Immunology & Vaccines

Chapter 13: Vaccines

 …Introduction
• Vaccines represent one of the most effective and cost-
effective medical and public health achievements of all time.

• Worldwide, vaccination programs are currently estimated to

save over 3 million lives each year also economic savings of
many billions of dollars each year.

• Vaccines are administered to healthy individuals (often to

entire age cohorts or populations), to prevent diseases caused
by infectious agents to which they might be exposed in the
future.
 Principles of Immunization…
• Vaccination specifically refers to efforts to induce protective
immune responses by administration of a vaccine.

• Immunization more generically refers to interventions either

active or passive that seek to confer immuneprotection.

• The actual mechanisms of immune protection induced by

either a natural infection or a vaccine are generally not
understood in detail for many infectious diseases.

• Vaccines depend on the induction of immunologic memory,

the magnitude, character, and duration of immune memory
differ between vaccines.
 …Principles of Immunization…
• Live attenuated vaccines:
• The fundamental concept of live attenuated vaccines is to
mimic the effective host immune responses that follow
natural infections.

• They are derived by propagation of initially pathogenic

organisms in culture on cells from different (nonhuman)
species, or at nonphysiologic temperatures, for prolonged
periods.

• Driving pathogen evolution in culture to select for variants

adapted to growth in heterologous cell types ex vivo often
leads to the derivation of pathogen variants that grow poorly
in vivo in humans and are unable to cause clinical symptoms.
 …Principles of Immunization…
• Vaccines developed via this approach include those used to
prevent a number of viral and bacterial infections, including
yellow fever, measles, mumps, rubella, polio (the “Sabin
vaccine”), varicella-zoster (used both for the prevention of
chickenpox and shingles) and rotavirus (one version of the
available vaccines), tuberculosis, and cholera.

• More recent technologies being applied to live attenuated

vaccine development include the application of reverse
genetic strategies and those involving genetic reassortment
with attenuated viral variants, as have been used to develop
polyvalent live attenuated vaccines against influenza and
rotavirus
 …Principles of Immunization…
• The live attenuated vaccines currently in use are highly
efficacious (> 90%) and protection is frequently durable.

• The efficacy of many live attenuated vaccines because they

replicate within vaccinated individuals, they can induce both
cellular (CD4 and CD8) and humoral (B cell) effector responses
and immunologic memory.

• They provide inherent adjuvant effects in augmenting

adaptive immune responses.
 …Principles of Immunization…
• A key consideration in the development of any live attenuated
vaccine relates to the relative balance between the ability to
induce sufficient immune responses in vivo to confer
protection (often associated with level of preserved
replicative ability in vivo), and the ability to cause symptoms
(which may also relate to the extent of in vivo replication).

• In addition, depending on the nature and number of genetic

mutations responsible for the attenuated phenotype, a
potential risk of reversion to a pathogenic form exists for
certain vaccines → The attenuating mutations are sufficiently
numerous or genetically stable.
 …Principles of Immunization…
• Example: live attenuated oral poliovirus vaccine. In this
instance, vaccine reversion to wild-type was shown to lead
rarely to cases of paralytic polio (approximately one case per
million doses administered).

• Based on these observations and the elimination of

endogenous polio transmission in many developed countries,
the inactivated polio vaccine was substituted for OPV
 …Principles of Immunization…
• Killed or inactivated organisms:
• Use of physical or chemical methods to kill or otherwise
inactivate a pathogenic organism.

• In most cases treatment with chemical agents such as β-

propiolactone or formaldehyde is used to eliminate pathogen
infectivity.

• Vaccines based on killed pathogens are believed to exert their

protective effects via elicitation of pathogen-neutralizing
antibodies and the induction of memory B-cell responses
(likely in concert with CD4 T-cell memory).
 …Principles of Immunization…
• Because inactivated pathogens cannot accomplish de novo
synthesis of pathogen derived gene products in antigen-
presenting cells (APCs), they do not typically induce CD8 T-cell
responses.

• killed vaccines are generally less immunogenic than live

attenuated vaccines. As a result, they are commonly
administered with an adjuvant (most often aluminium salts)
to augment their immunogenicity.

• Number of viral and bacterial vaccines currently in use are

killed/ inactivated vaccines, including whole-cell Bordetella
pertussis vaccine and the influenza virus, rabies virus, and
hepatitis A virus vaccines.
 …Principles of Immunization…
• Purified subunit vaccines:
• Detoxified versions of these toxins are referred to as
“toxoids,” and represent the purified components of vaccines
preventing diphtheria and tetanus, respectively.

• Toxoids have historically been produced by chemical

inactivation of toxins, but more recently, genetic inactivation
via targeted mutagenesis has been employed.

• The acellular pertussis vaccine is also a purified subunit

vaccine composed of a defined set of protein constituents
prepared from cultured Bordetella pertussis.
 …Principles of Immunization…
• The mechanism of immune protection conferred by purified
subunit vaccines is the antibody response elicited by
vaccination.

