Transcript SP1-Growth factor - in

Concept Cosmeceuticals
SP1- Growth Factor
: Skin
: permeable
: Want
Introduction of Cell
Transducible
Growth Factor for Skin
Renewal
SP1-EGF
& SP1-FGF1
Real Effective Cosmeceuticals
begin at
Aim of the Study
Development of Cell Transducible Growth Factors
1) Skin Permeable Epidermal Growth Factor (SP1-EGF)
2) Skin Permeable Acidic Fibroblast Growth Factor(SP1-FGF1)
3) Skin Permeable Superoxide Dismutase(SP1-SOD)
4) DDS technology of skin permeable therapeutic humanized
antibodies
Skin Care
Skin Regeneration
Before
After
Treatment of Burn Injury
Before
After
We aim to develop cosmeceuticals which cover from skin care, skin
regeneration to burn injury through the development of cell
transducible growth factors with PTD technology.
The Superiority and Creativity of PTD Technology
- Increase skin regeneration by improving skin permeability
No treated EGF
Product
Liposome linked EGF
Product
PTD Conjugated EGF
Product
Comparison with the third party product
Cosmetics containing even very small amounts of EGF, FGF1 is quite
expensive in the current market. Notwithstanding the high price,
protein itself is very difficult to permeate through the skin. So
liposome formulated one is widely used. However liposomal
formulation also has its limitation.
Thus we improved those limitation with PTD(Protein Transduction
Domain) Technology.
As shown in the figure, PTD conjugated product (protein) showed
better permeability compared to existing liposome. (Green
fluorescence represent protein permeated deeply into the skin)
EGF (Epidermal Growth Factor)
1. Structural Characteristics
- Consists of 53 amino acids
- Molecular weight 6.2 kDa
- 3 S-S bond(Disulfide bond)
(6-20, 14-31, 33-42)
2. Function
- Anti-aging & Improve Pre-made winkles by skin regeneration
- Improve skin tone
- Soft skin, Brightening and Rejuvenation
- Remove skin scar by forming new epidermis
FGF1 (Acidic Fibroblast Growth Factor)
1. Structural Characteristics
- Consists of 155 amino acids
- Molecular weight 17.3 kDa
2. Function
- Prevent winkles, aging, dry skin by synthesizing Collagen,
Elastin and Glucosaminoglucan under the skin.
- Promote skin elasticity by acting on the cells in the dermis.
Expression of SP1-EGF & SP1-FGF1
(A) SP1-EGF
(B) SP1-FGF1
SDS-PAGE
M
1
2
1
2
Western blot
SDS-PAGE
Western blot
M
1
2
1
2
SP1-EGF and SP1-FGF1 were analyzed by 15% SDS-PAGE and
Western bloting using specific antibodies for EGF and FGF1.
M : Size marker
1
: Before the expression of SP1-EGF & SP1-FGF1
2
: After the expression of SP1-EGF & SP1-FGF1
# SP1-EGF molecular masses : 12kDa, # SP1-FGF1 molecular masses : 19kDa
Activity of SP1-EGF & SP1-FGF1
(A) SP1-EGF
Phospho EGF receptor
(Tyr1068)
0
10
100ng/ml
Used Cell Line
A431
SP1-EGF
(B) SP1-FGF1
Phospho FRS2-alpha
(Tyr196)
0
10
100ng/ml
Used Cell Line
NIH3T3
SP1-FGF1
The data above are bioactivity test by specific growth factor
marker of SP1-EGF and SP1-FGF1
The data above shows the dose dependent activity of SP1-EGF,
SP1-FGF1 by showing phosphorlation status of EGF & FGF1
receptor as low as 10ng/ml concentration.
Effects of SP1-EGF & Sp1-FGF1 in cell proliferation
A
B
C
D
Used Cell Line : NIH3T3
Treated Concentration : each 100ng/ml
A : Control
B : SP1-EGF
C : SP1-FGF1
D : SP1-EGF + SP1-FGF1 Mixture
The data above shows the proliferative activity of SP1-EGF, SP1FGF1 in cells(NIH3T3). Compared to control A, all of the growth
factors showed proliferative effects.
Effects of SP1-EGF & Sp1-FGF1 in cell proliferation
SP1-EGF Proliferation
SP1-FGF1 Proliferation
SP1-EGF Conc. (ng/ml)
SP1-FGF1 Conc. (ng/ml)
Combination
SP1-EGF + SP1- FGF1 Conc. (ng/ml)
The figure above is graph of the proliferative activity of SP1-EGF
& SP1-FGF1 in cells(NIH3T3).
There
is
dose
dependent
concentration of 0.1ng/ml.
proliferative
effects
from
the
Ratio % vs GAPDH
Effects of SP1-EGF & Sp1-FGF1 in Collagen Synthesis
Used Cell Line : CCD-986SK
Treated Concentration : each 100ng/ml
Combination : SP1-EGF + SP1-FGF1 Mixture
The graph above represent the ability of SP1-EGF and SP1-FGF1 in
collagen synthesis. Collagen is distributed and acted in the dermis
layer of the skin. SP1-FGF1 showed best collagen synthesis when
measured its messenger RNA levels.
