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Biotechnology in Medicine > Biotechnology Modterm #3 > Flashcards

Biotechnology Modterm #3 Flashcards

1
Q

Goal of Regenerative Medicine

A

Accelerate the pace at which the body heals itself to a clinically relevant timescale

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2
Q

What is regenerative medicine

A

Process of replacing, engineering, or regenerating human cells, tissues or organs to restore or establish normal function

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3
Q

Bioreactor

A

Device that supports a biologically active environment, think of a vessel that hosts organic chemical processes

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4
Q

The regenerative medicine toolbox

A

Clinical translation—>Stem Cell Therapy, Tissue Engineering, Biomaterials

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5
Q

Scope of global stem cell clinical trials

A

As of January 2018, over 5000 stem cell clinical trials have been performed

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6
Q

Stem Cells

A

Foundational cells for every organ and tissue in the body—they replace cells that are injured or lost

Have 3 key properties:
1.) Ability to self-renew
2.) Unspecialized
3.) The ability to differentiate

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7
Q

Cell Potency

A

Cell’s ability to differentiate into other cell types

Totipotent (whole), Pluripotent (Many), Multipotent (Several)

Totipotent cells can give rise to the placenta and the embryo

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8
Q

3 types of stem cells

A

Embryonic stem cells, Adult stem cells, Induced pluripotent stem cells

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9
Q

Embryonic Stem cells

A

Pluripotent stem cells derived from the inner cell mass of a blastocyst–renew indefinitely

Can differentiate into any of the primary germ layers, which include 220 cell types in the body

Ectoderm (Exoskeleton)
Mesoderm (develop into organs)
Endoderm (form inner lining of organs)

But this can form teratomas (tissues from more than one germ layer)

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10
Q

1996 Dickey-Wicker Amendment

A

Prohibits the use of federal funds towards research involving the creation or destruction of human embryos

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11
Q

2000 NIH guidelines for human pluripotent stem cells

A

Human embryonic stem cells:

1.) Must be derived with private funds from frozen embryos from fertility clinics
2.) Must have been created for fertility treatment purposes
3.) Must be in excess of the donor’s clinical need
4.) Must be obtained with consent of the donor

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12
Q

Major events in stem cell research policy

A

President Bush 2001Executive Order
1.) Prohibited federal funding for human embryonic stem cell research
2.) Only cell lines derived prior to August 2001 could be used
3.) This did not affect private sector or state funding

In 2009, Obama reverses the 2001 Executive Order
1.) Removed barriers to responsible scientific barriers involving embryonic stem cells

2016 21st Century Cures Act
1.) Assures timely regulatory review of regenerative therapies, including cell therapies enabled by stem cell therapy research

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13
Q

First Human Embryonic Stem cell Clinical Trial Occurred in 2001

A

Transplantation of oligodendrocytes derived from human embryonic cells into spinal cord injured individuals

There have been neither adverse effects or improvements reported

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14
Q

VitaCyte

A

Developed through a regiment of adding and removing growth factors at precise developmental timepoints

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15
Q

Therapeutic Cloning

A

Somatic cell nuclear transfer harnesses the potential to use one’s own DNA–the same technique used during reproductive cloning

An enucleated egg has one’s genetic material inserted and then it develops into a blastocyst

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16
Q

Adult Stem Cells

A

Undifferentiated and multipoint cells existing within differentiated tissues

Can be tissue specific stem and progenitor cells, bone marrow stem cells, adult mesenchymal stem cells, or amniotic fluid and cord blood stem cells

Can renew themselves a number of times, but not indefinitely

Ethically uncomplicated

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17
Q

Hematopoietic stem cells

A

Give rise to all blood cell types-myeloid lineage and lymphoid lineage

Need 100 billion new blood cells each day, and HSCs are the only source of this new blood

Clinically used for blood disorders and types of leukemia

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18
Q

Stem Cell Niche

A

The microenvironment and the physical anatomical positions where stem cells are found

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19
Q

Autologous Hematopoietic Stem Cell Transplant

A

Immunosuppressive chemotherapy treatment combined with rein fusion of blood stem cells to rebuild the immune system

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20
Q

Mesenchymal Stem Cells

A

Typically derived from bone and marrow stroll cells, although there are several sources

Specialized cells of skeletal tissues
1.) Angiogenesis (blood vessels)
2.) Osteogenesis (bone)
3.) Chondrogenesis (Cartilage)
4.) Adipogenesis (fat)
5.) Myogenesis (muscle)

Important for bone defects, cardiac repair, cartilage repair

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21
Q

Umbilical Cord and Amniotic Stem Cells

A

Are stem cells harvested from younger sources more viable?

