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Introduction: Metastasis is a leading cause of cancer-related-deaths worldwide. Recently, cancer stem cells (CSCs) have been believed to be responsible for tumor initiation and metastasis, but till now, difference of cellular features and behaviors between CSCs from tumor tissues (TCSCs) and circulation (CCSCs) remains largely unknown, which hinders the progression of targeted therapies for metastasis. Methods and Results: Here, we provide the features of circulating gastric cancer stem cells (CGCSCs) isolated from human gastric adenocarcinoma. The CGCSCs and TGCSCs were culture in a same serum free stem cell culture medium, however the morphology are different with each other. EMT-associated markers were measured by Immunofluorescence, Western Blotting, and RT-PCR methods, and the results indicated that the CGCSCs and TGCSCs carry different epithelial-mesenchymal features. And then, proliferation and apoptosis assays revealed that the CGCSCs exhibited characteristics of higher proliferation and resistance to apoptosis in vitro. Soft agar assay and nude mice tumorigenicity assay displayed strong tumorigenicity of CGCSCs. Finally, Matrigel invasion assays and in vivo experimental metastasis assay were also performed, which demonstrated that CGCSCs carry high invasive and metastatic capabilities than TGCSCs. Conclusions: As expected, the CGCSCs indeed showed extremely invasive and metastatic properties. They also exhibited distinctive mesenchymal phenotypes, high self-renewal, proliferative capabilities, tumor induction and low apoptosis. Interestingly, CGCSCs show small cell-size than TGCSCs (tissue gastric cancer stem cells). The findings might help us to understand the biological characteristic of CGCSCs deeply, and give light to strategies for cancer therapies. [ABSTRACT FROM AUTHOR]

Purpose: Circulating tumor cells (CTCs) have been proved to be responsible for tumor metastasis and resistant to anticancer therapies. This study aims to isolate and characterize circulating tumor cells from human gastric cancer patients, and investigate characteristic differences between gastric CTCs and gastric cancer cell lines. Methods: We analyzed 31 cases of gastric cancer patients using anti-CD45 antibody-conjugated magnetic microbeads negative separation, combined with fluorescence activated cell sorter CD44 positive screening. Abilities of tumor formation, metastasis, invasion, migration, irradiation and drug sensitivity of CTCs and gastric cancer cell lines were detected and compared. Results: Of all the 31 patients, CD44/CD45CTCs were isolated in 14 patients, of which 3 cases were stage IIA, 2 cases stage IIB, 2 cases stage IIIC and 7 cases stage IV. The malignant behavior was demonstrated by both clonogenetic assay and tumor xenograft in nude mice. Compared with human gastric cancer cell lines, the migration and invasion abilities of CTCs increased to 3.21-12.6-fold and 2.3-6.7-fold, respectively (all p values <0.05). In addition, the metastatic potential of CTCs is much higher in vivo than that of the control. Furthermore, CTCs were found to be relatively sensitive to FU, cisplatin and paclitaxel, but relatively resistant to irradiation, oxaliplatin, cetuximab and trastuzumab. Conclusions: CD44/CD45 gastric CTCs were isolated and found to exhibit stronger malignant behavior when compared with human gastric cancer cell lines. Furthermore, CTCs cultured in vitro have potential implications in drug sensitivity screening for the future anticancer treatments. [ABSTRACT FROM AUTHOR]

Purpose: Hypoxia has been found to play an important role in regulating the biological characteristics of cancer stem cells (cCSCs). In this study, we tested whether a tumor hypoxic niche serves to the chemotherapeutic resistance of colon cCSCs. Methods: Each of 23 fresh samples of human colon adenocarcinoma was transplanted into nude mice. The tumor-bearing mice randomly and equally received (A) saline, (B) 5-fluorouracil (15 mg/kg), (C) oxaliplatin (10 mg/kg), and (D) oxaliplatin plus 5-fluorouracil when xenografts reached 250 mm ( n = 10). After 2-week treatment, tumor cells were quantified by flow cytometry for expression of CD133 and the hypoxic proportion of CD133 and CD133 cells which were also sorted and detected for ki67 and pimonidazole via immunofluorescence. Results: The hypoxic subpopulation of CD133 and CD133 cells was 66.5 and 26.4 %, respectively. Although there was no marked change for the hypoxic subpopulation of CD133 cells after treatment, the hypoxic fraction of proliferative CD133 cells was increased by 14.62, 16.45, and 20.46 % in groups B, C, and D, respectively. Furthermore, proliferative cells in CD133 and CD133 cells were reduced by 29.93 and 62.5 % in group C, and by 25.26 and 68.22 % in group D; in group B, however, the proliferative CD133 cells were increased by 37.09 %; the CD133 cells were unchanged. Conclusions: Most CD133 cCSCs are located in a hypoxic niche, where cCSCs are better at retaining proliferating property under chemotherapy. Oxaliplatin, rather than 5-FU, inhibits proliferation of cCSCs, which may be the mechanism underlying a better outcome by oxaliplatin in colon cancer patients. [ABSTRACT FROM AUTHOR]

