Granted Patent B2

US 12,428,461 · App. 17/997,363 · Granted Sep 30, 2025

Inventors: Nina Gunnarsson (Hellerup, DK); Matiss Maleckis (Hellerup, DK); David B. Rosen (Palo Alto, CA)

Assignee: ASCENDIS PHARMA ONCOLOGY DIVISION A/S

C07K14/55A61K47/542A61K47/60A61P35/00

Abstract

Claims (109)

1. A method of treating, controlling, or delaying a cancer, which can be treated with IL-2 in a mammalian patient in need thereof, comprising the step of administering to the patient in need thereof a therapeutically effective amount of a conjugate or a pharmaceutically acceptable salt thereof of formula (Ia) or (Ib)

wherein

-D comprises an IL-2 protein of formula (I)

Ala-SEQ A-Cys*-SEQ B (I),

wherein

SEQ A has at least 94% sequence identity to SEQ ID NO:1;

SEQ B has at least 94% sequence identity to SEQ ID NO:2;

Ala is an alanine residue; and

Cys* is a cysteine residue;

-L 1 - is a linker moiety covalently and reversibly attached to-D;

-L 2 - is a chemical bond or is a spacer moiety;

Z is a polymeric moiety or a substituted fatty acid moiety;

x is an integer selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15 or 16; and

y is an integer selected from the group consisting of 2, 3, 4 and 5.

2. The method of claim 1 , wherein SEQ A is selected from the group consisting of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO: 7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO: 10 and SEQ ID NO:11.

3. The method of claim 1 , wherein SEQ A has the sequence of SEQ ID NO:1.

4. The method of claim 1 , wherein SEQ B has the sequence of SEQ ID NO:2.

5. The method of claim 1 , wherein SEQ B has the sequence of SEQ ID NO:12.

6. The method of claim 1 , wherein the IL-2 protein has the sequence of SEQ ID NO: 14.

7. The method of claim 1 , wherein the IL-2 protein comprises a moiety M mod conjugated to the cysteine marked with the asterisk in the IL-2 protein of formula (I).

8. The method of claim 7 , wherein M mod is a polymeric moiety.

9. The method of claim 7 , wherein M mod is of formula (A-1)

wherein

-FG- is a linkage;

-SP- is a spacer moiety; and

-POL is a polymer.

10. The method of claim 7 , wherein M mod has a molecular weight ranging from 0.5 kDa to 120 kDa.

11. The method of claim 7 , wherein M mod is a PEG-based polymer.

12. The method of claim 7 , wherein attachment of M mod to the IL-2 protein is via a stable linkage.

13. The method of claim 7 , wherein M mod is of formula (A-1a)

wherein

the dashed line marked with the asterisk indicates attachment to the sulfur of the cysteine marked with the asterisk in formula (I);

b1 is an integer selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 and 20;

b2 is an integer selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 and 20; and

b3 is an integer ranging from 12 to 22700.

14. The method of claim 7 , wherein M mod is of formula (A-1d)

wherein

the dashed line marked with the asterisk indicates attachment to the sulfur of the cysteine marked with the asterisk in formula (I);

b1 is an integer selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 and 20;

b2 is an integer selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 and 20; and

b3 is an integer ranging from 12 to 22700.

15. The method of claim 7 , wherein M mod is of formula (A-1e)

wherein

the dashed line marked with the asterisk indicates attachment to the sulfur of the cysteine marked with the asterisk in formula (I);

b1 is an integer selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 and 20;

b2 is an integer selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 and 20; and

b3 is an integer ranging from 12 to 22700.

16. The method of claim 7 , wherein in a plurality of conjugates M mod comprises a mixture of formula (A-1d) and formula (A-1e).

17. The method of claim 1 , wherein the conjugate is a biased IL-2.

18. The method of claim 1 , wherein —Z is a PEG-based polymeric moiety.

19. The method of claim 1 , wherein —Z comprises a moiety of formula (A)

wherein

—BP1<, —BP2<, —BP3< are independently of each other selected from the group consisting of —N< and —C(R 8 )<;

R 8 is selected from the group consisting of H, C 1-6 alkyl, C 2-6 alkenyl and C 2-6 alkynyl;

—P1, —P2, —P3, —P4 are independently of each other a PEG-based chain comprising at least 40% PEG and having a molecular weight ranging from 3 to 40 kDa;

