Anti-mutant calreticulin (CALR) antibody-drug conjugates and uses thereof

Abstract

Anti-mutant calreticulin (mutCALR) antibody-drug conjugates comprising pyrrolobenzodiazepine dimers are disclosed. Also disclosed are related kits, and pharmaceutical compositions. Methods of treating myeloproliferative neoplasms with the anti-mutCALR antibody-drug conjugates are also disclosed.

Description

[0001] ANTI-MUTANT CALRETICULIN (CALR) ANTIBODY-DRUG CONJUGATES AND

[0002] USES THEREOF

[0003] CROSS-REFERENCE TO RELATED APPLICATIONS

[0004] This application claims the benefit of priority of U.S. Provisional Application No. 63 / 546,869, filed on November 1, 2023, the contents of which is incorporated by reference in its entirety herein.

[0005] SEQUENCE LISTING

[0006] The instant application contains a Sequence Listing which has been submitted electronically in XML format and is hereby incorporated by reference in its entirety. Said XML copy, created on October 29, 2024, is named 20443-0838W01_SL.xml and is 224,106 bytes in size.

[0007] BACKGROUND

[0008] Calreticulin (CALR) is a highly conserved chaperone protein that resides primarily in the endoplasmic reticulum and is involved in a variety of cellular processes including protein folding, calcium homeostasis, cell adhesion, and integrin signaling. CALR is also found in the nucleus, suggesting that it may have a role in transcription regulation. Mutations in the gene for CALR have been identified in patients with myeloproliferative neoplasms.

[0009] SUMMARY

[0010] The present disclosure is based, at least in part, on the development of antibody-drug conjugates that bind to mutant CALR (‘‘mutCALR”) on the cell surface, are internalized by the cell, and exhibit anti-proliferative effects.

[0011] Accordingly, aspects of the present application provide an antibody-drug conjugate comprising: an antibody that binds to human mutant calreticulin (CALR); and a pyrrolobenzodiazepine dimer, wherein the antibody comprises a heavy chain variable region (VH) comprising a VH CDR1; a VH CDR2; and a VH CDR3; wherein: the VH CDR1 comprises the amino acid sequence ELSMQ (SEQ ID NO:1);

[0012] 1

[0013] SUBSTITUTE SHEET (RULE 26) the VH CDR2 comprises the amino acid sequence GFDPDDX101ETMYAEX102X103QG (SEQ ID NO: 102); wherein X101 is D or G; wherein X102 is K or R; and wherein X103 is F or L; the VH CDR3 is SPGYDFFDY (SEQ ID NO: 18); wherein the antibody comprises a light chain variable region (VL) comprising a VL CDR1, a VL CDR2, and a VL CDR3, wherein: the VL CDR1 comprises the amino acid sequence GGX104X105X106GX107X108X109VX110 (SEQ ID NO: 103), wherein X104 is N, D, or S; wherein X105 is Y, N, or D; wherein X106 is I or T; wherein X107 is S, D, I, R, or T; wherein Xios is K, E, or I; wherein X109 is S, I, R, G, N, or A; and wherein Xno is H, F, or N; the VL CDR2 comprises the amino acid sequence DDX111DRPX112 (SEQ ID NO: 104), wherein X111 is G, S, or R; and wherein X112 is S or L; and the VL CDR3 comprises the amino acid sequence QVWDX113X114X115DX116X117X118 (SEQ ID NO: 105), wherein X113 is S or A; wherein X114 is I or S; wherein X115 is S, I, or N; wherein X116 is H, L, or Q; wherein X117 is V or L; and wherein Xns is V or I or the VL CDR1 comprises the amino acid sequence TGTSSDVGGYNYVS (SEQ ID NOTO); the VL CDR2 comprises the amino acid sequence X119VSX120RPS (SEQ ID NO: 106); wherein X119 is E or D; and wherein X120 is N or K; and the VL CDR3 comprises the amino acid sequence QVWDSSNDLLI (SEQ ID NO:71).

[0014] In some embodiments, the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of any one of SEQ ID NOs: 7-10 and 92-95; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of any one of SEQ ID NOs:26-41 and 118; the VL CDR2 comprises the amino acid sequence of any one of SEQ ID NOs:53-58; and the VL CDR3 comprises the amino acid sequence of any one of SEQ ID NOs:69-75.

[0015] In some embodiments, the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:26; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:53; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:69;

[0016] 2

[0017] SUBSTITUTE SHEET (RULE 26) the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:27; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:70; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 8; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:29; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:72; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:29; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ

[0018] 3

[0019] SUBSTITUTE SHEET (RULE 26) ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:73; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:56; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:31; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:57; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 9; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:32; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:33; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:73; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:34; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:56; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino

[0020] 4

[0021] SUBSTITUTE SHEET (RULE 26) acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:27; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:35; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:36; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:72; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:58; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:37; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:38; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:32; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71;

[0022] 5

[0023] SUBSTITUTE SHEET (RULE 26) the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:39; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:40; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 10; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:41; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:70; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:40; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:56; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ

[0024] 6

[0025] SUBSTITUTE SHEET (RULE 26) ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:74; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:75; the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO: 118; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 92; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 93; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 94; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 95; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 92; the VH CDR3 comprises the amino

[0026] 7

[0027] SUBSTITUTE SHEET (RULE 26) acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 93; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 94; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; or the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 95; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71.

[0028] In some embodiments, the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO: 30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71.

[0029] In some embodiments, the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:95; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO: 30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71.

[0030] In some embodiments, the VH is at least 80% identical to the amino acid sequence of any one of SEQ ID NOs: 165-174 and 188-208; and the VL is at least 80% identical to the amino acid sequence of any one of SEQ ID NOs:264-292 and 315-316.

[0031] In some embodiments, the VH comprises the amino acid sequence of any one of SEQ ID NOs:165-174 and 188-208; and the VL comprises the amino acid sequence of any one of SEQ ID NOs:264-292 and 315-316.

[0032] 8

[0033] SUBSTITUTE SHEET (RULE 26) In some embodiments, the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:264; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:265; the VH comprises the amino acid sequence of SEQ ID NO: 166 and the VL comprises the amino acid sequence of SEQ ID NO:266; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:266; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:267; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:269; the VH comprises the amino acid sequence of SEQ ID NO: 167 and the VL comprises the amino acid sequence of SEQ ID NO:270; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:271; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:272; the VH comprises the amino acid sequence of SEQ ID NO: 168 and the VL comprises the amino acid sequence of SEQ ID NO:273; the VH comprises the amino acid sequence of SEQ ID NO: 169 and the VL comprises the amino acid sequence of SEQ ID NO:274; the VH comprises the amino acid sequence of SEQ ID NO: 170 and the VL comprises the amino acid sequence of SEQ ID NO:275; the VH comprises the amino acid sequence of SEQ ID NO: 171 and the VL comprises the amino acid sequence of SEQ ID NO:276; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:277; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:278; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:279;

[0034] 9

[0035] SUBSTITUTE SHEET (RULE 26) the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:280; the VH comprises the amino acid sequence of SEQ ID NO: 172 and the VL comprises the amino acid sequence of SEQ ID NO:281; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:282; the VH comprises the amino acid sequence of SEQ ID NO: 173 and the VL comprises the amino acid sequence of SEQ ID NO:283; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:284; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:285; the VH comprises the amino acid sequence of SEQ ID NO: 174 and the VL comprises the amino acid sequence of SEQ ID NO:286; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:287; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:288; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:289; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:290; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:291; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:292; the VH comprises the amino acid sequence of SEQ ID NO: 188 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 189 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 190 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 191 and the VL comprises the amino acid sequence of SEQ ID NO:315;

[0036] 10

[0037] SUBSTITUTE SHEET (RULE 26) the VH comprises the amino acid sequence of SEQ ID NO: 192 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 193 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 194 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 195 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 196 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 188 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 189 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 190 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 191 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 192 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 193 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 194 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 195 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 196 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 197 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO: 198 and the VL comprises the amino acid sequence of SEQ ID NO:268;

[0038] 11

[0039] SUBSTITUTE SHEET (RULE 26) the VH comprises the amino acid sequence of SEQ ID NO: 199 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID N0:200 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO:201 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO: 197 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 198 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 199 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID N0:200 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO:201 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 197 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 198 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 199 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID N0:200 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO:201 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 170 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO:202 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO:203 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO:204 and the VL comprises the amino acid sequence of SEQ ID NO:268;

[0040] 12

[0041] SUBSTITUTE SHEET (RULE 26) the VH comprises the amino acid sequence of SEQ ID NO:205 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO:206 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO:207 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO:208 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO: 170 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO:202 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO:203 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO:204 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO:205 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO:206 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO:207 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO:208 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 170 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO:202 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO:203 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO:204 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO:205 and the VL comprises the amino acid sequence of SEQ ID NO:316;

[0042] 13

[0043] SUBSTITUTE SHEET (RULE 26) the VH comprises the amino acid sequence of SEQ ID NO:206 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO:207 and the VL comprises the amino acid sequence of SEQ ID NO:316; or the VH comprises the amino acid sequence of SEQ ID NO:208 and the VL comprises the amino acid sequence of SEQ ID NO:316.

[0044] In some embodiments, the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:268.

[0045] In some embodiments, the VH comprises the amino acid sequence of SEQ ID NO: 196 and the VL comprises the amino acid sequence of SEQ ID NO:315.

[0046] In some embodiments, the antibody comprises a heavy chain comprising the amino acid sequence of any one of SEQ ID NOs: 119-128 and 142-164; and a light chain comprising the amino acid sequence of any one of SEQ ID NOs:209-237 and 260-261.

[0047] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:209; the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:210; the heavy chain comprises the amino acid sequence of SEQ ID NO: 120 and the light chain comprises the amino acid sequence of SEQ ID NO:211 ; the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:211 ; the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:212; the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:214; the heavy chain comprises the amino acid sequence of SEQ ID NO: 121 and the light chain comprises the amino acid sequence of SEQ ID NO:215; the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:216; the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:217;

[0048] 14

[0049] SUBSTITUTE SHEET (RULE 26) the heavy chain comprises the amino acid sequence of SEQ ID NO: 122 and the light chain comprises the amino acid sequence of SEQ ID NO:218; the heavy chain comprises the amino acid sequence of SEQ ID NO: 123 and the light chain comprises the amino acid sequence of SEQ ID NO:219; the heavy chain comprises the amino acid sequence of SEQ ID NO: 124 and the light chain comprises the amino acid sequence of SEQ ID NO:220; the heavy chain comprises the amino acid sequence of SEQ ID NO: 125 and the light chain comprises the amino acid sequence of SEQ ID NO:221; the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:222; the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:223; the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:224; the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:225; the heavy chain comprises the amino acid sequence of SEQ ID NO: 126 and the light chain comprises the amino acid sequence of SEQ ID NO:226; the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:227; the heavy chain comprises the amino acid sequence of SEQ ID NO: 127 and the light chain comprises the amino acid sequence of SEQ ID NO:228; the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:229; the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:230; the heavy chain comprises the amino acid sequence of SEQ ID NO: 128 and the light chain comprises the amino acid sequence of SEQ ID NO:231; the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:232; the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:233; the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:234;

[0050] 15

[0051] SUBSTITUTE SHEET (RULE 26) the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:235; the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:236; the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:237; the heavy chain comprises the amino acid sequence of SEQ ID NO: 142 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 143 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 144 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 145 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 146 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 147 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 148 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 149 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 150 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 142 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 143 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 144 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 145 and the light chain comprises the amino acid sequence of SEQ ID NO:260;

[0052] 16

[0053] SUBSTITUTE SHEET (RULE 26) the heavy chain comprises the amino acid sequence of SEQ ID NO: 146 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 147 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 148 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 149 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 150 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 142 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 143 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 144 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 145 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 146 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 147 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 148 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 149 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 150 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 152 and the light chain comprises the amino acid sequence of SEQ ID NO:213;

[0054] 17

[0055] SUBSTITUTE SHEET (RULE 26) the heavy chain comprises the amino acid sequence of SEQ ID NO: 153 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 154 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 155 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 156 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 152 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 153 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 154 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 155 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 156 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 152 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 153 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 154 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 155 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 156 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 157 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 158 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 159 and the light chain comprises the amino acid sequence of SEQ ID NO:213;

[0056] 18

[0057] SUBSTITUTE SHEET (RULE 26) the heavy chain comprises the amino acid sequence of SEQ ID NO: 160 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 161 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 162 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 163 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 164 and the light chain comprises the amino acid sequence of SEQ ID NO:213; the heavy chain comprises the amino acid sequence of SEQ ID NO: 157 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 158 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 159 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 160 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 161 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 162 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 163 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 164 and the light chain comprises the amino acid sequence of SEQ ID NO:260; the heavy chain comprises the amino acid sequence of SEQ ID NO: 157 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 158 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 159 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 160 and the light chain comprises the amino acid sequence of SEQ ID NO:261;

[0058] 19

[0059] SUBSTITUTE SHEET (RULE 26) the heavy chain comprises the amino acid sequence of SEQ ID NO: 161 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 162 and the light chain comprises the amino acid sequence of SEQ ID NO:261; the heavy chain comprises the amino acid sequence of SEQ ID NO: 163 and the light chain comprises the amino acid sequence of SEQ ID NO:261; or the heavy chain comprises the amino acid sequence of SEQ ID NO: 164 and the light chain comprises the amino acid sequence of SEQ ID NO:261.

[0060] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:213.

[0061] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 142 and the light chain comprises the amino acid sequence of SEQ ID NO:213.

[0062] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:260.

[0063] In some embodiments, the human mutant CALR is human Type 1 mutant CALR comprising the amino acid sequence of SEQ ID NO: 320. In some embodiments, the human mutant CALR is human Type 2 mutant CALR comprising the amino acid sequence of SEQ ID NO:321.

[0064] In some embodiments, the antibody is a human or humanized antibody. In some embodiments, the antibody is a full-length antibody. In some embodiments, the antibody is an IgGl, IgG2, IgG3 or IgG4 antibody. In some embodiments, the antibody is an IgGl antibody. In some embodiments, the antibody is a bispecific antibody, a biparatopic antibody, a single chain antibody (scFv), an Fab fragment, an F(ab’)2 fragment, an Fab’ fragment, an Fsc fragment, an Fv fragment, an scFv, an sc(Fv)2, or a diabody.

[0065] In some embodiments, the pyrrolobenzodiazepine dimer is selected from the group consisting of PBD-1, SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD- 1882.

[0066] In some embodiments, the pyrrolobenzodiazepine dimer is selected from the group consisting of SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

[0067] In some embodiments, the pyrrolobenzodiazepine dimer is linked to a linker to form a payload. In some embodiments, the linker is cleavable. In some embodiments, the linker comprises a maleimide linker. In some embodiments, the linker comprises a peptide. In some embodiments, the linker comprises a polyethylene glycol spacer.

[0068] 20

[0069] SUBSTITUTE SHEET (RULE 26) In some embodiments, the payload is selected from the group consisting of SG3400, SG3227, SG3249, SG3710, SG3259, SG3584, SG3203, SG3231, SG3451, and SGD-1910.

[0070] In some embodiments, the pyrrolobenzodiazepine dimer comprises a cap. In some embodiments, the cap is cleavable. In some embodiments, the cap comprises a glucuronide group or derivative thereof or a galactoside group or derivative thereof. In some embodiments, the payload is SG3600.

[0071] In some embodiments, the antibody-drug conjugate has a drug loading of from about one to about five pyrrolobenzodiazepine dimers per antibody. In some embodiments, the antibody-drug conjugate has a drug loading of two pyrrolobenzodiazepine dimers per antibody.

[0072] In some embodiments, the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO: 30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of PBD-1, SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

[0073] In some embodiments, the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO: 30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

[0074] In some embodiments, the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO: 30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71, and wherein the pyrrolobenzodiazepine dimer is PBD-1.

[0075] In some embodiments, the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO: 30; the VL CDR2 comprises the amino acid sequence of SEQ

[0076] 21

[0077] SUBSTITUTE SHEET (RULE 26) ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71, and wherein the pyrrolobenzodiazepine dimer is SG2000.

[0078] In some embodiments, the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO: 30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71, and wherein the pyrrolobenzodiazepine dimer is SG3199.

[0079] In some embodiments, the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:268, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of PBD-1, SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

[0080] In some embodiments, the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:268, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

[0081] In some embodiments, the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:268, and wherein the pyrrolobenzodiazepine dimer is PBD-1.

[0082] In some embodiments, the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:268, and wherein the pyrrolobenzodiazepine dimer is SG2000.

[0083] In some embodiments, the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:268, and wherein the pyrrolobenzodiazepine dimer is SG3199.

[0084] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:213, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of PBD-1, SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

[0085] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:213, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

[0086] 22

[0087] SUBSTITUTE SHEET (RULE 26) In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:213, and wherein the pyrrolobenzodiazepine dimer is PBD-1.

[0088] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:213, and wherein the pyrrolobenzodiazepine dimer is SG2000.

