Importance of Protein Binding in Clinical Drug Development

This presentation discusses the importance of protein binding in clinical drug development, specifically in antimicrobial drug development. It covers the function of microbial proteins, potential drug targets for antimicrobial proteins, key points for drug targets in clinical development, and resistance mechanisms of bacteria. It also highlights the major drug binding proteins in plasma and their impact on drug development. The presentation provides insights into the importance of considering plasma protein binding during antimicrobial drug development.

Importance of
protein binding in
clinical drug
developmentNAME OF THE STUDENT
NAME OF THE UNIVERSITY
AUTHOR NOTE:

Protein in microbes
 Microbial proteins defines as the
protein requires for conducting
normal process of cell.
 The proteins also include the
normal cellular protein resembles
to the human to conduct normal
body cycle and proteins require
for the surviving in the host (4).
 examples , endotoxins,
exotoxins, leucocidine, plasma
protein, hymolysin
Figure: bacterial
protein
SOURCES: 6

FUNCTION OF MICROBIAL
PROTEIN
 The function of the
microbial protein is to
help in metabolism.
 To collect food from
other sources
 assist in cell division(3)
 to bind to the host
 survive inside the host
Figure: bacterial
protein
SOURCES

IMPORTANCE OF MICROBIAL PROTEIN
BINDING IN DRUG DEVELOPMENT
 A considerate number of antimicrobial
drug such as antibacterial drug target
extracellular or intracellular protein of the
bacteria to inhibit or static the growth of
bacteria
 The protein binding of bacteria also help
in reduction of deadly infections, assist in
evaluating the resistant mechanism of
bacteria which further showed the
direction of future research.
Figure: bacterial
protein
SOURCES: 7

antimicrobials
 Antimicrobials defined as a substance that either kills or
inhibits the growth of microorganism such as bacteria, fungi
and protozoa 8)
history:
in previous era various molds and plant extracts were used to
treat infections by some of the earliest civilisations such as
ancient Egyptians
After deaths of millions of individuals because of infections such
as such as pneumonia and diarrhea. Paul Ehrlich , the German
physicians suggested that certain chemical dyes colored have
ability to kill bacteria. Later, Alexander Fleming worked on
Penicillium notatum and accidentally discovered penicillin.
Figure:
antimicrobials
SOURCES: 7

History of antibiotics
Figure: History of antibiotics
SOURCES: 7

Protein target for antibacterial
drug development
 The potential drug target for antimicrobial
proteins
Bacterial
surface
protein
Bacterial
intracellular
proteins
lipoproteins
Trans
membrane
protein
Porin
proteins
DNA
polymerase
DNA
Gyrase

Key point of drug targets in
clinical development
 It should give have maximum drug effect
 It should not cause side effects in body
 it should be readily soluble in the body
 it should not target human proteins (9)
 it should not destroy by metabolic fluids of
human body(8)
 it should readily thatched to the bacterial
protein4
 it should have bacteriostatic or bactericidal
property
Figure: bacterial
protein
SOURCES: 7

Gram positive drug targets
(surface protein)
 Transmembrane protein of gram positive bacteria act as
potential drug target. Membrane proteins are used by cells to
transduce signals into cells, transport ions and molecules, bind
the cell to a surface or substrate, and catalyze reactions.
 Recent technologies allows drug conventionally to target
“undruggable” regions of membrane proteins, enabling
modulation of protein–protein, protein–lipid, and protein–nucleic
acid interactions
 () highlighted that Enzymes, transporters, ion channels, and
receptors are all common membrane protein (MP) used for drug
targets
 although no drug developed yet but researchers has been
conducted in the laboratories to develop drugs.

