Characterization of Vemurafenib-Resistant Melanoma Cell Lines Reveals Novel Hallmarks of Targeted Therapy Resistance

Regardless of the significant improvements in treatment of melanoma, the majority of patients develop resistance whose mechanisms are still not completely understood. Hence, we generated and characterized two melanoma-derived cell lines, primary WM793B and metastatic A375M, with acquired resistance to the RAF inhibitor vemurafenib. The morphology of the resistant primary WM793B melanoma cells showed EMT-like features and exhibited a hybrid phenotype with both epithelial and mesenchymal characteristics. Surprisingly, the vemurafenib-resistant melanoma cells showed a decreased migration ability but also displayed a tendency to collective migration. Signaling pathway analysis revealed the reactivation of MAPK and the activation of the PI3K/AKT pathway depending on the vemurafenib-resistant cell line. The acquired resistance to vemurafenib caused resistance to chemotherapy in primary WM793B melanoma cells. Furthermore, the cell-cycle analysis and altered levels of cell-cycle regulators revealed that resistant cells likely transiently enter into cell cycle arrest at the G0/G1 phase and gain slow-cycling cell features. A decreased level of NME1 and NME2 metastasis suppressor proteins were found in WM793B-resistant primary melanoma, which is possibly the result of vemurafenib-acquired resistance and is one of the causes of increased PI3K/AKT signaling. Further studies are needed to reveal the vemurafenib-dependent negative regulators of NME proteins, their role in PI3K/AKT signaling, and their influence on vemurafenib-resistant melanoma cell characteristics.     

Novel Systemic Treatment Modalities Including Immunotherapy and Molecular Targeted Therapy for Recurrent and Metastatic Head and Neck Squamous Cell Carcinoma

Head and neck squamous cell carcinomas (HNSCCs) are the sixth most common cancers worldwide. More than half of patients with HNSCC eventually experience disease recurrence and/or metastasis, which can threaten their long-term survival. HNSCCs located in the oral cavity and larynx are usually associated with tobacco and/or alcohol use, whereas human papillomavirus (HPV) infection, particularly HPV16 infection, is increasingly recognized as a cause of oropharyngeal HNSCC. Despite clinical, histologic, and molecular differences between HPV-positive and HPV-negative HNSCCs, current treatment approaches are the same. For recurrent disease, these strategies include chemotherapy, immunotherapy with PD-1-inhibitors, or a monoclonal antibody, cetuximab, that targets epidermal growth factor; these therapies can be administered either as single agents or in combination. However, these treatment strategies carry a high risk of toxic side effects; therefore, more effective and less toxic treatments are needed. The landscape of HNSCC therapy is changing significantly; numerous clinical trials are underway to test novel therapeutic options like adaptive cellular therapy, antibody-drug conjugates, new targeted therapy agents, novel immunotherapy combinations, and therapeutic vaccines. This review helps in understanding the various developments in HNSCC therapy and sheds light on the path ahead in terms of further research in this field.     

Discovery and in Vivo Evaluation of Novel RGD-Modified Lipid-Polymer Hybrid Nanoparticles for Targeted Drug Delivery

