BIOE 301

Lecture Nine

Engineering Design Method Designing a product to meet a practical

goal in the presence of constraints Six design steps:

1. Identify a need 2. Define the problem (goals, constraints) 3. Gather information 4. Develop solutions 5. Evaluate solutions 6. Communicate results

Papers, patents, marketing

Refine Design

SPECS

Three Case Studies Prevention of infectious disease

Early detection of cancer

Treatment of heart disease

Outline: Prevention of ID Science

Organisms that cause disease Immunity

Engineering How to make a vaccine

Vaccines: From idea to product Societal Impact

Health and economics Ethics of clinical trials Developed world/Developing world

Pathogens

How They Cause Disease

Types of Pathogens Bacteria

Cells with membrane and cell wall (usually) Can survive outside host Can reproduce without a host Can be killed or inhibited by antibiotics

Viruses Nucleic acid core with protein envelope Use host intracellular machinery to

reproduce Cannot be killed with antibiotics >50 different viruses that can infect humans

Bacteria

How do Bacteria Cause Disease?

Invade host Reproduce Produce toxins which disturb function

of normal cells

Virus

How do Viruses Cause Disease?

Virus invades host cell Binds to cell membrane receptors Endocytosis brings virus into cell

Virus takes over cell Use viral nucleic acid and host cell

resources to make new viral nucleic acid and proteins

More virus is released from host cell Virus causes host cell to lyse OR Viral particles bud from host cell surface

Pathophysiology of HIV/AIDS

Viruses

Three basic problems each must solve How to reproduce inside a human cell How to spread from one person to

another Inhale Eat During birth Intimate physical contact

How to evade the immune system

Influenza

Viral Reproduction - 1 Must get inside

human cell to use cell’s biosynthetic machinery

Influenza virus binds to cell receptor

Induces receptor mediated endocytosis

Influenza

Viral Reproduction - 2 pH slowly reduced in

endosome, virus releases its single stranded RNA and polymerase proteins

RNA segments and polymerase proteins enter nucleus of infected cell

Cell begins to make many copies of the viral RNA and viral coat proteins

Influenza

Viral Reproduction - 3 New viral particles exit

nucleus and bud from cell

Viral polymerase proteins don’t proofread reproduction

Nearly every virus produced in an influenza infected cell is a mutant

Influenza Viral Spread

Infected person sneezes or coughs Micro-droplets containing viral particles inhaled

by another person Penetrates epithelial cells lining respiratory

tract Influenza kills cells that it infects

Can only cause acute infections Cannot establish latent or chronic infections

How does it evade immune extinction? Antigenic drift Caused by point mutations

Yearly vaccination http://www.cdc.gov/flu/weekly/usmap.htm

http://students.washington.edu/grant/random/sneeze.jpg

Influenza How does the virus cause symptoms?

Cells of respiratory tract are killed by virus or immune system

Resulting inflammation triggers cough reflex to clear airways of foreign invaders

Influenza infection results in production of large quantities of interferon

Fever Muscle aches Headaches Fatigue

The Immune System

How Are We Protected Against

Pathogens?

Types of Immunity

Three layers of immunity: Physical Barriers Innate Immune System

All animals possess Adaptive Immune

System Vertebrates possess

Keep pathogens out Kill them if they get in

Types of Immunity Physical Barriers

Skin (2 square meters) Mucous Membranes (400 square meters)

Innate Immune System Produces general inflammatory response

when pathogens penetrate physical barriers

Adaptive Immune System Can adapt to defend against any invader Important when innate immune system

cannot defend against attack Provides immune system with “memory”

White Blood Cells

Neutrophil Lymphocyte Macrophage

What happens when you get a splinter?

Pathogen makes it past a physical barrier

Symptoms? Red, swollen, hot, pus

What causes these symptoms? Innate immune system is kicking into

gear Usually innate immune system can

take care of it

Innate Immune System

Macrophages eat bacteria on splinter Phagocytosis Produce chemicals which:

Increase local blood flow Redness Heat

Increase permeability of blood vessels Swelling

Recruit other phagocytes to site of infection Pus

Adaptive Immune System

Two main components Fight pathogens

outside of cells: 1) Antibodies

Fight pathogens inside of cells:

2) Killer T cells

What is an antibody? Bridge between:

Pathogen Tool to kill it

Antibodies have two important regions: Fab region:

Binds antigen Binds surface of virus

infected cell Fc region:

Binds macrophages and neutrophils, induces phagocytosis

Binds natural killer cell, induces killing

Antibodies How are antibodies made?