[0089] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 119 and the light chain comprises the amino acid sequence of SEQ ID NO:213, and wherein the pyrrolobenzodiazepine dimer is SG3199.

[0090] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 142 and the light chain comprises the amino acid sequence of SEQ ID NO:213, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of PBD-1, SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

[0091] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 142 and the light chain comprises the amino acid sequence of SEQ ID NO:213, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

[0092] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 142 and the light chain comprises the amino acid sequence of SEQ ID NO:213, and wherein the pyrrolobenzodiazepine dimer is PBD-1.

[0093] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 142 and the light chain comprises the amino acid sequence of SEQ ID NO:213, and wherein the pyrrolobenzodiazepine dimer is SG2000.

[0094] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 142 and the light chain comprises the amino acid sequence of SEQ ID NO:213, and wherein the pyrrolobenzodiazepine dimer is SG3199.

[0095] In some embodiments, the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:95; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO: 30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of PBD-1, SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

[0096] 23

[0097] SUBSTITUTE SHEET (RULE 26) In some embodiments, the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:95; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO: 30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

[0098] In some embodiments, the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:95; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO: 30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71, and wherein the pyrrolobenzodiazepine dimer is PBD-1.

[0099] In some embodiments, the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:95; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO: 30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71, and wherein the pyrrolobenzodiazepine dimer is SG2000.

[0100] In some embodiments, the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:95; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO: 30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71, and wherein the pyrrolobenzodiazepine dimer is SG3199.

[0101] In some embodiments, the VH comprises the amino acid sequence of SEQ ID NO: 196 and the VL comprises the amino acid sequence of SEQ ID NO:315, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of PBD-1, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

[0102] In some embodiments, the VH comprises the amino acid sequence of SEQ ID NO: 196 and the VL comprises the amino acid sequence of SEQ ID NO:315, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

[0103] 24

[0104] SUBSTITUTE SHEET (RULE 26) In some embodiments, the VH comprises the amino acid sequence of SEQ ID NO: 196 and the VL comprises the amino acid sequence of SEQ ID NO:315, and wherein the pyrrolobenzodiazepine dimer is PBD-1.

[0105] In some embodiments, the VH comprises the amino acid sequence of SEQ ID NO: 196 and the VL comprises the amino acid sequence of SEQ ID NO:315, and wherein the pyrrolobenzodiazepine dimer is SG2000.

[0106] In some embodiments, the VH comprises the amino acid sequence of SEQ ID NO: 196 and the VL comprises the amino acid sequence of SEQ ID NO:315, and wherein the pyrrolobenzodiazepine dimer is SG3199.

[0107] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:260, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of PBD-1, SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

[0108] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:260, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

[0109] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:260, and wherein the pyrrolobenzodiazepine dimer is PBD-1.

[0110] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:260, and wherein the pyrrolobenzodiazepine dimer is SG2000.

[0111] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:260, and wherein the pyrrolobenzodiazepine dimer is SG3199.

[0112] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:260, the linker is a group of formula L-l, and the pyrrolobenzodiazepine dimer is SG3199.

[0113] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:260, the linker is a group of formula L-2, and the pyrrolobenzodiazepine dimer is PBD-1.

[0114] 25

[0115] SUBSTITUTE SHEET (RULE 26) In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:260, the payload is PL-1.

[0116] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:260, and the payload is PL-2.

[0117] In some embodiments, the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:260, and the payload is PL-3.

[0118] Aspects of the present disclosure provide a pharmaceutical composition comprising an antibody-drug conjugate described herein, and a pharmaceutically acceptable carrier.

[0119] Aspects of the present disclosure provide a method of treating a myeloproliferative neoplasm in a human subject in need thereof, the method compnsing administering to the human subject an effective amount of an antibody-drug conjugate described herein or a pharmaceutical composition described herein.

[0120] In some embodiments, the myeloproliferative neoplasm is selected from the group consisting of chronic myelogenous leukemia, polycythemia vera, primary myelofibrosis, essential thrombocythemia, chronic neutrophilic leukemia, acute myelogenus leukemia, chronic eosinophilic leukemia, chronic myelomonocytic leukemia, myeloproliferative neoplasm and myelodysplastic syndrome, including myelodysplastic syndrome with refractory anaemia with ring sideroblasts, myelodysplastic syndrome with refractory anemia, and myelodysplastic syndrome with refractory anemia with excess blasts.

[0121] In some embodiments, methods described herein further comprise administering to the human subject an additional therapy selected from the group consisting of a Janus tyrosine kinase (JAK) inhibitor, a phosphoinositide 3-kinase (PI3K) inhibitor, a standard of care therapy, or a combination thereof.

[0122] In some embodiments, the JAK inhibitor is ruxolitinib and itaticinib. In some embodiments, the PI3K inhibitor is parsaclisib.

[0123] In some embodiments, the standard of care therapy is selected from the group consisting of IFN-alpha, hydroxyurea, thalidomide, lenalidomide, an androgen, an erythropoietin-stimulating agent, a chemotherapeutic agent, or a combination thereof.

[0124] Aspects of the present disclosure provide a kit comprising an antibody-drug conjugate described herein, and instructions for use in treating a myeloproliferative neoplasm in a

[0125] 26

[0126] SUBSTITUTE SHEET (RULE 26) human subject in need thereof, optionally with instructions for use in combination with an additional therapy.

[0127] BRIEF DESCRIPTION OF THE DRAWINGS

[0128] FIG. 1A includes a graph showing internalization of anti-mutCALR antibody (Clone 74) in 4F2 cells or MPL-6 cells in the absence or presence of dynasore or DMSO (vehicle control).

[0129] FIG. IB includes a graph showing internalization of anti-mutCALR antibody (Clone 55) in 4F2 cells or MPL-6 cells in the absence or presence of dynasore or DMSO (vehicle control).

[0130] FIG. 1C includes a graph showing internalization of anti-mutCALR antibody (Clone 6) in 4F2 cells or MPL-6 cells in the absence or presence of dynasore or DMSO (vehicle control).

[0131] FIG. ID includes a graph showing proliferation of 4F2 or MPL-6 cells in the presence of anti-mutCALR antibody Clone 74, Clone 55, or Clone 6. Proliferation was measured in the absence or presence of dynasore for each clone.

[0132] FIG. IE includes a graph showing internalization of the anti-mutCALR antibody B3 in 4F2 cells or MPL-6 cells in the absence or presence of dynasore or DMSO (vehicle control).

[0133] FIG. IF includes a graph showing internalization of a first isotype control antibody in 4F2 cells or MPL-6 cells in the absence or presence of dynasore or DMSO (vehicle control).

[0134] FIG. 1G includes a graph showing internalization of a second isotype control antibody in 4F2 cells or MPL-6 cells in the absence or presence of dynasore or DMSO (vehicle control).

[0135] FIG. 1H includes a graph showing proliferation of 4F2 or MPL-6 cells in the presence of the anti-mutCALR antibody B3 or isotype control antibodies. Proliferation was measured in the absence or presence of dynasore for each antibody.

[0136] FIG. 2A includes images showing time-dependent binding of fluorescently-labeled anti-mutCALR antibody (Clone 74) to Ba / F3-TPOR / CALRde152cells measured using high content imaging. Representative images show 20x magnification. Scale bar = 10 pm.

[0137] FIG. 2B includes a graph showing internalization of anti-mutCALR antibody (Clone 74) or an isotype control in Ba / F3-TPOR / CALRde152cells in the absence or presence of dynasore or DMSO (vehicle control).

[0138] 27

[0139] SUBSTITUTE SHEET (RULE 26) FIG. 2C includes a graph showing proliferation of Ba / F3-TPOR / CALRdeb2cells pretreated with dynasore or DMSO (vehicle control) in the presence of anti-mutCALR antibody (Clone 74) or an isotype control.

[0140] FIG. 2D includes a graph showing caspase induction in Ba / F3-TPOR / CALRde152cells pretreated with dynasore or DMSO (vehicle control) in the presence of anti-mutCALR antibody (Clone 74) or ab isotype control.

[0141] FIG. 2E includes a graph showing pSTAT5 inhibition in Ba / F3-TPOR / CALRde152cells pretreated with dynasore or DMSO (vehicle control) in the presence of anti-mutCALR antibody (Clone 74) or an isotype control. Inhibition of pSTAT5 was calculated after normalization to maximal (100%) inhibition achieved by the cell treatment with 2 pM of ruxolitinib and no inhibition (0%) in isotype-treated cells. Data show a representative of 3 independent experiments.

[0142] FIG. 2F includes an image showing that anti-mutCALR antibody (Clone 74) is internalized upon binding mutCALR on the cell surface. Arrow 1: cell membrane, Arrow 2: nucleus, Arrow 3: anti-mutCALR antibody.

[0143] FIG. 2G includes images showing that anti-mutCALR antibody (Clone 74) colocalizes with lysosome markers upon internalization. Light grey: LAMP-1, Dark grey: anti- mutCALR antibody.

[0144] FIG. 3A includes graphs from flow cytometry analysis of the surface expression of TPOR in the presence of anti-mutCALR antibody (Clone 74) or an isotype control. Graphs represent MFI mean ± SD of 3 (Ba / F3-TPOR) or 5 (Ba / F3-TPOR mutC ALRde152) independent experiments. Significance was determined using student’s t-test: ** p<0.01.

[0145] FIG. 3B includes an image from Western blot analysis of TPOR and pSTAT5 expression in the presence of anti-mutCALR antibody (Clone 74) or an isotype control. Data representative from 2 independent experiments.

[0146] FIG. 3C includes data from immunofluorescence analysis of internalization of TPOR / mutCALR complex upon anti-mutCALR antibody (Clone 74) binding. Areas showing both light grey (cell membrane) and dark grey (TPOR) were false colored for easier visualization of the pixels exhibiting colocalization.

[0147] FIG. 4A includes a graph showing in BAF3 / 4F2 cells internalization of anti- mutCALR antibody conjugated to PBD (Clone 6-PBD), unconjugated anti-mutCALR antibody (Clone 6), isotype control conjugated to PBD (Isotype-PBD), or unconjugated isotype control.

[0148] 28

[0149] SUBSTITUTE SHEET (RULE 26) FIG. 4B includes a graph showing caspase induction in BAF3 / 4F2 cells in the presence of anti-mutCALR antibody conjugated to PBD (Clone 6-PBD), unconjugated anti- mutCALR antibody (Clone 6), isotype control conjugated to PBD (Isotype-PBD), or unconjugated isotype control.

[0150] FIG. 4C includes a graph showing proliferation of BAF3 / 4F2 cells in the presence of anti-mutCALR antibody conjugated to PBD (Clone 6-PBD), unconjugated anti-mutCALR antibody (Clone 6), or isotype control conjugated to PBD (Isotype-PBD), or unconjugated isotype control.

[0151] FIG. 5A includes a graph showing viability of BAF3 / 4F2 cells in the presence of anti-mutCALR antibody conjugated to PBD (Clone 6-PBD), unconjugated anti-mutCALR antibody (Clone 6), or isotype control conjugated to PBD (Isotype-PBD).

[0152] FIG. 5B includes a graph showing viability of BAF3 / 4F2 cells in the presence of anti- mutCALR antibody conjugated to MMAE (Clone 6-MMAE), unconjugated anti-mutCALR antibody (Clone 6), or isotype control conjugated to MMAE (Isotype-MMAE).

[0153] FIG. 6A includes a graph showing viability of BAF3 / 3D3 cells in the presence of anti-mutCALR antibody conjugated to PBD (Clone 6-PBD), unconjugated anti-mutCALR antibody (Clone 6), or isotype control conjugated to PBD (Isotype-PBD).

[0154] FIG. 6B includes a graph showing viability of BAF3 / 3D3 cells in the presence of anti-mutCALR antibody conjugated to MMAE (Clone 6-MMAE), unconjugated anti- mutCALR antibody (Clone 6), or isotype control conjugated to MMAE (Isotype-MMAE).

[0155] FIG. 7A includes a graph showing viability of BAF3 / 4F2 cells treated with anti- mutCALR antibody conjugated to PBD (Clone 6-PBD), unconjugated anti-mutCALR antibody (Clone 6), or isotype control conjugated to PBD (Isotype-PBD) in the presence of 10 ng / mL human TPO.

[0156] FIG. 7B includes a graph showing viability of BAF3 / 4F2 cells treated with anti- mutCALR antibody conjugated to PBD (Clone 6-PBD), unconjugated anti-mutCALR antibody (Clone 6), or isotype control conjugated to PBD (Isotype-PBD) in the absence of human TPO.

[0157] FIG. 8A includes a graph showing tumor cell growth in mice administered anti- mutCALR antibody conjugated to PBD (Clone 6-PBD), unconjugated anti-mutCALR antibody (Clone 6), or isotype control, each at three doses (0.1 mg / kg, 0.3 mg / kg, 1 mg / kg). Untreated mice, or mice treated with ruxolitinib (90 mg / kg BID), or mice administered three doses of unconjugated Clone 6 (10 mg / kg IP 3X) were included as controls.

[0158] 29

[0159] SUBSTITUTE SHEET (RULE 26) FIG. 8B includes a graph showing platelet counts in mice administered anti- mutCALR antibody conjugated to PBD (Clone 6-PBD), unconjugated anti-mutCALR antibody (Clone 6), or isotype control, each at three doses (0.1 mg / kg, 0.3 mg / kg, 1 mg / kg). Untreated mice, or mice treated with ruxolitinib (90 mg / kg BID), or mice administered three doses of unconjugated Clone 6 (10 mg / kg IP 3X) were included as controls.

[0160] FIG. 9 includes a graph of mouse survival after treatment with anti-mutCALR antibody conjugated to PBD (Clone 6-PBD), unconjugated anti-mutCALR antibody (Clone 6), or isotype control.

[0161] DETAILED DESCRIPTION

[0162] Provided herein are anti-mutCALR antibody-drug conjugates comprising a pyrrolobenzodiazepine dimer, and related pharmaceutical compositions, and kits. The anti- mutCALR antibody-drug conjugates described herein are useful in the treatment or prevention of myeloproliferative neoplasms (e.g, chronic myelogenous leukemia, polycythemia vera, primary myelofibrosis, essential thrombocythemia, chronic neutrophilic leukemia, acute myelogenus leukemia, chronic eosinophilic leukemia, chronic myelomonocytic leukemia, myeloproliferative neoplasm and myelodysplastic syndrome, including myelodysplastic syndrome with refractory anaemia with ring sideroblasts, myelodysplastic syndrome with refractory anemia, and myelodysplastic syndrome with refractory anemia with excess blasts).

[0163] Mutant CALR and Anti-mutCALR Antibodies

[0164] CALR is a highly conserved chaperone protein that resides primanly in the endoplasmic reticulum and is involved in a variety of cellular processes including protein folding, calcium homeostasis, cell adhesion, and integrin signaling. Mutations in the CALR gene have been identified in patients with myeloproliferative neoplasms. The two most frequent CALR mutations are a 52 base pair (bp) deletion and a 5 bp insertion, which are referred to as Type 1 and Type 2 mutations, respectively. Type 1 and Type 2 mutations cause a +1 frameshift within exon 9 that generates a novel, positively-charged C-terminal amino acid sequence that lacks the KDEL domain (SEQ ID NO:347) of the WT protein, thereby enabling the mutCALR to escape the ER and activate the thrombopoietin receptor (MPL) and induce constitutive activation of Janus kinase 2 (JAK2) signaling. The amino acid sequences of human WT CALR, Type 1 and Type 2 mutCALR, and the novel C-terminal sequence are

[0165] 30

[0166] SUBSTITUTE SHEET (RULE 26) shown below. The frameshift amino acid residues in Type 1 and Type 2 mutCALR are shown in bold and the novel C-terminal sequence is marked by underlining.

