From DC18 to MR07: A Metabolically Stable 4,4'-Oxybisbenzoyl Amide as a Low-Nanomolar Growth Inhibitor of P. falciparum.

To improve the metabolic stability of a 4,4'-oxybisbenzoyl-based novel and potent (nanomolar-range IC50 ) antiplasmodial agent previously described by us, in silico-guided structure-activity relationship (SAR) campaigns have been conducted to substitute its peptide decorations with more metabolically stable residues. The effects of the various structural modifications were then correlated with the antiplasmodial activity in vitro in phenotypic assays. Among the several derivatives synthetized and compared with the 3D-pharmacophoric map of the original lead, a novel compound, characterized by a western tert-butyl glycine residue and an eastern 1S,2S-aminoacyclohexanol, showed low-nanomolar-range antiplasmodial activity, no signs of cross-resistance and, most importantly, 47-fold improved Phase I metabolic stability when incubated with human liver microsomes. These results highlight the efficacy of in silico-guided SAR campaigns which will allow us to further optimize the structure of the new lead aiming at testing its efficacy in vivo using different routes of administration.

Effects of Vitamin C Combined with Growth Inhibitors on Gastrointestinal Bleeding in Cirrhosis.

The purpose of this study was to investigate the effect of vitamin C combined with growth inhibitors on serum miR-130a, nitric oxide (NO), and hemostasis in the treatment of upper gastrointestinal bleeding (UGIB) in cirrhosis. Eighty patients with cirrhosis UGIB treated in our hospital from March 2021 to March 2022 were selected and divided into two groups using the random number table method. The control group received growth inhibitor treatment, while the observation group was given vitamin C combined with growth inhibitor treatment for 3 d. The hemostatic effect, serum laboratory indexes (miR-130a, NO), liver function indexes (aspartate aminotransferase (AST), alanine aminotransferase (ALT)), adverse effects, and 24 h hemostasis rate were compared between the two groups. The hemostasis time in the observation group was shorter than that in the control group, and the blood transfusion volume was lower than that in the control group. There was no statistical difference regarding the portal blood flow, miR-130a, NO, AST, and ALT indexes between the two groups before treatment. After treatment, the portal blood flow, miR-130a, NO, AST, and ALT indexes in both groups were lower than those before treatment, and all of them were lower in the observation group than in the control group. Adverse reactions showed no significant difference between the two groups of patients with cirrhosis UGIB,, while the 24 h hemostasis rate in the observation group (97.50%) was significantly higher than that in the control group (77.50%). Vitamin C combined with growth inhibitor was effective in the treatment of cirrhotic UGIB, which could effectively shorten the hemostasis time, reduce the transfusion volume and portal blood flow, and improve miR-130a, NO, and liver function levels of patients, with higher safety, and is worthy of clinical promotion.

Daidzein suppresses TGF-ß1-induced cardiac fibroblast activation via the TGF-ß1/SMAD2/3 signaling pathway.

Myocardial fibrosis is a concomitant bioprocess associated with many cardiovascular diseases (CVDs). Daidzein is an isoflavone that has been used for the treatment of CVDs. This study aimed to reveal its role in myocardial fibrosis. Our results indicate that daidzein had a nontoxic effect on cardiac fibroblasts and that TGF-ß1 and TGFßRI levels were gradually decreased by daidzein in a dose-dependent manner. In the current study, we show that daidzein significantly inhibited TGF-ß1-induced mRNA and protein expression of α-SMA, collagen I, and collagen III. Accordingly, immunofluorescence staining of α-SMA was performed. Daidzein also inhibited TGF-ß1-induced cardiac fibroblast proliferation and migration. Mechanistically, daidzein inhibited the TGF-ß/SMAD signaling pathway induced by TGF-ß1 in cardiac fibroblasts. Additionally, daidzein ameliorated MI-induced cardiac dysfunction and cardiac fibrosis in vivo. Based on these findings, we conclude that daidzein reduces TGF-ß1-induced cardiac fibroblast activation by partially regulating the TGF-ß1/SMAD2/3 signaling pathway.

A novel biphenyl diester derivative, AB38b, inhibits glioblastoma cell growth via the ROS-AKT/mTOR pathway.