B cells Lymphocytes that make antibodies Have B cell receptors on surface 100 million different types of B cells, each

with different surface receptors B cell receptors are so diverse they can

recognize every organic molecule When a B cell binds antigen:

Proliferates - In one week, clone of 20,000 identical B cells

Secretes antibody

Adaptive Immune System How do we kill virus once inside the cell?

Antibodies cannot get to it Need T cells

T Cells Recognize protein antigens When bind antigen, undergo clonal selection Three types of T Cells:

Killer T Cells (Cytotoxic T Lymphocytes – CTLs) Helper T Cells Regulatory T Cells

How do T Cells ID Virus Infected Cells?

Antigen Presentation All cells have MHC molecules on

surface When virus invades cell, fragments of viral

protein are loaded onto MHC proteins T Cells inspect MHC proteins and use this

as a signal to identify infected cells

Immunologic Memory

First time adaptive immune system is activated by an antigen: Build up a clone of B cells and T cells Takes about a week After infection is over, most die off Some remain – memory cells

Second time adaptive immune system is activated by that antigen: Memory cells are easier to activate Response is much faster – no symptoms

Summary

Pathogens: Bacteria and Virus Levels of Immunity:

Barriers First line of defense Innate Inflammation Adaptive Immunologic memory

Antibody mediated immunity Cell mediated immunity Pathogens within

cells Diversity to recognize 100 million antigens

Vaccines

How Are They Made?

Vaccination

Vaccination: Practice of artificially inducing immunity

Goal of vaccination: Make memory T helper cells, memory

killer T cells, and memory B cells that will protect the vaccinated person against future exposure to pathogen

Want the vaccine to have: Maximum realism Minimum danger

What is needed to make memory cells?

Memory B Cells & Memory Helper T Cells: B and T cell receptors must see virus

or viral debris

Memory Killer T Cells: Antigen Presenting Cells must be

infected with virus

Types of Vaccines

Non-infectious vaccines DTaP Pneumococcus

Live, attenuated bacterial or viral vaccines Chicken Pox

Carrier Vaccines DNA Vaccines

Non-infectious vaccines Killed bacterial or inactivated viral vaccines

Treat pathogen with chemicals (like formaldehyde) Impossible to guarantee that you have killed all the

pathogen Salk (inactivated) Polio vaccine, rabies vaccine

Subunit vaccines Use part of pathogen OR Use genetic engineering to manufacture pathogen protein No danger of infection Hepatitis A & B, Haemophilus influenza type b,

pneumonoccocal conjugate vaccines Toxoid vaccines

Bacterial toxins that have been made harmless Diphtheria, tetanus and pertussis vaccines

This approach will make memory B cells and memory helper T cells, but NOT memory killer T cells

Booster vaccines usually required

Live, attenuated vaccines Grow pathogen in host cells Produces mutations which:

Weaken pathogen so it cannot produce disease in healthy people

Pathogen still produces strong immune response that protects against future infection

Sabin Polio vaccine (oral Polio) Measles, mumps, rubella, varicella vaccines This approach makes memory B cells, memory

helper T cells, AND memory killer T cells Usually provide life-long immunity Can produce disease in immuno-compromised

host

Carrier Vaccines

Use virus or bacterium that does not cause disease to carry viral genes to APCs e.g. vaccinia for Smallpox vaccine http://www.bt.cdc.gov/agent/smallpox/vaccination/

facts.asp This approach makes memory B cells, memory

helper T cells, AND memory killer T cells Does not pose danger of real infection Immuno-compromised individuals can get

infection from carrier Carrier must be one that individuals are not

already immune to Cannot make booster vaccines with carrier (must

use different carrier for booster)

DNA Vaccines

DNA injections can produce memory B cells and memory T killer cells

Reasons are not fully understood Make a DNA vaccine from a few viral

genes No danger that it would cause

infection

Types of Vaccines Non-infectious vaccines

No danger of infection Does not stimulate cell mediated immunity Usually need booster vaccines

Live, attenuated bacterial or viral vaccines Makes memory B cells, memory helper T cells, AND

memory killer T cells Usually provides life-long immunity Can produce disease in immuno-compromised host

