Development of ABCG2-Sensitive Fluorescent Probe For Isolation of ABCG2low Neural Stem/Progenitor Cells

A method of visualizing a target cell, the method comprising:

• a) contacting a population of the target cell with a composition to form an incubation media;
• b) incubating the incubation media of step (a) for a period of time sufficient to stain the target cells;
• and
• c) visualizing the stained target cells of step (b) with fluorescence microscopy to visualize the target cell;
• wherein the composition is represented by structural formula (I):
• or a salt and/or a tautomer thereof, wherein
• n is a whole number selected from 1 to 5;
• X for each occurrence is independently selected from H, (C1-C20)alkyl, (C2-C20)alkenyl, (C2-C20)alkynyl, (C1-C20)alkoxy, (C1-C20)alkylamino, (C3-C10)cycloalkyl, —C(O)R1, —S(O)2R1, amino, pyridyl, nitrile, nitro or —C(O)N(R1)(R2);
• R1 is H, amino, (C1-C20)alkyl, (C2-C20)alkenyl, (C2-C20)alkynyl, (C1-C20)alkoxy, (C1-C20)alkylamino or (C3-C10)cycloalkyl, optionally substituted with one or more groups independently selected from (C1-C10)alkyl, (C3-C10)cycloalkyl, halo, (C6-C12)aryl, (5-12 atom) heteroaryl, (5-12 atom) heterocycle, or —C(O)O(C1-C3)alkyl, further optionally substituted with one or more groups selected from halo, (C6-C12)aryl, (C1-C3)alkyl, (C1-C3)alkoxy, —OCF3 or oxo;
• R2 is H, amino, (C1-C20)alkyl, (C2-C20)alkenyl, (C2-C20)alkynyl, (C1-C20)alkoxy, (C1-C20)alkylamino or (C3-C10)cycloalkyl, optionally substituted with one or more groups independently selected from (C1-C10)alkyl, (C3-C10)cycloalkyl, halo, (C6-C12)aryl, (5-12 atom) heteroaryl, (5-12 atom) heterocycle, or —C(O)O(C1-C3)alkyl, further optionally substituted with one or more groups selected from halo, (C6-C12)aryl, (C1-C3)alkyl, (C1-C3)alkoxy, —OCF3 or oxo;
• or R1 and R2 may be taken together to form a ring, wherein the ring is optionally substituted with one or more groups selected from (C1-C10)alkyl, (C3-C10)cycloalkyl, halo, (C6-C12)aryl, (5-12 atom) heteroaryl, (5-12 atom) heterocycle, —C(O)O(C1-C3)alkyl, or a 4-5 member polycyclyl fused to the ring, further optionally substituted with one or more groups selected from halo, (C6-C12)aryl, (C1-C3)alkyl, (C1-C3)alkoxy, —OCF3, or oxo.

A method of isolating a neural stem cell, the method comprising:

• a) visualizing the neural stem cell by contacting a population of the neural stem cells with a composition to form an incubation media, incubating the incubation media for a period of time sufficient to stain the neural stem cells, and visualizing the stained neural stem cells with fluorescence microscopy to visualize the neural stem cell;
• b) exciting the neural stem cells by exposing the incubation media to light of a wavelength of about 488 nm to about 561 nm;
• and
• c) separating the excited neural stem cells from the incubation media by fluorescence activated cell sorting using a bandpass filter configured to detect light emitted at about 529±28 nm;
• wherein the composition is represented by structural formula (I):
• or a salt and/or tautomer thereof, wherein
• n is a whole number selected from 1 to 5;
• X for each occurrence is independently selected from H, (C1-C20)alkyl, (C2-C20)alkenyl, (C2-C20)alkynyl, (C1-C20)alkoxy, (C1-C20)alkylamino, (C3-C10)cycloalkyl, —C(O)R1, —S(O)2R1, amino, pyridyl, nitrile, nitro or —C(O)N(R1)(R2);
• R1 is H, amino, (C1-C20)alkyl, (C2-C20)alkenyl, (C2-C20)alkynyl, (C1-C20)alkoxy, (C1-C20)alkylamino or (C3-C10)cycloalkyl, optionally substituted with one or more groups independently selected from (C1-C10)alkyl, (C3-C10)cycloalkyl, halo, (C6-C12)aryl, (5-12 atom) heteroaryl, (5-12 atom) heterocycle, or —C(O)O(C1-C3)alkyl, further optionally substituted with one or more groups selected from halo, (C6-C12)aryl, (C1-C3)alkyl, (C1-C3)alkoxy, —OCF3 or oxo;
• R2 is H, amino, (C1-C20)alkyl, (C2-C20)alkenyl, (C2-C20)alkynyl, (C1-C20)alkoxy, (C1-C20)alkylamino or (C3-C10)cycloalkyl, optionally substituted with one or more groups independently selected from (C1-C10)alkyl, (C3-C10)cycloalkyl, halo, (C6-C12)aryl, (5-12 atom) heteroaryl, (5-12 atom) heterocycle, or —C(O)O(C1-C3)alkyl, further optionally substituted with one or more groups selected from halo, (C6-C12)aryl, (C1-C3)alkyl, (C1-C3)alkoxy, —OCF3 or oxo;
• or R1 and R2 may be taken together to form a ring, wherein the ring is optionally substituted with one or more groups selected from (C1-C10)alkyl, (C3-C10)cycloalkyl, halo, (C6-C12)aryl, (5-12 atom) heteroaryl, (5-12 atom) heterocycle, —C(O)O(C1-C3)alkyl, or a 4-5 member polycyclyl fused to the ring, further optionally substituted with one or more groups selected from halo, (C6-C12)aryl, (C1-C3)alkyl, (C1-C3)alkoxy, —OCF3, or oxo.