L10 Introduction to Biologicals PDF

L10 Introduction to biologicals Introduction to Biologicals Dr Robbert Hoogeboom Lecturer in Lymphoma [email protected]  Learning outcomes Understand what biologica...

L10 Introduction to biologicals Introduction to Biologicals Dr Robbert Hoogeboom Lecturer in Lymphoma [email protected]  Learning outcomes Understand what biologicals are and what types you have Give examples of each type of biological Describe the main mechanisms of action of recombinant proteins, therapeutic antibodies and cellular therapies Understand how biologicals are produced and engineered Explain the main advantages and disadvantages of biologicals What are biologicals? Biologicals: drugs produced in or extracted from a biological source Also known as biopharmaceuticals Sources: Human Animal Fungi Bacteria Plant Biologicals can be: (recombinant) proteins, nucleic acids or living cells Source can be most living organisms e.g. animal, plant, fungi, bacterial 2  Why learn about biologicals? Rank Drug Trade name 2021 sales (billion USD) 1 adalimumab Humira 20.69 2 pembrolizumab Keytruda 17.18 3 lenalidomide Revlimid 12.82 4 apixaban Eliquis 10.76 5 ustekinumab Stelara 9.13 6 aflibercept Eylea 8.62 bictegravir/emtricitabine/ Biktarvy 7 7.81 tenofovir alafenamide 8 Rivaroxaban Xarelto 7.52 9 nivolumab Opdivo 6.94 10 ibrutinib Imbruvica 6.47 11 dulaglutide Trulicity 6.02 12 daratumumab Darzalex 5.97 Adapted from pharmashots.com They are the highest selling drugs. First uses of biologicals - Variolation Smallpox - 400.000 deaths/year in 18th century Europe Variolation - First described in China in the 10th century! - Inserting powdered scabs of smallpox infected individuals into superficial scratches of the skin - Smallpox eradicated in 1980 3  Variolation – inserting pathogen into small scratch in skin to trigger localized immune response. Biologicals are not a new thing James Gillray, 1802 neither is the anti-vaxx movement! 4  Early biologicals – Passive immunisation Diphtheria antitoxin First Nobel Prize in Medicine (1901): - Emil von Behring, “saviour of children” Taking serum provides immune protection. (Recombinant) proteins – The first industrial scale biologicals Proteins isolated from tissue or produced in vitro can be used as drugs when injected Mechanism of action: - Substitute function of protein that is not there or not functioning well Examples: Insulin Erythropoietin Blood factors Interferons Growth Factors 5  Protein used as drug to substitute when the protein lacks that protein. E.g. diabetics taking insulin. Insulin Produced by beta cells in the Islets of Langerhans in the Pancreas Hormone that regulates blood glucose levels Identified by Banting and Best (1920-1921) First patient treated with ox-pancreas extract in 1922: Allergic reaction due to impurity Cow/Pig insulin used as treatment for Type I Diabetes Mellitus until 1982 6  Recombinant insulin Recombinant technology: Biosynthetic human insulin available since 1982 Increased purity Less immunogenicity Less expensive Biosynthetic form reduced risk of immune response to animal proteins or other foreign proteins. Recombinant insulin is much easier to purify. 7  Recombinant proteins can be modified Very easy to introduce favourable mutations in recombinant proteins Development of insulin analogues Lispro (Humalog) does not dimerise, more easily absorbed -> Fast acting Glargine (Lantus) forms aggregates -> long acting Data taken from Diabetes Education Online, www.deo.ucsf.edu Recombinant proteins can be modified e.g. insulin can be modified so it doesn’t dimerise so it works more quickly in a hyperglycaemic patient (insulin lispro, glulisine). Can also be modified to work longer (glargine) replace glutamine with an arginine so it forms aggregates and is absorbed more slowly. 8  Blood factors X-linked recessive mutations in Factor VIII or IX cause haemophilia Blood does not clot Treated with clotting factors purified from human plasma since 1971 Replaced by recombinant form in 1984 Clotting factors isolated to replace the missing ones in haemophilia. Purifying proteins from human sources can have adverse effects Contamination! Plasma of up to 60,000 paid donors pooled together (including drug addicts, inmates) Contaminated with HIV and Hepatitis C 1,200 deaths in UK 9  Donors MUST be screened These vaccines are fully synthetic, so they are not biologicals. 10  Why learn about biologicals? Rank Drug Trade name 2021 sales (billion USD) 1 adalimumab Humira 20.69 2 pembrolizumab Keytruda 17.18 3 lenalidomide Revlimid 12.82 4 apixaban Eliquis 10.76 5 ustekinumab Stelara 9.13 6 aflibercept Eylea 8.62 bictegravir/emtricitabine/ Biktarvy 7 7.81 tenofovir alafenamide 8 Rivaroxaban Xarelto 7.52 9 nivolumab Opdivo 6.94 10 ibrutinib Imbruvica 6.47 11 dulaglutide Trulicity 6.02 12 daratumumab Darzalex 5.97 Adapted from pharmashots.com Above = therapeutic antibody Antibodies specifically bind foreign molecules Antibodies produced as part of the adaptive immune response Variable domain specifically binds to foreign molecules (antigen) Produced by B lymphocyte (B cell) Each B cell produces an antibody with different antigen binding domain When B cells encounter antigen it will produce soluble antibodies 11  Mechanism(s) of action Etanercept/Adalimumab (Anti-TNFα) Dulaglutide Agonist Trastuzumab (Anti-HER2) Tositumomab Rituximab (Anti-CD20) (Anti-CD20-I131) Adapted from Suzuki et al, J Toxicol Pathol 2016 Therapeutic antibodies can be used to mimic endogenous pathways for positive effects or block ligands from binding to receptors. E.g. trastuzumab prevents HER2 binding to cancer cells. Dulaglutide acts as an agonist at insulin receptors. Complement-dependent cytotoxicity – generate antibody to receptor and label this to attract chemotherapeutic agent. Antibodies can bind to targets to trigger destruction in the immune system. 