salinomycin and Stomach-Neoplasms

Chemotherapy is the main treatment for advanced gastric cancer. However, the emergence of multidrug resistance (MDR) has become a major obstacle in chemotherapy in many tumors, including gastric cancer. Epithelial-mesenchymal transition (EMT), which is considered an important process in cancer development, also contributes toward tumor MDR. Salinomycin, an EMT blocker, shows broad-spectrum antitumor and chemosensitization properties. Here, we hypothesized that salinomycin could reverse the MDR of SGC7901/cisplatin (CDDP) gastric cancer cell by inhibiting EMT and further explored its possible underlying mechanisms. Our results indicated higher 50% inhibiting concentration (IC50) and stronger migration capacity in SGC7901/CDDP than in SGC7901 cells, whereas salinomycin could reduce the IC50 (50% inhibition of the concentration of chemodrugs after 4 μmol/l salinomycin treatment) and migration capacity in SGC7901/CDDP cells. At the molecular level, we found that the expression of E-cadherin, ZO-1 decreased, whereas the expression of N-cadherin, Vimentin, ZEB-1, and Twist increased in SGC7901/CDDP cells, and that salinomycin potently blocked the EMT by enhancing the expression of E-cadherin, ZO-1 and reducing the expression of N-cadherin, Vimentin, ZEB-1, and Twist in the above MDR cells. In addition, we also found that long noncoding RNA HOTTIP, an oncogenic regulator, was upregulated in SGC7901/CDDP cells, whereas its downregulation could markedly attenuate the EMT, thereby reversing the MDR. Furthermore, our data showed that the salinomycin-elicited MDR-reversion effect was associated closely with suppression of EMT through inhibition of the expression of long noncoding RNA HOTTIP. Collectively, our findings suggest a new underlying mechanism and applicable therapeutic regimen for MDR gastric cancer.

Topics: Antineoplastic Agents; Apoptosis; Cell Line, Tumor; Cisplatin; Down-Regulation; Drug Resistance, Multiple; Drug Resistance, Neoplasm; Epithelial-Mesenchymal Transition; Gene Expression Regulation, Neoplastic; Humans; Pyrans; RNA, Long Noncoding; Signal Transduction; Stomach Neoplasms; Up-Regulation

The aim of the present study was to investigate the effects and mechanisms of 17‑AAG combined with salinomycin treatment on proliferation and apoptosis of the SGC‑7901 gastric cancer cell line. An MTT assay was used to detect the proliferation of SGC‑7901 cells. Morphological alterations of cells were observed under inverted phase‑contrast and fluorescence microscopes. Cell cycle and apoptosis were assessed by flow cytometry analysis. The protein expression of nuclear factor (NF)‑κB p65 and Fas‑ligand (L) were evaluated by immunocytochemistry. Salinomycin with a concentration range of 1‑32 µmol/l was demonstrated to inhibit growth of SGC‑7901 cells effectively, affect the morphology and apoptosis rate of cells, and arrest SGC‑7901 cells in S phase. Furthermore, salinomycin significantly increased the protein expression of Fas‑L and decreased the protein expression of NF‑κB p65. The alterations in SGC‑7901 cells co‑treated with salinomycin and 17‑AAG were more significant compared with cells treated with one drug only. In conclusion, the individual use of salinomycin and combined use with 17‑AAG may significantly inhibit SGC‑7901 gastric cancer cell proliferation and induce cell apoptosis. The potential mechanisms may be associated with upregulation of Fas‑L and downregulation of NF‑κB. These results provide a basis for the potential use of salinomycin in gastric cancer treatment.

Topics: Apoptosis; Benzoquinones; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Cell Shape; Cell Survival; Fas Ligand Protein; fas Receptor; Humans; Lactams, Macrocyclic; NF-kappa B; Pyrans; Signal Transduction; Stomach Neoplasms

The present study aimed to investigate the anticancer effects of cisplatin (DDP) combined with salinomycin (SAL) on the gastric cancer cell line SGC‑7901, as well as to explore the mechanisms underlying their actions. An MTT assay was used to evaluate the inhibitory effects of SAL, DDP and their combination on gastric cancer cell proliferation. Morphological alterations of cancer cells following treatment were observed under an inverted phase‑contrast microscope and a fluorescence microscope. Cell cycle progression and apoptosis were analyzed using flow cytometry. The expression of nuclear factor (NF)‑κB p65 and Fas protein ligand (L) in cancer cells was assessed using immunocytochemistry. The present results demonstrated that the combination of SAL and DDP significantly inhibited the proliferation (P<0.05) and altered the morphological characteristics of SGC‑7901 cells, thus suggesting that SAL may enhance the susceptibility of gastric cancer cells to DDP. In addition, treatment with a combination of SAL and DDP resulted in S phase‑arrest and increased the apoptotic rate of SGC‑7901 cells. Furthermore, marked FasL upregulation and NF‑κB p65 downregulation were observed in cancer cells treated with the combination of SAL and DDP. The results of the present study demonstrated that the combination of SAL and DDP induced the apoptosis of human gastric cancer cells, and suggested that the underlying mechanism may involve the upregulation of FasL and downregulation of NF‑κB p65.

