Original Article

Plasma derived versus biotechnological manufactured medicine

Paul F. W. Strengers MD, FFPM. Sanquin Blood Supply, Ams terdam, The Netherlands ,Email: [email protected]

Introduction these two institutes, two US based The research on alternatives for pharmaceutical companies accom- plasma products, whether biotech- plished scale-up, purification and nological developed medicinal prod- standardization of two recombinant ucts or recombinant alternatives to factor VIII products for clinical use. plasma products, with the devel- opment of recombinant factor VIII Recombinant technology started at the end of the 1980s. The first recombinant factor VIII There was considerable interest in products, produced by cell culture, the possibility of producing a ‘syn- were licensed in 1992-1993 and thetic’ form of factor VIII and factor induced a great change in the treat- IX in view of the high rate of trans- ment of patients with mission of blood-borne viruses by A. The high costs of haemophilia plasma derived factor concentrates treatment and the unavailability in the 1970s and early 1980s,. of clotting factor products for de- This objective became reality when veloping countries were stimuli for scientists at the Genetics Insti- researchers to study the basics of tute (Cambridge, Mass., US) and factor proteins. The expectations Downloaded from ijbc.ir at 12:07 +0330 on Sunday September 26th 2021 Genetech (Berkeley, Ca.,US) simul- and beliefs in new biotech products taneously announced the success- at that time were sky-high. While ful cloning of Factor VIII gene. In a many research groups focussed on landmark series of articles in a sin- recombinant factor VIII and IX, only gle issue of Nature in 1984, the two a few products reached the market. groups of scientists described the In 1997, recombinant factor IX was structure of the Factor VIII gene, the licensed, and in the same year re- isolation of cDNA clones encoding combinant factor VIIa. The current the complete factor VIII sequence, products are similar to natural co- and the in vitro expression of human agulation factors, but recombinant factor VIII in tissue culture (1, 2, 3, technology also allows for the de- 4). In collaboration with scientists of sign of bio-engineered proteins with

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improved biological properties. This tein alpha-1 antitrypsin and is also is therefore one of the most prom- in the development phase. The com- ising areas of research in haemo- pany has established founder trans- philia is related to factor VIII protein. genic animals that express recom- Also biotechnological research on binant human alpha-1 antitrypsin . other plasma proteins started and The plasma form of this product has resulted in 2001 in the licensing of been used as a treatment for cer- activated protein C. Since then how- tain forms of emphysema caused ever, on recombinant technology, no by a congenital deficiency of plasma new developments in plasma prod- alpha-1 antitrypsin. Recombinant ucts have reached the market. What human alpha-1 antitrypsin may also has changed is the formulation of be developed as an effective treat- the recombinant products by delet- ment for other diseases, potentially ing human plasma proteins in the including cystic fibrosis, chronic ob- formulations and as stabilisers. The structive pulmonary diseases, acute newest recombinant products lack respiratory syndrome, and severe any human or veterinarian protein. asthma. The research developments in this area are however still on going. Re- Monoclonal antibody based im- combinant is in phase III munoglobulins pre-registration phase, recombinant Monoclonal antibodies are antibody factor XIII in phase I, and recombi- proteins that bind to a specific tar- nant von Willebrand factor in pre- get molecule (antigen) at one spe- clinical phase. Factor VII, protein C cific site (antigenic site). In response

Downloaded from ijbc.ir at 12:07 +0330 on Sunday September 26th 2021 and are expressed in cell to either infection of immunization culture. with a foreign agent, the immune On recombinant human albumin system generates many different two companies has formed a joint antibodies that bind to the foreign venture to expand the commercial molecules. Individual antibodies development opportunities of a re- within this polyclonal antibody pool combinant human albumin program. bind to specific sites on a target The joint venture will manage devel- molecule known as epitopes. Isola- opment of the product for both the tion of an individual antibody within blood expander market and for the the polyclonal antibody pool would use in the excipient market. Recom- allow biochemical and biological binant human alpha-1 antitrypsin is characterization of a highly specific a recombinant form of the blood pro- molecular entity targeting only a sin-

