scholarly journals Preparation of an anti-NEK2 monoclonal antibody and its application in liver cancer

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Qiuli Chen ◽  
Hui Li ◽  
Lichao Yang ◽  
Sha Wen ◽  
Xuejing Huang ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Monoclonal Antibody ◽  
Liver Cancer ◽  
Cell Line ◽  
Prokaryotic Expression ◽  
Gene Sequence ◽  
Affinity Constant ◽  
Enzyme Linked Immunosorbent Assay ◽  
Prokaryotic Expression System ◽  
Hcc Cells

Abstract Background Never in mitosis gene-A (NIMA)-related expressed kinase 2 (NEK2) is a serine/threonine protein kinase regulated by the cell cycle. The purpose of this study was to obtain NEK2 protein to prepare an anti-NEK2 monoclonal antibody (mAb) and explore the application of the anti-NEK2 mAb of therapeutic and diagnostic in hepatocellular carcinoma (HCC). Results The NEK2 gene sequence was cloned from the normal liver cell line HL7702, and the full-length NEK2 gene sequence was cloned into the prokaryotic expression vector pET30a and transformed into Escherichia coli BL21 (DE3) cells. The recombinant fusion protein was obtained under optimized conditions and injected in BALB/c mice to prepare an anti-NEK2 mAb. By screening, we obtained a stable hybridoma cell line named 3A3 that could stably secrete anti-NEK2 mAb. Anti-NEK2 3A3 mAb was purified from ascites fluid. The isotype was IgG1, and the affinity constant (Kaff) was 6.0 × 108 L/mol. Western blot, indirect enzyme-linked immunosorbent assay (iELISA), immunofluorescence and immunocytochemical analyses showed that the mAb could specifically recognize the NEK2 protein. MTT assays showed that the mAb 3A3 could inhibit the proliferation of HCC cells. KEGG pathway analysis showed that NEK2 might affected pathways of the cell cycle. Moreover, NEK2-related genes were mainly enriched in the S and G2 phases and might act as tumor-promoting genes by regulating the S/G2 phase transition of HCC cells. Conclusions An anti-NEK2 mAb with high potency, high affinity and high specificity was prepared by prokaryotic expression system in this study and may be used in the establishment of ELISA detection kits and targeted treatment of liver cancer.

scholarly journals Anticancer and Immunomodulatory Activities of a Novel Water-Soluble Derivative of Ellipticine

Molecules ◽  
2020 ◽  
Vol 25 (9) ◽  
pp. 2130
Author(s):  
Regiane Costa de Oliveira ◽  
Gemilson Soares Pontes ◽  
Aleksandr Kostyuk ◽  
Gabriel B. Coutinho Camargo ◽  
Anamika Dhyani ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Line ◽  
K562 Cell ◽  
Water Soluble ◽  
Health Concern ◽  
Enzyme Linked Immunosorbent Assay ◽  
Immunomodulatory Activity ◽  
K562 Cell Line ◽  
Human Pbmc ◽  
Cancerous Cells

Cancer still remains a major public health concern around the world and the search for new potential antitumor molecules is essential for fighting the disease. This study evaluated the anticancer and immunomodulatory potential of the newly synthetized ellipticine derivate: sodium bromo-5,11-dimethyl-6H-pyrido[4,3-b]carbazole-7-sulfonate (Br-Ell-SO3Na). It was prepared by the chlorosulfonation of 9-bromoellipticine. The ellipticine-7-sulfonic acid itself is not soluble, but its saponification with sodium hydroxide afforded a water-soluble sodium salt. The cytotoxicity of Br-Ell-SO3Na was tested against cancerous (K562 cell line) and non-cancerous cells (Vero cell line and human peripheral blood mononuclear cells (PBMC)) using a Methylthiazoletetrazolium (MTT) assay. Cell cycle arrest was assessed by flow cytometry and the immunomodulatory activity was analyzed through an enzyme-linked immunosorbent assay (ELISA). The results showed that the Br-Ell-SO3Na molecule has specific anticancer activity (IC50 = 35 µM) against the K562 cell line, once no cytotoxicity effect was verified against non-cancerous cells. Cell cycle analysis demonstrated that K562 cells treated with Br-Ell-SO3Na were arrested in the phase S. Moreover, the production of IL-6 increased and the expression of IL-8 was inhibited in the human PBMC treated with Br-Ell-SO3Na. The results demonstrated that Br-Ell-SO3Na is a promising anticancer molecule attested by its noteworthy activity against the K562 tumor cell line and immunomodulatory activity in human PBMC cells.

scholarly journals Verbesina encelioides: cytotoxicity, cell cycle arrest, and oxidative DNA damage in human liver cancer (HepG2) cell line

2016 ◽  
Vol 16 (1) ◽  
Author(s):  
Mai M. Al-Oqail ◽  
Maqsood A. Siddiqui ◽  
Ebtesam S. Al-Sheddi ◽  
Quaiser Saquib ◽  
Javed Musarrat ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Liver Cancer ◽  
Cell Cycle Arrest ◽  
Cell Line ◽  
Human Liver ◽  
Oxidative Dna Damage ◽  
Hepg2 Cell Line ◽  
Cycle Arrest ◽  
Human Liver Cancer

