HLA superagonists and uses thereof

Abstract

The present disclosure is directed to methods of modifying an HLA binding pocket of an HLA molecule in a subject. Some embodiments are directed to HLA molecules comprising a modified HLA binding pocket, said HLA molecules having improved affinity for peptides such as antigens. Other embodiments are directed to compositions comprising said HLA molecules and methods of using same.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of priority to U.S. Provisional Patent Application No. 63 / 262,941, filed October 22, 2021, which is incorporated by reference herein in its entirety.

[0002] References to sequence listings submitted electronically via EFS-WEB The contents of the electronically submitted sequence listing in an ASCII text file filed with this application (Name: 4285_022PC01_Seqlisting_ST26, Size: 2,4484 bytes, Creation Date: October 21, 2022) are incorporated herein by reference in their entirety.

[0003] Technical Field The present disclosure provides methods for enhancing the immune response to an antigen by modifying the binding pocket of a human leukocyte antigen (HLA) expressed on an antigen presenting cell. [Background technology]

[0004] Immunotherapy has emerged as an important tool in the fight against various diseases, including cancer. T cell therapy is at the forefront of immunotherapy development, and adoptive transfer of antitumor T cells has been shown to induce clinical responses in cancer patients. Many T cell therapies target mutated tumor antigens, but the majority of neoantigens are not shared and are unique to each patient.

[0005] Immunogenicity of T cell antigens is a key driver of effective cancer immunotherapy and is directly correlated with the ability of antigen epitopes to bind HLA and be stably presented to T cells. Many tumor-associated epitopes derived from self-antigens elicit relatively weak T cell responses, which poses a limitation for the development of cancer immunotherapy. Traditionally, this problem has been overcome by the generation of “heterocritic” peptides, a laborious, trial-and-error-based, low-throughput approach, in which individual epitopes are engineered to bind HLA molecules with enhanced interaction strength.

[0006] Thus, there remains a need in the field of immunotherapy for novel means to improve the immunogenicity of potential target antigens. Summary of the Invention

[0007] Some embodiments of the present disclosure are directed to methods of conditioning a subject in need of treatment comprising modifying the human leukocyte antigen (HLA) binding pocket of an HLA molecule expressed on the subject's cells.

[0008] Some embodiments of the present disclosure are directed to methods of enhancing an immune response in a subject in need thereof comprising modifying the HLA-binding pocket of an HLA molecule expressed on a cell of the subject.

[0009] Some embodiments of the present disclosure are directed to a method of increasing the binding affinity of an antigen to an HLA molecule on a cell, comprising modifying an HLA binding pocket of the HLA molecule. In some embodiments, the cell is present in a subject in need of treatment.

[0010] Some embodiments of the present disclosure are directed to methods of increasing the immunogenicity of an antigen in a subject in need of treatment, comprising modifying an HLA-binding pocket of an HLA molecule on a cell in the subject, wherein the HLA molecule is capable of binding the antigen.

[0011] Some embodiments of the present disclosure are directed to a method of enhancing an immune response to a therapy in a subject in need thereof, comprising modifying an HLA-binding pocket of an HLA molecule in a cell of the subject. In some embodiments, the therapy comprises immunotherapy.

[0012] Some embodiments of the present disclosure are directed to methods of enhancing an immune response to an antigen in a subject in need thereof, comprising modifying an HLA-binding pocket of an HLA molecule in a cell of the subject, wherein the HLA molecule is capable of binding the antigen.

[0013] In some embodiments, the modification increases the binding affinity of the HLA binding pocket for the antigen.

[0014] In some embodiments, the cell is an antigen presenting cell, hi some embodiments, the cell is a dendritic cell.

[0015] In some embodiments, the HLA molecule is an HLA class I allele. In some embodiments, the HLA molecule is an allele selected from the group consisting of HLA-A, HLA-B, HLA-C, HLA-E, HLA-F, HLA-G, HLA-K, HLA-L, and any combination thereof.

[0016] In some embodiments, the HLA molecules include (a) HLA-A*01, HLA-A*02, HLA-A*03, HLA-A*11, HLA-A*23, HLA-A*24, HLA-A*25, HLA-A*26, HLA-A*29, HLA-A*30, HLA-A*31, HLA-A*32, HLA-A*33, HLA-A*34, HLA-A*36, HLA-A*43, HL A group consisting of A-A*66, HLA-A*68, HLA-A*69, HLA-A*74, and HLA-A*80; (b) HLA-B*07, HLA-B*08, HLA-B*13, HLA -B*14, HLA-B*15, HLA-B*18, HLA-B*27, HLA-B*35, HLA-B*37, HLA-B*38, HLA-B*39, HLA-B*40, HLA-B *41, HLA-B*42, HLA-B*44, HLA-B*45, HLA-B*46, HLA-B*47, HLA-B*48, HLA-B*49, HLA-B*50, HLA-B*5 1, HLA-B*52, HLA-B*53, HLA-B*54, HLA-B*55, HLA-B*56, HLA-B*57, HLA-B*58, HLA-B*59, HLA-B*67, (c) the group consisting of HLA-C*05:01, HLA-C*05:03, HLA-C*05:04, HLA-C*05:05, and HLA-C*05:06; and (d) any combination thereof.

[0017] The number of pages is HLA-A*23, HLA-A* 24. The HLA-A*32 is a high-speed snowflake Meanwhile, the ranges are HLA-A*23:01:01:01, HLA-A*23:01:01:02, HLA-A*2 3:01:01:03、SONG-A*23:01:01:04、SONG-A*23:01:01:05、SONG-A*23:01:01: 06、SONGS-A*23:01:01:07、SONGS-A*23:01:01:08、SONGS-A*23:01:01:09、SONGS-A* 23:01:01:10、SONG-A*23:01:01:11、SONG-A*23:01:01:12、SONG-A*23:01:01 :13、SONG-A*23:01:01:14、SONG-A*23:01:01:15、SONG-A*23:01:01:16、SONG-A *23:01:01:17、SONG-A*23:01:01:18、SONG-A*23:01:01:19、SONG-A*23:01:0 1:20, SONG-A*23:01:01:21, SONG-A*23:01:01:22, SONG-A*23:01:01:23, SONG- A*23:01:01:24、SONG-A*23:01:01:25、SONG-A*23:01:01:26、SONG-A*23:01: 02、SONG-A*23:01:03、SONG-A*23:01:04、SONG-A*23:01:05、SONG-A*23:01:06、 SONG-A*23:01:07、SONG-A*23:01:08、SONG-A*23:01:09、SONG-A*23:01:10、SONG A-A*23:01:11、SONG-A*23:01:12、SONG-A*23:01:13、SONG-A*23:01:14、SONG-A *23:01:15、SONGS-A*23:01:16、SONGS-A*23:01:17、SONGS-A*23:01:18、SONGS-A*2 3:01:19、SONGS-A*23:01:20、SONGS-A*23:01:21、SONGS-A*23:01:22、SONGS-A*23:0 1:23、SONG-A*23:01:24、SONG-A*23:01:25、SONG-A*23:01:26、SONG-A*23:01: 27、SONG-A*23:01:28、SONG-A*23:01:29、SONG-A*23:01:30、SONG-A*23:01:31、HLA-A*23:01:32、HLA-A*23:01:33、HLA-A*23:01:34、HLA-A*23:02、HLA-A*23:03:01、HLA-A*23:03:02:01、HLA-A*23:03:02:02、HLA-A*23:04、HLA-A*23:05、HLA-A*23:06、HLA-A*23:07、HLA-A*23:08、HLA-A*23:09、HLA-A*23:10、HLA-A*23:11、HLA-A*23:12、HLA-A*23:13、HLA-A*23:14:01、HLA-A* 23:14:02、HLA-A*23:15、HLA-A*23:16、HLA-A*23:17:01:01、HLA-A*23:17:01:02、HLA-A*23:17:01:03、HLA-A*23:17:02、HLA-A*23:17:03、HLA-A*23:18、HLA-A*23:19、HLA-A*23:20、HLA-A*23:21、HLA-A*23:22、HLA-A*23:23、HLA-A*23:24、HLA-A*23:25、HLA-A*23:26、HLA-A*23:27、HLA-A*23:28、 HLA-A*23:29、HLA-A*23:30、HLA-A*23:31、HLA-A*23:32、HLA-A*23:33、HLA-A*23:34、HLA-A*23:35、HLA-A*23:36、HLA-A*23:37:01、HLA-A*23:37:02、HLA-A*23:38、HLA-A*23:39、HLA-A*23:40、HLA-A*23:41、HLA-A*23:42、HLA-A*23:43、HLA-A*23:44、HLA-A*23:45、HLA-A*23:46、HLA-A*23:47、HLA -A*23:48、HLA-A*23:49、HLA-A*23:50、HLA-A*23:51、HLA-A*23:52、HLA-A*23:53、HLA-A*23:54、HLA-A*23:55、HLA-A*23:56、HLA-A*23:57、HLA-A*23:58、HLA-A*23:59、HLA-A*23:60、HLA-A*23:61、HLA-A*23:62、HLA-A*23:63、HLA-A*23:64、HLA-A*23:65、HLA-A*23:66、HLA-A*23:67、HLA-A*23:68、SONG-A*23:70, SONG-A*23:71, SONG-A*23:72, SONG-A*23:73, SONG-A*23:74, SONG-A*23:75, SONG-A *23:76、SONGS-A*23:77、SONGS-A*23:78、SONGS-A*23:79、SONGS-A*23:80、SONGS-A*23:81、SONGS-A*23:8 2, SONG-A*23:83, SONG-A*23:84, SONG-A*23:85, SONG-A*23:86, SONG-A*23:87, SONG-A*23:88, SONG -A*23:89、SONGS-A*23:90、SONGS-A*23:91、SONGS-A*23:92、SONGS-A*23:93、SONGS-A*23:94、SONGS-A*23 :95、SONGS-A*23:96、SONGS-A*23:97、SONGS-A*23:98、SONGS-A*23:99、SONGS-A*23:100、SONGS-A*23:1 、SONGS-A*23:102、SONGS-A*23:103、SONGS-A*23:104、SONGS-A*23:105、SONGS-A*23:106、SONGS-A*23:1 7, SONG-A*23:108, SONG-A*23:109, SONG-A*23:110, SONG-A*23:111, SONG-A*23:112, SONG-A*23:1 13. SONG-A*23:114; SONG-A*23:115; SONG-A*23:1 16. HLA-A*23:117 contains a wide range of materials.

