Stable heterodimeric antibody design with mutations in the Fc domain
The provided scaffolds have heavy chains that are asymmetric in the various domains (e.g. CH2 and CH3) to accomplish selectivity between the various Fc receptors involved in modulating effector function, beyond those achievable with a natural homodimeric (symmetric) Fc molecule, and increased stability and purity of the resulting variant Fc heterodimers. These novel molecules comprise complexes of heterogeneous components designed to alter the natural way antibodies behave and that find use in therapeutics.
1. An isolated heteromultimer comprising a heterodimer Fc region, the heterodimer Fc region comprising a variant CH3 domain comprising a first CH3 domain polypeptide and a second CH3 domain polypeptide, each of the first and second CH3 domain polypeptides comprising amino acid modifications that promote formation of the heterodimer Fc region, the heterodimer Fc region having a purity greater than 90% and the variant CH3 domain having a melting temperature (Tm) of 70° C. or greater,
wherein the heterodimer Fc region is an IgG Fc region, and wherein:
(a) the first CH3 domain polypeptide comprises the amino acid modifications L351Y and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications E357L, T366A, K409F and T411N (variant AZ15)
(b) the first CH3 domain polypeptide comprises the amino acid modification Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A and K409F (variant AZ63),
(c) the first CH3 domain polypeptide comprises the amino acid modifications L351Y and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A and K409F (variant AZ64),
(d) the first CH3 domain polypeptide comprises the amino acid modifications L351F and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A and K409F (variant AZ65),
(e) the first CH3 domain polypeptide comprises the amino acid modification Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366S and K409F (variant AZ66),
(f) the first CH3 domain polypeptide comprises the amino acid modification Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366C and K409F (variant AZ67),
(g) the first CH3 domain polypeptide comprises the amino acid modification Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366L and K409F (variant AZ68),
(h) the first CH3 domain polypeptide comprises the amino acid modification Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366M and K409F (variant AZ69),
(i) the first CH3 domain polypeptide comprises the amino acid modifications L351Y and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366V and K409F (variant AZ70),
(j) the first CH3 domain polypeptide comprises the amino acid modifications L351I, T366S, L368F and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A and K409F (variant AZ71),
(k) the first CH3 domain polypeptide comprises the amino acid modifications D399W and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A and K409F (variant AZ72),
(l) the first CH3 domain polypeptide comprises the amino acid modifications D399W, S400D and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A and K409F (variant AZ73),
(m) the first CH3 domain polypeptide comprises the amino acid modifications D399W, S400E and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A and K409F (variant AZ74),
(n) the first CH3 domain polypeptide comprises the amino acid modifications D399W, S400D and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A, K409F and T411R (variant AZ75),
(o) the first CH3 domain polypeptide comprises the amino acid modifications G371D, D399W and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A, K409F and T411R (variant AZ76),
(p) the first CH3 domain polypeptide comprises the amino acid modifications K370Q, G371D, D399W and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A, K409F and T411R (variant AZ77),
(q) the first CH3 domain polypeptide comprises the amino acid modifications D399Y, S400D and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A, N390R and K409F (variant AZ78),
(r) the first CH3 domain polypeptide comprises the amino acid modification Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications Q362R, T366A, K409F and T411K (variant AZ79),
(s) the first CH3 domain polypeptide comprises the amino acid modification Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications Q362K, T366A, K409F and T411R (variant AZ81),
(t) the first CH3 domain polypeptide comprises the amino acid modifications S400E and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A, N390K, K392R, K409F and T411R (variant AZ82),
(u) the first CH3 domain polypeptide comprises the amino acid modifications S400E and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A, N390K, K392R, K409F and T411K (variant AZ83),
(v) the first CH3 domain polypeptide comprises the amino acid modifications S400D and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A, N390K, K409F and T411R (variant AZ84),
(w) the first CH3 domain polypeptide comprises the amino acid modifications D399R and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A, K392L, K409F and T411D (variant AZ85),
(x) the first CH3 domain polypeptide comprises the amino acid modifications D399R and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A, K392L, K409F and T411E (variant AZ86),
(y) the first CH3 domain polypeptide comprises the amino acid modifications D399K and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A, K392L, K409F and T411D (variant AZ87),
(z) the first CH3 domain polypeptide comprises the amino acid modifications D399K and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A, K392L, K409F and T411E (variant AZ88),
(aa) the first CH3 domain polypeptide comprises the amino acid modifications D399R and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A, K392M, K409F and T411E (variant AZ89),
(bb) the first CH3 domain polypeptide comprises the amino acid modifications D399R, F405V and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A, K392F, K409F and T411D (variant AZ91),
(cc) the first CH3 domain polypeptide comprises the amino acid modifications D399R, S400E and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A, K409F and T411E (variant AZ92),
(dd) the first CH3 domain polypeptide comprises the amino acid modifications D399R, S400D and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A, K409F and T411E (variant AZ93),
(ee) the first CH3 domain polypeptide comprises the amino acid modifications D399R, S400R and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A, K392E, K409F and T411E (variant AZ94),
(ff) the first CH3 domain polypeptide comprises the amino acid modifications D399R, S400R and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366A, K392E, K409F and T411D (variant AZ95),
(gg) the first CH3 domain polypeptide comprises the amino acid modification Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications S364Y, T366A, K409F and T411R (variant AZ98),
(hh) the first CH3 domain polypeptide comprises the amino acid modifications L351Y, L368S and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366V and K409W (variant AZ100), or
(ii) the first CH3 domain polypeptide comprises the amino acid modifications L351Y and Y407A, and the second CH3 domain polypeptide comprises the amino acid modifications T366V and K409W (variant AZ101),
and wherein the numbering of amino acid residues is according to the EU index as set forth in Kabat.
2. The isolated heteromultimer according to claim 1 , wherein the heterodimer Fc region is a human IgG Fc region.
3. The isolated heteromultimer according to claim 2 , wherein the IgG Fc region is an IgG1 Fc region.
4. The isolated heteromultimer according to claim 1 , wherein the heteromultimer is an antibody.
5. The isolated heteromultimer according to claim 4 , wherein the heteromultimer is a bispecific or multispecific antibody.
6. The isolated heteromultimer according to claim 4 , wherein the antibody binds at least one cancer antigen.
7. The isolated heteromultimer according to claim 4 , wherein the antibody is conjugated to a therapeutic agent.
8. A composition comprising the isolated heteromultimer of claim 1 , and a pharmaceutically acceptable carrier.