INDUSTRY APPLICATIONS FOR 3D BIO- PRINTING - CONSUMER PRODUCTS

INDUSTRY APPLICATIONS FOR 3D BIO-PRINTING - CONSUMER PRODUCTS

ANDREW SCOTT,SAFETY & ENVIRONMENTAL ASSURANCE CENTRE

www.TT21C.org

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WHAT I’LL COVER TODAY

• About Unilever

• New technology and lead identification

• Assuring consumer safety without new animal tests

• A perspective on 3D-Bioprinting and where the bioprinting community can help

• Case study – human relevant skin models

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“MAKE CLEANLINESS COMMONPLACE” 3

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EUROPE

● €13.2 BILLION TURNOVER

● 0.2% UNDERLYING VOLUME GROWTH

● 27% OF GROUP TURNOVER

ASIA, AFRICA, CENTRAL & EASTERN EUROPE

● €19.7 BILLION TURNOVER

● 2.0% UNDERLYING VOLUME GROWTH

● 41% OF GROUP TURNOVER

THE AMERICAS

● €15.5 BILLION TURNOVER

● 0.7% UNDERLYING VOLUME GROWTH

● 32% OF GROUP TURNOVER

2014 TURNOVER = €48.4 BN

UNILEVER ISA GLOBAL COMPANY

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WE MAKE MANY OF THE WORLD’S FAVOURITE BRANDS

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INGREDIENT DISCOVERY FOR PERSONAL CARE

Mechanistic Insight

Target ID & Validation

ScreeningIngredient

testing in vitro

Validation in human studies

Representative in vitro models needed

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MICROCIRCULATION AND THE SKIN

• Skin is highly vascularised:

• Superficial plexus - below the skin

surface

• Sub papillary plexus - at the dermal-

subcutaneous junction.

• A complex molecular dialogue exists

between vascular derived cells and

mesenchymal skin cells impacting on skin

function.

• 3D bio-printing will help the investigation of

complex signalling pathways and cross talk

mechanisms apparent in vivo.

Li et al 2006

65 years old,

hand

Hembold et al 2006

old

young

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THE HAIR FOLLICLE – A COMPLEX MINI-ORGAN

(Schneider et al., 2009 Current Biology)

The hair follicle undergoes regular

cycles regeneration throughout life.

Hair follicle is a complex mini-organ

of the skin.

The Philpott hair follicle organ

culture model relies on availability

of waste donor-consented scalp

tissue following surgery.

Currently the above or variations of

the Philpott model is the only whole

hair follicle in vitro model available

despite attempts to grow hair

follicles in co-culture systems.

3D bio-printing combined with iPSC

technology may provide a solution

for overcoming limited supply of

tissue and recapitulation of hair

follicles that mimic those in vivo

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DEVELOPMENT OF SCALP SKIN MODELS

Scalp Skin

Development of scalp skin modelsReconstructed Models

Epiderm – skin equivalent Scalp epidermal model

Measure barrier

integrity (TEER)+ ‘SEBUM’

+ MICROBES

(lab grown or ex-vivo)

Microbes removed

‘microbially challenged’epidermal model

T-cell priming

Measure protein and mRNA biomarkers of

immune response

Measure epidermal inflammatory

response (transcript)

Measure epidermal inflammatory

response (protein)Epiderm embeddedwith Dendritic cells

Measure protein and mRNA biomarkers of

immune response

+ immune components

epidermal

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Can we use x% of ingredient yin product z?

CAN WE USE A NEW INGREDIENT OR SAFELY?

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PERSONAL CARE CONSUMER PRODUCTS

• Chemical ingredients not generally intended to be pharmacologically active (compare Pharmaceutical Co.)

• Low bioavailability and often topical exposure

• Open regulatory environment

Making an exposure-led safety decision based on confidence that the safe level is within or below the adaptive homeostasis response, captured by appropriate in vitro systems and complemented with network computational models

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NEW APPROACHES TO RISK ASSESSMENT WITHOUT ANIMALS

• Focus on non-animal approaches for consumer safety risk assessment

• Data required for safety decision should be driver

• Dose response information is essential

• Understanding the underpinning human biology

• We are not looking for a way to do the animal test without the animal

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ASSURING CONSUMER SAFETY

Start with exposure rather than toxicity

‘ ……. Human safety depends on exposure and toxicity. Indeed, the 2012 European Commission report on addressing the NewChallenges for Risk Assessment states, “ A paradigm shift is likely from a hazard-driven process to one that is exposure-driven” ….’

