© life_edu lecture 21 part ib. emergent technologies: dna-based biotechnology and pharmaceutical...

© life_edu

Lecture 21

Part Ib. Emergent Technologies: DNA-Based Biotechnology and

Pharmaceutical Drug Development

Issues in Biotechnology:The Way We Work With Life

Dr. Albert P. Kausch

life edu.us

Pharmaceutical Biotechnology

Issues in Biotechnology:Biotechnology, Our Society and Our Future

OnCampus LiveOnCampus LiveBCH 190, MIC 190, AFS 190, NRS 190, PLS 190BCH 190, MIC 190, AFS 190, NRS 190, PLS 190

OnLine BCH 190OnLine BCH 190

A Sweeping General Survey on Life and BiotechnologyA Public Access College Course

The University of Rhode Island

Kimberly Nelson



life edu.us

© life_edu

A Sweeping General Survey on Life and Biotechnology

The University of Rhode Island



life edu.us

BCH 190BCH 190 Section II.

The Applications of Biotechnology

• Where do our Medicines come from?• History• Alternative Therapies and Science

• How is DNA-based biotechnology used in current pharmaceutical drug development improvement?

• Small Molecule Drug Design• Recombinant DNA Drugs

• How is it done? What are the goals?• What as been done so far?• Antibody based drugs• Vaccine Development and Production• What is in the future?

• What are the controversies and concerns?

Pharmaceutical Biotechnology What is it?

Recombinant DNA Recombinant DNA And New Drugs And New Drugs

What Are the Implications

Of Gene CloningFor Pharma Now?

Genetic Constructs Now Make Proteins That Are Pharmaceuticals

Controlled expression“making protein”

Promoter

Your favorite gene

Insulin Enbrel Herceptin

Coding Sequence Terminator

Stop transcriptionMessage stability

DNA TechnologyDNA Technologyand Pharmaceuticalsand Pharmaceuticals

Allows precise treatments for:

Cancers Cardiovascular diseases Inflammation

Behavior Obesity Depression Schizophrenia

Cracking the Genetic Code (Nirumberg and Mathei 1962)

The information is the same in all living organisms

This fact is exploited for DNA based drugs

Biomanufacturing Process for Recombinant Proteins

Biomanufacturing Process for Recombinant Proteins

BiomanufacturingBiomanufacturing

Vaccines and AntibodiesHow Are They Made?Controversies: Vaccines and Human Health: Myth Understood

Vaccines DO NOT Cause AutismWhy ALL Children Should be Vaccinated?Can a Vaccine Compromise Your Immune System?

How Does Genomics Influence Vaccine Development?Are we Ready for Contagion?

Why Are There Broad Public Misgivings About Vaccination?

What happens when humans are invaded

The Immune System:

Antibodies are proteins encoded by genesImmunotherapy uses antibodies as drugs

The Immune System:

Antibody Based Drugs

Coding sequence for an antibody fragment to a specific antigen

Promoter Coding Sequence Terminator



Monoclonal antibody therapy is the use of monoclonal antibodies (or mAb) to specifically bind to target cells or proteins

Fv

Ancillaryfunctions

Fc

CH1

CH2

CH3

VH

VLAntigenBindingSite

Antibodiesversatile protein molecules capable of recognizing

foreign proteins

Antigen/pathogen specificity

Effector functions: • Neutralization/blocking effects• Complement fixation• Agglutination

ab ba

Ag Ag

The B cell receptor

Production of Hybridomas and Monoclonal Antibodies

Monoclonal antibodies are a homogeneouspopulation of antibodies that are specific to their antigen

Western Blot AnalysisWestern Blot AnalysisHome Pregnancy TestHome Pregnancy Test

Detection of specific proteins

proteins are separatedby gel electrophoresis

separated proteins are transferred to a filter

antibodies recognize proteins and are detected with specific dyes

Applications of monoclonal antibodies

ELISAEnzyme Linked

Immunosorbent Assay

Home Pregnancy Test -

an antibody test specific to human chorionic gonadotropin HCG is captured and detected with a color enzyme developed product

Vaccines and Vaccination

Facts and Fictions

Vaccines and AntibodiesHow Are They Made?Controversies: Vaccines and Human Health: Myth Understood

Vaccines DO NOT Cause AutismWhy ALL Children Should be Vaccinated?Can a Vaccine Compromise Your Immune System?

