Macrocyclic benzodiazepine dimers, conjugates thereof, preparation and uses

1. A compound having a structure represented by formula I

X is

X 1 is CH 2 , O, NH, S(O) 0-2 , 3- to 7-membered cycloalkylene or heterocycloalkylene unsubstituted or substituted with (CH 2 ) 0-5 X 2 or O(CH 2 ) 2-5 X 2 , or 5- to 6-membered arylene or heteroarylene unsubstituted or substituted with (CH 2 ) 0-5 X 2 or O(CH 2 ) 2-5 X 2 ;

each X 2 is independently Me, CO 2 H, NH 2 , NH(C 1 -C 5 alkyl), N(C 1 -C 5 alkyl) 2 , SH, CHO, N(CH 2 CH 2 ) 2 N(C 1 -C 3 alkyl), N(CH 2 CH 2 ) 2 NH, NHNH 2 , or C(═O)NHNH 2 ;

Y is (CH 2 ) 4-6 CH═CH(CH 2 ) 4-6 , (CH 2 ) 4-6 X 1 (CH 2 ) 4-6 , or (CH 2 ) 2 (OCH 2 CH 2 ) 2-3 ;

each R 1 and R 2 is independently H, F, Cl, Br, OH, C 1 -C 3 alkyl, O(C 1 -C 3 alkyl), cyano, (CH 2 ) 0-5 NH 2 , or NO 2 ;

each double line in a diazepine ring system independently represents a single bond or a double bond;

each R 3 is H if the double line to the N to which it is attached is a single bond and is absent if the double line is a double bond;

each R 4 is H, OH, SO 3 Na, or SO 3 K if the double line to the C to which it is attached is a single bond and is absent if the double line is a double bond;

A and B are independently according to formula Ia or Ib

wherein, in formula Ia

Y′ and Y″ are independently absent, CH 2 , C═O, or CHR 12 ; wherein each R 12 is independently F, Cl, Br, or C 1 -C 3 alkyl, with the proviso that Y′ and Y″ are not both absent;

each G is independently C or N, with the proviso that no more than two Gs are N; and

each R 5 , R 6 , R 7 , and R 8 is independently H, C 1 -C 5 alkyl, C≡C(CH 2 ) 1-5 X 2 , OH, O(C 1 -C 5 alkyl), cyano, NO 2 , F, Cl, Br, O(CH 2 CH 2 O) 1-8 (C 1-3 alkyl), (CH 2 ) 0-5 X 2 , O(CH 2 ) 2-5 X 2 , 3- to 7-membered cycloalkyl or heterocycloalkyl unsubstituted or substituted with (CH 2 ) 0-5 X 2 or O(CH 2 ) 2-5 X 2 , 5- to 6-membered aryl or heteroaryl unsubstituted or substituted with (CH 2 ) 0-5 X 2 or O(CH 2 ) 2-5 X 2 ,

or where a R 5 , R 6 , R 7 , or R 8 is attached to a G that is N, such R 5 , R 6 , R 7 , or R 8 is absent;

and

wherein, in formula Ib,

the dotted lines indicate the optional presence of a C1-C2, C2-C3, or C2-R 10 double bond;

R 10 is H, ═O, ═CH 2 , ═CH(C 1 -C 5 alkyl), CH═CH(CH 2 ) 1-5 X 2 , C≡C(CH 2 ) 1 —X 2 , C 1 -C 5 alkyl, OH, O(C 1 -C 5 alkyl), cyano, NO 2 , F, Cl, Br, O(CH 2 CH 2 O) 1-8 (C 1-3 alkyl), (CH 2 ) 0-5 X 2 , 4- to 7-membered cycloalkyl, or heterocycloalkyl unsubstituted or substituted with (CH 2 ) 0-5 X 2 , O(CH 2 ) 2-5 X 2 , 3- to 7-membered cycloalkyl or heterocycloalkyl unsubstituted or substituted with (CH 2 ) 0-5 X 2 or O(CH 2 ) 2-5 X 2 , 5- to 6-membered aryl or heteroaryl unsubstituted or substituted with (CH 2 ) 0-5 X 2 or O(CH 2 ) 2-5 X 2 ; and

R 9 is absent if a C1-C2, C2-C3, or C2-R 10 double bond is present and otherwise is H;

wherein a heterocycloalkyl group has 1 to 2 carbons replaced with a heteroatom independently selected from N, O, or S, where the N and S optionally may be oxidized and the N optionally may be quaternized; and

wherein a heteroaryl group has 1 to 4 heteroatoms independently selected from N, O, or S, where the N and S optionally may be oxidized and the N optionally may be quaternized;

or a pharmaceutically acceptable salt thereof.

