616 JOURNAL OF COSMETIC SCIENCE
RESULTS
From a preliminary screening of peptides performed on both NHK, NHF, and C. acnes,
we selected the pKTSKS peptide which modulated pre-inflammatory mediators release,
reduced the growth of C. acnes cells and improved extracellular matrix protein synthesis
without any observed cytotoxicity.
EFFECT OF PKTSKS AND ITS COMPOUNDS ON C. ACNES AND S. EPIDERMIDIS GROWTH
Growth of C. acnes with pKTSKS. C. acnes RT-1 cells grew regularly from 21–53 hours in
their cell culture medium with 0.1% DMSO (control). The presence of 0.1% DMSO had
no impact on C. acnes growth. On the other hand, pKTSKS (6–12 ppm) strongly reduced
C. acnes growth (Figure 2) with a dose-dependent effect. When treated with 6 ppm,
9 ppm and 12 ppm of peptide, C. acnes reached an OD
600nm
of 0.5 at respectively 52 hours,
77 hours, and 96 hours. All these values were significant compared to solvent control
(p 0.01). The peptide slows down the growth of this bacterium, limits its ability to reach
its quorum and to potentially acquire a more virulent phenotype.
In a second set of assays, RT-4, and RT-5 strains of C. acnes were tested in the same way
as RT-1. RT-5 and RT-4 strains required 3X and 2X more time respectively than RT-1 to
reach an OD
600nm
of 0.5 (data not shown). Therefore, it appears that, in vitro, the effects of
pKTSKS differ according to the strain used.
The in vitro activity was validated via a clinical trial. Analysis demonstrated that C. acnes
population of the pKTSKS treated zones of the face was reduced by 18.5% versus the zones
treated with the placebo cream (p =0.067), hence confirming the in vitro results.
Growth of C. acnes with KTSKS and/or palmitic acid. To further validate the interest of the
peptide pKTSKS, C. acnes RT-1 cells were put in contact with either the palmitic acid
alone (4 ppm), the “naked” peptide (8 ppm KTSKS), the association of 4 ppm palmitic acid
+8 ppm KTSKS (note that in the latter, palmitic acid and KTSKS do not form a single
INCI Placebo cream Active cream
%%
Water Qsp 100 Qsp 100
Squalane 3.00 3.00
C12-15 Alkyl Benzoate 2.00 2.00
Glycerin 2.00 2.00
Polysorbate 20 1.00 1.00
Phenoxyethanol 0.80 0.80
Caprylyl Glycol 0.50 0.50
Sodium Hydroxide 0.50 0.50
carbomer 0.45 0.45
Acrylates/C10-30
Alkyl Acrylate
Crosspolymer
0.20 0.20
Potassium Sorbate 0.15 0.15
Fragrance 0.10 0.10
pKTSKS 0.00 0.0012
Figure 1. Formulation composition of the placebo and active creams used for the two clinical studies.

617 Pentapeptide to Control Acne
molecule), or the solvent control. Cell growth was evaluated up to 72 hours (n =3 data
not shown). Neither the isolated compounds nor their combination exerts any significant
effect on bacteria growth compared to the solvent control, confirming the unique limiting
the growth of C. acnes activity of the pKTSKS peptide.
Remnant effects of pKTSKS on C. acnes. A study was conducted to evaluate the remnant effect
of the peptide on C. acnes RT-1 cells and to determine whether the inhibition can be rapidly
lifted in its absence. Bacteria were treated with either 9 ppm pKTSKS or 0.1% DMSO as
a control. After 48 hours, bacteria were centrifuged and rinsed to remove residual peptide
or solvent. Cells from both populations were seeded at same concentrations in fresh cell
culture medium (without pKTSKS nor solvent). Cell growth was estimated over time.
Figure 3 shows a clear increase in cell growth for control (OD
600nm
=0.650 after 48 hours)
0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
21h 29h 45h 53h
Time (h)
Effect of pKTSKS on C. acnes growth
Solvent control pKTSKS 6ppm pKTSKS 9ppm pKTSKS 12ppm
Figure 2. Effect of pKTSKS on C. acnes growth kinetic over time (n =3). Turbidity measurements (OD600nm)
on C. acnes (RT-1), to evaluate the effect of pKTSKS on C. acnes growth. Bacteria were treated with pKTSKS
in anaerobic conditions. OD600nm measurements were performed at different time-points (21 h, 29 h, 45 h,
and 53 h).
Figure 3. Evaluation of the remnant effect of pKTSKS on C. acnes growth after 48 h of contact (n =3).
C. acnes (RT-1) was treated with 9 ppm pKTSKS or its control for 48 h. Growth was monitored by OD600nm
measurements before and after pKTSKS wash-out.
OD
600nm