Immunotherapeutics Capstone Research Requirement

Submitted by: Louise T. Theunis, RPh

15th May 2019

APPLICATION OF IMMUNOTHERAPEUTICS IN VACCINE DEVELOPMENT

Infectious diseases have been a burden to our species since day one. Over the years, various
approaches to control, prevent, mitigate and cure infection have been developed - namely, public
health and personal hygiene, anti-infective drugs, dietary guidelines, herbal and food supplements, and
immune-directed therapies. The immune-directed approach is about lending the immune system a
hand in doing its job. This may be done by administering immunostimulatory agents (such as cytokines
and certain drugs like imiquimod) or other components of the immune response (such as antibodies
and antisera). Another method, vaccination, prepares the immune defenses via exposure to antigens.

The first attempts at vaccination were done in much ignorance of the causative pathogenic agents
as well as of the immune system. There was simply an observation, a notion, that those who had
recovered from an infectious disease could not (or seldom did) contract the disease again. Eventually,
through discovery, research and experimentation, we became more knowledgeable in this field and
applied our knowledge to further it.

The field of immunotherapy is concerned with using the immune system to combat disease. It is an
ingenius and undoubtedly successful application of the study of immunology to the field of medicine.
Immunotherapeutic agents work by activating or suppressing the immune system, and the use of such
agents depends on what is called for in the patient’s situation. Immunosuppressive therapy, which
reduces immune reactivity, is used in treating and/or preventing autoimmune disorders and reactions,
as well as graft or transplant rejection. Immunostimulation, on the other hand, is typically applied in
vaccination and cancer treatment.

Our knowledge of the immune system, I believe, has had its most successful application in
vaccination - thus far.

We know that there are two main branches of immune system. The first, the innate immune
system, launches rapid, broad-acting, nonspecific attacks (such as fever, inflammation and
phagocytosis) against foreign invaders that have managed to break through the body’s physical barriers,
both to eliminate them and simultaneously keep them at bay while alerting the other line of defense,
which is the adaptive or acquired immune system. The latter involves cells which recognize and adapt to
these foreign molecules (or rather, antigens), creating new and specified defenses (cell-mediated and
humoral) against them. On the first occasion an antigen is encountered by the body, the immune
response is rather slow; it usually takes a couple of days. Because the immune cells are naive and
unexperienced with this particular antigen, it takes a while for them to recognize and process the
antigen, and then expand up to sufficient numbers to eliminate it. This primary response is also quite
weak and short-lived. However, during this response, the immune cells adapt to that specific antigen to
customize specific antibodies and also differentiate into memory cells. Hence, during a subsequent
encounter with the same antigen, the immune cells can recognize the antigen immediately and thus
react faster and more effectively. The superior secondary response shows us two key features of the
adaptive immune system, namely specificity and memory. These are the underpinnings of vaccination.

The aim of vaccination is to prime the immune system to recognize and develop an attack against
an antigen (particularly the ones found on pathogens like bacteria and viruses), so that upon
encountering the actual antigen, the body can more easily eliminate it and thus survive the infection.
Now, with the continuous efforts in research and development, we have come a long way from simple
inoculation of pathogenic material from the source into the intended patient. Today’s vaccines may be
attenuated pathogens, toxoids, and other pathogen-derived antigens (peptides, polysaccharides,
conjugates), as well as live pathogens (where it has been deemed safe, effective and beneficial); and
there now exist guidelines for the appropriate dosing and use in different populations.