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gle epitope. Realization of the thera- least 213 companies active in vari- peutic potential of such specificity ous aspects of protein-based thera- launched research into the develop- peutics development. Immunothera- ment of methods to isolate and con- peutic drugs divide broadly into two tinuously generate a supply of a sin- groups: vaccines that elicit an active gle lineage of antibody, a monoclonal immune response (generating anti- antibody (mAb). Disease areas that bodies that target the disease), and currently are especially amenable to therapeutic antibodies that bypass antibody-based treatments include the immune system and directly tar- cancer, immune dysregulation, and get the disease (so-called passive infection. Depending upon the dis- immune treatment). ease and the biology of the target, By nature, the polyvalent immuno- therapeutic monoclonal antibodies globulins (Immunoglobulin IM, SC can have different mechanisms of and IV) are considered as not being action. A therapeutic monoclonal replaced by monoclonal antibody antibody may bind and neutralize products. The polyvalent immuno- the normal function of a target. For globulins derived from plasma of example, a monoclonal antibody at least 1,000 donations consist of that blocks the activity of the protein a great number and variety of an- needed for the survival of a cancer tibodies representing the normal cell causes the cell’s death. Another physiological status of the human therapeutic monoclonal antibody body. Plasma derived specific or hy- may bind and activate the normal per immune immunoglobulins how- function of a target. For example, a ever, could theoretically be replaced

Downloaded from ijbc.ir at 12:07 +0330 on Sunday September 26th 2021 monoclonal antibody can bind to a by monoclonal antibody products protein on a cell and trigger an apop- because these products have a tosis signal. Finally, if a monoclonal titre of one specific antibody. Re- antibody binds to a target expressed search on monoclonal anti Rhesus only on diseased tissue, conjugation (D) immunoglobulin is an example of a toxic payload (effective agent), of these efforts. Monoclonal anti- such as a chemotherapeutic or ra- Rhesus (D) antibodies (MAbs) are dioactive agent, to the monoclonal all derived from human anti-D pro- antibody can create a guided mis- ducing B cells from immunised do- sile for specific delivery of the toxic nors, as mice do not recognise the payload to the diseased tissue, re- D antigen. The first mAbs were pro- ducing harm to healthy tissue. duced by immortalizing B cells with At the present time, there are at Epstein-Barr virus then cloning and

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selecting the anti-D secreting EBV-B more than the amounts needed for lymphoblastoid cell lines. Although patients’ use. Van Cott suggests in a huge amount of information has his article that this excess volume been collected on the biological ac- can be used for another administra- tivity of antibodies directed to blood tion route for factor VIII and factor IX, cells, no products have reached the for instance oral administration even market due to break troughs during though this is less efficient than the clinical trials (5). traditional intravenous route (7). For transgenic production, admission to Transgenic technology the market by European and Ameri- The current scarcity of factor prod- can registration authorities will be of ucts is caused by the low concen- utmost importance. The first medi- tration of factor VII and factor IX in cine produced using genetically en- human plasma. Also in a bioreactor gineered animals was Atryn, which cultures of genetically engineered is the brand name of the anticoag- animal cells (Chinese Hamster Ova- ulant antithrombin manufactured ry -CHO- cells) only limited concen- from the milk of goats that have trations can be produced. In con- been genetically modified to pro- trast, milk of transgenic livestock is duce human antithrombin, a plasma a prodigious and expedient source protein with proper- of complex therapeutic proteins ties. According to the manufacturing such as factor VIII or factor IX. In company one genetically modified 2004, Van Cott et al calculated that goat can produce the same amount 1.2 million litres of plasma dona- of antithrombin in a year as 90,000

Downloaded from ijbc.ir at 12:07 +0330 on Sunday September 26th 2021 tions per year are needed to meet blood donations. Goats were se- the needs of prophylactic therapy of lected for the process because they all haemophilia B patients n the US reproduce more rapidly than cattle (6). For recombinant factor IX pro- and produce more protein than rab- duction the equivalent is 600.000 bits or mice. In 2009, ATryn was ap- litres of CJO Cell Supernatant per proved in the U.S. for treatment of year. An equivalent amount of factor patients with hereditary antithrom- IX can be produced from no more bin deficiency who are undergoing that 12,000 litres milk of transgen- surgical or childbirth procedures. ic pigs, the total milk yield from an In 2006, the European Medicines average sow being 100-300 litres Agency (EMEA) initially rejected and, a year. This enables the production after an appeal from GTC, approved of large quantities of factor IX, even the drug for use in the European