[0167] Human WT CALR (GenBank Accession No. NP_004334.1)

[0168] MLLSVPLLLGLLGLAVAEPAVYFKEQFLDGDGWTSRWIESKHKSDFGKFVLSSGKFYGDEEKDKGLQT SQDARFYALSASFEPFSNKGQTLWQFTVKHEQNIDCGGGYVKLFPNSLDQTDMHGDSEYNIMFGPDI CGPGTKKVHVIFNYKGKNVLINKDIRCKDDEFTHLYTLIVRPDNTYEVKIDNSQVESGSLEDDWDFLP PKKIKDPDASKPEDWDERAKIDDPTDSKPEDWDKPEHI PDPDAKKPEDWDEEMDGEWEPPVIQNPEYK GE WKPRQ I DNPDYKGTW IHPEIDNPEYSPDPS I YAYDNFGVLGLDLWQVKSGT I FDNFL I TNDE AYAE EFGNETWGVTKAAEKQMKDKQDEEQRLKEEEEDKKRKEEEEAEDKEDDEDKDEDEEDEEDKEEDEEED VPGQAKDEL ( SEQ ID NO : 319 )

[0169] Human Type 1 mutCALR

[0170] MLLSVPLLLGLLGLAVAEPAVYFKEQFLDGDGWTSRWIESKHKSDFGKFVLSSGKFYGDEEKDKGLQT SQDARFYALSASFEPFSNKGQTLWQFTVKHEQNIDCGGGYVKLFPNSLDQTDMHGDSEYNIMFGPDI CGPGTKKVHVIFNYKGKNVLINKDIRCKDDEFTHLYTLIVRPDNTYEVKIDNSQVESGSLEDDWDFLP PKKIKDPDASKPEDWDERAKIDDPTDSKPEDWDKPEHI PDPDAKKPEDWDEEMDGEWEPPVIQNPEYK GE WKPRQ I DNPDYKGTW IHPEIDNPEYSPDPS I YAYDNFGVLGLDLWQVKSGT I FDNFL I TNDE AYAE

[0171] EFGNETWGVTKAAEKQMKDKQDEEQRTRRMMRTKMRMRRMRRTRRKMRRKMSPARPRTSCREACLQGW TEA ( SEQ ID NO : 320 )

[0172] Human Type 2 mutCALR

[0173] MLLSVPLLLGLLGLAVAEPAVYFKEQFLDGDGWTSRWIESKHKSDFGKFVLSSGKFYGDEEK DKGLQTSQDARFYALSASFEPFSNKGQTLWQFTVKHEQNIDCGGGYVKLFPNSLDQTDMHG DSEYNIMFGPDICGPGTKKVHVI FNYKGKNVLINKDIRCKDDEFTHLYTLIVRPDNTYEVKI

[0174] DNSQVESGSLEDDWDFLPPKKIKDPDASKPEDWDERAKIDDPTDSKPEDWDKPEHIPDPDAK KPEDWDEEMDGEWEPPVIQNPEYKGEWKPRQIDNPDYKGTWIHPEIDNPEYSPDPSIYAYDN FGVLGLDLWQVKSGTIFDNFLITNDEAYAEEFGNETWGVTKAAEKQMKDKQDEEQRLKEEEE

[0175] DKKRKEEEEAEDNCRRMMRTKMRMRRMRRTRRKMRRKMSPARPRTSCREACLQGWTEA (SEQ ID NO : 321 ) mutCALR C-terminal consensus mutant sequence RMRRMRRTRRKMRRKMSPARPRTSCREACLQGWTEA ( SEQ ID NO : 322 )

[0176] This disclosure provides anti-mutCALR antibody-drug conjugates that are useful in treating myeloproliferative neoplasms. Note that in the present disclosure, unless stated otherwise, amino acid positions assigned to CDRs and frameworks in a variable region of the anti-mutCALR antibody-drug conjugates are specified according to Kabat; see EA Rabat, Sequences of Proteins of Immunological Interest, U.S. Dept, of Health and Human Sendees, Public Health Service, National Institutes of Health, 1991, (OCoLC)l 138727707.

[0177] 31

[0178] SUBSTITUTE SHEET (RULE 26) In some embodiments, the anti-mutCALR antibody-drug conjugate comprises an anti- mutCALR antibody that comprises one, two, three, four, five, and / or six CDRs of any one of the antibodies described herein. In some embodiments, an anti-mutCALR antibody-drug conjugate comprises (i) one, two, and / or three heavy chain CDRs of any one of the clones presented in Tables 1-2, and / or (ii) one, two, and / or three light chain CDRs from any one of the clones presented in Tables 1-2.

[0179] In some embodiments, an anti-mutCALR antibody-drug conjugate comprises (i) three heavy chain CDRs from any one of the clones presented in Tables 4-5, and (ii) three light chain CDRs from any one of the clones presented in Tables 4-5. In some embodiments, an anti-mutCALR antibody-drug conjugate comprises a Group

[0180] 1 anti-mutCALR antibody described in US20230272055, the content of which is incorporated herein by reference in its entirety.

[0181] 32

[0182] SUBSTITUTE SHEET (RULE 26) Table 1. CDR sequences.

[0183]

[0184] Table 2. CDRs of mutant clones.

[0185]

[0186]

[0187]

[0188]

[0189]

[0190]

[0191]

[0192]

[0193] Table 3. Consensus CDR sequences.

[0194]

[0195] Table 4. Heavy chain, heavy chain variable region, light chain, and light chain variable region sequences.

[0196]

[0197]

[0198]

[0199]

[0200]

[0201]

[0202]

[0203]

[0204]

[0205] Table 5. Heavy chain, heavy chain variable region, light chain, and light chain variable region sequences of Clone 6 and variants thereof

[0206]

[0207]

[0208]

[0209]

[0210]

[0211]

[0212]

[0213]

[0214]

[0215]

[0216]

[0217]

[0218]

[0219]

[0220]

[0221]

[0222] In some embodiments, an anti-mutCALR antibody-drug conjugate comprises a heavy chain CDR1, CDR2, and CDR3 and / or a light chain variable region CDR1, CDR2, and CDR3 from an antibody described herein. In some embodiments, an anti-mutCALR antibody-drug conjugate comprises a heavy chain CDR1, CDR2, and CDR3 and a light chain CDR1, CDR2, and CDR3 from an antibody described herein. In some embodiments, an anti- mutCALR antibody-drug conjugate comprises a mouse version, mouse variant, human version, human variant, humanized version, humanized variant, or affinity matured variant of an antibody described herein.

[0223] In some embodiments, an anti-mutCALR antibody-drug conjugate comprises a heavy chain CDR1, CDR2, and CDR3 and / or a light chain variable region CDR1, CDR2, and CDR3 from any clone disclosed herein, a humanized version thereof, or variants thereof (including affinity matured variants). In some embodiments, an anti-mutCALR antibody-drug conjugate comprises a heavy chain CDR1, a heavy chain variable region CDR2, and a heavy chain variable region CDR3 from any clone disclosed herein. In other embodiments, an anti- mutCALR antibody-drug conjugate comprises a light chain variable region CDR1, a light chain variable region CDR2, and a light chain variable region CDR3 from any clone disclosed herein. In certain embodiments, an anti-mutCALR antibody-drug conjugate comprises a heavy7chain CDR1, a heavy chain variable region CDR2, a heavy chain variable region CDR3, a light chain variable region CDR1, a light chain variable region CDR2, and a light chain variable region CDR3 from antibody any clone disclosed herein. In some embodiments, an anti-mutCALR antibody-drug conjugate comprises a mouse version of any clone disclosed herein. In some embodiments, an anti-mutCALR antibody-drug conjugate comprises a mouse variant of any clone disclosed herein. In some embodiments, an anti- mutCALR antibody-drug conjugate comprises a human version of any clone disclosed herein. In some embodiments, an anti-mutCALR antibody-drug conjugate comprises a human variant of any clone disclosed herein. In some embodiments, an anti-mutCALR antibodydrug conjugate comprises a humanized version of any clone disclosed herein. In some embodiments, an anti-mutCALR antibody-drug conjugate comprises a variant of any clone disclosed herein. In some embodiments, an anti-mutCALR antibody-drug conjugate comprises an affinity matured variant of any clone disclosed herein.

[0224] In some embodiments, the anti-mutCALR antibody-drug conjugate comprises a variant of an anti-mutCALR antibody described herein which comprises one to thirty conservative amino acid substitution(s), e.g., one to twenty-five, one to twenty, one to fifteen, one to ten, one to five, or one to three conservative amino acid substitution(s). In some

[0225] 69

[0226] SUBSTITUTE SHEET (RULE 26) embodiments, the conservative amino acid substitution(s) is in a CDR of the antibody. In some embodiments, the conservative amino acid substitution(s) is not in a CDR of the antibody. In some embodiments, the conservative amino acid substitution(s) is in a framework region of the antibody.

[0227] In some embodiments, a CDR comprises one amino acid substitution. In some embodiments, a CDR comprises two amino acid substitutions. In some embodiments, a CDR comprises three amino acid substitutions. In some embodiments, a CDR comprises four amino acid substitutions. In some embodiments, the one or more amino acid substitutions are conservative substitutions. In some embodiments, the CDR is a heavy chain CDR1. In some embodiments, the CDR is a heavy chain variable region CDR2. In some embodiments, the CDR is a heavy chain variable region CDR3. In some embodiments, the CDR is a light chain variable region CDR1. In some embodiments, the CDR is a light chain variable region CDR2. In some embodiments, the CDR is a light chain variable region CDR3. In some embodiments, the one or more substitutions are made as part of a humanization process. In some embodiments, the one or more substitutions are made as part of a germline humanization process. In some embodiments, the one or more substitutions are made as part of an affinity maturation process. In some embodiments, the one or more substitutions are made as part of an optimization process.

[0228] In some embodiments, an anti-mutCALR antibody-drug conjugate comprises a heavy chain variable region CDR1, a heavy chain variable region CDR2, and a heavy chain variable region CDR3, each of which correspond to the heavy chain variable region CDRs set forth in Tables 1-2 for a single clone, and a light chain variable region CDR1, a light chain variable region VL CDR2, and a light chain variable region VL CDR3, each of which correspond to the VL CDRs set forth in Tables 1-2 for a single clone.

[0229] In some embodiments, an anti-mutCALR antibody-drug conjugate comprises a heavy chain variable region CDR1, a heavy chain variable region CDR2, a heavy chain variable region CDR3, a light chain variable region CDR1, a light chain variable region CDR2, and a light chain variable region CDR3, each of which correspond to the VH and VL CDRs set forth in Tables 1-2 for a single clone.

[0230] In some embodiments, an anti-mutCALR antibody-drug conjugate comprises a heavy chain variable region (VH) comprising a VH CDR1 ; a VH CDR2; and a VH CDR3; wherein the VH CDR1 comprises the amino acid sequence ELSMQ (SEQ ID NO:1); the VH CDR2 comprises the amino acid sequence GFDPDDX101ETMYAEX102X103QG (SEQ ID NO: 102); wherein X101 is D or G; wherein X102 is K or R; and wherein X103 is F or L; the VH CDR3 is

[0231] 70

[0232] SUBSTITUTE SHEET (RULE 26) SPGYDFFDY (SEQ ID NO: 18); wherein the antibody comprises a light chain variable region (VL) comprising a VL CDR1, a VL CDR2, and a VL CDR3, wherein the VL CDR1 comprises the amino acid sequence GGX104X105X106GX107X108X109VX110 (SEQ ID NO: 103), wherein X104 is N, D, or S; wherein X105 is Y, N, or D; wherein X106 is I or T; wherein X107 is S, D, I, R, or T; wherein X108 is K, E, or I; wherein X109 is S, I, R, G, N, or A; and wherein Xno is H, F, or N; the VL CDR2 comprises the amino acid sequence DDX111DRPX112 (SEQ ID NO: 104), wherein X111 is G, S, or R; and wherein X112 is S or L; and the VL CDR3 comprises the amino acid sequence QVWDXi 13X114X115DX116X117X1 is (SEQ ID NO:105), wherein X113 is S or A; wherein X114 is I or S; wherein X115 is S, I, or N; wherein X116 is H, L, or Q; wherein X117 is V or L; and wherein Xus is V or I.

[0233] In some embodiments, an anti-mutCALR antibody-drug conjugate comprises a heavy chain variable region (VH) comprising a VH CDR1 ; a VH CDR2; and a VH CDR3; wherein the VH CDR1 comprises the amino acid sequence ELSMQ (SEQ ID NO: 1); the VH CDR2 comprises the amino acid sequence GFDPDDX101ETMYAEX102X103QG (SEQ ID NO: 102); wherein X101 is D or G; wherein X102 is K or R; and wherein X103 is F or L; the VH CDR3 is SPGYDFFDY (SEQ ID NO: 18); wherein the antibody comprises a light chain variable region (VL) comprising a VL CDR1, a VL CDR2, and a VL CDR3, wherein the VL CDR1 comprises the amino acid sequence TGTSSDVGGYNYVS (SEQ ID NO:30); the VL CDR2 comprises the amino acid sequence X119VSX120RPS (SEQ ID NO:106); wherein X119 is E or D; and wherein X120 is N or K; and the VL CDR3 comprises the ammo acid sequence QVWDSSNDLLI (SEQ ID NO:71).

[0234] In some embodiments, an anti-mutCALR antibody-drug conjugate comprises a heavy chain variable region (VH) comprising a VH CDR1 ; a VH CDR2; and a VH CDR3; wherein: the VH CDR1 comprises the amino acid sequence GYTLTELSMQ (SEQ ID NO:329); the VH CDR2 comprises the amino acid sequence GFDPDDX101ETMYAEX102X103QG (SEQ ID NO: 102); wherein X101 is D or G; wherein X102 is K or R; and wherein X103 is F or L; the VH CDR3 is SPGYDFFDY (SEQ ID NO: 18); wherein the antibody comprises a light chain variable region (VL) comprising a VL CDR1, a VL CDR2, and a VL CDR3, wherein: the VL CDR1 comprises the amino acid sequence TGTSSDVGGYNYVS (SEQ ID NO:30); the VL CDR2 comprises the amino acid sequence X119VSX120RPS (SEQ ID NO: 106); wherein X119 is E or D; and wherein X120 is N or K; and the VL CDR3 comprises the amino acid sequence QVWDSSNDLLI (SEQ ID NO:71).

[0235] In some embodiments, an anti-mutCALR antibody-drug conjugate comprises a heavy chain variable region with the C-terminal lysine removed. In some embodiments, an anti-

[0236] 71

[0237] SUBSTITUTE SHEET (RULE 26) mutCALR antibody-drug conjugate comprises a heavy chain variable region comprising an amino acid sequence that has the three VH CDRs of any anti-mutCALR clone disclosed herein and which has at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to any one of the VH sequences set forth in Tables 4-5 and a light chain variable region comprising an amino acid sequence that has the three VL CDRs of any anti-mutCALR clone disclosed herein and which has at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identity to any one of the VL sequences set forth in Tables 4-5.

[0238] In some embodiments, an anti-mutCALR antibody-drug conjugate comprises a heavy chain variable region comprising any one of the VH sequences set forth in Tables 4-5. In some embodiments, an anti-mutCALR antibody-drug conjugate comprises a light chain variable region comprising any one of the VL sequences set forth in Tables 4-5. In some embodiments, an anti-mutCALR antibody-drug conjugate comprises a heavy chain variable region comprising any one of the VH sequences set forth in Tables 4-5 and a light chain variable region comprising any one of the VL sequences set forth in Tables 4-5.

[0239] In some embodiments, an anti-mutCALR antibody-drug conjugate comprises a modification which modulates (e.g.. reduces or increases) the Fc region-mediated effector function, such as complement-dependent cytotoxicity (CDC), antibody-dependent cellular cytotoxicity (ADCC) and antibody-dependent cell phagocytosis (ADCP). Depending on the therapeutic antibody or Fc fusion protein application, it may be desired to either reduce or increase the effector function.

[0240] In certain embodiments, an anti-mutCALR antibody-drug conjugate has Fc effector function. In certain embodiments, an anti-mutCALR antibody-drug conjugate has enhanced Fc effector function. In certain embodiments, an anti-mutCALR antibody -drug conjugate exhibits antibody-dependent cell-mediated cytotoxicity (ADCC). An anti-mutCALR antibody-drug conjugate can be engineered to enhance the ADCC activity (for review, see Kubota T et al. Cancer Sci. 2009; 100(9): 1566-72). For example, ADCC activity of an antibody can be improved when the antibody itself has a low ADCC activity, by slightly modifying the constant region of the antibody (Junttila TT. et al. Cancer Res. 2010;70(l 1):4481-9). Changes are sometimes also made to improve storage or production or to remove C-terminal lysines (Kubota T et al. Cancer Sci. 2009;100(9): 1566-72). Another suitable method to improve ADCC activity of an antibody is by enzymatically interfering with the glycosylation pathway resulting in a reduced fucose (von Horsten HH. etal. Glycobiology. 2010;20(12): 1607-18). Alternatively, or additionally, other suitable methods can be used to achieve ADCC enhancement, for instance including glycoengineering (Kyowa

[0241] 72

[0242] SUBSTITUTE SHEET (RULE 26) Hakko / Biowa, GlycArt (Roche) and Eureka Therapeutics) and mutagenesis, all of which seek to improve Fc binding to low-affinity activating FcyRIIIa, and / or to reduce binding to the low affinity inhibitory FcyRIIb. In certain embodiments, a binding moiety of the present disclosure exhibits enhanced antibody-dependent cell-mediated cytotoxicity (ADCC). In certain embodiments, a binding moiety of the present disclosure is afucosylated.

