Fibrosarcoma Cell Line HT1080 Research Articles

Release of Exosomal PD-L1 in Bone and Soft Tissue Sarcomas and Its Relationship to Radiotherapy.

(1) Background: Exosomal PD-L1 has garnered attention owing to its role in instigating systemic immune suppression. The objective of this study is to elucidate whether bone and soft tissue sarcoma cells possess the capacity to secrete functionally active exosomal PD-L1 and whether radiotherapy (RT) induces the exosomal PD-L1 release. (2) Methods: Human osteosarcoma cell line 143B and human fibrosarcoma cell line HT1080 were utilized. Exosomes were isolated from the culture medium and blood via ultracentrifugation. The expression of PD-L1 on both tumor cells and exosomes was evaluated. The inhibitory effect on PBMC was employed to assess the activity of exosomal PD-L1. Post radiotherapy, changes in PD-L1 expression were compared. (3) Results: Exosomal PD-L1 was detected in the culture medium of tumor cells but was absent in the culture medium of PD-L1 knockout cells. Exosomal PD-L1 exhibited an inhibitory effect on PBMC activation. In tumor-bearing mice, human-derived exosomal PD-L1 was detected in the bloodstream. Following radiotherapy, tumor cells upregulated PD-L1, and human-derived exosomal PD-L1 were detected in the bloodstream. (4) Conclusions: Exosomal PD-L1 emanates from bone and soft tissue sarcoma cells and is disseminated into the circulatory system. The levels of PD-L1 in tumor cells and the release of exosomal PD-L1 were augmented after irradiation with RT.

Several first-line anti-hypertensives act on fibrosarcoma progression and PD1ab blockade therapy.

Patients are typically diagnosed with both hypertension and fibrosarcoma. Medical oncologists must prescribe suitable anti-hypertensive medications while considering anti-tumor drugs. Recently, immunotherapy has become prominent in cancer treatment. Nonetheless, it is unknown what role anti-hypertensive medications will play in immunotherapy. We examined the effects of six first-line anti-hypertensive medications on programmed cell death protein 1 antibody (PD1ab) in tumor treatment using a mouse model of subcutaneous fibrosarcoma. The drugs examined were verapamil, losartan, furosemide, spironolactone, captopril, and hydrochlorothiazide (HCTZ). The infiltration of CD8+ T cells was examined by immunohistochemistry. Additionally, several in vitro and in vivo assays were used to study the effects of HCTZ on human fibrosarcoma cancer cells to explore its mechanism. Verapamil suppressed tumor growth and showed an improved effect on the tumor inhibition of PD1ab. Captopril did not affect tumor growth but brought an unexpected benefit to PD1ab treatment. In contrast, spironolactone andfurosemide showed no effect on tumor growth but had an offset effect on the PD1ab therapy. Consequently, the survival time of mice was also significantly reduced. Notably, losartan and HCTZ, especially HCTZ, promoted tumor growth and weakened the effect of PD1ab treatment. Consistent results were observed in vivo and in vitro using the human fibrosarcoma cell line HT1080. We determined that the Solute Carrier Family 12 Member 3 (SLC12A3), a known target of HCTZ, may be the principal factor underlying its effect-enhancing properties through mechanism studies employing The Cancer Genome Atlas (TCGA) data and in vivo and in vitro assays. Verapamil and captopril potentiated the anti-tumor effect of PD1ab, whereas spironolactone andfurosemide weakened the effect of PD1ab on tumor inhibition. Alarmingly, losartan and HCTZ promoted tumor growth and impaired the effect of PD1ab. Furthermore, we preliminarily found that HCTZ may promote tumor progression through SLC12A3. Based on this study, futher mechanism researches and clinical trials should be conducted in the future.

Soluble CD147 regulates endostatin via its effects on the activities of MMP-9 and secreted proteasome 20S.

