Stem Cells and the Future of Medicine

Martin PeraEli and Edythe Broad

Center for Regenerative MedicineAnd Stem Cell Research at USC

What is regenerative medicine?

An emerging field of therapeutics that has as its goal the restoration of normal function in conditions characterised by cell loss, caused by disease or injury. Restoration may involve the administration of cells to replace damaged tissue, alone or in combination with synthetic scaffolds, or the administration of pharmaceuticals that help drive the patient’s tissue to repair itself.

What is a stem cell?

• A primitive cell with two key properties:• Self renewal, or the ability to divide many times to produce

more stem cells• The ability to undergo differentiation or specialisation to give

rise to mature functional cells• Stem cells have the potential to replace dead or damaged

cells in diseased tissue

Tissue Stem Cells

Clinical uses of tissue stem cells

• Bone marrow and cord blood-hematopoietic disorders, leukemias

• Mesenchymal stem cells-cartilage repair• Neural stem cells-early phase trials

Limitations with tissue stem cells

• Rare-minority population in most tissues ie less that 1/1000

• Usually have a limited repertoire-can only give rise to a few types of differentiated cell

• Not well characterised in many tissues• Difficult to propagate and expand in numbers

outside of the body

Nov 98- human embryonic stem cells discovered

Human embryonic stem cells

• Derived from spare embryos before specialised tissue of the body begin to form

• Can multiply indefinitely in laboratory cultures

• Retain the ability of embryonic cells to turn into any type of tissue

Early stages of human development

Teratoma formed by human ES cells shows differentiation into a wide variety of cell types

hESC differentiation

ES cell differentiation follows the road map

of embryogenesis

Partial list of cell types derived from human ES cells in vitro

• Nerve, astrocyte, oligodendrocyte• Hematopoietic stem cells• Insulin producing cells• Cardiomyocytes• Hepatocytes• Endothelial cells• Bone forming cells• Motor neurons• Trophoblast and yolk sac cells

Many species of mammal have now been cloned.Can cloning technology be used to surmount immunologicalbarriers to stem cell transplantation?

Somatic cell nuclear transfer and reprogramming

Somatic cell nuclear transfer

Induced pluripotent stem cellsMatching cells for patients

Skin cells taken from adult tissue are grown in a dish2-4 genes found in embryonic stem cells are introducedAdult cells are converted to pluripotent stem cellsThese stem cells provide a match for the patient

Induced Pluripotent Stem Cells: The potential

• Creation of large banks of stem cell lines of desired HLA haplotypes for tissue matching

• Development of in vitro models of diseases with complex genetic susceptibility

• Partial reprogramming to heal tissues: exocrine pancreas to endocrine pancreas

Induced Pluripotent Stem Cells: Challenges

• Can we make these cells without genetic modification?

• Are they really equivalent to human embryonic stem cells

• Embryonic stem cell research is still needed as is research in somatic cell nuclear transfer

Stem cell research will revolutionise medicine

• Powerful new tools to study human biology in health and disease

• Normal human cells to study in the laboratory-use to develop new drugs. Alternative to animal models or direct tests on human guinea pigs.

• Cells for replacement therapy in devastating conditions involving cell loss or injury

• New understand of how the body’s natural healing process, how and why it fails, and how to improve healing

Stem Cells to Study Disease

Marchetto et al. Cell Stem Cell 3: 649, 2008Amyelotrophic lateral sclerosis

Stem Cells to Study CNS Development

Cortical structures in vitro from human ES cellsEiraku et al. Cell Stem Cell 3: 519, 2008

The Eli and Edythe Broad CenterFor Regenerative Medicine and

Stem Cell Research

Broad CIRM Center Opening29 October 2010

The Center

• Established April 2006• Built on strengths in developmental biology,

clinical research, stem cell biology at Keck School and CHLA

• $50 million dollar commitment by USC to program development

• Now 12 PIs, over 100 staff, four core laboratories. Four administrative staff

Center Research StrategyDiscovery Technology Treatment

Basic discoveries in stem cell biology

Platform technologies:Large scale productionDrug discovery, functionalgenomics

Patient treatmentAnd clinical trials

Engineering, Biotech,pharma

KSOM clinical groups

Center Discovery Research ThemesStem Cell Biology

• Embryonic stem cell growth and differentiation; reprogramming adult cells

• Biology of tissue regeneration and repair; how stem cells are controlled in the body

• How cells are shaped to form organs

Embryonic stem cells from rat

Chimeric rat pups madefrom embryonic stemcells. Chimeras are blackand white.

An important new tool for basic research and drug discovery

• Workers have tried for 20 years to make rat embryonic stem cells• Rats are widely used in physiology and pharmacology and drug discovery• Until now there have been no tools to make specific modifications in the

rat genome to create disease models, like we can in mouse (Nobel prize 2007)

• Ying used his new discoveries about embryonic stem cell growth regulation (ES cell self renewal as a default pathway) to make rat ES cells for the first time

Science Magazine Top Ten Breakthroughs of 2010

Gene knockout rat technology developed by Dr. Qilong Yingnamed one of Top Ten Breakthroughs of 2010 by Science magazine

Understanding tissue repair and regeneration

Lower vertebrates can regenerate limbs, hearts and kidneys.How does this work?What stops this happeningin mammals?How do tissue stem cells functionin repair?

Skeletal regenerationIn Mammals

Dr. Francesca Mariani

Stem Cell Transplantation Biology

Dr. Gregor Adams and colleagues have identified a drug that promotes engraftment of blood stem cells in transplant recipients.The findings may lead to more effective treatment of blood disorders and cancers.

