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DNA Repair and Intracellular Communication Daniel B. Yarosh, Ph.D. SVP, Basic Science Research Estee Lauder Co.

DNA Repair and Intracellular Communication

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Page 1: DNA Repair and Intracellular Communication

DNA Repair and Intracellular Communication

Daniel B. Yarosh, Ph.D.SVP, Basic Science Research

Estee Lauder Co.

Page 2: DNA Repair and Intracellular Communication

1953: Birth of Molecular Genetics Watson and Crick

Heredity / Control of Cell FunctionDNA is a sensor for cell damage

Page 3: DNA Repair and Intracellular Communication

UVINDIRECT

Free Radicals

DIRECT

DNA-AGC^T-

DNA-AoGCT-

CellularOxidationProducts

ANTIOXIDANTS

UV DAMAGE TO DNA

Sunscreens

Page 4: DNA Repair and Intracellular Communication

Thymine CytosineCyclobutane pyrimidine dimer

Direct DNA Damage

Page 5: DNA Repair and Intracellular Communication

Can Antioxidants Prevent CPD?

Murray et al. (Pinnell) JAAD 59:418, 2008

15% Vitamin C, 1% Vitamin E and 0.5% Ferulic Acid (CEFer)

Page 6: DNA Repair and Intracellular Communication

Indirect DNA Damage

8-oxo-Guanine

ROS

Page 7: DNA Repair and Intracellular Communication

UV DAMAGE TO DNAType of Damage DNA Repair Enzyme

Direct:Cyclobutane pyrimidine T4 endonuclease V

dimer UV EndonucleasePhotolyase

Indirect:ROS → 8-oxo-Gua OGG1

Page 8: DNA Repair and Intracellular Communication

T4 Endonuclease V

Bacteriophage T4 recombinant protein

Recognition and incision in one polypeptide

Page 9: DNA Repair and Intracellular Communication

Liposome Delivery

DNA repair enzymes

LIPOSOME

Page 10: DNA Repair and Intracellular Communication

DNA Repair in Human Skin

Untreated

Single UVB dose (1,500J/m2)6h treatment with the DNA Repair Lotion (2mg/cm2)

DNA Repair Lotion

Zastrow and Olivier, 2004 ICP, Jeju Korea

Page 11: DNA Repair and Intracellular Communication

DNA Repair in Human Skin

UV

UV+Photosomes

Human skin: UVB

CPD (green)

Photosomes (1h) reduce CPD by 45%

(Stege et al., PNAS 97:1790, 2000)

Page 12: DNA Repair and Intracellular Communication

OGG1 Repair of 8-oxo-Gua

SKH‐1 mice: UV followed by OGG1 liposomes

Wulff et al., Photochem. Photobiol. 84:317, 2008

8oxoGua

Page 13: DNA Repair and Intracellular Communication

Organ Transplant Patientsand Skin Cancer

180,000 living transplant patients

Page 14: DNA Repair and Intracellular Communication

Nickel sulfate:wheal and flareimmune response

UV erythema and suppressionof immune response

Photosomes prevent UV-B erythemaand restore immune response

Stege et al., PNAS 97:1790, 2000

DNA Damage and Immunity

Page 15: DNA Repair and Intracellular Communication

Local Contact Hypersensitivity

FITC

FITCDNA

RepairUV-B

3 days 5 days 1 day

measureearswelling

Page 16: DNA Repair and Intracellular Communication

DNA Repair Prevents Immunosuppression at Remote Sites

FITC +UV +Placebo +T4N55

6

7

8

9

10

CH

S R

espo

nse

Mea

n Ea

r Sw

ellin

g

p<0.05

Kripke et al. Exp Derm 5:173, 1996

Page 17: DNA Repair and Intracellular Communication

Langerhans Cells Flee Skin

Kuchel et al. PPS 4:577, 2005

Dendritic cells in human skin (red)After 4 days of solar simulating light

Treated with Placebo liposomes (Empty)

orT4N5 Liposome Lotion

Page 18: DNA Repair and Intracellular Communication

T-cells in Lymph Node

Phase contrast FITC (antigen) DiI (Liposome marker)