During progression of rheumatoid arthritis (RA), angiogenesis provides oxygen and nutrients for the cells' increased metabolic demands and number. To turn on angiogenesis, pro-angiogenic factors must outweigh anti-angiogenic factors. We have previously shown that CD147/extracellular matrix metalloproteinase inducer (EMMPRIN) can induce the expression of the pro-angiogenic factors vascular endothelial growth factor (VEGF) and matrix metallopeptidase 9 (MMP-9) in a co-culture of the human HT1080 fibrosarcoma and U937 monocytic-like cell lines. However, whether CD147 influences anti-angiogenic factors was not known. We now show that relative to single cultures, the co-culture of these cells not only enhanced pro-angiogenic factors but also decreased the anti-angiogenic factors endostatin and thrombospondin-1 (Tsp-1), generally increasing the angiogenic potential as measured by a wound assay. Using anti-CD147 antibody, CD147 small interfering RNA (siRNA), and recombinant CD147, we demonstrate that CD147 hormetically regulates the generation of endostatin but has no effect on Tsp-1. Since endostatin is cleaved from collagen XVIII (Col18A), we applied different protease inhibitors and established that MMP-9 and proteasome 20S, but not cathepsins, are responsible for endostatin generation. MMP-9 and proteasome 20S collaborate to synergistically enhance endostatin generation, and in a non-cellular system, CD147 enhanced MMP-9 activity and hormetically regulated proteasome 20S activity. Serum samples obtained from RA patients and healthy controls mostly corroborated these findings, indicating clinical relevance. Cumulatively, these findings suggest that secreted CD147 mediates a possibly allosteric effect on MMP-9 and proteasome 20S activities and can serve as a switch that turns angiogenesis on or off, depending on its ambient concentrations in the microenvironment.

Endoplasmic Reticulum Stress Inducer Dithiothreitol Affects the Morphology and Motility of Cultured Human Dermal Fibroblasts and Fibrosarcoma HT1080 Cell Line

Endoplasmic Reticulum Stress Inducer Dithiothreitol Affects the Morphology and Motility of Cultured Human Dermal Fibroblasts and Fibrosarcoma HT1080 Cell Line

Danthron, an Anthraquinone Isolated from a Marine Fungus, Is a New Inhibitor of Angiogenesis Exhibiting Interesting Antitumor and Antioxidant Properties.

The role played by a sustained angiogenesis in cancer and other diseases stimulates the interest in the search for new antiangiogenic drugs. In this manuscript, we provide evidence that 1,8- dihydroxy-9,10-anthraquinone (danthron), isolated from the fermentation broth of the marine fungus Chromolaenicola sp. (HL-114-33-R04), is a new inhibitor of angiogenesis. The results obtained with the in vivo CAM assay indicate that danthron is a potent antiangiogenic compound. In vitro studies with human umbilical endothelial cells (HUVEC) reveal that this anthraquinone inhibits certain key functions of activated endothelial cells, including proliferation, proteolytic and invasive capabilities and tube formation. In vitro studies with human breast carcinoma MDA-MB231 and fibrosarcoma HT1080 cell lines suggest a moderate antitumor and antimetastatic activity of this compound. Antioxidant properties of danthron are evidenced by the observation that it reduces the intracellular reactive oxygen species production and increases the amount of intracellular sulfhydryl groups in endothelial and tumor cells. These results support a putative role of danthron as a new antiangiogenic drug with potential application in the treatment and angioprevention of cancer and other angiogenesis-dependent diseases.

Functional roles of FAP-α in metabolism, migration and invasion of human cancer cells.

Fibroblast activation protein-α (FAP-α) is a transmembrane serine protease that is attracting significant interest as it is expressed by a subgroup of cancer-associated fibroblasts that play a role in immune suppression and cancer metastasis. FAP-α is also expressed by some cancer cells, such as melanoma, colorectal and breast cancer cells. Triple negative breast cancer (TNBC) is an aggressive cancer that urgently requires identification of novel targets for therapy. To expand our understanding of the functional roles of FAP-α in TNBC we engineered a human TNBC cell line, MDA-MB-231, to stably overexpress FAP-α and characterized changes in metabolism by 1H magnetic resonance spectroscopy, cell proliferation, migration characterized by wound healing, and invasion. FAP-α overexpression resulted in significant alterations in myoinositol, choline metabolites, creatine, and taurine, as well as a significant increase of migration and invasion, although proliferation remained unaltered. The increase of migration and invasion are consistent with the known activities of FAP-α as an exopeptidase and endopeptidase/gelatinase/collagenase in tissue remodeling and repair, and in cell migration. We additionally determined the effects of FAP-α overexpression on the human fibrosarcoma HT1080 cell line that showed increased migration, accompanied by limited changes in metabolism that identified the dependency of the metabolic changes on cell type. These metabolic data identify a previously unknown role of FAP-α in modifying cancer cell metabolism in the TNBC cell line studied here that may provide new insights into its functional roles in cancer progression.