[0018] The timings are HLA-A*24:02:01:01, HLA-A*24:02:01:02, HLA-A*24: 02:01:03、SONG-A*24:02:01:04、SONG-A*24:02:01:05、SONG-A*24:02:01:0 、SONG-A*24:02:01:07、SONG-A*24:02:01:08、SONG-A*24:02:01:09、SONG-A*2 4:02:01:10、SONG-A*24:02:01:11、SONG-A*24:02:01:12、SONG-A*24:02:01: 13、SONGS-A*24:02:01:14、SONGS-A*24:02:01:15、SONGS-A*24:02:01:16、SONGS- A*24:02:01:17、SONG-A*24:02:01:18、SONG-A*24:02:01:19、SONG-A*24:02: 01:20, SONG-A*24:02:01:21, SONG-A*24:02:01:22, SONG-A*24:02:01:23, HL A-A*24:02:01:24、SONG-A*24:02:01:25、SONG-A*24:02:01:26、SONG-A*24:0 2:01:27、SONG-A*24:02:01:28、SONG-A*24:02:01:29、SONG-A*24:02:01:3 、SONG-A*24:02:01:31、SONG-A*24:02:01:32、SONG-A*24:02:01:33、SONG-A*2 4:02:01:34、SONG-A*24:02:01:35、SONG-A*24:02:01:36、SONG-A*24:02:01: 37、SONGS-A*24:02:01:38、SONGS-A*24:02:01:39、SONGS-A*24:02:01:40、SONGS-A *24:02:01:41、SONG-A*24:02:01:42、SONG-A*24:02:01:43、SONG-A*24:02: 01:44, SONG-A*24:02:01:45, SONG-A*24:02:01:46, SONG-A*24:02:01:47, HL A-A*24:02:01:48、SONG-A*24:02:01:49、SONG-A*24:02:01:50、SONG-A*24:0 2:01:51, SONG-A*24:02:01:52, SONG-A*24:02:01:53, SONG-A*24:02:01:54,HLA-A*24:02:01:55、HLA-A*24:02:01:56、HLA-A*24:02:01:57、HLA-A*24:02:01:58、HLA-A*24:02:01:59、HLA-A*24:02:01:60、HLA-A*24:02:01:61、HLA-A*24:02:01:62、HLA-A*24:02:01:63、HLA-A*24:02:01:64、HLA-A*24:02:01:65、HLA-A*24:02:01:66、HLA-A*24:02:01:67、HLA-A*24:02:01:6 8、HLA-A*24:02:01:69、HLA-A*24:02:01:70、HLA-A*24:02:01:71、HLA-A*24:02:01:72、HLA-A*24:02:01:73、HLA-A*24:02:01:74、HLA-A*24:02:01:75、HLA-A*24:02:01:76、HLA-A*24:02:01:77、HLA-A*24:02:01:78、HLA-A*24:02:01:79、HLA-A*24:02:01:80、HLA-A*24:02:01:81、HLA-A*24:02:01 :82、HLA-A*24:02:01:83、HLA-A*24:02:01:84、HLA-A*24:02:01:85、HLA-A*24:02:01:86、HLA-A*24:02:01:87、HLA-A*24:02:01:88、HLA-A*24:02:01:89、HLA-A*24:02:01:90、HLA-A*24:02:01:91、HLA-A*24:02:01:92、HLA-A*24:02:01:93、HLA-A*24:02:01:94、HLA-A*24:02:01:95、HLA-A*24:02: 01:96、HLA-A*24:02:01:97、HLA-A*24:02:01:98、HLA-A*24:02:01:99、HLA-A*24:02:01:100、HLA-A*24:02:01:101、HLA-A*24:02:01:102、HLA-A*24:02:01:103、HLA-A*24:02:01:104、HLA-A*24:02:01:105、HLA-A*24:02:01:106、HLA-A*24:02:01:107、HLA-A*24:02:01:108、HLA-A*24:02:01:109、HLA-A*24:02:01:110、HLA-A*24:02:02、HLA-A*24:02:03、HLA-A*24:02:04、HLA-A*24:02:05、HLA-A*24:02:06、HLA-A*24:02:07、HLA-A*24:02:08、HLA-A*24:02:09、HLA-A*24:02:10、HLA-A*24:02:11、HLA-A*24:02:12、HLA-A*24:02:13、HLA-A*24:02:14、HLA-A*24:02:15、HLA-A*24:02:16、HLA-A* 24:02:17、HLA-A*24:02:18、HLA-A*24:02:19、HLA-A*24:02:20、HLA-A*24:02:21、HLA-A*24:02:22、HLA-A*24:02:23、HLA-A*24:02:24、HLA-A*24:02:25、HLA-A*24:02:26、HLA-A*24:02:27、HLA-A*24:02:28、HLA-A*24:02:29、HLA-A*24:02:30、HLA-A*24:02:31、HLA-A*24:02:32、HLA-A*24:02:33、H LA-A*24:02:34、HLA-A*24:02:35、HLA-A*24:02:36、HLA-A*24:02:37、HLA-A*24:02:38、HLA-A*24:02:39、HLA-A*24:02:40:01、HLA-A*24:02:40:02、 HLA-A*24:02:41、HLA-A*24:02:42、HLA-A*24:02:43、HLA-A*24:02:44、HLA-A*24:02:45、HLA-A*24:02:46、HLA-A*24:02:47、HLA-A*24:02:48、HLA-A *24:02:49、HLA-A*24:02:50、HLA-A*24:02:51、HLA-A*24:02:52、HLA-A*24:02:53、HLA-A*24:02:54、HLA-A*24:02:55、HLA-A*24:02:56、HLA-A*24:02:57、HLA-A*24:02:58、HLA-A*24:02:59、HLA-A*24:02:60、HLA-A*24:02:61、HLA-A*24:02:62、HLA-A*24:02:63、HLA-A*24:02:64、HLA-A*24:02:65、HLA-A*24:02:66、HLA-A*24:02:67、HLA-A*24:02:68、HLA-A*24:02:69、HLA-A*24:02:70、HLA-A*24:02:71、HLA-A*24:02:72、HLA-A*24:02:73、HLA-A*24:02:74、HLA-A*24:02:75、HLA-A*24:02:76、HLA-A*24:02:77、HLA-A*24:02:78、HLA-A*24:02:79、HLA-A*24:02:80、HLA-A*24:02:81、HLA-A* 24:02:82、HLA-A*24:02:83、HLA-A*24:02:84、HLA-A*24:02:85、HLA-A*24:02:86、HLA-A*24:02:87、HLA-A*24:02:88、HLA-A*24:02:89、HLA-A*24:02:90、HLA-A*24:02:91、HLA-A*24:02:92、HLA-A*24:02:93、HLA-A*24:02:94、HLA-A*24:02:95、HLA-A*24:02:96、HLA-A*24:02:97、HLA-A*24:02:9 8、HLA-A*24:02:99、HLA-A*24:02:100、HLA-A*24:02:101、HLA-A*24:02:102:01、HLA-A*24:02:102:02、HLA-A*24:02:103、HLA-A*24:02:104、HLA-A*24:02:105、HLA-A*24:02:106、HLA-A*24:02:107、HLA-A*24:02:108、HLA-A*24:02:109、HLA-A*24:02:110、HLA-A*24:02:111、HLA-A*24:02:112 ,HLA-A*24:02:113,HLA-A*24:02:114,HLA-A*24:02:115:01,HLA-A*24:02:115:02,HLA-A*24:02:116,HLA-A*24:02:117,HLA-A*24:02:118,HLA-A*24:02:119,HLA-A*24:02:120,HLA-A*24:02:121,HLA-A*24:02:122,HLA-A*24:02:123,HLA-A*24:02:124,HLA-A*24:02:125,HLA-A*24:02:126,HLA-A*24:02:127、HLA-A*24:02:128、HLA-A*24:02:129、HLA-A*24:02:130、HLA-A*24:02:131、HLA-A*24:02:132、HLA-A*24:02:133、HLA-A*24:02:134、HLA-A*24:02:135、HLA-A*24:02:136、HLA-A*24:02:137、HLA-A*24:02:138、HLA-A*24:02:139、HLA-A*24:02:140、HLA-A*24:02:141、HLA-A*24:0 2:142、HLA-A*24:02:143、HLA-A*24:02:144、HLA-A*24:02:145、HLA-A*24:02:146、HLA-A*24:02:147、HLA-A*24:02:148、HLA-A*24:02:149、HLA-A*24:03:01:01、HLA-A*24:03:01:02、HLA-A*24:03:01:03、HLA-A*24:03:02、HLA-A*24:03:03、HLA-A*24:03:04、HLA-A*24:04、HLA-A*24:05:01、HLA-A* 24:05:02、HLA-A*24:06、HLA-A*24:07:01:01、HLA-A*24:07:01:02、HLA-A*24:07:01:03、HLA-A*24:07:02、HLA-A*24:07:03、HLA-A*24:07:04、HLA-A*24:08、HLA-A*24:09、HLA-A*24:10:01:01、HLA-A*24:10:01:02、HLA-A*24:10:02、HLA-A*24:11、HLA-A*24:13:01、HLA-A*24:13:02、HLA-A*24:14:0 1:01、HLA-A*24:14:01:02、HLA-A*24:14:01:03、HLA-A*24:14:01:04、HLA-A*24:15、HLA-A*24:17:01:01、HLA-A*24:17:01:02、HLA-A*24:18、HLA-A*24:19、HLA-A*24:20:01:01、HLA-A*24:20:01:02、HLA-A*24:20:02、HLA-A*24:21:01、HLA-A*24:21:02、HLA-A*24:22、HLA-A*24:23、HLA-A*24:24、HLA-A*24:25、HLA-A*24:26、HLA-A*24:27、HLA-A*24:28、HLA-A*24:29、HLA-A*24:30、HLA-A*24:31、HLA-A*24:32、HLA-A*24:33、HLA-A*24:34、HLA-A*24:35、H, LA-A*24:36, SONG-A*24:37, SONG-A*24:38, SONG-A*24:39, SONG-A*24:40, HL A-A*24:41, SONG-A*24:42, SONG-A*24:43, SONG-A*24:44, SONG-A*24:45, SONG- A*24:46, SONG-A*24:47, SONG-A*24:48, SONG-A*24:49, SONG-A*24:50, SONG-A* 24:51、SONGS-A*24:52、SONGS-A*24:53、SONGS-A*24:54、SONGS-A*24:55、SONGS-A*2 :56:01、SONG-A*24:56:02、SONG-A*24:57、SONG-A*24:58、SONG-A*24:59、SONG- A*24:60, SONG-A*24:61, SONG-A*24:62, SONG-A*24:63, SONG-A*24:64, SONG-A* 24:66、SONGS-A*24:67、SONGS-A*24:68、SONGS-A*24:69、SONGS-A*24:70、SONGS-A*2 :71、SONGS-A*24:72、SONGS-A*24:73、SONGS-A*24:74:01、SONGS-A*24:74:02、SONGS- A*24:75, SONG-A*24:76, SONG-A*24:77, SONG-A*24:78, SONG-A*24:79, SONG-A* 24:80、SONGS-A*24:81、SONGS-A*24:82、SONGS-A*24:83、SONGS-A*24:84、SONGS-A*2 :85、SONGS-A*24:86、SONGS-A*24:87、SONGS-A*24:88、SONGS-A*24:89、SONGS-A*24:9 0:01、SONG-A*24:90:02、SONG-A*24:91、SONG-A*24:92、SONG-A*24:93、SONG-A* 24:94、SONGS-A*24:95、SONGS-A*24:96、SONGS-A*24:97、SONGS-A*24:98、SONGS-A*2 :99、SONGS-A*24:100、SONGS-A*24:101、SONGS-A*24:102、SONGS-A*24:103、SONGS-A* 24:104, SONG-A*24:105, SONG-A*24:106, SONG-A*24:107, SONG-A*24:108, SONG -A*24:109, SONG-A*24:110, SONG-A*24:111, SONG-A*24:112, SONG-A*24:1HLA-A*24:114、HLA-A*24:115、HLA-A*24:116、HLA-A*24:117、HLA-A*24:118、HLA-A*24:119、HLA-A*24:120、HLA-A*24:121、HLA-A*24:122、HLA-A*24:123、HLA-A*24:124、HLA-A*24:125、HLA-A*24:126、HLA-A*24:127、HLA-A*24:128、HLA-A*24:129、HLA-A*24:130、HLA-A*24:131、HLA-A*24:132、H LA-A*24:133、HLA-A*24:134、HLA-A*24:135:01、HLA-A*24:135:02、HLA-A*24:136、HLA-A*24:137、HLA-A*24:138、HLA-A*24:139、HLA-A*24:140、HL A-A*24:141、HLA-A*24:142:01、HLA-A*24:142:02、HLA-A*24:143、HLA-A*24:144、HLA-A*24:145、HLA-A*24:146、HLA-A*24:147、HLA-A*24:148、HLA-A*24:148 -A*24:149, HLA-A*24:150, HLA-A*24:151, HLA-A*24:152, HLA-A*24:153, HLA-A*24:154, HLA-A*24:155, HLA-A*24:156, HLA-A*24:157, HLA-A*24:158, HLA-A*24:159, HLA-A*24:160, HLA-A*24:161, HLA-A*24:162, HLA-A*24:163, HLA-A*24:164, HLA-A*24:165, HLA-A*24:166, HLA-A*24:167, HLA- A*24:168、HLA-A*24:169、HLA-A*24:170、HLA-A*24:171、HLA-A*24:172:01、HLA-A*24:172:02、HLA-A*24:173、HLA-A*24:174、HLA-A*24:175、HLA-A*24:176、HLA-A*24:177、HLA-A*24:178、HLA-A*24:179、HLA-A*24:180、HLA-A*24:181、HLA-A*24:182、HLA-A*24:183、HLA-A*24:184、HLA-A-24:185、HLA-A*24:186、HLA-A*24:187、HLA-A*24:188、HLA-A*24:189、HLA-A*24:190、HLA-A*24:191、HLA-A*24:192、HLA-A*24:193、HLA-A*24:194、HLA-A*24:195、HLA-A*24:196、HLA-A*24:197、HLA-A*24:198、HLA-A*24:199、HLA-A*24:200、HLA-A*24:201、HLA-A*24:202、HLA-A*24:203、HLA-A*24:204、H LA-A*24:205、HLA-A*24:206、HLA-A*24:207:01、HLA-A*24:207:02、HLA-A*24:208:01、HLA-A*24:208:02:01、HLA-A*24:208:02:02、HLA-A*24:209、 HLA-A*24:210、HLA-A*24:212、HLA-A*24:213、HLA-A*24:214、HLA-A*24:215、HLA-A*24:216、HLA-A*24:217、HLA-A*24:218、HLA-A*24:219、HLA-A*24 :220、HLA-A*24:221、HLA-A*24:222、HLA-A*24:223、HLA-A*24:224、HLA-A*24:225:01、HLA-A*24:225:02、HLA-A*24:226:01、HLA-A*24:226:02、HLA-A*24:227、HLA-A*24:228、HLA-A*24:229、HLA-A*24:230、HLA-A*24:231、HLA-A*24:232、HLA-A*24:233、HLA-A*24:234、HLA-A*24:235、HLA-A*24:23 6、HLA-A*24:237、HLA-A*24:238、HLA-A*24:239、HLA-A*24:240、HLA-A*24:241、HLA-A*24:242、HLA-A*24:243、HLA-A*24:244、HLA-A*24:245、HLA-A*24:246、HLA-A*24:247、HLA-A*24:248、HLA-A*24:249、HLA-A*24:250、HLA-A*24:251、HLA-A*24:252、HLA-A*24:253、HLA-A*24:254、HLA-A*24:255、HLA-A*24:256、HLA-A*24:257、HLA-A*24:258、HLA-A*24:259、HLA-A*24:260、HLA-A*24:261、HLA-A*24:262、HLA-A*24:263、HLA-A*24:264、HLA-A*24:265、HLA-A*24:266、HLA-A*24:267、HLA-A*24:268、HLA-A*24:269、HLA-A*24:270、HLA-A*24:271、HLA-A*24:272、HLA-A*24:273、HLA-A*24:274、HLA -A*24:275, HLA-A*24:276, HLA-A*24:277, HLA-A*24:278, HLA-A*24:279, HLA-A*24:280, HLA-A*24:281, HLA-A*24:282, HLA-A*24:283, HLA-A*24:284, HLA-A*24:285, HLA-A*24:286, HLA-A*24:287, HLA-A*24:288, HLA-A*24:289, HLA-A*24:290, HLA-A*24:291, HLA-A*24:292, HLA-A*24:293, HLA-A* 24:294、HLA-A*24:295、HLA-A*24:296、HLA-A*24:297、HLA-A*24:298、HLA-A*24:299、HLA-A*24:300、HLA-A*24:301、HLA-A*24:302、HLA-A*24:303、HLA-A*24:304、HLA-A*24:305、HLA-A*24:306、HLA-A*24:307、HLA-A*24:308、HLA-A*24:309、HLA-A*24:310:01、HLA-A*24:310:02、HLA-A*24:311、HLA -A*24:312、HLA-A*24:313:01、HLA-A*24:313:02、HLA-A*24:314、HLA-A*24:315、HLA-A*24:316、HLA-A*24:317、HLA-A*24:318、HLA-A*24:319、HLA-A*24:320、HLA-A*24:321、HLA-A*24:322、HLA-A*24:323、HLA-A*24:324、HLA-A*24:325、HLA-A*24:326、HLA-A*24:327、HLA-A*24:328、HLA-A*24:329、HLA-A*24:330, HLA-A*24:331, HLA-A*24:332, HLA-A*24:333, HLA-A*24:334, HLA-A*24:335, HLA-A*24:336, HLA-A*24:337, HLA-A*24:338, HLA-A*24:339, HLA-A*24:340, HLA-A*24:341, HLA-A*24:342, HLA-A*24:343, HLA-A*24:344, HLA-A*24:345, HLA-A*24:346, HLA-A*24:347:01, HLA-A*24:34 7:02, HLA-A*24:348, HLA-A*24:349, HLA-A*24:350, HLA-A*24:351, HLA-A*24:352, HLA-A*24:353, HLA-A*24:354, HLA-A*24:355, HLA-A*24:356, HLA-A*24:357, HLA-A*24:358, HLA-A*24:359, HLA-A*24:360, HLA-A*24:361, HLA-A*24:362, HLA-A*24:363, HLA-A*24:364, HLA-A*24:365, HLA-A*24:3 66、HLA-A*24:367、HLA-A*24:368、HLA-A*24:369、HLA-A*24:370、HLA-A*24:371、HLA-A*24:372、HLA-A*24:373、HLA-A*24:374、HLA-A*24:375、HLA-A*24:376、HLA-A*24:377、HLA-A*24:378、HLA-A*24:379、HLA-A*24:380、HLA-A*24:381、HLA-A*24:382、HLA-A*24:383、HLA-A*24:384、HLA-A-24:38 5、HLA-A*24:386、HLA-A*24:387、HLA-A*24:388、HLA-A*24:389、HLA-A*24:390、HLA-A*24:391、HLA-A*24:392、HLA-A*24:393、HLA-A*24:394、HLA-A*24:395、HLA-A*24:396、HLA-A*24:397、HLA-A*24:398、HLA-A*24:399、HLA-A*24:400、HLA-A*24:401、HLA-A*24:402、HLA-A*24:403、HLA-A*24:404、HLA-A*24:405、HLA-A*24:406、HLA-A*24:407、HLA-A*24:408、HLA-A*24:409、HLA-A*24:410、HLA-A*24:411、HLA-A*24:412、HLA-A*24:413、HLA-A*24:414、HLA-A*24:415、HLA-A, *24:416、HLA-A*24:417、HLA-A*24:418、HLA-A*24:419、HLA-A*24:420、HLA-A*24:421、HLA-A*24:422、HLA-A*24:423、HLA-A*24:424、HLA-A*24:425、HLA-A*24:426、HLA-A*24:427、HLA-A*24:428、HLA-A*24:429、HLA-A*24:430、HLA-A*24:431:01、HLA-A*24:431:02、HLA-A*24:432、HLA-A*24:433、HL A-A*24:434、HLA-A*24:435、HLA-A*24:436、HLA-A*24:437、HLA-A*24:438、HLA-A*24:439、HLA-A*24:440、HLA-A*24:441、HLA-A*24:442、HLA-A*24:443、HLA-A*24:444、HLA-A*24:445、HLA-A*24:446、HLA-A*24:447、HLA-A*24:448、HLA-A*24:449、HLA-A*24:450、HLA-A*24:451、HLA-A*24:452、HLA-A *24:453、HLA-A*24:454、HLA-A*24:455、HLA-A*24:456、HLA-A*24:457、HLA-A*24:458、HLA-A*24:459、HLA-A*24:460:01:01、HLA-A*24:460:01:02、HLA-A*24:461、HLA-A*24:462、HLA-A*24:463、HLA-A*24:464、HLA-A*24:465、HLA-A*24:466、HLA-A*24:467、HLA-A*24:468、HLA-A*24:469、HLA-A*24: 470、HLA-A*24:472、HLA-A*24:473、HLA-A*24:474、HLA-A*24:475、HLA-A*24:476、HLA-A*24:477、HLA-A*24:478、HLA-A*24:479、HLA-A*24:480、HLA-A*24:481、HLA-A*24:482、HLA-A*24:483、HLA-A*24:484、HLA-A*24:485、HLA-A*24:486、HLA-A*24:487、HLA-A*24:488、HLA-A*24:489、HLA-A*24:490、SONG-A*24:491, SONG-A*24:492, SONG-A*24:493, SONG-A*24:494, SONG-A*24:495, SONG-A*24:496, SONG-A*24:497, SONG-A*24:49 8, SONG-A*24:499, SONG-A*24:500, SONG-A*24:501, SONG-A*24:502, SONG-A*24:503, SONG-A*24:504, SONG-A*24:505, SONG-A*24: 506、SONGS-A*24:507、SONGS-A*24:508、SONGS-A*24:509、SONGS-A*24:510、SONGS-A*24:511、SONGS-A*24:512、SONGS-A*24:513、SONGS-A*2 4:514, SONG-A*24:515, SONG-A*24:516, SONG-A*24:517, SONG-A*24:518, SONG-A*24:519, SONG-A*24:520, SONG-A*24:521, SONG-A *24:522、SONGS-A*24:523、SONGS-A*24:524、SONGS-A*24:525、SONGS-A*24:526、SONGS-A*24:527、SONGS-A*24:528、SONGS-A*24:529、SONGS -A*24:530, SONG-A*24:531, SONG-A*24:532, SONG-A*24:533, SONG-A*24:534, SONG-A*24:535, SONG-A*24:536, SONG-A*24:537, H LA-A*24:538、SONGS-A*24:539、SONGS-A*24:540、SONGS-A*24:541、SONGS-A*24:542、SONGS-A*24:543、SONGS-A*24:544、SONGS-A*24:545 、SONGS-A*24:546、SONGS-A*24:547、SONGS-A*24:548、SONGS-A*24:5 9. HLA-A*24:550 and HLA-A*24:5