How Does Genomics Influence Vaccine Development?Are we Ready for Contagion?

Why Are There Broad Public Misgivings About Vaccination?

Vaccines and Vaccination

A foreign protein (antigen), an attenuated, or killed virus will stimulate the immune system to cells with a memory for the antigen. The immune system is activated when the antigen is again encountered.

How Are Vaccines Made?Vaccine Production

MethodsExample: Attenuated Vaccineother Common methods

Flu vaccine timeline for productionPandemic fluContagion

Other VaccinesHIV VaccineHPV VaccineBacterial VaccinesMalaria Vaccine

How to Make a Vaccine - Six different ways

Similar-pathogen vaccine: smallpox virus

Attenuated vaccine: measles virus

Killed vaccine: polio virus

Toxoid vaccine: tetnus

Subunit vaccine: hepatitis B

Naked DNA vaccine: HIV virus

You are about to create a live-attenuated vaccine, which means that you need to alter a pathogen—in this case a measles virus—so that it will still invade cells in the body and use those cells to make many copies of itself, just as would any other live virus. The altered virus must be similar enough to the original measles virus to stimulate an immune response, but not so similar that it brings on the disease itself.

To create a new strain of the virus, you’ll need to let it grow in a tissue culture.

Attenuated vaccine: measles virus Step 1 Use the tissue culture to grow new viruses

The tissue culture is an artificial growth medium for the virus. You will intentionally make the environment of the culture different than that of the natural human environment. For this vaccine, you'll keep the culture at a lower temperature.

Attenuated vaccine: measles virus Step 2 Fill the syringe with a strain of the virus that has desirable characteristics

Over time, the virus will evolve into strains that grow better in the lower temperature. Strains that grow especially well in this cooler environment are selected and allowed to evolve into new strains. These strains are more likely to have a difficult time growing in the warmer environment of the human body. After many generations, a strain is selected that grows slow enough in humans to allow the immune system to eliminate it before it spreads.

Attenuated vaccine: measles virus Step 2 Fill the syringe with a strain of the virus that has desirable characteristics - continued

You have just produced a live-attenuated measles vaccine.

Like the smallpox vaccine, the virus within the vaccine will invade body cells, multiply within the cells, then spread to other body cells. The virus used in the measles vaccine today took almost ten years to create. The starting stock for the virus originated from a virus living in a child in 1954. Live-attenuated vaccines are also used to protect the body against mumps, rubella, polio, and yellow fever.

Attenuated vaccine: measles virus Step 3 Completed vaccine ready for use

How Are Vaccines Made?

‘You cannot develop vaccines against a bacterial pathogen without the genome.’

To Make a Bacterial Vaccine, First Sequence a Genome

Is this Science Fiction?

This couldn’t really happen-Right???

How accurate is this?

How worriedshould I be?

Pandemic: A Worldwide Outbreak of Influenza

How Are Vaccines Made?Flu Vaccine Production

Timeline

1. Surveillance (year round) ID emerging new pathogens (i.e. H5N1)2. Strain selection January - March3. Manufacturing and production January - July4. Purification and testing July - October5. Filing and packaging July - December6. Shipping August - November7. Vaccination October and beyond

Questions:Is this fast enough?Can it handle a fast emerging new pandemic?Can we make enough?Who should get it first?

Issues in Biotechnology

During your time in college you will have what number of sexual partners?

