Migration Of Human Cancer Cells Research Articles

G protein‑mediated EGFR transactivation is a common mechanism through which the CXCL12 receptors, CXCR4 and CXCR7, control human cancer cell migration.

C‑X‑C motif chemokine 12 (CXCL12) promotes metastasis of several tumors by affecting cell migration and invasion via its receptors, C‑X‑C chemokine receptor type (CXCR)4 and CXCR7. Current therapeutic approaches focus on the selective inactivation of either CXCR4 or CXCR7 in patients with cancer. Alternative strategies may emerge from the analysis of downstream events that mediate the migratory effects of CXCL12 in cancer cells. While CXCR4 activates cell signaling through both G proteins and arrestins, CXCR7 is believed to preferentially signal through arrestins. The present study analyzed the CXCL12‑dependent chemotaxis of A549, C33A, DLD‑1, MDA‑MB‑231 and PC‑3 cells, in which either the activity of G proteins, EGFR or Src kinase was inhibited pharmacologically or the expression of arrestins was inhibited by RNA interference. The results demonstrated that CXCL12‑induced migration of A549, C33A, DLD‑1, MDA‑MB‑231 and PC‑3 cells was attenuated by the Gαi/o‑inhibitor pertussis toxin (PTX), but was unaffected by small interfering RNA‑mediated gene silencing of β‑arrestin1/2. In particular, the sensitivity of DLD‑1 migration to PTX was unexpected, as it is solely dependent on the non‑classical chemokine receptor, CXCR7. Furthermore, chemotactic responses to CXCL12 were additionally prevented by inhibiting EGFR activity via AG1478 and Src kinase activity via Src inhibitor‑1. In conclusion, the results of the present study suggest that G protein‑ and Src‑dependent transactivation of EGFR is a common mechanism through which CXCL12‑bound CXCR4 and/or CXCR7 control cancer cell migration and metastasis. These findings highlight EGFR as a potential therapeutic target that interferes with CXCL12‑induced cancer expansion.

Contribution of the α5 nAChR Subunit and α5SNP to Nicotine-Induced Proliferation and Migration of Human Cancer Cells.

Nicotine in tobacco is known to induce tumor-promoting effects and cause chemotherapy resistance through the activation of nicotinic acetylcholine receptors (nAChRs). Many studies have associated the α5 nicotinic receptor subunit (α5), and a specific polymorphism in this subunit, with (i) nicotine administration, (ii) nicotine dependence, and (iii) lung cancer. The α5 gene CHRNA5 mRNA is upregulated in several types of cancer, including lung, prostate, colorectal, and stomach cancer, and cancer severity is correlated with smoking. In this study, we investigate the contribution of α5 in the nicotine-induced cancer hallmark functions proliferation and migration, in breast, colon, and prostate cancer cells. Nine human cell lines from different origins were used to determine nAChR subunit expression levels. Then, selected breast (MCF7), colon (SW480), and prostate (DU145) cancer cell lines were used to investigate the nicotine-induced effects mediated by α5. Using pharmacological and siRNA-based experiments, we show that α5 is essential for nicotine-induced proliferation and migration. Additionally, upon downregulation of α5, nicotine-promoted expression of EMT markers and immune regulatory proteins was impaired. Moreover, the α5 polymorphism D398N (α5SNP) caused a basal increase in proliferation and migration in the DU145 cell line, and the effect was mediated through G-protein signaling. Taken together, our results indicate that nicotine-induced cancer cell proliferation and migration are mediated via α5, adding to the characterization of α5 as a putative therapeutical target.

Vasoactive intestinal peptide-VIPR2 signaling regulates tumor cell migration.