Possible mode of action
bind proteins within
solvated regions outside
the membrane.
A hallmark of drug
ability is a solvent-
accessible hydrophobic
pockets often acts as a
active site of enzyme
( in case of orthosteric
drugs)

Gram positive drug targets
( bacterial DNA replication
protein) The two bacterial topoisomerase II enzymes – DNA gyrase and
TopoIV – modify the topology of DNA during replication acts as
potential protein binding site of antimicrobials.
 Despite structural similarities two enzymes perform distinct
functions in prokaryotes.
 () highlighted that DnaA and DnaC, Primase is targeted in the
bacteria such as Bacillus and Clostridium
 despite this acts as potential targets , only topoisomerase II acts
as drug target (5).
 the clinically available drug target aminocoumarin class and
fluoroquinolones and novobiocin (staphylococci and other
susceptible organisms)

Mode of action :
stabilize cleaved DNA–
topoisomerase II complexes
increasing the number of
double-stranded DNA breaks
in the bacterial cell(6)
Most FQs are able to inhibit
both gyrase and TopoIV
depending on efficiencies
and bacterial species

Other replicated protein
targets in gram positive
 adenosine analogues and 2-amino-[1,8]-naphthyridine-3-
carboxamides (ANCs) acts as drug which target the DNA ligase
as protein binding site for bacteria such as Staphylococcus
aureus.
Mode of action: All ligase inhibitors are competitive inhibitors that
bind a hydrophobic pocket near the binding site of the NAD+
substrate but shows potential toxicity because of resemblance (6).
 Pol iii inhibitor such as guanine inhibitors , 6-anilinouracils and
non-nucleobase inhibitors in bacteria such as B. subtilis.
Mode of action: These inhibitors interfere with the DNA base pairs
to disrupt the Bacterial DNA synthesis and consequently inhibit
growths (9).

other replicated protein
targets in gram positive
 sliding clamp, or β subunit f the bacterial
replications is targeted by Griselimycin in
Corynebacterineae suborder such as mycoplasma.
 () recent research suggested that small
antimicrobial molecules are molecules are proved
to either mimicking the structure of single
stranded binding protein or disrupting the
function of function. It specifically mimic C
terminal which is a platform for interactions of
other proteins(7).
 The importance of these bindings is that it can
disrupt bacterial replication which induce
inhibition of cells , acts a clinical development of
drug Figure: bacterial protein
Source: (10)

METABOLIC PROTEIN BINDING AS A
TARGET SITE OF ANTIMICROBIAL DRUG
 One of the famous metabolic protein that acts
as drug target from decade is transpeptidase, a
penicillin binding protein which involve the final
step of Peptidoglycan Biosynthesis pathway. It
was the one of the first conventional targets of
Beta-lactam antibiotics- Penicillin &
Cephalosporins in gram positive bacteria such
as staphylococcus, streptococcus (8).
 it also works against few gram negative
bacteria such as gonorrhea (4).
 The importance of these bindings is that it can
disrupt bacterial metabolism which induce
inhibition of cells.
Figure: Metabolic protein
source: (7)

Mode of action :
Penicillin shows
bactericidal effects by
binding with penicillin
binding protein
Disruption of function of
trans peptidase
Disruption of gram
positive cell wall
synthesis

Unique Gram negative drug
targets
 porin protein is unique to the gram negative bacteria which is a
beta barrel proteins which crosses the cellular membrane and
acts as a pore through which diffuse.
 although it is difficult to target porin proteins, few clinical drug
development laboratories are working .
 Porin loops are potential targets for adhesion to other cells and
binding of bactericidal compounds to the surface of Gram-
negative bacteria (6).
 Advance research proposed antimicrobial targets is histidine
kinases which is a common component of bacteria essential for
adaptation in the stress situation. Since in human the mechanism
is different , it can be used a potential drug (5).

antifungal drugs targeted the
fungal proteins
 Antifungal drugs are used for inhibition of fungal growths.
 Brown (2015), highlighted that lipid transfer protein Sec14p
as target for antifungal ergo lines.
 Tommasi (2015), highlighted that DNA topoisomerase is also a
potential target for the fungus such as candida sp. cytotoxic
tricyclic quinolone shows a slightly stronger DNA cleavage
enhancement effect with the Candida enzyme than with the
mammalian counterpart. Therefore, it is acts as potential drug
target for

Antiprozoan drugs targeted the
fungal proteins
 anti-protozoan drug
such as metronidazole
act on the DNA gyrase
function which in turn
disrupt the function of
the protozoan DNA
synthesis (6). Figure: protozoan
protein
Source: (5)

Anti-viral drugs targeted the viral
proteins
 the most common approach of the antiviral protein is inhibition
of reverse transcriptase used for generating complementary DNA
from the RNA strands (6).
 the most common drug that target the viral protein is acyclovir
effective against herpesvirus infections and HIV infection, chicken
pox.
 Another common target for the viral target is viral protease such
as amprenavir, darunavir used against the HIV and herpes.