[0019] The details are HLA-A*32:01:01:01, HLA-A*32:01:01:02, HLA-A*32: 01:01:03、SONG-A*32:01:01:04、SONG-A*32:01:01:05、SONG-A*32:01:01:0 、SONG-A*32:01:01:07、SONG-A*32:01:01:08、SONG-A*32:01:01:09、SONG-A*3 2:01:01:10、SONG-A*32:01:01:11、SONG-A*32:01:01:12、SONG-A*32:01:01: 13、SONG-A*32:01:01:14、SONG-A*32:01:01:15、SONG-A*32:01:01:16、SONG-A *32:01:01:17、SONG-A*32:01:01:18、SONG-A*32:01:01:19、SONG-A*32:01:0 1:20、SONGS-A*32:01:01:21、SONGS-A*32:01:01:22、SONGS-A*32:01:01:23、SONGS -A*32:01:01:24、SONG-A*32:01:01:25、SONG-A*32:01:01:26、SONG-A*32:0 :01:27、SONG-A*32:01:01:28、SONG-A*32:01:01:29、SONG-A*32:01:01:30、H LA-A*32:01:02、SONGS-A*32:01:03、SONGS-A*32:01:04、SONGS-A*32:01:05、SONGS -A*32:01:06、SONG-A*32:01:07、SONG-A*32:01:08、SONG-A*32:01:09、SONG-A *32:01:10、SONGS-A*32:01:11、SONGS-A*32:01:12、SONGS-A*32:01:13、SONGS-A*3 2:01:14、SONGS-A*32:01:15、SONGS-A*32:01:16、SONGS-A*32:01:17、SONGS-A*32: 01:18、SONGS-A*32:01:19、SONGS-A*32:01:20、SONGS-A*32:01:21、SONGS-A*32:01 :22、SONG-A*32:01:23、SONG-A*32:01:24、SONG-A*32:01:25、SONG-A*32:01:2 6、SONG-A*32:01:27、SONG-A*32:01:28、SONG-A*32:01:29、SONG-A*32:01:30、HLA-A*32:01:31、HLA-A*32:01:32、HLA-A*32:01:33、HLA-A*32:01:34、HLA-A*32:01:35、HLA-A*32:01:36、HLA-A*32:01:37、HLA-A*32:01:38、HLA-A*32:01:39、HLA-A*32:01:40、HLA-A*32:01:41、HLA-A*32:01:42、HLA-A*32:01:43、HLA-A*32:01:44、HLA-A*32:01:45、HLA-A*32:01:46、HLA-A*32: 01:47, HLA-A*32:02, HLA-A*32:03:01:01, HLA-A*32:03:01:02, HLA-A*32:04, HLA-A*32:05, HLA-A*32:06, HLA-A*32:07, HLA-A*32:08, HLA-A*32:09, HLA-A*32:10, HLA-A*32:11, HLA-A*32:12, HLA-A*32:13, HLA-A*32:14, HLA-A*32:15, HLA-A*32:16, HLA-A*32:17, HLA-A*32:18, HLA-A*32:19, HLA -A*32:20、HLA-A*32:21、HLA-A*32:22、HLA-A*32:23、HLA-A*32:24、HLA-A*32:25、HLA-A*32:26:01、HLA-A*32:26:02、HLA-A*32:27、HLA-A*32:28、HLA-A*32:29、HLA-A*32:30:01、HLA-A*32:30:02、HLA-A*32:31、HLA-A*32:32、HLA-A*32:33:01、HLA-A*32:33:02、HLA-A*32:33:03、HLA-A*32:34、HLA -A*32:35, HLA-A*32:36, HLA-A*32:37, HLA-A*32:38, HLA-A*32:39, HLA-A*32:40, HLA-A*32:41, HLA-A*32:42, HLA-A*32:43:01, HLA-A*32:43:02, HLA-A*32:44, HLA-A*32:45, HLA-A*32:46:01, HLA-A*32:46:02, HLA-A*32:47, HLA-A*32:48, HLA-A*32:49, HLA-A*32:50, HLA-A*32:51, HLA-A*32:52,HLA-A*32:53、HLA-A*32:54、HLA-A*32:55:01、HLA-A*32:55:02、HLA-A*32:55:03、HLA-A*32:56、HLA-A*32:57、HLA-A*32:58、HLA-A*32:59、HLA-A*32:60、HLA-A*32:61、HLA-A*32:62、HLA-A*32:63、HLA-A*32:64、HLA-A*32:65、HLA-A*32:66、HLA-A*32:67、HLA-A*32:68、HLA-A*32:69、HLA-A*32:70、 HLA-A*32:71、HLA-A*32:72、HLA-A*32:73、HLA-A*32:74、HLA-A*32:75、HLA-A*32:76、HLA-A*32:77、HLA-A*32:78、HLA-A*32:79、HLA-A*32:80、HLA-A*32:81、HLA-A*32:82、HLA-A*32:83、HLA-A*32:84、HLA-A*32:85、HLA-A*32:86、HLA-A*32:87、HLA-A*32:88、HLA-A*32:89、HLA-A*32:90、HLA-A*32:9 1、HLA-A*32:92、HLA-A*32:93、HLA-A*32:94、HLA-A*32:95、HLA-A*32:96、HLA-A*32:97、HLA-A*32:98、HLA-A*32:99、HLA-A*32:100、HLA-A*32:101、HLA-A*32:102、HLA-A*32:103、HLA-A*32:104、HLA-A*32:105、HLA-A*32:106:01:01、HLA-A*32:106:01:02、HLA-A*32:107、HLA-A*32:108、HLA-A*32:1 09、HLA-A*32:110、HLA-A*32:111、HLA-A*32:112、HLA-A*32:113、HLA-A*32:114、HLA-A*32:115、HLA-A*32:116、HLA-A*32:117、HLA-A*32:118、HLA-A*32:119、HLA-A*32:120、HLA-A*32:121、HLA-A*32:122、HLA-A*32:123、HLA-A*32:124、HLA-A*32:125、HLA-A*32:126、HLA-A*32:127、HLA-A*32:128、HLA-A*32:129, HLA-A*32:130, HLA-A*32:131, HLA-A*32:132, HLA-A*32:133, HLA-A*32:134, HLA-A*32:135, HLA-A *32:136, HLA-A*32:137, HLA-A*32:138, HLA-A*32:139, HLA-A*32:140, HLA-A*32:141, HLA-A*32:142, HLA-A*32:14 3, HLA-A*32:144, HLA-A*32:145, HLA-A*32:146, HLA-A*32:147, HLA-A*32:148, HLA-A*32:149, HLA-A*32:150, HLA-A*32:151, HLA-A*32:152, HLA-A*32:153, HLA-A*32:154, HLA-A*32:155, and HLA-A*32:156. In some embodiments, the HLA binding pocket is an A pocket, a B pocket, a C pocket, a D pocket, an E pocket, an F pocket, or any combination thereof. In some embodiments, the binding pocket is an F pocket. In some embodiments, more than one binding pocket is modified.