Phosphoinositide metabolism is critically involved in human cancer cell migration and metastatic growth. The formation of lamellipodia at the leading edge of migrating cells is regulated by metabolism of the inositol phospholipid PI(4,5)P2 into PI(3,4,5)P3. The synthesized PI(3,4,5)P3 promotes the translocation of WASP family verprolin homologous protein 2 (WAVE2) to the plasma membrane and regulates guanine nucleotide exchange factor Rac-mediated actin filament remodeling. Here, we investigated if VIPR2, a receptor for vasoactive intestinal peptide (VIP), has a potential role in regulating cell migration via this pathway. We found that silencing of VIPR2 in MDA-MB-231 and MCF-7 human breast cancer cells inhibited VIP-induced cell migration. In contrast, stable expression of exogenous VIPR2 promoted VIP-induced tumor cell migration, an effect that was inhibited by the addition of a PI3-kinase (PI3K)γ inhibitor or a VIPR2-selective antagonist. VIPR2 stably-expressing cells exhibited increased PI3K activity. Membrane localization of PI(3,4,5)P3 was significantly attenuated by VIPR2-silencing. VIPR2-silencing in MDA-MB-231 cells suppressed lamellipodium extension; in VIPR2-overexpressing cells, VIPR2 accumulated in the cell membrane on lamellipodia and co-localized with WAVE2. Conversely, VIPR2-silencing reduced WAVE2 level on the cell membrane and inhibited the interaction between WAVE2, actin-related protein 3, and actin. These findings suggest that VIP–VIPR2 signaling controls cancer migration by regulating WAVE2-mediated actin nucleation and elongation for lamellipodium formation through the synthesis of PI(3,4,5)P3.

Abstract 5125: Biological and clinical value of OCT4A/SPP1C axis in endometrial and lung adenocarcinoma

Abstract Background: It is largely unclear whether octamer-binding transcription factor 4A (OCT4A, OCT3/4A) functions physiologically or pathologically in adult somatic tissues and related cancers. We have recently identified OCT4A translation in a human cancer cell subpopulation that might exacerbate cell migration and invasion. Here, we focus on biological and clinical features of OCT4A and its relationship with secreted phosphoprotein 1(SPP1), which is strongly related to the malignant progression of numerous tumors. Methods: RT-PCR or qPCR analysis was used to identify full-length spliced forms of OCT4A and SPP1 transcripts in non-tumor and tumor human cells. An OCT4A-FLAG-tagged genomic transgene identified OCT4A-positive cancer cells. Cell-ablation of OCT4A-positive cells using promoter-driven diphtheria toxin A was used to examine the potential role of OCT4A in lung and endometrial adenocarcinoma cells. OCT4A and SPP1 transcripts in early-stage lung adenocarcinoma (stage I, fifty-eight cases) were analyzed and compared with pathohistological and prognostic features. Results: We found that OCT4A and SPP1C are co-expressed in highly aggressive human lung and endometrial adenocarcinoma cell lines but not in mesothelioma or non-tumor cell lines. Ablation of OCT4A-positive cells significantly decreased cancer-cell migration and SPP1C mRNA levels. The OCT4A/SPP1C axis was found in primary, early-stage, lung adenocarcinomas, and all four cases of recurrence and/or metastasis out of 58 cases were included in the OCT4A and SPP1C positive group. Conclusions: Co-expression of OCT4A and SPP1C may correlate with human cancer cell migration and the OCT4A/SPP1C axis may have a prognostic value in high-risk, early-stage lung adenocarcinomas.

Leaf Extract and Active Fractions of Dillenia pentagyna Roxb. Reduce In Vitro Human Cancer Cell Migration Via NF-κB Pathway.

Different parts of Dillenia pentagyna have long been used in traditional medicines to cure several diseases including cancer. However, the mechanism(s) of anti-cancer effects are still unknown. We aimed to elucidate the anti-metastatic potential of ethanolic extracts of leaves of D. pentagyna (EELDP) and active fractions of it in highly metastatic human cancer cells. We screened different HPLC fractions of EELDP based on their anti-metastatic effect. We used TLC and ESI-MS for determining the presence of various phytochemicals in EELDP and fractions. We monitored in vitro anti-metastasis effect of EELDP (0-0.6 mg/ml) and active fractions (0-0.050 mg/ml) on various human cancer cells like A549, HeLa, and U2OS. EELDP significantly reduced cell viability and cell migration in A549, HeLa, and U2OS cells. However, higher sensitivity was observed in A549 cells. We screened 2 active HPLC fractions F6 and F8 having anti-MMPs activity. EELDP and active fractions reduced metastasis via the NF-κB pathway, decreased the expression of Vimentin, N-cadherin, and increased the expression of Claudin-1. Significant reduction of metastasis by EELDP at a dose of 0.1 mg/ml or by active fractions at 0.050 mg/ml implicates that the active compound(s) present in crude or fractions are extremely potent to control highly metastatic cancer.