(A) 0-5(B) 5-10(C) 10-25(D) 25-50(E) over 50

Issues in Biotechnology Girls go First


(A) 0-5(B) 5-10(C) 10-25(D) 25-50(E) over 50

Issues in BiotechnologyNow the Boys


(A) 0-5(B) 5-10(C) 10-25(D) 25-50(E) over 50


I support a mandatory HPV vaccine at age 12:

(A) yes(B) no(C) undecided(D) there is still insufficient information

Recombinant Recombinant DNA TechnologyDNA Technologyand Pharmaceuticalsand Pharmaceuticals

Antibody TargetingAllows precise treatments for:

Cancers Cardiovascular diseases Inflammation

Behavior Obesity Depression Schizophrenia

Antibody Based Drugs

Coding sequence for an antibody fragment to a specific antigen

Promoter Coding Sequence Terminator



Monoclonal antibody therapy is the use of monoclonal antibodies (or mAb) to specifically bind to target cells or proteins

Antibody Based DrugsWhat do all the names of these drugs have in common?* Alemtuzumab * Gemtuzumab ozogamicin * Rituximab * Trastuzumab * Ibritumomab tioxetan

They all end in -mab, shorthand for monoclonal antibody

They are all names of monoclonal antibody based drugs targeted to cancer cells

Targeting proteins involved in disease with precise accuracy

Antibody Based DrugsDrug delivery to precise Targets

Lower side effectsHigher efficacy

Antibody Based Technologies

Antibody-nanoparticle drugsmagneticradioactivequantum dots

Antibody-nanoparticle devicescomputersoxygen deliverycommunications

Antibodies for the future

Rheumatoid Arthritis

Rheumatoid Arthritis is an inflammatory disease. It is an example of an AUTOIMMUNE DISEASE because the victim’s own immune system attacks a protein of its own.

Rheumatoid ArthritisAutoimmune diseases arise from an inappropriate immune response of the body against substances and tissues normally present in the body. The immune system of the patient ‘mistakes’ some protein as a pathogen and attacks its own cells. This may be restricted to certain organs (e.g. in autoimmune thyroiditis) or involve a particular tissue in different places (e.g. Goodpasture’s disease which may affect the basement membrane in both the lung and the kidney). The treatment of autoimmune diseases is typically with immunosuppression—medication which decreases the immune response.

Rheumatoid Arthritis is an Overreaction of the Immune System

Immuno-Suppressant TreatmentsIn addition to common anti-inflammatory drugs such as aspirin, which reduce pain, immuno-supressant drugs often induce remission

1. Gold Salts2. Anti-malerials3. Methotrexate (anti-metabolite)4. Corticosteroids5. Herbal treatments

Macrophages, white blood cells that engulf foreign invaders, play an important role in the body’s defense by making TNF alpha and other “cytokines” which cause the inflammation

Chronic Activation of the “Acute” response to infection is damaging to the body and must be halted

Overstimulation of the Immune System causes Inflammatory Disease

Painful

RECEPTORS are proteins on the surfaces of cells that enable them to recognize each other

In 1881, Surgeon William Coley noted that bacterial infections in cancer patients sometimes caused tumors to become “necrotic” or atrophy

In 1975 Anthony Cerami showed that bacteria induced release of a “wasting” factor in an infected host, a small protein that also had anti-tumor activity


Specialized proteins embedded in cell membranes which receive and transmit chemical messages are often desirable drug targets and are referred to as:

(A) random walkers (B) receptors (C) transgressors (D) retractors(E) diseases

Tumor Necrosis Factor - TNF α In 1881, Surgeon William Coley noted that bacterial

infections in cancer patients sometimes caused tumors to become “necrotic” or atrophy

In 1975 Anthony Cerami showed that bacteria induced release of a “wasting” factor in an infected host, a small protein that also had anti-tumor activity

A role in Rheumatoid Arthritis and inflammation (and other diseases)

ENBREL as an antibody mimics the soluble TNF Receptor

• By preventing the tumor necrosis factor from causing damage to the tissue, the progression of Rheumatoid arthritis is slowed down

• Pain secondary to the inflammatory process is reduced also

Antibody

B-cell Receptors

Antigen

Selected Recombinant Products For Medical Problems Affecting Large

Patient Populations

Hemophilia

Benefix recombinant (FIX) Wyeth

Rheumatoid Arthritis

Enbrel (etanercept) Amgen

Kineret (anakinra) Amgen

Remicade (infliximab) Centocor


Recombinate DNA technology has been able to make what class of compounds as a new class of effective drugs?