Blindness

Macular degeneration is a major cause of blindness in the aging

population

Retinal pigment epithelium and macular degeneration, a major cause of blindness

2000- hESC can form neural tissue in vitro. The eye formsas an outgrowth of the embryonic brain

Groppe et al. Nature420: 636, 2002

Treatment with the embryonic head inducer noggininduces differentiation of human ES cells into primitive neural tissueNestin and Sox-2, markers of early neurogenesis

2004-directed neural differentiation

Conservation of developmental mechanisms

Retinal pigment epithelium from human neural progenitorsDoheny Eye Institute and Center Collaboration

The road to the clinic:ES cells for eye disease

CIRM Macular Degeneration Disease Team: The California Project to Cure Blindness

• USC Doheny Eye Institute (Mark Humayun, PI; David Hinton Co-PI; Vas Sadda, Biju Thomas, Martin Pera)

• UCSB Macular Degeneration and Stem Cell Centers (Dennis Clegg, Co-PI; Lincoln Johnson)

• UCL London Project to Cure Blindness (Pete Coffey, Partner PI funded by MRC)

• Caltech Biology and Chemistry (Scott Fraser, Bob Grubbs, Yu-Chong Tai)

• City of Hope Center for Biomedicine and Genetics GMP Facility (Larry Couture)

Chemical genomics and stem cells

• Stem cells can be used in high throughput screens to discover new small molecules that modulate tissue regeneration or repair

• Important tool to understand stem cell control pathways-but also leads for drug development

Dr. Kahn has focused on WNT signaling, a key pathway in development and cancer

The Wingless mutation affects wing development in the fruit fly embryo.The Wnt gene, discovered as a virus integration site for mouse breast cancer induction, is involved in many cancers.Like many developmental pathways, Wnt is evolutionarily conserved and widely deployed in stem cell regulation in many tissues.

USC Center for Molecular Pathways and Drug Discovery

• A joint venture between the Broad Center at USC and the USC Norris Cancer Center

• Directed by Professor Michael Kahn (Broad) and Professor Heinz Lenz (Norris Cancer Center)

• Mission: discover new chemicals that modulate critical signaling pathways in stem cells and cancer and develop new therapeutics

PBS / PBS

PBS / Nilo (100mg)

ICG-001 / Nilo (100mg)

1.0E+05

1.0E+06

1.0E+07

1.0E+08

1.0E+09

1.0E+10

10 20 28 35 42 49 56 63 77 91

Days After BMT

Flux

(pho

tons

/sec

/cm

2 /sr)

CBP/Catenin Antagonists Effectively Eliminate Cancer Stem Cells When Used in Combination Chemotherapy

0 100 200 300 4000

25

50

75

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125

PBS / PBSPBS / NiloICG / Nilo

Days After BMT

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Mice are cancerfree

Cancer Therapy Program at NCI Considers the Development of PRI-724 its Highest Priority

“PRI-724 is a first in class and first in human agent. PRI-724 is a novel cancer stem cell targeting agent…” There are no other Wnt signaling pathway inhibitors targeting transcription.

Phase I clinical trial 7 day continuous infusion of C82 (primary endpoint MTD/biological activity, secondary endpoint proof of principle Survivin

expression in tumor and CTC)-later this year

No toxicity to normal tissues in dogs at 200X IC50 (the dose that kills

50% of tumor cells)Ph

Zinc finger nuclease-based stem cell therapy for AIDS

- $14 million CIRM Disease Team award

Paula CannonAssociate Professor, Microbiology & ImmunologyUSC Keck School of Medicine

Dave DiGiusto John Rossi John Zaia(PI)

City of Hope AIDS lymphoma group

The AIDS virus must enter cells to infect them.HIV-1 enters cells by binding to CD4 and a co-receptor, CCR5

CD4 and CCR5 are proteins on the surface of cells that the HIV-1 targets.-About 1% of people have 2 copies of a mutant version of CCR5 gene called CCR5D32, and theyare consequently extremely resistant to HIV-1.-The virus cannot enter their cells.

B cells

Erythrocytes platelets

CLP

CMP

ProB

GMP

MEP

T cells

Macrophages

HSC

HIV

Can we make AIDS resistant cells byblocking CCR5 expression in

hematopoietic stem cells and their progeny?

Bone marrow stem cellsare the source of T Cells and macrophagesthat the AIDS virusinfects

DNA binding domain

They cut the CCR5 gene

The cell repairs this break, but in a way that also destroys the CCR5 gene

DNA cleaving domain

Zinc finger nucleases to disrupt the CCR5 gene

ZFNs bind to the CCR5 gene

Zinc fingers targetspecific DNA sequences. The attachednucleases chop the DNA.

Day -1 Day 0

Transplant into special ‘NSG’ mice

Months 2 - 4

Human hematopoietic

stem cells

Human T cells in mouse blood HIV Infection

Pre-clinical testing

HIV-1

Treat with CCR5 ZFNs

ZFN-treated HSC generate human cells in the mice that are HIV-resistant

thymus

ZFNCtrl. Ctrl.

HIV-1 infectedCD

8

CD4

CD4

CD14

CD3

CD8

CD45

SSC

spleen

lung

Gut mucosa

Ctrl. HSC

ZFN HSC

0 2 4 6 8

CD4

T ce

lls

weeks post-HIV infection

Human cells are normal in ZFN-treated mice

Human cells killed by HIV-1 infectionin untreated mice

Holt et al. Nat Biotechnol. 2010 Aug;28(8):839-47

Other USC Bioscience Interdisciplinary Initiatives

• Neuroscience• Biomedical Nanoscience• Clinical and Translational Sciences InstituteFunded by NIH $60 million to facilitate

translational research