Radiosensitizing effect of gold nanoparticle loaded with small interfering RNA-SP1 on lung cancer: AuNPs-si-SP1 regulates GZMB for radiosensitivity

Radioresistance is a major challenge that largely limits the efficacy of radiotherapy in lung cancer. Gold nanoparticles (AuNPs) are emerging as novel radiosensitizers for cancer patients. Therefore, this study was designed to explore the radiosensitizing effect and mechanism of AuNPs loaded with small interfering RNA (siRNA)-SP1 (AuNPs-si-SP1) on lung cancer. AuNPs-si-SP1 was prepared by the noncovalent binding between AuNPs and siRNA-SP1. The adsorption capacity of AuNPs to siRNA-SP1 was analyzed by gel electrophoresis. The cell uptake of AuNPs-si-SP1 was observed under a laser confocal microscopy. Silencing efficacy of AuNPs-si-SP1 was validated by RT-qPCR and Western blot analysis. Cell viability was determined by CCK-8 assay, radiosensitization by plate colony formation assay, cell apoptosis and cell cycle by flow cytometry, and DNA double strand breaks by immunofluorescence in the presence or absence of AuNPs-si-SP1 or GZMB. The downstream mechanism of SP1 was predicted by bioinformatics analysis, followed by verification by Western blot analysis. Subcutaneous tumorigenesis in nude mice was established to verify the radiosensitization of AuNPs-si-SP1 and GZMB in vivo. AuNPs-si-SP1 effectively absorbed SP1 siRNA and was highly internalized by A549 cells to reduce SP1 protein expression. AuNPs-si-SP1 or GZMB overexpression promoted cells to G2/M phase, DNA double strand breaks, and enhanced radiosensitivity. SP1 could repress GZMB expression in lung cancer cells. In vivo experiments manifested that AuNPs-si-SP1 could inhibit the growth of solid tumor in nude mice to achieve radiosensitization by inhibiting SP1 to upregulate GZMB. AuNPs-si-SP1 might increase the radiosensitivity of lung cancer by inhibiting SP1 to upregulate GZMB.     

Quantitative analyses of amount and localization of radiosensitizer gold nanoparticles interacting with cancer cells to optimize radiation therapy

Previous studies showed that gold nanoparticles (AuNPs) are useful radiosensitizers which optimize radiation therapy under low-dose radiation. However, the mechanisms of AuNP radiosensitization, including the amount and localization of the AuNPs interacting with cancer cells, has not yet been quantified. To answer these questions, we prepared AuNPs conjugated with anti-human epidermal growth factor receptor type 2 (HER2) antibody via polyethylene glycol (PEG) chains (AuNP-PEG-HER2ab). AuNP-PEG-HER2ab specifically bound to the HER2-expressing cancer cells and entered the cells via endocytosis. Whether endocytosis of AuNP-PEG-HER2ab occurred had no effect on radiosensitization efficacy by AuNP-PEG-HER2ab in vitro. The radiosensitization efficacy in vitro depended on dose of AuNP-PEG-HER2ab or dose of X-ray. Moreover, AuNP-PEG-HER2ab administrated into tumor-bearing mice was localized to both the periphery of the tumor tissue and near the nuclei in cancer cells in tumor deep tissue. The localization of AuNP-PEG-HER2ab in tumor tissues was important factors for in vivo powerful radiosensitization efficacy.     

miR‐200c enhances radiosensitivity of human breast cancer cells

Due to the intrinsic resistance of many tumors to radiotherapy, current methods to improve the survival of cancer patients largely depend on increasing tumor radiosensitivity. It is well‐known that miR‐200c inhibits epithelial–mesenchymal transition (EMT), and enhances cancer cell chemosensitivity. We sought to clarify the effects of miR‐200c on the radiosensitization of human breast cancer cells. In this study, we found that low levels of miR‐200c expression correlated with radiotolerance in breast cancer cells. miR‐200c overexpression could increase radiosensitivity in breast cancer cells by inhibiting cell proliferation, and by increasing apoptosis and DNA double‐strand breaks. Additionally, we found that miR‐200c directly targeted TANK‐binding kinase 1 (TBK1). However, overexpression of TBK1 partially rescued miR‐200c mediated apoptosis induced by ionizing radiation. In summary, miR‐200c can be a potential target for enhancing the effect of radiation treatment on breast cancer cells. J. Cell. Biochem. 114: 606–615, 2013. © 2012 Wiley Periodicals, Inc.     

