US 12,410,472 B2
Method for evaluating the eye irritation potential of chemicals
Hervé Groux, Le Rouret (FR); Françoise Cottrez, Le Rouret (FR); Nathalie Alepee, Livry Gargan (FR); and Virginie Leblanc, Cachan (FR)
Assigned to IMMUNOSEARCH, Le Plan de Grasse (FR)
Appl. No. 16/334,269
Filed by IMMUNOSEARCH, Le Plan de Grasse (FR)
PCT Filed Sep. 19, 2017, PCT No. PCT/EP2017/073675
§ 371(c)(1), (2) Date Mar. 18, 2019,
PCT Pub. No. WO2018/050927, PCT Pub. Date Mar. 22, 2018.
Claims priority of application No. 1658746 (FR), filed on Sep. 19, 2016.
Prior Publication US 2019/0300952 A1, Oct. 3, 2019
Int. Cl. C12Q 1/68 (2018.01); C12Q 1/02 (2006.01); C12Q 1/6876 (2018.01); C12Q 1/6883 (2018.01)
CPC C12Q 1/6876 (2013.01) [C12Q 1/02 (2013.01); C12Q 1/6883 (2013.01); C12Q 2531/113 (2013.01); C12Q 2600/106 (2013.01); C12Q 2600/142 (2013.01); C12Q 2600/158 (2013.01)] 4 Claims
 
1. A method for evaluating the eye irritation potential of a test compound, comprising the steps of:
a) bringing a test compound into contact with an in vitro reconstructed corneal sample;
b) measuring the expression of at least 10 genes selected from the group consisting of: HSP90AA1, COL7A1, NOS3, MMP8, CASP1, ELN, IL-23R, DLK1, CLEC4D, IL-24, CCL22, SLIT2, ICAM2, MUC13, HSPA1A, FSHR, IL-1R2, ALB, CCND1, CXCL1, CXCR1, KL, COL6A2, MUC4, DDIT3, MMP3, HAS1, IRF1 and CYR61, wherein at least two of the genes are DLK1 and IL-1R2;
c) comparing the expression of each measured gene in the in vitro reconstructed corneal sample to a control sample in which the expression of said measured gene in the absence of said test compound is measured;
d) measuring an overexpression of the genes measured in step b) wherein an overexpression threshold value is assigned to each gene measured in step b), wherein the overexpression threshold value corresponds to a factor of increase in the expression of the gene when the in vitro reconstructed corneal sample is brought into contact with the test compound relative to the expression of said gene the control sample, and wherein an eye irritation index of the test compound corresponds to the overexpression threshold value,
e) categorizing the test compound as not irritant, reversible irritant, or irreversible irritant based on the eye irritation index of the test compound wherein when the test compound is brought into contact with the in vitro reconstructed corneal sample in a solid form at 60 mg/cm2, if the eye irritation index of the test compound is greater than or equal to 20, the test compound is an irreversible irritant, if the eye irritation index of the test compound is greater than or equal to 10 and less than 20 the test compound is a reversible irritant, and if the eye irritation index of the test compound is less than 10 the test compound is not an irritant, and
f) including the test compound in a pharmaceutic or cosmetic if the test compound is categorized as not irritant or reversible irritant and excluding the test compound in a pharmaceutic or cosmetic if the test compound is categorized as irreversible irritant.