Enhanced OXER1 expression is indispensable for human cancer cell migration

OXER1 is a recently identified receptor, binding the arachidonic acid metabolic product 5-oxo-ETE, considered an inflammatory receptor, implicated in chemoattraction of circulating mononuclear cells, Ca2+ surge in neutrophils, inflammation and cancer. Recently, we have shown that OXER1 is also a membrane androgen receptor in various cancer tissues. It was reported that the presence of OXER1 in leucocytes and the production and release of 5-oxo-ETE by wounded tissues is a wound sensing mechanism, leading to lymphocyte attraction. In view of the similarity of hallmarks of cancer and wound healing, we have explored the role of OXER1 and its endogenous ligand in the control of cell migration of human cancer epithelial cells (DU-145, T47D and Hep3B), mimicking the activation/migration phase of healing. We show that OXER1 is up-regulated only at the leading edge of the wound and its expression is up-regulated by its ligand 5-oxo-ETE, in a time-related manner. Knock-down of OXER1 or inhibition of 5-oxo-ETE synthesis led to decreased migration of cells and a prolongation of healing, in culture prostate cancer cell monolayers, with a substantial modification of actin cytoskeleton and a decreased filopodia formation. Inhibition of cell migration is a phenomenon mediated by Gβγ OXER1 mediated actions. These results provide a novel mechanism of OXER1 implication in cancer progression and might be of value for the design of novel OXER1 antagonists.

Lucidumones B-H, racemic meroterpenoids that inhibit tumor cell migration from Ganoderma lucidum

Seven new meroterpenoids, lucidumones B-H (1 and 3–8), along with one known meroterpenoid (2), were isolated from the fruiting bodies of Ganoderma lucidum. The structures of the new compounds were assigned by spectroscopic and computational methods. All the isolated compounds were tested for their inhibition on human cancer cell migration. It was found that compounds (–)-1, (+)-2, (–)-4, (+)-6, and (+)-8 could significantly inhibit cell migration in KYSE30 cell line. Further examination disclosed that cell migration inhibition of (+)-6 and (+)-8 might be related with downregulation of N-cadherin.

Heat shock protein 70 positively regulates transforming growth factor-α-induced hepatocellular carcinoma cell migration via the AKT signaling pathway

Heat shock proteins (HSPs) are induced in response to extracellular stress and manage the quality of proteins as molecular chaperones. HSP70, a highly conserved HSP, has been reported to correlate with the proliferation and migration of human cancer cells, such as oral, prostate, lung and liver cancer. Regarding hepatocellular carcinoma (HCC), the HSP70 levels in the tumor tissues from patients are significantly higher than those in the normal liver tissues. HSP70 reportedly upregulates the migration and invasion of HCC. The AKT, p38 mitogen-activated protein kinase (MAPK), c-jun N-terminal kinase (JNK) and Rho-kinase signaling pathways regulate the transforming growth factor (TGF)-α-induced migration of human HCC-derived HuH7 cells. However, the exact mechanism underlying the role of HSP70 in growth factor-induced HCC migration remains unclear. Therefore, in the present study, the mechanism underlying the involvement of HSP70 in TGF-α-induced HCC cell migration was investigated. Treatment with the HSP70 inhibitors VER155008 and YM-08 and the downregulation of HSP70 protein were confirmed to significantly suppress the TGF-α-induced cell migration of HuH7 cells. Both VER155008 and YM-08 reduced the TGF-α-induced phosphorylation of AKT without affecting the phosphorylation of p38 MAPK, JNK or Rho-kinase. These results strongly suggest that HSP70 positively regulates the TGF-α-induced migration of HCC cells via the AKT signaling pathway.