Cord blood: Hematopoietic stem cells
Cord Tissue: mesenchymal stem cells

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22
Q

Amniotic Stem Cells

A

Mixture of stem cells from amniotic fluid and membrane

Can differentiate into cells of all three germ layers

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23
Q

Induced Pluripotent Stem Cells

A

Identified in 2007. They are pluripotent stem cells derived from a non-pluripotent source, typically adult somatic cell, by inducing a forced expression of certain genes

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24
Q

Safety Concerns for unproven stem cell treatments

A

1.)Administration site reactions
2.) The ability of cells to move from the placement sites and changing into inappropriate cell types or multiply
3.) Failure of the cells to work as expected
4.) The growth of tumors

In August 2017, the FDA announced an increased enforcement of regulations and oversight of stem cell clinics–including administrative actions, judicial and criminal enforcement

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25
Q

Although over 5000 clinical trials the list of FDA approved is very limited

A

Only stem cell based products that are FDA approved in the U.S. consist of blood-forming stem cells (hematopoietic progenitor cells) derived from cord blood.

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26
Q

Great Demand for Tissue Engineering

A

Projected US market of around 33 billion USD in the coming decade

There are now over 20 FDA approved commercial products to date including skin and orthopedic replacements

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27
Q

Goal of tissue engineered products

A

To be implanted and regenerate function of the tissue or organ

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28
Q

After In Vitro Generation of the tissue

A

Implantation should lead to an in vivo regeneration of the lost/failing function

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29
Q

The biomaterial and tissue engineering toolbox

A

Signaling molecules (inductive)
1.) Signal cells with instructions
2.) Proteins and growth factors
3.) Mechanical stimulation

Cells (productive)
1.) Living part of tissue
2.) Provides function
3.) Produces proteins
4.) Gives tissue reparative properties

Scaffold (conductive)
1.) Provide structural support, shape, and place for cell
2.) Biocompatible
3.) Sometimes biodegradable

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30
Q

Three tools

A

Scaffold/matrix
1.) Usually degradable, porous

Soluble factors
1.) made by cells or synthetic
2.) various release profiles

Cells
1.) Precursors and/or differentiated
2.) Usually autologous

All this creates an integrated implantable or injectable device

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31
Q

Challenges to biomaterial and tissue engineering

A

Biological structures and functions are complex and require dynamic interactions between cells, scaffolds, and soluble molecules

Difficulties:
1.) Biomaterial rejection
2.) Density and variety of cells
3.) Cost of tissue and organ development
4.) Vascularization

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32
Q

Proof of Concept: Earmouse

A

Created in 1997

Biodegradable ear shaped scaffold

Seeded with cow chondrocytes

Immunosuppressed mouse strain

Mass protests erupted against genetic engineering–even though no form of genetic manipulation was performed

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33
Q

World’s first synthetic, tissue engineered organ transplant

A

In Sweden in 2011, a cancer patient needed a new trachea to survive

Although the transplant was a low hanging fruit and the trachea was purely mechanical, it was an important first step for the field

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34
Q

What is the current status of the artificial trachea

A

There have been 15 additional trachea transplants– all of appeared to be a short term fix

Research has considered different biomaterials for better long term success, although the misconduct cases have halted some of this progress

Are surgeries examples of successful tissue engineering that utilize the special abilities of stem cells or an elaborate temporary fix that is destined to fail

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35
Q

Understanding the design criteria

A

What type of tissues and functions are being replaced?
Structural v. Functional (like metabolism)

Is vascularization required? (biggest challenge to tissue engineering right now)
Cartilage v. Cardiac repair

Ex vivo or in situ or paracorporeal?
Prefabricated implant v. Stem cell infusion v. Bio-artificial liver
–in situ: stem cells, infuse them, growth/regenerations happens inside alongside physiological changes

Cell types, scaffolds, soluble factors?
Cell sources, Synthetic v. Natural, and Bound v. Free

