes in methylation of genes, including major imprinted genes in the placenta. The consequences of these epigenetic alterations in placenta during development deserve further investigation., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Background: Resistance to preoperative radiotherapy is a major clinical problem in the treatment for locally advanced rectal cancer. The role of NDRG1 in resistance to ionizing radiation in rectal cancer has not been fully elucidated. This study aimed to investigate the effect of the reduced intracellular NDRG1 expression on radio-sensitivity of human rectal cancer cells for exploring novel approaches for treatment of rectal cancer., Methods: Three radio-resistant human rectal cancer cell lines (SNU-61R80Gy, SNU-283R80Gy, and SNU-503R80Gy) were established from human rectal cancer cell lines (SNU-61, SNU-283, and SNU-503) using total 80 Gy of fractionated irradiation. Microarray analysis was performed to identify differently expressed genes in newly established radio-resistant human rectal cancer cells compared to parental rectal cancer cells., Results: A microarray analysis indicated the RNA expression of five genes (NDRG1, ERRFI1, H19, MPZL3, and UCA1) was highly increased in radio-resistant rectal cancer cell lines. Short hairpin RNA-mediated silencing of NDRG1 sensitized rectal cancer cell lines to clinically relevant doses of radiation by causing more DNA double strand breakages to rectal cancer cells when exposed to radiation., Conclusions: Targeting NDRG1 represents a promising strategy to increase response to radiotherapy in human rectal cancer.