(A) muscle fibers (B) antibodies and vaccines(C) lipids (D) Cox 2 inhibitors(E) stem cells

Selected Recombinant Products For Medical Problems Affecting Large

Patient PopulationsHepatitis B

Engerix-B (recombinant hepatitis B vaccine) GlaxoSmithKline

Intron A (interferon-a 2b) Schering Corp

Recombivax-HB (recombinant hepatitis B vaccine) Merck & Co., Inc

Acute Myocardial Infarction (heart attack)

Retavase (reteplase) Centocor

TNKase (tenecteplase) Genentech, Inc

Antibodies are used as drugs that are specific to their targets

The Immune System:


Rheumatoid arthritis is:

(A) an autoimmune disease (B) totally eradicated (C) curable with the correct diet (D) has been most effectively treated with homeopathic remedies(E) a consequence of poor health habits

Viruses Used as Drug Delivery Devices

Engineered to not be pathogenic

Exquisite cell or target specificity

(e.g. HIV specifically targets T4 Lymphocytes)

Suicide delivery agents

Viruses Used as Gene Delivery Devices

An approach to AIDS Treatment?

(e.g. HIV specifically targets T4 Lymphocytes)

Suicide delivery agents

Viruses for Gene TherapyAn approach to Gene Therapy?

Cell specific gene delivery

Replacement of defective genes

Cancer treatments

Addition of new genes

Genetic Surgery

8. Antibiotic resistant tuberculosis is on the rise world-wide. Which approach to research treatment development would not be the best choice? (A) develop a vaccine using recombinant DNA technologies (B) develop RNAi methods to target the tuberculosis bacteria(C) develop cheaper methods to make the antibiotic (D) develop early detection methods based on PCR(E) sequence the tuberculosis genome to look for new drug targets

9. Rheumatoid arthritis is: (A) an autoimmune disease (B) totally eradicated (C) curable with the correct diet and vitamins (D) has been most effectively treated with homeopathic remedies(E) best treated early with surgery

10. Specialized proteins embedded in cell membranes which receive and transmit chemical messages are often desirable drug targets and are referred to as: (A) random walkers (B) receptors (C) transgressors (D) retractors(E) transducers

11. Enzymes are: (A) only used in commercial detergents(B) genes involved with biochemical pathways (C) made primarily of lipid (D) not involved with energy production(E) usually proteins that catalyze reactions in cells

12. Recombinant DNA technology has been able to make what class of compounds as a new class of effective drugs? (A) muscle fibers (B) antibodies (C) lipids (D) Cox 2 inhibitors(E) homeopathic treatments

13. What are the implications of gene cloning for the pharmaceutical industry? (A) technically a good idea but all candidates have failed in Phase III trials (B) it might work but it will never gain public acceptance (C) drugs based on antibodies are now on the market made using this technology (D) technically a good idea but has yet to be proven (E) none, it’s the materials of science fiction and Hollywood movies

14. The ability to replace defective genes in a patient, as a sort of genetic surgery, has not yet been effectively achieved and is called: (A) gene therapy (B) chiral chemistry (C) combinatorial chemistry (D) recombinate drug technology(E) alternative therapy

15. HPV stands for: (A) High Purity Vaccine(B) Hallmark Pneumonia Vaccine(C) Henrietta’s Park Virus(D) Human Papilloma Virus(E) Human Pancreatic Virus

16. An influenza pandemic is a global outbreak of disease that occurs when a new influenza A virus appears or “emerges” in the human population, causes serious illness, and then spreads easily from person to person worldwide. Such a pandemic: (A) has only occurred once in recorded human history(B) is only the material of Hollywood movies such as “Contagion”(C) is only a matter of time before another occurrence (D) is totally preventable(E) proves that the theory of evolution is incorrect since viruses cannot evolve

© life_edu lecture 21 part ib. emergent technologies: dna-based biotechnology and pharmaceutical...

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