In silico prospective analysis of the medicinal plants activity on the CagA oncoprotein from Helicobacter pylori

BACKGROUND Colonization with Helicobacter pylori(H.pylori)has a strong correlation with gastric cancer,and the virulence factor CagA is implicated in carcinogenesis.Studies have been conducted using medicinal plants with the aim of eliminating the pathogen;however,the possibility of blocking H.pylori-induced cell differentiation to prevent the onset and/or progression of tumors has not been addressed.This type of study is expensive and time-consuming,requiring in vitro and/or in vivo tests,which can be solved using bioinformatics.Therefore,prospective computational analyses were conducted to assess the feasibility of interaction between phenolic compounds from medicinal plants and the CagA oncoprotein.AIM To perform a computational prospecting of the interactions between phenolic compounds from medicinal plants and the CagA oncoprotein of H.pylori.METHODS In this in silico study,the structures of the phenolic compounds(ligands)kaempferol,myricetin,quercetin,ponciretin(flavonoids),and chlorogenic acid(phenolic acid)were selected from the PubChem database.These phenolic compounds were chosen based on previous studies that suggested medicinal plants as non-drug treatments to eliminate H.pylori infection.The three-dimensional structure model of the CagA oncoprotein of H.pylori(receptor)was obtained through molecular modeling using computational tools from the I-Tasser platform,employing the threading methodology.The primary sequence of CagA was sourced from GenBank(BAK52797.1).A screening was conducted to identify binding sites in the structure of the CagA oncoprotein that could potentially interact with the ligands,utilizing the GRaSP online platform.Both the ligands and receptor were prepared for molecular docking using AutoDock Tools 4(ADT)software,and the simulations were carried out using a combination of ADT and AutoDock Vina v.1.2.0 software.Two sets of simulations were performed:One involving the central region of CagA with phenolic compounds,and another involving the carboxy-terminus region of CagA with phenolic compounds.The receptor-ligand complexes were then analyzed using PyMol and BIOVIA Discovery Studio software.RESULTS The structure model obtained for the CagA oncoprotein exhibited high quality(C-score=0.09)and was validated using parameters from the MolProbity platform.The GRaSP online platform identified 24 residues(phenylalanine and leucine)as potential binding sites on the CagA oncoprotein.Molecular docking simulations were conducted with the three-dimensional model of the CagA oncoprotein.No complexes were observed in the simulations between the carboxy-terminus region of CagA and the phenolic compounds;however,all phenolic compounds interacted with the central region of the oncoprotein.Phenolic compounds and CagA exhibited significant affinity energy(-7.9 to-9.1 kcal/mol):CagA/kaempferol formed 28 chemical bonds,CagA/myricetin formed 18 chemical bonds,CagA/quercetin formed 16 chemical bonds,CagA/ponciretin formed 13 chemical bonds,and CagA/chlorogenic acid formed 17 chemical bonds.Although none of the phenolic compounds directly bound to the amino acid residues of the K-Xn-R-X-R membrane binding motif,all of them bound to residues,mostly positively or negatively charged,located near this region.CONCLUSION In silico,the tested phenolic compounds formed stable complexes with CagA.Therefore,they could be tested in vitro and/or in vivo to validate the findings,and to assess interference in CagA/cellular target interactions and in the oncogenic differentiation of gastric cells.

Immune response modulation in inflammatory bowel diseases by Helicobacter pylori infection

Many studies point to an association between Helicobacter pylori(H.pylori)infection and inflammatory bowel diseases(IBD).Although controversial,this association indicates that the presence of the bacterium somehow affects the course of IBD.It appears that H.pylori infection influences IBD through changes in the diversity of the gut microbiota,and hence in local chemical characteristics,and alteration in the pattern of gut immune response.The gut immune response appears to be modulated by H.pylori infection towards a less aggressive inflammatory response and the establishment of a targeted response to tissue repair.Therefore,a T helper 2(Th2)/macrophage M2 response is stimulated,while the Th1/macrophage M1 response is suppressed.The immunomodulation appears to be associated with intrinsic factors of the bacteria,such as virulence factors-such oncogenic protein cytotoxin-associated antigen A,proteins such H.pylori neutrophil-activating protein,but also with microenvironmental changes that favor permanence of H.pylori in the stomach.These changes include the increase of gastric mucosal pH by urease activity,and suppression of the stomach immune response promoted by evasion mechanisms of the bacterium.Furthermore,there is a causal relationship between H.pylori infection and components of the innate immunity such as the NLR family pyrin domain containing 3 inflammasome that directs IBD toward a better prognosis.