Pastoor et al (2014), Crit Rev Toxicol, 44(S3): 1–5

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EXPOSURE – DRIVEN SAFETY ASSESSMENT

• Improving exposure estimates• Building on the current knowledge of consumer habits & practices• Focus on measurement and quantitative exposure science

Dermal kinetics

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WILL THESE EXPOSURES CAUSE ADVERSE OUTCOMES IN CONSUMERS?

BiologicInputs

NormalBiologicFunction

Adaptive StressResponses

Early CellularChanges

Exposure

Tissue Dose

Biologic Interaction

Perturbation

Low DoseHigher Dose

Morbidityand

Mortality

Cell Injury

Higher yet

(From Andersen & Krewski, 2009, Tox Sci, 107, 324)

Perturbations of Toxicity Pathways

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AN EXAMPLE PATHWAY

• Skin allergy

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Adapted from ‘Adverse Outcome Pathway (AOP) for Skin Sensitisation’, OECD

OPPORTUNITIES FOR BIO-PRINTING MAPPED TO SKIN SENSITISATION AOP

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OPPORTUNITIES FOR BIO-PRINTING MAPPED TO SKIN SENSITISATION AOP

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Replace ex vivo human skin for measurement of skin bioavailability (binding, metabolism,

penetration)

OPPORTUNITIES FOR BIO-PRINTING MAPPED TO SKIN SENSITISATION AOP

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Replace ex vivo human skin for measurement of skin bioavailability (binding, metabolism,

penetration)

Characterise impact of chemical/drug/product exposure on skin micro-environment

(e.g. induction of inflammatory response, interplay between skin microbiome and local immune cells)

OPPORTUNITIES FOR BIO-PRINTING MAPPED TO SKIN SENSITISATION AOP

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Replace ex vivo human skin for measurement of skin bioavailability (binding, metabolism,

penetration)

Characterise impact of chemical/drug/product exposure on skin micro-environment

(e.g. induction of inflammatory response, interplay between skin microbiome and local immune cells)

Develop skin models recreating different skin

disease pathologies to inform ‘sensitive’ sub-population

risk assessments

OPPORTUNITIES FOR BIO-PRINTING MAPPED TO SKIN SENSITISATION AOP

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TKTD MODEL OF HUMAN SKIN SENSITISATION: SCHEMATIC DIAGRAM

PANEL A = TOXICOKINETIC MODEL: DNCB kineticsin skin due to (1) diffusion and partitioning intothe stratum corneum and skin; (2) sensitiserclearance by dermal capillaries; (3) covalentmodification of protein nucleophiles by hapten.

PANEL B = TOXICODYNAMIC MODEL: biological response due to (4)proteasome processing of protein nucleophiles to form small peptides andtransport to the endoplasmic reticulum (ER); (5) binding of peptides andhapten-peptide complexes to MHCI and transport to plasma membrane; (6)binding of pMHC and hapten-pMHC to CD8+ T cell receptors and (7)activation and expansion of naïve specific CD8+ T cells.

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SUMMARY – OPPORTUNITIES FOR 3D BIO-PRINTED (SKIN) MODELS

• Improved speed and accuracy for identification of lead actives

• More predictive tissue models for investigate research and target identification

• More biologically relevant models for assessing effects on pathways

• Potential replacement for limited human tissue supply

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Unilever’s Safety & Environmental Assurance Centre (SEAC): helping to shape innovations that are safe for our consumers and workers, and better for the environment. SEAC was created 25 years ago by bringing together all relevant scientific expertise across Unilever in a single group.

1990 – 2015

AOP-BASED RISK ASSESSMENTSEXAMPLE: SKIN ALLERGY

Induction of skin allergy is a multi-stage process driven by toxicity pathways

- mechanistic understanding is captured in Adverse Outcome Pathway (AOP)

- non-animal test methods have been developed; each aims to predict impact of a chemical on one key event

- how can we make risk assessment decisions by integrating this scientific evidence?

Modified from ‘Adverse Outcome Pathway (AOP) for Skin Sensitisation’, OECD report

1. Skin Penetration

2. Electrophilicsubstance:

directly or via auto-oxidation or metabolism

3-4. Haptenation: covalent

modification of epidermal proteins

5-6. Activation of epidermal

keratinocytes & Dendritic cells

7. Presentation of haptenated protein by

Dendritic cell resulting in activation &

proliferation of specific T cells

8-11. Allergic Contact Dermatitis: Epidermal inflammation following

re-exposure to substance due to T cell-mediated cell

death

Key Event 1 Key Event 2 + 3 Key Event 4 Adverse Outcome