Specific targeting of RNA expanded repeat sequences
The present invention provides small molecule compounds that can form covalent adducts with specific sequences of RNA, such as the hairpin loop r(CUG) exp sequence which is a cause of myotonic dystrophy type 1 (DM1), or the r(CGG) exp sequence which is a cause of fragile X-associated tremor/ataxia syndrome (FXTAS); to methods of making the small molecule compounds; and to methods of using the small molecular compounds in the treatment of DM1 or of FXTAS in patients afflicted therewith. The invention further provides a method for identifying an RNA target of a small molecule drug in vivo, using a small molecule drug conjugated to an RNA-reactive crosslinker group and a reporter group, contacting a cell or nucleic acid extract with the small molecule drug conjugate, then separating RNA targets crosslinked to the small molecule drug conjugate by interaction of the affinity group with a complementary affinity group.
1. A compound of formula (I)
wherein
each individually selected R or R′ is H or (C1-C6)alkyl;
n1 is 2, 3, 4, 5, or 6;
Z is a (C1-C3)alkylene group, optionally substituted with a reporter or affinity group;
L is a linker group comprising an optionally substituted (C1-C6)alkylene, wherein one or two carbon atoms is optionally replaced by O;
XL is an RNA-reactive crosslinking group;
each individually selected Ht is a group of formula
wherein
n2 is 0, 1, 2, or 3;
n3 is 0, 1, 2, or 3;
Y is O or CH 2 ;
a wavy line indicates a point of bonding;
or any tautomer thereof; or any salt thereof, or any stereoisomer thereof.
2. The compound of claim 1 wherein the crosslinking group is an alkylating group.
3. The compound of claim 2 wherein the alkylating group is a chlorambucil derivative.
4. The compound of claim 1 wherein the crosslinking group is a triggered crosslinker using photochemical or shape-triggered catalysis.
5. The compound of claim 1 wherein the compound is of formula
or is of formula
wherein Ht, R, and R′, are as defined in claim 1 ;
or any tautomer thereof; or any salt thereof, or any stereoisomer thereof.
6. The compound of claim 1 wherein Ht is
wherein a wavy line indicates a point of bonding;
R is n-propyl; R′ is H; or any combination thereof;
or any tautomer thereof; or any salt thereof, or any stereoisomer thereof.
7. The compound of claim 1 wherein the compound is 2H-4-CA of formula
or is compound 2H-5-CA of formula
wherein Ht is a group of formula
wherein a wavy line indicates a point of bonding;
or any tautomer thereof; or any salt thereof, or any stereoisomer thereof.
8. The compound of claim 1 wherein Z is an unsubstituted alkylene group.
9. The compound of claim 1 wherein Z is an alkylene group substituted with an affinity group.
10. The compound of claim 9 wherein the affinity group is a biotin-comprising moiety.
11. The compound of claim 10 , wherein the compound is
or any tautomer thereof; or any salt thereof, or any stereoisomer thereof.
12. The compound of claim 11 , wherein n1 is 4 or 5.
13. The compound of claim 12 , wherein the compound is 2H-4-CA-biotin of formula
or is 2H-5-CA-biotin of formula
or any tautomer thereof; or any salt thereof, or any stereoisomer thereof.
14. A covalently-linked complex of the compound of formula (I) of claim 1 and a segment of RNA comprising a hairpin loop.
15. The complex of claim 14 wherein the RNA comprises an r(CUG) exp segment or wherein the RNA comprises an r(CGG) exp segment.
16. A method of inactivating an RNA, comprising contacting the RNA with a compound of claim 1 , wherein inactivating the RNA comprises interfering with molecular associations that the RNA makes in a living cell, wherein the RNA and the compound form a covalent bond to provide a covalently-linked complex.
17. The method of claim 16 wherein interfering with molecular associations comprises interfering with the association of the RNA comprising the r(CUG) exp segment and muscleblind-like 1 protein MBL1.
18. The method of claim 16 wherein interfering with molecular associations comprises interfering with the association of the RNA comprising the r(CGG) exp segment and DiGeorge Syndrome Critical Region 8 protein (DGCR8), or Src-associated in mitosis, 68 kDa protein (Sam68).
19. A method of a treatment of a patient suffering from DM1, comprising administering to the patient an effective dose of the compound of claim 1 .
20. A method of a treatment of a patient suffering from fragile X-associated tremor/ataxia syndrome (FXTAS), comprising administering to the patient an effective dose of the compound of claim 1 .