Abstract A016: Selective delivery of fibroblast activation protein conjugated dual phosphoinositide 3-kinase–AKT Kinase-mTOR inhibitor associated with decreased tumor proliferation and on-target toxicity in high-grade soft-tissue sarcomas

Abstract Soft-tissue sarcomas (STS) are a group of lethal mesenchymal tumors that are a major cause of cancer related morbidity for those under the age of 20.PI3K inhibitors have been found to be effective against subset of STS but are associated with significant toxicities limiting their use.We have thought to test a selective fibroblast activation protein (FAP) targeted delivery of PI3K/AKT/mTOR inhibitor omipalisib to STS in pre-clinical setting as FAP is overwhelmingly expressed agressive soft tissue malignancies. Experimental Design: Patient derived fibrosarcomas (aSMA-,FAP+,pAKT+) were isolated by flow cytometry.FAP(-)HT1080 fibrosarcoma cell lines were used as a negative control whereas methylcoanthrene induced mouse fibrosarcoma transfected with human FAP cDNA were used as positive control.Cell expression of FAP,AKT,mTOR was compared using SDS Page,IHC,and flow cytometry.FAP ligand coupled to omipalisib(FAPL-PI3Ki)was then evaluated in-vitro and in small animal models. Results: Cell lines with known FAP expression demonstrated significant uptake of FAPL conjugated NIR fluorochrome.Wild type cells(hFAP-) showed no ligand binding both in flow cytometry and fluorescent microscopy(p<0.05).Escalating doses of FAP-PI3Ki suppressed AKT/mTOR activity in HT-1080 hFAP cells with 10 nM being the smallest effective dose.In hFAP+cell lines 10 nM of FAP-PI3Ki inhibited the AKT/mTOR signaling but inhibition was not observed in hFAP- cell lines demonstrating the need for FAP presence for FAPL-PI3Ki activity(p<0.05).Pan non-targeted PI3Ki inhibited the AKT/mTOR pathway in both hFAP+ and hFAP-cell lines.Densitometric analysis demonstrated that in hFAP+ cells,FAP-PI3Ki at above 10 nM suppresses the AKT/mTOR activity with ratio of pAKT/AKT and pmTOR/mTOR decreasing 11 and 4-fold respectively(p<0.05).FAP-PI3Ki activity significantly reduced when hFAP+ cells were competitively inhibited with free FAPL(4.6x increase in AKT activity,p<0.05).hFAP+ cells treated with FAP-PI3Ki had significant decrease in cell migration.In small animal models,SCID Mice injected with 25 umol/kg of either FAP-PI3Ki and non-targeted PI3Ki showed marked elevation of serum insulin levels in the non-targeted PI3Ki group (0.91 ng/mL vs 0.58 ng/mL,p<0.05).Xenograft tumor digest demonstrated mice treated with FAP-PI3Ki had Akt/mTOR pathway downregulated only in hFAP+ cell lines and this was evident in mice which received 10 nM or higher dosing(pAkt/Akt ratio of 1.1 WT mice vs 0.55 in hFAP+ mice,p<0.05).There was near complete suppression of the mTOR expression in the hFAP+ treatment group (pmTOR/mTOR 1.05 vs 0.087,p<0.05).There was no treatment related mortality in FAPL-PI3Ki group as compared to non-targeted PI3Ki delivery (p<0.05). Conclusion: Our study demonstrated that FAP conjugated modified omipalisib binds FAP expressing soft-tissue sarcomas both in-vitro and in xenograft models in concordance with FAP receptor density.FAP-PI3Ki induces PI3K/AkT/mTOR pathway inhibition similar to non-targeted PI3K inhibitors without significant systemic toxicities observed in pan nontargeted PI3K inhibitors. Citation Format: Feredun Azari, Gregory T. Kennedy, Ashley Chang, Bilal Nadeem, Azra Din, Isvita Marfatia, Steven Albelda, Philip Low, Madduri Srinivasarao, Ramesh Mukkamala, Neil T. Sullivan, Evgeniy Eruslanov, Sunil Singhal. Selective delivery of fibroblast activation protein conjugated dual phosphoinositide 3-kinase–AKT Kinase-mTOR inhibitor associated with decreased tumor proliferation and on-target toxicity in high-grade soft-tissue sarcomas [abstract]. In: Proceedings of the AACR Special Conference: Sarcomas; 2022 May 9-12; Montreal, QC, Canada. Philadelphia (PA): AACR; Clin Cancer Res 2022;28(18_Suppl):Abstract nr A016.

