lecture 4 notes

Lecture 4: 

Why are proteins useful as drugs  

• Proteins catalyses biochemical reactions, form receptors and channels in membranes, provide scaffolding support and has many more functions. Diseases can result when proteins contain mutations or other abnormalities and/or are present in an abnormally high or low concentrations. Therefore by utilising proteins, we can target or use proteins to alleviate diseases.  

Antibodies 

• Large proteins produced by immune cells in response to detecting foreign antigens  

• These directly binds to and block the action of that antigen  

• Antibody therapies be used to eliminate disease-causing proteins 

Making antibodies 

• In the body, B cells recognise foreign antigens through surface antibodies  

• The antigen is then presented to T cells which release cytokines to activate more B cells 

• B cells expand in number and change into memory B cells and plasma cells 

• Plasma cells secrete antibodies that target the original antigen  

Making antibodies in the lab 

• Hybridoma technology --> antibody-producing B cells fused with cancer cell line in lab; enables production of unlimited quantities of monoclonal antibodies against any protein target  

Recombinant DNA technologies  

• Most therapeutic antibodies and proteins are now produced recombinantly in bacterial, yeast or mammalian expression systems  

• Transformation --> amplification --> transfection --> expression  

Recombinant antibody technologies 

• Recombinant DNA technologies enabled lab production of chimeric, humanised and human monoclonal antibodies  

• LC, Fd and Fc domains are cloned from animal and/or human antibodies, then produced from mammalian cells  

Recombinant protein technologies 

• Bioreactors used for large scale production  

• Special culture medias and growth conditions needed to grow large batches of protein/antibody producing cells  

• Must then be purified from the cells or their media  

Antibody purification 

• Done in serum or cell media containing antibodies and mixture of other biological materials  

• Antibodies are purified using chromatography with beads containing protein A; unwanted molecules are washed out, then purified antibodies are eluted off the beads  

Protein purification 

• Secreted proteins are collected in cell media, intracellular proteins needs to undergo cell lysis first  

• Proteins can require more than one chromatography technique to purify; depends on protein structure --> can be done by separation by charges, by size or by polarity 

• Protein drugs require many rounds of filtration to remove cellular and microbial contaminants meaning each step increases the time and costs of manufacturing  

Biologics  

• Antibodies and protein based drugs --> medicine made from living organisms or their product  

• Most are monoclonal antibodies and their conjugates  

• Others are hormones, enzymes and other proteins