Immuno 19

Question 1

What is variolation?

Answer 1

term for delivery of initial smallpox vaccine either intranasally or intradermally (scratch on the arm)

Question 2

What is vaccinia?

Answer 2

another name for the virus used to vaccinate against smallpox (vaccinia virus)

Question 3

What is vaccination?

Answer 3

the deliberate administration of either killed or living forms of a pathogen, or antigens derived from that pathogen in an effort to elicit an immune response that will prevent infection upon exposure

Question 4

What is immunization?

Answer 4

the deliberate provocation of an acquired immune response by introducing antigen into the body

Question 5

What are killed or inactivated vaccines?

Answer 5

any vaccine that employs killed pathogens as the antigen

the pathogens can be killed either by chemical treatment, heat, or irradiation

Question 6

What are attenuated vaccines?

Answer 6

any vaccine protocol that employs “weakened” pathogens as the immunogen;

pathogens can be weakened by several methods, and the result is that the attenuated pathogen’s ability to cause disease is either weakened or destroyed

Question 7

What is a subunit vaccine?

Answer 7

vaccines that employ only part of the pathogen as the vaccine immunogen toxoid: an inactivated toxin used as a vaccine immunogen whose toxic activity has been destroyed (usually by chemical treatment)

Question 8

Before the 1800s, smallpox epidemics were fairly routine and were devastating because the virus was easily transmitted from person-to- person, and because smallpox killed approximately 25% of those that became infected.

Answer 8

Before the 1800s, smallpox epidemics were fairly routine and were devastating because the virus was easily transmitted from person-to- person, and because smallpox killed approximately 25% of those that became infected.

Question 9

How was smallpox first addressed/vaccinated?

Answer 9

The first efforts to prevent deadly smallpox infections was to take dried pustules from people that suffered relatively mild symptoms from a smallpox infection. These pustules were then used to inoculate uninfected people by scratching their skin with these pustules; the process was known as variolation.

Variolation killed anywhere from 1-10 percent of people that took this “vaccine”, but people were still willing to play the odds.

Question 10

When was the first vaccine produced? By whom?

Answer 10

Edward Jenner in the 1780s

Question 11

How did Kylie Jenner (and Tyga) make the first vaccine?

Answer 11

Jenner noticed that milkmaids seldom became infected with smallpox, and those that did suffered very mild disease symptoms. He somehow made the connection that milkmaids were being infected with a virus known as cowpox, and he believed that the cowpox infection was immunizing them against smallpox. It turned out that he was right.

Cowpox is a virus that is very similar to smallpox, except that it is not very virulent in humans. The surface proteins of cowpox are very similar to those expressed on the surface of smallpox.

It turned out that immunization (or infection) with cowpox elicited neutralizing antibodies specific for the surface proteins of the
virus. These antibodies are the most important immune response for clearing/preventing infection with cowpox.

It turned out that the antibodies raised against cowpox surface determinants would also neutralize smallpox virions.

This vaccine was the first and only vaccine to ever be completely effective.

Question 12

By 1975, no cases of smallpox were being seen anywhere in the world. In 1979, smallpox was declared completely eradicated, and vaccine programs for smallpox were discontinued. Before 1979, everyone in this country was required to take the smallpox vaccine. I had mine when I was about 6 years old. I still have the characteristic scar at the vaccination site on my shoulder.

Answer 12

By 1975, no cases of smallpox were being seen anywhere in the world. In 1979, smallpox was declared completely eradicated, and vaccine programs for smallpox were discontinued. Before 1979, everyone in this country was required to take the smallpox vaccine. I had mine when I was about 6 years old. I still have the characteristic scar at the vaccination site on my shoulder.

Question 13

What is an example of a killed-subunits/toxoid vaccine?

Answer 13

1) Diptheria-tetanus-pertussis

Question 14

What is an example of a whole killed virion vaccine?

Answer 14

inactivated polio vaccine

Question 15

What are some examples of live attenuated virus vaccines?

Answer 15

1) measles, mumps, and rubella
2) varicella
3) Rotavirus
4) Influenza (can also be killed attenuated)
5) yellow fever

Question 16

What is an example of a heptavalent/diphtheria vaccine?

Answer 16

Pneumococcal conjugate

Question 17

What is an example of a subunit vaccine?

Answer 17

Hep B

Question 18

What is an example of a conjugated capsule subunit vaccine?

Answer 18

Meningococcus C

Question 19

What is an example of a Gardasil: virus-like particles vaccine?

Answer 19

HPV

Question 20

How do live attenuated viruses work?

