2. EVALUATION SEMINAR ON
PROTEIN THERAPEUTICS
PRESENTED BY:
Ms. PRIYANKA SHRESTHA
1ST M. PHARM
DEPT. OF PHARMACOLOGY
FACILIATED TO:
DR. SHIVALINGE GOWDA K.P.
H.O.D, DEPARTMENT OF
PHARMACOLOGY

3. TABLE OF CONTENTS
 DEFINITION
 ATTRACTIVE FEATURES OF PROTEIN MOEITY
 PROTEIN DISCOVERY: HISTORY AND EVOLUTION
 SCOPE OF PROTEIN THERAPEUTICS
 CLASSIFICATION OF PROTEIN THERAPEUTICS
 PRODUCTION OF THERAPEUTIC PROTEINS
 OBSTACLES IN PRODUCTION
 SOURCES
 EXAMPLES OF PRODUCTION
 DELIVERY SYSTEM
 EXAMPLES OF SOME MARKETED PROTEIN THERAPEUTICS
 APPLICATION AND MARKET OF PROTEIN THERAPEUTICS
 REFERENCE

4. THERAPEUTIC PROTEIN
 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.

5. PROTEIN THERAPEUTICS
 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.

6. ATTRACTIVE FEATURES
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.

7. PROTEIN DISCOVERY: HISTORY
 Proteins were accepted as a distinct class of biological
molecules in the 18th century by Antoine Fourcroy and other.
 They are found to be able to coagulate in distinct conditions
e.g. albumen from egg whites, blood serum albumin, fibrin,
and wheat gluten.
 The elemental analysis of protein by Gerhardus Johannes
Mulder & use the name ‘Protein’ coined by JönsJakob
Berzelius in ~1839 in his papers.

8. The evolution of Protein therapeutics
 1953: First accurate model of DNA was suggested
 1982: Recombinant DNA technology was used to create human insulin
 1986: Approval of Interferon Alfa and muromonab-CD3
 1993: CBER's Office of Therapeutics Research and Review (OTRR) was
formed
 1997: First whole chimeric antibody, Rituximab, and first humanized
Antibody, Daclizumab, approved
 2002: Market for biotechnology products represented approximately
$30 Billions to $400 billion in yearly worldwide pharmaceutical sales
 2006: An inhaled form of insulin (Exubera) was approved, expanding
protein Products into a new dosage form

9. 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.

10. CLASSIFICATION
 Classification based on pharmacological action:
 Group I: protein therapeutics with enzymatic or regulatory
activity
I a: Replacement of a protein that is deficient or abnormal: e.g. -
Exubera, Increlex
I b: Augmentation of an existing pathway: e.g. - Ovidrel ,
Neupogen
I c: Provides a novel function or activity: e.g. - Myoblock

11.  Group II : protein therapeutics with special targeting activity
II a: Interferes with a molecule or organism: e.g. - Avastin
II b: Delivers other compounds or proteins (such as radionuclie,
cytotoxic drug or effector protein): e.g. – Ontak
 Group III : Protein vaccines
IIIa : Protecting against a deleterious foreign agent: e.g. -
Engerix
IIIb : Treating an autoimmune disease. : e.g. - Rophylac
 Group IV : Protein diagnostics: e.g. – Geref

12.  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

13. PRODUCTION OF THERAPEUTIC
PROTEINS
OBSTACLES IN PRODUCTION:
 Protein solubility, distribution, stability
 Delivery route (blood or digestive juice may degrade it)
 Physiologically active-post translational modifications
 Cost

14. SOURCES
Various types of cells like
 bacterial cells,
 yeast cells,
 insect cells,
 microbes (Bacillus sp., Actinomycetes sp., E coli),
 plants,
 animals,
 also mammalian cells
are used for producing therapeutic proteins under defined
conditions.

15. EXAMPLE OF PRODUCTION
MANUFACTURING SYNTHETIC HUMAN
INSULIN
51 amino acids = two chains
linked by disulfide bonds
21 in chain A
30 in chain B

16. DIAGRAMMATIC REPRESENTATION

17.  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

18.  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

19. Example of production explained
 Production of recombinant protein therapeutics in
cultivated mammalian cells
 Production of recombinant protein therapeutics in
milk of transgenic animals and schematic
representation of the process used to purify atryn
from the milk of transgenic goats

20. DELIVERY SYSTEM
 Proteins with a peptide sequence that shows the capability
to translocate membrane rapidly, termed as ‘‘cell
penetrating peptide (CPP) or protein transduction
domain(PTD) are covalently modified by methods
mentioned below:
1. Recombinant fusion protein from a vector containing DNA
sequence of the CPP sequence are directly expressed
2. Linker such as disulfide bond linkage that is cleavable
under reductive environment are used for protein or
chemical conjugation of CPP to the protein

21.  Peptide based biomaterial are used for delivery to protect
protein from protease degradation & Strategy to improve
delivery efficiency for the following reasons :
-Easy to synthesize -Easy characterization -Less toxic & has
higher immunogenicity than high molecular wt polymers.
 Due to its amphipathic character of peptides can associate
rapidly with protein cargos in solution in self-assembly manner,
possibly through noncovalent hydrophobic interaction.

22. Protein polymers conjugate for Targeted delivery
 Most commonly employed polymer : Polyethylene
glycol(PEG) & Poly(N-isopropyl acrylamide) (PNIPAM)
 These polymers that alter their solubility or propensity for
self-assembly when exposed to changes in pH or
temperature allow their responsive nature to be conferred
to the protein to which they are attached.
 Functionalizable with active esters & hence can be
conjugated with protein amine.

23. MARKETED PRODUCTS
rhDNase I
FOLLICLE STIMULATING
HORMONE (FSH)

24. APPLICATIONS

25. The therapeutic proteins in market and clinical trials today are
produced by recombinant method.
Higher efficiency and fewer side effects of protein therapeutics are
driving the protein therapeutics market comprising 16% of prescription
drug sales in 2011 of which antibody-based drug accounts to be the
fastest growing class of protein therapeutics.

26. REFERENCES: Protein therapeutics: a summary and pharmacological classification Benjamin Leader,
Quentin J. Baca and David E. Gola
 Production of recombinant protein therapeutics in cultivated mammalian cells Florian M
Wurm
 Recombinant Proteins Amith Reddy Eastern New Mexico University
 Recombinant therapeutic proteins Beenish Choudhary
 DESIGN AND PURIFICATION OF PROTEINS Biotechnology project, 18/05/09 Marielle
Brockhoff, Aurore Lacas , Raphael Lieberherr Sebastian Olényi, Morgane Perdomini,
Zrinka Raguz.
 production of recomb. Proteins BRUNO THADEUS
 Protein therapeutics Soma Mukherjee SMU, Chemistry 5th April’2011
 http://en.wikipedia.org/wiki/Protein_therapy
 Protein_Therapeutics--Xiao_Yu.ppt

27. THANK
YOU..!