Topics: Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Cell Shape; Cisplatin; Fas Ligand Protein; Humans; Pyrans; Stomach Neoplasms; Transcription Factor RelA

Cancer stem cells (CSCs) in gastric cancer (GC) have been established recently as key therapeutic targets for the successful treatment of GC. Emerging evidence suggests that both CSCs and cancer cells should be eradicated to achieve optimal therapeutic efficacy. In the present study, salinomycin, which has been reported to kill CSCs, was used in combination with docetaxel, a chemotherapeutic drug that is used as first-line therapy in GC, to eradicate both GC stem cells (SCs) and cancer cells. Salinomycin and docetaxel were loaded separately into poly(D,L-lactic-co-glycolic acid)-poly(ethylene glycol) nanoparticles of ∼140 nm with a narrow size distribution, high drug loading, and sustained drug release. GC SCs were isolated by magnetic-activated cell sorting on the basis of CD44 expression as the CSC phenotype. CD44 GC SCs showed the characteristics of CSCs, including increased SC gene expression, tumorsphere formation capacity, and tumorigenicity in nude mice. We found that both salinomycin and salinomycin-loaded nanoparticles (salinomycin-NPs) could selectively eradicate GC SCs, as reflected by reduced tumorsphere formation capacity and the frequency of CD44 GC cells, whereas docetaxel and docetaxel-loaded nanoparticles (docetaxel-NPs) could significantly eradicate GC cells. In nude mice bearing GC xenografts, salinomycin-NPs and salinomycin significantly decreased the intratumor population of GC SCs. Notably, salinomycin-NPs combined with docetaxel-NPs suppressed tumor growth more effectively than did salinomycin combined with docetaxel, single salinomycin-NPs, or docetaxel-NPs. Therefore, salinomycin-NPs combined with docetaxel-NPs represent a promising strategy for the treatment of GC by eradicating both GC SCs and cancer cells.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Cell Line, Tumor; Delayed-Action Preparations; Docetaxel; Female; Humans; Male; Mice; Mice, Inbred BALB C; Mice, Nude; Nanoparticles; Neoplastic Stem Cells; Polyethylene Glycols; Polyglactin 910; Pyrans; Stomach Neoplasms; Taxoids; Xenograft Model Antitumor Assays

Anti-angiogenesis targeting VEGFR2 has been an attractive strategy for cancer therapy for its role in promoting cancer growth and metastasis. However, the currently available drugs have unexpected side effects. Therefore, development of novel VEGFR2 inhibitors with less toxicity would be of great value. In this study, we describe a novel and safely VEGFR2 inhibitor, Salinomycin (Sal), which was screened from the drug libraries of Food and Drug Administration (FDA) and prohibited the binding of the ATP at its binding pocket of VEGFR2 using molecular docking model. Sal could interfere a series of VEGF-induced angiogenesis processes including proliferation, migration, and tube formation in HUVECS in vitro. Matrigel plug model demonstrated Sal strongly inhibited angiogenesis in vivo. We found that Sal significantly decreased VEGF-induced phosphorylation of VEGFR2 and its downstream STAT3 in dose- and time-dependent manner in HUVECs. Besides, Sal could directly reduce the cell viability and induce apoptosis in SGC-7901 cancer cells in vitro. Sal inhibited constitutive STAT3 activation by blocking its DNA binding and reduced various gene products including Bcl-2, Bcl-xL and VEGF both at mRNA and protein levels. Intra-peritoneal injection of Sal at doses of 3 and 5 mg/kg/day markedly suppressed human gastric cancer xenografts angiogenesis and growth without causing obvious toxicities. Taken together, Sal inhibits tumor angiogenesis and growth of gastric cancer; our results reveal unique characteristics of Sal as a promising anticancer drug candidate.

Topics: Angiogenesis Inhibitors; Animals; Antineoplastic Agents; Cell Movement; Cell Proliferation; Cell Survival; Human Umbilical Vein Endothelial Cells; Humans; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Nude; Neovascularization, Pathologic; Pyrans; Stomach Neoplasms; Vascular Endothelial Growth Factor Receptor-2; Xenograft Model Antitumor Assays