AB38b is a novel biphenyl diester derivative synthesized in our laboratory, and it has been shown to improve the pathology of nephropathy and encephalopathy in diabetic mice. Glioblastoma (GBM) is the most lethal brain tumor, without effective drugs to date. The present study aims at investigating the role of AB38b in GBM growth and revealing the underlying molecular mechanisms. We found that AB38b administration showed a dose- and time-dependent inhibition on cell proliferation in multiple immortalized and primary GBM cell lines, but it had no significant effects on human astrocyte cell line. More importantly, AB38b blocked cell cycle progression, induced early apoptosis, decreased the activity of AKT/mTOR pathway, and increased the generation of reactive oxygen species (ROS) in GBM cells. Interestingly, antioxidant treatments could reverse the AB38b-mediated abovementioned effects; overexpression of constitutively active AKT could partially rescue the suppressive effects of Ab38b on GBM cell proliferation. In addition, AB38b administration inhibited the tumor growth, decreased the activity of AKT/mTOR pathway, and prolonged the survival time in GBM animal models, without any adverse influences on the important organs. These findings suggest that AB38b exerts anti-glioma activity via elevating the ROS generation followed by inhibiting the activity of AKT/mTOR pathway.

Cediranib, a pan-inhibitor of vascular endothelial growth factor receptors, inhibits proliferation and enhances therapeutic sensitivity in glioblastoma cells.

AIMS: Glioblastoma (GB) is the most aggressive type of brain tumor. Rapid progression, active angiogenesis, and therapy resistance are major reasons for its high mortality. Elevated expression of members of the vascular endothelial growth factor (VEGF) family suggests that anti-VEGF therapies may be potent anti-glioma therapeutic approaches. Here, we evaluated the anti-tumor activity of cediranib, a pan inhibitor of the VEGF receptors, on GB cells. MATERIALS AND METHODS: Anti-proliferative effects of cediranib were determined using MTT, crystal-violet staining, clonogenic and anoikis resistance assays. Apoptosis induction was assessed by Annexin V/PI staining and Western blot analysis and aggressive abilities of GB cells were investigated using cell migration/invasion assays and zymography. Small-interfering RNA (siRNA)-mediated Knockdown was used to study resistance mechanisms. The anti-proliferative and apoptotic effects of cediranib in combination with radiotherapy, temozolomide, bevacizumab were also evaluated using MTT, Annexin V/PI staining and Western blot analysis for cleaved PARP-1. KEY FINDINGS: Cediranib reduced GB cell proliferation, induced apoptotic cell death and inhibited the aggressive abilities of GB cells. Cediranib synergistically increased the anti-proliferative and apoptotic effects of radiotherapy and bevacizumab and augmented the sensitivity of GB cells to temozolomide chemotherapy. In addition, knockdown of MET and AKT potentiated cediranib sensitivity in cediranib-resistant GB cells. SIGNIFICANCE: These findings suggest that cediranib, alone or in combination with other therapeutics, is a promising strategy for the treatment of GB and provide a rationale for further investigation of the therapeutic potential of cediranib for the treatment of this fatal malignancy.

Improvement of PD-1 Blockade Efficacy and Elimination of Immune-Related Gastrointestinal Adverse Effect by mTOR Inhibitor.

During the past decades, immunotherapy, especially the antibody-mediated immune checkpoint blockade (ICB) has shown durable tumor inhibition and changed the paradigm of cancer treatment. However, a growing body of evidence suggests that ICB treatment induces severe immune-related adverse events (irAEs), and the side effect even leads to the discontinuation of lifesaving treatment. Here, we found that ICB treatment induces colitis in melanoma patients and promotes the infiltration of CD8+ effector T cells into colitic lesions. Further transcriptomic dissection indicated the PI3K-AKT-mTOR pathway was highly activated in CD8+ effector T cells of colitic lesions. Moreover, we developed a mouse melanoma model to recapitulate the gastrointestinal toxicity of anti-PD-1 treatment in clinical settings. Anti-PD-1 treatment significantly contributed to the infiltration of CD8+ T cells, and correspondingly induced severe enteritis. Immunohistochemistry experiments showed that the PI3K-AKT-mTOR pathway of T cells was activated by anti-PD-1 treatment. Blockade of the pathway with mTOR inhibitor sirolimus not only inhibits tumor growth but also suppresses the T cell infiltration in colitic lesions. More importantly, combination with sirolimus and anti-PD-1 synergistically inhibits tumor growth via inducing the immunogenic cell death of tumor cells in vivo. In summary, our research demonstrated the principle of mTOR inhibitor and anti-PD-1 combinatorial therapeutic regimen, which provided a novel therapeutic strategy for irAEs in clinics.