[0020] In some embodiments, the modification increases the presentation of the antigen on the cell surface. In some embodiments, the modification increases antigen-specific T cell responses. In some embodiments, the antigen is a tumor antigen. In some embodiments, the modification increases tumor antigen-specific T cell proliferation.

[0021] 26. The method of any one of claims 1 to 25, wherein the modifying comprises mutating an amino acid in the HLA binding pocket of the HLA molecule. In some embodiments, the mutating comprises substitution of an amino acid. In some embodiments, the modifying comprises substitution of an amino acid, wherein the substituted amino acid is alanine. In some embodiments, the new amino acid replacing the original amino acid is selected from the group consisting of leucine, valine, isoleucine, methionine, phenylalanine, tyrosine, and tryptophan. In some embodiments, the new amino acid replacing the original amino acid is leucine. In some embodiments, the modification comprises mutating an amino acid residue selected from amino acid residue 79, amino acid residue 80, amino acid residue 81, amino acid residue 82, or amino acid residue 83, wherein the position corresponds to the amino acid sequence shown in SEQ ID NO:1. In some embodiments, the modification comprises mutating amino acid residue 81, which corresponds to the amino acid sequence shown in SEQ ID NO:1. In some embodiments, the modification comprises substitution of the amino acid residue at position 81, which corresponds to the amino acid sequence shown in SEQ ID NO:1. In some embodiments, the modification comprises an A81L substitution, which corresponds to the amino acid sequence shown in SEQ ID NO:1.