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36
Q

Diabetic Ulcers

A

Occurs in 15% of patients with diabetes. The condition precedes 84% of all diabetes related leg amputations

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37
Q

Most of the cells found in a wound closed with a loving skin equivalent are from the…

A

host, not the graft

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38
Q

Future of wound healing

A

Look at picture

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39
Q

Cartilage

A

Has a complex 3 dimensional shape, but also has low cell density and is avascular

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40
Q

CARTICEL and MACI

A

Look at slide and 2 slides underneath

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41
Q

There remains a need for partial and whole bladder regeneration

A

Pediatric population:
1.) Abnormal development
2.) Neurogenic bladder associated with spina bifida

Adults:
1.) Prone to anatomical and functional loss (surgery, radiation, repeated infections, cancer)

Aging Population:
1.) Muscle under activity

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42
Q

Complex Layer of the bladder

A

Urothelium: impenetrable barrier that allows for urine containment

Lamina propria: houses the vasculature needed for oxygen and nutrients

Muscle: fascinates urine storage and coordinates the movements needed to expel urine

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43
Q

Therapeutic management can range from conservative to minimally invasive to surgery

A

Current standard of care consists of augmenting the bladder with portions of GI tissue

This approach, however, lacks barrier properties and can lead to:

1.) Tendency to absorb waste products
2.) Infections
3.) Potential malignancy

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44
Q

The regenerative medicine approach consists of urinary bladder matrix being ECM derived from porcine urinary bladder

A

The urinary bladder matrix consists of epithelial membrane and lamina propria

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45
Q

Urinary Bladder Matrix increases bladder compliance

A

Compliance: Increase in pressure per unit of volume. Normally the bladder is very compliant and may be filled to large volumes with very little increase in pressure

Bladder compliance increases by the same percentage comparing ileum augmentation to urinary bladder matrix

Urinary bladder matrix does not have associated risks like GI tissue

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46
Q

Autologous cell regenerative approach

A

Scaffold made of collagen + polyglycolic acid, seeded with autologous cells

Engineered bladder connected to failing bladder

Covered with fibrin glue and omentum

36 months did not improve compliance—perhaps because autologous cells were already failing

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47
Q

Understanding decellularized scaffolds

A

Decellularization typically consists of both physical (temperature, force, pressure)
and enzymatic steps (immersion or perfusion)

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48
Q

Perfusion decellularization is primary method for whole organ decellularization

A

Cadaveric organ–kidney (decellularization), decellularized organ–kidney scaffold (repopulation), bioengineered organ–regenerated kidney (transplantation)

LOOK AT SLIDE

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49
Q

Bladder Acellular Matrix

A

Direct implantation of this scaffold allows for regenerationof urothelium–but insufficient muscle development.

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50
Q

Lung tissue engineering

A

Design considerations:
1.) Extensive surface area
2.) Very thin alveolar-capillary membranes
3.) Viscoelastic behavior

Synthetic scaffolds? Not enough control to recapitulate hierarchy needed

3D bioprinters? Do not have the resolution to create the gas exchange capillary-alveolar capillary network

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51
Q

The future of lungs

A

Look at picture

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52
Q

Complex bioprinting process

A

Look at picture

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53
Q

Comparison of Bioprinter Types

A

Look at picture

Step 1: Imaging
Step 2: Design Approach
Step 3: Material Selection
Step 4: Cell Selection
Step 5: Bioprinting
Step 6: Application

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54
Q

A Comparison of Bioprinter Types

A

Look at picture

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55
Q

Examples of human-scale bio-printed tissues

A

Inkjet–Skin and Cartilage
less cell density

Microextrusion–vasculature, valve, and kidney

Laser–tracheal splint

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56
Q

Studies demonstrate that we can use tissue engineering to produce grafts for medium to large vasculature, creating microvasculature remains elusive

A

Scaffolds were composed of polyactic acid (PLA) and spun into porous nano fiver conduits using electro spinning technology

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57
Q

Microvasculature to engraft organs

A

Resolution for capillaries remains a problem a problem—we can incorporate angiogenic growth factors such as vascular endothelial growth factor (VEGF) to nurture capillary growth

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58
Q

Can they perfuse?

