Stem cells and cell therapy. lecture

Transcription

1 Stem cells and cell therapy lecture

2 What is a stem cell? Stem cell can differentiate into two or more cell types Progenitor cells differentiates into single cell type

3 Classification of SCs 1. Based on the tissue commitment/ differentation capacity Totipotent SCs - zygote Pluripotent SCs Multipotent SCs Unipotent SCs (Tissue progenitors) 2. Based on their origin Embryonic SCs (ESCs) Fetal SC ( adult SC) Umbilical cord blood stem cells ( adult ) Postnatal - Adult SCs (tissue-specific SCs) Reprogrammed SCs (Induced Pluripotent SCs = ips cells)

4 What are embryonic stem cells? - develop from eggs fertilized in vitro - derived from 4-5 days old embryos - isolated from ~ 8 cell embryo or inner cell mass

5 Human embryonic stem cells Obtained in 1998 by J. Thompson The National Institutes of Health resource for stem cell research

6 Embryonic stem cells are pluripotent 9 Human ESC, after being differentiated to a specific cell type, can be potentially used for the therapy

7 Stem cells in therapy examples

8 Neuronal stem cells and Parkinson Disease Neurodegenerative disease - loss of dopamine neurons Fourth most common neurodegenerative disease Who is under the risk? 1% of people > 65 years 0.4% of people > 40 years

9 Clinical study Transplantation of embryonic dopamine neurons for severe Parkinson s Disease Dopamine neurons in embryonic tissue In vitro culture for 4 weeks Transplantation Control group: sham surgery

10 Results Transplanted cells survived Growth of the transplants was observed

11 Clinical study Freed CR et al. N Engl J Med Mar 8;344(10):710-9.

12 Embryonic stem cells in therapy of human diseases

13 Important! In therapy no one will use un-differentiated embryonic stem cells! The ESC before application to human are differentiated to the specific cell type However, the risk of contamination with un-diiferentiated (not fully differentiated) ESC exists There is always a risk of rejection of transplanted cells due to genetic differences with the host

14 Oligodendrocytes from hesc - Oligodendrocytes have several functions: * produce myelin * produce neurotrophic factors - Oligodendrocytes are lost in spinal cord injury, resulting in myelin and neuronal loss that cause paralysis in many patients with spinal cord injuries. oligodendrocytes grown on axons in culture

15 Embryonic stem cell-based therapy - GRNOPC1 contains hesc-derived oligodendrocyte progenitor cells. - The cells have demonstrated remyelinating and nerve growth stimulating properties leading to restoration of function in animal models of acute spinal cord injury. - Preclinical studies showed that administration of GRNOPC1 significantly improved locomotor activity and kinematic scores of animals with spinal cord injuries when injected seven days after the injury. - Histological examination of the injured spinal cords treated with GRNOPC1 showed improved axon survival and extensive remyelination surrounding the rat axons.

16 Embryonic stem cells in therapy Geron Corp. - USA clinical trial in patients after severe spinal cord injury ESCs differentiated to oligodendrocytes (halted: The Scientist, ) Adcanced Cell Technology (ACT) Statdgard s blindness affects humans at years, uncurable ESCs differentiated to pigment epithelial cells Science 30 September 2011 Problem: genetic disparity of stem cells and host

17 Human ESC ethical and practical problems 1. Classic ESC are genetically different from the host 2. Side effects teratocarcinoma formation 3. ESC are obtained from the surpluss embryos generated by in vitro fertilisation (for reproductive purposes) (in vitro fertiliation only for research pursposes is forbidden in most countries) 4. Ethical problems for belivers that human entity starts from the conception

18 Producing embryonic stem cells by somatic cell nuclear transfer (SCNT) Byrne JA et al. (Mitalipov), Nature 22 November 2007

19 How to obtain embryonic stem cells identical with the patient? Therapeutic cloning=scnt somatic cell nuclear transfer (to enucleated oocyte)

20 Traditional SCNT is not (was not ) effective in human G.O. Daley, Nature, 6th October 2011 Triploid human ESCs have been recently obtained by S. Noggle et al. (D. Egli) Nature, 6t October 2011

21 Human embryonic stem cells obtained by SCNT M. Tchibana (S. Mitalipov), Cell, 15 maja 2013

22 What about adult stem cells in therapy?

23 Stem cells 1. Somatic ( adult ) stem cells 1.1. Fetal, tissue specific stem cells 1.2. Umbilical cord bloods stem cells 1.3. bone marrow hematopoietic mesenchymal proangiogenic (endothelial) progenitor cells (?) 1.4. skin 1.5. nervous (brain) 1.6. intestine 1.7. other adult, organ specific (heart, fat tissue) 1.8. embryonal carcinoma cells