[0022] In some embodiments, modifying is performed by a gene editing tool. In some embodiments, the gene editing tool comprises CRISPR technology, zinc finger nuclease, TALEN, shRNA, siRNA, miRNA, antisense oligonucleotide, meganuclease, restriction endonuclease, or any combination thereof.

[0023] In some embodiments, the method further comprises administering immunotherapy to the subject.

[0024] In some embodiments, the immunotherapy comprises immune cell therapy. In some embodiments, the immune cell therapy comprises administering T cells, NK cells, tumor infiltrating lymphocytes, or any combination thereof. In some embodiments, the immune cell therapy comprises administering engineered T cells, wherein the engineered T cells comprise a nucleic acid molecule encoding a chimeric antigen receptor (CAR), a heterologous T cell receptor (TCR), an engineered TCR, or any combination thereof.

[0025] In some embodiments, the immunotherapy comprises administering a checkpoint inhibitor. In some embodiments, the immunotherapy comprises administering a PD-1 inhibitor, a PD-L1 inhibitor, a CTLA-4 inhibitor, a LAG3 inhibitor, a TIM3 inhibitor, or any combination thereof. In some embodiments, the immunotherapy comprises administering an antibody or antigen-binding portion thereof that specifically binds to PD-1 and inhibits the interaction of PD-1 and PD-L1. In some embodiments, the antibody or antigen-binding portion thereof that specifically binds to PD-1 and inhibits the interaction of PD-1 and PD-L1 is selected from nivolumab and pembrolizumab. In some embodiments, the immunotherapy comprises administering an antibody or antigen-binding portion thereof that specifically binds to and inhibits CTLA-4. In some embodiments, the antibody or antigen-binding portion thereof that specifically binds to and inhibits CTLA-4 is selected from ipilimumab and tremelimumab.

[0026] In some embodiments, the immunotherapy comprises administering an antibody or antigen-binding portion thereof that specifically binds and inhibits LAG3, hi some embodiments, the immunotherapy comprises a cancer vaccine.

[0027] In some embodiments, the subject is suffering from cancer. In some embodiments, the cancer is melanoma, bone cancer, kidney cancer, prostate cancer, breast cancer, colon cancer, lung cancer, cutaneous or intraocular malignant melanoma, pancreatic cancer, skin cancer, head and neck cancer, uterine cancer, ovarian cancer, rectal cancer, anal cancer, gastric cancer, testicular cancer, uterine cancer, fallopian tube cancer, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, Hodgkin's disease, non-Hodgkin's lymphoma (NHL), primary mediastinal large B-cell lymphoma (PMBC), diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), transformed follicular lymphoma, splenic marginal zone lymphoma (SMZL), esophageal cancer, small intestine cancer, endocrine system cancer, thyroid cancer, parathyroid cancer, adrenal cancer, sarcoma of soft tissue, The cancer is selected from the group consisting of urethral cancer, penile cancer, chronic or acute leukemia, acute myeloid leukemia (AML), chronic myeloid leukemia, acute lymphoblastic leukemia (ALL) (including non-T cell ALL), chronic lymphocytic leukemia (CLL), solid tumors of childhood, lymphocytic lymphoma, bladder cancer, kidney or ureter cancer, renal pelvis cancer, neoplasms of the central nervous system (CNS), primary CNS lymphoma, tumor angiogenesis, spinal axis tumor, brain stem glioma, pituitary adenoma, Kaposi's sarcoma, epidermoid carcinoma, squamous cell carcinoma, T cell lymphoma, environmentally induced cancers including those due to asbestos, other B cell malignancies, and combinations of the above cancers.

[0028] Some embodiments of the present disclosure are directed to HLA molecules that comprise a modified HLA binding pocket.

[0029] In some embodiments, the HLA molecule is an HLA class I allele. In some embodiments, the HLA molecule is an allele selected from the group consisting of HLA-A, HLA-B, HLA-C, HLA-E, HLA-F, HLA-G, HLA-K, HLA-L, and any combination thereof. In some embodiments, the HLA molecule is an allele selected from the group consisting of: (a) HLA-A*01, HLA-A*02, HLA-A*03, HLA-A*11, HLA-A*23, HLA-A*24, HLA-A*25, HLA-A*26, HLA-A*29, HLA-A*30, HLA-A*31, HLA-A*32, HLA-A*33, HLA-A*34, HLA-A*36, HLA-A*43, HLA-A*44, HLA-A*45, HLA-A*46, HLA-A*47, HLA-A*48, HLA-A*49, HLA-A*50, HLA-A*51, HLA-A*52, HLA-A*53, HLA-A*54, HLA-A*55, HLA-A*56, HLA-A*57, HLA-A*58, HLA-A*59, HLA-A*60, HLA-A*61, HLA-A*62, HLA-A*63, HLA-A*64, HLA-A*65, HLA-A*66, HLA-A*67, HLA-A*68, HLA-A*69, HLA-A*70, HLA-A*71, HLA-A*72, HLA-A*73, HLA-A*74, HLA-A*75, HLA-A*76, HLA-A*77, HLA-A*78, H A group consisting of A-A*66, HLA-A*68, HLA-A*69, HLA-A*74, and HLA-A*80; (b) HLA-B*07, HLA-B*08, HLA-B*13, HLA -B*14, HLA-B*15, HLA-B*18, HLA-B*27, HLA-B*35, HLA-B*37, HLA-B*38, HLA-B*39, HLA-B*40, HLA-B *41, HLA-B*42, HLA-B*44, HLA-B*45, HLA-B*46, HLA-B*47, HLA-B*48, HLA-B*49, HLA-B*50, HLA-B*5 1, HLA-B*52, HLA-B*53, HLA-B*54, HLA-B*55, HLA-B*56, HLA-B*57, HLA-B*58, HLA-B*59, HLA-B*67, (c) the group consisting of HLA-B*73, HLA-B*78, HLA-B*79, HLA-B*81, HLA-B*82, and HLA-B*83; (c) the group consisting of HLA-C*05:01, HLA-C*05:03, HLA-C*05:04, HLA-C*05:05, and HLA-C*05:06; and (d) any combination thereof. In some embodiments, the HLA molecule is an allele selected from the group consisting of HLA-A*23, HLA-A*24, HLA-A*25, HLA-A*31, and HLA-A*32. In some embodiments, the HLA molecule is an HLA-A*24 allele.

