Cellular Therapies

Question 1

what is the clinical need for cellular therapies?

Answer 1

> 130 million individuals globally
Chronic/degenerative/ acute diseases
NO TREATMENTS

this presents significant challenges for the health care systems and for patients/families, suffering, social and economic losses

Question 2

what potential do cellular therapies have?

Answer 2

potential to provide curative treatments for many diseases with unmet needs

Question 3

define cellular therapy

Answer 3

Administration of live human cells to a patient for repair/replacement/ regeneration of damaged tissue and/or cells

Question 4

what kinds disorder are cellular therapies being invistagted to treat?

Answer 4

Baldness
Brain injury - stroke, MS, cerebral palsy, parkisons, AD
Blindness
Hearing Loss
Spinal Cord Injury
Bone fractures
CF
Heart disease
Liver Failure
Diabetes
Cancer

Question 5

what therapies are currently licenced?

Answer 5

ChondroCelect- repair of knee cartilage (

MACI - repair of knee cartilage

Provenge - Prostat cancer

Holoclar - cornea epithelium replacement

Stimvelis - adenosine deaminase deficiency

Zalmoxis - haematological cancers

Question 6

what are the aims of cell and gene therapy catapult?

Answer 6

Bridge the gap between business, academia, research and government

Accelerate the growth of cell therapy industry in UK

Question 7

what are the 2 classifications of cell therapy?

Answer 7

Autologous (patients own cells)

Allogenic (donor cells)

Question 8

explain autologous cell therapy

Answer 8

Immunological compatibility

No HLA matching/immunosuppression required

Resembles an individualised procedure

Heterogeneous - donor variability

Imprecisely characterised

Stringent traceable logistics:
Collection
Transport
Manufacture/manipulation
Administration back to patient

Manual process- high production work load

Expensive

Question 9

explain allogenic cell therapy

Answer 9

Risks of: immune response/rejection/ graft-versus-host disease

Immunosuppression

May ↓ product function, other risks

Standardised product- guaranteed dose
Cell bank

Simplified supply: off-the-shelf product

Automated process- less labour intensive

Lower costs

Question 10

what are the cell types under clinical development?

Answer 10

1) stem cells
2) terminally differentiated cells
3) genetically modified cells

Question 11

what are stem cells?

Answer 11

single cells with unique characteristics such as:

replicate itself to maintain stem cell pool
retain its undiff state

can diff into many cell types by switching on specific genes in response to external/internal chemical signals

replace dead/damaged cells throughout life

Question 12

what is the stem cell hierarchy?

Answer 12

classified according to potential to develop into other cell types (plasticity:
1)totipotent - most versatile, produced in first few cell division in embryonic development, differentiate into any adult cell type, give rise to entire organism

2) pluripotent - originate from 5 day old embryo, can differentiate into any adult cell types
3) mulitpotent ie. blood stem cells and other stem cells (muscle, nerve, bone)

Question 13

how are stem cells relevant to therapies?

Answer 13

ESC (pluripotent) - grown in lab from cells of early embryo

iPSC (pluripotent) - made from adult specialised using lab technique

adult/somatic stem cell (multipotent) - found throuhghout body, found in children and adults

Question 14

what are embryonic stem cells?

Answer 14

derived from inner cell mass of 5 day old embryo

ability to differentiate into cell types of three primary germ layers

equates to >200 diff cell types in adult human

Question 15

ethical dilemma of using ESC?

Answer 15

derived from embryos produced via IVF

donated for research with informed consent of donors

balance of preventing/alleviating suffering vs respecting the value of life

research is tightly regulated, illegal in austria, denmark, france, germany and ireland

used in UK strictly controlled by HEFEA

Question 16

what disorders are ESC being used for in trials currently?

Answer 16

age-related macular degeneration - retinal pigment epithelium

parkinsons disease - a9 dopaminergic neuron

spinal cord injury-oligodendrocyte progenitor

diabetes-pancreatic islet b-cell progenitor

MI- cardiomyocytes

Question 17

what are iPSCs?

Answer 17

produced by reprogramming terminally differentiated cells:

1) terminally diff cells removed from patient
2) transduced with stemm cell associated genes using viral vectors
3) cells reprogrammed
4) directed to differentiate

iPSCs behave like embryonic stem cells-can differentiate into a variety of cells types

Question 18

how can iPSC be used therapeutically?

Answer 18

1) patient skin biposy or other tissue
2) cells cultured in vivo
3) cells reprogrammed back to expandable iPSC
4) Directed to differentiate into clinically useful cell types
5) autologous transfer to treat original patient
6) treat individual with:disease, injury, inherited disorder or age related tissue degeration

Question 19

what are the pros of iPSCs?