[0243] In certain embodiments, an anti-mutCALR antibody-drug conjugate has reduced Fc effector function. In certain embodiments, an anti-mutCALR antibody-drug conjugate exhibits reduced or substantially no complement-dependent cytotoxicity (CDC), antibodydependent cellular cytotoxicity (ADCC) or antibody -dependent cell phagocytosis (ADCP). In certain embodiments, an anti-mutCALR antibody-drug conjugate exhibits reduced or substantially no antibody-dependent cell-mediated cytotoxicity (ADCC). An anti-mutCALR antibody-drug conjugate can be engineered to reduce effector function, for example ADCC activity, by any suitable method including removal of glycosylation sites in the Fc region. In certain embodiments, an anti-mutCALR antibody-drug conjugate that has a reduced Fc effector function (e.g., a reduced ADCC effector function) comprises an N297A mutation on the heavy chain.

[0244] In some embodiments, an anti-mutCALR antibody-drug conjugate comprises an IgGl isotype (e.g., IgGl, IgG2, IgG3 or IgG4). In some embodiments, an anti-mutCALR antibody - drug conjugate comprises an IgGl. In some embodiments, an IgGl, IgG2, IgG3 or IgG4 anti- mutCALR antibody-drug conjugate has Fc-effector function. In some embodiments, an IgGl, IgG2, IgG3 or IgG4 anti-mutCALR antibody-drug conjugate is Fc-effector function null. In some embodiments, an IgGl anti-mutCALR antibody-drug conjugate has Fc-effector function. In some embodiments, an IgGl anti-mutCALR antibody-drug conjugate is Fc- effector function null.

[0245] In some instances, the anti-mutCALR antibody-drug conjugate comprises an antibody fragment. Fragments of the antibodies described herein (e.g., Fab, Fab', F(ab')2, Facb, and Fv) may be prepared by proteolytic digestion of intact antibodies. For example, antibody fragments can be obtained by treating the whole antibody with an enzyme such as papain, pepsin, or plasmin or the FabRICATOR® (IdeS) recombinant enzyme (Genovis AB) that digests IgG antibodies to produce a homogeneous pool of F(ab')2 and Fc / 2 fragments. Papain digestion of whole antibodies produces F(ab)2 or Fab fragments; pepsin digestion of whole antibodies yields F(ab')2 or Fab'; and plasmin digestion of whole antibodies yields Facb fragments.

[0246] 73

[0247] SUBSTITUTE SHEET (RULE 26) Alternatively, antibody fragments can be produced recombinantly. For example, nucleic acids encoding the antibody fragments of interest can be constructed, introduced into an expression vector, and expressed in suitable host cells. See, e.g., Co, M.S. et al., J. Immunol., 152:2968-2976 (1994); Better, M. and Horwitz, A.H., Methods in Enzymology, 178:476-496 (1989); Plueckthun, A. and Skerra, A., Methods in Enzymology, 178:476-496 (1989); Lamoyi, E., Methods in Enzymology, 121 :652-663 (1989); Rousseaux, J. et al., Methods in Enzymology, (1989) 121:663-669 (1989); and Bird, R.E. et al, TIBTECH, 9: 132-137 (1991)). Antibody fragments can be expressed in and secreted from A. coll, thus allowing the facile production of large amounts of these fragments. Antibody fragments can be isolated from the antibody phage libraries. Alternatively, Fab'-SH fragments can be directly recovered from A. coli and chemically coupled to form F(ab)2 fragments (Carter et al., Bio / Technology, 10: 163-167 (1992)). According to another approach, F(ab')2 fragments can be isolated directly from recombinant host cell culture. Fab and F(ab')2 fragment with increased in vivo half-life comprising a salvage receptor binding epitope residues are described in U.S. Pat. No. 5,869,046.

[0248] In some instances, the anti-mutCALR antibody-drug conjugate comprises a minibody. Minibodies of anti-mutCALR antibodies include diabodies, single chain (scFv), and singlechain (Fv)2 (SC(FV)2).

[0249] A “diabody” is a bivalent minibody constructed by gene fusion (see, e.g, Holliger, P. et al, Proc. Natl. Acad. Sci. U. S. A., 90:6444-6448 (1993); EP 404,097; WO 93 / 11161). Diabodies are dimers composed of two polypeptide chains. The VL and VH domain of each polypeptide chain of the diabody are bound by linkers. The number of amino acid residues that constitute a linker can be between 2 to 12 residues (e.g., 3-10 residues or five or about five residues). The linkers of the polypeptides in a diabody are typically too short to allow the VL and VH to bind to each other. Thus, the VL and VH encoded in the same polypeptide chain cannot form a single-chain variable region fragment, but instead form a dimer with a different single-chain variable region fragment. As a result, a diabody has two antigenbinding sites.

[0250] An scFv is a single-chain polypeptide antibody obtained by linking the VH and VL with a linker (see, e.g., Huston et al., Proc. Natl. Acad. Sci. U. S. A., 85:5879-5883 (1988); and Plickthun, “The Pharmacology of Monoclonal Antibodies” Vol.113, Ed Resenburg and Moore, Springer Verlag, New York, pp.269-315, (1994)). The order of VHs and VLs to be linked is not particularly limited, and they may be arranged in any order. Examples of arrangements include: [VH] linker [VL]; or [VL] linker [VH], The heavy chain variable

[0251] 74

[0252] SUBSTITUTE SHEET (RULE 26) domain and light chain variable domain in an scFv may be derived from any anti-mutCALR antibody described herein.

[0253] An SC(FV)2 is a minibody in which two VHs and two VLs are linked by a linker to form a single chain (Hudson, et al., J. Immunol. Methods, (1999) 231: 177-189 (1999)). An SC(FV)2 can be prepared, for example, by connecting scFvs with a linker. The sc(Fv)2 of the present invention include antibodies preferably in which two VHs and two VLs are arranged in the order of: VH, VL, VH, and VL ([VH] linker [VL] linker [VH] linker [VL]), beginning from the N terminus of a single-chain polypeptide; however the order of the two VHs and two VLs is not limited to the above arrangement, and they may be arranged in any order.

[0254] In some instances, the anti-mutCALR antibody-drug conjugate comprises a bispecific antibody. Bispecific antibodies are antibodies that have binding specificities for at least two different epitopes. Exemplary bispecific antibodies may bind to two different epitopes of the mutCALR protein. Other such antibodies may combine a mutCALR binding site with a binding site for another antigen. Bispecific antibodies can be prepared as full length antibodies or low molecular weight forms thereof (e.g., F(ab')2 bispecific antibodies, sc(Fv)2 bispecific antibodies, diabody bispecific antibodies).

[0255] Traditional production of full length bispecific antibodies is based on the coexpression of two immunoglobulin heavy chain-light chain pairs, where the two chains have different specificities (Millstein et al., Nature, 305:537-539 (1983)). In a different approach, antibody variable domains with the desired binding specificities are fused to immunoglobulin constant domain sequences. DNAs encoding the immunoglobulin heavy chain fusions and, if desired, the immunoglobulin light chain, are inserted into separate expression vectors, and are co-transfected into a suitable host cell. This provides for greater flexibility in adjusting the proportions of the three polypeptide fragments. It is, however, possible to insert the coding sequences for two or all three polypeptide chains into a single expression vector when the expression of at least two polypeptide chains in equal ratios results in high yields.

[0256] According to another approach described in U.S. Pat. No. 5,731,168, the interface between a pair of antibody molecules can be engineered to maximize the percentage of heterodimers that are recovered from recombinant cell culture. The preferred interface comprises at least a part of the CH3 domain. In this method, one or more small amino acid side chains from the interface of the first antibody molecule are replaced with larger side chains (e.g., tyrosine or tryptophan). Compensatory “cavities” of identical or similar size to the large side chain(s) are created on the interface of the second antibody molecule by replacing large amino acid side chains with smaller ones (e.g, alanine or threonine). This

[0257] 75

[0258] SUBSTITUTE SHEET (RULE 26) provides a mechanism for increasing the yield of the heterodimer over other unwanted endproducts such as homodimers.

[0259] Bispecific antibodies include cross-linked or “heteroconjugate” antibodies. For example, one of the antibodies in the heteroconjugate can be coupled to avidin, the other to biotin. Heteroconjugate antibodies may be made using any convenient cross-linking methods.

[0260] The “diabody” technology provides an alternative mechanism for making bispecific antibody fragments. The fragments comprise a VH connected to a VL by a linker which is too short to allow pairing between the two domains on the same chain. Accordingly, the VH and VL domains of one fragment are forced to pair with the complementary VL and VH domains of another fragment, thereby forming two antigen-binding sites.

[0261] In certain embodiments, the bispecific anti-mutCALR antibody-drug conjugate comprises a biparatopic antibody. A biparatopic antibody is antibody which recognizes two non-identical epitopes (overlapping or non-overlapping epitopes) on the same target antigen (e.g. the C-terminal of mutCALR domain). A biparatopic anti-mutCALR antibody-drug conjugate can comprise two immunoglobulin heavy chain-light chain pairs or one immunoglobulin heavy chain-light chain pair. In some embodiments, a biparatopic anti- mutCALR antibody-drug conjugate comprises one immunoglobulin heavy-chain-lighl chain pair. In some embodiments, a biparatopic anti-mutCALR antibody-drug conjugate comprises a full-length antibody comprising one immunoglobulin heavy -chain-light chain pair.

[0262] In some instances, the anti-mutCALR antibody-drug conjugate comprises a multivalent antibody. A multivalent antibody may be internalized (and / or catabolized) faster than a bivalent antibody by a cell expressing an antigen to which the antibodies bind. The antibodies describe herein can be multivalent antibodies with three or more antigen binding sites (e.g., tetravalent antibodies), which can be readily produced by recombinant expression of nucleic acid encoding the polypeptide chains of the antibody. The multivalent antibody can comprise a dimerization domain and three or more antigen binding sites. An exemplary dimerization domain comprises (or consists of) an Fc region or a hinge region. A multivalent antibody can comprise (or consist of) three to about eight (e.g, four) antigen binding sites. The multivalent antibody optionally comprises at least one polypeptide chain (e.g, at least two polypeptide chains), wherein the polypeptide chain(s) comprise two or more variable domains. For instance, the polypeptide chain(s) may comprise VDl-(Xl)n-VD2-(X2)n-Fc, wherein VD1 is a first variable domain, VD2 is a second variable domain, Fc is a polypeptide chain of an Fc region, XI and X2 represent an amino acid or peptide spacer, and n is 0 or 1.

[0263] 76

[0264] SUBSTITUTE SHEET (RULE 26) Pyrrolobenzodiazepine Dimers

[0265] The present disclosure provides an anti-mutCALR antibody-drug conjugate comprising any one of the anti-mutCALR antibodies described herein and a pyrrolobenzodiazepine dimer, e.g., a pyrrolobenzodiazepine dimer described herein.

[0266] Pyrrolobenzodiazepines are sequence-selective DNA minor-groove binding agents based on the naturally occurring anthramycin family of antitumor antibiotics.

[0267] Pyrrolobenzodiazepine has a general structure shown below:

[0268] Pyrrolobenzodiazepines differ in the number, type and position of substituents, in both their aromatic A-rings and pyrrolo C-rings, and in the degree of saturation of the C-ring. In the B-ring there is either an imine (N=C), a carbinolamine (NH-CH(OH)), or a carbinolamine methyl ether (NH-CH(OMe)) at the N10-C11 position, which is the electrophilic center responsible for alkylating DNA. The biological activity of pyrrolobenzodiazepines can be enhanced by joining two pyrrolobenzodiazepines monomers together through their C8 / C'-hydroxyl functionalities via a flexible alkylene linker to form a pyrrolobenzodiazepine dimer (see, e.g., Bose, D.S. et al., “Rational Design of a Highly Efficient Irreversible DNA Interstrand Cross-Linking Agent Based on the Pyrrolobenzodiazepine Ring System,” J. Am. Chem. Soc., 114, 4939-4941 (1992); Thurston, D.E. et al., “Synthesis of Sequence-selective C8-linked Pyrrolo [2,l-c][l,4] Benzodiazepine DNA Interstrand Cross-linking Agent,” J. Org. Chem., 61:8141-8147 (1996)).

[0269] Any pyrrolobenzodiazepine dimer (e.g., a pyrrolobenzodiazepine dimer described herein) can be included in an anti-mutCALR antibody-drug conjugate described herein. Nonlimiting examples of pyrrolobenzodiazepine dimers that can be used in anti-mutCALR antibody-drug conjugates are provided in Mantaj et al., “From Anthramycin to Pyrrolobenzodiazepine (PBD)-Containing Antibody-Drug Conjugates (ADCs),” Angew. Chem. Int. Ed. 56, 462 - 48, (2017); Varvounis, “An update on the Synthesis of Pyrrolo[l,4]benzodiazepines,” Molecules, 21(2), 154 (2016); US7,528,126; W02005085260; W02006111759; WO2011 / 130598; WO2013055987; W02014057074; the contents of each of which is incorporated herein by reference in its entirety.

[0270] 77

[0271] SUBSTITUTE SHEET (RULE 26) Non-limiting examples of pyrrolobenzodiazepine dimers include PBD-1, SG2000,

[0272] SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882. In some embodiments, the pyrrolobenzodiazepine dimer has a structure selected from the group consisting of:

[0273] SUBSTITUTE SHEET (RULE 26)

[0274] In some embodiments, pyrrolobenzodiazepine dimer PBD-1 further comprises a cleavable cap. In some embedments, pyrrolobenzodiazepine dimer PBD-1 further comprises a cleavable cap and is of the following formula: Non-limiting examples of pyrrolobenzodiazepine dimers include SG2000, SG2219,

[0275] SG3199, SG3312, SG3650, SG2057, and SGD-1882. In some embodiments, the pyrrolobenzodiazepine dimer has a structure selected from the group consisting of:

[0276] 79

[0277] SUBSTITUTE SHEET (RULE 26)

[0278] In some embodiments, the pyrrolobenzodiazepine dimer is linked to a linker to form a pay load. The pyrrolobenzodiazepine dimer can be conjugated to the antibody by performing chemical and / or enzymatic modifications on the antibodies, respectively, or the lower molecular weight forms thereof described herein. Methods for conjugating payloads are known in the art (e.g., lysine amide coupling, cysteine coupling, non-natural amino acid incorporation by genetic engineering, transpeptidation using sortase or microbial transglutaminase, and N-glycan engineering) and provided in, e.g., Tsuchikama and An, “Antibody-drug conjugates: recent advances in conjugation and linker chemistries,” Protein Cell, 9(l):33-46 (2018); WO2011 / 130613; WO2011 / 130616; WO2016 / 054315; and WO2011 / 130598; the contents of each of which is incorporated herein by reference in its entirety.

[0279] 80

[0280] SUBSTITUTE SHEET (RULE 26) Any payload compnsing a pyrrolobenzodiazepine dimer and a linker (e.g. , a linker described herein) can be included in an anti-mutCALR antibody-drug conjugate described herein. Non-limiting examples of payloads that can be used in anti-mutCALR antibody -drug conjugates are provided in Mantaj et al, “From Anthramycin to Pyrrolobenzodiazepine (PBD)-Containing Antibody-Drug Conjugates (ADCs),” Angew. Chem. Int. Ed. 56, 462 - 48, (2017); Christie et at., “Pyrrolobenzodiazepine Antibody-Drug Conjugates Designed for Stable Thiol Conjugation, Antibodies (Basel), 6(4):20 (2017); W02011 / 130613;

[0281] WO2011 / 130616; WO2011 / 130598; W02014 / 057073; WO2015 / 052321; and

[0282] WO2015052322; the contents of each of which is incorporated herein by reference in its entirety.

[0283] Non-limiting examples of payloads include PL-1, PL-2, PL-3, SG3400, SG3227, SG3249, SG3710, SG3259, SG3584, SG3203, SG3231, SG3451, and SGD-1910. In some embodiments, the payload has a structure selected from the group consisting of:

[0284] 81

[0285] SUBSTITUTE SHEET (RULE 26)

[0286]

[0287] 82

[0288] SUBSTITUTE SHEET (RULE 26)

[0289]

[0290] 83

[0291] SUBSTITUTE SHEET (RULE 26) SGD

[0292] The above payloads, and other payloads described herein, are depicted in their structural form prior to conjugation with an antibody with the understanding that the payloads would attach to an antibody through a reactive group on the linker moiety such as a terminal cyclooctynyl group, terminal maleimide group, terminal iodo group, or other reactive group.

[0293] Examples of cyclooctynyl reaction chemistry that can be used to connect the payload and the antibody can be found, for example, in U.S. Patent Nos.: 7,807,619; 8,431,558; 8,461,298; 9,260,371; 10,434,111; 11,278,554; 7,807,619; 8,431,558; 8,461,298; 9,260,371; 10,434,111; and 11,278,554; and European Patent No.: 1812031B1, the disclosures of which are incorporated herein by reference in their entireties.

[0294] Non-limiting examples of payloads include SG3400, SG3227, SG3249, SG3710, SG3259, SG3584, SG3203, SG3231, SG3451, and SGD-1910. In some embodiments, the payload has a structure selected from the group consisting of:

[0295] 84

[0296] SUBSTITUTE SHEET (RULE 26)

[0297]

[0298] 85

[0299] SUBSTITUTE SHEET (RULE 26)

[0300] 1910

[0301] 86

[0302] SUBSTITUTE SHEET (RULE 26) wherein the wavy line (where present) represents a connection point to an antibody or wherein the terminal maleimide (when present) reacts with a thiol group in the antibody to connect the payload and the antibody.