[0030] In some embodiments, the HLA binding pocket is the A pocket, the B pocket, the C pocket, the D pocket, the E pocket, the F pocket, or any combination thereof. In some embodiments, the HLA binding pocket is the F pocket.

[0031] In some embodiments, the HLA molecule comprises more than one modified HLA binding pocket.

[0032] In some embodiments, the HLA molecule has a higher affinity for the antigen than an HLA molecule that comprises an unmodified HLA binding pocket, hi some embodiments, the antigen is a tumor antigen.

[0033] In some embodiments, the modified HLA binding pocket comprises an amino acid substitution compared to the unmodified HLA binding pocket. In some embodiments, the original amino acid replaced by the substitution is alanine. In some embodiments, the new amino acid replacing the original amino acid is selected from the group consisting of leucine, valine, isoleucine, methionine, phenylalanine, tyrosine, and tryptophan. In some embodiments, the new amino acid replacing the original amino acid is leucine.

[0034] In some embodiments, the HLA binding pocket comprises an amino acid substitution at a residue selected from amino acid residue 79, amino acid residue 80, amino acid residue 81, amino acid residue 82, or amino acid residue 83, wherein said positions correspond to the amino acid sequence set forth in SEQ ID NO: 1. In some embodiments, the HLA binding pocket comprises an amino acid substitution at amino acid residue 81, which corresponds to the amino acid sequence set forth in SEQ ID NO: 1. In some embodiments, the HLA binding pocket comprises an A81L substitution, which corresponds to the amino acid sequence set forth in SEQ ID NO: 1.

[0035] Some aspects of the present disclosure are directed to a nucleic acid or a set of nucleic acids that encode the HLA molecule disclosed herein.Some aspects of the present disclosure are directed to a vector or a set of vectors that comprise the nucleic acid disclosed herein.In some aspects, the vector is a viral vector.

[0036] Some embodiments of the present disclosure are directed to an HLA-antigen complex comprising an HLA and an antigen as disclosed herein. In some embodiments, the antigen is a tumor antigen.

[0037] Some aspects of the present disclosure are directed to antigen-presenting cells that comprise the HLA molecule disclosed herein, the nucleic acid or set of nucleic acids disclosed herein, the vector or set of vectors disclosed herein, or the HLA-antigen complex disclosed herein.In some aspects, the antigen-presenting cell is a dendritic cell.In some aspects, the antigen-presenting cell is an artificial antigen-presenting cell.

[0038] Some aspects of the present disclosure are directed to a pharmaceutical composition comprising an HLA molecule disclosed herein, a nucleic acid or set of nucleic acids disclosed herein, a vector or set of vectors disclosed herein, an HLA-antigen complex disclosed herein, or an antigen-presenting cell disclosed herein, and a pharma- ceutical acceptable excipient.

[0039] Some aspects of the present disclosure are directed to a vaccine comprising an HLA molecule disclosed herein, a nucleic acid or set of nucleic acids disclosed herein, a vector or set of vectors disclosed herein, an HLA-antigen complex disclosed herein, or an antigen-presenting cell disclosed herein.

[0040] Some aspects of the present disclosure are directed to a method of treating a subject in need of treatment comprising administering to the subject an HLA molecule disclosed herein, a nucleic acid or set of nucleic acids disclosed herein, a vector or set of vectors disclosed herein, an HLA-antigen complex disclosed herein, an antigen-presenting cell disclosed herein, a pharmaceutical composition disclosed herein, or a vaccine disclosed herein.

[0041] In some embodiments, the method further comprises administering immunotherapy to the subject.

[0042] In some embodiments, the immunotherapy comprises immune cell therapy. In some embodiments, the immune cell therapy comprises administering T cells, NK cells, tumor infiltrating lymphocytes, or any combination thereof. In some embodiments, the immune cell therapy comprises administering engineered T cells, wherein the engineered T cells comprise a nucleic acid molecule encoding a chimeric antigen receptor (CAR), a heterologous T cell receptor (TCR), an engineered TCR, or any combination thereof.

[0043] In some embodiments, the immunotherapy further comprises administering a checkpoint inhibitor. The method of any one of claims 80-84, wherein the immunotherapy comprises administering a PD-1 inhibitor, a PD-L1 inhibitor, a CTLA-4 inhibitor, a LAG3 inhibitor, a TIM3 inhibitor, or any combination thereof. In some embodiments, the immunotherapy comprises administering an antibody or antigen-binding portion thereof that specifically binds to PD-1 and inhibits the interaction of PD-1 and PD-L1, an antibody or antigen-binding portion thereof that specifically binds to and inhibits CTLA-4, an antibody or antigen-binding portion thereof that specifically binds to and inhibits LAG3, or any combination thereof. In some embodiments, (i) the antibody or antigen-binding portion thereof that specifically binds to PD-1 and inhibits the interaction of PD-1 and PD-L1 is selected from nivolumab and pembrolizumab, (ii) the antibody or antigen-binding portion thereof that specifically binds to and inhibits CTLA-4 is selected from ipilimumab and tremelimumab, or (iii) any combination of (i) and (ii).

[0044] In some embodiments, the subject is suffering from cancer. In some embodiments, the cancer is melanoma, bone cancer, kidney cancer, prostate cancer, breast cancer, colon cancer, lung cancer, cutaneous or intraocular malignant melanoma, pancreatic cancer, skin cancer, head and neck cancer, uterine cancer, ovarian cancer, rectal cancer, anal cancer, gastric cancer, testicular cancer, uterine cancer, fallopian tube cancer, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, Hodgkin's disease, non-Hodgkin's lymphoma (NHL), primary mediastinal large B-cell lymphoma (PMBC), diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), transformed follicular lymphoma, splenic marginal zone lymphoma (SMZL), esophageal cancer, small intestine cancer, endocrine system cancer, thyroid cancer, parathyroid cancer, adrenal cancer, sarcoma of soft tissue, The cancer is selected from the group consisting of urethral cancer, penile cancer, chronic or acute leukemia, acute myeloid leukemia (AML), chronic myeloid leukemia, acute lymphoblastic leukemia (ALL) (including non-T cell ALL), chronic lymphocytic leukemia (CLL), solid tumors of childhood, lymphocytic lymphoma, bladder cancer, kidney or ureter cancer, renal pelvis cancer, neoplasms of the central nervous system (CNS), primary CNS lymphoma, tumor angiogenesis, spinal axis tumor, brain stem glioma, pituitary adenoma, Kaposi's sarcoma, epidermoid carcinoma, squamous cell carcinoma, T cell lymphoma, environmentally induced cancers including those due to asbestos, other B cell malignancies, and combinations of the above cancers.