EFFECT OF DRUG-RESISTANCE REVERSORS ON EXPRESSIONS OF ONCOGENES OR TUMOR SUPPRESSOR ONCOGENES OF HUMAN TUMOR CELL LINES

MIT method was applied to assay the cytotoxicityof three reversors, verapamil (VER), dipyridamole (DPM)and cyclosporin (CSA) in K562, K562/ADM and KB celllines. S-P immunocytochemical technique was applied todetect cxpressions of oncoproteins or tumor suppressoroncoproteins in these tumor cell lines before or aftertreatment with these reversors. Results showed that threereversors were capable of inhibiting to a certain extentgrowth of K562 or KB cell lines and reversing greatlyadriamycin (ADM)-resistance in K562/ADM cell line.DPM and CsA were observed to inhibit, partly or wholly,expressions of p53, p16, bcl-2, p21 or cerbB-2oncoproteins. VER showed whole inhibition ofexpressions of P53, p16, p21 and bcl-2 and partlyexpression of p53 oncoprotein in K562 cell line. Theseresults suggest that growth inhibition in K562 or KB celllines by the reversors may be associated with partial orwhole inhibition or expressions of p53, p16, p21 or c-erbB-2 oncoproteins. Inhibitions of expressions p53, p16,p21 oncoproteins by VER but not DPM or CsA, may berespossible for reversing activity of VER for ADM-resistance in K562/ADM cell line.

人微管蛋白失稳剂Oncoprotein18原核表达载体的构建及表达

目的:原核表达人Oncoprotein18(Op18)蛋白,为 制备抗Op18的mAb作准备.方法:从人皮肤组织中用RT PCR扩增Op18cDNA的全长编码序列,克隆入表达载体pR SETA中,构建Op18的高表达工程菌,并以IPTG诱导表达目 的蛋白,通过亲和层析法纯化表达的Op18融合蛋白.结果: 酶切及测序鉴定证明,获得含有目的基因片段的重组质粒,表 达的Op18融合蛋白以可溶性的形式存在.结论:所获Op18 融合蛋白以可溶性的形式存在,为制备其mAb及进一步研究 Op18在创伤愈合及瘢痕形成中的作用打下了基础.

麻黄果实总黄酮含量的光学分析

近年来我们对麻黄果实进行了较为全面的研究,结果表明该果实具有抗凝血、降血压等作用。为此我们采用以芦丁为标准,用分光光度法对其总黄酮类物质含量进行了测定。

Apoptosis induced by norcantharidin in human tumor cells

INTRODUCTIONThe antitumor activity of norcantharidin (NCTD),the demethylated analogue of cantharidin,wasstudied in the early 1980s in China.NCTD has noside effects on urinary organs which cantharidin hasshown and is easier to synthesize,and it can inhibitthe proliferation of several tumor cell lines as wellas transplanted tumors.Clinical trials with NCTD asa monotherapeutic agent indicated that NCTD hadbeneficial effects in patients with different kinds

Dys-psychological Stress Effect on Expressions of P53 and NFκBp65 in Human Ovarian Carcinoma In Vivo

Objective： To investigate the dys-psychological stress effect on the growth of subcutaneous xenotransplanted tumor in nude mice bearing human epithelium ovarian carcinoma, and the influence on P53 and NFκBp65 expressions. Methods： The subcutaneous tumor xenografts were established by implanting human epithelium ovarian carcinoma tissues into nude mice and the dys-psychological stress model was established with restraint. The mice were randomized into the following four treatment groups with each group six mice respectively： tumor group （group A）, normal saline intraperitoneal injection; tumor with stress group （group B）, normal saline intraperitoneal injection; tumor therapy group （group C）, cisplatin intraperitoneal injection; and tumor therapy with stress group （group D）, cisplatin intraperitoneal injection. The expressions of P53 and NFκBp65 in tumor tissues were determined by Western blotting. Results： The expressions of P53 and NFκBp65 in each restraint group were enhanced compared with the control groups （P0.05）. Conclusion： The dys-psychological stress may induce the high expressions of P53 and NFκBp65 proteins and further promote tumor growth.