[0303] Any pyrrolobenzodiazepine dimer (e.g, a pyrrol obenzodiazepine dimer described herein) can comprise a cap, which can also be referred to as a trigger. In some embodiments, the pyrrolobenzodiazepine dimer includes a cap at the N10 position of the pyrrolobenzodiazepine ring system. Alternatively, or in addition to, a linker between the pyrrolobenzodiazepine dimer and the antibody can include a cap.

[0304] In some embodiments, the cap comprises a cleavable cap, e.g., an enzyme-cleavable cap. For example, the pyrrolobenzodiazepine dimer can comprise a glucuronide group that is cleaved by P-glucuronidase, which is overexpressed in many cancers, but not present in general circulation. In such instances, the cap can mask the cytotoxic effects of the pyrrolobenzodiazepine dimer until the cap is cleaved by a factor in a cancer cell.

[0305] Non-limiting examples of caps include a glucuronide group or derivative thereof and a galactoside group or derivative thereof. See, also, Gregson et al., “Synthesis and evaluation of pyrrolobenzodiazepine dimer antibody-drug conjugates with dual P-glucuronide and dipeptide triggers,” European Journal of Medicinal Chemistry 179, 591-607 (2019) and Su et al., “Antibody-drug conjugates: Recent advances in linker chemistry ,” Acta Pharmaceutica Sinica B, 11 (12) : 3889-3907 (2021), the contents of each of which is incorporated herein by reference in its entirety.

[0306] In some embodiments, the pyrrolobenzodiazepine dimer (e.g, PBD-1, SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, or SGD-1882) comprises a P-glucuronidase- cleavable cap, e.g., a glucuronide group or derivative thereof. In some embodiments, the pyrrolobenzodiazepine dimer (e.g., PBD-1, SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, or SGD-1882) comprises a P-galactosidase-cleavable cap, e.g., a galactoside group or derivative thereof.

[0307] In some embodiments, the payload includes a pyrrolobenzodiazepine dimer (e.g., PBD-1, SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, or SGD-1882) comprising a cap, e.g., a glucuronide group or derivative thereof and a galactoside group or derivative thereof.

[0308] In some embodiments, the payload has a structure of:

[0309] 87

[0310] SUBSTITUTE SHEET (RULE 26)

[0311] SG3600.

[0312] In some embodiments, the payload has a structure of:

[0313] In some embodiments, the payload has a structure of: wherein CAP is a cleavable cap as described herein.

[0314] In some embodiments, the payload has a structure of:

[0315] PL-3.

[0316] 88

[0317] SUBSTITUTE SHEET (RULE 26) A linker (e.g., a linker described herein) can be used to link the anti-mutCALR antibody and the pyrrolobenzodiazepine dimer. Non-limiting examples of linkers are provided in Tsuchikama and An, “Antibody-drug conjugates: recent advances in conjugation and linker chemistries.” Protein Cell, 9(1): 33-46 (2018); WO2015 / 052322; WO2015 / 095124; WO2016 / 054315; WO2017 / 137555; and WO2022 / 079211; the contents of each of which is incorporated herein by reference in its entirety.

[0318] Non-limiting examples of linkers include peptide linkers (e.g., cathepsin B-cleavable peptide linker such as valine-citrulline-p-aminobenzyloxy carbonyl (PABC) and valine- alanine-PABC), hydrozone linkers, disulfide linkers, and pyrophosphate diester linkers.

[0319] The linker can be cleavable or non-cleavable. In some embodiments, the linker is cleavable. In such instances, the linker can be cleaved upon internalization of the anti- mutCALR antibody-drug conjugate into the tumor cell expressing mutCALR, thereby releasing the pyrrolobenzodiazepine dimer, which is cytotoxic, into the tumor cell and minimizing its exposure to non-tumorigenic cells and tissues. In some embodiments, the linker is non-cleavable. In some embodiments, the non-cleavable linker comprises a maleimidocaproyl linker, a 4-maleimidomethyl cyclohexane-l-carboxylate linker, or both. In some embodiments, the anti-mutCALR antibody-drug conjugate comprises a cleavable linker, a non-cleavable linker, or both.

[0320] In some embodiments, the peptide linker comprises a dipeptide linker (e.g., valinecitrulline (vc or val-cit), alanine-phenylalanie (af or ala-phe)), a tripeptide linker e.g, glycine - valine-citrulline (gly-val-cit) and glycine-glycine-glycine (gly-gly-gly )), a tetrapeptide linker, or a pentapeptide linker. In some embodiments, the peptide linker comprises naturally occurring amino acids, non-naturally occurring amino acids, or a combination thereof.

[0321] In some embodiments, the linker comprises a functional group that reacts with a thiol group of an antibody to form an antibody-drug conjugate. Non-limiting examples of functional groups that are reactive with a thiol group of an antibody include bromoacetamide, disulfide, iodoacetamide, isocynate, isothiocynate, maleimide, pyridyl disulfide, and vinyl pyridine.

[0322] In some embodiments, the linker comprises a terminal maleimide group that can be used to conjugate the pyrrolobenzodiazepine dimer to the anti-mutCALR antibody. In some embodiments, the linker comprises a terminal iodoacetamide group that can be used to conjugate the pyrrolobenzodiazepine dimer to the anti-mutCALR antibody.

[0323] In some embodiments, the linker comprises a self-immolative moiety that generates free pyrrolobenzodiazepine dimer via an immolative elimination. Non-limiting examples of

[0324] 89

[0325] SUBSTITUTE SHEET (RULE 26) self-immolative moieties include 2-aminoimidazol-5-methanol derivatives, orthoaminobenzylacetals, para-aminobenzylacetals, para-amino-benzyloxy carbonyl (PAB), and thiazoles.

[0326] In some embodiments, the anti-mutCALR antibody-drug conjugate comprises a spacer. A spacer (e.g., a spacer described herein) can be included in the anti-mutCALR antibody-drug conjugates described herein. In some embodiments, the anti-mutCALR antibody-drug conjugate comprises a polyethylene glycol spacer, a sulfamide spacer, or a combination thereof.

[0327] In some embodiments, the linker provided herein is selected from a linker disclosed in any of U.S. Patent Nos.: 8,859,629, 9,636,421, 11,951,175, 11,168,085, 10,131,682, 10,456,479, 10,646,585, 10,975,112; U.S. Publication Nos.: 20230364262A1, 20230114866A1, 20230226208A1; International Publication No.: WO2024121123A1; and European Publication No.: 4450489A1, the disclosures of which are incorporated herein by reference in their entireties.

[0328] In some embodiments, the linker has a structure of: wherein - indicates the bond between the linker and the pyrrolobenzodiazepine dimer. In some embodiments, linker L-l is connected to the antibody via the cyclooctynyl moiety.

[0329] In some embodiments, the linker has a structure of: wherein - indicates the bond between the linker and the pyrrolobenzodiazepine dimer. In some embodiments, linker L-2 is connected to the antibody via the cyclooctynyl moiety'.

[0330] 90

[0331] SUBSTITUTE SHEET (RULE 26) In some embodiments, the anti-mutCALR antibody-drug conjugate comprises an anti- mutCALR antibody that is modified with a moiety that improves its stabilization and / or retention in circulation, e.g, in blood, serum, or other tissues, e.g, by at least 1.5, 2, 5, 10, or 50 fold. For example, the anti-mutCALR antibody can be associated with (e.g, conjugated to) a polymer, e.g, a substantially non-antigenic polymer, such as a polyalkylene oxide or a polyethylene oxide. Suitable polymers will vary substantially by weight. Polymers having molecular number average weights ranging from about 200 to about 35,000 Daltons (or about 1,000 to about 15,000, and 2,000 to about 12,500) can be used. For example, the anti- mutCALR antibody can be conjugated to a water soluble polymer, e.g. , a hydrophilic polyvinyl polymer, e.g, polyvinylalcohol or polyvinylpyrrolidone. Examples of such polymers include polyalkylene oxide homopolymers such as polyethylene glycol (PEG) or polypropylene gly cols, poly oxy ethylenated polyols, copolymers thereof and block copolymers thereof, provided that the water solubility of the block copolymers is maintained. Additional useful polymers include poly oxyalkylenes such as polyoxyethylene, polyoxypropylene, and block copolymers of polyoxyethylene and polyoxypropylene; polymethacrylates; carbomers; and branched or unbranched polysaccharides.

[0332] The above-described modified antibodies can be prepared by performing chemical and / or enzymatic modifications on the antibodies, respectively, or the lower molecular weight forms thereof described herein. Methods for modifying antibodies are well known in the art (see, e.g., US 5,057,313 and US 5,156,840).

[0333] An antibody-drug conjugate described herein can include a drug loading of one or more pyrrolobenzodiazepine dimers per anti-mutCALR antibody, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more pyrrolobenzodiazepine dimers per anti-mutCALR antibody. Drug loading refers to the average number of pyrrolobenzodiazepine dimers per anti-mutCALR antibody.

[0334] Methods of Producing Antibodies

[0335] Antibodies described herein may be produced in bacterial or eukaryotic cells. Some antibodies, e.g., Fabs, can be produced in bacterial cells, e.g., E. coli cells. Antibodies can also be produced in eukaryotic cells such as transformed cell lines (e.g, CHO, 293E, COS). In addition, antibodies (e.g, scFvs) can be expressed in a yeast cell such as Pichia (see, e.g, Powers et al., J Immunol Methods. 251: 123-35 (2001)), Hansenula, or Saccharomyces . To produce the antibody of interest, a polynucleotide encoding the antibody is constructed, introduced into an expression vector, and then expressed in suitable host cells. Standard

[0336] 91

[0337] SUBSTITUTE SHEET (RULE 26) molecular biology techniques are used to prepare the recombinant expression vector, transfect the host cells, select for transformants, culture the host cells and recover the antibody.

[0338] If the antibody is to be expressed in bacterial cells (e.g, E. coli), the expression vector should have characteristics that permit amplification of the vector in the bacterial cells. Additionally, when E. coli such as JM109, DH5a, HB101, or XLl-Blue is used as a host, the vector must have a promoter, for example, a lacZ promoter (Ward et al., 341:544-546 (1989), araB promoter (Better et al., Science, 240: 1041-1043 (1988)), or T7 promoter that can allow efficient expression in E. coli. Examples of such vectors include, for example, M13-series vectors, pUC-series vectors, pBR322, pBluescript, pCR-Script, pGEX-5X-l (Pharmacia), “QIAexpress system” (QIAGEN), pEGFP, and pET (when this expression vector is used, the host is preferably BL21 expressing T7 RNA polymerase). The expression vector may contain a signal sequence for antibody secretion. For production into the periplasm of E. coli, the pelB signal sequence (Lei et al., J. Bacteriol., 169:4379 (1987)) may be used as the signal sequence for antibody secretion. For bacterial expression, calcium chloride methods or electroporation methods may be used to introduce the expression vector into the bacterial cell.

[0339] If the antibody is to be expressed in animal cells such as CHO, COS, and NIH3T3 cells, the expression vector includes a promoter necessary for expression in these cells, for example, an SV40 promoter (Mulligan et al., Nature, 277: 108 (1979)), MMLV-LTR promoter, EFla promoter (Mizushima et al., Nucleic Acids Res., 18:5322 (1990)), or CMV promoter. In addition to the nucleic acid sequence encoding the immunoglobulin or domain thereof, the recombinant expression vectors may carry additional sequences, such as sequences that regulate replication of the vector in host cells (e.g., origins of replication) and selectable marker genes. The selectable marker gene facilitates selection of host cells into which the vector has been introduced (see, e.g, U.S. Pat. Nos. 4,399,216, 4,634,665 and 5,179,017). For example, typically the selectable marker gene confers resistance to drugs, such as G418, hygromycin, or methotrexate, on a host cell into which the vector has been introduced. Examples of vectors with selectable markers include pMAM, pDR2, pBK-RSV, pBK-CMV, pOPRSV, and pOP13.

[0340] In one embodiment, antibodies are produced in mammalian cells. Exemplary mammalian host cells for expressing an antibody include Chinese Hamster Ovary (CHO cells) (including dhfr- CHO cells, described in Urlaub and Chasin (1980) Proc. Natl. Acad. Sci. USA 77:4216-4220, used with a DHFR selectable marker, e.g, as described in Kaufman and Sharp (1982) Mol. Biol. 159:601 621), human embryonic kidney 293 cells (e.g., 293,

[0341] 92

[0342] SUBSTITUTE SHEET (RULE 26) 293E, 293T), COS cells, NIH3T3 cells, lymphocytic cell lines, e.g., NSO myeloma cells and SP2 cells, and a cell from a transgenic animal, e.g., a transgenic mammal. For example, the cell is a mammary' epithelial cell.

[0343] In an exemplary system for antibody expression, a recombinant expression vector encoding both the antibody heavy chain and the antibody light chain of an anti-mutCALR antibody is introduced into dhfr- CHO cells by calcium phosphate-mediated transfection. Within the recombinant expression vector, the antibody heavy and light chain genes are each operatively linked to enhancer / promoter regulatory elements e.g., derived from SV40, CMV, adenovirus and the like, such as a CMV enhancer / AdMLP promoter regulatory element or an SV40 enhancer / AdMLP promoter regulatory element) to drive high levels of transcription of the genes. The recombinant expression vector also carries a DHFR gene, which allows for selection of CHO cells that have been transfected with the vector using methotrexate selection / amplifi cation. The selected transformant host cells are cultured to allow for expression of the antibody heavy and light chains and the antibody is recovered from the culture medium.

[0344] Antibodies can also be produced by a transgenic animal. For example, U.S. Pat. No. 5,849,992 describes a method of expressing an antibody in the mammary gland of a transgenic mammal. A transgene is constructed that includes a milk-specific promoter and nucleic acids encoding the antibody of interest and a signal sequence for secretion. The milk produced by females of such transgenic mammals includes, secreted-therein, the antibody of interest. The antibody can be purified from the milk, or for some applications, used directly. Animals are also provided comprising one or more of the nucleic acids described herein.

[0345] The antibodies of the present disclosure can be isolated from inside or outside (such as from the medium) of the host cell and purified as substantially pure and homogenous antibodies. Methods for isolation and purification commonly used for antibody purification may be used for the isolation and purification of antibodies, and are not limited to any particular method. Antibodies may be isolated and purified by appropriately selecting and combining, for example, column chromatography, filtration, ultrafiltration, salting out, solvent precipitation, solvent extraction, distillation, immunoprecipitation, SDS- polyacrylamide gel electrophoresis, isoelectnc focusing, dialysis, and recrystallization. Chromatography includes, for example, affinity chromatography, ion exchange chromatography, hydrophobic chromatography, gel filtration, reverse-phase chromatography, and adsorption chromatography (Strategies for Protein Purification and Charactenzation: A Laboratory Course Manual. Ed Daniel R. Marshak et al., Cold Spring Harbor Laboratory

[0346] 93

[0347] SUBSTITUTE SHEET (RULE 26) Press, 1996). Chromatography can be carried out using liquid phase chromatography such as HPLC and FPLC. Columns used for affinity chromatography include protein A column and protein G column. Examples of columns using protein A column include Hyper D, POROS, and Sepharose FF (GE Healthcare Biosciences). The present disclosure also includes antibodies that are highly purified using these purification methods.

[0348] Indications

[0349] The anti-mutCALR antibody -drug conjugates of the present disclosure can inhibit the activity of mutCALR, inhibit the activity of one or more signaling pathways downstream of MPL, increase internalization of an anti-mutCALR antibody and / or an associated cytotoxin, inhibit oncogenic cell proliferation and / or cause oncogenic cell death, increase caspase induction, increase platelet count, prolong survival, inhibit dimerization of MPL, compete with MPL for binding to mutCALR, or a combination thereof.

[0350] As used herein, an anti-mutCALR antibody-drug conjugate that competes with MPL for binding to mutCALR means that the anti-mutCALR antibody-drug conjugate binds to mutCALR with a greater affinity than MPL, the anti-mutCALR antibody in the unconjugated form, or both. In some embodiments, the anti-mutCALR antibody-drug conjugate binds to mutCALR with about 10-fold, 50-fold, 100-fold, 500-fold or 1000-fold greater affinity than MPL, the anti-mutCALR antibody in the unconjugated form, or both. In some embodiments, the anti-mutCALR antibody-drug conjugate binds to mutCALR with an IC50 of about 10- fold, 50-fold, 100-fold, 500-fold or 1000-fold less than MPL, the anti-mutCALR antibody in the unconjugated form, or both. In some embodiments, the anti-mutCALR antibody-drug conjugate binds to mutCALR with an IC50 of between about 0.1 and 1 nM. In some embodiments, the anti-mutCALR antibody-drug conjugate binds to mutCALR with an IC50 of about 0.1 nM, 0.2 nM, 0.3 nM, 0.4 nM, 0.5 nM, 0.6 nM, 0.7 nM, 0.8 nM, 0.9 nM or 1 nM.

