Use of ide as a biomarker for a scalp condition

Abstract

The subject matter of the present invention is in particular the use of an amino acid sequence of IDE, or of an analogue or fragment thereof, or of at least one nucleic acid sequence encoding this sequence, as a biomarker, or as an active agent, with regard to a dandruff condition of the scalp.

Claims

1. An in vitro or ex vivo process for detecting insulin-degrading enzyme (IDE) in an individual having dandruff of the scalp, comprising: a) obtaining an isolated scalp sample from the individual; and b) performing, in the isolated sample of the scalp taken from said individual, a quantitative measurement of the expression of an amino acid sequence represented by a sequence consisting of SEQ ID No.: 8, or an analog of said amino acid sequence, said analog having a sequence identity of at least 85% with said amino acid sequence, having a biological activity of the same nature and being detectable by antibodies directed against an amino acid sequence represented by a sequence consisting of SEQ ID No.: 8.

2. An in vitro or ex vivo process for detecting insulin-degrading enzyme (IDE) in an individual having dandruff of the scalp, comprising: a) obtaining a first isolated scalp sample from the individual; b) performing, before implementing the cosmetic treatment of step c), in a first isolated scalp sample taken from said individual, at least a first quantitative measurement of the expression of at least one amino acid sequence represented by a sequence consisting of SEQ ID No.: 8, or an analog of said amino acid sequence, said analog having a sequence identity of at least 85% with said amino acid sequence, having a biological activity of the same nature and being detectable by antibodies directed against an amino acid sequence represented by sequence consisting of SEQ ID No.: 8, c) implementing a cosmetic treatment on the scalp of said individual, and d) performing, after implementing the cosmetic treatment of step c), in a second isolated scalp sample taken from said individual, at least a second quantitative measurement of the expression of said amino acid sequence represented by a sequence consisting of SEQ ID No.: 8, or an analog of said amino acid sequence, said analog having a sequence identity of at least 85% with said amino acid sequence, having a biological activity of the same nature and being detectable by antibodies directed against an amino acid sequence represented by a sequence consisting of SEQ ID No.: 8.

Description

FIGURES

(1) FIG. 1: shows the magnification of a protein spot, obtained by 2D difference electrophoresis proteomic analysis, identified as being IDE.

(2) FIG. 2: is a graph of the intensity of the volume of the spots observed in FIG. 1.

(3) FIG. 3: shows the immunodetection by Western blotting of IDE.

(4) FIG. 4: shows the mean intensity of the bands detected during the Western blotting illustrated by FIG. 3.

(5) For the purposes of the present invention, one should be understood, unless otherwise indicated, in the sense of at least one.

(6) The examples and figures hereinafter are presented by way of illustration and without implied limitation of the invention.

EXAMPLES

Example 1

(7) Expression of IDE in the Scalp

(8) 1Materials and Methods

(9) A 2D difference electrophoresis proteomic analysis performed using noninvasive samples of dandruff-free and dandruff-bearing scalps was performed.

(10) The study is performed on 6 dandruff-free volunteers (adherent dandruff grade from 0 to 0.25) and 6 dandruff-bearing volunteers (adherent dandruff grade from 3.25 to 4). The volunteers are 36 to 39 year-old men with short hair. The grading of the dandruff condition of the volunteers is performed by an expert according to the following standard classification which provides a scale of scores of adherent dandruff of between 0 and 5.

(11) The samples are taken by corneodisc (reference GODS 100, CuDerm) on an area of 2 cm. Four saturated corneodiscs are produced for each sample. The soluble proteins are extracted into a native buffer (TBS, 1M NaCl, 1% Triton X100). After filtration, the proteins are precipitated by addition of acetone. The protein pellet is dissolved in the extraction buffer III (BioRad; Ref.: 163-2104) supplemented with 40 mM of DTT. For each sample corresponding to a volunteer, two-dimensional electrophoresis is performed along a first dimension by IEF separation pH 3-11 on an 11 cm strip (GE-Healthcare) and along a second dimension on a 10.5-14% Criterion gradient gel (Bio-Rad) according to the suppliers' recommendations.

(12) After staining the gels with SyproRuby (reference S4942, Invitrogen) used according to the supplier's protocol, an image analysis is performed with the Progenesis software after image acquisition according to the following parameters: Excitation: 460/80; Emission: 620/30; Resolution 50 m; Exposure 3 s; Flat field: 1 s; Exposure 1.

(13) In a first step, the images are aligned together and the intensities normalized. In a second step, after having defined groups, a statistical analysis is performed with the Progenesis Samespots software (NonLinear Dynamics). A selection of spots is made as a function of the p value (less than 0.05), the fold (greater than 2, ratio of intensity between the most intense spot of one group and the least intense spot of another group) and the q value (greater than 0.8).

(14) The selected spots are then cut out and the proteins are digested with trypsin and analyzed by LC-MS/MS.

(15) The protein databank UniRef100.15.3.9606.homo-sapiens was interrogated to identify the proteins.

(16) 2Results

(17) IDE is identified among the spots selected in favor of an overexpression in the dandruff-bearing group according to the criteria defined previously.

(18) Magnification of this protein spot for each volunteer and the graphical representation of the volume of the spots is given by FIGS. 1 and 2.

(19) These results validate the use of IDE as a biomarker for the skin, in particular for skin desquamation, and more particularly for a dandruff condition of the scalp. IDE may thus be advantageously used for the screening of active agents for the treatment of a dandruff condition of the scalp or for characterizing the efficacy of a cosmetic skin treatment.

Example 2

(20) Confirmation by Western Blotting of the Expression of IDE in the Skin

(21) 1Materials and Methods

(22) The concentration of the samples described previously in example 1 is aligned, and 2 g are deposited in the wells of a 10-20% Criterion gel (BioRad). The proteins are thus separated by SDS-PAGE electrophoresis. After semidry transfer onto a PVDF membrane according to a standard protocol, the proteins are incubated with a primary antibody directed against the protein of interest, overnight at 4 C. A second incubation is then performed with a secondary antibody (coupled to a peroxidase) directed against the first antibody.

(23) An electrochemiluminescence kit is used for revealing the target proteins.

(24) The image is acquired with Fluor Smax (Biorad) and the bands are quantified using the Quantity-One software (Biorad). A commercial anti-IDE antibody was used at a dilution of 1/5000 (reference AB28560, AbCam).

(25) 2- Results

(26) FIG. 3 shows the Western blot image obtained. The mean intensity of the detected bands is represented by the histogram in FIG. 4.

(27) The immunodetection of IDE reveals an overexpression in the dandruff-bearing individuals. This observation confirms the differential identifications observed in example 1.

LITERATURE

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