An Approach to Treatment of Liver Cancer by Novel Glycyrrhizin Derivative

2019 ◽  
Vol 19 (15) ◽  
pp. 1863-1873 ◽  
Author(s):  
Fardous F. El-Senduny ◽  
Mahmoud M. Zidane ◽  
Magdy M. Youssef ◽  
Farid A. Badria
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Cycle ◽  
Methyl Ester ◽  
Liver Cancer ◽  
Cell Line ◽  
Cancer Cell ◽  
Cancer Cell Line ◽  
Normal Lung ◽  
Liver Cancer Cell ◽  
Liver Cancer Cell Line

Background: Liver cancer is a life threating disease as it is the fifth most common cancer and the third most common cause of death worldwide with no safe, efficient, and economic drug available for treatment. Methods: This study intended to investigate glycyrrhizin and its derivatives for possible use as a cytotoxic agent and as a drug for liver cancer treatment. Thus, after treatment of liver cancer cell line HepG-2 with 50 μM of each compound, cell viability was determined. Results: The cytotoxicity assay showed glycyrrhizin derivatives ME-GA (18β-Glycyrrhetinic-30-methyl ester) and AKBA (3-acetyl-11- keto-β-Boswellic acid) to be the most potent drug against liver cancer cell line HepG-2 with IC50 values 25.50 ± 1.06 and 19.73 ± 0.89 μM, respectively. Both the compounds showed higher selectivity towards hepatocellular carcinoma rather than the normal lung fibroblast cell line WI-38. The presence of methyl ester at C-30 greatly increased the cytotoxicity of ME-GA which might be attributed to its higher activity and selectivity. Both ME-GA and AKBA contributed to inhibit cancer cell migration in the wound healing assay and impeded colony formation. The use of flow cytometry to carry out cell cycle analysis and the determination of possible mechanisms of action for apoptosis revealed that ME-GA arrested the cell cycle at G2/M that led to the inhibition of hepatocellular carcinoma and induced apoptosis via the extrinsic pathway and its ability to increase p53 transactivation. Conclusion: This work highlights the cytotoxicity of glycyrrhizin and its derivatives for possible use as a chemotherapeutic agent against hepatocellular carcinoma cells HepG-2. The most cytotoxic compound was ME-GA (18β-Glycyrrhetinic-30-methyl ester) with no cytotoxic effect on the normal cell line. In summary, this new derivative may be used as an alternative or complementary medicine for liver cancer.

5-Nitro-Thiophene-Thiosemicarbazone Derivatives Present Antitumor Activity Mediated by Apoptosis and DNA Intercalation

2019 ◽  
Vol 19 (13) ◽  
pp. 1075-1091 ◽  
Author(s):  
Karla Mirella Roque Marques ◽  
Maria Rodrigues do Desterro ◽  
Sandrine Maria de Arruda ◽  
Luiz Nascimento de Araújo Neto ◽  
Maria do Carmo Alves de Lima ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Antitumor Activity ◽  
Cell Line ◽  
Mitochondrial Membrane ◽  
In Vitro Cytotoxicity ◽  
Dna Intercalation ◽  
Phase Cell ◽  
Fluorescence Studies ◽  
In Silico Studies

Background: Considering the need for the development of new antitumor drugs, associated with the great antitumor potential of thiophene and thiosemicarbazonic derivatives, in this work we promote molecular hybridization approach to synthesize new compounds with increased anticancer activity. Objective: Investigate the antitumor activity and their likely mechanisms of action of a series of N-substituted 2-(5-nitro-thiophene)-thiosemicarbazone derivatives. Methods: Methods were performed in vitro (cytotoxicity, cell cycle progression, morphological analysis, mitochondrial membrane potential evaluation and topoisomerase assay), spectroscopic (DNA interaction studies), and in silico studies (docking and molecular modelling). Results: Most of the compounds presented significant inhibitory activity; the NCIH-292 cell line was the most resistant, and the HL-60 cell line was the most sensitive. The most promising compound was LNN-05 with IC50 values ranging from 0.5 to 1.9 µg.mL-1. The in vitro studies revealed that LNN-05 was able to depolarize (dose-dependently) the mitochondrial membrane, induceG1 phase cell cycle arrest noticeably, promote morphological cell changes associated with apoptosis in chronic human myelocytic leukaemia (K-562) cells, and presented no topoisomerase II inhibition. Spectroscopic UV-vis and molecular fluorescence studies showed that LNN compounds interact with ctDNA forming supramolecular complexes. Intercalation between nitrogenous bases was revealed through KI quenching and competitive ethidium bromide assays. Docking and Molecular Dynamics suggested that 5-nitro-thiophene-thiosemicarbazone compounds interact against the larger DNA groove, and corroborating the spectroscopic results, may assume an intercalating interaction mode. Conclusion: Our findings highlight 5-nitro-thiophene-thiosemicarbazone derivatives, especially LNN-05, as a promising new class of compounds for further studies to provide new anticancer therapies.

Sign in / Sign up

Export Citation Format

Share Document