[0351] Accordingly, the antibody-drug conjugates or compositions described herein can be used in methods of inhibiting activity of mutCALR, inhibiting the activity of one or more signaling pathways downstream of MPL, increasing internalization of an anti-mutCALR antibody and / or an associated cytotoxin, inhibiting oncogenic cell proliferation and / or cause oncogenic cell death, increasing caspase induction, increasing platelet count, prolonging survival, inhibiting dimerization of MPL, inhibiting the binding of MPL to mutCALR, or a combination thereof in an individual / patient in need of the inhibition by administering an effective amount of an antibody-drug conjugate described herein.

[0352] 94

[0353] SUBSTITUTE SHEET (RULE 26) Non-limiting examples of signaling pathways downstream of MPL include Janus tyrosine kinase (JAK) and signal transducers and activators of transcription (STAT) signaling, mitogen-activated protein kinase (MEK) and extracellular signal-regulated kinase (ERK) signaling, serine / threonine kinase (AKT) signaling, and mammalian target of rapamycin (mTOR) signaling.

[0354] Another aspect of the present disclosure pertains to methods of treating a mutCALR- associated disease or disorder in an individual (e.g, patient) by administering to the individual in need of such treatment a therapeutically effective amount or dose of one or more antibody-drug conjugates of the present disclosure or a pharmaceutical composition thereof. Other aspects of the present disclosure provide an antibody-drug conjugate described herein for use in the treatment of a mutCALR-associated disease or disorder, or the use of an antibody-drug conjugate described herein for the manufacture of a medicament for the treatment of a mutCALR-associated disease or disorder. A mutCALR-associated disease or disorder can include any disease, disorder or condition that is directly or indirectly linked to expression or activity of mutCALR.

[0355] Another aspect of the present disclosure pertains to methods of treating a myeloproliferative neoplasm in an individual (e.g., patient) by administering to the individual in need of such treatment a therapeutically effective amount or dose of one or more antibodydrug conjugates of the present disclosure or a pharmaceutical composition thereof. Other aspects of the present disclosure provide an antibody-drug conjugate described herein for use in the treatment of a myeloproliferative neoplasm, or the use of an antibody-drug conjugate described herein for the manufacture of a medicament for the treatment of a myeloproliferative neoplasm.

[0356] Non-limiting examples of a myeloproliferative neoplasm include chronic myelogenous leukemia, polycythemia vera, primary myelofibrosis, essential thrombocythemia, chronic neutrophilic leukemia, acute myelogenus leukemia, chronic eosinophilic leukemia, chronic myelomonocytic leukemia, myeloproliferative neoplasm and myelodysplastic syndrome, including myelodysplastic syndrome with refractory' anaemia with ring sideroblasts, myelodysplastic syndrome with refractory anemia, and myelodysplastic syndrome with refractory anemia with excess blasts.

[0357] Anti-mutCALR antibody-drug conjugates disclosed herein can be used to treat, alone or in combination with other therapies, a myeloproliferative neoplasm, or can be used, alone or in combination with other therapies, for the manufacture of a medicament for the treatment of a myeloproliferative neoplasm. Non-limiting examples of other therapies include a JAK

[0358] 95

[0359] SUBSTITUTE SHEET (RULE 26) inhibitor (e.g., ruxolitinib, itaticinib), a PI3K inhibitor (e.g, parsaclisib), a standard of care therapy (e.g, IFN-alpha, hydroxyurea, thalidomide, lenalidomide, an androgen, an erythropoietin-stimulating agent, a chemotherapeutic agent), or a combination thereof.

[0360] Non-limiting examples of JAK inhibitors for use as described herein are provided in U.S. Pat No. 7,335,667; U.S. Pat. No. 9,359,358; U.S. Pat. No. 8,691,807; U.S. Pat. No. 9,181,271; and U.S. Pat. No. 9,034,884, each of which is incorporated herein by reference in its entirety.

[0361] Non-limiting examples of PI3K inhibitors for use as described herein are provided in U.S. Pat. No. 9,108,984; U.S. Pat. No. 9,062,055; U.S. Pat. No. 8,759,359; and U.S. Pat. No. 9,434,746, each of which is incorporated herein by reference in its entirety.

[0362] The terms “individual” or “patient” or “subject”, used interchangeably, refer to any animal, including mammals, preferably mice, rats, other rodents, rabbits, dogs, cats, swine, cattle, sheep, horses, or primates, and most preferably humans (i.e. , a human subject).

[0363] The phrase “therapeutically effective amount” refers to the amount of active antibodydrug conjugate that elicits the biological or medicinal response in a tissue, system, animal, individual or human that is being sought by a researcher, veterinarian, medical doctor or other clinician.

[0364] As used herein, the term “treating” or “treatment” refers to one or more of (1) inhibiting the disease; e.g., inhibiting a disease, condition or disorder in an individual who is experiencing or displaying the pathology or symptomatology of the disease, condition or disorder (i.e., arresting further development of the pathology and / or symptomatology); and (2) ameliorating the disease; e.g., ameliorating a disease, condition or disorder in an individual who is experiencing or displaying the pathology or symptomatology of the disease, condition or disorder i.e., reversing the pathology and / or symptomatology) such as decreasing the severity of disease.

[0365] In some embodiments, the antibody-drug conjugates of the invention are useful in preventing or reducing the risk of developing any of the diseases referred to herein; e.g., preventing or reducing the risk of developing a disease, condition or disorder in an individual who may be predisposed to the disease, condition or disorder but does not yet experience or display the pathology or symptomatology of the disease.

[0366] Pharmaceutical Compositions

[0367] An anti-mutCALR antibody-drug conjugate described herein can be formulated as a pharmaceutical composition for administration to a subject, e.g., to treat a disease or disorder

[0368] 96

[0369] SUBSTITUTE SHEET (RULE 26) described herein. Typically, a pharmaceutical composition includes a pharmaceutically acceptable carrier. As used herein, “pharmaceutically acceptable carrier" includes any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like that are physiologically compatible. The composition can include a pharmaceutically acceptable salt, e.g., an acid addition salt or a base addition salt (see, e.g, Berge, S.M., et al. (1977) J. Pharm. Sci. 66: 1-19).

[0370] Pharmaceutical formulation is a well-established art, and is further described, e.g, in Gennaro (ed.), Remington: The Science and Practice of Pharmacy, 20thed., Lippincott, Williams & Wilkins (2000) (ISBN: 0683306472); Ansel et al., Pharmaceutical Dosage Forms and Drug Delivery Systems, 7th Ed., Lippincott Williams & Wilkins Publishers (1999) (ISBN: 0683305727); and Kibbe (ed.), Handbook of Pharmaceutical Excipients American Pharmaceutical Association, 3rd ed. (2000) (ISBN: 091733096X).

[0371] The pharmaceutical compositions may be in a variety of forms. These include, for example, liquid, semi-solid and solid dosage forms, such as liquid solutions (e.g, injectable and infusible solutions), dispersions or suspensions, tablets, pills, powders, liposomes and suppositories. The preferred form can depend on the intended mode of administration and therapeutic application. Typically compositions for the agents described herein are in the form of injectable or infusible solutions.

[0372] The pharmaceutical compositions may be in a variety of forms. These include, for example, liquid, semi-solid and solid dosage forms, such as liquid solutions (e.g, injectable and infusible solutions), dispersions or suspensions, and liposomes. A suitable form can depend on the intended mode of administration and therapeutic application. Typically compositions for the agents described herein are in the form of injectable or infusible solutions.

[0373] The composition can be formulated as a solution, microemulsion, dispersion, liposome, or other ordered structure suitable for stable storage at high concentration or as a lyophilized preparation. Sterile injectable solutions can be prepared by incorporating an anti- mutCALR antibody-drug conjugate described herein in the required amount in an appropriate solvent with one or a combination of ingredients enumerated above, as required, followed by filtered sterilization. Generally, dispersions are prepared by incorporating an agent described herein into a sterile vehicle that contains a basic dispersion medium and the required other ingredients from those enumerated above. In the case of sterile powders for the preparation of sterile injectable solutions, the preferred methods of preparation are vacuum drying and freeze drying that yield a powder of an agent described herein plus any additional desired

[0374] 97

[0375] SUBSTITUTE SHEET (RULE 26) ingredient from a previously sterile-filtered solution thereof. Prolonged absorption of injectable compositions can be brought about by including in the composition an agent that delays absorption, for example, monostearate salts.

[0376] Any of the anti-mutCALR antibody-drug conjugates described herein can also be formulated as liposomes prepared by any suitable method known in the art.

[0377] Pharmaceutical compositions formulated for subcutaneous administration may be suitable in some circumstances because the subject can self-administer the pharmaceutical composition. Pharmaceutical formulations for subcutaneous administration can further comprise protein formulations comprising arginine-HCl, histidine, and / or polysorbate, which may confer increased potency, improved serum half-life, or enhanced solubility to the anti- mutCALR antibodies.

[0378] Administration

[0379] The anti-mutCALR antibody-drug conjugate can be administered to a subject, e.g, a subject in need thereof, for example, a human subject, by a variety of methods. For many applications, the route of administration can be intravenous injection or infusion (IV), subcutaneous injection (SC), intraperitoneally (IP), or intramuscular injection.

[0380] The route and / or mode of administration of the antibody can also be tailored for the individual case, e.g, by monitoring the subject, e.g, using tomographic imaging, e.g, to visualize a tumor.

[0381] The anti-mutCALR antibody-drug conjugate can be administered as a fixed dose, or in a mg / kg patient weight dose. The dose can also be chosen to reduce or avoid production of antibodies against the anti-mutCALR antibody-drug conjugate. Dosage regimens are adjusted to provide the desired response, e.g. , a therapeutic response or a combinatorial therapeutic effect. Generally, doses of the anti-mutCALR antibody-drug conjugate (and optionally a second agent) can be used in order to provide a subject with the agent in bioavailable quantities.

[0382] Dosage unit form or “fixed dose” or “flat dose” as used herein refers to physically discrete units suited as unitary dosages for the subjects to be treated; each unit contains a predetermined quantity of active compound calculated to produce the desired therapeutic effect in association with the required pharmaceutical carrier and optionally in association with the other agent. Single or multiple dosages may be given. Alternatively, or in addition, the antibody-drug conjugate may be administered via continuous infusion.

[0383] 98

[0384] SUBSTITUTE SHEET (RULE 26) The disclosure also provides a kit comprising one or more containers of an anti- mutCALR antibody or a pharmaceutical formulation thereof, optionally with one or more other prophylactic or therapeutic agents useful for the treatment of a disease or disorder, and optionally with instructions for using the anti-mutCALR antibody-drug conjugate or a pharmaceutical formulation thereof.

[0385] The instructions relating to the use of an anti-mutCALR antibody-drug conjugate generally include information as to dosage, dosing schedule, and route of administration for the intended treatment. The containers can be unit doses, bulk packages (e.g., multi-dose packages) or sub-unit doses. Instructions supplied in the kits of the disclosure are typically written instructions on a label or package insert. The label package insert indicates that an anti-mutCALR antibody-drug conjugate is used for treating, delaying the onset, and / or alleviating a myeloproliferative neoplasm.

[0386] The following are examples of the practice of the invention. They are not to be construed as limiting the scope of the invention in any way.

[0387] EXAMPLES

[0388] The following examples are provided to better illustrate the claimed invention and are not to be interpreted as limiting the scope of the invention. To the extent that specific materials are mentioned, it is merely for purposes of illustration and is not intended to limit the invention. One skilled in the art can develop equivalent means or reactants without the exercise of inventive capacity and without departing from the scope of the invention.

[0389] Example 1: Internalization of Anti-mutCALR Antibodies and Dose-Dependent Inhibition of Cell Proliferation

[0390] Two different cell lines were used to test the internalization and the effect on proliferation of various anti-mutCALR antibodies including Clone 6, Clone 55, and Clone 74. Two different isotype control antibodies and the anti-mutCALR antibody B3 were used as controls. The cell lines tested included MPL-6 cells expressing endogenous, mouse CALR and human TPO-R, and 4F2 cells expressing human mutCALRde152and human MPL.

[0391] Cells were seeded onto poly-D-lysine coated plates and allowed to attach overnight. Prior to the addition of antibodies, cells were treated with 20 pM dynasore, an inhibitor of dynamin GTPase function, which is required for some forms of endocytosis. Antibodies were labeled with FabFluor pH sensitive dye that produces a red fluorescence once the antibody is

[0392] 99

[0393] SUBSTITUTE SHEET (RULE 26) internalized. Internalization was measured by quantifying the area of fluorescent cells as a percentage of total cell area in the well. Proliferation was measured as the area of cells in each well (confluence) Plates were scanned hourly for 24h using the IncuCyte® live-cell analysis system.

[0394] Clone 74 (FIG. 1A), Clone 55 (FIG. IB), and Clone 6 (FIG. 1C) were internalized at the two highest concentrations (5 pg / mL and 2 pg / mL) tested in 4F2 cells, however, no internalization was observed in MPL6 cells. For each clone, antibody internalization was inhibited in the presence of dynasore (FIGs. 1A-1C). As shown in FIG. ID, proliferation of 4F2 cells, but not MPL6 cells, was inhibited in the presence of Clone 74, Clone 55, and Clone 6. No internalization or inhibition of cell proliferation was observed when cells were treated with the anti-mutCALR antibody B3 (FIG. IE and FIG. 1H) or the two isotype control antibodies (FIGs. 1F-1H).

[0395] Taken together, these results demonstrate that the anti-mutCALR antibodies Clone 74, Clone 55, and Clone 6 are internalized by cells expressing mutCALRde152and MPL, and that inhibition of proliferation of such cells by the anti-mutCALR antibodies is dependent upon antibody internalization.

[0396] Example 2: Antagonistic Effect of Anti-mutCALR Antibody is Dependent on Cell Internalization

[0397] Anti-mutCALR antibody (Clone 74) was conjugated with Alexa Fluor™ 647 fluorophore using lysine amine chemistry. The resulting fluorophore to protein ratio was 5.53. Ba / F3-TPOR / CALRde152cells, which are also referred to as 4F2 cells or BAF3 / 4F2 cells, were added in an amount of 500 cells / well to a retronectin-coated plate (Greiner Bio-One) and incubated at 37 °C for three days. Cells were then treated with 0.2 pg / mL Alexa Fluor- 647-Clone-74 and fixed with 8 % paraformaldehyde (Electron Microscopy Sciences) at various time points. Cells were washed twice with PBS, incubated in 2 pg / mL Hoechst 33342 (Thermo Fisher) for 1 hour, followed by additional washing and addition of PBS. The plate was then scanned at 20X magnification using the Celllnsight™ CX7 LZR High Content Analysis Platform (Thermo Fisher). Nuclei stained with Hoechst 33342 were detected at 405 nm and used to identify cells and Alexa Fluor-647 -Clone-74 was detected at 650 nm. A minimum of 1000 cells from each well were analyzed. Representative data from each time point were compiled for display.

[0398] Alternatively, cells (5000 cells / well) were added to retronectin-coated, clear, cell culture plates (Coming) and incubated at 37 °C overnight. Clone 74 and isotype were labeled

[0399] 100

[0400] SUBSTITUTE SHEET (RULE 26) as a 20 pg / mL stock with IncuCyte® Human Fabfluor-pH antibody labeling dye (Sartorius) according to the manufacturer’s instructions. Cells were then pretreated with either 20 pM dynasore (EMD Millipore) or DMSO. Next, the labeled antibodies were added to a final concentration of 5 pg / mL and the plate was placed in the IncuCyte® S3 Live-Cell Analysis Instrument (Sartorius) and scanned at 10X magnification every hour for 24 hours using brightfield and red fluorescence to monitor proliferation and antibody internalization, respectively. Proliferation was calculated as cell confluence and expressed as percent. Antibody internalization was measured at the percentage of cells containing red fluorescence (internalized antibody) among the cell population, then normalized to the signal at time 0.

[0401] Internalization of Clone 74 in Ba / F3-CALRde152cells was time-dependent (FIG. 2A).

[0402] The endocytosis of Clone 74 was dynamin-dependent as the presence of the dynamin inhibitor dynasore completely abrogated the internalization of Clone 74 (FIG. 2B), abrogated mutCALR-induced inhibition of cell proliferation (FIG. 2C), and inhibited mutCALR- induced caspase activation (FIG. 2D). Dynasore treatment showed only a partial effect in reducing mutCALR-induced inhibition of pSTAT5 (FIG. 2E).

[0403] Internalization of Clone 74 was analyzed using confocal microscopy. Ba / F3- TPOR / CALRde152cells were treated with Alexa Fluor™-647-Clone-74 for 18 h and microscopy images were collected. As shown in FIG. 2F, confocal microscopy confirmed the complete endocytosis of Clone 74 at 18 hours post-cell treatment.