A

Media can indeed circulate through

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59
Q

Another potential solution

A

Look at picture

3D printed filament network->encapsulate network and living cells-> dissolve network-> place in media-> fibrin, collagen, material, agarose, ECM Mimics-> seed endothelial or stem cells into the network

60
Q

The Magnitude of The Human Genome

A

Each somatic cell in the human body (except for mature red blood cells and platelets, which do not contain a nucleus) contains a complete copy of the human genome:

1.) Approximately 3.2 billion base pairs
2.) Organized into 23 pairs of chromosomes
3.) An estimated total of 30,000 genes

61
Q

Gene therapies

A

Products that harness the Central Dogma–the principles of transcription and translation–to modify genetic information and integrate it to a patient as nucleic acids, viruses, or genetically engineered microorganisms

The products may be used to modify cells in vivo or transferred to cells ex vivo prior to administration to the recipient

62
Q

Gene Therapies are regarded as a potential revolution in the health sciences and pharmaceutical fields

A

The number of clinical trials investigating gene therapies is increasing worldwide, despite the limited number of products that have successfully reached the market

63
Q

To treat a genetic disease, we must first identify the….

A

causing gene, but not all diseases are that simple

64
Q

Monogenic diseases

A

Result from changes to a single gene- occurring in all cells of the body

Examples:
1.) Thalassaemia
2.) Sickle cell anemia
3.) Hemophilia
4.) Cystic Fibrosis
5.) Tay Sachs Disease
6.) Spinal muscular atrophy
7.) Huntington’s Disease

Scientists estimate that over 10,000 human diseases are monogenic disorders

65
Q

Somatic vs Germ Cell Gene Therapy

A

Look at table

66
Q

For a gene therapy to be approved, two important criteria must be met:

A

1.) There must be effective therapeutic genes that can be expressed at specific target sites
————-Specificity
————-Effective transfer to target cell and target cell nucleus

2.) There must be an efficient and safe deliver system
———Protection against premature degradation in blood and intracellular (endosomal) degradation
———Entry through cytoplasmic and nuclear membranes

67
Q

Gene therapies can’t be administered as a simple over-the counter pill

A

Special vectors must be used

68
Q

Adenovirus vectors release the gene freely into the nucleus

A

The gene is not incorporated and readministration is often required

69
Q

Retrovirus vectors insert RNA and enzymes

A

these incorporate randomly into genome (integrase)

70
Q

Comparison of commonly used viral vectors in gene therapy

A

Look at table

71
Q

In vivo gene therapy

A

1.) The therapeutic gene is cloned into the genome of a recombinant virus

2.) The recombinant virus carrying the therapeutic gene is infused into the patient

3.) The cells that are transducer by the recombinant virus produce the therapeutic protein

72
Q

Ex vivo gene therapy

A

1.) Cells from the patient are extracted via a biopsy and expanded in culture

2.) Cells are transduced using a recombinant virus carrying the therapeutic gene

3.) Transduced cells that produce the therapeutic protein are infused back into the patient

73
Q

There are benefits and drawbacks to both methods –which is used ultimately depends on the specific therapy

A

In vivo
1.) Less invasive
2.) Technically simple
3.) Vectors introduced directly
4.) Safety check not possible
5.) Decreased control over target cells

Ex Vivo
1.) More invasive
2.) Technically Complex
3.) No vectors introduced directly
4.) Safety check possible
5.) Close control possible

74
Q

Immunotherapy

A

Treatment that harnesses a person’s immune system to fight diseases such as cancer

This can be done in a couple of ways:
1.) Stimulating your own immune system to work harder or smarter to attack cancer cells
2.) Providing your immune system with extra components–such as man-made immune system proteins

While traditional cancer therapies kill both cancerous and healthy cells, cancer immunotherapies can selectively kill cancerous cells and minimize side effects

75
Q

Process of immunotherapy

A

Look at picture

76
Q

CAR-T cell therapy

A

Chimeric Antigen receptor-T cell therapy is a type of treatment in which a patient’s T cells are genetically modified to attack cancer cells.

T cells focus on recognizing and attacking specific foreign antigens or particles

77
Q

CD-19

A

Molecule exclusive to B-cells—lymphocytes that produce antibodies.

Since B-cells are a common marker for certain cancers and are also not necessary for survival, CAR-T can target B-Cells for patients with CD19+ lymphoma and leukemia

Look at picture for CD-19 targeted CAR-T cell therapy

78
Q

Kymirah

A

First gene therapy treatment approved for the treatment of patients up to 25 years of age with B-Cell precursor acute lymphoblastic lymphoma that is refractory or in second or later relapse.