The propensity for tumorigenesis in human induced pluripotent stem cells is related with genomic instability

The discovery of induced pluripotent stem cells (iPSCs) is a promising advancement in the field of regenerative medicine. Previous studies have indicated that the teratoma-forming propensity of iPSCs is variable; however, the relationship between tumorigenic potential and genomic instability in human iPSCs (HiPSCs) remains to be fully elucidated. Here, we evaluated the malignant potential of HiPSCs by using both colony formation assays and tumorigenicity tests. We demonstrated that HiPSCs formed tumorigenic colonies when grown in cancer cell culture medium and produced malignancies in immunodeficient mice. Furthermore, we analyzed genomic instability in HiPSCs using whole-genome copy number variation analysis and determined that the extent of genomic instability was related with both the cells′ propensity to form colonies and their potential for tumorigenesis. These findings indicate a risk for potential malignancy of HiPSCs derived from genomic instability and suggest that quality control tests, including comprehensive tumorigenicity assays and genomic integrity validation, should be rigorously executed before the clinical application of HiPSCs. In addition, HiPSCs should be generated through the use of combined factors or other approaches that decrease the likelihood of genomic instability.

The Expression of the Apoptosis-Suppressive Gene bcl-2 in the Tissue of Colon Carcinoma

The role of bcl 2 in the pathogenesis of colorectal tumor were studied. The exp ression of bcl 2 in the colorectal carcinoma and incisional edge tissue of tumo r was detected by using SABC method. The results showed that the positive rate o f bcl 2 was 69.6 % in colorectal carcinoma and 47.6 % in the incisional edge ti ssue respectively, with the difference being very significant ( P =0.001). B cl 2 positive rate was associated with Dukes' stage, but had nothing to do with histological classification. It was concluded that bcl 2 might play a signific ant role in the development of colorectal carcinoma.

API2-MALT1 oncoprotein promotes lymphomagenesis via unique program of substrate ubiquitination and proteolysis

Lymphoma of mucosa-associated lymphoid tissue(MALT lymphoma) is the most common extranodal B cell tumor and accounts for 8% of non-Hodgkin's lymphomas. Gastric MALT lymphoma is the best-studied example and is a prototypical neoplasm that occurs in the setting of chronic inflammation brought on by persistent infection or autoimmune disease. Cytogenetic abnormalities are commonly acquired during the course of disease and the most common is chromosomal translocation t(11;18)(q21;q21), which creates the API2-MALT1 fusion oncoprotein. t(11;18)-positive lymphomas can be clinically aggressive and have a higher rate of dissemination than t(11;18)-negative tumors. Many cancers, including MALT lymphomas, characteristically exhibit deregulated over-activation of cellular survival pathways, such as the nuclear factor-κB(NF-κB) pathway. Molecular characterization of API2-MALT1 has revealed it to be a potent activator of NF-κB, which is required for API2-MALT1-induced cellular transformation, however the mechanisms by which API2-MALT1 exerts these effects are only recently becoming apparent. The API2 moiety of the fusion binds tumor necrosis factor(TNF) receptor associated factor(TRAF) 2 and receptor interacting protein 1(RIP1), two proteins essential for TNF receptor induced NF-κB activation. By effectively mimicking ligand-bound TNF receptor, API2-MALT1 promotes TRAF2-dependent ubiquitination of RIP1, which then acts as a scaffold for nucleating and activating the canonical NF-κB machinery. Activation occurs, in part, through MALT1 moiety-dependent recruitment of TRAF6, which can directly modify NF-κB essential modulator, the principal downstream regulator of NF-κB. While theintrinsic MALT1 protease catalytic activity is dispensable for this canonical NF-κB signaling, it is critical for noncanonical NF-κB activation. In this regard, API2-MALT1 recognizes NF-κB inducing kinase(NIK), the essential upstream regulator of non-canonical NF-κB, and cleaves it to generate a stable, constitutively active fragment. Thus, API2-MALT1 harnesses multiple unique pathways to achieve deregulated NF-κB activation. Emerging data from our group and others have also detailed additional gain-of-function activities of API2-MALT1 that extend beyond NF-κB activation. Specifically, API2-MALT1 recruits and subverts multiple other signaling factors, including LIM domain and actin-binding protein 1(LIMA1) and Smac/DIABLO. Like NIK, LIMA1 represents a unique substrate for API2-MALT1 protease activity, but unlike NIK, its cleavage sets in motion a major NF-κB-independent pathway for promoting oncogenesis. In this review, we highlight the most recent results characterizing these unique and diverse gain-of-function activities of API2-MALT1 and how they contribute to lymphomagenesis.

