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Screening and evaluation of anti-glypican 3 antibodies for liver cancer treatment
China (IFMSA-China) - Nanjing Medical University, Nanjing
Department of Cell Biology, School of Basic Medical Sciences
Type of Research Project
- Basic science
What is the background of the project?
Liver cancer is the fifth most prevalent neoplasm worldwide. Both the incidence and mortality of liver cancer are rising. Most patients do not respond to most chemotherapy drugs and are not applicable to radical surgery. The development of antibody-based drugs targeting specific antigens derived from tumor cells is a new promising strategy. In view of this, it has been suggested that glypican-3 (GPC3) represents an attractive target for liver cancer therapy because it is highly expressed in liver cancers. GPC3 is a member of the glypican family of heparan sulfate proteoglycans that are attached to the cell surface. Although our group and others have developed several specific antibodies (such as clone YP7, 1G12, GC33) targeting GPC3, there is still an urgent need for more anti-GPC3 monoclonal antibodies with new characteristics, due to the complex tumor microenvironment or for certain clinical applications. (YP7, 1G12, GC33 are the clone names of monoclonal antibodies)
What is the aim of the project?
To develop new GPC3 antibodies with optimized affinity to GPC3 in tumor.
What techniques and methods are used?
1. Expression of antibodies in eukaryotic cells and purification of protein by protein A agarose column. Protein purification is determined by gel electrophoresis and western blotting. Protein concentration is tested by a colorimetric method using spectrophotometer. (about 10 days) 2. Evaluation of antibody affinity by sandwich enzyme linked immunosorbent assay (ELISA) for molecular level binding in 96-well plate and flow cytometry for binding with GPC3 on cell membrane. (about one week) 3. Evaluation of anti-tumor activity in vitro by Cell Counting Kit-8 (CCK-8) assay. IC50 (half maximal inhibitory concentration) is calculated for compare between different groups. (about one week) 4. Data analysis and presentation with PowerPoint.
What is the role of the student?
- The student will observe the practical experiments but will be highly involved in the analysis of the results
- The tasks will be done under supervision
What are the tasks expected to be accomplished by the student?
1. construction of recombinant plasmid with the antibodies sequence which was determined by phage display experiment. 2.Transformation of the plasmid into E. coli cells, and extraction of plasmid DNA after bacteria culture. 3. Expression of antibodies in eukaryotic cells and purification of protein by protein agarose column. 4. Evaluation of antibody affinity to purified GPC3 protein by enzyme linked immunosorbent assay (ELISA). 5. Evaluation of antibody affinity to GPC3 on cell membrane by flow cytometry. 6. Evaluation of anti-tumor activity in vitro by Cell Counting Kit-8 (CCK-8) assay.
Will there be any theoretical teaching provided (preliminary readings, lectures, courses, seminars etc)
Preliminary readings will be provided by the tutor. In addition, the tutor will discuss and explain in detail about the project that students will be conducting. Students are encouraged to take part in the group meeting and seminars
What is expected from the student at the end of the research exchange? What will be the general outcome of the student?
- The student will prepare a presentation - The student will prepare a scientific report
What skills are required of the student? Is there any special knowledge or a certain level of studies needed?
literature searching, bacteria culture, nucleic acid analysis, data analysis. Subjects passed: biology and biochemistry
Are there any legal limitations in the student’s involvement
Type of students accepted
This project accepts: - Medical students - Graduated students (less than 6 months) - Pre-Medical students from the American-British system - Students in biomedical fields
- Gao W; Tang Z; Zhang YF; Feng M; Qian M; Dimitrov DS; Ho M. Immunotoxin targeting glypican3 regresses liver cancer via dual inhibition of Wnt signalling and protein synthesis. Nat Commun. 2015;11;6:6536.
- Gao W; Xu Y; Liu J; Ho M. Epitope mapping by a Wnt-blocking antibody: evidence of the Wnt binding domain in heparan sulfate. Sci Rep. 2016;17;6:26245.
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