As knowledge of immunotherapy deepens, it has been found that there is more to the vaccination
than just giving the body antigens to which we expect it to respond. The timing, the accompanying
ingredients, the routes of administration and other details are also very important to inducing the
optimal immune response. There are appropriate dosing regimens for different vaccines, based on age,
sex, health status, level of exposure, and other characteristics or certain situations. For example, we
know that the immune systems of newborns and babies are immature, and we must also consider the
presence of maternal antibodies in addition to their own as their immune systems develop. For the
elderly we have to take into account the weakening of their immune systems. Live vaccines often
cannot be given to pregnant women due to the risk of harm to the fetus. Some vaccines, such as
tetanus, have booster shots since we know that the immune response for that antigen will slowly
decline over the years. The other ingredients in vaccine formulations are also of major importance.
Adjuvants are ingredients in vaccines that aid their action, usually by reducing antigen solubility and
slowly releasing small amounts of the antigens over an extended time period to allow the immune
system time to react. There are certain molecules common to many pathogens, called pathogen-
associated molecular patterns (PAMPs), which may be used as adjuvants because they can stimulate
both the innate and adaptive immune systems. One such example of PAMPs as a potential adjuvant is
unmethylated cytosine phosphate guanosine (CpG) dinucleotides which are found commonly in
prokaryotic deoxyribonucleic acid (DNA) (bacterial DNA) and rarely in eukaryotic DNA (e.g. our DNA).
These CpG dinucleotides bind with certain receptors on immune cells (toll-like receptors (TLR),
specifically TLR-9) and trigger additional immune reactivity for enhanced pathogen elimination. This
immunostimulatory activity has been recreated artificially with synthetic CpG oligodeoxynucleotides
(ODNs). Preclinical and clinical studies have demonstrated their safety and efficacy as vaccine adjuvants,
with an additional important benefit of being able to boost immune response even in populations with
reduced immune function (such as the immunosuppressed, elderly, and newborns or infants). Many
new adjuvants are still being developed and tested, as they offer potential benefits in the efficacy of
vaccines. And not all vaccines need to be injected. A welcome development, since the need to inject
vaccines (and the storage requirements) can be a challenge such as when big groups of people in
remote areas need to be reached. Some vaccines, such as the flu vaccine, may be administered
intranasally. Other vaccines, such as the rotavirus and oral polio virus, are administered orally because
of the benefit of superior mucosal immunity and herd immunity.

So, by using our knowledge of our immune system, not only have we managed to create vaccines
that induce the desired immune responses without causing major harm, but we have also gone into the
details of the frequency of dosing, the best routes of administration, the possible adverse effects, why
they happen and how to avoid or treat them, different effects for different populations (age, ethnicity,
genetics), other warnings and precautions, and more. That is, of course, not to say that we have
mastered it all. Vaccine development today faces a number of challenges: emerging and re-emerging
diseases (such as severe acute respiratory syndrome or SARS); constantly mutating pathogens (such as
the influenza virus); pathogens that we have not been able to develop vaccines for up to now (such as
HIV/AIDS and malaria); and, despite all progress, our still limited knowledge (of certain immune
mechanisms). Take for example the challenges posed by the variety of immunoregulatory factors at play
in our bodies, differing in various populations and stages in life. Newborns and infants in particular,
having exited the bubble of protection that is the womb and entered a world full of foreign matter and
pathogens, need such immunoregulation to prevent their bodies from having hypersensitive reactions
to almost everything. As they are still relying on their maternal antibodies and innate immunity, they
also need to develop adaptive immune responses to pathogens. To prevent potentially fatal encounters
with such pathogens, a number of vaccines are given at this age. However, immunoregulatory factors
may act in opposition to the desired immune response to the vaccines. One such factor is the regulatory
T cells. As their name suggests, they play a role in limiting the immune response and they may have an
effect on vaccine immunogenicity. There is hardly any literature on their effects on vaccines, but,
theoretically, transient inactivation of these cells may enhance vaccine efficacy. A better understanding
of such immunoregulatory mechanisms would most probably have an impact on vaccine development.

A very new and radical approach illustrates again the importance of immunotherapeutics in vaccine
development. This method involves directly injecting a plasmid containing the DNA sequence that
encodes the antigen of interest, which will then be produced by the body’s cells and subsequently
trigger an immune response. This approach provides a number of potential advantages over the
traditional approaches - stimulation of both humoral (B cell) and cellular (T cell) immune responses,
absence of infectious agent in the formulation, improved product stability, and relative ease of mass
manufacture. Although the field of DNA vaccination is developing rapidly, many aspects of the immune
response generated by this kind of vaccine are not yet understood. Researchers continue to study the
immune mechanisms and processes involved, as well as the range of immune activity and other effects
that may be elicited. The studies also cover the vaccine formulations, including possible adjunct
ingredients and adjuvants. All this in the hopes of gathering data that may be applied in the design of
better DNA vaccines and guidelines for their use.