2. A compound according to claim 1 , having a structure represented by formula IIa:

wherein

x is 3 or 5;

Y is (CH 2 ) 7-12 , (CH 2 ) 2 (OCH 2 CH 2 ) 1-3 , (CH 2 ) 2-4 NH(CH 2 ) 2-4 or (CH 2 ) 2-4 NH((CH 2 ) 0-1 phenyl)(CH 2 ) 2-4 where the phenyl group is optionally substituted with NH 2 ;

A is A1, A2, or A3

B is B1, B2, or B3

3. A compound according to claim 1 , having a structure represented by formula IIb:

wherein

x is 3 or 5;

each Y′ is independently absent or CH 2 ;

Y is (CH 2 ) 7-12 , (CH 2 ) 2 (OCH 2 CH 2 ) 1-3 , (CH 2 ) 2-4 NH(CH 2 ) 2-4 or (CH 2 ) 2-4 NH((CH 2 ) 0-1 phenyl)(CH 2 ) 2-4 where the phenyl group is optionally substituted with NH 2 ; and

R 40 and R 41 are independently H, Cl, Br, OH, O(C 1-3 alkyl), NH 2 , or C 1-3 alkyl.

4. A compound according to claim 1 , having a structure represented by formula IIc

wherein

x is 3 or 5;

Y is (CH 2 ) 7-12 , (CH 2 ) 2 (OCH 2 CH 2 ) 1-3 , (CH 2 ) 2-4 NH(CH 2 ) 2-4 or (CH 2 ) 2-4 NH((CH 2 ) 0-1 phenyl)(CH 2 ) 2-4 where the phenyl group is optionally substituted with NH 2 ; and

R 42 and R 43 are independently H, OMe, NH 2 , OCH 2 CH 2 OMe, N(CH 2 CH 2 )O, N(CH 2 CH 2 )NMe, or N(CH 2 CH 2 )NH.

5. A compound according to claim 1 , having a structure represented by formula IIb-6, IIc-8, IIc-9, IIc-10, IIc-11, IId-1, IId-2, or IId-3

6. A compound having a structure represented by formula III:

wherein

R 60 is according to formula IIIa, IIIa′, or IIIa″

Y is (CH 2 ) 6-10 ;

x is 3 or 5;

each y is independently 2, 3, or 4;

A and B are independently according to formula Ia or Ib

wherein, in formula Ia

Y′ and Y″ are independently absent, CH 2 , C═O, or CHR 12 ; wherein each R 12 is independently F, Cl, Br, or C 1 -C 3 alkyl, with the proviso that Y′ and Y″ are not both absent;

each G is independently C or N, with the proviso that no more than two Gs are N; and

each R 5 , R 6 , R 7 , and R 8 is independently H, Cl, Br, C 1-3 alkyl, NO 2 , CN, NH 2 , O(C 1-3 alkyl), or (OCH 2 CH 2 ) 1-2 O(C 1-3 alkyl);

or where a R 5 , R 6 , R 7 , or R 8 is attached to a G that is N, such R 5 , R 6 , R 7 , or R 8 is absent;

and

wherein, in formula Ib,

the dotted lines indicate the optional presence of a C1-C2, C2-C3, or C2-R 10 double bond;

R 9 is absent if a C1-C2, C2-C3, or C2-R 10 double bond is present and otherwise is H;

R 10 is H, Cl, Br, ═CH 2 , ═CH(C 1-5 alkyl), C 1-3 alkyl, NO 2 , CN, or NH 2 ;

A′ is

R 50 is H, Cl, Br, C 1-3 alkyl, NO 2 , CN, NH 2 , O(C 1-3 alkyl), or (OCH 2 CH 2 ) 1-2 O(C 1-3 alkyl);