In the current study, the lipid-shell and polymer-core hybrid nanoparticles (lpNPs) modified by Arg–Gly–Asp(RGD) peptide, loaded with curcumin (Cur), were developed by emulsification-solvent volatilization method. The RGD-modified hybrid nanoparticles (RGD–lpNPs) could overcome the poor water solubility of Cur to meet the requirement of intravenous administration and tumor active targeting. The obtained optimal RGD-lpNPs, composed of PLGA (poly(lactic-co-glycolic acid))–mPEG (methoxyl poly(ethylene- glycol)), RGD–polyethylene glycol (PEG)–cholesterol (Chol) copolymers and lipids, had good entrapment efficiency, submicron size and negatively neutral surface charge. The core-shell structure of RGD–lpNPs was verified by TEM. Cytotoxicity analysis demonstrated that the RGD–lpNPs encapsulated Cur retained potent anti-tumor effects. Flow cytometry analysis revealed the cellular uptake of Cur encapsulated in the RGD–lpNPs was increased for human umbilical vein endothelial cells (HUVEC). Furthermore, Cur loaded RGD–lpNPs were more effective in inhibiting tumor growth in a subcutaneous B16 melanoma tumor model. The results of immunofluorescent and immunohistochemical studies by Cur loaded RGD–lpNPs therapies indicated that more apoptotic cells, fewer microvessels, and fewer proliferation-positive cells were observed. In conclusion, RGD–lpNPs encapsulating Cur were developed with enhanced anti-tumor activity in melanoma, and Cur loaded RGD–lpNPs represent an excellent tumor targeted formulation of Cur which might be an attractive candidate for cancer therapy.     

The Role of E-Cadherin and microRNA on FAK Inhibitor Response in Malignant Pleural Mesothelioma (MPM)

Malignant pleural mesothelioma (MPM) is an aggressive malignancy with limited effective treatment options. Focal adhesion kinase (FAK) inhibitors have been shown to efficiently suppress MPM cell growth initially, with limited utility in the current clinical setting. In this study, we utilised a large collection of MPM cell lines and MPM tissue samples to study the role of E-cadherin (CDH1) and microRNA on the efficacy of FAK inhibitors in MPM. The immunohistochemistry (IHC) results showed that the majority of MPM FFPE samples exhibited either the absence of, or very low, E-cadherin protein expression in MPM tissue. We showed that MPM cells with high CDH1 mRNA levels exhibited resistance to the FAK inhibitor PND-1186. In summary, MPM cells that did not express CDH1 mRNA were sensitive to PND-1186, and MPM cells that retained CDH1 mRNA were resistant. A cell cycle analysis showed that PND-1186 induced cell cycle disruption by inducing the G2/M arrest of MPM cells. A protein−protein interaction study showed that EGFR is linked to the FAK pathway, and a target scan of the microRNAs revealed that microRNAs (miR-17, miR221, miR-222, miR137, and miR148) interact with EGFR 3′UTR. Transfection of MPM cells with these microRNAs sensitised the CHD1-expressing FAK-inhibitor-resistant MPM cells to the FAK inhibitor.     

Novel miRNA Targets and Therapies in the Triple-Negative Breast Cancer Microenvironment: An Emerging Hope for a Challenging Disease

Treatment of triple-negative breast cancer (TNBC) remains challenging because of the heterogeneity of the disease and lack of single targetable driving mutations. TNBC does not rely on estrogen, progesterone or epidermal growth factor receptors and is associated with aggressive disease progression and poor prognosis. TNBC is also characterized by resistance to chemotherapeutics, and response to immunotherapies is limited despite promising results in a subset of TNBC patients. MicroRNAs (miRNAs) have emerged as significant drivers of tumorigenesis and tumor progression in triple-negative breast cancer (TNBC) and present unique opportunities to target various components of the TNBC microenvironment for improved efficacy against this difficult to treat cancer. Effects of miRNAs on multiple targets may improve response rates in the context of this genetically and biologically heterogeneous disease. In this review, we offer a comprehensive view of miRNA regulation in TNBC, treatment challenges presented by TNBC in the context of the tumor microenvironment and stem cell subpopulations, and current and emerging miRNA-based therapeutic strategies targeting various components of the TNBC microenvironment. In addition, we offer insight into novel targets that have potential for treating TNBC through multiple mechanisms in the tumor microenvironment simultaneously and those that may be synergistic with standard chemotherapies.     

Cell Therapy for Colorectal Cancer: The Promise of Chimeric Antigen Receptor (CAR)-T Cells

Colorectal cancer (CRC) is a global public health problem as it is the third most prevalent and the second most lethal cancer worldwide. Major efforts are underway to understand its molecular pathways as well as to define the tumour-associated antigens (TAAs) and tumour-specific antigens (TSAs) or neoantigens, in order to develop an effective treatment. Cell therapies are currently gaining importance, and more specifically chimeric antigen receptor (CAR)-T cell therapy, in which genetically modified T cells are redirected against the tumour antigen of interest. This immunotherapy has emerged as one of the most promising advances in cancer treatment, having successfully demonstrated its efficacy in haematological malignancies. However, in solid tumours, such as colon cancer, it is proving difficult to achieve the same results due to the shortage of TSAs, on-target off-tumour effects, low CAR-T cell infiltration and the immunosuppressive microenvironment. To address these challenges in CRC, new approaches are proposed, including combined therapies, the regional administration of CAR-T cells and more complex CAR structures, among others. This review comprehensively summarises the current landscape of CAR-T cell therapy in CRC from the potential tumour targets to the preclinical studies and clinical trials, as well as the limitations and future perspectives of this novel antitumour strategy.     

PD-1/PD-L1, MDSC Pathways,and Checkpoint Inhibitor Therapy in Ph(-) Myeloproliferative Neoplasm: A Review

There has been significant progress in immune checkpoint inhibitor (CPI) therapy in many solid tumor types. However, only a single failed study has been published in treating Ph(-) myeloproliferative neoplasm (MPN). To make progress in CPI studies on this disease, herein, we review and summarize the mechanisms of activation of the PD-L1 promoter, which are as follows: (a) the extrinsic mechanism, which is activated by interferon gamma (IFN γ) by tumor infiltration lymphocytes (TIL) and NK cells; (b) the intrinsic mechanism of EGFR or PTEN loss resulting in the activation of the MAPK and AKT pathways and then stat 1 and 3 activation; and (c) 9p24 amplicon amplification, resulting in PD-L1 and Jak2 activation. We also review the literature and postulate that many of the failures of CPI therapy in MPN are likely due to excessive MDSC activities. We list all of the anti-MDSC agents, especially those with ruxolitinib, IMID compounds, and BTK inhibitors, which may be combined with CPI therapy in the future as part of clinical trials applying CPI therapy to Ph(-) MPN.     

Effect of electronegativity and steric hindrance of the cocatalyst on the activity and selectivity of butadiene polymerization catalyzed by molybdenum

Molybdenum-based random 1,2-polybutadiene (1,2-PB) rubber, with its excellent wet skid resistance, low rolling resistance, and low heat buildup, can be used for green tread rubber, and it has significant benefits for the enhancement of the mechanical strength of 1,2-PB. In this study, we investigated in detail the effects of the electronegativity and steric hindrance of the cocatalyst on the stereoselectivity of the resulting polymers. In the case of cocatalysts with moderate electronegativities, the active centers were more stable; this resulted in a larger molecular weight and a higher conversion, whereas a cocatalyst with excessive electronegativity tended to initiate cationic polymerization. The high sterically hindered substituent restricted the coordination configuration of the monomer; this ultimately resulted in the increase in the 1,2 structure to 90%. Fourier transform infrared analysis showed that the content of trans-1,4 structure increased from 7 to 66%, and its crystallinity reached 59.2%, as calculated with differential scanning calorimetry when triethyl aluminum was used as the cocatalyst. Consequently, trans-1,4-polybutadiene crystals were introduced into the 1,2-PB elastomer in one step; this could be a promising facile route for strengthening 1,2-PB. © 2018 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2019 , 136, 46906.     