Degrees of Skin permeability
Epidermis
Epidermis
A : Control, B : Normal EGF, C : SP1-EGF, D : Normal FGF1, E : SP1-FGF1
Used skin : Porcine skin
Treated Concentration : each 100ng/ml
Treated Time : 3hr
The results above represent microscopically observed cross
section of skin after fluorescent labeled SP1-EGF, SP1-FGF1
treatment to see the degree of tissue permeability .
In contrast to other EGF and FGF1, SP1-EGF and SP1-FGF1 showed
superior permeability compared to others.
SP1-EGF
(Skin Permeable - Epidermal Growth Factor)
Characteristics of SP1-EGF
PTD & EGF Fusion Protein
The PTD fusion protein with EGF makes skin permeability better than with
protein itself so maximize its effects by deep penetration into the cells.
High Purity and High Activity
With purity more than 95% , suitable as a cosmetic
Ingredient and more active than normal EGF.
Safety and Stability
Free from various microbial contamination and allergen. Suitable for the
specification. Through the toxicity test, safety is assured. It is proved to be
stable up to 12 months through long term stability study.
Trade Name
SP1-EGF
(INCI name : processing)
Source
E. coli
Purity
>95% (HPLC)
Amino Acid
99
Molecular Weight
11.9 kDa
pH value
8.0
Activity unit
>5 X 105 IU/mg
Formulation
Lyophilized or Solution stock
Concentration
10, 50, 100 ppm
SP1-FGF1
(Skin Permeable – Acidic Fibroblast Growth Factor)
Characteristics of SP1-FGF1
PTD & FGF1 Fusion Protein
The PTD fusion protein with FGF1 makes skin permeability better than with
protein itself so maximize its effects by deep penetration into the cells.
High Purity and High Activity
With purity more than 95% , suitable as a cosmetic
Ingredient and more active than normal FGF1.
Safety and Stability
Free from various microbial contamination and allergen. Suitable for the
specification. Through the toxicity test, safety is assured. It is proved to be
stable up to 12 months through long term stability study.
Trade Name
SP1-FGF1
(INCI name : processing)
Source
E. coli
Purity
>95% (HPLC)
Amino Acid
171
Molecular Weight
19.9 kDa
pH value
8.0
Activity unit
>1 X 106 IU/mg
Formulation
Lyophilized or Solution stock
Concentration
10, 50, 100 ppm
Purity of SP1-EGF
SDS-PAGE
Western blot
HPLC Analysis of SP1-EGF
SP1-EGF
98.2%
SP1-EGF
Marker
Purity of SP1-FGF1
SDS-PAGE Western blot
HPLC Analysis of SP1-FGF1
SP1-FGF1
96.4%
SP1-FGF1
Marker
Stability of SP1-EGF
SP1-EGF long term storage stability
(12 months)
SP1-FGF1 Solution Product
SP1-FGF1 Lyophilized Product
1
2
3
4
1
2
3
4
Phospho-EGF
receptor
(Tyr1068)
1. No treatmetnt
2. Normal EGF
3. Store at -60℃
4. Store at 4℃
# Treat Conc. : 100ng/ml
All stable up to 12 months
Continuous up date
Stability of SP1-FGF1
SP1-FGF1 long term storage stability
(12 months)
SP1-FGF1 Solution Product
SP1-FGF1 Lyophilized Product
1
2
3
4
1
2
3
Phospho
FRS2-alpha
(Tyr196)
1. No treatment
2. Normal FGF1
3. Store at -60℃
4. Store at 4℃
# Treat Conc. : 100ng/ml
All stable up to 12 months
Continuous up data
4
Prototype of SP1-EGF containing cosmetics
Phospho-EGF
receptor
Used Cell Line : A431
M : Size ladder
A : Growth Factor Non Treated Group
B : 100ng/ml of Normal EGF Treated Group
0.1 ~ 100ng/ml : Diluted SP1-EGF Containing Cosmetics
This proves that SP1- EGF contains proper function which can be used
as a cosmetics.
Prototype of SP1-FGF1 containing cosmetics
Phospho-FGF1
receptor
Used Cell Line : NIH3T3
M : Size ladder
A : Growth Factor Non Treated Group
B : 100ng/ml of Normal FGF1 Treated Group
0.1 ~ 100ng/ml : Diluted SP1-FGF1 Containing Cosmetics
This proves that SP1- FGF1 contains proper function which can be used
as a cosmetics.
Clinical evidence of effect for skin renewal
Day 0
Day 28
Dermal Density TEST
Test Sample
Cosmetics containing SP1-EGF and SP1-FGF1
Contents
SP1-EGF : 10ppm / SP1-FGF1 : 10ppm
Subjects
Information
Wrinkle degree who SOP standards than Class 4,
36 to 48-year-old(average 43.3±3.1) female subjects 21
Test Period
8 weeks
Test Method
Crow’s feet using a three-dimensional image analysis, dermis
density, moisture content measurements facial sag,
viscoelastic, and evaluate the effectiveness by improving the
product through survey evaluation by subjects
Real Effective Cosmeceuticals
begin at
MANUFACTURER / SUPPLIER
BIOCELTRAN. INC.
RM# 305, BIO 2BD, Hupyong-Dong, Chuncheon, Gangwon-Do, South Korea
Tel : +82-33-258-6780 Fax : +82-33-258-6781
www.bioceltran.com([email protected])
Print version 20120917