There is a life threatening risk of cytokine release syndrome and neurological effects

79
Q

Yescarta

A

2nd gene therapy treatment approved (2017) and the first approved to treat Non-Hodgkin Lymphoma (NHL)

There are now over 30 approved gene therapy treatments

80
Q

Understanding the Retina

A

Layer in the back of the eye containing cells that are sensitive to light and trigger nerve impulses that pass via the optic nerve to the brain–where a visual image is formed

The retinal pigment epithelium is a single layer of cells that acts as a barrier to the retina as well as providing protection and stability

81
Q

RPE65 and Rhodopsin

A

RPE65 is critical to resetting the visual cycle and responsible for rhodopsin production–the most abundant protein in the rod cells that is the primary photoreceptor molecule of vision ‘
When RPE65 is mutated photoreceptor cells (rods and cones) die

In degenerated retina (picture below) the cycle is stuck in trans

82
Q

The proposed gene therapy–RPE65 Leber Congenital Amaurosis

A

The FDA advisory committee voted unanimously to approve this therapy in 2017. 90% of study participants report increased sight within days

83
Q

Epidermolysis Bullosa patients have skin as delicate as butterfly wings–it falls apart with the slightest touch

A

Several forms of this disease result from different genes-all of which affect either cell adhesion molecules, basement membrane proteins, or matrix proteins

Patients often have “mitten hands” due to frequent wound healing and the formation of scar tissue

84
Q

Genetically modified cells for an unmutated LAMB3 regenerate the skin—a gene responsible for a majority of epidermolysis bulls cases.

A

Lamin is the protein network foundation for most cells and organs

85
Q

Bacteria’s First Line of Defense

A

Bacteriophages are a type of virus that infect bacteria

To prevent viral disease, CRISPR evolved as sequences of DNA that are derived from bacteriophage genes that have previously infected the bacteria

When the bacteriophage attacks again, their newly inserted genes are recognized and removed by CRISPR-Cas9

86
Q

CRISPR-Cas system gives bacteria acquired immunity… but it can also be used for other purposes

A

Evolution has given us a tool that can recognize specific sequences of nucleotides and remove them from the genome

We have genetic scissors at our disposal

87
Q

CRISPR-Cas9 can be used to select specific genes and entirely remove them from a genome

A

The Cas9 enzyme takes in a guide RNA of our gene of interest and the target DNA

After unwinding the DNA, Cas9 checks for sites complementary to the 20 base pair spacer region of the guide RNA

If the DNA substrate is complementary to the guide RNA, Cas9 cuts the DNA

88
Q

While bacterial Cas9 requires of two different RNA strands–crRNA to serve as reference and tracrRNA to activate the CRISPR mechanism

A

Engineering has allowed for a linker loop to bind the two RNA’s into a singular strand. This is the form of CRISPR we use today

89
Q

Cut-Copy-Paste

A

In the cases of genetic diseases caused by a single gene, it may be useful to cut out a portion of the genome–yet we must often replace DNA as opposed to just removing it

Look at image below

90
Q

There are two ways in which the DNA strand can be repaired after the cut

A

Non-Homologous End joining mends the broken strand right back together

Homology-Directed Repair adds a new serious of nucleotides to the cut area

91
Q

The He Jiankui Affair

A

In a secret experiment, Dr. He Jiankui used CRISPR to genetically edit two twin girls known as Nana and Lulu

The two embryos were edited of their CCR5 gene in an attempt to confer genetic resistance to HIV—this is the first and only recorded case of a genome edited human babies

Although the twins were born healthy, the experiment was met with outrage from the scientific community. Jianjui was arrested in December of 2019 for 3 years and must pay a fine of around half a million dollars

92
Q

Understanding the CCR5 Gene

A

Codes for CCR5 receptors that act as the pathway for the HIV virus into the cell. Typically, HIV binds to the receptors and injects its virulent DNA through CCR5 gene

By removing the CCR5 gene, these receptors are not made and HIV can no longer infect cells

Why the outrage?

Apart from ethical and social considerations, the CCR5 gene is also linked to improved memory function, enhanced recovery from strokes, and cancer. Additionally, the gene was only deleted—leading to further risk of infection and unintended mutations.