Potential roles of longan flower and seed extracts for anticancer

Polyphenol-rich plants are known to possess benefits to human health. Recent studies have revealed that many Traditional Chinese Medicines(TCMs) are rich sources of polyphenols and exhibit antioxidant and anti-inflammatory activities, and these TCMs have been shown experimentally to overcome some chronic diseases, including cancer. Longan flowers and seeds, two TCMs traditionally used for relieving pain and urinary diseases, have been revealed in our recent reports and other studies to possess rich amounts of polyphenolic species and exhibit strong anti-oxidant activity, and these could be applied for the treatment of diabetes and cancer. Herein, we review the recent findings regarding the benefits of these two TCMs in the treatment of humancancer and the possible cellular and molecular mechanisms of both substances.

EXPRESSIONS OF ESTROGEN OCCUPIED RECEPTOR（EoR） AND PROGESTERONEOCCUPIED RECEPTOR（PoR） AND C－erbB－2ONCOPROTEININ HUMANNASOPHAR

It is first reported here that estrogen occupied receptor(EoR)and progesterone occupied receptor (RoR)expressed in cancerous tissues (59.57% and 82.98% respectively)and morphologically normal epithlium(50 77.78% and 70-88.89%respectively) in nasoplharyngeal carcinomas(NPCs)with insignificant difference(P>0.05).Positive rates of EoR and PoR increased greatly in clinical stage Ⅲ and Ⅳ, compared with in Ⅱ(P<0.05), and exhibited insignificant difference between female cases and male ones(P>0.05).Positive rate of C-erbB-2 was 19.15% in cancerous cells, and 9.68% in stage Ⅲand 66.67% in Ⅳin NPCs(P<0.05).Significant difference of C-erbB-2expression was observed between bilateral cervical lymph node metastasis(BCLM)and unilateral ones(P<0.05)but not for EoR or PoR(P>0.05)These findings suggest that EoR or PoR may be correlated with aggravation but not genesis and node metastasis in NPCs and that C-erbB-2may be correlated with aggravation and promotion of formation of node metastasis in NPCs.

MD Simulations of the P53 oncoprotein structure: the effect of the Arg273→His mutation on the DNA binding domain

A comparative molecular dynamics (MD) simulation study was performed on the p53 oncoprotein to investigate the effect of the Arg273His (R273H) mutation on the p53→DNA Binding Domain (DBD). The two p53 dimer structures of the wild-type and mutant Arg273His (R273H) were simulated with the same thermodynamic and environmental parameters. The obtained results demonstrate that the induced Arg273His mutation has a considerable effect on the p53→DNA close contact interaction and changes the picture of hydrogen formation. The Arg273His mutation, in some cases, destroys the existing native hydrogen bond, but, in other cases, forms a strong p53→DNA hydrogen bond, which is not proper for the native protein. The MD simulation results illustrate some molecular mechanism of the conformational changes of the Arg273His key amino acid residue in the p53→DNA binding domain, which might be important for the understanding of the physiological functioning of the p53 protein and the origin of cancer.

微管失稳剂oncoprotein18/stathmin基因在深Ⅱ度烧伤愈合前创面及增生性瘢痕中的表达

目的 观察深Ⅱ度烧伤创面愈合前及愈合后增生性瘢痕组织中微管失稳剂oncopro tein18 stathmin(Op18)基因的表达。 方法 提取临床深Ⅱ度烧伤愈合前与愈合后不同时期增生性瘢痕中的总RNA,以GADPH基因表达为内参照 ,采用逆转录酶 多聚酶链式反应 (RT PCR)方法 ,半定量检测上述基因表达水平。 结果 与正常组织比较 ,伤口愈合前Op18基因表达显著降低 (P <0.0 5 ),愈合后表达迅速增高 (P <0.0 5 ),在 4～ 32个月增生性瘢痕中Op18基因稳定高表达。 结论 在伤后早期Op18基因的表达水平降低与细胞增殖、组织修复的需要相适应 。