The Wnt1 protein, a secreted ligand that activates Wnt signaling pathways, contributes to the self-renewal of cancer stem cells (CSCs) and thus may be a major determinant of tumor progression and chemoresistance. In a series of gastric cancer specimens, we found strong correlations among Wnt1 expression, CD44 expression, and the grade of gastric cancer. Stable overexpression of Wnt1 increased AGS gastric cancer cells' proliferation rate and spheroids formation, which expressed CSC surface markers Oct4 and CD44. Subcutaneous injection of nude mice with Wnt1-overexpressing AGS cells resulted in larger tumors than injection of control AGS cells. Salinomycin, an antitumor agent, significantly reduced the volume of tumor caused by Wnt1-overexpressing AGS cells in vivo. This is achieved by inhibiting the proliferation of CD44+Oct4+ CSC subpopulation, at least partly through the suppression of Wnt1 and β-catenin expression. Taken together, activation of Wnt1 signaling accelerates the proliferation of gastric CSCs, whereas salinomycin acts to inhibit gastric tumor growth by suppressing Wnt signaling in CSCs. These results suggest that Wnt signaling might have a critical role in the self-renewal of gastric CSCs, and salinomycin targeting Wnt signaling may have important clinical applications in gastric cancer therapy.

Topics: Adult; Aged; Animals; beta Catenin; Cell Line, Tumor; Cell Proliferation; Female; Humans; Male; Mice; Mice, Nude; Middle Aged; Neoplastic Stem Cells; Pyrans; Signal Transduction; Stomach Neoplasms; Tumor Cells, Cultured; Wnt1 Protein

Gastric cancer stem cells (CSCs) play a crucial role in the initiation, development, relapse and metastasis of gastric cancer because they are resistant to a standard chemotherapy and the residual CSCs are able to proliferate indefinitely. Therefore, eradication of this cell population is a primary objective in gastric cancer therapy. Here, we report a gastric CSCs-specifically targeting drug delivery system (SAL-SWNT-CHI-HA complexes) based on chitosan(CHI) coated single wall carbon nanotubes (SWNTs) loaded with salinomycin (SAL) functionalized with hyaluronic acid (HA) can selectively eliminate gastric CSCs. Gastric CSCs were identified as CD44+ cells and cultured in serum-free medium. SAL-SWNT-CHI-HA complexes were capable of inhibiting the self-renewal capacity of CD44+ population, and decrease mammosphere- and colon-formation of CSCs. In addition, the migration and invasion of gastric CSCs were significantly blocked by SAL-SWNT-CHI-HA complexes. Quantitative and qualitative analysis of cellular uptake demonstrated that HA functionalization facilitated the uptake of SWNTs in gastric CSCs while free HA competitively inhibited cellular uptake of SAL-SWNT-CHI-HA delivery system, revealing the mechanism of CD44 receptor-mediated endocytosis. The SAL-SWNT-CHI-HA complexes showed the strongest antitumor efficacy in gastric CSCs by inducing apoptosis, and in CSCs mammospheres by penetrating deeply into the core. Taken altogether, our studies demonstrated that this gastric CSCs-targeted SAL-SWNT-CHI-HA complexes would provide a potential strategy to selectively target and efficiently eradicate gastric CSCs, which is promising to overcome the recurrence and metastasis of gastric cancer and improve gastric cancer treatment.

Topics: Apoptosis; Cell Line, Tumor; Cell Movement; Cell Proliferation; Drug Delivery Systems; Humans; Hyaluronic Acid; Microscopy, Confocal; Nanotubes, Carbon; Neoplastic Stem Cells; Pyrans; Stomach Neoplasms; Wound Healing

Salinomycin is a novel identified cancer stem cells (CSCs) killer. Higher ALDH activity represents CSCs characterization. Here, we screened ALDH activities on several gastric cancer cell lines and divided them into ALDH(high) and ALDH(low) gastric cancer groups. ALDH(high) cancer cells (NCI-N87 and SNU-1) disclosed more CSCs characteristics, such as higher levels of Sox2, Nanog and Nestin, more floating spheroid bodies, more colony formation and more resistance to conventional chemotherapeutic drugs 5-Fu and CDDP, compared to these parameters observed in ALDH(low) cancer cells (P<0.01). Importantly, ALDH(high) cancer cells are relatively sensitive to salinomycin when compared to ALDH(low) cancer cells (P<0.01). Our results confirmed ALDH as functional marker of CSCs population on gastric cancer. Salinomycin might be selective therapy for CSCs fraction, which is resistant to conventional anticancer drugs 5-Fu and CDDP.

Topics: Aldehyde Dehydrogenase; Aldehyde Dehydrogenase 1 Family; Antibiotics, Antineoplastic; Antimetabolites, Antineoplastic; Antineoplastic Agents, Alkylating; Cisplatin; Enzyme Induction; Fluorouracil; Gene Expression Regulation, Neoplastic; Homeodomain Proteins; Humans; Intermediate Filament Proteins; Nanog Homeobox Protein; Neoplasm Proteins; Neoplastic Stem Cells; Nerve Tissue Proteins; Nestin; Pyrans; Retinal Dehydrogenase; RNA, Messenger; RNA, Neoplasm; SOXB1 Transcription Factors; Spheroids, Cellular; Stomach Neoplasms; Tumor Cells, Cultured; Tumor Stem Cell Assay