• Initial successful vaccine efforts against S. pneumoniae and N.

meningitidis utilized purified preparations of capsular
polysaccharides but they are poorly immunogenic in children
under 2 years of age.

• Purified polysaccharides are chemically conjugated to a

carrier protein (such as diphtheria toxoid or an outer-
membrane protein complex (OMPC) derived from N.
meningitidis).
 …Principles of Immunization…
• The carrier protein augments CD4 T-cell helper responses to
the polysaccharide antigens, and enables elicitation of
durable protective antibody responses even in young children.

• Polysaccharide-conjugate vaccines have been produced that

protect against Haemophilus influenzae b, Streptococcus
pneumoniae, and N. meningitidis infections.
 …Principles of Immunization…
• Recombinant protein subunit vaccines:
• They facilitating the identification and expression of
pathogen-derived protective antigens, techniques were
developed that enabled their large-scale manufacture as
vaccines.

• The first recombinant vaccine developed, the recombinant

hepatitis B surface antigen (HBsAg) prepared in yeast, was
developed in hopes of avoiding safety concerns related to the
plasma-derived HBsAg vaccine.

• The recombinant vaccine, when combined with adjuvant

(aluminium salts), elicits favorable immune responses, is
highly efficacious and is well tolerated.
 …Principles of Immunization…
• More recently, recombinant technology-derived purified
subunit vaccines have been developed that consist of virus-
like particles (VLPs) that self-assemble when the L1 protein of
HPV is produced in isolation of other viral proteins.

• The L1 protein is the target of virus-neutralizing antibodies

and vaccines consisting of a mixture of types 16 and 18 (the
cause of 70% of cases of cervical cancer) and 6 and 11 (the
cause of 90% of cases of genital warts) or of HPV types 16 and
18 alone are highly efficacious and well tolerated.
 Vaccine development & evaluation…
• First, animal studies must show that the vaccine candidate
raises the desired type and magnitude of immune response
against the infectious agent.

• Second, the vaccine needs to protect animals against death or

disease in an appropriate challenge model.

• Third, the vaccine should be relatively free of serious

discernible toxicities and side effects in animals when
administered by the route intended for humans.
…Vaccine development & evaluation…
• Fourth, it is necessary to demonstrate that the vaccine can be
produced in a consistent manner by a process that follows the
current good manufacturing practices (cGMP) process by
which the first clinical trial materials will be produced.

• Phase I studies primarily focus on detailed assessment of the

safety and tolerability of a vaccine, but evaluation of its
immunogenicity is also frequently conducted.

• A phase I study includes fewer than 100 healthy volunteers

divided unequally between those who receive vaccine or
placebo (2 or 3 vaccines per placebo recipient).
…Vaccine development & evaluation…
• Blood samples are taken at prescribed intervals and analyzed
for laboratory evidence of potential toxicity, as well as for
evidence of vaccine-elicited immune responses.

• A phase I study is considered successful if it demonstrates that

the candidate vaccine is well tolerated or identifies any
immediate safety concerns that will need to be closely
monitored in potential future clinical studies.

• Ideally, phase I studies also provide an initial indication of the

optimal dose level and number of doses required.
…Vaccine development & evaluation…
• A phase II study typically includes several hundred to a few
thousand volunteers (randomized between vaccine and
placebo) and can assume two general design types.

• Phase IIa studies provide additional safety data on a larger

number of individuals of the intended age who receive the
intended vaccine dose, as well as provide additional data on
vaccine immunogenicity.

• Larger phase IIb studies can provide additional data on

vaccine safety and immunogenicity in subjects generally
representative of those for whom the vaccine might be
recommended → Does this vaccine work in humans?
…Vaccine development & evaluation…
• Vaccines that have been shown to be immunogenic and well
tolerated in phase II studies can then advance to pivotal phase
III.

• Phase III studies are intended to expand further the safety

database in a larger number of individuals (who are
representative of the specific populations for which the
vaccine will ultimately be used).

• Phase III studies include 10 000 or more subjects in a blinded,

placebo-controlled design. This size trial allows the
identification of less frequent safety events. It also provides
an opportunity to capture data on health care utilization, cost,
and impact of the vaccine on these parameters.
…Licensure & recommendation of vaccines
• The US CDC has responsibility for making recommendations
about the use of licensed vaccines, and it relies on its Advisory
Committee on Immunization Practices (ACIP) for guidance.

• ACIP recommendation concerning vaccine storage, handling

and administration as well as Vaccine Information Statements
(VISs) that explain both the benefits and risks of each
recommended vaccine and that are to be provided to all
vaccine recipients.