24 Adult stem cells bone marrow hematopoietic stem cells

25 Adult stem cells bone marrow mesenchymal stem cells hematopoietic stem cells

26 HSC hematopoietic stem cells K.Szade, 2012 Szade et al., PLoS One, 17 maja 2013

27 Functional capacity Differentiation potential The hierarchical structure of differentiation stem cells progenitor cells terminally differentiated cells

28 Limbal stem cells in treatment of blindess caused by damage (mostly burn-caused) The autologous limbal cells were used for the transplantation Permanent restoration of a transparent, renewing corneal epithelium was attained in 76.6% of eyes. The failures occurred within the first year. Restored eyes remained stable over time, with up to 10 years of follow-up (mean, 2.91±1.99; median, 1.93). P. Rama (G. Pellegrini) et al., NEJM 2010

29 Hematopoietic stem cells in vivo bone marrow cord blood peripheral blood

30 Isolation of HSC

31 Isolation of HSC 2. Peripheral blood mobilization continuos blood flow up to 20 L are proceed during one aspiration

32 Autologous transplantation of HSC - procedure

33 HSC in cardiac regeneration Coronary heart disease Ischemia limitation of blood flow - cell starvation Myocardial infarction (heart attack) lack of blood - death of myocardial muscles Occlusion of the artery Rapid ischemia Apoptosis of myocytes Collagen replacement - fibrous tissue Expansion of the infarct zone

34 Delivery Application of stem cells

35 HO-1 overexpression in pig bone marrow mesenchymal stem cells Separation of BM mononuclear cells differentiation and expansion of cells GFP and HO-1 adenoviral transfer CD105, CD29, CD34, CD31 Injection into ischemic myocardium Wojakowski et al., Thrombosis & Haemostasis, 2012; Aug 7;108(3).

36 Modification of pig mesenchymal stem cells improves recovery after myocardial infarction Number of arms HMEC (48 h) MSC-GFP (26 h) tube formation * 150 MSC (48 h) MSC-HO-1 (26 h) GFP HO-1 8 h Wojakowski et al., Thrombosis & Haemostasis, 2012; Aug 7;108(3).

37 HO-1 positive and GFP positive cells in the infarct border zone HO-1 GFP GFP, DAPI 5.2. Wojakowski et al. Thromb Haemos. 2012

38 Healing the burn or diabetic wounds Stem cells proliferation is the key step in wound healing Supply of stem cells: basal layer of epidermis bulge sebaceous glands

39 SC and burns treatment Cells characterisation: Easy to isolate Easy to culture human keratinocytes Dr Justyna Drukała

40 Tissue engineering - Tissue engineering combines advances in cell biology and materials science. These tissues consist of biocompatible scaffolding material, plus living cells grown using cell culture techniques. * Simple tissues, such as skin and cartilage, were the first to be engineered successfully.

41 Adult stem cells in therapy P. Rama (G. Pellegrini) et al., NEJM 2010 P. Bianco et al., EMBO J, 3 May, 2013

42 embryonic vs adult in terms of cell therapy embryonic SC adult SC potency telomerase expression pluripotent yes unipotent multipotent no culturing easily grown hard to obtain large numbers of cells stem cell therapy rejection problem no rejection problem

43 Is the other way possible? Human Dermal Fibroblasts Stem Cell

44 Induced pluripotent stem cells (ipscs) Shinya Yamanaka 2006

45 Nobel Prize 2012 Sir John Gurdon & Shinya Yamanaka Rossant & Mummery, Nature, 6th December 2012

46 J. Stępniewski i wsp Reprogramming of murine fibroblasts Mysie IPS uzyskane po transdukcji fibroblastów genami OSMK w wektorze lentiwirusowym SSEA-1

47 Application of ipscs not only regenerative medicine Annals of Internal Medicine, 155:L 117: 2011

48 ips-based gene therapy

49 Therapeutic capacities of IPSC 1. Odróżnicowanie fibroblastów myszy z anemią sierpowatą do ips 2. Naprawa mutacji poprzez transfer genu 3. Różnicowanie naprawionych ips do komórek hematopoetycznych (HSC) 4. Przeszczepienie naprawionych HSC myszom po uprzednim naświetleniu 5. Korekta układu krwiotwórczego Rudolf Jaenisch Jaenisch i wsp, Nature 2008

50 Turystyka komórkowa - Stem cells tourism China cracks down on stem cell tourism 00:01 04 September 2009 by Andy Coghlan But not only China. Not only in exotic countries Warnings are being issued by experts of the dangers of medical tourism saying that unproven stem cell therapy overseas could leave patients worse off.