China Anti-cancer AssociationActa Medica Okayama2095-39412026SPRED2 suppresses the stemness of hepatocellular carcinoma through the p53/miR-506-3p/KLF4 pathwayENTongGaoDepartment of Pathology and Experimental Medicine, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversitySachioItoDepartment of Pathophysiology and Drug Discovery, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityAyeMoh-Moh-AungDepartment of Pathology and Experimental Medicine, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityTianyiWangDepartment of Pathology and Experimental Medicine, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityMasayoshiFujisawaDepartment of Pathology and Experimental Medicine, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityToshiakiOharaDepartment of Pathology and Experimental Medicine, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityTeizoYoshimuraDepartment of Pathology and Experimental Medicine, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityAkihiroMatsukawaDepartment of Pathology and Experimental Medicine, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityObjective: We previously reported that endogenous Sprouty-related, EVH1 domain-containing protein 2 (SPRED2), an inhibitor of the Ras/Raf/ERK-MAPK pathway, controls hepatocellular carcinoma (HCC) cell stemness by downregulating the expression of pluripotency factors, such as Nanog, c-Myc, and KLF4, in an ERK-dependent fashion. However, the exact mechanisms by which SPRED2 regulates HCC cell stemness have not been established.<br>
Methods: Three human HCC cell lines [HepG2 (parental and SPRED2-deficient), HLE, and Hep3B] were used. Cells were transfected to downregulate or overexpress proteins. Western blot and RT-qPCR were used to evaluate the level of protein and mRNA expression. Co-immunoprecipitation and ChIP-qPCR were used to examine protein-protein interactions and the activation of gene transcription. Clinical HCC tissues were also used to validate in vitro data.<br>
Results: KLF4 was identified as the major pluripotency factor responsible for SPRED2-mediated downregulation of HCC cell stemness and KLF4 expression was regulated by miR-506-3p. SPRED2 formed a protein complex with the tumor suppressor (p53) and upregulated miR-506 gene transcription by binding to the promoter region, resulting in subsequent downregulation of KLF4 mRNA expression. There was a negative correlation between KLF4 expression and miR-506-3p and a positive correlation between miR-506-3p expression and SPRED2 in human HCC samples, highlighting the relevance of the study findings.<br>
Conclusions: The current study revealed a novel SPRED2/p53/miR-506-3p/KLF4 axis through which SPRED2 contributes to the suppression of HCC cell stemness and provides a potential new target to prevent HCC progression.No potential conflict of interest relevant to this article was reported.Springer Science and Business Media LLCActa Medica Okayama1573-72092842025Cancer-associated fibroblast-derived SOD3 enhances lymphangiogenesis to drive metastasis in lung adenocarcinoma51ENMay WathoneOoDepartment of Pathophysiology and Drug Discovery, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityTakaoHikitaDepartment of Pathophysiology and Drug Discovery, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityTomohaMashimaDepartment of Pathophysiology and Drug Discovery, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityKosukeTorigataSchool of Medicine, Kobe UniversityYin MinThuDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityTomohiroHabuDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityHotakaKawaiDepartment of Oral Pathology and Medicine, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityToshiakiOharaDepartment of Pathology and Experimental Medicine, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityShutaTomidaCenter for Comprehensive Genomic Medicine, Okayama University HospitalSachioItoDepartment of Pathophysiology and Drug Discovery, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityKenSuzawaDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityHitoshiNagatsukaDepartment of Thoracic Surgery, National Hospital Organization, Shikoku Cancer CenterShinichiToyookaDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityMasanoriNakayamaDepartment of Pathophysiology and Drug Discovery, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityDespite advancements in diagnostic and therapeutic strategies, lung adenocarcinoma (LUAD) remains a leading cause of cancer-related mortality due to its aggressive metastatic potential. Extracellular superoxide dismutase (SOD3) is an antioxidant enzyme that regulates oxidative stress and is regarded as a tumor suppressor. However, studies have demonstrated that SOD3 can either promote or inhibit cell proliferation and survival in various cancers, and its molecular mechanisms within the tumor microenvironment are poorly understood. In this study, we report a breakthrough in uncovering the role of SOD3 derived from