1. Stem Cell Therapies for Inherited and
Acquired Disorders
Tim Townes, Ph.D.
Director, UAB Stem Cell Institute
Chairman, Department of Biochemistry & Molecular
Genetics
UAB School of Medicine

2. Hematopoiesis

3. Sickle Cell Disease
GAG to GTG at codon 6 of the beta-globin gene
Glutamic Acid to Valine substitution produces hydrophobic projection
Hemoglobin polymers cause red blood cell sickling and severe anemia
Occlusions of small vessels cause:
ke, kidney disease, liver disease, acute chest syndrome, retinopathy, painful crises, e

4. Sickle Hemoglobin Polymers
sagittal transverse
In vivo sickling

5. Left Internal Carotid Artery Occlusion
In UAB Sickle Patient

6. Mouse Model of Sickle Cell
Disease
Science 245: 971-973 (1989)
Science 247: 566-568 (1990)
Science 278: 873-876 (1997)
Science 318:1920-1923 (2007)

7. Hematopoiesis

8. Isolation of Highly Purified Hematopoietic
Stem Cells
Sca+
c-Kit+
Lin-

9. HumanAS3-Globin Lentiviral SIN Vector
432
SD
AS3globin
LCR
WPRE
5’ LTR
SA
RRE
3’ Enh
y cPPT
3’ D LTR
SIN
e yG A
45 123
Globin
Locus

10. Correction of Sickle Cell Disease
in Mice
Sickle Control Lentivirus AS3

11. Sickle Cell Disease
GAG to GTG at codon 6 of the beta-globin gene
Glutamic Acid to Valine substitution produces hydrophobic projection
Hemoglobin polymers cause red blood cell sickling and severe anemia
Occlusions of small vessels cause:
ke, kidney disease, liver disease, acute chest syndrome, retinopathy, painful crises, e

12. Modified from
Murry and Keller
Cell 132:661-680
Feb. 22, 2008
ntES cells derived from skin

13. Mouse Model of Sickle Cell
Disease
Science 245: 971-973 (1989)
Science 247: 566-568 (1990)
Science 278: 873-876 (1997)
Science 318:1920-1923 (2007)

14. Human A-Globin Knock-In To Correct SCD
Atk HygVect
Targeting Vector
Human AKnock-In
5’ 3’
1.7 7.0
1234567
ey h0 h1 h2 S
1234567
ey h0 h1 h2 S
1234567
ey h0 h1h2 A Hyg
1234567
ey h0 h1 h2 S

15. Correction of Sickle Cell Disease by
Homologous Recombination
Control Sickle Corrected
hA/hA hS/hS hA/hS
Blood,108:1183 (2006)

16. Modified from
Murry and Keller
Cell 132:661-680
Feb. 22, 2008
iPS cells derived from skin

17. Scheme for Gene and Cell Therapy
of Humanized Sickle Mouse
Science 318:1920-1923
Dec 21, 2007

18. Wright-Giemsa Stained Blood Smears
Science 318:1920-1923
Dec 21, 2007

19. Correction of Sickle Disease

20. Sickle Cell Disease
GAG to GTG at codon 6 of the beta-globin gene
Glutamic Acid to Valine substitution produces hydrophobic projection
Hemoglobin polymers cause red blood cell sickling and severe anemia
Occlusions of small vessels cause:
ke, kidney disease, liver disease, acute chest syndrome, retinopathy, painful crises, e

21. Human Sickle Skin Fibroblasts
Gene Replacement in Sickle Skin
Fibroblasts?
45 year old sickle patient

22. Homologous Recombination in Human
Sickle Fibroblasts
Sickle Human Fibroblasts Corrected Sickle Human Fibroblasts
A

23. hESC-H1 c-iPSC-1
anti-TRA-1-60
anti-TRA-1-81
anti-SSEA-4
DAPI
DAPI
DAPI
hESC-H1 c-iPSC-1
2.5 X 10 X
iPSC Derived From Corrected Human Sickle
Fibroblasts

24. Corrected Human Sickle iPS Cells Form Tissue Derived
From All Three Germ Layers
Intestine Smooth Muscle and Gland Neural Ectoderm

25. Normal Karyotype of Corrected Human Sickle iPS Cells

26. Skin cell
biopsy
Derive iPS
cells
Correct Sickle
Gene in
patient
fibroblasts
Culture into blood
stem cells
Transplant
corrected blood
stem cells
into patient
Sickle
Patient
Gene Replacement Therapy for Sickle
Cell Disease

27. Hematopoiesis

29. GMP Cell Processing Lab Larry Lamb and Shin Mineishi
Sanyo/Panasonic Cell Processing Work Station (CPWS)

30. Modified from
Murry and Keller
Cell 132:661-680
Feb. 22, 2008
iPS cells derived from skin