93
Q

While the NaLu experiment was done in vivo, most medical applications are being designed to be ex vivo

A

Look at picture

94
Q

As of today…

A

CRISPR clinical trials only edit somatic tissue cells without affecting germ cells, meaning that no DNA changes can be passed onto future generations

95
Q

Vertex and Blood Disease-Casgevy

A

Red blood cells use hemoglobin to pick up oxygen in the lungs and carry it to all the tissues of the body. Genetic mutations in Hemoglobin leads to different disorders: beta thalassemia and sickle cell disease

Current CRISPR trials these aim to increase levels of fetal hemoglobin…a form of hemoglobin only made by fetuses in the womb that can replace defective adult hemoglobin in red blood cells

96
Q

In the first use of an ex vivo CRISPR-based therapy to cure a genetic disease

A

One patient with beta thalassemia and one with SCD have been successfully treated

97
Q

Leber congenital amaurosis 10

A

Caused by mutations in the CEP290 gene

Since LCA-10 is same condition as caused to the RPE65 gene and it has an approved gene therapy, why can’t we administer gene therapy for CEP290 gene the same way

98
Q

A refresher on RPE65 gene therapy

A

RPE65 is critical to resetting the visual cycle and responsible for rhodopsin production—the most abundant protein in the rod cells that is the primary photoreceptor molecule of vision

When RPE65 is mutated, photoreceptor cells (rods and cones) die

99
Q

Gene Editing Scheme for CEP290 gene

A

In march 2020, a CRISPR-modified virus was injected into a set of patients eyes in an attempt to treat LCA.

While the procedure didn’t work for all of the patients and none of the patients have regained normal vision, there have been notable improvements with no significant side effects have occurred

100
Q

Somatic v Germline

A

Look at Venn diagram

101
Q

Editing An Embryo 101

A

in vitro fertilisation (IVF)–> CRISPR/Cas9 gene-editing–>Implantation of gene-edited embryo into mother

102
Q

Prenatal Screens and Genetic Abnormalities

A

Even without genetic editing, a certain type of selective pressure has been exerted with the rise of prenatal screens that test of genetic diseases

The suspicion of a genetic defect often leads to the termination of a pregnancy (around 92% of pregnancies in Europe in which Down syndrome is detected are terminated)

What if we could just prevent it?

103
Q

Why edit ourselves?

A

Prevent genetic diseases that cause suffering and death based on heredity and genetics

For all those that follow by removing disease-causing genes from the gene pool

Cure diseases that are currently only treatable such as herpes and HIV

Drive our evolution towards a healthier and disease free future, exerting absolute control over out health

104
Q

Although CRISPR shows great promise, there are several hurdles to be resolved before it can become widespread

A

Immune responses to the bacterial parts of CRISPR may render the technology useless. A majority of tested blood samples showed existing immune responses to Cas9

Cas9 is large, so its gene is difficult to deliver to cells via vectors such as adeno-associated viruses commonly used in gene therapy

Off-target effects must be mediated to minimize unintended mutations

The need for PAM sequences limits potential target sequences

105
Q

Sometimes genes differ not only between individuals in a population, but also among the cells of an individual. The advent of cheap and rapid genome sequencing has revealed that this mosaicism is incredibly common

A

Mosaicism is a serious problem

How that affects the health of the resulting child would depend on which cells were edited and which were not–something that could be difficult to predict in advance

106
Q

Repairing the Broken

A

The cell’s repair processes are unpredictable

The first studies to try using the CRISPR genome-editing technique to alter the DNA of human embryos revealed edits corrected mutations in a small proportion of embryos—only 1 in 10 cells were successfully repaired

Since these embryos were very genetically abnormal, however, these experiments may not have given an accurate indication of how well the technique would work in healthier embryos

107
Q

Cas9 has been known to cut DNA at unintended non-target sites, particularly when there are DNA sequences in the genome similar to the target

A

These off target cuts can result in health problems: a change to a gene that suppresses tumor growth, for example, might lead to cancer

108
Q

p53 and Cancer

A

Following CRISPR edits, cells have inherent mechanisms that respond quickly to this type of DNA damage–and the transcription factor p53 is at the center of these