Suppression of type 1 Insulin-like growth factor receptor expression by small interfering RNA inhibits A549 human lung cancer cell invasion in vitro and metastasis in xenograft nude mice

Cancer invasion and metastasis, involving a variety of pathological processes andcytophysiological changes,contribute to the high mortality of lung cancer.The type 1 insulin-like growthfactor receptor (IGF-1R),associated with cancer progression and invasion,is a potential anti-invasion andanti-metastasis target in lung cancer.To inhibit the invasive properties of lung cancer cells,we successfullydown-regulated IGF-1R gene expression in A549 human lung cancer cells by small interfering RNA (siRNA)technology,and evaluated its effects on invasion-related gene expression,tumor cell in vitro invasion,andmetastasis in xenograft nude mice.A549 cells transfected with a plasmid expressing hairpin siRNA forIGF-1R showed a significantly decreased IGF-1R expression at the mRNA level as well as the proteinlevel.In biological assays,transfected A549 cells showed a significant reduction of cell-matrix adhesion,migration and invasion.Consistent with these results,we found that down-regulation of IGR-1Rconcomitantly accompanied by a large reduction in invasion-related gene expressions,including MMP-2,MMP-9,u-PA,and IGF-1R specific downstream p-Akt.Direct tail vein injections of plasmid expressinghairpin siRNA for IGF- 1R significantly inhibited the formation of lung metastases in nude mice.Our resultsshowed the therapeutic potential of siRNA as a method for gene therapy in inhibiting lung cancer invasionand metastasis.     

TUSC1, a Putative Tumor Suppressor Gene,Reduces Tumor Cell Growth In Vitro and Tumor Growth In Vivo

We previously reported the identification of TUSC1 (Tumor Suppressor Candidate 1), as a novel intronless gene isolated from a region of homozygous deletion at D9S126 on chromosome 9p in human lung cancer. In this study, we examine the differential expression of TUSC1 in human lung cancer cell lines by western blot and in a primary human lung cancer tissue microarray by immunohistochemical analysis. We also tested the functional activities and mechanisms of TUSC1 as a tumor suppressor gene through growth suppression in vitro and in vivo. The results showed no expression of TUSC1 in TUSC1 homozygously deleted cells and diminished expression in some tumor cell lines without TUSC1 deletion. Interestingly, the results from a primary human lung cancer tissue microarray suggested that higher expression of TUSC1 was correlated with increased survival times for lung cancer patients. Our data demonstrated that growth curves of tumor cell lines transfected with TUSC1 grew slower in vitro than those transfected with the empty vector. More importantly, xenograph tumors in nude mice grew significantly slower in vivo in cells stably transfected with TUSC1 than those transfected with empty vector. In addition, results from confocal microscopy and immunohistochemical analyses show distribution of TUSC1 in the cytoplasm and nucleus in tumor cell lines and in normal and tumor cells in the lung cancer tissue microarray. Taken together, our results support TUSC1 has tumor suppressor activity as a candidate tumor suppressor gene located on chromosome 9p.     