Mode of action of acyclovir
Nucleoside analogues
acts on the reverse
transcriptase(7)
Disrupts the
complimentary DNA
synthesis from RNA
Inhibit the DNA
replication of
herpes and HIV

Mode of action of amprenavir
Disrupt the function of
protease by binding to it
Disrupt the
multiplication of HIV
and Herpes
Figure: amprenavir
Source: (5)

Resistance mechanism of
bacteria
 to stop the antibiotic from reaching its
target bacteria:
Bacterial cells produce efflux pump in the
cell membrane to eliminate the internalize
drug. Bacteria decrease the permeability of
membrane (7).
Bacteria destroy the antibiotic through
bacterial enzymes. Bacteria produce
enzymes that are capable of adding
different chemical groups to antibiotics and
therefore modify the bacteria. Figure: Resistance
mechanism of
protein
Source: (5)

Resistance mechanism of
bacteria
 To modify or bypass the target that the
antibiotic cannot identify target:
Some bacteria can hide the target by modifying
the drug binding site of protein and therefore
antimicrobials failed to identify the drug target.
Some bacteria able to express alternative
proteins so that drug failed to identify
proteins. for example, the
bacterium Staphylococcus aureus can acquire
the resistance gene mecA which produce
alternative PPB (8).
Figure: Resistance
mechanism of protein
Source: (9)

Resistance mechanism of
bacteria
 Some bacteria can produce
different variant if structure of
protein so that anti microbial drug
cannot identify the protein (7).
 Some bacteria carry resistant
genes in plasmid through
conjugation that antimicrobials
failed to understand the target
site.
Figure: Resistance
mechanism of
protein
Source: (7)

Importance of protein binding in clinical drug
development ( alternative perspective)
 Majority of the drug is bound to plasma protein then only the
unbound fraction can have a biological effect or be
metabolized/excreted. Therefore, the degree of binding to the
plasma membrane influences the pharmacokinetic and
pharmacodynamic properties of a drug.
 The efficacy of the anti-microbial drug will depend on the
exposure to the amount of unbound drug in plasma.
 The binding portion can be also affected if two drug are
consumed at the same time because each drug may dislocate a
proportion of the other from being bound to the plasma protein.

MAJOR DRUG BINDING
PROTEINS IN PLASMA
 Major drug binding protein in plasma is
albumin which inhibit the function of the
fluroqunolones and therefore it is crucial for
the bacterial drug development.
 the unique feature of this major drug binding
protein is it interferes with the protein bind to
the free drug and therefore, it is crucial for
researchers to consider the plasma protein
binding during antimicrobial drug
development.
 Each anti-microbial drug development
follows ADME principle.

The importance of protein
binding in clinical trials
 During the development of
antimicrobial drugs it is
crucial for researchers to
look after each plasma
protein interactions with
antimicrobial protein.
 The prime reason behind it
is that Drug transport and
disposition is influenced by
the non specific reversible
drug binding to plasma and
tissue.
Figure: protein binding
mechanism
Source: (9)

Significance of protein binding of
drug
 Absorption :
conventional dose form follows first order kinetics. The more proteins are
present in the blood , it distributed to absorption equilibrium.
 Distribution:
protein bound drug hinders the drug from penetrating blood brain barriers,
placental barriers and therefore, it decrease the distribution of drug.
 Metabolism:
Protein binding decreases the blood metabolism and reduces the half life.
 Elimination:
the protein drug complex sometimes remain in the blood and provide free
supply of blood.

Common plasma protein influence
the efficacy of drug
human
serum lipoprotein,
glycoprotei
n
α, β‚ and γ
globulins

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