Answer 20

These are all vaccines against viral pathogens, and because the vaccine formulation is an attenuated virus, the goal of the vaccine is to deliver the immunogens of the pathogen into the MHC class I pathway with the goal of activating a CD8+ T cell effector response.

Question 21

For the vaccines that are either killed, subunit, or conjugated capsular vaccines, what is the goal?

Answer 21

the goals are to deliver the immunogen(s) into the MHC class II pathway, and to elicit a potent B cell (antibody) response against the pathogen.

Question 22

What are some factors that make a ‘good’ vaccine?

Answer 22

1) safe
2) protective
3) sustained protection (An ideal vaccine is one that will elicit protective immune responses that will last a lifetime. This is a very high standard.)
4) Induced neutralizing antibodies (whether or not this is important clearly depends on the pathogen being vaccinated against. Neutralizing antibodies can be very important, but for some pathogens they have little if any role in protective immune responses)
5) Induces protective T cells (same as for neutralizing Abs. Can be critically important, sometimes T cells have no protective role.)

Question 23

T or F. The immune system is designed to respond to physiological amounts of antigen/mmunogen.

Answer 23

T.

(Left-Hand Panel): In the left hand panel, you see a graph that shows what happens when subjects are immunized with increasing concentrations of antigen (note that this is on a log10 scale). The immunogen dose is shown on the X-axis and the antibody response intensity is shown on the Y-axis. You can see that a threshold amount of antigen had to be administered before any real immune response was elicited. As the dose of immunogen increased, the immune response that resulted increased too, until really high doses were administered. Once the dose got high enough, the response to vaccination actually went down.

(Right-Hand Panel): Now, looking at the right-hand panel: in this panel, the vaccination that was done is the left-hand panel was repeated, and then each study participant was given a booster immunization with the 10e3 dose. The secondary immune response was then measured. What is very clear is that the participants that received very low primary doses responded very poorly to the booster immunization. This is known as low-zone tolerance, and it indicates that the immune system is designed to not respond to very low concentration of antigens, and in fact, when really low doses of antigens are administered, any responding lymphocytes are subject to being removed from the repertoire. Participants that received medium doses upon primary immunization responded very well to the booster immunization. Those that received very high primary doses responded very poorly to the booster immunization. This indicates that the immune system is designed to tolerize itself against any antigen that is supplied in amounts that are too high to be physiological for an infection. This is known as high zone tolerance.

Question 24

Effective vaccination is also difficult because the route of vaccination needs to match the normal route of infection for maximum benefit.

Answer 24

Effective vaccination is also difficult because the route of vaccination needs to match the normal route of infection for maximum benefit.

Question 25

The mucosal and systemic immune systems appear to be separated in some way. How is this known?

Answer 25

When a vaccine is administered via a parenteral route (IV, sub-Q, IM), a strong systemic immune response can be elicited, but there will be very little immune response that develops in the mucosal immune system.

In contrast, when an effective vaccine is administered via a mucosal route, the resulting immune response can be measured in both the mucosal compartment and the parenteral compartment.

Question 26

What is the relative mucosal response to different parenteral immunization routes?

Answer 26

1) subcutaneous
2) intraperitoneal
3) intramuscular
4) intravenous

Effective vaccination via the mucosal route has been difficult to date, but it is clearly the route that should be used for any pathogen that gains access via a mucosal route.

If you want a mucosal immune response, gotta give immunization through a mucosal route

Question 27

T or F. The immune system tailors immune responses depending on whether the pathogen likes to live extracellularly (TH2-type responses) or intracellularly (TH1-type responses).

Answer 27

T.

Question 28

What else must a vaccine accomplish?

Answer 28

The vaccine must provide danger signals that can be recognized by PRRs on phagocytes so that APCs will express the co- stimulator (B7) on their surface.

Question 29

What will happen if a peptide derived from a vaccine immunogen is recognized by a naïve T cell, but no co-stimulation occurs?

Answer 29

the T cell will become anergic and die.

Similarly, B7 binding to CD28 in the absence of peptide recognition has no effect on the naïve T cell.

Question 30

One other thing that makes vaccination difficult is that protective determinants must be supplied in the vaccine. Why?

Answer 30

It is difficult to know which determinants of a pathogen can actually serve as protective determinants. It is possible to make a very strong immune response against a particular determinant, but those responses may not be protective.

If protective determinants (i.e. bad to us) can be identified, those determinants should be supplied in the vaccine.

Question 31

There are two broad classification for vaccines:

Answer 31

Whole pathogen vaccines: these can be a whole wild-type bug (e.g. Cowpox), they can be live but attenuated, or they can be whole but killed organisms.