Tumor-targeted pH-low insertion peptide delivery of theranostic gadolinium nanoparticles for image-guided nanoparticle-enhanced radiation therapy

Tumor targeting studies using metallic nanoparticles (NPs) have shown that the enhanced permeability and retention effect may not be sufficient to deliver the amount of intratumoral and intracellular NPs needed for effective in vivo radiosensitization. This work describes a pH-Low Insertion Peptide (pHLIP) targeted theranostic agent to enable image-guided NP-enhanced radiotherapy using a clinically feasible amount of injected NPs. Conventional gadolinium (Gd) NPs were conjugated to pHLIPs and evaluated in vitro for radiosensitivity and in vivo for mouse MRI. Cultured A549 human lung cancer cells were incubated with 0.5 mM of pHLIP-GdNP or conventional GdNP. Mass spectrometry showed 78-fold more cellular Gd uptake with pHLIP-GdNPs, and clonogenic survival assays showed 44% more enhanced radiosensitivity by 5 Gy irradiation with pHLIP-GdNPs at pH 6.2. In contrast to conventional GdNPs, MR imaging of tumor-bearing mice showed pHLIP-GdNPs had a long retention time in the tumor (>9 h), suitable for radiotherapy, and penetrated into the poorly-vascularized tumor core. The Gd-enhanced tumor corresponded with low-pH areas also independently measured by an in vivo molecular MRI technique. pHLIPs actively target cell surface acidity from tumor cell metabolism and deliver GdNPs into cells in solid tumors. Intracellular delivery enhances the effect of short-range radiosensitizing photoelectrons and Auger electrons. Because acidity is a general hallmark of tumor cells, the delivery is more general than antibody targeting. Imaging the in vivo NP biodistribution and more acidic (often more aggressive) tumors has the potential for quantitative radiotherapy treatment planning and pre-selecting patients who will likely benefit more from NP radiation enhancement.     

Competitive but Not Allosteric mTOR Kinase Inhibition Enhances Tumor Cell Radiosensitivity

The mechanistic target of rapamycin (mTOR) is a critical kinase in the regulation of gene translation and has been suggested as a potential target for radiosensitization. The goal of this study was to compare the radiosensitizing activities of the allosteric mTOR inhibitor rapamycin with that of the competitive mTOR inhibitor PP242. On the basis of immunoblot analyses, whereas rapamycin only partially inhibited mTOR complex 1 (mTORC1) activity and had no effect on mTOR complex 2 (mTORC2), PP242 inhibited the activity of both mTOR-containing complexes. Irradiation alone had no effect on mTORC1 or mTORC2 activity. Clonogenic survival was used to define the effects of the mTOR inhibitors on in vitro radiosensitivity. In the two tumor cell lines evaluated, PP242 treatment 1 hour before irradiation increased radiosensitivity, whereas rapamycin had no effect. Addition of PP242 after irradiation also enhanced the radiosensitivity of both tumor lines. To investigate the mechanism of radiosensitization, the induction and repair of DNA double-strand breaks were evaluated according yH2AX foci. PP242 exposure did not influence the initial level of yH2AX foci after irradiation but did significantly delay the dispersal of radiationinduced yH2AX foci. In contrast to the tumor cell lines, the radiosensitivity of a normal human fibroblast cell line was not influenced by PP242. Finally, PP242 administration to mice bearing U251 xenografts enhanced radiationinduced tumor growth delay. These results indicate that in a preclinical tumor model PP242 enhances tumor cell radiosensitivity both in vitro and in vivo and suggest that this effect involves an inhibition of DNA repair.     

M867, a novel selective inhibitor of caspase-3 enhances cell death and extends tumor growth delay in irradiated lung cancer models

Background

Lung cancer remains the leading cause of cancer death worldwide. Radioresistance of lung cancer cells results in unacceptable rate of loco-regional failure. Although radiation is known to induce apoptosis, our recent study showed that knockdown of pro-apoptotic proteins Bak and Bax resulted in an increase in autophagic cell death and lung cancer radiosensitivity in vitro. To further explore the potential of apoptosis inhibition as a way to sensitize lung cancer for therapy, we tested M867, a novel chemical and reversible caspase-3 inhibitor, in combination with ionizing radiation in vivo and in vitro.