134 IRANIAN JOURNAL OF BLOOD AND CANCER Plasma derived versus biotechnological...

Union countries. sealant this product is in phase II A second transgenic product is C1- research. Such developments may inhibitor. C1-inhibitor a protease lead to production of cheaper plas- inhibitor belonging to the ma products on a larger scale in the superfamily is also produced from future. plasma. Deficiency of this protein is associated with hereditary angioe- Pro and Cons dema (“hereditary angioneurotic The main advantage of biotechno- edema”), or swelling due to leak- logical products is viral safety. While age of fluid from blood vessels into the current plasma-derived plasma connective tissue. Its main function products are safe and transmission is the inhibition of the complement of blood borne infections have not system to prevent spontaneous ac- occurred since 1996, any discovery tivation.Transgenic C1-esterase in- of an emergent virus that potentially hibitor is produced in glycosylated can be transmitted by blood requires form using transgenic rabbits. The evidence by validation studies and product is licensed in Europe, but risk assessment analyses that the not in the US. plasma product is regarding the po- Transgenic haemophilia products tential transmission of transfusions are still in an early research phase transmittable infections. Maximum so the products cannot be expected safety is the objective. The transmis- within ten years. Nevertheless, this sion of viruses which are potentially timeframe implicates that trans- present in cell lines which produce genic haemophilia products could the recombinant proteins has never

Downloaded from ijbc.ir at 12:07 +0330 on Sunday September 26th 2021 mean serious competition for re- been reported. search on gene therapy. Factor VIII The disadvantages of these prod- is expressed in genetically-modified ucts are the development of anti- tilapia, a fresh fish farmed for food, bodies against the biotechnological and in transgenic sheep, factor IX produced proteins since these pro- in transgenic sheep and in preclini- teins are not identical to the self pro- cal phase, and with factor VIIa the tein. Currently a prospective study developments have been started. is ongoing where the incidence of On thrombin, factor XIII, factor VII, antibody formation against recombi- protein C, fibrinogen, the protein nant factor VIII is compared to the involved has been expressed in antibody formation against plasma transgenic animals, while with fibrin derived factor VIII. Another disadvan-

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tage of these products is the high a new generation of improved re- costs. Also with transgenic produced combinant products with extended proteins the occurrence of antibody half-life, less immunogenic, and formation is a matter of concern. In more resistant to inactivation. The particular because of the treatment question whether the new transgen- with these products of chronic dis- ic substitutes will be less expensive eases, a life long concern on anti- and enable larger supplies is not body formation and thus neutralisa- solved. At the same time efforts will tion of the administered protein ask be made to get new indications for for intensive post marketing follow existing products, and new thera- up. Although biotechnological man- pies (e.g. gene therapy). ufactured proteins inherently have For the hyper-immune immunoglob- the promise that abundant produc- ulins substitution developments tion implies low costs, this promise are ongoing to manufacture new had not become truth over the last vaccines, new anti-virals, and new years. monoclonals. Recombinant factor VIIa has been proposed as alternative to pro- References thrombin complex concentrates 1. Gotschier J, Wood WI, Goralka TM et al. for reversal the effects of vitamin Characterization of the human factor VIII gene. Nature 1984; 312:326-330. K antagonists such as warfarin or 2. Toole JT, Knopf JL, Wozney JM et al. cumadin. However, its short half life Molecular cloning of a cDNA encoding hu- may result in a need for repeated man aanti-hemphilic factor. Nature 1984; doses, increasing the risk of throm- 312:342-347.

Downloaded from ijbc.ir at 12:07 +0330 on Sunday September 26th 2021 bosis. Given recent reports of its 3. Vehar GA, Keyt B, Eaton D et al. Struc- usefulness in antagonist ture of human factor VIII. Nature 1984; 312:337-342. reversal, trials assessing safety are 4. Wood WI, Capon DJ, Simonsen CC et al. needed. Expression of active human factor VIII from recombinant DNA clones. Nature 1984; Future 312:342-347. The developments on new plasma 5. Kumpel BM. Efficacy of RhD monoclo- nal antbodies in clinicval trials as replace- products manufactured by biotech- ment therapy for prophylactic anti-D immu- nological production techniques will noglobulin: more questions than answers continue and will result in new re- Vox Sanquinis 2007;93:99-111. combinant substitutes. In particular 6. Van Cott KE, Monahan PE, Nichols TC, efforts will be made to manufacture Velander WH. Haemophilic factor pro- duced by transgenic livestock: abundance

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that can enable alternative therapies Zwaan RF. Oral treatment of haemophilia worldwide. Haemophilia 2004; 10 Suppl A by gastro-intestinal absorption of factor 4:70-76. VIII entrapped in liposomes. Lancet 1980; 7. Hemker HC, Hermens WT, Muller AD, 1(8159):70-71. Downloaded from ijbc.ir at 12:07 +0330 on Sunday September 26th 2021

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