A novel trifluoromethyl 2-phosphonopyrrole analogue inhibits human cancer cell migration and growth by cell cycle arrest at G1 phase and apoptosis

Pyrroles, an important class of heterocyclic compounds found in naturally occurring products, represent an interesting biologically active scaffold for drug design. Recently we have synthetized a series of five new fluorinated pyrrole derivatives for potential anticancer applications. All new compounds contain a trifluoromethyl and N-benzyl group, but they are different from each other by bearing a phenyl, ethoxycarbonyl or carboxylic moiety, with two of them possessing an additional phosphonyl function.The aim of this study was to evaluate anticancer activity of the new compounds in human lung and breast cancer cells. We found that compound 3, bearing a phosphonyl and phenyl group, was the most effective in attenuating growth of A549 and MCF-7 cells in a dose dependent manner with IC50 36.5 μM ± 1.80 and 27.9 μM ± 1.68, respectively. Compound 3 inhibited cancer cell proliferation by cell cycle arrest at G1 phase as detected by flow cytometry analysis. Furthermore, compound 3 induced apoptosis of A549 cells by activation of caspase-3. Cancer cell migration rate was significantly inhibited when A549 and MCF-7 cells were cultured in the presence of the compound. These results demonstrate that a novel trifluoromethyl-functionalized phosphonopyrrole with a phenyl group might be a promising pyrrole analogue with anticancer potential.

Comparison of Chitosan Nanoparticles and Soluplus Micelles to Optimize the Bioactivity of Posidonia oceanica Extract on Human Neuroblastoma Cell Migration.

Posidonia oceanica (L.) Delile is a marine plant endemic of Mediterranean Sea endowed with interesting bioactivities. The hydroalcholic extract of P. oceanica leaves (POE), rich in polyphenols and carbohydrates, has been shown to inhibit human cancer cell migration. Neuroblastoma is a common childhood extracranial solid tumor with high rate of invasiveness. Novel therapeutics loaded into nanocarriers may be used to target the migratory and metastatic ability of neuroblastoma. Our goal was to improve both the aqueous solubility of POE and its inhibitory effect on cancer cell migration. Methods: Chitosan nanoparticles (NP) and Soluplus polymeric micelles (PM) loaded with POE have been developed. Nanoformulations were chemically and physically defined and characterized. In vitro release studies were also performed. Finally, the inhibitory effect of both nanoformulations was tested on SH-SY5Y cell migration by wound healing assay and compared to that of unformulated POE. Results: Both nanoformulations showed excellent physical and chemical stability during storage, and enhanced the solubility of POE. PM-POE improved the inhibitory effect of POE on cell migration probably due to the high encapsulation efficiency and the prolonged release of the extract. Conclusions: For the first time, a phytocomplex of marine origin, i.e., P. oceanica extract, has enhanced in terms of acqueous solubility and bioactivity once encapsulated inside nanomicelles.

Effect of silencing phosphatidylinositol 3-kinase regulatory subunit gamma gene by small interfering RNA on proliferation and migration capability of gallbladder carcinoma cells

