Background
The attainment of drug resistance in gastric cancer (GC) is a problematic issue. Although many studies have shown that cancer stem cells (CSCs) play an important role in the acquisition of ...drug resistance, there is no clinically available biomarker for predicting oxaliplatin (L-OHP) resistance in relation to CSCs. Organoid technology, a novel 3D cell culture system, allows harboring of patient-derived cancer cells containing abundant CSCs using niche factors in a dish.
Methods
In this study, we established L-OHP-resistant gastric cancer organoids (GCOs) and evaluated their gene expression profile using microarray analysis. We validated the upregulated genes in the L-OHP-resistant GCOs compared to their parental GCOs to find a gene responsible for L-OHP resistance by qRT-PCR, immunohistochemistry, in vitro, and in vivo experiments.
Results
We found myoferlin (MYOF) to be a candidate gene through microarray analysis. The results from cell viability assays and qRT-PCR showed that high expression of MYOF correlated significantly with the IC50 of L-OHP in GCOs. Immunohistochemistry of MYOF in GC tissue samples revealed that high expression of MYOF was significantly associated with poor prognosis, T grade, N grade, and lymphatic invasion, and showed MYOF to be an independent prognostic indicator, especially in the GC patients treated with platinum-based chemotherapy. The knockdown of MYOF repressed L-OHP resistance, cell growth, stem cell features, migration, invasion, and in vivo tumor growth.
Conclusions
Our results suggest that MYOF is highly involved in L-OHP resistance and tumor progression in GC. MYOF could be a promising biomarker and therapeutic target for L-OHP-resistant GC cases.
5-FU is one of the key drugs in the treatment of gastric cancer (GC). Much evidence has shown that cancer stem cells (CSCs) play a key role in the acquisition of drug resistance. The organoid is a ...novel 3D cell culture system technology that sustains stem-cell-driven formation of near-physiological, self-renewing tissues using specific niche factors in a dish. In this study, we established GC organoids (GCOs) and gradually treated them with higher concentrations of 5-FU. We successfully harvested four 5-FU-resistant GCOs, which were supported by significant changes in the expression of molecules related to 5-FU metabolism. We then performed microarray analysis using three normal gastric organoids and three pairs of 5-FU-resistant and parental GCOs. Through the comparison of expression profiles and further validation, we chose KHDRBS3 as a target gene. We found KHDRBS3 to be an independent prognostic factor in GC patients, especially in GC patients treated with 5-FU chemotherapy. We also determined that KHDRBS3 might play an important role in the acquisition of stem cell-like features, such as multi-drug resistance and organoid formation, by regulating CD44 variant expression. We found KHDRBS3, which is thought to play an important role in the acquisition of characteristics of CSCs in GC, to be a promising candidate marker for predicting therapeutic effect and prognosis in GC patients.
DNA-damaging agents include first-line drugs such as platinum (cisplatin, carboplatin), topoisomerase inhibitors (etoposide, doxorubicin), and replication inhibitors (cytarabine, gemcitabine). ...Despite their wide and long usage, there is no clinically available biomarker to predict responses to these drugs. Schlafen 11 (SLFN11), a putative DNA/RNA helicase, recently emerged as a dominant determinant of sensitivity to these drugs by enforcing the replication block in response to DNA damage. Since the clinical importance of SLFN11 is implicated, a comprehensive analysis of SLFN11 expression across human organs will provide a practical resource to develop the utility of SLFN11 in the clinic. In this study, we established a scoring system of SLFN11 expression by immunohistochemistry (IHC) and assessed SLFN11 expression in ~ 700 malignant as well as the adjacent non-tumor tissues across 16 major human adult organs. We found that the SLFN11 expression is tissue specific and varies during tumorigenesis. Although The Cancer Genome Atlas (TCGA) is a prevailing tool to assess gene expression in various malignant and normal tissues, our IHC data exhibited obvious discrepancy from the TCGA data in several organs. Importantly, SLFN11-negative tumors, potentially non-responders to DNA-damaging agents, were largely overrated in TCGA because TCGA samples are a mixture of infiltrating immune cells, including T cells, B cells, and macrophages, which have strong SLFN11 expression. Thus, our study reveals the significance of immunohistochemical procedures for evaluating expression of SLFN11 in patient samples and provides a robust resource of SLFN11 expression across adult human organs.