Answer 19

Eliminate ethical issues of ESC use 

Derived from patient’s own cells: ↓ immuno-rejection

Ability to differentiate → any cell type

Unlimited proliferation capacity 

Readily accessible (skin, blood cells)

Opportunities for personalised treatments

Potential for preservation (cell banks)

Possibilities of gene correction therapies

Question 20

what are the cons of iPSCs?

Answer 20

Low rate of reprogramming

Cells from patient with genetic disease still carry same defective gene

High level of genetic instability in culture

Pluripotency genes (c-Myc) is oncogene- overexpression could cause cancer

Retroviral vectors insert pluripotency genes randomly → genome = undesirable mutations

Question 21

what are the challenges to overcome before iPSC suitable for clinical use?

Answer 21

Regulatory requirement: cells with stable/consistent characteristics.

2014: Japan, patient administered autologous iPSC-derived retinal pigment epithelium cells for treatment of age-related macular degeneration
2015: trial halted - mutations identified in iPSCs produced for second patient- arose during reprogramming process

Concern: transplantation of genetically unstable cells → uncontrolled cell growth/tumour formation

Question 22

what are adult or somatic stem cells?

Answer 22

multipotent: limited to differentiating into specialised cell types within tissue of origin

found in many organs and tissues: stem cell niches

remain undifferntiated until activated: maintan tissue homeostasis, disease, tissue injury.

Question 23

adult stem cells in current clinical practice

Answer 23

haematopoietic stem cell translplantation - most frequently used cell therapy. Obtained from peripheral blood, bone marrow and umbilical cord blood

SC, diff into all the types of blood cell.
SC can be autologous or allogenic

treatment of: malignant/non-malignant blood disorders
genetic disorders of immune system

Question 24

what are mesenchymal cells (MSC)?

Answer 24

example of adult stem cell, derived from: bone marrow, adipose tissue, placenta and umbilical cord, peripheral blood.

Capable of:
differentiating into cell types of mesenchymal tissues
transdifferentiating into non-mesenchymal cells (neurons, epithelial)

Question 25

what is the therapeutic potential of MSC?

Answer 25

Comes from their ability to: secrete solutble factors crucial for cell survival and proliferation

modulate immune response

differentiate into various cell types

transdifferentiate-non mesoderm

migrate to sites of injurt in response to cell signals

Question 26

MSC clinical trials

Answer 26

Globally: 463 registered clinical trials

296 employ autologous MSC

Cover wide range of therapeutic applications:
Cardiovascular
Autoimmune
Osteoarthritis
Liver disorders
GvHD

Major sources of MSC in clinical trials:
Bone marrow
Umbilical cord
Adipose tissue

No approved treatments using MSC

Question 27

how can we exploit use of adult stem cells?

Answer 27

Commercial companies provide services to isolate/store stem cells
UK: Future Health Biobank, Cell4life, Precious cells

Biological insurance: individual/child/ family member develops a disease in the future treatable by stem cells
One off charge (+maintenance fees)

Cells sources:
umbilical cord
placenta blood
adipose tissues
dental pulp from child's teeth

Question 28

what are terminally differentiated cells?

Answer 28

Specialized cells unable to proliferate and have reached the final stage of development

Question 29

what current practice involve TDCs?

Answer 29

donor platelet transfusions

platelet deficiency due:
disease
treatment related- chemotherapy
life threatening bleeding - injur injury or trauma

autologous platelet rich plasma therapy

Question 30

what is platelet rich plasma therapy?

Answer 30

platelets rich source of growth factors which stimulate development soft tissue/bone cells

procedure claims to initiate faster healing response

blood collected, centrifuged to seperte blood, plateley rich plasma collected, injected into injured area

used in treatment of ortho conditions:
Osteoarthritis
	Tendonitis
	Tendon tears
	Nerve injury
	Professional athletes with muscle and ligament injuries (Tiger Woods

Question 31

what other practice involves TDCs?

Answer 31

red cell transfusions

treat anaemia:
heavy blood loss
bone marrow not producing enough red cells (chemo, leukaemia/sickle cell)

autologous erthyrocyte encapuslated enzyme replacement therapy

Question 32

what is autologous erythrocyte encapuslated enzyme replacement therapy?

Answer 32

Under clinical development at SGUL for treatment of diseases due to inherited enzyme deficiencies

1) deficient enzyme encapsulated in patients erythrocytes in vitro
2) intravenously administered to the patient
3) permit elimination of pathological plasma metabolites:

pathologically elevated plasma metabolite permeate cell membrane

encapsulated enzyme catalyses substrate–> normal product

normal product exits cell to enter usual metabolic pathway

this is applicable to disorders where pathologically high metabolite permeate erythrocyte membrane

Question 33

what are the benefits of a therapeutic approach?

Answer 33

majority licesence enzyme replacement therapies (ERT)

High enzyme activity half-life:

• short plasma half-life, frequent infusions:maintain therapeutic effective levels

low immunogenic reactions and production of anti-enzyme antibodies:

anti-enzyme antibody production leads to loss of therapeutic efficacy
allergic reactions

Question 34

what is the encapsulating process?