Targeting glutamine metabolism as an effective means to promote allograft acceptance while inhibit tumor growth.

Recently the role of metabolic signaling pathways has emerged as playing a critical role in dictating the outcome of T cell responses. The uptake and metabolism of the amino acid glutamine is essential for effector T cell activation. Since the growth and expansion of tumor cells relies on similar anabolic and metabolic requirements, we hypothesized that glutamine blockage might represent a promising strategy to promote allograft survival while inhibit tumor growth. 6-Diazo-5-oxo-L-norleucine (DON) was used as a glutamine antagonist. First, an in vitro study of T cell proliferation was performed to examine the ability of glutamine antagonism to inhibit T cell proliferation. Then we investigated whether DON could prolong allograft survival and inhibit tumor growth by using a fully MHC-mismatched mice full thickness skin transplantation model and a mice TC-1 tumor-bearing model. The proliferation study demonstrated that DON inhibited effector T cells proliferation in a dose-dependent manner. We found a marked prolonged graft median survival time and significant tumor inhibition for mice that received DON compared to those that received no treatment. These results highlight that targeting glutamine metabolism can promote allograft acceptance in a long tumor-free period.

A novel small molecule STAT3 inhibitor SLSI-1216 suppresses proliferation and tumor growth of triple-negative breast cancer cells through apoptotic induction.

Triple-negative breast cancer (TNBC) is the most aggressive type of breast cancer, characterized by the lack of expression of estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2. Owing to the absence of molecular targets, there are limited treatment options, and TNBC patients exhibit high mortality rates. Signal transducer and activator of transcription 3 (STAT3) is overexpressed and aberrantly activated in TNBC cells. Therefore, inhibition of STAT3-mediated signaling provides a potential strategy for the treatment of TNBC. In this study, A series of synthetic derivatives of SLSI-1 (a STAT3 inhibitor) were designed and evaluated for antitumor activity in TNBC cells. A novel derivative (SLSI-1216) exhibited the most potent anti-proliferative activity. SLSI-1216 effectively inhibited STAT3 activity and activation of STAT3, leading to the downregulation of AXL, a downstream target of STAT3 and epithelial-mesenchymal transition (EMT) progression. The inhibition of EMT by SLSI-1216 was associated with modulation of E-cadherin and N-cadherin. Furthermore, SLSI-1216 induced apoptosis by targeting STAT3 and effectively inhibited tumor growth in vivo. These findings suggest that SLSI-1216, as a potential inhibitor of STAT3, may be a promising therapeutic agent for TNBC.

Anticancer effects of Daidzein against the human melanoma cell lines involves cell cycle arrest, autophagy and deactivation of PI3K/AKT signalling pathways.

PURPOSE: Melanoma is one of the fatal human malignancies. Its incidence in humans is increasing constantly and therefore there is urgent need to develop effective therapies for its management. This study was therefore undertaken to investigate the anticancer effects of Daidzein on human melanoma cells and also an attempt was made to decipher the underlying mechanisms. METHODS: MTT assay was used to determine the melanoma A-375 cells viability. Αcridine orange (AO)/ Εthidium bromide (EB) and Annexin V/propidium iodide (PI) assays were used to detect the cell apoptosis. Autophagy was detected by electron microscopy and cell cycle analysis was performed by flow cytometry. The protein expression was determined by western blot analysis. RESULTS: The results of MTT assay showed that Daidzein causes significant decrease in the proliferation of the melanoma A-375 cells and showed an IC50 of 18 µM. However, the IC50 of Daidzein was very high against the normal HEMn-LP cells, indicative of low cytotoxicity. Flow cytometry showed significant arrest of the A-375 cells at the G0/G1 phase of the cell cycle. Western blot analysis showed that the molecule suppressed the expression cell cycle regulatory proteins such as cyclin D1, CDK4, CDK6 and p27. DAPI and annexin V/PI staining assays showed that Daidzein prompted apoptosis in A-375 melanoma cells which was concomitant with depletion of Bcl-2, increase of Bax and activation of cleavage of caspase-3 and caspase-9. Electron microscopic analysis showed that the molecule led to the development of autophagosomes in A-375 cells, which was also concomitant with increase in the expression of LC3B II and decrease in the expression of p62. Finally, Daidzein also suppressed the phosphorylation of PI3K and AKT, causing deactivation of the PI3K/AKT signalling pathway. CONCLUSION: Daidzein may prove beneficial in the development of melanoma systemic therapy.