[0404] Confocal microscopy was also used to analyze co-localization of Clone 74 with the lysosomal marker LAMP-1. Ba / F3-TPOR / CALRde152cells were pre-treated with Alexa Fluor™-647-Clone-74 for 2 h on ice, followed by washing and additional incubation for 6 h at 37 °C. Lysosomal compartments were stained with anti-LAMP-1, and cells were evaluated by microscopy. As shown in FIG. 2G, confocal microscopy images showed that internalized Clone 74 co-localized with the lysosomal marker LAMP-1, suggesting that the Clone 74 / mutCALR / TPOR complex is localized to the cell lysosome upon endocytosis and is likely targeted for degradation.

[0405] Flow cytometry analysis of the surface expression of TPOR in the presence of anti- mutCALR antibody (Clone 74) or an isotype control was performed. Ba / F3-TPOR and Ba / F3-TPOR / mutCALRde152cells were treated overnight with 10 pg / mL of anti-mutCALR antibody (Clone 74) or isotype and the surface expression of TPOR was analyzed by flow cytometry. As shown in FIG. 3A, flow cytometry analysis confirmed the Clone 74-induced internalization and degradation of TPOR in Ba / F3 cells expressing mutCALR.

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[0407] SUBSTITUTE SHEET (RULE 26) Western blot analysis of TPOR and pSTAT5 expression in the presence of anti- mutCALR antibody (Clone 74) or an isotype control was performed. HAP1-JAK2 KO cells were transiently transfected with wtJAK2, TPOR and CALRde152, treated with 10 pg / mL of anti -mutC ALR antibody (Clone 74) or isotype for 2 h or overnight and then evaluated by Western blot regarding expression of TPOR and pSTAT5. P-actin staining was used as a protein loading control. As shown in FIG. 3B, Western blot analysis confirmed the Clone 74- induced internalization and degradation of TPOR in HAP1 cells expressing mutCALR.

[0408] Immunofluorescence analysis of internalization of TPOR / mutCALR complex upon anti-mutCALR antibody (Clone 74) binding was performed. CD34+cells from a MF patient expressing CALRde152were treated with 2 pg / mL of anti-mutCALR antibody (Clone 74) for 12 h. Internalization of TPOR was assessed by a fluorescence labeled TPOR antibody after cells were fixed and permeabilized. Treatment with 10 ng / ml of TPO was used as a positive control of TPOR internalization. Image analysis was performed using colocalization module in ZEN software on a pixel by pixel basis. Single label control samples were used to set up compensation crosshair. As shown in FIG. 3C, confocal microscopy analysis confirmed the Clone 74-induced internalization and degradation of TPOR in primary CD34+cells expressing mutCALR.

[0409] Taken together, these results demonstrate that the anti-mutCALR antibody Clone 74 is internalized and that the antagonistic effects of the antibody are dependent on its cellular internalization.

[0410] Example 3: Preparation of Anti-mutCALR Antibody-Drug Conjugates

[0411] This Example describes conjugation of an anti-mutCALR antibody (Clone 6) to monomethyl auristatin E (MMAE) or SG3249 (also known as tesirine).

[0412] MMAE Conjugation

[0413] Anti-mutCALR antibody (Clone 6) and a control antibody were labelled with MMAE using a maleimide-activated valine-citruline p-aminobenzylcarbamate (VC-PAB) linker.

[0414] Samples were prepared as shown below:

[0415] 102

[0416] SUBSTITUTE SHEET (RULE 26) A solution of TCEP (tris(2-carboxyethyl)phosphine) was freshly prepared and 4 molar equivalents of TCEP was added to each sample to selectively reduce cysteine disulfide bridges. Samples were incubated at 37 °C for 80 minutes.

[0417] After incubation with TCEP, 5 molar equivalents of mc-vc-PAB-MMAE in DMA (dimethylacetamide) was added to each sample. Samples were incubated at room temperature for 2 hours. After 2 hours, samples were analyzed for aggregation by size-exclusion chromatography. The percent monomer and aggregation in each sample are shown below:

[0418] Samples were dialyzed to remove excess mc-vc-PAB-MMAE. Conjugated antibody was purified by size-exclusion chromatography using a HiLoad® 26 / 600 Superdex® 200 pg column and lx PBS as the mobile phase and concentrated into lx PBS using a 10 kDa molecular weight cut-off (MWCO) membrane.

[0419] Concentration of antibody-drug conjugate was determined by spectroscopic absorbance at 280 nm. Drug-per-antibody ratio (DAR) was determined using liquid chromatography-mass spectrometry (LC / MS) and reverse-phase high-performance liquid chromatography (RP-HPLC). Characteristics of the antibody-drug conjugates are shown below:

[0420] SG3249 Conjugation

[0421] Anti-mutCALR antibody (Clone 6) and a control antibody were labelled with

[0422] SG3249. Samples were prepared as shown below:

[0423] A solution of TCEP was freshly prepared and 3.2 molar equivalents of TCEP was added to each sample to selectively reduce cysteine disulfide bridges. Samples were incubated at 37 °C for 80 minutes.

[0424] 103

[0425] SUBSTITUTE SHEET (RULE 26) After incubation with TCEP, 4 molar equivalents of SG3249 in DMA was added to each sample. Samples were incubated at room temperature for 2 hours. After 2 hours, samples were analyzed for aggregation by size-exclusion chromatography. The percent monomer and aggregation in each sample are shown below:

[0426] Samples were dialyzed to remove excess SG3249. Conjugated antibody was purified by size-exclusion chromatography using a HiLoad® 26 / 600 Superdex® 200 pg column and lx PBS as the mobile phase and concentrated into lx PBS using a 30 kDa MWCO membrane.

[0427] Concentration of antibody-drug conjugate was determined by spectroscopic absorbance at 280 nm. Drug-per-antibody ratio (DAR) was determined using liquid chromatography-mass spectrometry (LC / MS) and reverse-phase high-performance liquid chromatography (RP-HPLC). Characteristics of the antibody-drug conjugates are shown below:

[0428] Example 4: Improved Properties for Anti-mutCALR Antibody-Drug Conjugate Compared to Unconjugated Anti-mutCALR Antibody

[0429] The internalization rate of an anti-mutCALR antibody (Clone 6) conjugated to SG3249 (referred to as Clone 6-PBD) and unconjugated Clone 6 was tested as described in

[0430] Examples 1-2. An isotype control antibody (Isotype-PBD) and unconjugated isotype control antibody were used as controls.

[0431] As shown in FIG. 4A, the internalization rate of Clone 6-PBD in cells expressing Type 1 mutant CALR was significantly increased compared to unconjugated Clone 6 or isotype controls. As shown in FIG. 4B, caspase induction in cells was significantly increased in the presence of Clone 6-PBD compared to unconjugated Clone 6 or isotype controls. As shown in FIG. 4C, increased inhibition of cell proliferation was observed in the presence of Clone 6-PBD compared to unconjugated Clone 6 or isotype controls.

[0432] 104

[0433] SUBSTITUTE SHEET (RULE 26) Taken together, these results demonstrate that the anti-mutCALR antibody Clone 6 conjugated to PBD has improved internalization, caspase induction, and inhibition of cell proliferation compared to the unconjugated anti-mutCALR antibody.

[0434] Example 5: Anti-mutCALR Antibody Conjugated to PBD Increased Cell Killing Compared to Anti-mutCALR Antibody Conjugated to MMAE

[0435] Cell killing by an anti-mutCALR antibody (Clone 6) conjugated to either SG3249 or monomethyl auristatin E (MMAE) was tested. The conjugates were prepared as described in Example 3.

[0436] BAF3 / 4F2 cells (Type 1 mutant CALR) or BAF3 / 3D3 cells (Type 2 mutant CALR) were plated at 5000 cells / well in a 96-well plate. Cells were then treated with anti-mutCALR antibody (Clone 6) conjugated to PBD or MMAE, an isotype control antibody conjugated to PBD or MMAE, or unconjugated anti-mutCALR antibody, for 72 hours. Following incubation, cells were mixed 1 : 1 with Cell Titer-Gio (Promega) and measured for luminescence via SpectraMAX plate reader. Percent viability was normalized to the viability of an IgGl control.

[0437] As shown in FIG. 5A, killing of cells expressing Type 1 mutant CALR was increased in the presence of Clone 6-PBD compared to unconjugated Clone 6 or isotype antibody conjugated to PBD. By contrast, as shown in FIG. 5B, cell killing was increased in the presence of unconjugated Clone 6 compared to Clone 6-MMAE or isotype antibody- conjugated to MMAE. Similar results were observed in cells expressing Type 2 mutant CALR (FIGs. 6A-6B). A summary of EC50 values for cell killing in the presence of Clone 6 conjugated to PBD or MMAE are shown below:

[0438] As shown in FIGs. 7A-7B, cell killing by anti-mutCALR antibody (Clone 6) conjugated to PBD was equally potent in cells which were cultured either with 10 ng / mL thrombopoietin (TPO) (FIG. 7A) or without TPO (FIG. 7B). In contrast, the activity of unconjugated antibody was reduced in the presence of 10 ng / nL TPO (FIGs. 7A-7B). These

[0439] 105

[0440] SUBSTITUTE SHEET (RULE 26) data demonstrate that cell killing with the mutCALR antibody conjugated to PBD (Clone 6) is not dependent on the presence of 10 ng / mL TPO.

[0441] Taken together, these results demonstrate that the anti-mutCALR antibody Clone 6 conjugated to PBD more effectively kills cells with Type 1 or Type 2 mutant CALR than the anti-mutCALR antibody conjugated to MMAE or unconjugated anti-mutCALR antibody.

[0442] Example 6: Effect of Anti-mutCALR Antibody Conjugates in Mice Injected with Tumor Cells Expressing Type 1 Mutant CALR

[0443] To test the functional ability of an anti-mutCALR antibody (Clone 6) conjugate in vivo, the antibody-drug conjugate was evaluated in a mouse model of tumor growth. Female NSG mice, aged 6 to 8 weeks, were inoculated via tail vein with 1x105BA / F3 4F2 cells on Day 0. Six days following inoculation, the mice were either administered anti-mutCALR antibody (Clone 6)-PBD (single dose i.v. on Day 6), unconjugated Clone 6 (single dose i.v. on Day 6), unconjugated Clone 6 at 10 mg / kg i.p. (Day 6, 9, and 12), or ruxohtinib twice daily per orally for 7 days. Hematology analysis for tumor cells and platelet count was conducted on the Sysmex instrument.

[0444] As shown in FIG. 8A, tumor cell growth was inhibited in mice administered anti- mutCALR antibody (Clone 6) conjugated to PBD compared to mice administered dose matched unconjugated antibody. As shown in FIG. 8B, platelet count was increased in mice administered anti-mutCALR antibody (Clone 6) conjugated to PBD compared to mice administered dose matched unconjugated antibody. Platelet counts in mice administered anti- mutCALR antibody (Clone 6) were similar to mice administered ruxolitinib. As shown in FIG. 9, mice administered a single dose of Clone 6-PBD at 1 mg / kg exhibited increased survival compared to mice administered 3 doses of unconjugated antibody at 10 mg / kg.

[0445] Taken together, these results demonstrate that administration of the anti-mutCALR antibody Clone 6 conjugated to PBD inhibited tumor growth, increased platelet count, and prolonged survival of mice compared to administration of the unconjugated anti-mutCALR antibody.

[0446] OTHER EMBODIMENTS

[0447] It is to be understood that while the invention has been described in conjunction with the detailed description thereof, the foregoing description is intended to illustrate and not limit the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages, and modifications are within the scope of the following claims.

[0448] 106

[0449] SUBSTITUTE SHEET (RULE 26)

Claims

WHAT IS CLAIMED IS:

1. An antibody-drug conjugate comprising: an antibody that binds to human mutant calreticulin (CALR); and a pyrrolobenzodiazepine dimer, wherein the antibody comprises a heavy chain variable region (VH) comprising a VH CDR1; a VH CDR2; and a VH CDR3; wherein: the VH CDR1 comprises the amino acid sequence ELSMQ (SEQ ID NO:1); the VH CDR2 comprises the amino acid sequence GFDPDDX101ETMYAEX102X103QG (SEQ ID NO: 102); wherein X101 is D or G; wherein X102 is K or R; and wherein X103 is F or L; the VH CDR3 is SPGYDFFDY (SEQ ID NO: 18); wherein the antibody comprises a light chain variable region (VL) comprising a VL CDR1, a VL CDR2, and a VL CDR3, wherein: the VL CDR1 comprises the amino acid sequence GGX104X105X106GX107X108X109VX110 (SEQ ID NO: 103), wherein X104 is N, D, or S; wherein X105 is Y, N, or D; wherein X106 is I or T; wherein X107 is S, D, I, R, or T; wherein Xios is K, E, or I; wherein X109 is S, I, R, G, N, or A; and wherein X110 is H, F, or N; the VL CDR2 comprises the amino acid sequence DDX111DRPX112 (SEQ ID NO: 104), wherein X111 is G, S, or R; and wherein X112 is S or L; and the VL CDR3 comprises the amino acid sequence QVWDX113X114X115DX116X117X118 (SEQ ID NO: 105), wherein X113 is S or A; wherein X114 is I or S; wherein X115 is S, I, or N; wherein X116 is H, L, or Q; wherein X117 is V or L; and wherein Xus is V or I or the VL CDR1 comprises the amino acid sequence TGTSSDVGGYNYVS (SEQ ID NO:30); the VL CDR2 comprises the amino acid sequence X119VSX120RPS (SEQ ID NO: 106); wherein X119 is E or D; and wherein X120 is N or K; and the VL CDR3 comprises the amino acid sequence QVWDSSNDLLI (SEQ ID NO:71).

2. The antibody-drug conjugate of claim 1, wherein: the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1;107SUBSTITUTE SHEET (RULE 26)the VH CDR2 comprises the amino acid sequence of any one of SEQ ID NOs:7-10 and 92-95; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of any one of SEQ ID NOs:26-41 and 118; the VL CDR2 comprises the amino acid sequence of any one of SEQ ID NOs:53-58; and the VL CDR3 comprises the amino acid sequence of any one of SEQ ID NOs:69-75.

3. The antibody-drug conjugate of claim 1, wherein: the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:26; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:53; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:69; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:27; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:70; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 8; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ108SUBSTITUTE SHEET (RULE 26)ID NO:29; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:72; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:29; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:73; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:56; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:31; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:57; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 9; the VH CDR3 comprises the amino109SUBSTITUTE SHEET (RULE 26)acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:32; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:33; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:73; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:34; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:56; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:27; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:35; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:36; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:72; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:58; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; noSUBSTITUTE SHEET (RULE 26)the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:37; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:38; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:32; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:39; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:40; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 10; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ illSUBSTITUTE SHEET (RULE 26)ID N0:41; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:70; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:40; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:56; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:74; the VH CDR1 comprises the amino acid sequence of SEQ ID NO:1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:75; the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO: 118; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 92; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 93; the VH CDR3 comprises the amino112SUBSTITUTE SHEET (RULE 26)acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 94; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 95; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 92; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 93; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 94; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71; or the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO: 95; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:28; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:54; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71.113SUBSTITUTE SHEET (RULE 26)4. The antibody-drug conjugate of claim 1, wherein the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID N0:71.

5. The antibody-drug conjugate of claim 1, wherein the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:95; the VH CDR3 comprises the amino acid sequence of SEQ ID NO:18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID N0:71.

6. The antibody-drug conjugate of any one of claims 1-5, wherein: the VH is at least 80% identical to the amino acid sequence of any one of SEQ ID NOs: 165-174 and 188-208; and the VL is at least 80% identical to the amino acid sequence of any one of SEQ ID NOs:264-292 and 315-316.

7. The antibody-drug conjugate of any one of claims 1-5, wherein: the VH comprises the amino acid sequence of any one of SEQ ID NOs: 165-174 and 188-208; and the VL comprises the amino acid sequence of any one of SEQ ID NOs:264-292 and 315-316.