Purpose: The mammalian Janus kinase (JAK) family plays a critical role in cytokine/growth factor signalling pathways and is associated with human cancers. In this study, we explored the role of JAK1 in the non-small cell lung cancer (NSCLC) cell line A549 and its molecular crosstalk with the phosphatidyl inositol-3-kinase (PI3K)/mammalian target of the rapamycin (mTOR) pathway. Methods: One hundred and two NSCLC and 50 normal lung specimens were collected after surgical resection. JAK1 expression and phosphorylation were determined via immunohistochemical staining (IHC) assay. A stable knockdown of JAK1 was performed in A549 cells by RNA interference. Stable cell proliferation, cell cycle, apoptosis, and invasion were characterised in vitro. Tumourigenicity was analysed in vivo. The NSCLC xenograft protein expression of PI3K/mTOR pathway molecules was determined by Western blot assay. Results: JAK1 expression was higher in NSCLC tissues than in normal lung tissues ( P < 0.01). JAK1 knockdown in A549 cells significantly inhibited cell proliferation and invasion while promoting cell arrest at G0/G1 phase (all P < 0.05). The xenograft model showed that JAK1 suppression inhibited tumour growth compared with normal control ( P < 0.05). Moreover, JAK1 knockdown inhibited mTOR or P70 ribosomal protein S6 kinase (P70S6K) phosphorylation, but increased glycogen synthase kinase-3α (GSK-3α) and B-cell lymphoma-extra large (Bcl-xl) phosphorylation. Total protein expression and Akt1/2 phosphorylated status remained unchanged. Conclusion: Our study suggests that JAK1 expression and phosphorylation is abnormal in NSCLC tissues. The knockdown of JAK1 significantly inhibits tumourigenicity of the A549 cell line and demonstrates that crosstalk between the JAK1 and PI3K/mTOR pathways is involved. [ABSTRACT FROM AUTHOR]

Purpose: Pyruvate kinase isoenzyme M2 (PKM2) is a key enzyme in aerobic glycolysis; inhibition of PKM2 leads to the tumor growth inhibition. In this study, the effects of combined treatment with cisplatin (DDP) and a plasmid that expresses a short hairpin RNA (shRNA) targeting PKM2 on the growth of human A549 xenograft lung cancer model were investigated. Methods: The expression of PKM2 in A549 cells was determined by immunofluorescence. PKM2 expression levels were evaluated by Western blot analysis. In a human A549 lung cancer xenograft model, the effects of treatment with shRNA, with or without cisplatin, on tumor volume were determined. Apoptosis and cell proliferation status were examined to determine the mechanisms of tumor growth inhibition. Results: Expression of shRNA targeting PKM2 resulted in inhibition of PKM2 expression in A549 cells. In the lung cancer xenograft model, average tumor volume in the group treated with both cisplatin and shRNA was statistically lower than those of other groups ( P < 0.05). The levels of apoptotic cells were significantly higher in samples from animals in the combined treatment group than those from untreated animals ( P < 0.05). The cell proliferation rate, as determined by counting cells labeled with an anti-phospho-histone H3, a marker for mitosis, was lower in samples from animals treated with both cisplatin and shRNA than in samples from other groups ( P < 0.05). Conclusions: Use of RNA interfering (RNAi) targeting PKM2 significantly inhibited tumor growth when combined with cisplatin in a human A549 lung cancer xenograft model. The enhanced antitumor activity of the combined treatment compared to treatment with shRNA alone may result in part from increased induction of apoptosis and augmented inhibition of cancer cell proliferation. [ABSTRACT FROM AUTHOR]