—C 1 —, —C 2 — are independently of each other selected from the group consisting of C 1-50 alkyl, C 2-50 alkenyl, and C 2-50 alkynyl; wherein C 1-50 alkyl, C 2-50 alkenyl, and C 2-50 alkynyl are optionally substituted with one or more R 9 , which are the same or different and wherein C 1-50 alkyl, C 2-50 alkenyl, and C 2-50 alkynyl are optionally interrupted by one or more groups selected from the group consisting of -T-, —C(O)O—, —O—, —C(O)—, —C(O)N(R 10 )—, —S(O) 2 N(R 10 )—, —S(O)N(R 10 )—, —S(O) 2 —, —S(O)—, —N(R 10 ) S(O) 2 N(R 10a )—, —S—, —N(R 10 )—, —OC(OR 10 ) (R 10a )—, —N(R 10 )C (O)N(R 10a )—, and —OC(O)N(R 10 )—;

each T is independently selected from the group consisting of phenyl, naphthyl, indenyl, indanyl, tetralinyl, C 3-10 cycloalkyl, 3- to 10-membered heterocyclyl, 8- to 11-membered heterobicyclyl, 8- to 30-membered carbopolycyclyl, and 8- to 30-membered heteropolycyclyl; wherein each T is independently optionally substituted with one or more R 9 , which are the same or different;

each R 9 is independently selected from the group consisting of halogen, —CN, oxo (═O), —COOR 11 , —OR 11 , —C(O) R 11 , —C(O)N(R 11 R 11a ), —S(O) 2 N(R 11 R 11a ), —S(O)N(R 11 R 11a ), —S(O) 2 R 11 , —S(O)R 11 , —N(R 11 )S(O) 2 N(R 11a R 11b ), —SR 11 , —N(R 11 R 11a ), —NO 2 , —OC(O)R 11 , —N(R 11 )C(O)R 11a , —N(R 11 )S(O) 2 R 11a , —N(R 11 )S(O)R 11a , —N(R 11 )C(O)OR 11a , —N(R 11 )C(O)N(R 11a R 11b ), —OC(O)N(R 11 R 11a ), and C 1-6 alkyl; wherein C 1-6 alkyl is optionally substituted with one or more halogen, which are the same or different; and

each R 10 , R 10a , R 11 , R 11a and R 11b is independently selected from the group consisting of —H, and C 1-6 alkyl, wherein C 1-6 alkyl is optionally substituted with one or more halogen, which are the same or different.

20. The method of claim 19 , wherein C 1 and C 2 are of formula (A-a)

wherein

the dashed line marked with the asterisk indicates attachment to BP 1 ;

the unmarked dashed line indicates attachment to BP 2 or BP 3 , respectively;

q1 is selected from the group consisting of 1, 2, 3, 4, 5, 6, 7 and 8;

q2 is selected from the group consisting of 1, 2, 3, 4, and 5;

q3 is selected from the group consisting of 1, 2, 3, 4, 5, 6, 7 and 8; and

q4 is selected from the group consisting of 1, 2 and 3.

21. The method of claim 19 , wherein P 1 , P 2 , P 3 and P 4 are independently of each other of formula (A-b)

wherein

the dashed line indicates attachment to the remainder of —Z;

m is 0 or 1;

p is an integer ranging from 70 to 900; and

q is selected from the group consisting of 1, 2, 3, 4, 5, and 6.

22. The method of claim 1 , wherein -L 1 - is of formula (IX-a):

wherein

the dashed line marked with the asterisk indicates attachment to a nitrogen of-D and the unmarked dashed line indicates attachment to -L 2 -Z;

n is 0, 1, 2, 3, or 4;

═Y, is selected from the group consisting of ═O and ═S;

—Y 2 — is selected from the group consisting of —O— and —S—;

—Y 3 — is selected from the group consisting of —O— and —S—;

—Y 4 — is selected from the group consisting of —O—, —NR 5 — and —C(R 6 R 6a )—;

═Y 5 is selected from the group consisting of ═O and ═S;

—R 3 , —R 5 , —R 6 , —R 6a are independently of each other selected from the group consisting of —H, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, 2-methylbutyl, 2,2-dimethylpropyl, n-hexyl, 2-methylpentyl, 3-methylpentyl, 2,2-dimethylbutyl, 2,3-dimethylbutyl and 3,3-dimethylpropyl;

—R 4 is selected from the group consisting of methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, 2-methylbutyl, 2,2-dimethylpropyl, n-hexyl, 2-methylpentyl, 3-methylpentyl, 2,2-dimethylbutyl, 2,3-dimethylbutyl and 3,3-dimethylpropyl;

—W— is selected from the group consisting of C 1-20 alkyl optionally interrupted by one or more groups selected from the group consisting of C 3-10 cycloalkyl, 8- to 30-membered carbopolycyclyl, 3- to 10-membered heterocyclyl, —C(O)—, —C(O)N(R 7 )—, —O—, —S— and —N(R 7 )—;

—Nu is a nucleophile selected from the group consisting of —N(R 7 R 7a ), —N(R 7 OH), —N(R 7 )—N(R 7a R 7b ), —S(R 7 ), —COOH,

—Ar— is selected from the group consisting of

wherein

dashed lines indicate attachment to the remainder of -L 1 -,

—Z 1 — is selected from the group consisting of —O—, —S— and —N(R 7 )—, and

—Z 2 — is —N(R 7 )—; and

—R 7 , —R 7a , —R 7b are independently of each other selected from the group consisting of —H, C 1-6 alkyl, C 2-6 alkenyl and C 2-6 alkynyl;

wherein -L 1 - is optionally further substituted.