12  Antibodies can also be used to treat infections (antisera) By Ant, The Cartoon Movement Antibodies can be used to bind to infective agents such as viruses to stop them being able to bind to their target receptors. 13  Regen-COV – anti-COVID antibody cocktail https://www.nature.com/articles/d43747-020-01115-y Antibodies bind to coronavirus particles to stop them binding to their target receptors on lung epithelial cells. 14  Development of monoclonal antibodies Immune system of mouse sees antigen as foreign and produces antibodies against it. Can also be used for experimental purposes (identifying proteins)> 15  Evolution of therapeutic antibodies Example: Few approved Rituximab Ocrelizumab Ofatumumab (65% Human) (95% Human) (100% Human) Risk: production of anti-mouse antibodies. You only need the part of the protein that binds the ligand to be the same as the model. You can genetically engineer the mouse to encode human antibodies. 16  Next generation therapeutic antibodies They can be used to deliver cytotoxic drugs. If you want to block something, then you don’t want it to be recognized by the immune system so in that case it may be better to have a fragment of the antibody. If you do want the immune system to recognize it, then it is better to include features such as glycosylation sites to avoid being recognized by complement. 17  What type of antibody are these blockbusters? Drug Trade name Type adalimumab Humira nivolumab Opdivo pembrolizumab Keytruda rituximab Rituxan, MabThera What type of biological are these blockbusters? Drug Trade name Type adalimumab Humira Human antibody nivolumab Opdivo Human antibody pembrolizumab Keytruda Humanised Antibody rituximab Rituxan, MabThera Chimeric antibody 18  Cellular therapies Bone marrow transplantation Transfer of haematopoietic stem cells to treat blood cancers (Leukaemia, Lymphoma, Myeloma) and other blood disorders (anemia) Patient’s own cells (autologous) or donor-derived (allogenic) Stem cell transplantation Few approved uses CAR T cells Immune cells trained to kill tumours You can substitute cancerous bone marrow for healthy bone marrow. Autologous (patient uses their own bone marrow from an unaffected part of their body). Allogenic = from donor. 19  CAR T cells CAR: Chimeric antigen receptor Problem: immune cells do not recognize cancer as foreign. CAR T cells, take target on cancer cells. Find antibody that binds that target, fuse this onto receptors on T cells that recognize foreign targets. Bind these together so it forms a unique receptor that signals like a t cell antigen receptor but binds to your intended target. 20  Production of autologous CAR T cells T cells extracted and sent to lab to have chimeric antigen receptor made. Antibodies against their form of cancer is manufactured, and fused onto the T cell as a chimeric antigen receptor. These are then grown under sterile conditions, and transferred back into the patient. 21  Allogenic CAR T cell design CAR recognising CD19 expressed by B cell leukaemia and lymphoma Kill switch Deplete patient T cells with anti-CD52 antibody to prevent Host vs Graft Disease Eliminate TCR to prevent Graft vs Host Disease Problem with allogenic – can cause immune reactions as the t cells would be foreign to the patient. T cells must be treated with anti-CG52. Treatment can be stopped by adding rituximab recognition domains (kill switch). 22  Advantages of biologicals Highly specific, less off-target effects Well-tolerated (as they look just like endogenous proteins) Can perform functions that cannot be mimicked with small molecules Replace proteins Specifically activate immune cells Off target effects due to receptors in unexpected places. Disadvantages of biologicals Biologicals are enormous compared too small molecules Poor absorption in gut Need to be injected Biologicals more complex May be a mixture Multiple functional domains Can only target extracellular protein/molecule Production via mammalian cells is a laborious and slow process Costly Small scale of production (few kg/year) Shorter shelf-life 23  One dose is around £20,000 They need to be stored very carefully as the protein will degrade. Pharmacology of biologicals Variable half-life Short for insulin Very long for therapeutic antibodies Unquantifiable for cellular therapies Clearance by intracellular lysosomal degradation Efficacy may depend on proteins/cells of patient Complement/Immune cells Anti-drug antibodies Patient T cells may be less functional Safety Less toxicity (only target-related adverse effects) No drug-drug interactions Increased immunogenicity (allergic reactions) Some must expand in order to work (cellular therapies). Proteins are degraded in lysosomes so every cell would be able to metabolise the drug. 24  Biologicals have a different regulatory status Very difficult to generate products that are always the same Batch-to-batch variation Not all components of a biologic may be known No time to characterise all possible components in a cell mixture Production process has to be the same every time, not the product Production process part of the patent Biosimilars, the generic variant of biologicals Not necessary to be identical as long as safety and efficacy is the same Further reading Hill & Rang, Drug Discovery and Development, 2012. Chapter 3: Therapeutic Modalities, p33-40 Greenstein and Brook, Biological Therapeutics, 2011. Chapters 1 and 2 Suzuki M, Kato C, Kato A. Therapeutic antibodies: their mechanisms of action and the pathological findings they induce in toxicity studies. J Toxicol Pathol. 2015;28(3):133–139. doi:10.1293/tox.2015-0031 Castelli MS, McGonigle P, Hornby PJ. The pharmacology and therapeutic applications of monoclonal antibodies. Pharmacol Res Perspect. 2019 Dec; 7(6): e00535. doi: 10.1002/prp2.535 25