TIM-3 targeted heterodimeric fusion proteins containing IL-15/IL-15RA Fc-fusion proteins and TIM-3 antigen binding domains
The present invention is directed to novel targeted heterodimeric fusion proteins comprising an IL-15/IL-15Rα Fc-fusion protein and a TIM-3 antibody fragment-Fc fusion protein.
1. A heterodimeric fusion protein comprising:
a) a first monomer comprising, from N- to C-terminal:
i) an IL-15Rα(sushi) domain;
ii) a first domain linker;
iii) an IL-15 variant;
iv) a hinge; and
v) a first variant Fc domain comprising CH2-CH3; and
b) a second monomer comprising, from N- to C-terminal, VH-CH1-hinge-CH2-CH3, wherein the CH2-CH3 is a second variant Fc domain; and
c) a third monomer comprising a VL-CL,
wherein the VH and VL are a variable heavy domain and a variable light domain, respectively, that form a human T-cell immunoglobulin and mucin-domain containing-3 (TIM-3) antigen binding domain,
wherein the first variant Fc domain comprises skew variants L368D/K370S and the second variant Fc domain comprises skew variants S364K/E357Q,
wherein the first and second variant Fc domains each comprise FcKO variants E233P/L234V/L235A/G236del/S267K,
wherein the first variant Fc domain comprises pI variants Q295E/N384D/Q418E/N421D, and wherein numbering is according to EU numbering, and
wherein the first variant Fc domain and the second variant Fc domain are each a variant of a human IgG Fc domain.
2. A heterodimeric fusion protein according to claim 1 , wherein the hinge of the first monomer comprises amino acid substitution C220S, and wherein numbering is according to EU numbering.
3. A heterodimeric fusion protein according to claim 1 , wherein the first and second variant Fc domains each further comprise half-life extension variants M428L/N434S.
4. A heterodimeric fusion protein according to claim 1 , wherein the IL-15 variant is a variant human IL-15 comprising amino acid substitution(s) selected from the group consisting of N1D, N4D, D8N, D30N, D61N, E64Q, N65D, Q108E, N4D/N65D, D30N/N65D, and D30N/E64Q/N65D.
5. A heterodimeric fusion protein according to claim 4 , wherein the IL-15 variant comprises amino acid substitutions N4D/N65D, D30N/N65D, or D30N/E64Q/N65D.
6. A heterodimeric fusion protein according to claim 1 , wherein the variable heavy domain and the variable light domain of the TIM-3 antigen binding domain is selected from the group consisting of SEQ ID NOS:56 and 60, SEQ ID NOS:64 and 68, SEQ ID NOS:72 and 76, SEQ ID NOS:80 and 84, SEQ ID NOS:88 and 92, SEQ ID NOS:96 and 100, SEQ ID NOS:104 and 108, SEQ ID NOS:112 and 116, SEQ ID NOS:120 and 124, SEQ ID NOS:128 and 132, SEQ ID NOS:136 and 140, SEQ ID NOS:144 and 148, SEQ ID NOS:152 and 156, SEQ ID NOS:160 and 164, SEQ ID NOS:168 and 172, SEQ ID NOS:176 and 180, and SEQ ID NOS:184 and 188.
7. A heterodimeric fusion protein according to claim 6 , wherein the variable heavy domain and the variable light domain of the TIM-3 antigen binding domain is 3H3_H1_L2.1 (SEQ ID NOS:72 and 76).
8. A heterodimeric fusion protein according to claim 1 , wherein the heterodimeric fusion protein is selected from the group consisting of: XENP27974 (SEQ ID NOS:241, 242 and 243), XENP27979 (SEQ ID NOS:246, 247 and 248), XENC1000 (SEQ ID NOS:267, 268 and 269), XENC1001 (SEQ ID NOS:270, 271 and 272), XENC1002 (SEQ ID NOS:273, 274 and 275), and XENC1003 (SEQ ID NOS:276, 277 and 278).
9. A nucleic acid composition comprising:
a) a first nucleic acid encoding said first monomer of claim 1 ;
b) a second nucleic acid encoding said second monomer of claim 1 ; and
c) a third nucleic acid encoding said third monomer of claim 1 ; respectively.
10. An expression vector composition comprising:
a) a first expression vector comprising said first nucleic acid of claim 9 ;
b) a second expression vector comprising said second nucleic acid of claim 9 ;
c) a third expression vector comprising said third nucleic acid of claim 9 .
11. A host cell comprising the expression vector composition according to claim 10 .
12. A method of making a heterodimeric fusion protein comprising culturing the host cell of claim 11 and recovering the heterodimeric fusion protein from the cell culture.