Carrier Vaccines Makes memory B cells, memory helper T cells, AND

memory killer T cells Does not pose danger of real infection Immuno-compromised individuals can get infection from

carrier DNA Vaccines

Effectiveness of Vaccines Vaccination Effectiveness

About 1-2 of every 20 people immunized will not have an adequate immune response to a vaccine

Herd Immunity Vaccinated people have antibodies against a

pathogen They are much less likely to transmit that germ

to other people Even people that have not been vaccinated are

protected About 95% of community must be vaccinated to

achieve herd immunity Does not provide protection against non-

contagious diseases – eg tetanus

Vaccines

How Are They Tested?

Vaccine Testing

Laboratory testing Animal Model

Animal must be susceptible to infection by agent against which vaccine is directed

Animal should develop same symptoms as humans

Vaccine Testing Human Trials

Phase I Small number of volunteers (20-100) Usually healthy adults Last few months Determine vaccine dosages that produce

levels of memory B or T cells that are likely to be protective

Evaluate side effects at these dosages FDA must approve the vaccine as an

Investigational New Drug (IND) NPR Story – Ebola Vaccine Trials

http://www.npr.org/rundowns/segment.php?wfId=1513230

Vaccine Testing

Human Trials Phase II

Larger number of volunteers (several hundred)

Last few months to few years Controlled study, with some volunteers

receiving: Vaccine Placebo (or existing vaccine)

Endpoints: Effectiveness, safety

Vaccine Testing

Human Trials Phase III

Large number of volunteers (several hundred to several thousand)

Last years Controlled double blind study, with some

volunteers receiving: Vaccine Placebo (or existing vaccine)

Neither patients nor physicians know which was given

Vaccine Testing Role of the FDA:

Licensure by FDA required before a company can market the vaccine (about a decade)

Each batch of vaccine must be tested for safety, potency, purity and sample lot must be sent to FDA

Post-licensure surveillance Doctors must report adverse reactions after

vaccination to FDA and CDC Vaccine Adverse Events Reporting System

(VAERS) As many as 12,000 reports per year, 2,000 serious Most are unrelated to the vaccine

Vaccine Testing Recommendations by health departments

and expert physician groups When should vaccine be used Who should receive it Weigh: risks and benefits of the vaccine, costs

of vaccination Legislation:

States determine which vaccines are required by law

All 50 states have school immunization laws Can be exempted based on:

Medical reasons (50 states) Religious reasons (48 states) Philosophical reasons (15 states)

Recommended Vaccine Schedule

History of the Rotavirus Vaccine

Withdrawn from the market after post-licensure surveillance indicated small number of adverse effects

http://www.npr.org/templates/story/story.php?storyId=3262013

Vaccines

Are They Effective?

Effects of Vaccination in USDisease Max # of

Cases# Cases in

2000%

Diphtheria

206,929 (1921)

2 -99.99

Measles 894,134 (1941)

63 -99.99

Mumps 152,209 (1968)

315 -99.80

Pertussis 265,269 (1952)

6,755 -97.73

Polio 21,269 (1952) 0 -100

Rubella 57,686 (1969) 152 -99.84

Tetanus 1,560 (1923) 26 -98.44

HiB ~20,000 (1984)

1,212 - 93.14

Hep B 26,611 (1985) 6,646 -75.03

Effects of Vaccination Smallpox

First human disease eradicated from the face of the earth by a global immunization campaign

1974 Only 5% of the world’s children received

6 vaccines recommended by WHO 1994

>80% of the world’s children receive basic vaccines

Each year: 3 million lives saved

Childhood Immunization

1977: Goal to immunize at least 80% of

world’s children against six antigens by 1990

http://www.npr.org/templates/story/story.php?storyId=849775

http://www.npr.org/templates/story/story.php?storyId=3870193

Vaccines

What is Still Needed?

What Vaccines are needed?

The big three: HIV Malaria Tuberculosis

Summary How do vaccines work?

Stimulate immunity without causing disease How are vaccines made?

Non-infectious vaccines Live, attenuated bacterial or viral vaccines Carrier Vaccines DNA Vaccines

How are vaccines tested? Lab/Animal testing Phase I-III human testing Post-licensure surveillance

Impact of vaccines

Assignments Due Next Time

None