Objective To observe the effect of small interfering RNA (siRNA) silencing phosphatidylinositol 3-kinase regulatory subunit gamma (PIK3R3) gene on the proliferation and migration of gallbladder cancer cells NOZ and GBC-SD. Methods Small interfering RNA (si-PIK3R3) against PIK3R3 was constructed and real-time quantitative polymerase chain reaction (Real-time PCR) and Western blotting were used to detect PIK3R3 mRNA and protein in transfected gallbladder cancer cells NOZ and GBC-SD. The expression of cell counting kit-8 (CCK-8) proliferation assay was used to detect the proliferation of transfected 0-5 d cells. Transwell chamber assay was used to detect the migration of gallbladder cancer cells 48h after transfection. Results The si-PIK3R3 transfection group and the si-NC negative control group were successfully transfected into the NOZ and GBC-SD of gallbladder cancer cells. The expression of PIK3R3 mRNA (0.119±0.447) in the NOZ cell si-PIK3R3 group was significantly lower than that of NC group (1.000±0.398) (t=165.100, P<0.05), the PIK3R3 mRNA (0.085±0.593) in the si-PIK3R3 group of GBC-SD cells was significantly lower than that in the si-NC group (1.000±0.669) (t=500.200, P<0.05); the expression of PIK3R3 protein in the transfected group was significantly lower than that in the negative control group. The results of CCK-8 in NOZ cells showed that the absorbance values of 0-5 d si-PIK3R3 transfection group were 0.185±0.007, 0.163±0.013, 0.229±0.019, 0.240±0.017, 0.273±0.012, 0.350±0.069 (t=11.960, P<0.05); The results of 48 h Transwell showed that the number of cells migrated in the si-PIK3R3 transfection group [(69.0±4.5) cells] was significantly lower than that in the si-NC group (t=25.120, P<0.05). The results of CCK-8 in GBC-SD cells showed that the absorbance values of the 0-5 d si-PIK3R3 transfection group were 0.153±0.003, 0.158±0.002, 0.201±0.010, 0.240±0.012, 0.297±0.018, 0.483±0.024 (t=14.160, P<0.05); 48 h Transwell results showed that the number of cells migrated in the si-PIK3R3 transfection group [(91.0±3.5) cells] was significantly lower than that in the si-NC group (t=25.390, P<0.05). Conclusion si-PIK3R3 can inhibit the proliferation and migration of human gallbladder cancer cells, suggesting that PIK3R3 plays an important role in the development of gallbladder carcinoma. Key words: Gallbladder cancer; Small interfering RNA; Phosphatidylinositol 3-kinase regulatory subunit gamma

Effect of macrophage capping protein expression on proliferation and migration of prostate cancer cells

Objective To investigate the expression and explore the biological function of macrophage capping protein (CapG) in prostate cancer (PCa). Methods Immunohistochemistry was used to detect the expression of CapG in 115 cases of prostate cancer and 42 cases of adjacent normal prostate in the Second Affiliated Hospital of Wenzhou Medical University from January 2013 to June 2018. The correlation between CapG expression and clinicopathological features in PCa was evaluated by statistical methods. The expression of CapG in 3 prostate cancer tissues was detected by real-time quantitative reverse transcriptase-polymerase chain reaction (RT-qPCR) and Western blotting. CapG expression was knocked down by small interfering RNA (siRNA), and the effect of CapG on proliferation and migration of human prostate cancer cells was detected by methyl thiazol tetrazolium (MTT) (5 g/L) and Transwell chamber experiments. Results The positive rates of CapG expression in prostate cancer and adjacent normal prostate tissues were 55.6% and 16.7%, respectively, and the difference was statistically significant (χ2=18.879, P<0.05); CapG expression was significantly correlated with prostate specific antigen (PSA) level, Gleason score, T stage and lymph node metastasis of prostate cancer (P<0.05). Down-regulation of CapG significantly inhibited the proliferation and migration of PC3 and DU145 cells (P<0.05). Conclusion CapG is highly expressed in prostate cancer and affects the proliferation and migration of prostate cancer cells. Key words: Prostate cancer; Macrophage capping protein; Proliferation; Metastasis

Effects of neddylation modification inhibitor MLN4924 on proliferation and migration of human gastric cancer cells

