Will stem cells be a key resource in the future for regulating inflammation

and treatment of IBD?

Thaddeus Stappenbeck MD, PhD

Will stem cells graduate and move down the hall to the ‘big’ room?

Thaddeus Stappenbeck MD, PhD

Will stem cells graduate and move down the hall to the ‘big’ room?

1) Cellular therapy – addition of stem cells or their progeny

2) Use of cells for patient stratification

Will stem cells graduate and move down the hall to the ‘big’ room?

1) Cellular therapy – addition of stem cells or their progeny – HSCs, MSCs, epithelial stem cells, iPSCs

2) Use of cells for patient stratification

Direct incorporation of epithelial stem cells

Mesenchymal stem cells

Direct incorporation of hematopoietic stem cells

iPS cells

Will stem cells graduate and move down the hall to the ‘big’ room?

1) Cellular therapy – addition of stem cells or their progeny

2) Use of cells for patient stratification – epithelial stem cells from patients

Dark Ages (1950s-2008)Caco-2 HT-29 SW480

Advantages:Easy to cultureAbility to rapidly expand>21K papers in Pubmed

Disadvantages:AneuploidLimited ability to differentiate

Dark AgesCaco-2 HT-29 SW480

Advantages:Easy to cultureAbility to rapidly expand>21K papers in Pubmed

Disadvantages:AneuploidLimited ability to differentiate

Renaissance (2008-current)

Renaissance (2008-2013)Critical factors for in vitro crypt culture primary cells Wnt ligands (e.g. Wnt3a) R-spondin (e.g. Rspo1) Noggin (BMP inhibitor)

(Sato, T. et al. Nature 459, 262-266, 2009)

Organoids aka ‘miniguts’

Expanded Platforms• 3D - Organoid (stem cells and

multilineage)• 3D - Spheroid (stem cell enriched)• 2D – sheet of differentiated• Co-culture pathogens• Co-culture anaerobic commensals• Co-culture with mesenchymal cells

Industrial Revolution Individualized Medicine

Oootani et al., Nat Med 15, 701-6 (2009)Miyoshi et al., Nat Prot (2013)Wang et al., Gastroenterology (2013)Yin et al., Nat Methods (2013)

The Goal of Understanding Inter-Individual Variation: Personalized Medicine

Patient 1 Drug A = best response

Drug B = best response

Drug A & B = best response

Patient 2

Patient 3

Exhibit similar

symptoms or have same diagnosis

Development of in vitro Systems and Assays Is Critical for Individualized Medicine Approaches

Patient 1 Drug A = best

Drug B = best

Drug A & B = best

Patient 2

Patient 3

Collect sample

Cell-based assay

Inter-individual Variation in Intestinal Epithelial Cells

Epithelial Cell Function

Genetics

Epigenetics EnvironmentMicrobiome

Other host cells

Infection susceptibility

DrugresponseDisease

susceptibility

ExposomeGenome

ExposomeGenome

Hypothesis: Functional variation will be present in primary intestinal epithelial cell

lines from different individuals

Question: What is driving the inter-individual

variation observed for particular readouts?

Variations in epithelial cell functional readouts

Establishment of Human Intestinal Epithelial Cell Cultures

Proliferative Stem cells

Established primary cell line

1) Collagenase treatment

2) Filtration

Tissue culture plate well

Matrigel

Epithelial spheroid

lumen

Bar, 100 µm

Bar, 100 µm

Human Intestinal Epithelial Cell Cultures Can be Rapidly Expanded

50% L-WRN CMConditioned

medium containing stem

cell growth factors

lumen

Spheroids are enriched for stem cells in

50% L-WRN CM

Intestinal stem cell marker

Same spheroid imaged dailyBar, 100 µm

Generation of a Biobank of Human Intestinal Epithelial Cell Lines

• 153 cell lines generated to date• Includes 45 lines from other

IBD centers (Cedars-Sinai, Harvard)

• 105 individuals with different genetic landscapes

• No disease and disease

Human Epithelial Cells Form a Differentiated, Polarized Monolayer on Transwell Membranes

Monolayers contain the major differentiated cell types of the

intestinal epithelium

• Lines can be established within 2-3 weeks• Epithelial cells can be rapidly expanded as

stem cells or differentiated• Epithelial cells can form a polarized monolayer

covered by a mucus layer• Allows for repeated experiments

Stappenbeck labKelli VandussenHiroyuki MiyoshiClara MoonStacy RyuNaomi SonnekChristina HickeyHaerin JungGerard KaikoTa-Chiang LiuRC LaiChristine LuoNick ManieriNikki MalvinGreg RippbergerAshley SteedLulu SunMarta Wegorzewska

Matt CiorbaJeff Marinshaw

Phil TarrNurmohammad

Shaikh

Dave SibleyGeorgia Wilke

Skip VirginMegan BaldridgeDebbie Lenschow

Helen and David Piwnica-Worms

DDRCCRodney NewberryDarren Nix

Cedars-SinaiDermot McGovern

Harvard/Broad InstituteRamnik Xavier

GTACRich HeadMike HeinzChris Sawyer

Acknowledgements