CELLS AND METHODS FOR SELECTION BASED ASSAY
Cells and methods for screening inhibitors against a heterologous target protein are disclosed.
1 . A cell comprising: i) one or more exogenous nucleic acids expressing one or more targets and ii) one or more genes native to the cell genetically modified and/or deleted, wherein the combination of the one or more targets with the genetic modification and/or deletion of one or more genes native to the cell is toxic to the cell.
2 . The cell of claim 1 , wherein genetic modifications and/or deletion of the one or more genes native to the cell provides a synthetic sick or synthetic lethal interaction to the cell.
3 . The cell of claim 1 , wherein the cell is a eukaryotic cell.
4 . The cell claim 3 , wherein the cell is a yeast cell, optionally wherein the yeast cell is Saccharomyces cerevisiae.
5 . (canceled)
6 . The cell of claim 1 , wherein the one or more targets comprises a disease target, optionally wherein said disease target is a human disease target.
7 . (canceled)
8 . The cell of claim 7 , wherein the disease target comprises or consists of MMSET.
9 . The cell of claim 1 , wherein the modified and/or deleted one or more genes native to the cell are selected from the group consisting of SET2, SWR1, and LGE1.
10 . (canceled)
11 . The cell of claim 1 , further comprising one or more nucleic acids encoding enzymes that produce candidate inhibitor compounds.
12 . The cell of claim 1 , wherein the one or more targets comprises a mixture of hyperactive targets and/or catalytically dead targets, the hyperactive targets and/or catalytically dead targets varied in relative abundance to calibrate relative toxicity to the cell.
13 . The cell of claim 12 , wherein the mixture of hyperactive targets and/or catalytically dead targets comprises one or more MMSET proteins, each having at least one or more of the following mutations: F1177A, Y1118A, Y1179A, and/or Y1092A, wherein the residues are numbered according to SEQ ID NO: 1.
14 . A method of detecting inhibitors of one or more targets, comprising:
a) providing a cell comprising one or more exogenous nucleic acids expressing the one or more targets;
b) genetically modifying and/or deleting one or more genes native to the cell, wherein the combination of the one or more targets with the genetic modification and/or deletion of the one or more genes native to the cell is toxic to the cell;
c) exposing the cell to candidate inhibitor compounds;
d) growing the cell under growth conditions; and
e) measuring growth of the cell,
wherein growth of the cell detects a candidate inhibitor compound as an inhibitor of the one or more targets.
15 . The method of claim 14 , wherein the combination of the one or more targets with genetic modification and/or deletion of the one or more genes native to the cell provides a synthetic sick or synthetic lethal interaction to the cell.
16 . The method of claim 14 , wherein the cell is a eukaryotic cell.
17 . The method of claim 16 , wherein the cell is a yeast cell, optionally wherein said yeast cell is Saccharomyces cerevisiae.
18 . (canceled)
19 . The method of claim 14 , wherein the one or more targets comprises a disease target, optionally wherein said disease target is a human disease target.
20 . (canceled)
21 . The method of claim 20 , wherein the disease target comprises or consists of MMSET.
22 . The method of claim 14 , wherein the modified and/or deleted one or more genes native to the cell are selected from the group consisting of SET2, SWR1, and LGE1.
23 . (canceled)
24 . The method of claim 14 , wherein exposing the cell to candidate inhibitor compounds comprises expressing in the cell nucleic acids encoding enzymes that produce the candidate inhibitor compounds.
25 . The method of claim 14 , wherein exposing the cell to candidate inhibitor compounds comprises contacting the cell with the candidate inhibitor compounds.
26 . The method of claim 25 , wherein contacting the cell with candidate inhibitor compounds comprises adding the candidate inhibitor compounds to a cell culture.
27 . The method of claim 14 , wherein the growth conditions omit one or more of histidine, uracil, and/or lysine.
28 . The method of claim 14 , wherein the growth conditions comprise growing the cell at a temperature of less than about 30° C.
29 . The method of claim 28 , wherein the growth conditions comprise growing the cell at a temperature of less than about 25° C.
30 . The method of claim 14 , wherein measuring growth of the cell comprises calculating population size using a Z-factor or Hedge's effect.
31 . The method of claim 14 , wherein the one or more targets comprises a mixture of hyperactive targets and/or catalytically dead targets varied in relative abundance to calibrate toxicity to the cell.
32 . The method of claim 31 , wherein the mixture of hyperactive targets and/or catalytically dead targets comprises one or more MMSET proteins having at least one or more of the following mutations: F1177A, Y1118A, Y1179A, and/or Y1092A, wherein the residues are numbered according to SEQ ID NO: 1.