DROSOPHILA TUMOR STEM CELL MODEL AND USES THEREOF

• US 20130096067A1
• Filed: 09/14/2012
• Published: 04/18/2013
• Est. Priority Date: 09/14/2011
• Status: Abandoned Application

• Current Assignee: Michele Markstein, Norbert Perrimon
• Original Assignee: Michele Markstein, Norbert Perrimon
• Inventors: Perrimon, Norbert, Markstein, Michele
• Application Number:
  US13/617,049
• Publication Number:
  US 20130096067A1
• Time in Patent Office: Days
• US Class Current: 514/20.1
• CPC Class Codes: A01K 2217/052 inducing gain of function
• CPC Class Codes: A01K 2217/15 Animals comprising multiple alterations of the genome, by transgenesis or homologous recombination, e.g. obtained by cross-breeding
• CPC Class Codes: A01K 2217/203 Animal model comprising inducible/conditional expression system, e.g. hormones, tet
• CPC Class Codes: A01K 2217/206 Animal model comprising tissue-specific expression system, e.g. tissue specific expression of transgene, of Cre recombinase
• CPC Class Codes: A01K 2267/0331 Animal model for proliferative diseases
• CPC Class Codes: A01K 2267/0393 Animal model comprising a reporter system for screening tests
• CPC Class Codes: A01K 67/0339 Genetically modified insects, e.g. Drosophila melanogaster, medfly
• CPC Class Codes: C12N 9/12 transferring phosphorus containing groups, e.g. kinases (2.7)

Abstract

The present invention provides a transgenic Drosophila tumor stem cell model and uses thereof. Also provided is a method to selectively inhibit cancer stem cells in a mammal having refractory intestinal cancer or a relapse of intestinal cancer.

Claims

1. A transgenic Drosophila comprising a plurality of expression cassettes, which plurality comprises at least four of the following expression cassettes:

a first expression cassette having a promoter operably linked to an open reading frame for a heterologous transcription factor;

a second expression cassette having a transcriptional regulatory element comprising a nucleotide sequence that binds the transcription factor and a promoter operably linked to an oncogene;

a third expression cassette having a transcriptional regulatory element comprising a nucleotide sequence that binds the transcription factor and a promoter operably linked to a first marker gene that encodes an optically detectable gene product;

a fourth expression cassette having a promoter operably linked to an open reading frame encoding a gene product that regulates the heterologous transcription factor; and

a fifth expression cassette having a transcriptional regulatory element comprising a nucleotide sequence that binds the transcription factor and a promoter operably linked to a second marker gene that encodes an optically detectable gene product.

2. The transgenic Drosophila of claim 1 wherein the first expression cassette has a gut-specific promoter operably linked to the open reading frame and the fourth expression cassette has a constitutive promoter operably linked to an open reading frame encoding a repressor of the transcription factor, which repressor is a conditional mutant.

3. The transgenic Drosophila of claim 1 which comprises all five expression cassettes.

4. The transgene Drosophila of claim 1 which comprises the first, third, fourth and fifth expression cassettes.

5. The transgenic Drosophila of claim 2 wherein the conditional mutant is a temperature sensitive mutant.

6. The transgenic Drosophila of claim 2 wherein the repressor represses the transcription factor at room temperature.

7. The transgenic Drosophila of claim 1 wherein one of the optically detectable gene products is a luminescent protein and the other is a fluorescent protein.

8. The transgenic Drosophila of claim 1 wherein one of the optically detectable markers is a luciferase.

9. The transgenic Drosophila of claim 1 wherein the oncogene is Raf^gof, myr-AKT or Notch^DN.

10. A method to screen compounds, comprising:

providing the transgenic Drosophila of claim 1 which expresses the transcription factor under conditions that allow for binding of the transcription factor to the nucleotide sequence;

exposing the transgenic Drosophila to one or more compounds; and

detecting at least one of the optically detectable gene products in or in a portion of the transgenic Drosophila exposed to the one or more compounds, wherein an alteration in the presence or amount of the at least one optically detectable gene product relative to the presence or amount of the corresponding optically detectable gene product in a corresponding control transgenic Drosophila that is not exposed to the one or more compounds or a portion of the control transgenic Drosophila, is indicative of a compound that alters the number of intestinal stem cells.

11. The method of claim 10 wherein the one or more compounds are ingested or inhaled by the transgenic Drosophila.

12. The method of claim 10 wherein the presence or amount of at least one of the optically detectable gene products is detected in the intestine.

13. The method of claim 10 wherein the presence or amount of at least one of the optically detectable gene products is detected in homogenates.

14. The method of claim 10 wherein one of the compounds decreases the number of intestinal tumor stem cells but not wild-type intestinal stem cells.

15. The method of claim 10 wherein one of the compounds enhances the number of wild-type intestinal stem cells.

16. The method of claim 10 wherein an increase in one of the optically detectable products is detected.

17. The method of claim 10 wherein a decrease in one of the optically detectable products is detected.

18. The method of claim 10 wherein the transgenic Drosophila is further exposed to an agent that activates the JNK-stress-related pathway and/or induces expression of IL-6 related cytokines.

19. The method of claim 10 wherein one of the detected compounds is a pathway-specific compound.

20. A method to selectively inhibit cancer stem cells in a mammal having refractory intestinal cancer or a relapse of intestinal cancer, comprising selecting a compound that alters the activity of intestinal tumor stem cells but not wild-type stem cells;

and administering to the mammal an effective amount of the compound.