Vimentin Cytoskeleton Architecture Analysis on Polylactide and Polyhydroxyoctanoate Substrates for Cell Culturing

Polylactide (PLA), widely used in bioengineering and medicine, gained popularity due to its biocompatibility and biodegradability. Natural origin and eco-friendly background encourage the search of novel materials with such features, such as polyhydroxyoctanoate (P(3HO)), a polyester of bacterial origin. Physicochemical features of both P(3HO) and PLA have an impact on cellular response 32, i.e., adhesion, migration, and cell morphology, based on the signaling and changes in the architecture of the three cytoskeletal networks: microfilaments (F-actin), microtubules, and intermediate filaments (IF). To investigate the role of IF in the cellular response to the substrate, we focused on vimentin intermediate filaments (VIFs), present in mouse embryonic fibroblast cells (MEF). VIFs maintain cell integrity and protect it from external mechanical stress, and also take part in the transmission of signals from the exterior of the cell to its inner organelles, which is under constant investigation. Physiochemical properties of a substrate have an impact on cells’ morphology, and thus on cytoskeleton network signaling and assembly. In this work, we show how PLA and P(3HO) crystallinity and hydrophilicity influence VIFs, and we identify that two different types of vimentin cytoskeleton architecture: network “classic” and “nutshell-like” are expressed by MEFs in different numbers of cells depending on substrate features.     

Novel Inhibitor for Downstream Targeting of Transforming Growth Factor-β Signaling to Suppress Epithelial to Mesenchymal Transition and Cell Migration

Cancer metastasis accounts for most of the mortality associated with solid tumors. However, antimetastatic drugs are not available on the market. One of the important biological events leading to metastasis is the epithelial to mesenchymal transition (EMT) induced by cytokines, namely transforming growth-factor-β (TGF-β). Although several classes of inhibitors targeting TGF-β and its receptor have been developed, they have shown profound clinical side effects. We focused on our synthetic compound, HPH-15, which has shown anti-fibrotic activity via the blockade of the TGF-β Smad-dependent signaling. In this study, 10 μM of HPH-15 was found to exhibit anti-cell migration and anti-EMT activities in non-small-cell lung cancer (NSCLC) cells. Although higher concentrations are required, the anti-EMT activity of HPH-15 has also been observed in 3D-cultured NSCLC cells. A mechanistic study showed that HPH-15 inhibits downstream TGF-β signaling. This downstream inhibition blocks the expression of cytokines such as TGF-β, leading to the next cycle of Smad-dependent and -independent signaling. HPH-15 has AMPK-activation activity, but a relationship between AMPK activation and anti-EMT/cell migration was not observed. Taken together, HPH-15 may lead to the development of antimetastatic drugs with a new mechanism of action.     

Novel membranes based on poly(5‐(methacrylamido)tetrazole) and sulfonated polysulfone for proton exchange membrane fuel cells

Proton‐exchange membrane fuel cells (PEMFC)s are increasingly regarded as promising environmentally benign power sources. Heterocyclic molecules are commonly used in the proton conducting membranes as dopant or polymer side group due to their high proton transfer ability. In this study, 5‐(methacrylamido)tetrazole monomer, prepared by the reaction of methacryloyl chloride with 5‐aminotetrazole, was polymerized via conventional free radical mechanism to achieve poly(5‐(methacrylamido)tetrazole) homopolymer. Novel composite membranes, SPSU‐PMTetX, were successfully produced by incorporating sulfonated polysulfone (SPSU) into poly(5‐(methacrylamido)tetrazole) (PMTet). The sulfonation of polysulfone was performed with trimethylsilyl chlorosulfonate and high degree of sulfonation (140%) was obtained. The homopolymers and composite membranes have been characterized by NMR, FTIR, thermogravimetric analysis (TGA), and differential scanning calorimetry (DSC). ¹H‐NMR and FTIR confirmed the sulfonation of PSU and the ionic interaction between sulfonic acid and poly(5‐(methacrylamido)tetrazole) units. TGA showed that the polymer electrolyte membranes are thermally stable up to ～190°C. Scanning electron microscopy analysis indicated the homogeneity of the membranes. This result was also supported by the appearance of a single T_g in the DSC curves of the blends. Water uptake and proton conductivity measurements were, as well, carried out. Methanol permeability measurements showed that the composite membranes have similar methanol permeability values with Nafion 112. The maximum proton conductivity of anhydrous SPSU‐PMTet0.5 at 150°C was determined as 2.2 × 10^?6 S cm^?1 while in humidified conditions at 20°C a value of 6 × 10^?3 S cm^?1 was found for SPSU‐PMTet2. © 2013 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2014 , 131, 40107.     