Objective To explore the effects of NEDD8 activating enzyme (NAE) inhibitor MLN4924 on gastric cancer cell proliferation and migration. Methods Human gastric cancer cell lines NCI-N87 and SGC-7901 were treated with different concentrations of MLN4924, and the inhibitory concentrations were detected by cell counting kit-8 (CCK-8) method. 20% maximal inhibitory concentration (IC20) was used to treat gastric cancer cells. The cell proliferation was measured by CCK-8 and clone formation assay, and cell migration was determined by cell scratch test. Results When human gastric cancer cell lines NCI-N87 and SGC-7901 were treated with 0.528 and 0.375 μmol/L respectively, the inhibitory rate reached to 20%. CCK-8 method showed that MLN4924 inhibited the proliferation of human gastric cells in a dose- and time-dependent manner. In the clone formation assay, the clone number of NCI-N87 cells was 71.00±10.19 in MLN4924 treated group and 215.33±9.56 in the control group. The clone number of SGC-7901 cells was 23.66±18.66 in MLN4924-treated group, and 32.33±34.12 in the control group.In vitro wound scratch assay revealed that the healing rate of MLN4924-treated NCI-N87 cells was (5.30±0.78)%, and (51.05±6.35)% in the control group, and that of MLN4924-treated SGC-7901 cells was (3.12±0.52)%, and (23.05±2.01)% in the control group. Conclusion Neddylation modification inhibitor MLN4924 suppresses the growth and migration of gastric cancer cells. Key words: Gastric cancer; Neddylation modification inhibitor; Proliferation; Migration

Down regulated Linc00152 expression affects proliferation and migration of human gastric cancer cells

Down regulated Linc00152 expression affects proliferation and migration of human gastric cancer cells

Inhibitory effect of interleukin -11 receptor α-targeted 131I-CGRRAGGSC on human breast cancer cells

Objective To establish interleukin -11 receptor α (IL-11Rα)-based 131I-CGRRAGGSC by chloramines-T method, and explore the inhibitory effects of 131I-CGRRAGGSC on human breast cancer cells overexpressing IL-11Rα. Methods The tyrosine modified CGRRAGGSC protein was labeled with 131I by chloramines-T method. Methyl thiazol tetrazolium (MTT), Transwell and wound-healing assay were used to detect the effects of 131I-CGRRAGGSC on the migration and proliferation of human breast cancer cells. Western blotting was used to detect the protein expression of IL-11Rα, phosphorylated signal transducer and activators of transcription 3 (p-STAT3) and signal transducer and activators of transcription 3 (STAT3) in breast cancer cells after treatment with 0, 2.775 and 5.550 MBq/ml 131I-CGRRAGGSC. Results 131I-CGRRAGGSC was radiolabeled successfully and radiochemical purity after purification was over 95%. The inhibition ratio of breast cancer cells (MDA-MB-231 and MCF-7) was (21.46±1.08)%, (37.95±2.59)%, (49.18±0.77)%, (60.34±0.19)% (F=389.4, P=0.000) and (27.59±2.15)%, (41.67±0.92)%, (54.53±1.07)%, (64.84±1.69)% (F=328.6, P=0.000) after 131I-CGRRAGGSC treatment (4.625, 9.250, 18.500 and 37.000 MBq/ml, respectively). Transwell demonstrated that the migration ratios of 2.775, 5.55 MBq/ml 131I-CGRRAGGSC were (32.05±4.44)%, (45.92±2.55)% (t=4.688, P=0.009) for MDA-MB-231 cells and (21.99±1.18)%, (39.45±1.36)% (t=16.760, P=0.000) for MCF-7 cells, respectively. Wound-healing assay also suggested that 131I-CGRRAGGSC inhibited MDA-MB-231 cells, but not MCF-7 cells. The cellular IL-11Rα and p-STAT3 levels were decreased in a dose-dependent manner whereas the STAT3 level had no significant change in 131I-CGRRAGGSC-treated breast cancer cells after 24 h in compared to those in the control cells. Conclusion 131I-CGRRAGGSC could inhibit the migration and proliferation of human breast cancer cells by down-regulating the IL-11Rα to inhibit STAT3 activation. Key words: Breast cancer; Radiolabeling; Migration; 131I

EB1 and cytoplasmic dynein mediate protrusion dynamics for efficient 3-dimensional cell migration.