Vaccines not only prevent bacterial and viral diseases but are also used to treat allergic conditions.
An allergy is an immune response to a foreign molecule that is typically harmless, like seafood, nuts,
pollen, and dust. Pathogenic organisms have antigens that are recognized by the immune system and
similarly, these foreign molecules or ‘allergens’ are also seen by the immune system as a threat. Based
on the current understanding of immunotherapeutics, ‘allergen immunotherapy’ has been developed.
Commonly known as allergy shots, this long-term treatment works like a vaccine. Formulations of a
particular allergen are injected subcutaneously a couple of times a week, starting with a very low dose
which is gradually increased. This results in the body becoming ‘hyposensitized,’ meaning it develops a
tolerance to the allergen. The immune response is no longer be triggered, and most allergic symptoms
are resolved.

Straying farther from the direct or indirect introduction of antigens to the body, one of the latest
applications of immunotherapeutics in vaccine development is in the making of ‘anticancer vaccines.’
Aptly called vaccines, they follow the same underlying principle of vaccination, basically priming the
immune system against disease. But there are major differences: the disease (i.e. cancer) is already
present in the body; and, there is no causative agent (such as a pathogen), rather it is the body’s very
own cells that have gone haywire. Years and years of research and experimentation have brought us
towards a still vastly incomplete but better understanding of cancer. We know that one of the reasons
why cancer cells thrive is because they are able to hide from the body’s defenses. They downregulate
their surface receptors and antigens, and send inappropriate signals to immune cells, all of which
prevent the immune system from recognizing them as a threat. Hence, the immunotherapeutic
approaches to this problem include marking the cancer cells (so the immune system can easily detect
and destroy them), monoclonal antibodies, adoptive cell transfer (ex vivo expansion of tumor-reactive T
cells and subsequent reinfusion), immunostimulatory cytokines (to modify immune cell activity), and
dendritic cell vaccines.

Antigen presentation is a crucial initial step in inducing an immune response. Dendritic cells are
cells that capture foreign molecules and present the processed antigen, in the form of peptide-major
histocompatibility complex molecule complexes, to T lymphocytes and also B lymphocytes. This allows
the effector lymphocytes to recognize the cancer cells and work to eliminate them. Vaccinologists have
figured out a way to harness the potent antigen-presenting capabilities of dendritic cells in the fight
against cancer. To prepare the dendritic cell vaccine, peripheral blood mononuclear cells and tumor
cells or tumor-specific antigens are harvested from the patient and purified. In the laboratory, the
mononuclear cells are enriched, matured and multiplied (with the use of cytokines), and then mixed
together with the tumor cells/antigens. This ‘cross-priming’ enables the matured dendritic cells to
process the tumor antigen/s and present them as surface molecules. After the incubation process, the
now antigen-pulsed dendritic cells are re-infused into the patient, and can then evoke a strong response
against cells expressing the antigen/s. Research is continuously being carried out on the dendritic cell
vaccination approach to cancer. Promising clinical outcomes have been seen in trials for the treatment
of various forms of cancer such as prostate, lymph, colorectal, brain and breast cancer. Treatment with
dendritic cell vaccines has shown low toxicity, with the common side effect being local injection site
reactions that are generally mild and self-limiting.

In conclusion, it is obvious, that I am fascinated by the existing knowledge of the immune system
and the gaps in our knowledge, the simplicity and complexity, the functionality and flaws, and the
unrealized potential. A detailed understanding of the many components of the immune system, their
types and subsets, their roles and functions, and the ways they work and interconnect, leads us to a
better understanding of vaccination. This information will provide us with better, more strategic
approaches to vaccines. The combination of knowledge and modern technology promises more exciting
developments in immunotherapeutics and vaccination in the (near) future.