Transcription biomarkers of biological responses and methods

• US 20110065100A1
• Filed: 09/16/2009
• Published: 03/17/2011
• Est. Priority Date: 09/16/2009
• Status: Active Grant

• Current Assignee: Active Motif, Inc.
• Original Assignee: SwitchGear Genomics, Inc. (Active Motif, Inc.)
• Inventors: Collins, Patrick, Trinklein, Nathan D., Aldred, Shelley Force, Rose, Michael
• Granted Patent: US 8,815,779 B2
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: C12N 15/1037 Screening libraries presented on the surface of microorganisms, e.g. phage display, E. coli display
• CPC Class Codes: C12Q 1/6876 Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
• CPC Class Codes: C12Q 1/6883 for diseases caused by alterations of genetic material
• CPC Class Codes: C12Q 1/6886 for cancer immunoassay for cancer G01N33/574
• CPC Class Codes: C12Q 2600/136 Screening for pharmacological compounds
• CPC Class Codes: C12Q 2600/158 Expression markers
• CPC Class Codes: G01N 33/5011 for testing antineoplastic activity

Abstract

This invention provides transcription regulatory control sequences, the activity of which function as biomarkers for a variety of biological responses. This invention also provides expression constructs in which a biomarker transcription regulatory sequence is operably linked with a sequence for a reporter. Cells that comprise these expression constructs can be used in assays to identify conditions that modulate activity of the biological response.

Claims

1. A collection comprising a plurality of expression constructs, each expression construct comprising a transcriptional regulatory sequence operatively linked with a heterologous sequence encoding a reporter, wherein at least 20% of the expression constructs in the collection each comprise a transcription regulatory sequence of a different gene of Table 1.

2. The collection of claim 1 wherein the transcription regulatory sequences of the different genes of Table 1 are biomarkers of the same biological response.

3. The collection of claim 1 wherein the transcription regulatory sequences of the different genes of Table 1 comprise biomarkers of different biological responses.

4. The collection of claim 1 wherein the expression construct is comprised in a plasmid, a virus, artificial chromosome, or a transposon vector or is integrated into a chromosome in a cell or maintained as a stable episomal vector.

5. The collection of claim 1 wherein the reporter is a light-emitting reporter, a fluorescent reporter or a colorimetric reporter.

6. A collection of a plurality of cells comprising expression constructs, each expression construct comprising a transcriptional regulatory sequence operatively linked with a heterologous sequence encoding a reporter, wherein at least 20% of the expression constructs each comprise a transcription regulatory sequence of a different gene of a section of Table 1.

7. A cell line comprising at least one expression construct that is stably integrated into a chromosome or stably maintained in a episomal vector wherein the expression construct comprises a transcriptional regulatory sequence operatively linked with a heterologous sequence encoding a reporter, wherein the expression construct comprises a transcription regulatory sequence of a gene of Table 1.

8. A device comprising a plurality of receptacles, wherein each receptacle comprises an expression construct comprising a transcriptional regulatory sequence operatively linked with a heterologous sequence encoding a reporter, wherein at least 20% of the expression constructs each comprise a transcription regulatory sequence of a different gene of Table 1.

9. A device comprising a plurality of receptacles, wherein each receptacle contains at least one cell comprising an expression construct, each expression construct comprising a transcriptional regulatory sequence operatively linked with a heterologous sequence encoding a reporter, wherein at least 20% of the expression constructs each comprise a transcription regulatory sequence of a different gene of Table 1.

10. A method comprising:

a) exposing at least one cell to a test condition, wherein the at least one cell comprises at least one expression construct comprising a transcriptional regulatory sequence operatively linked with a heterologous sequence encoding a reporter, wherein the transcription regulatory sequence is a transcription regulatory sequence of a gene of Table 1;

b) measuring expression of the reporter; and

c) correlating the measurement with a biological response for which the transcription regulatory sequence is a biomarker.

11. The method of claim 11 further comprising

i) measuring the activity of the reporter in the absence of the test condition or a control condition;

and

ii) measuring the activity of the reporter in the presence of the test condition; and

iii) measuring the difference or ratio of reporter activity between the cells exposed to the test condition and cells that were not; and

iv) correlating the difference or ratio between treated and untreated measurements with a biological response.

12. The method of claim 11 wherein the transcription regulatory sequence comprises a sequence selected from the group consisting of

(i) any of SEQ ID NO:

1 to SEQ ID NO;

174

(ii) a sequence of at least 50 nucleotides having at least 95% sequence identity to a sequence of (i)

(iii) a fragment of a sequence of (i).

13. The method of claim 11 wherein the test condition is selected from the group consisting of contacting the cell with a test compound, exposing the cell to an environmental condition and inducing or repressing expression of one or more genes in the cell.

14. The method of claim 11 wherein the test condition comprises contacting the cell with a test compound selected from:

(1) a small organic molecule;

(2) a nucleic acid derivative (e.g., a small interfering RNA, micro RNA mimic or micro RNA inhibitor); and

(3) an expression construct that contains an open reading frame of a gene.

15. The method of claim 11 wherein each expression construct is comprised in a different cell.

16. The method of claim 11 wherein the biological responses are responses selected from the hypoxia response, glucocorticoid response, estrogen response, PPAR-mediated response, retinoic acid response, heat shock response, p53-mediated response, NFkB-mediated response, STAT1-mediated response, CREB-mediated response, serum response and SREBP-mediated response.

17. The method of claim 11 further comprising, if the measurement is at a target level;

i) exposing at least one second cell to the test condition, wherein the at least one second cell comprises at least one expression construct comprising a transcriptional regulatory sequence operatively linked with a heterologous sequence encoding a reporter, wherein the transcription regulatory sequence is a transcription regulatory sequence of a different gene of the same biological response of Table 1; and

;

ii) measuring expression of the reporter sequence in the at least one second cell.