8. The antibody-drug conjugate of claim 1, wherein the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:264; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:265; the VH comprises the amino acid sequence of SEQ ID NO: 166 and the VL comprises the amino acid sequence of SEQ ID NO:266;114SUBSTITUTE SHEET (RULE 26)the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:266; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:267; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:269; the VH comprises the amino acid sequence of SEQ ID NO: 167 and the VL comprises the amino acid sequence of SEQ ID NO:270; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:271; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:272; the VH comprises the amino acid sequence of SEQ ID NO: 168 and the VL comprises the amino acid sequence of SEQ ID NO:273; the VH comprises the amino acid sequence of SEQ ID NO: 169 and the VL comprises the amino acid sequence of SEQ ID NO:274; the VH comprises the amino acid sequence of SEQ ID NO: 170 and the VL comprises the amino acid sequence of SEQ ID NO:275; the VH comprises the amino acid sequence of SEQ ID NO: 171 and the VL comprises the amino acid sequence of SEQ ID NO:276; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:277; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:278; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:279; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:280; the VH comprises the amino acid sequence of SEQ ID NO: 172 and the VL comprises the amino acid sequence of SEQ ID NO:281; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:282;115SUBSTITUTE SHEET (RULE 26)the VH comprises the amino acid sequence of SEQ ID NO: 173 and the VL comprises the amino acid sequence of SEQ ID NO:283; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:284; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:285; the VH comprises the amino acid sequence of SEQ ID NO: 174 and the VL comprises the amino acid sequence of SEQ ID NO:286; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:287; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:288; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:289; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:290; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:291; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:292; the VH comprises the amino acid sequence of SEQ ID NO: 188 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 189 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 190 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 191 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 192 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 193 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 194 and the VL comprises the amino acid sequence of SEQ ID NO:315;116SUBSTITUTE SHEET (RULE 26)the VH comprises the amino acid sequence of SEQ ID NO: 195 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 196 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 188 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 189 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 190 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 191 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 192 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 193 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 194 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 195 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 196 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 197 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO: 198 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO: 199 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID N0:200 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO:201 and the VL comprises the amino acid sequence of SEQ ID NO:268;117SUBSTITUTE SHEET (RULE 26)the VH comprises the amino acid sequence of SEQ ID NO: 197 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 198 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 199 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID N0:200 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO:201 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 197 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 198 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 199 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID N0:200 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO:201 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO: 170 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO:202 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO:203 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO:204 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO:205 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO:206 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO:207 and the VL comprises the amino acid sequence of SEQ ID NO:268;118SUBSTITUTE SHEET (RULE 26)the VH comprises the amino acid sequence of SEQ ID NO:208 and the VL comprises the amino acid sequence of SEQ ID NO:268; the VH comprises the amino acid sequence of SEQ ID NO: 170 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO:202 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO:203 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO:204 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO:205 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO:206 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO:207 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO:208 and the VL comprises the amino acid sequence of SEQ ID NO:315; the VH comprises the amino acid sequence of SEQ ID NO: 170 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO:202 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO:203 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO:204 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO:205 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO:206 and the VL comprises the amino acid sequence of SEQ ID NO:316; the VH comprises the amino acid sequence of SEQ ID NO:207 and the VL comprises the amino acid sequence of SEQ ID NO:316; or the VH comprises the amino acid sequence of SEQ ID NO:208 and the VL comprises the amino acid sequence of SEQ ID NO:316.119SUBSTITUTE SHEET (RULE 26)9. The antibody-drug conjugate of claim 1, wherein the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:268.

10. The antibody-drug conjugate of claim 1, wherein the VH comprises the amino acid sequence of SEQ ID NO: 196 and the VL comprises the amino acid sequence of SEQ ID NO:315.

11. The antibody-drug conjugate of claim 1, wherein the antibody comprises: a heavy chain comprising the amino acid sequence of any one of SEQ ID NOs: 119- 128 and 142-164; and a light chain comprising the amino acid sequence of any one of SEQ ID NOs:209-237 and 260-261.

12. The antibody-drug conjugate of claim 11 : wherein the heavy chain comprises the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:209; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:210; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 120 and the light chain comprises the amino acid sequence of SEQ ID NO:211; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:211; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:212; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:214; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 121 and the light chain comprises the amino acid sequence of SEQ ID NO:215; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:216;120SUBSTITUTE SHEET (RULE 26)wherein the heavy chain compnses the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:217; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 122 and the light chain comprises the amino acid sequence of SEQ ID NO:218; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 123 and the light chain comprises the amino acid sequence of SEQ ID NO:219; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 124 and the light chain comprises the amino acid sequence of SEQ ID NO:220; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 125 and the light chain comprises the amino acid sequence of SEQ ID NO:221; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:222; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:223; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:224; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:225; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 126 and the light chain comprises the amino acid sequence of SEQ ID NO:226; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:227; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 127 and the light chain comprises the amino acid sequence of SEQ ID NO:228; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:229; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:230; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 128 and the light chain comprises the amino acid sequence of SEQ ID NO:231; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:232; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:233;121SUBSTITUTE SHEET (RULE 26)wherein the heavy chain compnses the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:234; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:235; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:236; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:237; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 142 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 143 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 144 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 145 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 146 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 147 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 148 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 149 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 150 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 142 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 143 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 144 and the light chain comprises the amino acid sequence of SEQ ID NO:260;122SUBSTITUTE SHEET (RULE 26)wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 145 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 146 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 147 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 148 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 149 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 150 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 142 and the light chain comprises the amino acid sequence of SEQ ID NO:261; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 143 and the light chain comprises the amino acid sequence of SEQ ID NO:261; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 144 and the light chain comprises the amino acid sequence of SEQ ID NO:261; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 145 and the light chain comprises the amino acid sequence of SEQ ID NO:261; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 146 and the light chain comprises the amino acid sequence of SEQ ID NO:261; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 147 and the light chain comprises the amino acid sequence of SEQ ID NO:261; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 148 and the light chain comprises the amino acid sequence of SEQ ID NO:261; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 149 and the light chain comprises the amino acid sequence of SEQ ID NO:261; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 150 and the light chain comprises the amino acid sequence of SEQ ID NO:261; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:261;123SUBSTITUTE SHEET (RULE 26)wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 152 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 153 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 154 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 155 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 156 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 152 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 153 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 154 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 155 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 156 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 152 and the light chain comprises the amino acid sequence of SEQ ID NO:261; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 153 and the light chain comprises the amino acid sequence of SEQ ID NO:261; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 154 and the light chain comprises the amino acid sequence of SEQ ID NO:261; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 155 and the light chain comprises the amino acid sequence of SEQ ID NO:261; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 156 and the light chain comprises the amino acid sequence of SEQ ID NO:261; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 157 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO:158 and the light chain comprises the amino acid sequence of SEQ ID NO:213;124SUBSTITUTE SHEET (RULE 26)wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 159 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 160 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO:161 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 162 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 163 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 164 and the light chain comprises the amino acid sequence of SEQ ID NO:213; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 157 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO:158 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 159 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 160 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO:161 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 162 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 163 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 164 and the light chain comprises the amino acid sequence of SEQ ID NO:260; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 157 and the light chain comprises the amino acid sequence of SEQ ID NO:261; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO:158 and the light chain comprises the amino acid sequence of SEQ ID NO:261; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 159 and the light chain comprises the amino acid sequence of SEQ ID NO:261;125SUBSTITUTE SHEET (RULE 26)wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 160 and the light chain comprises the amino acid sequence of SEQ ID NO:261; wherein the heavy chain compnses the amino acid sequence of SEQ ID NO:161 and the light chain comprises the amino acid sequence of SEQ ID NO:261; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 162 and the light chain comprises the amino acid sequence of SEQ ID NO:261; wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 163 and the light chain comprises the amino acid sequence of SEQ ID NO:261; or wherein the heavy chain compnses the amino acid sequence of SEQ ID NO: 164 and the light chain comprises the amino acid sequence of SEQ ID NO:261.

13. The antibody-drug conjugate of claim 11, wherein the heavy chain comprises the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:213.

14. The antibody-drug conjugate of claim 11, wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 142 and the light chain comprises the amino acid sequence of SEQ ID NO:213.

15. The antibody-drug conjugate of claim 11, wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:260.

16. The antibody-drug conjugate of any one of claims 1-15, wherein the human mutant CALR is human Type 1 mutant CALR comprising the amino acid sequence of SEQ ID NO:320.

17. The antibody-drug conjugate of any one of claims 1-15, wherein the human mutant CALR is human Type 2 mutant CALR comprising the amino acid sequence of SEQ ID NO:321.

18. The antibody-drug conjugate of any one of claims 1-17, wherein the antibody is a human or humanized antibody.126SUBSTITUTE SHEET (RULE 26)19. The antibody-drug conjugate of any one of claims 1-18, wherein the antibody is a full-length antibody.

20. The antibody-drug conjugate of any one of claims 1-19, wherein the antibody is an IgGl, IgG2, IgG3 or IgG4 antibody.

21. The antibody-drug conjugate of claim 20, wherein the antibody is an IgGl antibody.

22. The antibody-drug conjugate of any one of claims 1-18, wherein the antibody is a bispecific antibody, a biparatopic antibody, a single chain antibody (scFv), an Fab fragment, an F(ab’)2 fragment, an Fab’ fragment, an Fsc fragment, an Fv fragment, an scFv, an sc(Fv)2, or a diabody.

23. The antibody-drug conjugate of any one of claims 1-22, wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of PBD-1, SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

24. The antibody-drug conjugate of any one of claims 1-23, wherein the pyrrolobenzodiazepine dimer is linked to a linker to form a payload.

25. The antibody-drug conjugate of claim 24, wherein the linker is cleavable.

26. The antibody-drug conjugate of claim 24 or claim 25, wherein the linker comprises a maleimide linker.

27. The antibody-drug conjugate of any one of claims 24-26, wherein the linker comprises a peptide.

28. The antibody-drug conjugate of any one of claims 24-27, wherein the linker comprises a polyethylene glycol spacer.

29. The antibody-drug conjugate of any one of claims 24-28, wherein the linker is selected from the group consisting of:127SUBSTITUTE SHEET (RULE 26)L-2; wherein - indicates the bond between the linker and the pyrrolobenzodiazepine, and wherein the linker is connected to the antibody via the cyclooctynyl moiety.

30. The antibody-drug conjugate of any one of claims 24-29, wherein the payload is selected from the group consisting of PL-1, PL-2, PL-3, SG3400, SG3227, SG3249, SG3710, SG3259, SG3584, SG3203, SG3231, SG3451, and SGD-1910.

31. The antibody-drug conjugate of any one of claims 1-30, wherein the pyrrolobenzodiazepine dimer comprises a cap.

32. The antibody-drug conjugate of claim 31, wherein the cap is cleavable.

33. The antibody-drug conjugate of claim 31 or claim 32, wherein the cap comprises a glucuronide group or derivative thereof or a galactoside group or derivative thereof.

34. The antibody-drug conjugate of any one of claims 31-33, wherein the payload is SG3600.

35. The antibody-drug conjugate of any one of claims 1-34, wherein the antibody-drug conjugate has a drug loading of two pyrrolobenzodiazepine dimers per antibody.128SUBSTITUTE SHEET (RULE 26)36. The antibody-drug conjugate of claim 1, wherein the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of PBD-1, SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

37. The antibody-drug conjugate of claim 1, wherein the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71, and wherein the pyrrolobenzodiazepine dimer is SG2000.

38. The antibody-drug conjugate of claim 1, wherein the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:7; the VH CDR3 comprises the amino acid sequence of SEQ ID NO: 18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71, and wherein the pyrrolobenzodiazepine dimer is SG3199.

39. The antibody-drug conjugate of claim 1, wherein the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:268, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting ofPBD-1, SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

40. The antibody-drug conjugate of claim 1, wherein the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:268, and wherein the pyrrolobenzodiazepine dimer is SG2000.129SUBSTITUTE SHEET (RULE 26)41. The antibody-drug conjugate of claim 1, wherein the VH comprises the amino acid sequence of SEQ ID NO: 165 and the VL comprises the amino acid sequence of SEQ ID NO:268, and wherein the pyrrolobenzodiazepine dimer is SG3199.

42. The antibody-drug conjugate of claim 11, wherein the heavy chain comprises the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:213, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of PBD-1, SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

43. The antibody-drug conjugate of claim 11, wherein the heavy chain comprises the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:213, and wherein the pyrrolobenzodiazepine dimer is SG2000.

44. The antibody-drug conjugate of claim 11, wherein the heavy chain comprises the amino acid sequence of SEQ ID NO:119 and the light chain comprises the amino acid sequence of SEQ ID NO:213, and wherein the pyrrolobenzodiazepine dimer is SG3199.

45. The antibody-drug conjugate of claim 11, wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 142 and the light chain comprises the amino acid sequence of SEQ ID NO:213, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of PBD-1, SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

46. The antibody-drug conjugate of claim 11, wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 142 and the light chain comprises the amino acid sequence of SEQ ID NO:213, and wherein the pyrrolobenzodiazepine dimer is SG2000.

47. The antibody-drug conjugate of claim 11, wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 142 and the light chain comprises the amino acid sequence of SEQ ID NO:213, and wherein the pyrrolobenzodiazepine dimer is SG3199.

48. The antibody-drug conjugate of claim 1, wherein the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ130SUBSTITUTE SHEET (RULE 26)ID NO:95; the VH CDR3 comprises the amino acid sequence of SEQ ID NO:18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of PBD-1, SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

49. The antibody-drug conjugate of claim 1, wherein the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:95; the VH CDR3 comprises the amino acid sequence of SEQ ID NO:18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71, and wherein the pyrrolobenzodiazepine dimer is SG2000.

50. The antibody-drug conjugate of claim 1, wherein the VH CDR1 comprises the amino acid sequence of SEQ ID NO: 1; the VH CDR2 comprises the amino acid sequence of SEQ ID NO:95; the VH CDR3 comprises the amino acid sequence of SEQ ID NO:18; the VL CDR1 comprises the amino acid sequence of SEQ ID NO:30; the VL CDR2 comprises the amino acid sequence of SEQ ID NO:55; and the VL CDR3 comprises the amino acid sequence of SEQ ID NO:71, and wherein the pyrrolobenzodiazepine dimer is SG3199.

51. The antibody-drug conjugate of claim 1, wherein the VH comprises the amino acid sequence of SEQ ID NO: 196 and the VL comprises the amino acid sequence of SEQ ID NO:315, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting ofPBD-1, SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

52. The antibody-drug conjugate of claim 1, wherein the VH comprises the amino acid sequence of SEQ ID NO: 196 and the VL comprises the amino acid sequence of SEQ ID NO:315, and wherein the pyrrolobenzodiazepine dimer is SG2000.

53. The antibody-drug conjugate of claim 1, wherein the VH comprises the amino acid sequence of SEQ ID NO: 196 and the VL comprises the amino acid sequence of SEQ ID NO:315, and wherein the pyrrolobenzodiazepine dimer is SG3199.131SUBSTITUTE SHEET (RULE 26)54. The antibody-drug conjugate of claim 11, wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:260, and wherein the pyrrolobenzodiazepine dimer is selected from the group consisting of PBD-1, SG2000, SG2219, SG3199, SG3312, SG3650, SG2057, and SGD-1882.

55. The antibody-drug conjugate of claim 11, wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:260, and wherein the pyrrolobenzodiazepine dimer is SG2000.

56. The antibody-drug conjugate of claim 11, wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:260, and wherein the pyrrolobenzodiazepine dimer is SG3199.

57. The antibody-drug conjugate of claim 24, wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:260, the linker is a group of formula L-l, and the pyrrolobenzodiazepine dimer is SG3199.

58. The antibody-drug conjugate of claim 24, wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 151 and the light chain comprises the amino acid sequence of SEQ ID NO:260, the linker is a group of formula L-2, and the pyrrolobenzodiazepine dimer is PBD-1.

59. A pharmaceutical composition comprising the antibody-drug conjugate of any one of claims 1-58, and a pharmaceutically acceptable carrier.

60. A method of treating a myeloproliferative neoplasm in a human subject in need thereof, the method comprising administering to the human subject an effective amount of the antibody-drug conjugate of any one of claims 1-58 or the pharmaceutical composition of claim 59.

61. The method of claim 60, wherein the myeloproliferative neoplasm is selected from the group consisting of chronic myelogenous leukemia, polycythemia vera, primary132SUBSTITUTE SHEET (RULE 26)myelofibrosis, essential thrombocythemia, chronic neutrophilic leukemia, acute myelogenus leukemia, chronic eosinophilic leukemia, chronic myelomonocytic leukemia, myeloproliferative neoplasm and myelodysplastic syndrome, including myelodysplastic syndrome with refractory anaemia with ring sideroblasts, myelodysplastic syndrome with refractory anemia, and myelodysplastic syndrome with refractory anemia with excess blasts.

62. The method of claim 60 or claim 61, further comprising administering to the human subject an additional therapy selected from the group consisting of a Janus tyrosine kinase (JAK) inhibitor, a phosphoinositide 3-kinase (PI3K) inhibitor, a standard of care therapy, or a combination thereof.

63. The method of claim 62, wherein the JAK inhibitor is ruxolitinib and itaticinib.

64. The method of claim 62, wherein the PI3K inhibitor is parsaclisib.

65. The method of claim 62, wherein the standard of care therapy is selected from the group consisting of IFN-alpha, hydroxyurea, thalidomide, lenalidomide, an androgen, an erythropoietin-stimulating agent, a chemotherapeutic agent, or a combination thereof.

66. A kit comprising the antibody-drug conjugate of any one of claims 1-58, and instructions for use in treating a myeloproliferative neoplasm in a human subject in need thereof, optionally with instructions for use in combination with an additional therapy.133SUBSTITUTE SHEET (RULE 26)

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