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PURPOSE: Histological transformation from epidermal growth factor receptor (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 (TKIs). However, transcriptomic changes between NSCLC and transformed SCLC (t-SCLC) remain unexplored. EXPERIMENTAL DESIGN: We conducted whole transcriptome analysis of 59 regions of interest (ROIs) through the spatial profiling of FFPE tissues obtained from ten patients (lung adenocarcinoma, 22; combined SCLC/NSCLC, 7; t-SCLC, 30 ROIs). Transcriptomic profiles and differentially expressed genes (DEGs) were compared between pre- and post-transformed tumors. RESULTS: Following EGFR-TKI treatment, 93.7% (15/16) of transformed-SCLC (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 (HDAC) inhibitors restored EGFR expression in SNU-2962A cells and their organoid model. The synergistic effects of third-generation EGFR-TKI osimertinib and the HDAC inhibitor fimepinostat were validated in both in vitro and in vivo models. CONCLUSIONS: Our study demonstrated most t-SCLC showed neuronal subtypes with low EGFR expression. DEGs analysis and t-SCLC preclinical models identified an epigenetic modifier as a promising treatment strategy for t-SCLC.
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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.
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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.
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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.
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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 G0/G1 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.
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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.
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Neoplasias Encefálicas , Linhagem Celular Tumoral , Genes Neoplásicos , Glioblastoma , Humanos , Neoplasias Encefálicas/genética , Genômica , Glioblastoma/genética , MutaçãoRESUMO
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.
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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.
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Carcinoma Ductal Pancreático , Carcinoma de Células Escamosas , Neoplasias Pancreáticas , Proteogenômica , Animais , Camundongos , Humanos , Neoplasias Pancreáticas/genética , Carcinoma Ductal Pancreático/genética , Biomarcadores , Neoplasias PancreáticasRESUMO
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.
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Neoplasias Encefálicas , Glioblastoma , Humanos , Temozolomida/uso terapêutico , Glioblastoma/tratamento farmacológico , Forma Celular , Metabolismo dos Lipídeos , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismo , Resistencia a Medicamentos Antineoplásicos , Lipídeos , Linhagem Celular Tumoral , Neoplasias Encefálicas/tratamento farmacológico , Antineoplásicos Alquilantes/farmacologiaRESUMO
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 (Pprogress = 0.011, Pdecease = 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.
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Neoplasias Colorretais , Prevotella , Humanos , Prevotella/genética , Estudos Prospectivos , Fezes , Bactérias/genética , Fusobacterium nucleatum/genética , Neoplasias Colorretais/cirurgiaRESUMO
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.
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Glioma , Telomerase , Glioma/genética , Humanos , Mutação , Telomerase/genética , Telômero/genética , Homeostase do TelômeroRESUMO
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.
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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/ß-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/ß-catenin in ovarian cancer treatment. This study aims to investigate the effect of CWP232291, a small molecular Wnt/ß-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 ß-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/ß-catenin pathway inhibition as a novel therapeutic strategy against ovarian cancer.
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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.
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Antineoplásicos , Neoplasias do Colo , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/patologia , Ensaios de Seleção de Medicamentos Antitumorais , Ensaios de Triagem em Larga Escala , Humanos , Organoides/patologiaRESUMO
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.
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Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Derrame Pleural Maligno , Derrame Pleural , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Mutação , Derrame Pleural/complicações , Derrame Pleural Maligno/diagnósticoRESUMO
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.