• The approach to developing vaccine recommendations in the

European Union (EU) is somewhat more complicated, as
multiple countries are involved. At present, vaccines are
evaluated for registration through a centralized procedure by
the European Medicines Agency.
 New antigen discovery methods…
• With the advent of molecular biology in the late 1970s, a new
set of tools allowed a more directed approach for the
discovery of pathogen virulence factors, vaccine antigen
discovery, and vaccine development.

• The earliest rDNA technology-enabled methods of antigen

discovery involved the expression of individual pathogen
derived gene products (or fragments thereof) in bacterial
hosts (typically Escherichia coli) using rDNA expression
vectors.
 New antigen discovery methods…
• DNA sequencing became more efficient and scalable,
determining the entire sequence of the genomes of viruses,
bacteria, and parasites has become routine allowing
identification of previously unknown genes (and predicted
gene products) that can be evaluated as vaccine immunogens.

• Over 300 bacterial genomes have now been sequenced, and

hundreds more are currently in process. Genome- based
antigen discovery is being applied to a wide range of bacteria,
including streptococci, pneumococci, staphylococci and
Chlamydia, as well as nonbacterial pathogens such as
Plasmodium falciparum. This general technique is now called
“reverse vaccinology.
 Adjuvants…
• The term “adjuvant” (derived from the Latin adjuvare, to help)
refers to any substance added as a component of a vaccine
preparation—in addition to the vaccine antigens
themselves— that improves the immunological response to
the antigen.

• Most adjuvants were derived empirically and the mechanisms

by which they augmented immune responses were unknown.

• Particularly promising advance emerged from the discovery

that pathogen sensing by the innate immune system is
mediated by recognition PAMPs by PRRs such as TLRs that are
expressed on dendritic cells and other hemato-lymphoid and
some epithelial cells.
 …Adjuvants…

• Aluminium salts (Alum):

• Classical adjuvant most often used in vaccines in humans,
includes a range of salts of aluminum precipitated under basic
conditions, usually aluminum sulfate mixed with sodium or
potassium hydroxide plus a variable amount of phosphate.

• Alum is utilized as an adjuvant in many of the currently

available vaccines composed of inactivated toxins or
recombinant proteins (live attenuated vaccines do not include
alum or other adjuvants).
 …Adjuvants…
• Alum serves two main purposes as an adjuvant. First, it acts as
an antigen depot. Vaccine antigens adsorb to alum and elute
from it following injection into the host.

• Second, alum acts a mild irritant, causing the recruitment of

leukocytes necessary for generation of an immune response
to the site of injection.

• Alum does not typically enhance CD8 T-cell responses. Alum

has been a component of many vaccines for decades and has
an excellent safety record.
 …Adjuvants…

• Liposomes:
• Using lipids with polar head groups (e.g., triglycerides) and
differing types of hydrophobic tails, one can form either
micelles (spheres) or multilamellar sheets in aqueous
environments.

• Antigens can be incorporated into the spheres or between

layers of the sheets, providing a potential slow-release depot
system.

• Immunopotentiators such as QS21 or detoxified LPS

derivatives (such as monophosphoryl lipid A (MPL)) can be
added to the lipid mix.
 …Adjuvants…

• Immune-stimulating complexes (ISCOMs):

• Proprietary forms of liposomes made of cholesterol, saponins
from quillaia bark, and phospholipids that form cage-like
structures into which antigens can be entrapped or
intercalated.

• ISCOM complexes can provide a depot function, as well as

facilitating the delivery, uptake, and processing of vaccine
immunogens by APCs.
 …Adjuvants…

• Emulsions:
• Numerous oil-in-water and water-in-oil emulsions have been
tested as adjuvants. One such emulsion, MF59, is used in a
licensed influenza vaccine.

• MF59 consists of squalane, a metabolizable shark oil and two

surfactants, polyoxyethylene sorbitan monooleate and
sorbitan trioleate, in an oil-in-water emulsion.

• Cytokines:
• Several cytokines are being tested as potential vaccine
adjuvants, including granulocyte macrophage colony-
stimulating factor (GM-CSF), interleukin-2 (IL-2), and IL-12.
 …Adjuvants…
• Virosomes:
• Purified influenza virus hemagglutinin (HA) and
neuraminidase mixed with phosphatidyl choline and
phosphatidyl ethanolamine (polar lipids) will form empty
particles that have the surface properties of influenza virus.

• Adding an antigen in solution before mixing the lipids results

in the incorporation of the antigen inside the particle.

• This provides a vehicle for delivering antigens to the interior

of a cell, via the influenza HA membrane fusion process,
thereby enabling antigen processing and presentation via
both major histocompatibility complex (MHC) class 1 and 2
pathways.
 …Adjuvants

• Toll-receptor agonists:
• Of the defined TLR agonists being explored as vaccine
adjuvants, LPS and its partially detoxified form, MPL, which
activate TLR 4, have been most thoroughly explored in clinical
trials.
Questions?

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