CRISPR has been shown to work best in p53-deficient cells, deleting or reducing p53 increased the number of surviving cells

By selects for p53-deficient cells, edited cells are vulnerable to mutagenesis and can result in tumors

109
Q

Criteria for a Safe Edit

A

In 2017, the US National Academies of Sciences, Engineering, and Medicine outlined the conditions that should be met before editing a human embryo that is destined for implantation

110
Q

USNASEM Criteria

A

1.) Absence of reasonable alternatives
2.) Restriction to preventing a serious disease
3.) Restriction to editing genes that strongly predispose to the disease or condition
4.) Availability of credible preclinical or clinical data on risks and health benefit of the procedure
5.) Ongoing oversight to the effects of the procedure on health and safety
6. Concrete plans for long-term multigenerational follow up while still respecting personal autonomy
7.) Absolute transparency consistent with patient privacy
8.) Continued reassessment of health and societal benefits and risk, with participation and input by the public
9.) Oversight mechanisms to precent extension to uses other than preventing disease

111
Q

The Future: Enter RNA and Base Editing

A

REPAIR release on CAS13 to edit RNA nucleosides involved in single-base diseases

REPAIR allows for edits to be temporary (and even reversible) and the genome to remain untouched

Research has shown success in editing RNA in human kidney cells to fix mRNA, leading to functional proteins. Cas 13, just like Cas9, can be edited to select its target gene.

112
Q

World Population Age 80 or Older

A

1950: 14.5 million

2014: 70 million

2023: 167 million

2050: 394.7 million

Eventually individuals may live to 120 years old

113
Q

What is aging?

A

Evolutionary natural selection has no reason to keep our “mechanisms” working once we have passed the reproductive years–our only purpose genetically speaking is the propagation of our genes

114
Q

Scientifically

A

Aging is a process that converts an optimally healthy, fit organism into a less healthy, less fit organism

1.)Reduced tissue/physiological function
2.)Increased susceptibility to disease (age-related diseases)
3.)Decreased resistance to stress (physical and psychological)

Aging is a biological process-not a disease

115
Q

The case for calories and primates

A

Look at Picture

116
Q

Restricting calorie intake enhances lifespan in animals

A

Studies have shown that 30-40% calorie restriction without malnutrition extends a healthy lifespan by 40-50% in worms, flies, mice, rats, and potentially monkeys

117
Q

Aging

A

A combination of our chromosomes, genes, and environment

118
Q

Cellular senescence

A

Refers to a process in which cell growth and development is irreversible halted. Senescent cells no longer divide, nut maintain metabolically active

119
Q

Senscence Process

A

Look at Picture

120
Q

Telomere

A

Region of repetitive nucleotide sequences at each end of a chromosome, which protects the end of the chromosome from deterioration or from fusion with neighboring chromosomes

121
Q

Hayflick limit

A

The number of cell divisions a normal cell can undergo before its telomeres are too short—typically 50 to 70 cell divisions

122
Q

Removing senescent cells to prevent aging

A

Proxofim is a small peptide that clears senescent cells and stimulates surrounding stem cells to create new tissue, with results seen within 10 days

123
Q

Species-specific longevity genes

A

Life spans ranging from 2-3 weeks to 100-200 years

Flies (weeks)
Nematodes (months)
Mice (<4 years)
Humans (<100 years)
Turtles

Look at picture above

124
Q

Aging in Mice and Men

A

look at graph

125
Q

Genes that have been shown to affect stress resistance and lifespan in laboratory models

A

Human homologs do not show the same relationship

126
Q

Sirtuins

A

Longevity gene, originally studied in yeast (SIR2), that has been shown to be significant to cellular processes in mammals (SIRT1)

In yeast:

Increased sirtuins=Increased lifespan

Decreased sirtuins= Decreased lifespan

127
Q

Role of sirtuins and aging

A

1.) Energy expenditure
2.) Senescence
3.) Stress Resistance
4.) Histone acetylation/ deacetylation

Sirtuins respond to decreases in nutrient availability and cellular stress to promote cell survival by deacetylating histone and non-histone targets in the nucleus.