cancer-associated fibroblasts (CAFs) in LUAD. Using LUAD xenograft models co-implanted with SOD3-overexpressing CAFs (CAFSOD3), we observe an aggressive tumor phenotype characterized by increased lymphangiogenesis and lymphatic vessel invasion (LVI) of the tumor. Additionally, LUAD patients with elevated SOD3 levels exhibit a higher incidence of LVI and metastasis. Notably, RNA sequencing of CAFSOD3 reveals that SOD3-mediated VEGF-dependent tumor progression and lymphangiogenesis are up-regulated. Furthermore, single-cell transcriptomic analysis of LUAD clinical samples confirms a strong correlation between SOD3 expression in fibroblasts and characteristics of tumor exacerbation, such as lymphangiogenesis and metastasis. These findings underscore new insights into the role of CAF-derived SOD3 in LUAD progression and highlight its potential as a biomarker and therapeutic target.No potential conflict of interest relevant to this article was reported.Spandidos PublicationsActa Medica Okayama2049-94341352020Human NINEIN polymorphism at codon 1111 is associated with the risk of colorectal cancer45ENYukikoYasudaDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesAkikoSakaiDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesSachioItoDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKaoriSasaiDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesAkisadaIshizakiDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesYoshiyaOkanoDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesSeitoKawaharaDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesYoshimiJitsumoriDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesHiromasaYamamotoDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesNagahideMatsubaraDepartment of Gastroenterological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKenjiShimizuDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesHiroshiKatayamaDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesNINEIN serves an essential role in centrosome function as a microtubule organizing center, and in the reformation of the interphase centrosome architecture following mitosis. In the present study, the association between NINEIN Pro1111Ala (rs2236316), a missense single nucleotide polymorphism, and the risk of colorectal cancer (CRC), related to smoking and alcohol consumption habits in 200 patients with CRC and 1,141 cancer‑free control participants were assessed in a case‑control study performed in Japan. The results showed that the NINEIN Ala/Ala genotype compared with the Pro/Pro genotype was significantly more associated with an increased risk of CRC, and the males with the Ala/Ala genotype exhibited a significantly increased risk of CRC compared with those with Pro/Pro and Pro/Ala genotypes. Stratified analyses of the Ala/Ala genotype with CRC risk further showed an increased association in never/light drinkers (<23 g of ethanol/day), in male never/light drinkers and in male patients with rectal cancer. These findings suggest that the genetic variant of the NINEIN Pro1111Ala polymorphism has a significant effect on CRC susceptibility in the Japanese population.No potential conflict of interest relevant to this article was reported.Okayama University Medical SchoolActa Medica Okayama0386-300X7112017Human RAD 17 Polymorphism at Codon 546 Is Associated with the Risk of Colorectal Cancer5968ENYukikoYasudaDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesAkikoSakaiDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesSachioItoDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKaoriSasaiDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesHiromasaYamamotoDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesNagahideMatsubaraDepartment of Gastroenterological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesMamoruOuchidaDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesHiroshiKatayamaDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKenjiShimizuDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesOriginal Article10.18926/AMO/54826Human RAD17 acts as an activator of checkpoint signals in response to DNA damage. Here we evaluated the association of hRAD17 Leu546Arg (rs1045051), a missense single nucleotide polymorphism, with the risk of colorectal cancer (CRC) in relation to smoking and alcohol consumption habits in 212 CRC patients and 1,142 cancer-free controls in a case-control study conducted in Japan. The results showed that the hRAD17 Leu/Arg genotype compared to the Leu/Leu genotypes was significantly associated with the protective effect on CRC risk with the adjusted odds ratio (OR) of 0.68 [95% confidence interval (CI): 0.49|0.95, p=0.024], and the males with the Arg/Arg genotype had a greater risk of CRC compared to those with the Leu/Leu and Leu/Arg genotypes (OR=1.87, 95%CI 1.03|3.40, p=0.04). In stratified studies, the protective effect of the Leu/Arg genotype on CRC risk was markedly higher in the light smokers (< 20 pack years) (OR=0.61, 95%CI 0.40|0.94, p=0.024) and