Biotechnology Modterm #3 Flashcards
Goal of Regenerative Medicine
Accelerate the pace at which the body heals itself to a clinically relevant timescale
What is regenerative medicine
Process of replacing, engineering, or regenerating human cells, tissues or organs to restore or establish normal function
Bioreactor
Device that supports a biologically active environment, think of a vessel that hosts organic chemical processes
The regenerative medicine toolbox
Clinical translation—>Stem Cell Therapy, Tissue Engineering, Biomaterials
Scope of global stem cell clinical trials
As of January 2018, over 5000 stem cell clinical trials have been performed
Stem Cells
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
Cell Potency
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
3 types of stem cells
Embryonic stem cells, Adult stem cells, Induced pluripotent stem cells
Embryonic Stem cells
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)
1996 Dickey-Wicker Amendment
Prohibits the use of federal funds towards research involving the creation or destruction of human embryos
2000 NIH guidelines for human pluripotent stem cells
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
Major events in stem cell research policy
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
First Human Embryonic Stem cell Clinical Trial Occurred in 2001
Transplantation of oligodendrocytes derived from human embryonic cells into spinal cord injured individuals
There have been neither adverse effects or improvements reported
VitaCyte
Developed through a regiment of adding and removing growth factors at precise developmental timepoints
Therapeutic Cloning
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
Adult Stem Cells
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
Hematopoietic stem cells
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
Stem Cell Niche
The microenvironment and the physical anatomical positions where stem cells are found
Autologous Hematopoietic Stem Cell Transplant
Immunosuppressive chemotherapy treatment combined with rein fusion of blood stem cells to rebuild the immune system
Mesenchymal Stem Cells
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
Umbilical Cord and Amniotic Stem Cells
Are stem cells harvested from younger sources more viable?
Cord blood: Hematopoietic stem cells
Cord Tissue: mesenchymal stem cells
Amniotic Stem Cells
Mixture of stem cells from amniotic fluid and membrane
Can differentiate into cells of all three germ layers
Induced Pluripotent Stem Cells
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
Safety Concerns for unproven stem cell treatments
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
Although over 5000 clinical trials the list of FDA approved is very limited
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.
Great Demand for Tissue Engineering
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
Goal of tissue engineered products
To be implanted and regenerate function of the tissue or organ
After In Vitro Generation of the tissue
Implantation should lead to an in vivo regeneration of the lost/failing function
The biomaterial and tissue engineering toolbox
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
Three tools
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
Challenges to biomaterial and tissue engineering
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
Proof of Concept: Earmouse
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
World’s first synthetic, tissue engineered organ transplant
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
What is the current status of the artificial trachea
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
Understanding the design criteria
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
Diabetic Ulcers
Occurs in 15% of patients with diabetes. The condition precedes 84% of all diabetes related leg amputations
Most of the cells found in a wound closed with a loving skin equivalent are from the…
host, not the graft
Future of wound healing
Look at picture
Cartilage
Has a complex 3 dimensional shape, but also has low cell density and is avascular
CARTICEL and MACI
Look at slide and 2 slides underneath
There remains a need for partial and whole bladder regeneration
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
Complex Layer of the bladder
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
Therapeutic management can range from conservative to minimally invasive to surgery
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
The regenerative medicine approach consists of urinary bladder matrix being ECM derived from porcine urinary bladder
The urinary bladder matrix consists of epithelial membrane and lamina propria
Urinary Bladder Matrix increases bladder compliance
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
Autologous cell regenerative approach
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
Understanding decellularized scaffolds
Decellularization typically consists of both physical (temperature, force, pressure)
and enzymatic steps (immersion or perfusion)
Perfusion decellularization is primary method for whole organ decellularization
Cadaveric organ–kidney (decellularization), decellularized organ–kidney scaffold (repopulation), bioengineered organ–regenerated kidney (transplantation)
LOOK AT SLIDE
Bladder Acellular Matrix
Direct implantation of this scaffold allows for regenerationof urothelium–but insufficient muscle development.
Lung tissue engineering
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
The future of lungs
Look at picture
Complex bioprinting process
Look at picture
Comparison of Bioprinter Types
Look at picture
Step 1: Imaging
Step 2: Design Approach
Step 3: Material Selection
Step 4: Cell Selection
Step 5: Bioprinting
Step 6: Application
A Comparison of Bioprinter Types
Look at picture
Examples of human-scale bio-printed tissues
Inkjet–Skin and Cartilage
less cell density
Microextrusion–vasculature, valve, and kidney
Laser–tracheal splint
Studies demonstrate that we can use tissue engineering to produce grafts for medium to large vasculature, creating microvasculature remains elusive
Scaffolds were composed of polyactic acid (PLA) and spun into porous nano fiver conduits using electro spinning technology
Microvasculature to engraft organs
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
Can they perfuse?
Media can indeed circulate through
Another potential solution
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
The Magnitude of The Human Genome
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
Gene therapies
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
Gene Therapies are regarded as a potential revolution in the health sciences and pharmaceutical fields
The number of clinical trials investigating gene therapies is increasing worldwide, despite the limited number of products that have successfully reached the market
To treat a genetic disease, we must first identify the….
causing gene, but not all diseases are that simple
Monogenic diseases
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
Somatic vs Germ Cell Gene Therapy
Look at table
For a gene therapy to be approved, two important criteria must be met:
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
Gene therapies can’t be administered as a simple over-the counter pill
Special vectors must be used
Adenovirus vectors release the gene freely into the nucleus
The gene is not incorporated and readministration is often required
Retrovirus vectors insert RNA and enzymes
these incorporate randomly into genome (integrase)
Comparison of commonly used viral vectors in gene therapy
Look at table
In vivo gene therapy
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
Ex vivo gene therapy
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
There are benefits and drawbacks to both methods –which is used ultimately depends on the specific therapy
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
Immunotherapy
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
Process of immunotherapy
Look at picture
CAR-T cell therapy
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
CD-19
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
Kymirah
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
Yescarta
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
Understanding the Retina
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
RPE65 and Rhodopsin
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
The proposed gene therapy–RPE65 Leber Congenital Amaurosis
The FDA advisory committee voted unanimously to approve this therapy in 2017. 90% of study participants report increased sight within days
Epidermolysis Bullosa patients have skin as delicate as butterfly wings–it falls apart with the slightest touch
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
Genetically modified cells for an unmutated LAMB3 regenerate the skin—a gene responsible for a majority of epidermolysis bulls cases.