Subunit vaccines: these can be purified components or mixtures of pathogen-derived antigens, they can be recombinant forms of pathogen components, or they can be genetically engineered into delivery vector organisms.

The decision to use each of these different formats depends on what type of response is needed, and the route of administration that will be best for the particular pathogen.

Question 32

What are the subcategories of whole pathogen vaccines?

Answer 32

1) smallpox vaccine (vaccinia)
2) live-attenuated vaccines (an attenuated virus is a virus strain that has been mutated so that it no longer causes disease)
3) whole killed vaccines

Question 33

What are the subcategories of subunit vaccines?

Answer 33

1) individual or mixtures of pathogen-derived components
2) recombinant forms of pathogen-derived components
3) recombinant live vectors expressing component(s) of a pathogen

Question 34

What is an adjuvant?

Answer 34

An adjuvant is a material that is used to enhance the adaptive immune response that will be made to a vaccine immunogen. Usually, the adjuvant must be co-administered with the vaccine immunogen to be effective.

in many cases, adjuvants turn a soluble protein antigen into a particulate antigen (absorbed onto alum, emulsified in mineral oil) which is more readily ingested by antigen-presenting cells

Question 35

What are some common adjuvants?

Answer 35

1) Freund’s incomplete adjuvant
2) Freund’s complete adjuvant
3) Freund’s adjuvant with MDP
4) Alum (aluminum hydroxide)
5) Alum plus Bordetella pertussis
6) Immune stimulatory complexes (ISCOMs)
7) MF59

Question 36

What is the composition and mechanism of action of Freund’s incomplete adjuvant?

Answer 36

oil in water emulsion and works by delayed release of antigen causing enhanced uptake by macrophages

Question 37

What is the composition and mechanism of action of Freund’s complete adjuvant?

Answer 37

oil in water emulsion with dead mycobacterium

works by delayed release of antigen causing enhanced uptake by macrophages and via induction of co-stimulators in macrophages

Question 38

What is the composition and mechanism of action of Freund’s adjuvant with MDP?

Answer 38

oil in water emulsion with morally dipeptide (MDP), a constituent of mycobacteria (a source of PAMPs for induction of inflammation)

similar to Fruend’s complete adjuvant

Question 39

What is the composition and mechanism of action of Alum?

Answer 39

Aluminum hydroxide gel

an aluminum hydroxide gel that immunogens can be complexed with. It is also a depot former, resulting in delayed release of antigen. Alum is approved for use in humans, but it is not terribly effective. It works better when used in combination with Bordetella pertussis PAMPs which promotes co-stimulator expression.

Question 40

What is the composition and mechanism of action of Alum plus Bordetella pertussis?

Answer 40

Aluminum hydroxide gel with killed B. pertussis

works by delayed release of antigen causing enhanced uptake by macrophages and via induction of co-stimulators in macrophages

Question 41

What is the composition and mechanism of action of MF59?

Answer 41

another depot former that is a squalene, oil, and water emulsion.

***each of the adjuvants listed above will deliver antigen efficiently into the MHC class II
processing and presentation pathway.

Question 42

What is the composition and mechanism of action of ISCOMs?

Answer 42

a matrix of lipid micelles that can be loaded with the immunogen of choice. The micelles fuse with the cytoplasmic membrane of host cells, including APCs, dumping the immunogen into the cytoplasm of the cell. Therefore, ISCOMs is used for delivery of immungens into the MHC class I processing and presentation pathway (only one). There are many other lipid micelle like formulations (liposomes) that are being tested for vaccine efficacy.`

Question 43

Fruend’s adjuvant notes

Answer 43

all very effective, but they have too many side effects for use in humans. They are commonly used for research purposes, and they apply a couple of important vaccine strategies.. Each of them is an oil—in-water emulsion, and when antigen is mixed into the Freund’s, it becomes a sticky paste like substance. When it is injected into a host (typically sub-Q), it causes slow release of the antigen over several weeks (this is known as depot formation—similar to the “pegylated” drugs you have talked about in pharmacology). The “complete” and “with MDP” versions of Freund’s also include mycobacterial PAMPs that can be recognized by APCs, causing them to upregulate B7 expression.

Question 44

What are some bacterial toxins (that can be used as adjuvants) that have high potential for promoting immune responses to immunogens that are delivered via the mucosal route?

Answer 44

1) Cholera toxin- best studied
2) E. coli heat-labile lymphotoxin
3) Pertussis toxin

Cholera toxin B is the most well-studied of these agents, but E. coli heat-labile toxin and pertussis toxin also have high potential.

Question 45

What is the primary problem with bacterial toxin used as adjuvant candidates?