R 51 is H, Cl, Br, C 1-3 alkyl, NO 2 , CN, or NH 2 ;

T is a self-immolating group;

t is 0 or 1;

AA a and each AA b are independently selected from the group consisting of alanine, β-alanine, γ-aminobutyric acid, arginine, asparagine, aspartic acid, γ-carboxyglutamic acid, citrulline, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, norleucine, norvaline, ornithine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine;

p is 1, 2, 3, or 4;

q is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10;

r is 1, 2, 3, 4, or 5;

s is 0 or 1; and

R 31 is

7. A compound according to claim 6 , wherein R 60 is IIIa.

8. A compound according to claim 6 , wherein R 60 is IIIa′.

9. A compound according to claim 6 , wherein R 60 is IIIa″.

10. A compound according to claim 6 , having a structure represented by formula IIIa-1, IIIa-2, IIIa-3, IIIa-4, IIIa-5, IIIa-6, or IIIa-7

11. A conjugate comprising a compound according to claim 1 conjugated to an antibody.

12. A conjugate according to claim 11 , having a structure represented by formula IV:

wherein

Ab is an antibody;

m is 1, 2, 3, or 4;

R 40 is

where the open valence of R 40 that is bonded to Ab is denoted by an asterisk (*) and the open valence of R 40 that is bonded to (CH 2 ) r is denoted by a wavy line ( );

R 60 is according to formula IIIa, IIIa′, or IIIa″

Y is (CH 2 ) 6-10 ;

x is 3 or 5;

each y is independently 2, 3, or 4;

A and B are independently according to formula Ia or Ib

wherein, in formula Ia

Y′ and Y″ are independently absent, CH 2 , C═O, or CHR 12 ; wherein each R 12 is independently F, Cl, Br, or C 1 -C 3 alkyl, with the proviso that Y′ and Y″ are not both absent;

each G is independently C or N, with the proviso that no more than two Gs are N; and

each R 5 , R 6 , R 7 , and R 8 is independently H, Cl, Br, C 1-3 alkyl, NO 2 , CN, NH 2 , O(C 1-3 alkyl), or (OCH 2 CH 2 ) 1-2 O(C 1-3 alkyl);

or where a R 5 , R 6 , R 7 , or R 8 is attached to a G that is N, such R 5 , R 6 , R 7 , or R 8 is absent;

and

wherein, in formula Ib,

the dotted lines indicate the optional presence of a C1-C2, C2-C3, or C2-R 10 double bond;

R 9 is absent if a C1-C2, C2-C3, or C2-R 10 double bond is present and otherwise is H;

R 10 is H, Cl, Br, ═CH 2 , ═CH(C 1-5 alkyl), C 1-3 alkyl, NO 2 , CN, or NH 2 ;

A′ is

R 50 is H, Cl, Br, C 1-3 alkyl, NO 2 , CN, NH 2 , O(C 1-3 alkyl), or (OCH 2 CH 2 ) 1-2 O(C 1-3 alkyl);

R 51 is H, Cl, Br, C 1-3 alkyl, NO 2 , CN, NH 2 ;

T is a self-immolating group;

t is 0 or 1;

AA a and each AA b are independently selected from the group consisting of alanine, β-alanine, γ-aminobutyric acid, arginine, asparagine, aspartic acid, γ-carboxyglutamic acid, citrulline, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, norleucine, norvaline, ornithine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine;

p is 1, 2, 3, or 4;

q is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10;

r is 1, 2, 3, 4, or 5; and

s is 0 or 1.

13. A conjugate according to claim 12 , wherein R 60 is IIIa.

14. A conjugate according to claim 12 , wherein R 60 is IIIa′.

15. A conjugate according to claim 12 , wherein R 60 is IIIa″.

16. A pharmaceutical formulation comprising a conjugate according to claim 12 and a pharmaceutically acceptable excipient.

17. A method of treating a cancer in a subject suffering from such cancer, comprising administering to the subject a therapeutically effective amount of a conjugate according to claim 12 , wherein the cancer is lung, gastric, or ovarian cancer.