Microtubules have long been implicated to play an integral role in metastatic disease, for which a critical step is the local invasion of tumor cells into the 3-dimensional (3D) collagen-rich stromal matrix. Here we show that cell migration of human cancer cells uses the dynamic formation of highly branched protrusions that are composed of a microtubule core surrounded by cortical actin, a cytoskeletal organization that is absent in cells on 2-dimensional (2D) substrates. Microtubule plus-end tracking protein End-binding 1 and motor protein dynein subunits light intermediate chain 2 and heavy chain 1, which do not regulate 2D migration, critically modulate 3D migration by affecting RhoA and thus regulate protrusion branching through differential assembly dynamics of microtubules. An important consequence of this observation is that the commonly used cancer drug paclitaxel is 100-fold more effective at blocking migration in a 3D matrix than on a 2D matrix. This work reveals the central role that microtubule dynamics plays in powering cell migration in a more pathologically relevant setting and suggests further testing of therapeutics targeting microtubules to mitigate migration.—Jayatilaka, H., Giri, A., Karl, M., Aifuwa, I., Trenton, N. J., Phillip, J. M., Khatau, S., Wirtz, D. EB1 and cytoplasmic dynein mediate protrusion dynamics for efficient 3-dimensional cell migration.

Suppression of cell migration by phospholipase C-related catalytically inactive protein-dependent modulation of PI3K signalling

The metabolic processes of phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] into PI(3,4,5)P3 and the subsequent PI(3,4,5)P3 signalling are involved in cell migration. Dysfunctions in the control of this pathway can cause human cancer cell migration and metastatic growth. Here we investigated whether phospholipase C-related catalytically inactive protein (PRIP), a PI(4,5)P2-binding protein, regulates cancer cell migration. PRIP overexpression in MCF-7 and BT-549 human breast cancer cells inhibited cell migration in vitro and metastasis development in vivo. Overexpression of the PRIP pleckstrin homology domain, a PI(4,5)P2 binding motif, in MCF-7 cells caused significant suppression of cell migration. Consistent with these results, in comparison with wild-type cells, Prip-deficient mouse embryonic fibroblasts exhibited increased cell migration, and this was significantly attenuated upon transfection with a siRNA targeting p110α, a catalytic subunit of class I phosphoinositide 3-kinases (PI3Ks). PI(3,4,5)P3 production was decreased in Prip-overexpressing MCF-7 and BT-549 cells. PI3K binding to PI(4,5)P2 was significantly inhibited by recombinant PRIP in vitro, and thus the activity of PI3K was downregulated. Collectively, PRIP regulates the production of PI(3,4,5)P3 from PI(4,5)P2 by PI3K, and the suppressor activity of PRIP in PI(4,5)P2 metabolism regulates the tumour migration, suggesting PRIP as a promising target for protection against metastatic progression.

Effects of upregulatioon of Forkhead box protein M1 on migration, invasion and epithelial-mesenchymal transition of human ovarian cancer cell

Objective To explore the effects and mechanism of over-expression of Forkhead box protein M1 (FoxM1) gene on the migration, and invasion and epithelial-mesenchymal transition (EMT) of human ovarian cancer cells. Methods FoxM1 over-expression of the virus vector was constructed. All HO-8910 cells were divided into three groups, the control group (Con-A), Lentiviral vector(LV) control group (Con-B) and over-expression group FoxM1. After three group HO-8910 cells was transfected with phosphate buffer (PBS), Lentiviral vector (LV), and LV-FoxM1, the protein levels of FoxM1 were determined by Western blotting, respectively. The migration and invasion ability was evaluated by wound healing assay and Transwell assay, respectively. The E-Cadherin and N-Cadherin of protein of cancer cells was examined by Western blotting assay. Results The results from Western blotting assay showed that the levels protein of FoxM1 were greatly increased in HO-8910 cancer cells transfected with LV-FoxM1. The results of the wound healing test showed that the migration distance of Con-A, Con-B, and LV-FoxM1 group was(156.80±2.26), (161.60±1.81) and (278.60±2.62) μm, respectively (P<0.01); The results of the Transwell invasion assay showed that the membrane cell number of Con-A, Con-B, and LV-FoxM1 group was (68.56±1.69) and (67.27±1.36) and (126.38±1.56), respectively (P<0.01); Compared with the Con-A, there were E-cadherin protein expression decreased; however, N-cadherin protein increased in HO-8910 cancer cells transfected with LV-FoxM1. Conclusion FoxM1 overexpression can promote human ovarian cancer cell migration and invasion, and its mechanism may be related to the promotion of epithelial-mesenchymal transition. Key words: Ovarian carcinoma; Forkhead box protein M1; Migration; Invasion; Epithelial-mesenchymal transition

CLC-3 Cl- channel-mediated invasion and migration of human ovarian cancer cells.