5‐Fluorouracil (5‐FU) is one of the most frequently used pharmacological agents in the treatment of colorectal cancer (CRC). Resistance to chemotherapy is a major cause of treatment failure of CRC, ...and it is a well known fact that cancer stem cells play a significant role in the acquisition of drug resistance. In this study, we focused on the KHDRBS3 gene that encodes KH RNA Binding Domain Containing, Signal Transduction Associated 3. We first clarified the relationship between KHDRBS3 and 5‐FU resistance. We then observed higher expression levels of KHDRBS3 in KRAS‐mutant organoids and cell lines in comparison with KRAS wild‐type organoids and cell lines. Immunohistochemical analysis using CRC cases revealed that the prognosis of KHDRBS3‐positive patients was significantly worse compared with that of KHDRBS3‐negative patients. Univariate and multivariate Cox proportional hazards analyses showed that KHDRBS3 was an independent prognostic factor in patients with CRC. We determined that KHDRBS3 might play a crucial role in the acquisition of stem cell properties, such as drug resistance and spheroid/organoid formation, by regulating CD44 variant expression and the Wnt signaling pathway. In an immunodeficient mouse model, KHDRBS3‐positive cells showed efficient tumor formation and formed metastatic lesions in the lungs. These results indicated that KHDRBS3 plays a crucial role in drug resistance and anchorage‐independent growth by maintaining stem cell‐like features in CRC cells. KHDRBS3 could be a promising candidate marker for predicting chemotherapeutic effect and prognosis in CRC patients.
KHDRBS3 plays a crucial role in drug resistance and anchorage‐independent growth by maintaining stem cell‐like features in CRC cells. KHDRBS3 could be a promising candidate marker for predicting chemotherapeutic effect and prognosis in CRC patients.
Background
Transcribed ultraconserved regions (T-UCRs) are a novel class of noncoding RNAs that are highly conserved among the orthologous regions in most vertebrates. It has been reported that ...T-UCRs have distinct signatures in human cancers. We previously discovered the downregulation of T-UCR expression in gastric cancer (GC), indicating that T-UCRs could play an important role in GC biology. Uc.160+, a T-UCR reported to be downregulated in human cancer, has not been examined in GC.
Methods
We analyzed the expression pattern of Uc.160+ in nonneoplastic and tumor tissues of the stomach by using uantitative reverse transcription polymerase chain reaction (qRT-PCR) and in situ hybridization (ISH), specifically focusing on the mechanism of transcriptional regulation and target genes that are regulated by T-UCRs. We also attempted to determine the effect of Uc.160+ expression on biological features of GC cell lines by Western blotting.
Results
On the basis of the qRT-PCR and ISH results, Uc.160+ expression in adenoma and GC tissues was clearly downregulated compared with that in nonneoplastic mucosa tissues of the stomach. Cancer-specific DNA methylation in the promoter region of Uc.160 was observed by bisulfite genomic DNA sequencing analysis. The effect of DNA methylation on Uc.160+ expression was further confirmed by reporter gene assay. We also revealed that Uc.160+ inhibited the phosphorylation of Akt by regulating phosphatase and tensin homolog (PTEN) expression.
Conclusions
These results indicate that Uc.160+ could possibly have a tumor suppressive role in GC.
Background
The transcribed ultraconserved regions (T-UCRs) are a novel class of long non-coding RNAs and are involved in the development of several types of cancer. Although several different papers ...have described the oncogenic role of Uc.63+, there are no reports mentioning its importance in gastric cancer (GC) biology.
Methods
In this study, we evaluated Uc.63+ expression using clinical samples of GC by qRT-PCR, and also assessed the correlation between Uc.63+ expression and clinico-pathological factors.