Methods and Findings

M867 reduced clonogenic survival in H460 lung cancer cells (DER = 1.27, p = 0.007) compared to the vehicle-treated treated cells. We found that administration of M867 with ionizing radiation in an in vivo mouse hind limb lung cancer model was well tolerated, and produced a significant tumor growth delay compared to radiation alone. A dramatic decrease in tumor vasculature was observed with M867 and radiation using von Willebrand factor staining. In addition, Ki67 index showed >5-fold reduction of tumor proliferation in the combination therapy group, despite the reduced levels of apoptosis observed with terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling staining. Radiosensitizing effect of M867 through inhibiting caspases was validated using caspase-3/-7 double-knockout (DKO) mouse embryonic fibroblasts (MEF) cell model. Consistent with our previous study, autophagy contributed to the mechanism of increased cell death, following inhibition of apoptosis. In addition, matrigel assay showed a decrease in in vitro endothelial tubule formation during the M867/radiation combination treatment.

Conclusions

M867 enhances the cytotoxic effects of radiation on lung cancer and its vasculature both in vitro and in vivo. M867 has the potential to prolong tumor growth delay by inhibiting tumor proliferation. Clinical trials are needed to determine the potential of this combination therapy in patients with locally advanced lung cancer.     

Suberoylanilide Hydroxamic Acid,an Inhibitor of Histone Deacetylase,Enhances Radiosensitivity and Suppresses Lung Metastasis in Breast Cancer In Vitro and In Vivo

Triple-negative breast cancer (TNBC), defined by the absence of an estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2 expression, is associated with an early recurrence of disease and poor outcome. Furthermore, the majority of deaths in breast cancer patients are from metastases instead of from primary tumors. In this study, MCF-7 (an estrogen receptor-positive human breast cancer cell line), MDA-MB-231 (a human TNBC cell line) and 4T1 (a mouse TNBC cell line) were used to investigate the anti-cancer effects of ionizing radiation (IR) combined with suberoylanilide hydroxamic acid (SAHA, an inhibitor of histone deacetylase (HDAC)) and to determine the underlying mechanisms of these effects in vitro and in vivo. We also evaluated the ability of SAHA to inhibit the metastasis of 4T1 cells. We found that IR combined with SAHA showed increased therapeutic efficacy when compared with either treatment alone in MCF-7, MDA-MB-231 and 4T1 cells. Moreover, the combined treatment enhanced DNA damage through the inhibition of DNA repair proteins. The combined treatment was induced primarily through autophagy and ER stress. In an orthotopic breast cancer mouse model, the combination treatment showed a greater inhibition of tumor growth. In addition, SAHA inhibited the migration and invasion abilities of 4T1 cells and inhibited breast cancer cell migration by inhibiting the activity of MMP-9. In an in vivo experimental metastasis mouse model, SAHA significantly inhibited lung metastasis. SAHA not only enhances radiosensitivity but also suppresses lung metastasis in breast cancer. These novel findings suggest that SAHA alone or combined with IR could serve as a potential therapeutic strategy for breast cancer.     

SIRT1 Expression Is Associated with the Chemotherapy Response and Prognosis of Patients with Advanced NSCLC

Aim

The role of Sirtuin 1 (SIRT 1) in carcinogenesis is controversial. This study was to explore the association between the SIRT1 expression and the clinical characteristics, the responsiveness to chemotherapy and prognosis in Non-small cell lung cancer (NSCLC).

Methods

We enrolled 295 patients with inoperable advanced stage of NSCLC, namely, stage III (A+B) and IV NSCLC. All patients had received platinum-based chemotherapy after diagnosis and the chemotherapy response were evaluated. All patients were followed up for overall survival (OS) and progression free survival (PFS). In vitro, H292 cells were tranfected with SIRT1 small interfering RNA (siRNA). The cell biological behaviors and chemosensitivity to cisplatin treatment were studied. The in vivo tumorgenesis and metastasis assays were performed in nude mice.