Yarosh et al, JID 103:461, 1994

Page 19: DNA Repair and Intracellular Communication

Cell to Cell Communication

2 MED (UVA,B) 6 hours

anti-IL-10 antibody staining

T4N5 LiposomeLotion

Placebo

Wolf et al. JID 114:149-156, 2000

DNA Damage induces IL-10

DNA repair reduces expression of IL-10

Human Skin

Page 20: DNA Repair and Intracellular Communication

Wound Healing Protease: MMP-1

Direct UV damageto collagen

UV to fibroblast

UV to keratinocytereleases cytokine

MMP-1 release

collagen

Page 21: DNA Repair and Intracellular Communication

Induction of MMP-1 in Skin

Modified from: Fisher et al., New England Journal of Medicine, 337:1419, 1997

Page 22: DNA Repair and Intracellular Communication

Cell to Cell Communication

UVB 500 J/m2

Keratinocytes+ DNA Repair

24h media

Fibroblasts

48 h media

MMP-1Collagen

Page 23: DNA Repair and Intracellular Communication

MMP-1 Gene Expression

Normal human fibroblasts treated with media from NHEKNHEK: 500 J/m2 UV-B + T4N5 Liposomes (1 µg/ml) for 1 hr post-UV.MMP-1 mRNA expression measured by Real-Time PCR (RT-PCR)

1

1.5

2

2.5

3

3.5

4

0 10 20 30

UV-BUV-B+T4N5None

Hours Media Exposure

Rel

ativ

e In

crea

se M

MP-

1Human Fibroblasts Receiving NHEK Media

Page 24: DNA Repair and Intracellular Communication

50

100

MM

P-1

/GA

PD

H (%

Con

trol)

Significance: * p<0.05

0.0 0.1 1.0 0.1 1.0

150

200

(%)

- UVB

+ UVB

0.0 0.1 1.0

MMP-1

GAPDH

0.1 1.0Blank

LiposomeUV

Endonuclease(liposomal M. luteus

extract)

(%)

- UVB

+ UVB BlankLiposome

UVEndonuclease

(liposomal M. luteus extract)

UV endonuclease significantly suppressed the Kc-induced MMP-1 expression in Fb

Page 25: DNA Repair and Intracellular Communication

OGG1 Inhibition of MMP-1MMP1 in Fb

48h after UV-Kc media

0.0 1.0 2.50.00

0.25

0.50

0.75

1.00

1.25 Empty LiposomesOGG1 Liposomes

Liposomes (µg/ml)

MM

P-1

(ng/

ml)

Page 26: DNA Repair and Intracellular Communication

DNA Damage and Cell Communication

Kinase CascadeSelective transcript activation

CYTOKINES:IMMUNE SUPPRESSIONWOUND HEALINGVIRAL ACTIVATION

DNA PK (PI3K family)ATM, ATR, mTOR

RNA-Pol

Page 27: DNA Repair and Intracellular Communication

The New Synthesis• DNA damage injures a cell• Injured cells communicate

– Cytokines release– Immune suppression– Wound healing response– Viral activation

• Undamaged cells respond to DNA damage• Extracellular signals contribute to skin

cancer

Page 28: DNA Repair and Intracellular Communication

Acknowledgements

AGI Dermatics Cosmos ResearchKen Smiles, PhD Hitoshi Masaki, PhDMatt Canning Yasunobu OchiaiKelly Dong Kuri ObayashiStephanie Nay Yuri OkanoNoah DamaghiStephaney Picart THE XP STUDY GROUPNelli Markova, PhD Margaret Kripke labNeveena Juruvoska, PhD Jean Krutmann lab