Answer 34

1) erythrocytes + exogenous enzyme undergo hypo-osmotic dialysis
2) erythrocytes swell; formation of pores in membran
3) exogenous enzyme enters cell and iso-osmotic dialysis occurs.
4) erthyrocytes reseal, encapsulating therapeutic agent

Question 35

what are the applications for erythrocyte mediated enzyme replacement therapy?

Answer 35

treatment two autosomal recessive metabolic disorders:

1)Mitochondrial NeuroGastroIntestinal Encephalomyopathy (MNGIE)
Product: erythrocyte encapsulated thymidine phophorylase (EE-TP)

2) Adenosine deaminase deficiency
product: erythrocyte encapsulated adenosine deaminase (EE-ADA)

Question 36

explain the metabolic defect in MNGIE

Answer 36

mutatuon in the nuclear gene (TYMP) which encodes for thymidine phosphorylase (TP)

mitochondiral deoxynucleotide pool imbalance leading to:

1) increased [plasma] of thymidine and deoxyuridine
2) impaired replication/repair leading to multiple deletions, point mutations and depletion

Ultimarley loss of mitochondiral respiratory chain function.

Question 37

what are the symptoms of MNGIE?

Answer 37

Mainly effects GI and nervous system.

GI symptoms:
Severe gastrointestinal dysmotility 
Psuedo-obstruction
Nausea/vomiting
Chronic abdominal pain
Premature satiety
LEADS TO:
Malabsorption
Bacterial over-growth
Intestinal perforation
Loss of muscle mass

Neuro symptoms:
Leukoencephalopathy
Demyelination nerve fibres
Initially patchy

Ocular symptoms
Ptosis
Loss of vision

Peripheral neuropathy
Numbness
Muscle weakness
Hearing loss

patients die avg age 37 combo of nut and neuromusc failure

Question 38

clinical experience with EE-TP in MNGIE

Answer 38

Bax et al (2013) demonstrated:
Clinical improvements in sensory ataxia (balance and gait) & fine finger functioning
Increase in body weight
Walk longer distances, climb stairs without assistance, tie shoe laces
Returned to public performances as guitar player in a band
Previously reported numbness in hands/feet resolved

Question 39

what types of genetically modified cells are under clinical investigation?

Answer 39

1) gene modified autologous stem cell therapies

2) engineered T-cell therapies

Question 40

what are gene modified autologous stem cell therapies?

Answer 40

autologous stem cells collected from patient and genetically corrected prior to reinfusion

eg: HSC-directed gene therapy
HSCs are multipotent, capable of generating entire epctrum of blood/lymphoid cells

suitable target therapies for: haematological malignancies
iherited blood disorders

Question 41

steps of gene modified autologoues stem cell therapies

Answer 41

approved 2016: treatment of inherited severe combined immunodeficiency disease: adenosine deaminase deficiency

1)HSCs isolated from bone marrow

2)CD34+ cells expanded in culture
Transduces with retroviral vector expressing fucntional copy of defective gene (adenosine deaminase)

3) endogenous bone marrow progenitors eliminated to favour engraftment
Modified stem cells infused.

Gene-corrected stem cells reconsititute lymphoid lineages/restore immune function

Question 42

what are enginereed t cell therapies?

Answer 42

tumour cells often recognised as ‘self’ - prevents T cells from recognizing tumour proteins

solution: genetically alter T-cells to create recognition receptors unique to patients tumour
altered t cells expanded in culture and infused into patient which seek out/destroy tumour cells

two approached:
t-cell receptor therapies (TCR)

chimeric antigen receptor (CAR-T) therapies

Question 43

steps of TCR therapy

Answer 43

1) peripheral blood lymphocytes removed from patient
2) cells transduced with viral vector containing contruct encoding for tumour reactive TCR
Expanded in culture
Infused into patient
3) Infused cells express modified TCR on surface
4) recognise tumour-specific proteins on the inside of cell through encountering tumour antigen peptide processed and presented on cell surface

5)tumour destruction

Question 44

what is CAR-T?

Answer 44

chimeric=composed of parts from two or more different sources

CAR= artificial receptor that links:

1)antibody molecule:polypeptide sequences of light and heavy chain from antibody recognises target proteins expressed on cancer cell surface

and

2)t-cell signalling machinery of t-cell receptor:
on binding to target- clonal expansion, secretion of cytokines to recruit immune system, destruction of tumour cell

Question 45

steps of CAR-T therapy

Answer 45

1) peripheral blood lympocytes removed from patient

2)cells transduced with viral vector containing CAR construct
expanded in culture
infudes into patient

3)expressed CAR recognises external antigen leading to death of targeted cancer cells

Question 46

what are the challenges faceing cell therapy commercialisation?

Answer 46

Turning cells into effective/safe/ affordable therapies poses many challenges: