Therapeutic proteins are proteins engineered for pharmaceutical use. They are produced through recombinant DNA technology and delivered to replace proteins deficient in certain illnesses. Therapeutic proteins are classified based on their pharmacological action, molecular type, and mechanism. They can be produced through microbial bioreactors, mammalian cell culture bioreactors, or by expressing the protein in the milk of transgenic animals. Common therapeutic proteins include hormones, clotting factors, vaccines, and monoclonal antibodies used to treat diseases like cancer, infections, hemophilia, and hepatitis.

1. THERAPEUTIC PROTEINS
PRESENTED BY :RAFA ZUBAIR N.V
FIRST M.PHARM
DEPARTMENT OF PHARMACOLOGY
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2. INTRODUCTION
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“Proteins which are engineered in
the laboratory for pharmaceutical use are
referred to as therapeutic proteins”
 Proteins which are absent or low in individuals
with an illness such as Cancer, Infectious
diseases, Hemophilia, Anemia, Multiple
sclerosis, Hepatitis B/C, etc. are artificially
synthesized on large scale through genetically
modified host cells and delivered.

3.  This therapeutic approach in treating diseases using
proteins and peptides is termed protein therapeutics.
 Protein therapy is similar to gene therapy, but unlike
gene therapy, protein therapy delivers protein to the
body in specific amounts (as would be ordinarily
present), to help repair illness, treat pain or remake
structures.
 Introduced in 1920’s Human insulin is considered to
be the first therapeutic protein.
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4.  Proteins have been considered for the following facts:
 Diversity of functional groups: free thiols (on cysteine residue) &
amine (on the N-terminus or on lysine residue)
 Imitations by simple chemical compounds are less
 Lower side effects: due to high specificity there’s less potential
for protein to interrupt the normal biological processes
 Less likely for the body to evoke immune responses as the body
naturally produces many of the proteins
 Clinical development and FDA approval time are comparatively
faster than that for small molecule drugs.
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5. SCOPE OF PROTEIN THERAPEUTICS
 The hope is that the protein, which is not present in
adequate levels, will function as it is designed to do.
 For example, use of certain proteins in addressing
cardiovascular disease has been evaluated in some
studies. Especially when veins or arteries become
blocked, the right types of proteins might help here by
building new passages for blood flow.
 Some doctors suggest that protein therapy of this type
might eventually be so successful that it could eliminate
the need for complicated surgeries like bypass surgery. 5

6. CLASSIFICATION
 Classification based on pharmacological action:
 Group I: protein therapeutics with enzymatic or
regulatory activity
a: Replacement of a protein that is deficient or abnormal:
e.g. - Exubera, Increlex
b: Augmentation of an existing pathway: e.g. - Ovidrel ,
Neupogen
c: Provides a novel function or activity: e.g. - Myoblock
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7.  Group II : protein therapeutics with special targeting activity
a: Interferes with a molecule or organism: e.g. - Avastin
b: Delivers other compounds or proteins (such as radionuclie,
cytotoxic drug or effector protein): e.g. – Ontak
 Group III : Protein vaccines
a : Protecting against a deleterious foreign agent: e.g. - Engerix
b : Treating an autoimmune disease. : e.g. - Rophylac
 Group IV : Protein diagnostics: e.g. – Geref
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8.  Classification based on molecular types:
Antibody based drugs, Fc fusion proteins, anticoagulants,
blood factors, growth factors, hormones, interferon, bone
morphogenetic proteins, interleukins and thrombolytic.
 Classification based on molecular mechanism:
Binding non-covalently to target e.g. –mAbs
Affecting covalent bonds e.g. – enzymes
Exerting activity without specific interactions e.g. - serum
albumin
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9. METHODS USED TO PRODUCE
RECOMBINANT PROTEINS
(i) Production of recombinant proteins in microbial bioreactors
Examples
 E.coli expression system
 Saccharomyces cerevisiae
(ii) Mammalian cell derived bioreactors
 E.g. Chinese Hamster Ovary cell (CHO) bioreactors.
(iii) Animal Bioreactors “Pharming”
Production of Recombinant Therapeutic Proteins in the Milk
of Transgenic Animals Eg, Cows, sheep, pigs etc.
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rDNA
Protein

11. (i) Microbial bioreactors
 The first microbial bioreactors, in particular
Escherichia coli (bacterial) and Saccharomyces
cerevisiae & Pichia pastoris (yeasts) were found to
production of simple polypeptides such as insulin and
human growth hormone
 However, microbial bioreactors were found to be
unsuitable for proteins with complex post-
translational modifications or intricate folding
requirements, such as the coagulation factors, or
monoclonal antibodies.
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12.  This led to the development of large-scale
mammalian cell culture, for example, the use of
Chinese Hamster Ovary (CHO) cell cuture
bioreactors.
Limitations of microbial bioreactors
 Bacteria often improperly fold complex proteins,
leading to involved and expensive refolding
processes and ;
 Both bacteria and yeast lack adequate post-
translational modification machinery for mammalian-
specific N- and O-linked glycosylation, γ-
carboxylation, and proteolytic processing
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14.  Synthesis of the DNA containing the nucleotide sequences of
the A and B polypeptide chains of insulin.
 Plasmid + restriction enzyme Insertion of the insulin gene into
plasmid (circular DNA)
 Restriction enzymes cut plasmidic DNA
 DNA ligase agglutinates the insulin gene and the plasmidic DNA
Plasmid + insulin gene
 Introduction of recombinant plasmids into bacteria: E. coli
 E.coli = factory for insulin production
 Using E. coli mutants to avoid insulin degradation
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15.  Bacterium reproduces  the insulin gene replicates along with
plasmid E. Coli
 Formed protein partly of a byproduct the A or B chain of insulin
 Extraction and purification of A and B chain
 Connections of A- and B-chain by reaction forming disulfide
cross bridges
results in Pure synthetic human insulin
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16. (ii) Cell culture bioreactors
 These technologies permitted the development of
numerous monoclonal antibodies, cytokines, and
other complex bioactive biomolecules.
 However, there are proteins that, due to a
combination of complex structure and large
therapeutic dosing have until now eluded (fail to be
attained) recombinant production using traditional
bacterial and cell culture bioreactors
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17.  E.g Commercial recombinant production of complex
molecules, such as antithrombin and alpha1-
antitrypsin, has not yet been achieved in microbial or
mammalian cell derived bioreactors
 Cell culture systems require high initial capital
expenditures, lack scale-up (or down) flexibility and
use large volumes of culture media
 This led to development of transgenics technology i.e
Production of Recombinant Therapeutic Proteins in
the Milk of Transgenic Animals
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18. (iii) Production of Recombinant Therapeutic
Proteins in the Milk of Transgenic Animals
What is a transgenic animal?
 A transgenic animal is one which has been
genetically altered to have specific characteristics
(genes) it otherwise would not have.
 Different types of transgenic animals have been
invented to carter to specific societal needs.
 It includes; transgenic disease models, transgenic
pharmers, xenotransplanters and transgenic food
source.
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19. How are transgenic animals
produced?
The foreign DNA can be inserted in three ways:
(i) DNA microinjection
Fusing an expression vector, comprising a gene that is
encoded for the human or humanized target protein with
mammary gland-specific regulatory sequences, and then
inserting into the germline of the selected production
species.
 When integrated, the milk-specific expression construct
becomes a dominant genetic characteristic that is
inherited by the progeny of the founder animal.
 This general strategy makes it possible to harness the
ability of dairy animal mammary glands to produce large
quantities of complex proteins.
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21.  (ii) Retrovirus-Mediated Gene Transfer
 A retrovirus is a virus that carries its genetic material
in the form of RNA rather than DNA
 retroviruses used as vectors to transfer genetic
material into the host cell, resulting in a chimera
 chimeras are inbred for as many as 20 generations
until homozygous transgenic offspring are born
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22. (iii) Embryonic Stem Cell-Mediated Gene Transfer
 This method involves isolation of totipotent stem cells
from embryos
 The desired gene is inserted into these cells
 Cells containing the desired DNA are incorporated
into the host’s embryo, resulting in a chimeric animal
Advantage of transgenic systems
•Transgenic livestock can be maintained and
scaled-up in relatively inexpensive facilities
• Use animal feed as raw material
•Can achieve impressive yields of recombinant
proteins.
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23. APPLICATIONS
 Several proteins are created from recombinant
DNA (recombinant proteins) and are used in
medical applications.
 Hematopoietic growth factor.
 Interferon’s
 Hormones
 Recombinant protein vaccines
 Tissue/bone growth factors and clotting factors
 Biological response modifiers
 Monoclonal/Diagnostic/Therapeutic antibodies
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24.  Recombinant proteins is extensively used in
biotechnology, medicine and research.
 Hematopoietic growth factor
 Product of blood cells in bone marrow of central axial
skeleton is referred to as medullary hematopoiesis.
 While the mechanism of early stages of lineage
commitment by bone marrow to particular type of
blood cells remains elusive, the later stages of this
process is driven by hematopoietic growth factor.
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25. •List of factors of recombinant origin
 Recombinant technology is mostly used in production of insulin,
human growth hormone, vaccines ,Interferons etc.
 Recombinant proteins are used in medical applications, particularly
as medications and vaccines.
 Development of improved drug delivery system..
Product Company Indication
Thrombopoietin Phamacia Thrombocytopenia
Erythropoietin Amgen Anaemia
Ancestim Amgen Blood cell
transplantation
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26.  List of products of recombinant origin
Product Company Indication
Alpha-glucosidase Genzyme Pompe’s disease
Interleukin-4 receptor Immunex Asthma
Tumor necrosis factor
receptor
Immunex Rheumatoid arthritis
Vascular endothelial growth
factor
Genvec Cardiovascular
disorders
HIV vaccine Chiron AIDS
Prostvac Therion Prostate cancer
Neurex Xoma Cystic fibrosis
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27. List of hormones of recombinant origin
Hormones Company Indication
Human chronic
gonadotropin
Sereno Breast cancer
Leptin Amgen Diabetes mellitus
Thyroid stimulating
hormone
Genzyme Recurrent thyroid
cancer
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