Rituximab Molecular Pharming: Designing a vector that yields high monoclonal antibody Rituximab levels through plant transformation
Abstract
Plant transformation for protein expression is a rapidly expanding method in biomedical sciences, providing a means of replicating therapeutic proteins that hold limits in their reproducibility. One category of these proteins is monoclonal antibodies, known for their capabilities in the diagnosis and treatment of cancers and autoimmune diseases, but holding limits in their replicability given they exist as clones from one original, lab-synthesized protein. The importance of these antibodies and the expensive nature of their formulation creates demand for a measure of reproducibility. Rituximab, used in the treatment of leukemia and lymphoma, is an example of this and is used in this experiment. The objective of this project is to design a protein expression vector with a high yield of Rituximab in Nicotiana tabacum using an agrobacterium-mediated molecular cloning system. The first step is engineering the plant expression vector (pCNHP) for the agrobacterium-mediated plant transformation process. An optimized promoter for maximal transcription and an effective terminator, as well as an experimental 2A peptide linker to ensure that the heavy and light chains are expressed in tandem with high expression, were used. The co-expression of P19 and the vector inhibits gene silencing. Gibson Assembly merged DNA constructs and assembles them within a single vector without the use of restriction enzymes. Electroporation was used to introduce the plasmid to GV3101 agrobacterium electro-competent cells. We infiltrated hydroponically-raised Nicotiana tabacum plants with the agrobacterium. To purify the protein, we will utilize Fast Protein Liquid Chromatography (FPLC) to calculate our total soluble protein. The anticipated outcomes for this experiment are implementing a strategy to express as much of Rituximab (used in treatment of lymphoma and leukemia) in a transformed plant as possible. Rituximab is synthetic, so expressing this antibody maximally through plants is a convenient and cost-efficient way to produce the therapeutic antibody wide-scale.
Start Time
16-4-2025 1:30 PM
End Time
16-4-2025 4:00 PM
Presentation Type
Poster
Presentation Category
Science, Technology and Engineering
Student Type
Undergraduate Student
Faculty Mentor
Tianhu Sun
Faculty Department
Biological Sciences
Rituximab Molecular Pharming: Designing a vector that yields high monoclonal antibody Rituximab levels through plant transformation
Plant transformation for protein expression is a rapidly expanding method in biomedical sciences, providing a means of replicating therapeutic proteins that hold limits in their reproducibility. One category of these proteins is monoclonal antibodies, known for their capabilities in the diagnosis and treatment of cancers and autoimmune diseases, but holding limits in their replicability given they exist as clones from one original, lab-synthesized protein. The importance of these antibodies and the expensive nature of their formulation creates demand for a measure of reproducibility. Rituximab, used in the treatment of leukemia and lymphoma, is an example of this and is used in this experiment. The objective of this project is to design a protein expression vector with a high yield of Rituximab in Nicotiana tabacum using an agrobacterium-mediated molecular cloning system. The first step is engineering the plant expression vector (pCNHP) for the agrobacterium-mediated plant transformation process. An optimized promoter for maximal transcription and an effective terminator, as well as an experimental 2A peptide linker to ensure that the heavy and light chains are expressed in tandem with high expression, were used. The co-expression of P19 and the vector inhibits gene silencing. Gibson Assembly merged DNA constructs and assembles them within a single vector without the use of restriction enzymes. Electroporation was used to introduce the plasmid to GV3101 agrobacterium electro-competent cells. We infiltrated hydroponically-raised Nicotiana tabacum plants with the agrobacterium. To purify the protein, we will utilize Fast Protein Liquid Chromatography (FPLC) to calculate our total soluble protein. The anticipated outcomes for this experiment are implementing a strategy to express as much of Rituximab (used in treatment of lymphoma and leukemia) in a transformed plant as possible. Rituximab is synthetic, so expressing this antibody maximally through plants is a convenient and cost-efficient way to produce the therapeutic antibody wide-scale.