The inhibitory effect of ECG and EGCG dimeric procyanidins on colorectal cancer cells growth is associated with their actions at lipid rafts and the inhibition of the epidermal growth factor receptor signaling.

Colorectal cancer (CRC) is one of the most common cancers worldwide. Epidemiological studies indicate that consumption of fruits and vegetables containing procyanidins is associated with lower CRC risk. This study investigated the capacity of two dimeric procyanidins composed of epicatechin gallate (ECG) or epigallocatechin gallate (EGCG) isolated from persimmons, to inhibit CRC cell growth and promote apoptosis, characterizing the underlying mechanisms. ECG and EGCG dimers reduced the growth of five human CRC cell lines in a concentration (10-60 µM)- and time (24-72 h)-dependent manner, with a 72 h-IC50 value in Caco-2 cells of 10 and 30 µM, respectively. ECG and EGCG dimers inhibited Caco-2 cell proliferation by arresting the cell cycle in G2/M phase and by inducing apoptosis via the mitochondrial pathway. In addition, ECG and EGCG dimers inhibited cell migration, invasion, and adhesion, decreasing the activity of matrix metalloproteinases (MMP-2/9). Mechanistically, ECG and EGCG dimers inhibited the activation of lipid raft-associated epidermal growth factor (EGF) receptor (EGFR), without affecting its localization at lipid rafts. In particular, ECG and EGCG dimers reduced EGFR phosphorylation at Tyr1068 residue, prevented EGFR dimerization and activation upon stimulation, and induced EGFR internalization both in the absence and presence of EGF. Furthermore, ECG and EGCG dimers increased EGFR phosphorylation at Tyr1045 residue, providing a docking site for ubiquitin ligase c-Cbl and induced EGFR degradation by the proteasome. Downstream of EGFR, ECG and EGCG dimers inhibited the activation of the MEK/ERK1/2 and PI3K/AKT signaling pathways, downregulating proteins involved in the modulation of cell survival. In conclusion, ECG and EGCG dimers reduced CRC cell growth by inhibiting EGFR activation at multiple steps, including the disruption of lipid rafts integrity and promoting EGFR degradation. These results shed light on a potential molecular mechanism on how procyanidins-rich diets may lower CRC risk.

Penfluridol: An antipsychotic agent suppresses lung cancer cell growth and metastasis by inducing G0/G1 arrest and apoptosis.

Lung cancer remains the leading cause of cancer mortality because of highly malignant and metastatic potential. The current status of lung cancer treatment is limited, and more treatment options are needed. Interesting, antipsychotic drugs have been reported to show anti-cancer effects. In this present study, we investigated the anticancer potential of penfluridol (PF), an anti-schizophrenic drug, in lung cancer and its underlying mechanism in vitro and in vivo. In vitro, it could inhibit the viability of various lung cancer cells with G0/G1 phase arrest via increasing the expression level of p21/p27 and decreasing the expression levels of cyclin-CDK complex. Meanwhile, cell-cycle arrest causes DNA repair in the nucleus, which was associated with the upregulation of H2A.X and p-H2A.X. Moreover, PF could also decrease mitochondrial membrane potential and increase reactive oxygen species levels in the lung cancer cells. These results implied that PF might induce the mitochondria-mediated intrinsic apoptosis. In addition, PF inhibits the migration and invasion of lung cancer cells via downregulation of FAK-MMP signaling. In vivo, oral administration of PF at concentration of 10 mg/kg inhibited tumor growth in A549 xenograft model. Notably, PF is an approved drug and the price is exceedingly cheap, so this study demonstrates the potential of PF to treat lung cancer.

Chlorogenic acid inhibits esophageal squamous cell carcinoma growth in vitro and in vivo by downregulating the expression of BMI1 and SOX2.

Esophageal squamous cell carcinoma (ESCC) is one of the most common cancers in China, accompanied by an extremely high mortality rate. Chlorogenic acid (CGA) is a small-molecule compound, that has been shown to have a wide range of biological activities, including antitumor. However, the efficacy and molecular mechanism of CGA on ESCC remains unknown. In this study, we confirmed the inhibition of proliferation by CGA in ESCC cells, as well as the reduction of ESCC xenograft volume by CGA in vivo. In addition, CGA also suppressed both the migration and invasion of ESCC cells in vitro. In a carcinogen-induced murine model of ESCC, hyperplasia of the esophagus was slowed by CGA, while mice suffering from ESCC that were treated with CGA had longer survival times than mice in the control group. The measurement of pluripotency factors (BMI1, SOX2, OCT4 and Nanog) that are related to poor prognosis revealed reduced expression of both BMI1 and SOX2, but not of OCT4 or Nanog, in ESCC cells, in both a dose- and time-dependent manner. Together, our initial findings demonstrate that CGA suppresses ESCC progression, downregulates the expression of BMI1 and SOX2, and provide an anti-tumor candidate for ESCC therapy.

VISAGE Reveals a Targetable Mitotic Spindle Vulnerability in Cancer Cells.

There is an unmet need for new antimitotic drug combinations that target cancer-specific vulnerabilities. Based on our finding of elevated biomolecule oxidation in mitotically arrested cancer cells, we combined Plk1 inhibitors with TH588, an MTH1 inhibitor that prevents detoxification of oxidized nucleotide triphosphates. This combination showed robust synergistic killing of cancer, but not normal, cells that, surprisingly, was MTH1-independent. To dissect the underlying synergistic mechanism, we developed VISAGE, a strategy integrating experimental synergy quantification with computational-pathway-based gene expression analysis. VISAGE predicted, and we experimentally confirmed, that this synergistic combination treatment targeted the mitotic spindle. Specifically, TH588 binding to ß-tubulin impaired microtubule assembly, which when combined with Plk1 blockade, synergistically disrupted mitotic chromosome positioning to the spindle midzone. These findings identify a cancer-specific mitotic vulnerability that is targetable using Plk1 inhibitors with microtubule-destabilizing agents and highlight the general utility of the VISAGE approach to elucidate molecular mechanisms of drug synergy.

The Antiproliferative and Apoptotic Effects of Capsaicin on an Oral Squamous Cancer Cell Line of Asian Origin, ORL-48.

Background and Objectives: The antitumor activities of capsaicin on various types of cancer cell lines have been reported but the effect of capsaicin on oral cancer, which is prevalent among Asians, are very limited. Thus, this study aimed to investigate the effects of capsaicin on ORL-48, an oral cancer cell line of Asian origin. Materials and Methods: Morphological changes of the ORL-48 cells treated with capsaicin were analyzed using fluorescence microscopy. The apoptotic-inducing activity of capsaicin was further confirmed by Annexin V-Fluorescein isothiocyanate / Propidium iodide (V-FITC/PI) staining using flow cytometry. In order to establish the pathway of apoptosis triggered by the compound on ORL-48 cells, caspase activity was determined and the mitochondrial pathway was verified by mitochondrial membrane potential (MMP) assay. Cell cycle analysis was also performed to identify the cell cycle phase of ORL-48 cells being inhibited by the capsaicin compound. Results: Fluorescence microscopy exhibited the presence of apoptotic features in capsaicin-treated ORL-48 cells. Apoptosis of capsaicin-treated ORL-48 cells revealed disruption of the mitochondrial-membrane potential, activation of caspase-3, -7 and -9 through an intrinsic apoptotic pathway and subsequently, apoptotic DNA fragmentation. The cell cycle arrest occurred in the G1-phase, confirming antiproliferative effect of capsaicin in a time-dependent manner. Conclusion: This study demonstrated that capsaicin is cytotoxic against ORL-48 cells and induces apoptosis in ORL-48 cells possibly through mitochondria mediated intrinsic pathway resulting in cell cycle arrest.

A suite of phenotypic assays to ensure pipeline diversity when prioritizing drug-like Cryptosporidium growth inhibitors.

Cryptosporidiosis is a leading cause of life-threatening diarrhea in children, and the only currently approved drug is ineffective in malnourished children and immunocompromised people. Large-scale phenotypic screens are ongoing to identify anticryptosporidial compounds, but optimal approaches to prioritize inhibitors and establish a mechanistically diverse drug development pipeline are unknown. Here, we present a panel of medium-throughput mode of action assays that enable testing of compounds in several stages of the Cryptosporidium life cycle. Phenotypic profiles are given for thirty-nine anticryptosporidials. Using a clustering algorithm, the compounds sort by phenotypic profile into distinct groups of inhibitors that are either chemical analogs (i.e. same molecular mechanism of action (MMOA)) or known to have similar MMOA. Furthermore, compounds belonging to multiple phenotypic clusters are efficacious in a chronic mouse model of cryptosporidiosis. This suite of phenotypic assays should ensure a drug development pipeline with diverse MMOA without the need to identify underlying mechanisms.

Growth suppression of human oral cancer cells by candidate agents for cetuximab-side effects.

Cetuximab, an inhibitor of the epidermal growth factor receptor that is used widely to treat human cancers including oral squamous cell carcinoma (OSCC), has characteristic side effects of skin rash and hypomagnesemia. However, the mechanisms of and therapeutic agents for skin rashes and hypomagnesemia are still poorly understood. Our gene expression profiling analyses showed that cetuximab activates the p38 MAPK pathways in human skin cells (human keratinocyte cell line [HaCaT]) and inhibits c-Fos-related signals in human embryonic kidney cells (HEK293). We found that while the p38 inhibitor SB203580 inhibited the expression of p38 MAPK targets in HaCaT cells, flavagline reactivated c-Fos-related factors in HEK293 cells. It is noteworthy that, in addition to not interfering with the effect of cetuximab by both compounds, flavagline has additive effect for OSCC growth inhibition in vivo. Collectively, our results indicate that combination of cetuximab and these potential therapeutic agents for cetuximab-related toxicities could be a promising therapeutic strategy for patients with OSCC.

FS-7 inhibits MGC-803 cells growth in vitro and in vivo via down-regulating glycolysis.

In this study, we investigated the anticancer effects of FS-7, a flavonoid salicylate derivative, in human gastric carcinoma MGC-803 cell line and studied its preliminary anticancer effects. FS-7 displayed greater in vitro cytotoxicity against MGC-803 cell line compared with 5-Fu and had a certain extent of selectivity to cancer cells. The flow cytometry analysis revealed that FS-7 induced apoptosis MGC-803 cells and mainly caused cells arrest in the G2/M phase in a concentration-dependent manner. Additionally, FS-7 inhibited the colony formation and cell migration in a concentration-dependent manner. Notably, FS-7 noticeably down-regulated glycolysis-related protein HIF-1α, HK-II and PFKP expression in a concentration-dependent manner, possibly causing glycolysis inhibition. Importantly, compared with 5-Fu, FS-7 showed better anticancer activity in the MGC-803 xenograft murine tumor models. Collectively, the present study provided a promising anticancer drug candidate for gastric cancer therapy.

Growth-inhibitory effects of TGFαL3-SEB chimeric protein on colon cancer cell line.

BACKGROUND: TGFαL3-SEB chimeric protein is a synthetic protein, which is produced by combining the third loop (L3) of transforming growth factor-α (TGF-α) with staphylococcal enterotoxin type B. To the best of our knowledge, anti-cancer activity of this chimeric protein against colon cancer that overexpresses epidermal growth factor receptor (EGFR) has not yet been studied. Thus, in the present study, the anti-tumor effects of TGFαL3-SEB chimeric protein on HT-29 colon cancer cells were evaluated. MATERIALS AND METHODS: The TGFαL3-SEB chimeric protein was previously designed and cloned in Escherichia coli (E. coli) [1,2]. The level of expression and the purity of this novel protein were examined for further analysis. For this purpose, the cells were treated with different concentrations (25, 50 and 75 µg/ml) of TGFαL3-SEB and then the proliferation was detected using the MTT assay. The apoptosis-inducing potential of TGFαL3-SEB in HT-29 and HEK-293 cells was evaluated by flow cytometry using Annexin V/PI double staining method; in addition, bax/bcl2 mRNA ratio, caspase-3 and caspase-9 activity were also assessed. RESULTS: In the present study, TGFαL3-SEB chimeric protein was produced in E. coli. After effective purification, its growth inhibitory effect was evaluated. Our results indicated that the incubation of HT-29 colon cancer cell with 25, 50 and 75 µg/ml of TGFαL3-SEB for 24 h leads to significant reduction of proliferation in a dose-dependent manner (P < 0.05). Further analysis indicated that exposure of EGFR expressing HT-29 cells to TGFαL3-SEB leads to significant increase of the caspase-3 and caspase-9 activity in a concentration-dependent manner (P < 0.05). Bax/bcl-2 ratio also confirmed that TGFαL3-SEB has the pro-apoptotic effect. Flow cytometry analysis of TGFαL3-SEB treated cells showed that in addition to apoptotic cells, necrotic cells were also increased significantly at the concentration of 25, 50 and 75 µg/ml (P < 0.05). CONCLUSION: In conclusion, our results demonstrated that TGFαL3-SEB chimeric protein induced cell death through both mechanisms of apoptosis and necrosis in HT-29 colon cancer cells. This paper has highlighted that TGFαL3-SEB has the potential to target EGFR expressing cancer cell.

Oncostatin M-induced upregulation of SDF-1 improves Bone marrow stromal cell migration in a rat middle cerebral artery occlusion stroke model.

Bone marrow-derived mesenchymal stem cells (BMSCs) exhibit potential regenerative effects on the injured brain. However, these effects are constrained by their limited ability to migrate to the injured site. Oncostatin M (OSM) has been shown to affect the proliferation and migration of mesenchymal stem cells. Therefore, in the present study, we explored whether OSM improves BMSC migration and secretion of growth factors and cytokines in a rat middle cerebral artery occlusion (MCAO) stroke model. The effect of OSM on the proliferation and apoptosis of rat BMSCs was first assessed in vitro, and the gene and secretion levels of factors related to cell nutrition and migration, such as SDF-1 and VEGF, were detected. To further explore underlying pathways triggered by OSM, BMSCs were treated with OSM in the presence or absence of inhibitors of the STAT3 and ERK pathways. Effects of OSM on SDF-1 expression in astrocytes and BMSC migration were also evaluated. In the rat MCAO model, OSM secretion levels were detected in the brain for up to 72 h after model establishment. Ventricle injection of OSM alone or OSM combined with caudal vein graft of BMSCs was then performed in MCAO stroke rats. After 72 h, production of SDF-1 and grafted BMSCs was detected in the lesion areas of the brain, and the nerve function score was evaluated. We found that the production of OSM continually increased in the brains of MCAO rats from 12 h to 72 h. OSM significantly upregulated SDF-1 in BMSCs via the STAT3 and ERK pathways and significantly promoted the expression of VEGF and MMP-2. OSM also promoted the secretion of SDF-1 in astrocytes through the STAT3 and ERK pathways to in turn enhance BMSC migration. Combination treatment with OSM and BMSCs in MCAO rats increased the migration efficiency of BMSCs in the brain, which significantly improved neurofunctional recovery while reducing the expression of inflammatory mediators and promoting the secretion of nutrition factors. Overall, these results show that OSM is highly expressed in the brains of MCAO stroke rats and can upregulate SDF-1 to promote BMSC migration. Thus, combination treatment with OSM and BMSCs improves the graft efficiency of BMSCs and neurofunctional recovery.

N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) attenuates silicotic fibrosis by suppressing apoptosis of alveolar type II epithelial cells via mediation of endoplasmic reticulum stress.

Damage to alveolar epithelial cells (AECs) caused by long-term inhalation of large amounts of silica dust plays a significant role in the pathology of silicosis. The present study was undertaken to investigate the regulatory mechanism(s) involved in type II AEC damage from silicon dioxide (SiO2) as well as the mechanism(s) related to the prevention of silicosis by the antifibrotic tetra peptide, N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP). The 2-DE results showed that SiO2 induced endoplasmic reticulum (ER) stress in A549 cells. In addition, typical apoptotic characteristics were observed using a transmission electron microscope (TEM) in A549 cells stimulated by SiO2 and in type II AECs from silicotic rats. Mechanistic study showed that both Ac-SDKP and 4-phenylbutyrate (4-PBA), an inhibiter of ER stress, attenuated GRP78, phosphor-PERK, phosphor-eIF2α, CHOP and Caspase-12 protein expression in A549 cells stimulated by SiO2 and in type II AECs from silicotic rats. Treatment with Ac-SDKP and 4-PBA in vivo effectively inhibited collagen deposition in the lungs of silicotic rats. In summary, ER stress is involved in the apoptosis of type II AECs both in vitro and in vivo. Ac-SDKP effectively suppresses SiO2-induced apoptosis in type II AECs by attenuating the Caspase-12 and PERK/eIF2α/CHOP pathway activation caused by ER stress, thus preventing silicotic fibrosis.