Results

We found that the SIRT1 expressions were significantly associated with the tumor stage, tumor size and differentiation status. Patients with high SIRT 1 expressions had a significantly higher chance to be resistant to chemotherapy than those with low SIRT 1 expression. Patients with high expression of SIRT1 had significantly shorter OS and DFS than those with low expression. Cox analyses confirmed that the SIRT 1 expression was a strong predictor for a poor OS and PFS in NSCLC patients underwent Platinum-based chemotherapy. In vitro studies revealed that the reduced expression SIRT 1 by siRNA technique significantly inhibited cell proliferation, migration and invasion. More importantly, SIRT1 si-RNA significantly enhanced the chemosensitivity of H292 cells to cisplatin treatment. The in vivo tumorgenesis and metastasis assays showed that SIRT1 knockdown dramatically reduced the tumor volume and the metastatic ability in nude mice.

Conclusion

Collectively, our data suggest that the SIRT1 expression may be a molecular marker associated with the NSLCLC clinical features, treatment responsiveness and prognosis of advanced NSCLC.     

ABCA1, ABCG1 and SR-BI: hormonal regulation in primary rat hepatocytes and human cell lines

ATM and PARP-1 are two of the most important players in the cell's response to DNA damage. PARP-1 and ATM recognize and bound to both single and double strand DNA breaks in response to different triggers. Here we report that ATM and PARP-1 form a molecular complex in vivo in undamaged cells and this association increases after γ-irradiation. ATM is also modified by PARP-1 during DNA damage. We have also evaluated the impact of PARP-1 absence or inhibition on ATM-kinase activity and have found that while PARP-1 deficient cells display a defective ATM-kinase activity and reduced γ-H2AX foci formation in response to γ-irradiation, PARP inhibition on itself is able to activate ATM-kinase. PARP inhibition induced γ H2AX foci accumulation, in an ATM-dependent manner. Inhibition of PARP also induces DNA double strand breaks which were dependent on the presence of ATM. As consequence ATM deficient cells display an increased sensitivity to PARP inhibition. In summary our results show that while PARP-1 is needed in the response of ATM to gamma irradiation, the inhibition of PARP induces DNA double strand breaks (which are resolved in and ATM-dependent pathway) and activates ATM kinase.     

Radiosensitization of Chinese hamster V79 cells by bromodeoxyuridine substitution of thymidine: enhancement of radiation-induced toxicity and DNA strand break production by monofilar and bifilar substitution

Bromodeoxyuridine (BrdU) competes with thymidine (TdR) for incorporation into DNA of exponentially growing V79-171 cells. Such cells show an enhancement of the radiation response as determined by clonogenic survival and DNA damage measured by filter elution techniques after doses up to 15 Gy. The degree of radiosensitization for both survival and rates of alkaline and neutral elution are dependent on percentage BrdU substitution and independent of whether BrdU is in one strand only (monofilar) or both strands (bifilar) of the DNA duplex: e.g., for 16% BrdU substitution distributed either monofilarly or partially bifilarly, there is an enhancement factor for Do of 1.55. At this percentage substitution, the enhancement factor for the rate of alkaline elution is 1.75 and that for the rate of neutral elution is 1.54. The greater the percentage BrdU substitution, the larger was the enhancement ratio for survival and radiation-induced strand breaks in both monofilarly and bifilarly substituted cells. The increase in cell radiosensitivity caused by BrdU substitution shows a better correlation with the increase in radiation-induced double-strand breaks than with the increase in radiation-induced single-strand breaks.     

Evaluation of Different Biomarkers to Predict Individual Radiosensitivity in an Inter-Laboratory Comparison–Lessons for Future Studies