Stathmin/Oncoprotein18(Op18)基因在人骨肉瘤中的表达及其意义

目的:探讨stathmin/oncoprotein 18(Op18)基因在人骨肉瘤中的表达规律及意义。方法:分别采用免疫组化法和Western blot法检测10例正常骨组织和56例骨肉瘤中stathmin蛋白的表达。结果:免疫组化法检测stathmin蛋白在正常骨组织、低级别骨肉瘤(G1级组)及高级别骨肉瘤(G2级组)中阳性表达率分别为20%、65%、100%。正常骨组织分别与G1级组、G2级组比较,均有显著性差异(P﹤0.05);G1级组与G2级组比较,有显著差异(P<0.05)。Western blot法检测显示,stathmin蛋白在正常骨组织、G1级组、G2级组表达相对值分别为(0.34±0.15)、(0.68±0.21)、(0.90±0.17)。正常骨组织分别与G1级级组、G2级组比较,差异均有显著性(P<0.01),G1级组与G2级组比较,差异有显著性(P<0.01)。结论:Stathmin在骨肉瘤中过表达,与骨肉瘤的发生及发展密切相关。

Transcriptional repression of hDaxx enhanced by adenovirus 12 E1B 55-kDa oncoprotein interacting with hDaxx

Background Daxx has been identified as a nuclear protein that involves in apoptosis and transcriptional repression. Daxx co-localizes with the promyelocytic leukemia (PML) protein and regulates transcription. Human Daxx (hDaxx) is a protein that functions as a transcriptional regulation through its interaction with some DNA-associated proteins. The aim of this study was to explore the transcriptional regulatory effect of hDaxx interacting with adenovirus (Ad) 12 E1B (Ad12E1B) 55-kDa oncoprotein Methods The co-localization of hDaxx-Ad12E1B or hDaxx-PML protein in the nucleus was observed under a confocal microscope Interaction of hDaxx and Ad12E1B was analyzed by yeast two-hybrid assay Direct binding of hDaxx and Ad12E1B was analyzed using coimmunoprecipitation and Western blot in vivo and in vitro The activity of a luciferase reporter gene, which was regulated by an hDaxx modulated thymidine kinase (TK) promoter, was detected in an automat luminometer Results Ad12E1B, which co-localized with hDaxx in the nuclei of G401-CC3 cells, disrupted the co-localization of hDaxx and PML in the PML oncogenic domains (PODs) hDaxx bound directly to Ad12E1B in vivo and in vitro hDaxx interacted with Ad12E1B along its full length Ad12E1B enhanced transcriptional repression activity of hDaxx Conclusion Ad12E1B disrupts the co-localization of hDaxx with PML in PODs and enhances transcriptional repression activity of hDaxx

IMMUNOCYTOCHEMICAL ANALYSIS OF ras RELATED PROTEINS WITH ANTISERUM AGAINST PEPTIDE FRAGMENT OF ras ONCOPROTEIN P21 (56—66) IN HUMAN CANCER TISSUES

The c-ras and its product P21 have been shown to play an important role in the initiation and development of some malignancies. The activation of c-ras may be associated with two mechanisms: (ⅰ) a point mutation of the codon at position 12 or 61 of the genome leading to a single amino acid alteration in the corresponding product P21; (ⅱ) enhanced expression of ras family encode proteins with

Molecular mechanisms of leukemia-associated proteindegradation

Chemical biology,using small molecules as probes to study the cellular signaling network,has developed rapidly in recent years.The interaction between chemistry and biology not only provides new insight into the understanding of cellular activities,but also generates new lead compounds for the treatment of diseases.Transcription factors and kinases such as retinoic acid receptor-alpha(RARα),acute myeloid leukemia 1(AML1),CAAT/enhancer-binding proteinα(C/EBPα),c-myc,and c-abl play important roles in the differentiation of hematopoietic stem/progenitor cells.Abnormalities in these proteins may cause the dysregulation of hematopoi-esis and even the occurrence of leukemia.Ubiquitin-mediated protein degradation represents a critical mechan-ism in regulating the cellular levels and functions of these proteins.Thus,targeting protein degradation has been emerging as an important strategy to conquer malignant diseases.In this review,we will summarize the recent advances in the understanding of the roles of protein degradation in leukemia,with an emphasis on the mechanisms revealed by small molecules.