Lamin is the protein network foundation for most cells and organs
Bacteria’s First Line of Defense
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
CRISPR-Cas system gives bacteria acquired immunity… but it can also be used for other purposes
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
CRISPR-Cas9 can be used to select specific genes and entirely remove them from a genome
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
While bacterial Cas9 requires of two different RNA strands–crRNA to serve as reference and tracrRNA to activate the CRISPR mechanism
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
Cut-Copy-Paste
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
There are two ways in which the DNA strand can be repaired after the cut
Non-Homologous End joining mends the broken strand right back together
Homology-Directed Repair adds a new serious of nucleotides to the cut area
The He Jiankui Affair
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
Understanding the CCR5 Gene
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.
While the NaLu experiment was done in vivo, most medical applications are being designed to be ex vivo
Look at picture
As of today…
CRISPR clinical trials only edit somatic tissue cells without affecting germ cells, meaning that no DNA changes can be passed onto future generations
Vertex and Blood Disease-Casgevy
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
In the first use of an ex vivo CRISPR-based therapy to cure a genetic disease
One patient with beta thalassemia and one with SCD have been successfully treated
Leber congenital amaurosis 10
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
A refresher on RPE65 gene therapy
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
Gene Editing Scheme for CEP290 gene
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
Somatic v Germline
Look at Venn diagram
Editing An Embryo 101
in vitro fertilisation (IVF)–> CRISPR/Cas9 gene-editing–>Implantation of gene-edited embryo into mother
Prenatal Screens and Genetic Abnormalities
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?
Why edit ourselves?
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
Although CRISPR shows great promise, there are several hurdles to be resolved before it can become widespread
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
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
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
Repairing the Broken
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
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
These off target cuts can result in health problems: a change to a gene that suppresses tumor growth, for example, might lead to cancer
p53 and Cancer
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
Criteria for a Safe Edit
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
USNASEM Criteria
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
The Future: Enter RNA and Base Editing
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.
World Population Age 80 or Older
1950: 14.5 million
2014: 70 million
2023: 167 million
2050: 394.7 million
Eventually individuals may live to 120 years old
What is aging?
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
Scientifically
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
The case for calories and primates
Look at Picture
Restricting calorie intake enhances lifespan in animals
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
Aging
A combination of our chromosomes, genes, and environment
Cellular senescence
Refers to a process in which cell growth and development is irreversible halted. Senescent cells no longer divide, nut maintain metabolically active
Senscence Process
Look at Picture
Telomere
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
Hayflick limit
The number of cell divisions a normal cell can undergo before its telomeres are too short—typically 50 to 70 cell divisions
Removing senescent cells to prevent aging
Proxofim is a small peptide that clears senescent cells and stimulates surrounding stem cells to create new tissue, with results seen within 10 days
Species-specific longevity genes
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
Aging in Mice and Men
look at graph
Genes that have been shown to affect stress resistance and lifespan in laboratory models
Human homologs do not show the same relationship
Sirtuins
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
Role of sirtuins and aging
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
High SIRT1 activity promotes deaceytlation of…
Ku70, FOXO and p53, (transcription factors) promotes DNA repair and cell-cycle arrest, and blocks apoptosis induction
Resveratrol
Molecule present in red wine and manufactured by other plants in response to stress
High concentration pill of resveratrol extended mouse lifespan 44%
Putting it all together
look at 3 slides
Has modern medicine changed the demographics of aging?
Virtually all of the topics that have been covered in this course have been developed either to enhance or prolong life
Cancer immunoediting role in evading the immune system
Elimination, Equilibrium, Escape
Unleashing the power of the immune system to defeat cancer
Immunotherapy mobilizes the body’s own natural defense system to fight diseases and is revolutionizing the way cancer Is treated
Immunomodulators
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
Immunomodulator targets relevant to the immune response to cancer
Immunomodulator targets can be characterized as checkpoint inhibitors, cytokines, adjuvants
Checkpoint inhibitors
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
The use of immunomodulators extends beyond therapeutic uses for cancer
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
Engineering Oncolytic Virus to Target Cancer
(+) INSERT: Immune-stimulating genes
(-) REMOVE: Disease causing genes (selective targeting of tumors)
Oncolytic viruses represent a promising approach to treating cancer for several reasons
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
Oncolytic Viruses target cancer cells
Look at picture and picture below
The first oncolytic virus to receive FDA approval was a treatment for melanoma known as talimogene laherparepvec, or T-VEC
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
Cancer vaccines can be classified as preventative, therapeutic, or personalized
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
Cancer Vaccine targets relevant to the immune response to cancer
look at table
Therapeutic Vaccines
Can be cell-based, protein based, or nucleic acid based
There are two FDA approved therapeutic vaccines
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
Neoantigens (New antigens)
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
How can you make immunotherapy the most efficient?
Look at picture below
