METHODS UTILIZING DIFFERENTIAL SPLICING EVENTS IN BLOOD CELLS FOR THE DETECTION OF PATHOLOGICAL EVENTS

• US 20090264301A1
• Filed: 06/29/2009
• Published: 10/22/2009
• Est. Priority Date: 09/16/1999
• Status: Abandoned Application

• Current Assignee: Exonhit Therapeutics SA (Eurobio-Scientific SA)
• Original Assignee: Exonhit Therapeutics SA (Eurobio-Scientific SA)
• Inventors: Schweighoffer, Fabien, Bracco, Laurent, Tocque, Bruno
• Application Number:
  US12/494,039
• Publication Number:
  US 20090264301A1
• Time in Patent Office: Days
• US Class Current: 506/7
• CPC Class Codes: C12Q 1/6809 Methods for determination or identification of nucleic acids involving differential detection
• CPC Class Codes: G01N 33/56972 White blood cells

Abstract

The present invention concerns new compositions and methods for the detection of pathological events. It more specifically concerns methods for the detection in vitro of the presence of a pathology or a pathological event in a subject, comprising taking a sample of blood cells from the subject and determining, in this sample, the presence of blood cells presenting a physiological state characteristic of the pathology. The invention also concerns the tools, kits and compositions for the implementation of such methods, as well as their uses in the field of human and animal health, or in experimental research for example.

Claims

1. A method for in vitro detection of a given, predefined pathological condition associated with deregulation of a cell signaling pathway in a human subject, wherein said given, predefined pathological condition causes disease in a tissue distinct from nucleated blood cells of said human subject, said method comprising:

(i) providing a sample comprising nucleated blood cells from the subject being tested for the presence of said pathological condition,

(ii) preparing nucleic acid molecules from the sample of step (i),

(iii) hybridizing all or part of the nucleic acid molecules from step (ii) to at least one nucleic acid library capable of detecting the presence of said given, predefined pathological condition when contacted with a diverse population of nucleic acid molecules prepared from nucleated blood cells from a human subject having said given, predefined pathological condition under conditions allowing hybridization to occur, said library having an ordered arrangement on a support and comprising a plurality of nucleic acid molecules that specifically hybridize to differentially spliced ribonucleic acid molecules (RNAs) expressed in nucleated blood cells from human subjects known to have said given, predefined pathological condition, wherein said differentially spliced RNAs are characteristic of said given, predefined pathological condition that causes disease in a tissue distinct from nucleated blood cells, and

(iv) detecting hybridization between a plurality of said nucleic acid molecules of said subject being tested and said nucleic acid molecules of said library, wherein said hybridization allows determination of the presence or absence of said given, predefined pathological condition in said subject being tested.

2. The method of claim 1, wherein the nucleic acid molecules prepared from the sample are total or messenger RNA or complementary deoxyribonucleic acid (cDNA) derived therefrom.

3. The method of claim 2, wherein the nucleic acid molecules prepared from the sample are amplified.

4. The method of claim 1, wherein the nucleic acid molecules prepared from the sample are labeled.

5. The method of claim 1, for the detection in vitro of the stage of progression of said given, predefined pathological condition in said subject.

6. The method of claim 1, wherein said support is a membrane, a glass plate, or a biochip.

7. The method of claim 1, wherein said pathological condition is characterized by excessive cell proliferation.

8. The method of claim 1, wherein said nucleated blood cells comprise lymphocytes, macrophages, monocytes or dendritic cells.

9. The method of claim 1, wherein said nucleic acid library further comprises a control nucleic acid molecule.

10. A method for in vitro detection of a given, predefined pathological condition characterized by excessive cell proliferation in a human subject, wherein said given, predefined pathological condition causes disease in a tissue distinct from nucleated blood cells of said human subject, said method comprising:

(i) providing a sample comprising nucleated blood cells from the subject being tested for the presence of said pathological condition

(ii) preparing nucleic acid molecules from the sample of step (i),

(iii) hybridizing all or part of the nucleic acid molecules from step (ii) to at least one nucleic acid library capable of detecting the presence of said given, predefined pathological condition when contacted with a diverse population of nucleic acid molecules prepared from nucleated blood cells from a human subject having said given, predefined pathological condition under conditions allowing hybridization to occur, said library having an ordered arrangement on a support and comprising a plurality of nucleic acid molecules that specifically hybridize to differentially spliced ribonucleic acid molecules (RNAs) expressed in nucleated blood cells from human subjects known to have said given, predefined pathological condition, wherein said differentially spliced RNAs are characteristic of said given, predefined pathological condition that causes disease in a tissue distinct from nucleated blood cells, and

(iv) detecting hybridization between a plurality of said nucleic acid molecules of said subject being tested and said nucleic acid molecules of said nucleic acid library, wherein said hybridization allows determination of the presence or absence of said given, predefined pathological condition in said subject being tested.

11. The method of claim 10, wherein said given, predefined pathological condition is stenosis.

12. The method of claim 10, wherein said nucleated blood cells comprise lymphocytes, macrophages, monocytes or dendritic cells.

13. The method of claim 10, wherein said nucleic acid library further comprises a control nucleic acid molecule.