A time-resolved fibrosis model – in vitro assessment of antifibrotic implant coatings

Abstract Wound healing after implant surgery represents a spontaneous repair process that, when disturbed, can lead to a pathological situation known as fibrosis, characterized by excessive proliferation, increased extracellular matrix deposition, and scarring of the tissue. Tissue-specific cell culture models simulating physiological or pathophysiological cell responses are valuable research tools in the field of fibrosis, among others. In such in vitro systems, endpoint measurements are typically used to assess cell viability and/or proliferation. However, this approach is usually time consuming, costly and often leads to the destruction of the sample. Additionally, these conventional in vitro systems provide only a single snapshot of the respective sample at the end of the experiment. Therefore, we aimed to evolve the fibrotic disease in vitro model established by Stahnke et al. To this end, we chose a time-resolved, non-invasive approach based on cell impedance measurements and bright-field microscopy that allows continuous monitoring of the cells. Such measurements can be directly conducted using the xCELLigence RTCA eSight system. This system was used to characterize the cellular responses of the fibrosarcoma cell line HT-1080 and primary fibroblasts isolated from the Tenon’s capsule of human donors to TGF-β1, a key cytokine involved in the pathogenesis of fibrosis. The data shows that compared to Tenon’s fibroblasts the response of HT-1080 cells to TGF- β1 is less pronounced and only detectable within a narrow time frame. Thus, HT-1080 do not appear to be a suitable cell culture model for the assessment of antifibrotic implant coatings.

The Distinct Roles of LKB1 and AMPK in p53-Dependent Apoptosis Induced by Cisplatin.

Liver kinase B1 (LKB1) is a serine/threonine protein kinase that acts as a key tumor suppressor protein by activating its downstream kinases, such as AMP-activated protein kinase (AMPK). However, the regulatory actions of LKB1 and AMPK on DNA damage response (DDR) remain to be explored. In this study, we investigated the function of LKB1 in DDR induced by cisplatin, a representative DNA-damaging agent, and found that LKB1 stabilizes and activates p53 through the c-Jun N-terminal kinase (JNK) pathway, which promotes cisplatin-induced apoptosis in human fibrosarcoma cell line HT1080. On the other hand, we found that AMPKα1 and α2 double knockout (DKO) cells showed enhanced stabilization of p53 and increased susceptibility to apoptosis induced by cisplatin, suggesting that AMPK negatively regulates cisplatin-induced apoptosis. Moreover, the additional stabilization of p53 and subsequent apoptosis in AMPK DKO cells were clearly canceled by the treatment with the antioxidants, raising the possibility that AMPK suppresses the p53 activation mediated by oxidative stress. Thus, our findings unexpectedly demonstrate the reciprocal regulation of p53 by LKB1 and AMPK in DDR, which provides insights into the molecular mechanisms of DDR.

KIF20A promotes the development of fibrosarcoma via PI3K-Akt signaling pathway

Adult fibrosarcoma is an aggressive subtype of soft tissue sarcoma (STS), in which high expression of KIF20A indicates a poor prognosis. However, the precise role of KIF20A in fibrosarcoma progression remains unknown. In this study, we initially examined KIF20A expression and function in the human fibrosarcoma cell line HT-1080. The results showed that KIF20A was highly expressed in HT-1080, knockdown of KIF20A impaired cell proliferation, migration, invasion and induced G2/M arrest and cell apoptosis. Transcriptome study suggested that PI3K-Akt signal pathway was involved in these biological changes. We confirmed that PI3K-Akt and NF-κB signaling pathways were impaired after the down-regulation of KIF20A, which can be reversed by the Akt activator SC79 in HT-1080 in vitro. In a xenograft mouse model, knockdown of KIF20A inhibited tumor growth, Ki67 expression and liver metastasis. Taken together, our results suggested that KIF20A promoted fibrosarcoma progression via PI3K-Akt signaling pathway and might be a potential therapeutic target for fibrosarcoma.

