WO2012148297A4 - Method for determining skin sensitisation potential - Google Patents
Method for determining skin sensitisation potential Download PDFInfo
- Publication number
- WO2012148297A4 WO2012148297A4 PCT/PT2012/000018 PT2012000018W WO2012148297A4 WO 2012148297 A4 WO2012148297 A4 WO 2012148297A4 PT 2012000018 W PT2012000018 W PT 2012000018W WO 2012148297 A4 WO2012148297 A4 WO 2012148297A4
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- genes
- predictor
- compounds
- sensitizers
- cell line
- Prior art date
Links
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/5005—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells
- G01N33/5008—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics
- G01N33/502—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics for testing non-proliferative effects
- G01N33/5041—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics for testing non-proliferative effects involving analysis of members of signalling pathways
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/5005—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells
- G01N33/5008—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics
- G01N33/5044—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics involving specific cell types
- G01N33/5047—Cells of the immune system
Abstract
The present invention provides an in vitro method for predicting the in vivo skin sensitization potential of chemical compounds using a combination of mammalian cell models with multiple endpoints analysis. The method comprises the steps of: 1- selection of the chemicals concentrations to be tested using cell viability assays 2-detection of expression levels of genes implicated in skin sensitization 3-detection of the activation status of intracellular signaling pathways 4- outputs from points 2 and 3 are assessed for a group of chemical compounds comprising known sensitizers and irritants (training compounds) and used to build a statistical classification model trained and optimized to classify unknown chemicals compounds (test compounds) as either sensitizers or non-sensitizers.
Claims
1. Method for determining skin sensitisat.ion potential of chemical3 based, on the combined analysis of the expression levels of a set of genes and on the activation status of a set of intracellular signaling pathways involved in immunization response events, using cell models with dendritic cell charsc' er.ist.ics, Compi-'isihg the steps:
(a), culturing; a uman nr. mouse monocytic cell line with features of antigen presenting cells;
(b). applying selected Coric.en Lxatians of known sensitizers and irritants (training compounds) during a predeterminec period of time, to the cells of step (a) based 011 the results obtained from viability assays;
(c) measuring the expression level of one or more predictor key genes in the cells exposed to training compounds of step (b) ;
(d) measuring the activation status of a set of intracellular signalling pathways involved in immunization response events., such as skin inflammation or llergic events, which are selected from p33 MAPE, J , ERK, P A/AMPc, calcium release, PKC.,. ART/PI 3 , X.rf-2, A?-l, CR;!B , F-kB and JAK STAT in the cells exposed to training compou ds of step b) ;
(e) conducting a statistical discriminant analysis where the expression level (s) measured in step (c) and the phosphorylation levels measured in step (d) were used to build a statistical discrimant classification model trained and optimized to classify chemical compounds as either sensitizers or non-sensi Lizers ;
(£:) applying selected .concentrations oi a test compound during a predetermined period of time, wherein the concentrations of chemical compounds were chosen based an induced cell mortality between 10-50%, to the cel s of step (.a) based on the results obtained from viability assays;
(g). measuring the expression level of one or mere key predicto genes in the cells exposed to test compound of step (f) ;
ih) measuring the activation status. of predictor intracellular signalling pathways in the cells exposed to test compound of step (f)
(i) classifying the test compound through the statistical discriminant classifica ion model from step (e) as Dolonging either to sensitizer or nor.-s.ensitizer class.
2. The method according, to claim 1 which comprises the analysis of the expression of key gene markers associated with aki sens tiz tion., which are 3Ql.ec.ted from FABP4, I.L-1.7F, GXCL10 and genes related to elec ophilic. stress,.
3. The method according to claim 1 or 2, wherein the suitable cell culture is a human or mouse ceil line with feature of skin dendritic cells selected from FSDC cell line, MUTZ-3. cell line, ΤΉΡ-1 cell line and U937 cell line.
4. The method according to the any of the preceding claims, wherei the preferable cell culture model is a mouse fetal sk'i rs-detived dendritic cell line FSDC.
5. The method according to any of the preceding claims, wherein the statistical discriminant classification model is built based on the simultaneous analysis of the expression level of one or more key predictor genes and on the activation, of one. o more predictor intracellular signalling pathwa s .
6. The method according to the claim 5, wherein the
29 number of key predictor genes analysed is at least 1, preferably is at least' '.3:.
7. The method according to the claim 6, wherein one of the predictor genes is CXCL-10.
8. The method according to- the claim 5 , wherein the number of predictor signalling pathways analysed, is at leas 1., prefera l is at least 2.
9. The method, according to the claim; 8 , wherein the preferential red ctor signalling pathways analysed are p38 MAPK and JNK..
10. The method according, to the. claim 9, wherein JNK signalling pathway is used to discriminate .sensitizers from non- s:eiisit.izers
11. The method according to any of the preceding claims, wherein the tested chemical could be a synthetic chemical, a natural occurring chemical, a botanical fragrance or a drug .
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| PT105663 | 2011-04-29 | ||
| PT10566311 | 2011-04-29 |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| WO2012148297A1 WO2012148297A1 (en) | 2012-11-01 |
| WO2012148297A4 true WO2012148297A4 (en) | 2013-02-14 |
Family
ID=46317475
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| PCT/PT2012/000018 WO2012148297A1 (en) | 2011-04-29 | 2012-04-27 | Method for determining skin sensitisation potential |
Country Status (1)
| Country | Link |
|---|---|
| WO (1) | WO2012148297A1 (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP5997543B2 (en) * | 2012-08-13 | 2016-09-28 | 富士フイルム株式会社 | Reagent for skin sensitization |
Family Cites Families (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP2059803A4 (en) * | 2006-09-12 | 2011-02-09 | Entelos Inc | Apparatus and method for computer modeling chemical sensitivity of skin |
| EP2294412A1 (en) * | 2008-06-04 | 2011-03-16 | Ceetox Inc. | Method for predicting skin sensitizing activity of compounds |
-
2012
- 2012-04-27 WO PCT/PT2012/000018 patent/WO2012148297A1/en active Application Filing
Also Published As
| Publication number | Publication date |
|---|---|
| WO2012148297A1 (en) | 2012-11-01 |
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