Deacetylation promotes survival rather than apoptosis

128
Q

High SIRT1 activity promotes deaceytlation of…

A

Ku70, FOXO and p53, (transcription factors) promotes DNA repair and cell-cycle arrest, and blocks apoptosis induction

129
Q

Resveratrol

A

Molecule present in red wine and manufactured by other plants in response to stress

High concentration pill of resveratrol extended mouse lifespan 44%

130
Q

Putting it all together

A

look at 3 slides

131
Q

Has modern medicine changed the demographics of aging?

A

Virtually all of the topics that have been covered in this course have been developed either to enhance or prolong life

132
Q

Cancer immunoediting role in evading the immune system

A

Elimination, Equilibrium, Escape

133
Q

Unleashing the power of the immune system to defeat cancer

A

Immunotherapy mobilizes the body’s own natural defense system to fight diseases and is revolutionizing the way cancer Is treated

134
Q

Immunomodulators

A

Substances that can help support immune function by modifying, generally in a beneficial way, the immune system’s response to a threat

Immunomodulators act as the gas and breaks of the immune system

135
Q

Immunomodulator targets relevant to the immune response to cancer

A

Immunomodulator targets can be characterized as checkpoint inhibitors, cytokines, adjuvants

136
Q

Checkpoint inhibitors

A

Such as PD-1/PDL-1 and CTLA-4 are among the most understood

Immune checkpoint inhibitors work by blacking checkpoint proteins from binding with their partner proteins. This prevents the “off” signal from being sent, allowing the T cells to kill cancer cells

Used to treat a range of cancer including melanoma, lung, kidney, liver, bladder, cervical, and colorectal cancer

137
Q

The use of immunomodulators extends beyond therapeutic uses for cancer

A

Research has shown that there is a complex immune response in most individuals, and even a severe cytokine storm in some so it is not surprise that a diverse group of immunomodulators are being investigated

138
Q

Engineering Oncolytic Virus to Target Cancer

A

(+) INSERT: Immune-stimulating genes

(-) REMOVE: Disease causing genes (selective targeting of tumors)

139
Q

Oncolytic viruses represent a promising approach to treating cancer for several reasons

A

1.) Cancer seeks often have impaired antiviral defenses that make them susceptible to infection

2.) These natural viruses can be engineered to give them advantageous properties, including decreasing their ability to infect healthy cells as well as granting them the ability to deliver therapeutic payloads specifically to tumors and produce immune-boosting molecules once they infect tumor cells

3.)After infection, these oncolytic viruses can cause cancer cells to burst, killing the cancer cells and releasing cancer antigens. These antigens can then stimulate immune responses that can seek out and eliminate any remaining tumor cells nearby and potentially anywhere else in the body

140
Q

Oncolytic Viruses target cancer cells

A

Look at picture and picture below

141
Q

The first oncolytic virus to receive FDA approval was a treatment for melanoma known as talimogene laherparepvec, or T-VEC

A

T-VEC has a two phase mechanism, consisting of primary ablation and secondary tumor-specific immune response

Melanocyte: specialized skin cell that produces the protective skin-darkening pigment melanin

142
Q

Cancer vaccines can be classified as preventative, therapeutic, or personalized

A

Preventive Vaccines: play an important role in reducing risk for viral infections that can aid the development of several cancers

Therapeutic cancer vaccine: Vaccine which is administered after the cancer already occurred. It activates the immune system of the patient to fight towards the already developed cancer

Personalized cancer vaccines: Create an immune response directed precisely against patients’ unique antigens on their tumor cells while sparring their healthy cells from immune attack, thus possibly preventing side effects

143
Q

Cancer Vaccine targets relevant to the immune response to cancer

A

look at table

144
Q

Therapeutic Vaccines

A

Can be cell-based, protein based, or nucleic acid based

145
Q

There are two FDA approved therapeutic vaccines

A

Sipuleucel-T (Provenge): Vaccine composed of patients’ own stimulated dendritic cells; approved for prostate cancer

Bacillus Calmette-Guérin (BCG): a vaccine that uses weakened bacteria to stimulate the immune system; approved for patients with early-stage bladder cancer

146
Q

Neoantigens (New antigens)

A

Antigens exclusive to tumor cells and not by healthy cells

With neoantigen vaccines, it is conceivable that immune responses could be directed precisely against patients’ tumor cells while sparing their healthy cells from immune attack, thus possibly preventing side effects

147
Q

How can you make immunotherapy the most efficient?

A

Look at picture below

Biotechnology in Medicine (1 decks)

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