Radiotherapy is a powerful cure for several types of solid tumours, but its application is often limited because of severe side effects in individual patients. With the aim to find biomarkers capable of predicting normal tissue side reactions we analysed the radiation responses of cells from individual head and neck tumour and breast cancer patients of different clinical radiosensitivity in a multicentric study. Multiple parameters of cellular radiosensitivity were analysed in coded samples of peripheral blood lymphocytes (PBLs) and derived lymphoblastoid cell lines (LCLs) from 15 clinical radio-hypersensitive tumour patients and compared to age- and sex-matched non-radiosensitive patient controls and 15 lymphoblastoid cell lines from age- and sex- matched healthy controls of the KORA study. Experimental parameters included ionizing radiation (IR)-induced cell death (AnnexinV), induction and repair of DNA strand breaks (Comet assay), induction of yH2AX foci (as a result of DNA double strand breaks), and whole genome expression analyses. Considerable inter-individual differences in IR-induced DNA strand breaks and their repair and/or cell death could be detected in primary and immortalised cells with the applied assays. The group of clinically radiosensitive patients was not unequivocally distinguishable from normal responding patients nor were individual overreacting patients in the test system unambiguously identified by two different laboratories. Thus, the in vitro test systems investigated here seem not to be appropriate for a general prediction of clinical reactions during or after radiotherapy due to the experimental variability compared to the small effect of radiation sensitivity. Genome-wide expression analysis however revealed a set of 67 marker genes which were differentially induced 6 h after in vitro-irradiation in lymphocytes from radio-hypersensitive and non-radiosensitive patients. These results warrant future validation in larger cohorts in order to determine parameters potentially predictive for clinical radiosensitivity.     

Src-Mediated Cross-Talk between Farnesoid X and Epidermal Growth Factor Receptors Inhibits Human Intestinal Cell Proliferation and Tumorigenesis

Besides its essential role in controlling bile acid and lipid metabolism, the farnesoid X receptor (FXR) protects against intestinal tumorigenesis by promoting apoptosis and inhibiting cell proliferation. However, the mechanisms underlying these anti-proliferative actions of FXR remain to be elucidated. In the present study, we examined the effects of FXR activation (FXR overexpression and treatment with an FXR agonist GW4064) and inactivation (treatment with FXR siRNA and an FXR antagonist guggulsterone) on colon cancer cell proliferation in vitro using human colon cancer cell lines (H508, SNU-C4 and HT-29) and in vivo using xenografts in nude mice. Blocking FXR activity with guggulsterone stimulated time- and dose-dependent EGFR (Tyr845) phosphorylation and ERK activation. In contrast, FXR overexpression and activation with GW4064 attenuated cell proliferation by down-regulating EGFR (Tyr845) phosphorylation and ERK activation. Treatment with guggulsterone and GW4064 also caused dose-dependent changes in Src (Tyr416) phosphorylation. In stably-transfected human colon cancer cells, overexpression of FXR reduced EGFR, ERK, Src phosphorylation and cell proliferation, and in nude mice attenuated the growth of human colon cancer xenografts (64% reduction in tumor volume; 47% reduction in tumor weight; both P<0.01). Moreover, guggulsterone-induced EGFR and ERK phosphorylation and cell proliferation were abolished by inhibiting activation of Src, EGFR and MEK. Collectively these data support the novel conclusion that in human colon cancer cells Src-mediated cross-talk between FXR and EGFR modulates ERK phosphorylation, thereby regulating intestinal cell proliferation and tumorigenesis.     

MTA1-TJP1 interaction and its involvement in non-small cell lung cancer metastasis

Distant metastasis is the main cause of death in non-small cell lung cancer (NSCLC) patients. The mechanism of metastasis-associated protein 1(MTA1) in NSCLC has not been fully elucidated. This study aimed to reveal the mechanism of MTA1 in the invasion and metastasis of NSCLC.Bioinformatics analysis and our previous results showed that MTA1 was highly expressed in NSCLC tissues and correlated with tumor progression. Knockout of MTA1 by CRISPR/Cas9 significantly inhibited the migration and invasion of H1299 cells, but enhanced cell adhesion. Stable overexpression of MTA1 by lentivirus transfection had opposite effects on migration, invasion and adhesion of A549 cells. The results of in vivo experiments in nude mouse lung metastases model confirmed the promotion of MTA1 on invasion and migration. Tight junction protein 1 (TJP1) was identified by immunoprecipitation and mass spectrometry as an interacting protein of MTA1 involved in cell adhesion. MTA1 inhibited the expression level of TJP1 protein and weakened the tight junctions between cells. More importantly, the rescue assays confirmed that the regulation of MTA1 on cell adhesion, migration and invasion was partially attenuated by TJP1.In Conclusion, MTA1 inhibits the expression level of TJP1 protein co-localized in the cytoplasm and membrane of NSCLC cells, weakens the tight junctions between cells, and changes the adhesion, migration and invasion capabilities of cells, which may be the mechanism of MTA1 promoting the invasion and metastasis of NSCLC. Thus, targeting the MTA1-TJP1 axis may be a promising strategy for inhibiting NSCLC metastasis.     

