USE OF BACTERIAL TOXINS AS

VACCINES

General introduction
Bacteria generate toxins which can be classified as either
exotoxinsorendotoxins.

Exotoxins are generated and actively secreted ,whose

toxicity has been inactivated or suppressed either by
chemical (formalin)or heat treatment.
endotoxins remain part of the bacteria. Usually, an
endotoxin is part of thebacterial outer membrane, and it is
not released until the bacterium is killed by the
immune system

 Genes for toxins are usually on plasmids.

Toxoids are used as vaccines because they induce
an immune response to the original toxin.
Example:

Tetnus toxoid,derived from the

tetanospasmin produced by clostridium tetani

causes tetanus and is vaccinated against by the
DTaP vaccine.

Bacterial protein toxins penetrate cells via a four-step mechanism

INTRODUCTION
In the case of bacterial toxins, this basic knowledge

has resulted in many applications Toxins are powerful tools to study cellular functions.

Several toxins are now used in the treatment of

various tumors

after coupling to suitable tumor-

specific vectors.
The botulinum neurotoxins are used in the therapy of a

series of dystonias.

(1) BINDING
The clinically active concentrations of toxins are very low.

Moreover, some of them are released in a rapidly cleared

environment such as the gastrointestinal tract.
Hence, they have to bind rapidly and firmly to the cell

surface.

They do so via two different modes of binding and this

corresponds

to

two

different

toxin

structural

organizations.

Oligomeric B toxins are composed of a pentameric discshaped binding protomer with a small central cavity.

Each B subunit of heat-labile Escherichia coli toxin (LT), cholera

toxin (CLT) and Shiga toxin (ST), contain a low affinity binding
site for the oligosaccharide of a glycolipid molecule .

These toxins are thought to bind to cells via a first interaction

with one glycolipid molecule, rapidly followed, in the twodimensional plane of the plasma membrane, by encounters with
other glyocolipid molecules.
The final result is a high-affinity cell association due to a
pentavalent binding. Pertussis toxin (PT) acts in the upper
respiratory

tract

and

its

protomer

has

the

same

oligosaccharide binding fold with the addition of two lateral

projections,

that

glycoproteins.

are

believed

to

be

involved

in

binding

 The catalytic domain A has little protein-protein

contacts with B to which is linked via a long alpha
helical segment and a segment which penetrates
in to the central hole of the B oligomers.

INTERNALIZATOIN
While binding does not depend on temperature,the toxin recaptor

complex is internalized inside membrane vesicles only at

permissive temperature.
This stage is very relevant in the immunotherapy of tetanus or

botulism, because, after internalization, the toxin is no longer

neutralized by anti-toxin antibodies.
Endocytosis may take place via coated vesicles, as it is the case

for DT and ETA, or via non-coated vesicles,as found for CLT and
TeNT.
Morphological studies indicate that ST and ETA undergo a

retrograde transport to the TGN, Golgi and ER.

MEMBRANE TRANSLOCATION

Since the targets of the toxins are located in the cytosol (or
are membrane-bound and face the cytosol),at least the
catalytic A subunit of the toxin has to cross the lipid bilayer.

Membrane

translocation

internalization because it is

is

clearly

distinct

from

the now known that only a

minor proportion of the internalized toxin molecules is

actually able to translocate the A moiety in the cytosol.

After internalization, toxins appear to be able to participate

in elaborated vesicular trafficking processes.

TARGET MODIFICATION

This fourth step is the final goal of the overall intoxication

process.

USING MODIFIED BACTERIAL TOXINS

TO DELIVER VACCINES ANTIGENS
Microbial proteins in the cytosol of host cells activate CD8
cytotoxic T lymphocytes (CTL).
Once activated, CTL lyse infected cells and secrete
cytokines that stimulate other immune cells at the site of
infection.
Because CTL are key components in protecting against
intracellular microbes, researchers are interested in
developing safe and effective vaccines that specifically
stimulate protective CTL.
The challenge in designing vaccines that stimulate CTL is to
deliver antigens not only into the bloodstream of the
recipient, but also into the cytosol of host cells.

ANTHRAX TOXIN CAN DELIVER CTL

ANTIGENS INTO HOST CELLS
AT(anthrax toxins) consists of three proteins: protective
antigen (PA), lethal factor (LF), and edema factor (EF).
Pair-wise combinations of PA and either LF or EF
generate functional toxin molecules.
As the first step in cellular entry, PA binds to surfaceexpressed anthrax toxin receptor (ATR) and is cleaved
by a furin-like protease to generate an activated form
of PA (PA63).

 PA63 oligomerizes on the cell surface and then can

bind to LF or EF.
The toxin complex then enters the cell via receptormediated endocytosis
When the endosome is acidified, a heptameric PA
pore facilitates translocation of the catalytic LF or EF
molecules into the cytosol.

As a strategy for vaccination, AT fusions have

been used to deliver a 287-amino-acid fragment
of LCMV NP, as well as HIV gp120 and p24.

SEVERAL TOXINS OF THE AB TYPE

CAN DELIVER ANTIGENS
A number of other bacterial toxins of the AB class,
including Bordetella pertussis adenylate cyclase toxin
(ACT), pertussis toxin (PT), Pseudomonas exotoxin A
(PE), and ST translocate their toxic catalytic domains
into host cytosol.
As we have done with AT, others have using the ability
of these toxins to translocate, using them to deliver
epitopes into host cells where they can be processed,
presented, and used to stimulate CTL.

 To ensure safety, a toxins enzymatic activity

must be inactivated before it is used to deliver
epitopes into host cells.
Methods of inactivating these molecules to
produce safe vaccines include deleting the
catalytic portion, removing the entire active
domain, and mutating key residues to block or
reduce enzymatic activity.

USING TOXIN FUSIONS TO

VACCINATE HUMANS
Although all these toxin systems sensitize target cells
for lysis by CTL in vitro, the ability to stimulate CTL in
animals so far is limited to AT, PT, ST, and ACT.
When toxins such as these are used in vivo as tools
for immunization, it is important to determine
whether the recipients have preexisting antibodies
that are specific to the toxin being evaluated.
For example ,toxins themselves are often used as
vaccines.

 The neutralizing antibodies resulting from such a

vaccination inhibit toxin binding and therefore
protect the immunized individual against the effects
of bacterial infections.
However, these antibodies might also render toxinfusion immunization ineffective. Even in the
absence of preexisting immunity, the toxin-fusion
dose required to stimulate CTL may also stimulate a
toxin-specific, neutralizing antibody response,
limiting the efficiency of repeated immunizations
with the same vector.

 Although any one toxin-fusion delivery strategy may face

these problems, alternatives will likely be found among
the other toxin-based systems under development.
For many of the pathogens for which there is no effective
vaccine, it will be necessary to use methods that
stimulate multiple arms of the adaptive immune
response including CTL, helper T cells,and antibodies.

Because of the technical challenges of stimulating

CTL, researchers are likely to continue developing
new methods to introduce antigens into the cytosol
of cells

TETANUS TOXOID VACCINES

INDICATIONS AND USAGE
Tetanus Toxoid is indicated for booster injection
only for persons 7 years of age or older against
tetanus. This vaccine is
NOT indicated for primary immunization.

PRECAUTIONS
Care is to be taken by the health-care provider for
the safe and effective use of Tetanus Toxoid.

 Since the discovery and productions of the tetanus

vaccines, the occurrence of tetanus, diphtheria, and

pertussis has decreased,100% people are prtected from
Tetanus.

ADVERSE REACTIONS
Adverse reactions may be local and include

redness, warmth, edema, induration with or without

tenderness as well as urticaria, and rash. Malaise,
transient fever, pain, hypotension, nausea and
arthralgia may develop in some patients after the
injection.

NEUROLOGICAL REACTIONS
The peripheral nervous system is affected more
often than the central nervous system.
The cranial nerves can be affected.

CARDIAC COMPLICATIONS
Myocardial infarction was noticed.

RHEUMATIC REACTIONS
Swelling of the joints, Persistent joint pains in 1
leg.

Model for anthrax toxin-mediated delivery of epitopes to stimulate cytotoxic T cells. (a) Toxin binding. Protective
antigen (PA) binds to its
cellular receptor, anthrax toxin receptor (ATR), expressed on host cells. Proteolytic cleavage of PA generates PA63.
PA63 then oligomerizes
and is able to bind a recombinant fusion protein containing the PA-binding domain of lethal factor (LFn) and a
cytotoxic T-cell (CTL) epitope.
(b) Cytoplasmic delivery. After LFn fusion protein binding, the entire complex is endocytosed via receptor-mediated
endocytosis. Following
endosome acidification, a heptameric PA pore mediates translocation of the LFn-epitope fusion protein into the host
cytoplasm. (c) Epitope
processing and presentation. Once in the cytosol, the fusion protein is processed by the proteasome into peptides.
The peptides are then
transported into the endoplasmic reticulum (ER) by the antigen-processing (TAP) complex, where they bind nascent