Preparation of thermo-responsive polymer encapsulated exosomes and its role as a therapeutic agent for blood clot lysis

Tissue plasminogen activators induce enzymatic activation of plasminogen to plasmin that cleaves fibrin strands in blood clots. In the present study, extracellular vesicles such as exosomes from fibrosarcoma cell line HT1080 were utilized as clot-busting agents. These exosomes were being used for clot lysis of whole blood which showed 28% lysis within 10 h, which was comparable to that of the streptokinase (commercial plasmin activator) with no significant difference. These exosomes were able to facilitate the migration of endothelial cells in a scratch wound assay where normalized wound area remaining was 7.5% at 18 h. Also, exosomes aided in attenuation of oxidative stress generated on the cells, thereby maintaining cell viability. These exosomes were further encapsulated in a thermo-responsive polymer for better localized delivery that showed no cytotoxic effects, and sustained delivery was achieved up to a concentration of 117 µg/mL in 25 days, which corresponds to around 65% of the total amount of exosomes added. When a combination of exosomes and thermo–responsive polymer was utilized, the clot lysis activity reached to around 22% in 72 h. Thus, it proves the potential of this combinatorial approach which can be effectively used for thrombus degradation and healing of endothelium lining in damaged blood vessels.

In vitro antimicrobial and anti-proliferative activity of crude methanolic extract of pigment from Streptomycetes spp. on HT-1080 fibro sarcoma cell line

The ability to produce secondary metabolites like antibiotics is the characteristic feature of Streptomyces. Besides antibiotics, they are also seen to produce other bioactive compounds of economic importance like pigments which are potentially used in industries with remarkably appealing results. Under this backdrop, extraction of such priceless molecule from microbial source was the aim of this work. Actinomycetes strain isolated from water sample was identified as Streptomycetes with conventional and molecular approach. The metabolite exhibited antibacterial, antioxidant and anticancer activity as evident from the liquid fermentation. The crude ethanolic pigment extract was shown to be antibacterial on gram negative human pathogens. Also it exhibited antioxidant and anticancer activity against two cancer cell lines i.e. HeLa and HT-1080. Pigments from bacteria can be effectively used as natural source of treatment against various human ailments. Our results provide insights into effective role of pigment as potent bioactive metabolites with effective pharmaceutical properties.

Analysis of Active Compounds Using Target Protein Cofilin-Cucurbitacins in Cytotoxic Plant Bryonia cretica.

We examined a two-step target protein binding strategy that uses cofilin as the target protein to analyze the active constituents in Bryonia cretica. In the first step, we prepared the target protein, and used it to analyze the compounds binding to it in the second step. We used the methanolic extract of B. cretica as a library of possible active compounds. We conducted LC–MS analysis using information from our previous study. The peaks in the HPLC profile were identified as cucurbitacin D, isocucurbitacin D, and cucurbitacin I. As far as we know, there is no known study of the activity of isocucurbitacin D in this research field. Therefore, we examined the effects of isocucurbitacin D on cell proliferation and cofilin protein in human fibrosarcoma cell line HT1080 to confirm the effectiveness of this strategy. The cytotoxicity assay, the fibrous/globular actin ratio assay, and the immunoblotting analysis revealed that isocucurbitacin D showed a cytotoxic effect with disruption of target protein cofilin. The target protein binding strategy is a direct and straightforward method for finding new drug seeds from crude sources, such as natural plant extracts.

DNA-Directed Assembly of a Cell-Responsive Biohybrid Interface for Cargo Release.

The development of a DNA-based cell-responsive biohybrid interface that can be used for spatially confined release of molecular cargo is reported. To this end, tailored DNA-protein conjugates are designed as gatekeepers that can be specifically cleaved by matrix metalloproteases (MMPs), which are secreted by many cancer cells. These gatekeepers can be installed by DNA hybridization on the surface of mesoporous silica nanoparticles (MSNs). The MSNs display another orthogonal DNA oligonucleotide that can be exploited for site-selective immobilization on solid glass surfaces to yield micropatterned substrates for cell adhesion. Using the human fibrosarcoma cell line HT1080 that secretes MMPs, it is demonstrated that the biohybrid surface is specifically modified by the cells to release both MSN-bound gatekeeper proteins and the encapsulated cargo peptide KLA. In view of the enormously high modularity of the system presented here, this approach promising for applications in drug delivery, tissue engineering, or other areas of nanobiotechnology is considered.