To investigate the potential role of CLC-3, a member of the voltage-gated chloride channel (CLC) superfamily, in invasion and migration of ovarian cancer cell line SKOV3. CLC-3 antisense oligonucleotides were transfected into ovarian cancer cell line SKOV3, and its effects on cell invasion and migration were analyzed by using Transwell chamber assay and wound healing assay in vitro. The efficiency of CLC-3 antisense was determined with RT-PCR and Western blotting. The protein concentrations of matrix metalloproteinase (MMP)-2, MMP-9, and vascular endothelial growth factor (VEGF) were determined using ELISA kits. Cell volume measurements were performed. Studies in vitro revealed that the CLC-3 antisense inhibited invasion and migration of ovarian cancer cell line SKOV3. CLC-3 antisense treatment decreased protein levels of MMP-2, MMP-9, and VEGF in culture medium of SKOV3 cells. In addition, the authors found that the capability for regulatory volume decrease (RVD) was much attenuated in SKOV3 cells transfected with CLC-3 antisense. These results strongly suggest that CLC-3 may get involved in proliferation, invasion, and migration of ovarian cancer cells and thus may be a useful therapeutic target.

Abstract LB-058: Linker variation and structure-activity relationship analyses of salicylic, benzoic and hydroxamic acid-based small molecule Stat3 inhibitors

Abstract Benzoic acid, salicylic acid and hydroxamic acid-based small molecules have been reported that inhibit Stat3 activation. However, the structural basis for their Stat3-inhibitory activities are not clearly defined. To investigate the molecular determinants for activity and to derive structurally optimized analogs of improved pharmacokinetic parameters and potency, we conducted a computer-aided molecule optimization program and synthesized new analogs of the previously reported lead Stat3 inhibitors, BP-1-102 (IC50, 6.8 ± 2.5), SH4-54 (IC50 4.4 ± 0.3), and SH5-07 (IC50 3.9 ± 0.6). All three leads are based on N-methylglycinamide scaffold with its two amine moieties condensed with three different functionalities. The three functionalities or the CH2 group of the glycolamide scaffold were separately modified to generate a series of compounds, some of which show improved potency compared to the corresponding parental compounds. Specifically, H012, H070 and H072 had improved inhibitory activity against Stat3 DNA-binding activity in vitro, with IC50 of 3.0 ± 0.9, 2.4 ± 0.4, and 2.2 ± 0.9 μM, respectively. The replacement of the pentafluorobenzene or the cyclohexylbenzene with other components was detrimental to activity and led to decreased potency of the new analogues, as did the substitution of the benzene of the pTyr-mimetic moiety with heterocyclic components containing nitrogen and oxygen elements. Pharmacokinetic study showed that analog H058 has improved permeability in Caco-2 bidirectional permeability studies, compared to the lower relative permeability of the lead agents. Ala-based analogs, such as H048 and H058, were low in microsomal stability, presumably due to the labile methyl groups of the Ala. Select active analogs inhibited constitutive Stat3 activation and the DNA-binding activity of Stat3 in cancer cells, and they further decreased the growth, colony formation, and migration of human cancer cells harboring persistently active Stat3. Studies together demonstrate a significant progress towards developing a more suitable Stat3 inhibitor with drug-like features. Additional efforts are focused on refining the physicochemical properties to further improve the activity and efficacy and ultimately furnish suitable Stat3-inhibiting candidates for clinical application. Citation Format: Peibin Yue, Francisco Lopez-Tapia, Christine Brotherton-Pleiss, Marcus Tius, James Turkson. Linker variation and structure-activity relationship analyses of salicylic, benzoic and hydroxamic acid-based small molecule Stat3 inhibitors. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr LB-058.