Stem Cells and Neuroregeneration

Douglas Kerr MD/PhD Associate Professor, Neurology Johns Hopkins Hospital Director, JH Transverse Myelitis Center

Outline of Talk: 3 Stem Cell Vignettes
• Using bone marrow stem cells to reconstitute immune tolerance in patients with aggressive multiple sclerosis • ES cell-derived motor neurons as a model of Spinal Muscular Atrophy • Glial Progenitors for Demyelinating Disorders

HiCy: Immune Ablation without Transplantation
• A single course of a high dose chemotherapy agent used to reset the immune system to a tolerized state • HiCy eliminates mature and maturing WBC. • Bone marrow Stem Cells not killed by HiCy. • Aldehyde dehydrogenase inactivates Cy. • Immune System reconstituted from endogenous stem cells. “Re-booted” Immune system may be tolerant of autoantigen.

HiCy Protocol
• Cy: 50 mg/Kg/day x 4 days. Hydration, mesna, ondansetron. Anti-infection prophylaxis. • Follow daily until hematologic recovery: Give RBC, platelets, etc. as needed. • No additional immunomodulatory therapy • Re-immunize at 1 year.

Advantages of HiCy
• Unlike HSCT, peripheral blood cells are not re-infused. Less likely contamination. • Much lower cost (30K vs. 300 K) • Lower mortality (< 0.5% vs 5-7%)

Use of HiCy in aggressive RRMS
• Phase 1/2 Clinical Trial with patients failing other therapies
– 2 exacerbations in last year AND – 2 gad enhancing lesions on each of two baseline MRIs AND – Increase in disability (>1.0 EDSS) – ALL WHILE ON CONVENTIONAL THERAPY

Hematologic Recovery after HiCy

EDSS Change After HiCy

Summary Disability Scores

P <0.02

Summary MRI Activity

P < 0.01

Conclusions: HiCy Phase I/II
– HiCy is safe and induces rapid and sustained clinical and radiologic improvement – Patients with advanced disease continue to have progression of disability while those with early disease have an improvement in function – 42% Reduction in disability at 24 months compared to baseline in entire cohort – Phase III Trial Being Considered
• HiCy + glatiramer vs. glatiramer vs. HiCy • Multi-Center

The ultimate stem cells: self-renew indefinitely and differentiate into all types of human cells

Fischbach et al, JCI, 114:2004

Differentiation of ES cells to Motor Neurons
ES cells/ Fibroblasts (undifferentiated) ES cells without Fibroblasts SHH/RA Day +1 Sox2+ Islet1/2+ Hb9+ NeuN+ Day +2 Day +3 Day +4
Wichterle et al, Cell

Day -1 Day 0

Day +5

Harper et al. Proc Natl Acad Sci U S A. 2004 May 4;101(18):7123-8.

C2C12 myoblasts


Axon outgrowth in vitro

Neuromuscular Junction Formation In Vitro

• Developmental biology of motor unit formation • Pathologic motor unit formation

2) In Vivo Transplantation of ES CellDerived Motor Neurons in Paralyzed Rats

• ..\stem cells\StemCell.wmv

• Transplanted motor neurons reach muscle given the right cues • they form synapses with muscle • These connections are electrically active • Rats recover 40% of hind limb grip strength

Status: Currently in Large Mammal Studies with hES Cells
• Ricin injection for focal weakness
– Focal fulminant weakness with α-MN loss

• Non-federally approved hES cell-derived motor neurons
– Cells produced under FDA-compliant GLP conditions

Initial Clinical Population: Type 1 SMA
• • • • Universally fatal without life support ‘pure’ α-MN loss Little myelin Developmental cues guiding axonal growth still present • Shorter distance

4) Glial Restricted Precursors for Demyelinating Disorders
Embryonic Stem Cells Somatic Stem Cells (Fetal or Adult Tissue)




GRPs Naturally restricted fate means high efficacy and control with fewer side effects


(product growth factors)

(produce myelin)

Results: Robust Myelination by Implanted hGRPs-Oligos
A single injection resulted in widespread myelination of the entire forebrain
1 mm

CC Striatum LV

• Shiverer is a mouse with a mutation that
produces defective MBP, resulting in defective myelin. • Human GRPs mature into oligos that produce normal MBP (green) and myelinate extensive regions of shiverer mouse brain.
• All myelin seen here is produced by human

100 µm


GRPs that matured into oligos (red nuclei).

Data from Goldman Lab, Nature Medicine 2004

Histology of the Inflammatory Demyelinated Lesion

Focal Demyelination
H and E Myelin Stain

Imaging of TM in a Rat

3 days after injection in vivo

GFAP 5 days

Blocked Differentiation of GRPs Post Transplantation

Myelin Repair
Neural Stem Cell Oligodendrocyte  Progenitor Premyelinating Oligodendrocyte Myelinating Oligodendrocyte








MBP Expression after LINGO Treatment of GRP Cells

Rat 424077


Ex Vivo LINGO Enhances GRP Differentiation In Vivo


• Involved in the recruitment of immune cells from the blood to inflammation sites in chronic inflammatory diseases (an antibody against alpha 4 is used for the treatment of MS – trade name Tysabri)

Very Late Antigen 4 (VLA-4)

Initial Indication: Transverse Myelitis
• Single focal lesion in spinal cord – Acute, monophasic and monofocal lesion – Pathology of demyelination and axonal loss – Biomarkers for prognosis (IL-6) – Easy surgery, small target. ?Vascular delivery? – Surrogate markers: SSEPs, DTI, MTw – Straightforward path to determination of clinical efficacy • Remyelination and axon survival • Function recovery – Benefits of orphan indication while establishing proof of concept for larger disease targets

• Planned IND in 2009 (Q Therapeutics)