基于动态优化和反馈控制的聚合反应过程牌号切换策略

针对单反应器多牌号聚合物生产过程, 提出一种结合动态优化和反馈控制的牌号切换策略。以实验室规模的连续搅拌釜式反应器中苯乙烯聚合牌号切换为对象, 以原料消耗最少为优化目标, 利用迭代动态规划求得切换过程中反应条件和产品性能指标的优化轨迹。引入针对反应温度的路径约束, 使优化后的切换轨迹更易跟踪实现, 防止过渡过程中变量的剧烈波动。仿真结果表明, 这一切换策略可以显著减少牌号切换过渡时间及过渡过程中原料的消耗量, 并能够有效克服进料温度变化的干扰。     

猪繁殖与呼吸综合征弱毒疫苗的安全性及新型疫苗的研究进展

猪繁殖与呼吸综合征(Porcine reproductive and respiratory syndrome,PRRS)是影响养猪业的重大传染病,应用PRRS弱毒疫苗免疫是主要的预防措施。尽管各种PRRS弱毒疫苗已在我国使用,但PRRS仍持续流行,每年均有不同程度的暴发,且造成巨大的经济损失。最新研究提示,基因缺失疫苗能有效提升猪对PRRS的特异性免疫力。本文就PRRS弱毒疫苗在使用过程中可能存在的安全隐患及新型疫苗的研究进展作一综述。     

人C-反应蛋白重组真核表达质粒的构建及其对人脐静脉内皮细胞LOX-1和TF表达的影响

目的构建人C-反应蛋白(C-reactive protein,CRP)重组表达质粒pTracer CMV2-CRP,并观察其在人脐静脉内皮细胞(Human umbilical vein endothelical cells,HUVEC)中的表达及其对凝集素样氧化型低密度脂蛋白受体-1(Lectin-type oxidizedLDL receptor-1,LOX-1)、组织因子(Tissue factor,TF)表达的影响。方法以质粒pCR-BluntⅡ-TOPO-CRP为模板,PCR扩增CRP基因CDS序列,克隆至pTracer CMV2载体中,转化感受态大肠杆菌DH5α,构建重组表达质粒pTracer CMV2-CRP,转染HUVEC,设实验组(转染pTracer CMV2-CRP)、阴性对照组(转染pTracer-CMV2)及正常对照组,各组细胞经G418抗性筛选,RT-PCR及Western b1ot检测CRP基因的过表达效应及CRP的表达对HUVEC中LOX-1和TF转录及蛋白水平的影响。结果重组真核表达质粒pTracer CMV2-CRP经双酶切鉴定及测序证明构建正确。实验组细胞中,CRP基因过表达,且LOX-1和TF基因的转录及蛋白水平明显高于正常对照组和阴性对照组(P<0.05)。结论已成功构建人CRP基因重组真核表达质粒pTracerCMV2-CRP,并在HUVEC中过表达CRP,且明显上调HUVEC中LOX-1和TF的表达,为进一步阐述CRP在动脉粥样硬化形成过程中的作用提供了新的实验依据。     