Microrna-145 Inhibits EMT and Fibrosarcoma Cell Proliferation, Migration and Invasion by Targeting HMGA2

1.1 Aim: Fibrosarcoma is a rare malignant tumor of subcutaneous tissue characterized by slow infiltrative growth and a high rate of local recurrence. The participation of microRNAs (miRNAs/miRs) in the pathogenesis of human diseases and the vital role of miR-145 in different types of cancer. However, the role of miR-145 in fibrosarcoma remains unclear. The present study aimed to investigate the effect of miR-145 in the progression of fibrosarcoma and its potential application as a new therapeutic target. 1.2. Methods: The present study screened out a differential gene named high mobility group protein A2 (HMGA2) via bioinformatics analysis. In the present study, the gene expression profiles of fibrosarcoma cells and normal cells were downloaded from GEO Datasets GSE10021 and GSE1774. The human fibrosarcoma cell line HT1080 were transfected with miR-145 mimics, mimic negative control (NC), pCMV6 vector, overexpression plasmid vector of HMGA2, (si)RNA-HMGA2 or siRNA-NC. Reverse transcription-quantitative PCR and western blot analysis were performed to examine the mRNA (miR-145) and protein (HMGA2, N-cadherin, Vimentin and E-cadherin) expression levels in HT1080 cells. Cell Counting Kit-8, wound healing and Transwell assays were used to examine the TH1080 cell viability, proliferation and migratory capacities, respectively. Bioinformatics prediction and luciferase reporter assay were performed to investigate the relationship between miR145 and its potential target gene (HMGA2). 1.3. Results: The results demonstrated that overexpression of HMGA2 promoted cell proliferation, migration and invasion of HT1080 fibrosarcoma cells, whereas HMGA2 knockdown had no effect on the proliferation of HT1080 fibrosarcoma cells. However, overexpression of miR-145 reversed the facilitating effects of HMGA2 in tumor progression via epithelial-to-mesenchymal transition (EMT) modulation. Mechanistically, miR-145 exerted its inhibitory role in fibrosarcoma by directly binding to the 3’-untranslated region of HMGA2. 1.4. Conclusion: miR-145 plays an inhibitory role in fibrosarcoma progression by directly suppressing HGMA2 expression and EMT progression. Thus, miR-145 may act as a potential therapeutic target for malignant fibrosarcoma therapy

Intrinsic Oncolytic Activity of Hoshino Mumps Virus Vaccine Strain Against Human Fibrosarcoma and Cervical Cancer Cell Lines

Background: The use of oncolytic viruses as therapeutic agents is a promising treatment for various human cancers. Several viruses have been extensively examined to achieve tumor cell death. Objectives: This study aimed at evaluating the natural oncolytic activity of mumps Hoshino vaccine strain against two human cancer cell lines, that is, HT1080 fibrosarcoma and HeLa cervical adenocarcinoma cell lines. Methods: The cytolytic activity of the virus was evaluated using an MTT assay. Apoptosis was detected by Annexin-V/propidium iodide (PI) staining and analyzed via flow cytometry. To indicate viral replication in vivo, nude mice with HeLa heterografts were treated with the Hoshino strain of mumps virus. Results: It was found that human fibrosarcoma and cervical cells were more sensitive to the mumps Hoshino strain, even at a very low multiplicity of infection (MOI) compared to normal human diploid cells. The results also showed that the Hoshino strain induced apoptosis in both cancer cells. A preliminary in vivo study revealed the significant suppression of tumor growth in the group treated with the mumps Hoshino strain compared to the control group. Conclusions: The Hoshino vaccine strain of mumps virus showed promising oncolytic activities against human fibrosarcoma and cervical adenocarcinoma cells.

Potential contrasting effects of platelets on the migration and invasion of sarcomas versus carcinomas

The ability of platelets to promote carcinoma and melanoma progression has been thoroughly studied and occurs in numerous ways. In contrast, the effect of platelets on sarcomas, tumors arising from mesenchymal cells, has received very little attention. This study was undertaken to simultaneously compare the effects of platelets on murine and human sarcomas and carcinomas. In contrast to their effect on carcinomas, platelets inhibited the invasion of some murine- and all human sarcomas tested in vitro. Further invasion studies with TGFβ treatment only partially recapitulated the results seen with whole platelets. In a spontaneous tumor growth and lung metastasis model, platelets promoted 4T1 mammary carcinoma metastasis but not MCA-1 fibrosarcoma metastasis. Gene expression analysis of the platelet-promoted MDA-MB-231 breast carcinoma, and the platelet-inhibited HT1080 fibrosarcoma cell lines revealed that exposure of MDA-MB-231 to platelets, resulted in upregulation of oncogenes and EMT-associated genes whereas in HT1080 a tumor-suppressor gene was significantly upregulated. Thus, this study has revealed a potential diametrically opposing effect of platelets on mesenchymal and epithelial cancers, a finding that warrants further investigation.