23. The method of claim 1 , wherein -L 1 - is of formula (IX-c)

wherein

the dashed line marked with the asterisk indicates attachment to a nitrogen of -D;

the unmarked dashed line indicates attachment to -L 2 -Z; and

s1 is an integer selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9 and 10.

24. The method of claim 1 , wherein -L 2 — is selected from the group consisting of -T-, —C(O)O—, —O—, —C(O)—, —C(O)N(R y1 )—, —S(O) 2 N(R y1 )—, —S(O)N(R y1 )—, —S(O) 2 —, —S(O)—, —N(R y1 )S(O) 2 N(R y1a )—, —S—, —N(R y1 )—, —OC(OR y1 )(R y1a )—, —N(R y1 )C(O)N(R y1a )—, —OC(O)N(R y1 )—, C 1-50 alkyl, C 2-50 alkenyl, and C 2-50 alkynyl; wherein -T-, C 1-50 alkyl, C 2-50 alkenyl, and C 2-50 alkynyl are optionally substituted with one or more —R y2 , which are the same or different and wherein C 1-50 alkyl, C 2-50 alkenyl, and C 2-50 alkynyl are optionally interrupted by one or more groups selected from the group consisting of -T-, —C(O)O—, —O—, —C(O)—, —C(O)N(R y3 )—, —S(O) 2 N(R y3 )—, —S(O)N(R y3 )—, —S(O) 2 —, —S(O)—, —N(R y3 )S(O) 2 N(R y3 a)-, —S—, —N(R y3 )—, —OC (OR y3 ) (R y3a )—, —N(R y3 )C(O)N(R y3a )—, and —OC(O)N(R y3 )—;

—R y1 and —R y1a are independently of each other selected from the group consisting of —H, -T, C 1-50 alkyl, C 2-50 alkenyl, and C 2-50 alkynyl; wherein -T, C 1-50 alkyl, C 2-50 alkenyl, and C 2-50 alkynyl are optionally substituted with one or more —R y2 , which are the same or different, and wherein C 1-50 alkyl, C 2-50 alkenyl, and C 2-50 alkynyl are optionally interrupted by one or more groups selected from the group consisting of -T-, —C(O)O—, —O—, —C(O)—, —C(O)N(R y4 )—, —S(O) 2 N(R y4 )—, —S(O)N(R y4 )—, —S(O) 2 —, —S(O)—, —N(R y4 )S(O) 2 N(R y4a )—, —S—, —N(R y4 )—, —OC (OR y4 ) (R y4a )—, —N(R y4 )C(O)N(R y4a )—, and —OC(O)N(R y4 )—;

each T is independently selected from the group consisting of phenyl, naphthyl, indenyl, indanyl, tetralinyl, C 3-10 cycloalkyl, 3- to 10-membered heterocyclyl, 8- to 11-membered heterobicyclyl, 8- to 30-membered carbopolycyclyl, and 8- to 30-membered heteropolycyclyl; wherein each T is independently optionally substituted with one or more -R y2 , which are the same or different;

each —R y2 is independently selected from the group consisting of halogen, —CN, oxo (═O), —COOR y5 , —OR y5 , —C(O)R y5 , —C(O)N(R y5 R y5a ), —S(O) 2 N(R y5 R y5a ), —S(O)N(R y5 R y5a ), —S(O) 2 R y5 , —S(O) R y5 , —N(R y5 )S(O) 2 N(R y5a R y5b ), —SR y5 , —N(R y5 R y5a ), —NO 2 , —OC(O)R y5 , —N(R y5 )C(O)R y5a , —N(R y5 )S(O) 2 R y5a , —N(R y5 ) S(O) R y5a , —N(R y5 )C(O)OR y5a , —N(R y5 )C(O)N(R y5a R y5b ), —OC(O)N(R y5 R y5a ), and C 1-6 alkyl; wherein C 1-6 alkyl is optionally substituted with one or more halogen, which are the same or different; and

each —R y3 , —R y3a , —R y4 , —R y4a , —R y5 , —R y5a and —R y5b is independently selected from the group consisting of —H, and C 1-6 alkyl, wherein C 1-6 alkyl is optionally substituted with one or more halogen, which are the same or different.

25. The method of claim 1 , wherein -L 2 - is of formula (IX-e)

wherein

the dashed line marked with the asterisk indicates attachment to -L 1 -;

the unmarked dashed line indicates attachment to —Z; and

s2 is an integer selected from the group consisting of 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 and 20.

26. The method of claim 1 , wherein the mammalian patient is human.

Assignments (1)

Priority Claims (5)

Continuity (2)

References Cited (121)