Results
The upregulation of Uc.63+ was significantly correlated with advanced clinico-pathological features. Knockdown of Uc.63+ significantly repressed GC cell growth and migration, whereas overexpression of Uc.63+ conversely promoted those of GC cells. In situ hybridization of Uc.63+ revealed its preferential expression in poorly differentiated adenocarcinoma. We further conducted a microarray analysis using MKN-1 cells overexpressing Uc.63- and found that NF-κB signaling was significantly upregulated in accordance with Uc.63+ expression.
Conclusion
Our results suggest that Uc.63+ could be involved in GC progression by regulating GC cell growth and migration via NF-κB signaling
The transcribed ultraconserved regions (T-UCRs) are a novel class of non-coding RNAs that are absolutely conserved across species and are involved in carcinogenesis in some cancers. However, the ...expression and biological role of T-UCRs in renal cell carcinoma (RCC) remain poorly understood. This study aimed to examine the expression and functional role of Uc.416 + A and analyze the association between Uc.416 + A and epithelial-to-mesenchymal transition in RCC.
Expression of Uc.416 + A in 35 RCC tissues, corresponding normal kidney tissues and 13 types of normal tissue samples was determined by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). We performed a cell growth and migration assay in RCC cell line 786-O transfected with negative control and siRNA for Uc.416 + A. We evaluated the relation between Uc.416 + A and miR-153, which has a complimentary site of Uc.416 + A.
qRT-PCR analysis revealed that the expression of Uc.416 + A was higher in RCC tissues than that in corresponding normal kidney tissues. Inhibition of Uc.416 + A reduced cell growth and cell migration activity. There was an inverse correlation between Uc.416 + A and miR-153. Western blot analysis showed Uc.416 + A modulated E-cadherin, vimentin and snail. The expression of Uc.416 + A was positively associated with the expression of SNAI1, VIM and inversely associated with the expression of CDH1.
The expression of Uc.416 + A was upregulated in RCC and especially in RCC tissues with sarcomatoid change. Uc.416 + A promoted epithelial-to-mesenchymal transition through miR-153. These results suggest that Uc.416 + A may be a promising therapeutic target.
The term “non-coding RNA” (ncRNA) is generally used to indicate RNA that does not encode a protein and includes several classes of RNAs, such as microRNA and long non-coding RNA. Several lines of ...evidence suggest that ncRNAs appear to be involved in a hidden layer of biological procedures that control various levels of gene expression in physiology and development including stem cell biology. Stem cells have recently constituted a revolution in regenerative medicine by providing the possibility of generating suitable cell types for therapeutic use. Here, we review the recent progress that has been made in elaborating the interaction between ncRNAs and tissue/cancer stem cells, discuss related technical and biological challenges, and highlight plausible solutions to surmount these difficulties. This review particularly emphasises the involvement of ncRNAs in stem cell biology and in vivo modulation to treat and cure specific pathological disorders especially in cancer. We believe that a better understanding of the molecular machinery of ncRNAs as related to pluripotency, cellular reprogramming, and lineage-specific differentiation is essential for progress of cancer therapy.
Although unresectable or recurrent gastric cancers (GC) are frequently treated with platinum-based chemotherapy, response to treatment remains unpredictable. Because Schlafen 11 (SLFN11) is recently ...identified as a critical determinant of platinum sensitivity, we investigated the potential clinical utility of SLFN11 in the treatment of GC.
We analysed the correlation between SLFN11 expression and overall survival in 169 GC patients by our established immunohistochemical approach. The impact of SLFN11 expression on the response to platinum and transition of SLFN11 expression upon long-term treatment with platinum were examined using GC cell lines and organoids.
GC patients with high-SLFN11 expression exhibited significantly better survival than those with low-SLFN11 expression, and the significance increased when we selected patients treated with platinum-based chemotherapy. Knockout of SLFN11 and reactivation of SLFN11 in GC cells conferred resistance and sensitivity to platinum, respectively. In GC cells and organoids, long-term treatment with oxaliplatin suppressed SLFN11 expression while imparting drug resistance. The acquired resistance to oxaliplatin was reversed by reactivation of SLFN11 with epigenetic modifying drugs.
This is the first report revealing definitive clinical implications of SLFN11 in the treatment of GC patients and providing novel strategies for the drug selection based on SLFN11 expression.
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