Pro-apoptotic functions of TRAF2 in p53-mediated apoptosis induced by cisplatin.

Tumor necrosis factor receptor-associated factor 2 (TRAF2) is an essential component of tumor necrosis factor-α (TNF-α) signaling that regulates nuclear factor-κB (NF-κB) and c-Jun N-terminal kinase (JNK) pathways, and compelling evidence has demonstrated that TRAF2 suppresses TNF-α-induced cytotoxicity. On the other hand, it has been reported that oxidative stress-induced cytotoxicity is potentiated by TRAF2, indicating that TRAF2 both positively and negatively regulates stress-induced cytotoxicity in a context-specific manner. However, the causal role of TRAF2 in DNA damage response (DDR) remains to be explored. In this study, we assessed the function of TRAF2 in DDR induced by cisplatin, a representative DNA-damaging agent, and found that TRAF2 exerts pro-apoptotic activity through p53-dependent mechanisms at least in human fibrosarcoma cell line HT1080. TRAF2 deficient cells exhibit significant resistance to cell death induced by cisplatin, accompanied by the reduction of both p53 protein level and caspase-3 activation. Moreover, cisplatin-induced JNK activation was attenuated in TRAF2-deficient cells, and pharmacological inhibition of JNK signaling suppressed p53 stabilization. These results suggest that TRAF2 promotes p53-dependent apoptosis by activating the JNK signaling cascade in HT1080 cells. Thus, our data demonstrate a novel function of TRAF2 in cisplatin-induced DDR as a pro-apoptotic protein.

Protect-seq: genome-wide profiling of nuclease inaccessible domains reveals physical properties of chromatin.

In metazoan cell nuclei, heterochromatin constitutes large chromatin domains that are in close contact with the nuclear lamina. These heterochromatin/lamina-associated domains (LADs) domains are difficult to profile and warrants a simpler and direct method. Here we report a new method, Protect-seq, aimed at identifying regions of heterochromatin via resistance to nuclease degradation followed by next-generation sequencing (NGS). We performed Protect-seq on the human colon cancer cell line HCT-116 and observed overlap with previously curated LADs. We provide evidence that these protected regions are enriched for and can distinguish between the repressive histone modification H3K9me3, H3K9me2 and H3K27me3. Moreover, in human cells the loss of H3K9me3 leads to an increase in chromatin accessibility and loss of Protect-seq signal. For further validation, we performed Protect-seq in the fibrosarcoma cell line HT1080 and found a similar correlation with previously curated LADs and repressive histone modifications. In sum, Protect-seq is an efficient technique that allows rapid identification of nuclease resistant chromatin, which correlate with heterochromatin and radial positioning.

SLFN5 suppresses cancer cell migration and invasion by inhibiting MT1-MMP expression via AKT/GSK-3β/β-catenin pathway

Human SLFN5 inhibits invasions of IFNα-sensitive renal clear-cell carcinoma and melanoma cells. However, whether this inhibition is confined to these IFNα-sensitive cancers is unclear. Here we show that SLFN5 expressions on both mRNA and protein levels are significantly higher in non/low-invasive cancer cell lines (breast cancer cell line MCF7, colorectal cancer cell line HCT116 and lung cancer cell line A549) than in highly-invasive cancer cell lines (fibrosarcoma cell line HT1080 and renal clear cell cancer cell line 786-0). SLFN5 knockdown in non/low-invasive cancer cell lines enhanced MT1-MMP expression and increased migration and invasion in vitro, and in vivo. Furthermore, SLFN5 overexpression in HT1080 and 786-0 inhibited MT1-MMP expression and repressed migration and invasion. MT1-MMP is instrumental in SLFN5-controlled inhibition of cancer cell migration and invasion, as shown by MT1-MMP-knockdown and -overexpression analyses. SLFN5 knockdown activated AKT/GSK-3β/β-catenin pathway by promotion AKT phosphorylation and subsequent GSK-3β phosphorylation, further β-catenin translocation into nucleus as un-phosphorylated protein at Ser33, 37 and 45 and Thr41 sites. This is the first study to report that SLFN5 inhibits cancer migration and invasiveness in several common cancer cell lines by repressing MT1-MMP expression via the AKT/GSK-3β/β-catenin signalling pathway, suggesting that SLFN5 plays wide inhibitory roles in various cancers.