the rectal cancer patients (OR=0.49, 95%CI 0.31|0.78, p=0.003). The risk of the Arg/Arg genotype was associated with heavy smoking (≥ 20 pack-years) (OR=2.24, 95%CI 1.09|4.61, p=0.03). These findings suggest that the genetic variant of hRAD17 Leu546Arg polymorphism has a significant effect on CRC susceptibility in Japanese.No potential conflict of interest relevant to this article was reported.Public Library ScienceActa Medica Okayama1932-62038102013Truncated SSX Protein Suppresses Synovial Sarcoma Cell Proliferation by Inhibiting the Localization of SS18-SSX Fusion ProteinENYasushiYonedaSachioItoToshiyukiKunisadaYukiMorimotoHirotakaKanzakiAkiYoshidaKenjiShimizuToshifumiOzakiMamoruOuchidaSynovial sarcoma is a relatively rare high-grade soft tissue sarcoma that often develops in the limbs of young people and induces the lung and the lymph node metastasis resulting in poor prognosis. In patients with synovial sarcoma, specific chromosomal translocation of t(X; 18) (p11.2; q11.2) is observed, and SS18-SSX fusion protein expressed by this translocation is reported to be associated with pathogenesis. However, role of the fusion protein in the pathogenesis of synovial sarcoma has not yet been completely clarified. In this study, we focused on the localization patterns of SS18-SSX fusion protein. We constructed expression plasmids coding for the full length SS18-SSX, the truncated SS18 moiety (tSS18) and the truncated SSX moiety (tSSX) of SS18-SSX, tagged with fluorescent proteins. These plasmids were transfected in synovial sarcoma SYO-1 cells and we observed the expression of these proteins using a fluorescence microscope. The SS18-SSX fusion protein showed a characteristic speckle pattern in the nucleus. However, when SS18-SSX was co-expressed with tSSX, localization of SS18-SSX changed from speckle patterns to the diffused pattern similar to the localization pattern of tSSX and SSX. Furthermore, cell proliferation and colony formation of synovial sarcoma SYO-1 and YaFuSS cells were suppressed by exogenous tSSX expression. Our results suggest that the characteristic speckle localization pattern of SS18-SSX is strongly involved in the tumorigenesis through the SSX moiety of the SS18-SSX fusion protein. These findings could be applied to further understand the pathogenic mechanisms, and towards the development of molecular targeting approach for synovial sarcoma.No potential conflict of interest relevant to this article was reported.Acta Medica Okayama1535-71631092011Liposomal Delivery of MicroRNA-7-Expressing Plasmid Overcomes Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitor-Resistance in Lung Cancer Cells17201727ENKammeiRaiNagioTakigawaSachioItoHiromiKashiharaEikiIchiharaTatsujiYasudaKenjiShimizuMitsuneTanimotoKatsuyukiKiuraEpidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKI) have been strikingly effective in lung cancers harboring activating EGFR mutations. Unfortunately, the cancer cells eventually acquire resistance to EGFR-TKI. Approximately 50% of the acquired resistance involves a secondary T790M mutation. To overcome the resistance, we focused on EGFR suppression using microRNA-7 (miR-7), targeting multiple sites in the 30-untranslated region of EGFR mRNA. Two EGFR-TKI-sensitive cell lines (PC-9 and H3255) and two EGFR-TKI-resistant cell lines harboring T790M (RPC-9 and H1975) were used. We constructed miR-7-2 containing miR-7-expressing plasmid. After transfection of the miR-7-expressing plasmid, using cationic liposomes, a quantitative PCR and dual luciferase assay were conducted to examine the efficacy. The antiproliferative effect was evaluated using a cell count assay and xenograft model. Protein expression was examined by Western blotting. The miR-7 expression level of the transfectants was approximately 30-fold higher, and the luciferase activity was ablated by 92%. miR-7 significantly inhibited cell growth not only in PC-9 and H3255 but also in RPC-9 and H1975. Expression of insulin receptor substrate-1 (IRS-1), RAF-1, and EGFR was suppressed in the four cell lines. Injection of the miR-7-expressing plasmid revealed marked tumor regression in a mouse xenograft model using RPC-9 and H1975. EGFR, RAF-1, and IRS-1 were suppressed in the residual tumors. These findings indicate promising therapeutic applications of miR-7-expressing plasmids against EGFR oncogene-addicted lung cancers including T790M resistance by liposomal delivery. Mol Cancer Ther; 10(9); 1720-7.No potential conflict of interest relevant to this article was reported.ͺRγwοActa Medica Okayama0030-155812432012EGFR`VLi[[jQςΟ«xΰΧEΙΞ·ιmicroRNA-7»vX~hΜ|\[πp’½±όΙζιΜ’207210ENKammeiRaiNagioTakigawaSachioItoHiromiKashiharaEikiIchiharaTatsujiYasudaKenjiShimizuMitsuneTanimotoKatsuyukiKiuraNo potential conflict of interest relevant to this article was reported.Acta Medica Okayama2001A Novel WD40 Repeat Protein, WDC146, Highly Expressed during Spermatogenesis in a Stage-specific MannerENNo potential conflict of interest relevant to this article was reported.