Identifying new therapeutic targets via modulation of protein corona formation by engineered nanoparticles

Background

We introduce a promising methodology to identify new therapeutic targets in cancer. Proteins bind to nanoparticles to form a protein corona. We modulate this corona by using surface-engineered nanoparticles, and identify protein composition to provide insight into disease development.

Methods/Principal Findings

Using a family of structurally homologous nanoparticles we have investigated the changes in the protein corona around surface-functionalized gold nanoparticles (AuNPs) from normal and malignant ovarian cell lysates. Proteomics analysis using mass spectrometry identified hepatoma-derived growth factor (HDGF) that is found exclusively on positively charged AuNPs (⁺AuNPs) after incubation with the lysates. We confirmed expression of HDGF in various ovarian cancer cells and validated binding selectivity to ⁺AuNPs by Western blot analysis. Silencing of HDGF by siRNA resulted s inhibition in proliferation of ovarian cancer cells.

Conclusion

We investigated the modulation of protein corona around surface-functionalized gold nanoparticles as a promising approach to identify new therapeutic targets. The potential of our method for identifying therapeutic targets was demonstrated through silencing of HDGF by siRNA, which inhibited proliferation of ovarian cancer cells. This integrated proteomics, bioinformatics, and nanotechnology strategy demonstrates that protein corona identification can be used to discover novel therapeutic targets in cancer.     

AKT2 contributes to increase ovarian cancer cell migration and invasion through the AKT2-PKM2-STAT3/NF-κB axis

Multiple studies have shown that protein kinase Bβ (AKT2) is involved in the development and progression of ovarian cancer, however, its precise role remains unclear. Here we explored the underlying molecular mechanisms how AKT2 promotes ovarian cancer progression. We examined the effects of AKT2 in vitro in two ovarian cancer cell lines (SKOV3 and HEY), and in vivo by metastasis assay in nude mice. The migration and invasion ability of SKOV3 and HEY cells was determined by transwell assay. Overexpression and knockdown (with shRNA) experiments were carried out to unravel the underlying signaling mechanisms induced by AKT2. Overexpression of AKT2 led to increased expression of pyruvate kinase (PKM2) in ovarian cancer cells and in lung metastatic foci from nude mice. Elevated AKT2/PKM2 expression induced cell migration and invasion in vitro, as well as lung metastasis in vivo; silencing AKT2 blocked these effects. Meanwhile, PKM2 overexpression was unable to increase AKT2 expression. The expressions of p-PI3K, p-AKT2, and PKM2 were increased when stimulated by epidermal growth factor (EGF); however, these expressions were blocked when inhibited the PI3K by LY294002. STAT3 expression was elevated and NF-κB p65 nuclear translocation was activated both in vitro and in vivo when either AKT2 or PKM2 was overexpressed; and these effects were inhibited when silencing AKT2 expression. Taken together, AKT2 increases the migration and invasion of ovarian cancer cells in vitro and promotes lung metastasis in nude mice in vivo through PKM2-mediated elevation of STAT3 expression and NF-κB activation. In conclusion, we highlight a novel mechanism of the AKT2-PKM2-STAT3/NF-κB axis in the regulation of ovarian cancer progression, and our work suggested that both AKT2 and PKM2 may be potential targets for the treatment of ovarian cancer.