Answer 45

Their inherent toxicity. Numerous researchers have been trying to separate the adjuvanticity of these materials from their toxic properties, but it has been quite difficult. It may be that their adjuvanticity in some way depends on their toxicity.

Question 46

What is an attenuated pathogen?

Answer 46

a strain of a pathogen that will grow and replicate in the host and will elicit an immune response, but it will not cause disease.

Question 47

Why are Attenuated vaccines are typically more effective than killed or subunit vaccines?

Answer 47

because they:

1) replicate within the proper body compartment or infect the appropriate cells and replicate within them, generating physiologically appropriate amounts of antigen.
2) They also deliver their antigens into the appropriate processing and presentation pathway, and they elicit appropriate cytokine/chemokine signaling that ultimately promotes the right type of immune response to deal with that particular pathogen.

Unfortunately, live attenuated vaccines almost always have safety concerns.

Question 48

Attenuating a pathogen…in this case a virus.

Answer 48

(Left-Hand Panel): A viral pathogen that is isolated from a patient is initially cultured in human cells in vitro (in the lab in a dish).

(Left-Middle Panel): The cultured viruses are then collected and transferred onto monkey cells.

(Right-Middle Panel): The viruses that are generated during the replicative cycle are now adapted for growth in monkey cells, and will not be able to grow as well in human cells.

(Right-Hand Panel): Since they cannot grow as well in human cells, they do not cause as severe of a disease in humans.

(All Panels): Unfortunately, once a person has been “vaccinated” with this monkey adapted virus, some of the progeny virus in the vaccinee will be readapted for growth in human cells. If that virus is transmitted to a naïve patient, it could potentially have regained all of its virulence.

Question 49

What is a better way to produce safe attenuated vectors?

Answer 49

The use of recombinant DNA techniques to either mutate or excise one or more virulence genes from the genome of the pathogen. Once the gene(s) are either mutated or removed, it is very difficult for the virus to revert to its highly pathogenic wild-type form.

The problem with this method is that it is difficult to identify one or more virulence genes, that when mutated, cause attenuation of the pathogen without dramatically altering its infectivity and/or its replicative capacity.
If appropriate virulence genes can be deactivated, the attenuated pathogen can potentially be a very efficacious vaccine.

critical point: for this method to be effective, a virulence gene that does not play a role in the ability of the virus to infect cells or replicate within host cells must be identified

Question 50

What is Rotavirus?

Answer 50

a pathogen that was discovered in 1973 and found to be an important cause of childhood diarrhea. Worldwide, there are over 2 million hospital visits and over 600K deaths each year that are attributed to this nasty little virus.

Question 51

Rotavirus has some similarities to influenza virus. Name them.

Answer 51

Like Flu, antibody responses directed at the surface unit glycoproteins are the most important and protective immune responses to rotavirus.

The surface proteins of rotavirus (VP4 and VP7) are rapidly mutated during its replicative cycle (antigenic drift), resulting is distinct serotypes

It has a segmented genome (11 dsRNAs) that can be exchanged during co- infection (antigenic shift).

There are 42 known serotypes, but 5 of them are responsible for >90% of all morbidity/mortality.

Question 52

There are two attenuated vaccines for rotavirus:

Answer 52

(Left-Hand Panel): rotarix: this attenuated vaccine strain expresses the two most important variants of the two surface proteins (VP4 and VP7).

(Right-Hand Panel): RotaTeq: this is a mixture of 5 bovine rotavirus strains that are non-pathogenic in humans. They have been genetically engineered to express the most important human variants of VP4 and VP7.

(Both Panels): These attenuated viral vaccines are between 85 and 98% effective for preventing severe rotaviral diarrhea.

Question 53

Cytokines can be administered therapeutically to alter the outcome of infection. How?

Answer 53

Leishmania is an intracellular parasite that has differential outcomes in different strains of inbred mice.

Upon infection, one strain of mice (C57Bl/6) makes an anti-Leishmania immune response that can be characterized as a TH1 response; these animals survive the infection. A second strain of mice (Balb/c) routinely produces a TH2 response following infection with Leishmania, and these mice cannot clear the infection and they die.

o if however, the susceptible strain of mice receives therapeutic injection with the cytokine IL-12 (which promotes development of TH1 effector cells), the mice produce a TH1 response and survive the infection

Question 54

Removal or inhibition of cytokines can also alter outcome of infection. How?

Answer 54

using the same model described above: if the susceptible strain of mice (Balb/c) is infected with Leishmania, but is given
an injection of anti-IL-4 antibodies (remember that IL-4 promotes development of TH2 effector T cells), the mice will develop a TH1 type immune response and they will be able to clear the infection