氧糖剥夺条件下PC12细胞RAGE的表达及对其损伤的影响

目的探讨大鼠肾上腺嗜铬细胞瘤(PC12)细胞在氧糖剥夺(OGD)的条件下,晚期糖基化终产物受体(RAGE)的表达及其对PC12细胞损伤的影响。方法PC12细胞随机分为3组:OGD培养组(PC12细胞在无血清无糖的DMEM培养液中厌氧培养);封闭RAGE的OGD培养组(PC12细胞在加5μg/mlRAGE抗体的无血清无糖DMEM培养液中厌氧培养);对照组(PC12细胞在无血清无糖DMEM培养液中培养)。免疫组化法检测RAGE表达。收集细胞上清液,检测乳酸脱氢酶(LDH)活性和一氧化氮(NO)含量,并对细胞死亡率进行检测。结果在OGD条件下PC12细胞培养8、11、20h均有RAGE表达,与对照组相比表达均明显增加,OGD培养组与封闭RAGEOGD培养组相比,LDH活性差异显著。随着厌氧时间的延长,PC12细胞死亡率明显增高,封闭RAGEOGD培养组与OGD培养组相比,PC12细胞死亡率明显降低。NO含量差异无显著意义,但与对照组相比差异显著。结论在OGD条件下,PC12细胞RAGE表达增高。RAGE的表达对细胞的损伤起促进作用。     

Selective Anti-Cancer Effects of Plasma-Activated Medium and Its High Efficacy with Cisplatin on Hepatocellular Carcinoma with Cancer Stem Cell Characteristics

Hepatocellular carcinoma (HCC) is a major histological subtype of primary liver cancer. Ample evidence suggests that the pathological properties of HCC originate from hepatic cancer stem cells (CSCs), which are responsible for carcinogenesis, recurrence, and drug resistance. Cold atmospheric-pressure plasma (CAP) and plasma-activated medium (PAM) induce apoptosis in cancer cells and represent novel and powerful anti-cancer agents. This study aimed to determine the anti-cancer effect of CAP and PAM in HCC cell lines with CSC characteristics. We showed that the air-based CAP and PAM selectively induced cell death in Hep3B and Huh7 cells with CSC characteristics, but not in the normal liver cell line, MIHA. We observed both caspase-dependent and -independent cell death in the PAM-treated HCC cell lines. Moreover, we determined whether combinatorial PAM therapy with various anti-cancer agents have an additive effect on cell death in Huh7. We found that PAM highly increased the efficacy of the chemotherapeutic agent, cisplatin, while enhanced the anti-cancer effect of doxorubicin and the targeted-therapy drugs, trametinib and sorafenib to a lesser extent. These findings support the application of CAP and PAM as anti-cancer agents to induce selective cell death in cancers containing CSCs, suggesting that the combinatorial use of PAM and some specific anti-cancer agents is complemented mechanistically.     

湿法磷酸过滤设备和管道结垢原因分析及控制研究

分析湿法磷酸过滤设备与管道结垢的主要原因是氟硅酸钾、钠盐因磷酸液相温度下降而沉积下来。提出采用添加剂,抑制或提前析出氟硅酸钾、钠盐,同时进行了5种添加剂的实验室研究:均有较好的阻垢效果,但最终选择添加剂E(氟硅酸),阻垢率可达70%以上,添加量以0.5g/100g磷矿为宜。     

《材料科学基础（双语）》课程 CDIO 教学模式的研究

根据《材料科学基础（双语）》课程在教学中出现的畏惧语言心理、理论抽象难以理解、不理解实验内涵、理论与实践脱节等问题,提出采用CDIO教学模式对其进行教学改革。在教学过程中,通过克服“畏惧语言”的心理、增强理论分析能力、增强实验内涵的理解以及理论联系实际等教学改革,使得学生对具体理论知识点有了深刻的认识,提高学生的英语水平,充分发挥了学生学习的自主性和能动性,取得了较好的教学效果。     

工业循环冷却水成套水处理技术的研究与应用

中海油天津化工研究设计院设计开发的缓蚀阻垢水处理技术,能够使腐蚀性水质的海水淡化水成功地作为补充水应用于循环冷却水系统,保证循环水稳定运行,确保正常生产。