[0045] Some embodiments of the present disclosure are directed to methods of enriching a population of T cells obtained from a human subject comprising contacting the T cells with an HLA molecule disclosed herein, an HLA-antigen complex disclosed herein, or an antigen-presenting cell disclosed herein. [Brief description of the drawings]

[0046] [Figure 1] This shows a comparison of the amino acid sequences of the α1 domain of the most commonly expressed HLA class I alleles in the population. It shows that the amino acid sequence of HLA-A*24:02 differs from other HLA class I alleles at positions 81-83. The box represents the amino acid residues 81-83. [Diagram 2]Graphical representation of the surface formation of peptide-HLA complexes. (A) Surface expression of ΔNGFR, HLA class I and HLA-A*24:02 genes in T2 cells transduced with ΔNGFR-tagged full-length HLA-A*24:02 gene, analyzed by flow cytometry and subsequently stained with the indicated antibodies (open curves) or isotype control (filled curves). (B) Mean fluorescence intensity (MFI) of biotinylated peptides with streptavidin-PE on T2-HLA-A*24:02 (wild type or A81L) cells. The peptides used are shown in Table 1. B7-tethered HPVE75-13 and C7-tethered MART151-61 peptides were used as relevant controls. For data analysis, the MFI of samples for non-biotinylated peptides was subtracted from the MFI of samples for biotinylated peptides. P values ​​were determined by repeated measures one-way analysis of variance (ANOVA) with Tukey's multiple comparison test for each peptide (F = 1128 (adenovirus 5 hexon), 45.78 (CMV-IE1), 453.3 (CMVpp65), 484.2 (HIV env), 20.08 (EpCAM), and 430.2 (KM-HN-1), degrees of freedom = 11). [Table 1] [Figure 3A] Figure 8 shows that position 81 is important for surface expression of HLA-A*02:01. Graphical representation of surface expression of endogenous β2m and HLA class I molecules in T2 cells. β2m and HLA class I derived from transduced β2m-HLA-A*02:01 (wild type or L81A) in T2 / β2mKO cells were analyzed by flow cytometry after staining with anti-β2m and HLA class I mAbs (open curves) and isotype control (filled curves). Surface expression after fixation and permeabilization is shown. [Figure 3B]Graphical representation of intracellular expression of endogenous β2m and HLA class I molecules in T2 cells. β2m and HLA class I derived from transduced β2m-HLA-A*02:01 (wild type or L81A) in T2 / β2mKO cells were analyzed by flow cytometry after staining with anti-β2m and HLA class I mAbs (open curves) and isotype control (filled curves). Intracellular expression after fixation and permeabilization is shown. [Figure 3C] Jurkat76 / CD8 cells transduced with A2 / NY-ESO-1157-165TCR (clone 1G4LY) were used as responder cells in IFN-γ ELISPOT assays. [Figure 3D] Jurkat76 / CD8 cells transduced with A2 / gp100154-162 TCR were used as responder cells in IFN-γ ELISPOT assays. The indicated T2 cells pulsed with 10 μg / ml of heteroctic NY-ESO-1157-165, gp100154-162, or HIV pol476-484 control peptides were used as stimulator cells. Data shown represent the mean ± SD of experiments performed in triplicate. **P<0.01, ***P<0.001 (two-tailed Welch's t-test). [Figure 4]Graphical representation showing that HLA-A*24:02(A81L) enhances antigen presentation to T cells. (A) Jurkat76 / CD8 cells individually transduced with the indicated HLA-A*24:02-restricted WT1235-243 TCR were used as responder cells in IL-2 ELISPOT assays. T2 cells transduced with HLA-A*24:02 (wild type or A81L) and pulsed with graded concentrations of A24 / heterocritic WT1235-243 peptide were co-cultured with T cells in the presence of peptide (top) or in the presence of peptide-pulsed T2 cells (bottom). IL-2 secretion relative to maximal response was analyzed. (B) Primary T cells transduced with HLA-A*24 / gp100-intron 4 170-178 TCR were used as responder cells in IFN-γ ELISPOT assays. T2 cells or T2 cells transduced with HLA-A*24:02 (wild type or A81L) were pulsed with the indicated gp100-intron 4 peptides in serum-free medium and served as stimulator cells. HIV env584-592 peptide and non-transduced primary T cells were used as negative controls. *P<0.05, **P<0.01 (two-tailed Welch's t-test). [Diagram 5]1 is a graphical representation showing that peptide-loaded HLA-A*24:02 (A81L) multimers can be used to stain low affinity TCRs. (A) Peptide exchange efficiency in soluble monomeric HLA-A*24:02 wild type as measured by peptide competitive binding assay and ELISA. (B) Peptide exchange efficiency in soluble monomeric HLA-A*24:02 wild type with A81L substitution as measured by peptide competitive binding assay and ELISA. The HLA-A*24:02-restricted peptides employed are shown in Table 1. HLA-B*18-restricted MAGE-A3167-176 (No. 65), HLA-B*27-restricted VEGF (No. 66), and HLA-C*16-restricted MAGE-A4293-301 (No. 67) peptides were used as irrelevant controls. Data shown represent the mean ± SD of experiments performed in triplicate. (C) HLA-A*24:02 multimers with A81L substitution efficiently stain the cognate TCR with low affinity. Jurkat76 / CD8 cells transduced with three different A24 / WT1 TCRs or A24 / gp100-intron 4170-178 TCR as a control were stained with HLA-A*24:02Q115E-Kb / wild type WT1 (top) or HLA-A*24:02Q115E / A81L-Kb / wild type WT1 (bottom) multimers. The percentage of multimer+ cells among CD8+ T cells is shown. [Figure 6] 1 is a graphical representation showing that aAPC / HLA-A*24:02(A81L) enhances proliferation of tumor antigen-specific T cells. CD8+ T cells isolated from melanoma TILs were stimulated with aAPC / HLA-A*24:02 (wild type or A81L) pulsed with 10 μg / ml of gp100-intron 4170-178 or HTLV-1 tax301-309 peptides. (A) Shows staining with the indicated multimers before stimulation (day 0) and after 14 days of stimulation. Gates indicate percentage of multimer+ CD8+ T cells. (B) Shows cumulative frequency of gp100-intron 4170-178-multimer+ CD8+ T cells after 14 days of expansion culture. (C) Shows cumulative fold change of gp100-intron 4170-178-multimer+ CD8+ T cells after 14 days of expansion culture. Data shown represent mean ± SD. *p<0.05 (paired test). [Figure 7] Protein models showing the A81L substitution in the HLA-A*24:02 peptide complex. (A) Top view of the peptide binding cleft of HLA-A*24:02 complexed with Flu PB1498-505 peptide containing a side chain at position 81 (RCSB PDB: 4F7T). Modeled with an alanine (A81) at position 81. (B) Top view of the peptide binding cleft of HLA-A*24:02 complexed with Flu PB1498-505 peptide containing a side chain at position 81 (RCSB PDB: 4F7T). Modeled with a leucine (81L) at position 81. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0047] Some aspects of the present disclosure are directed to a method of conditioning a subject in need of treatment, comprising modifying a human leukocyte antigen (HLA) binding pocket expressed on the subject's cells. Some aspects of the present disclosure are directed to a method of enhancing an immune response in a subject in need thereof, comprising modifying a binding pocket of an HLA binding of an HLA molecule expressed on the subject's cells. Other aspects of the present disclosure are directed to a method of increasing the binding affinity of an antigen to an HLA molecule on a cell, comprising modifying a HLA binding pocket of an HLA molecule. In some aspects, the binding pocket of the HLA is modified to increase the binding affinity of the HLA to the antigen. Further aspects of the present disclosure are directed to engineered antigen-presenting cells comprising a binding pocket that is modified to increase the affinity of the HLA to the antigen.

[0048] Some embodiments of the present disclosure are further directed to a method of identifying a novel T cell receptor (TCR) capable of binding to a target antigen-HLA complex, comprising: (i) contacting a target antigen with an engineered antigen-presenting cell, where the engineered antigen-presenting cell comprises a binding pocket modified to increase the affinity of the HLA for the antigen; and (ii) contacting a plurality of TCRs with the target antigen-HLA complex.

[0049] Another aspect of the present disclosure is directed to a vaccine comprising an engineered antigen-presenting cell complexed with a target antigen, where the engineered antigen-presenting cell comprises a binding pocket that is modified to increase the affinity of HLA for the antigen.

[0050] I. Terminology In order that this disclosure may be more readily understood, certain terms are first defined. Unless otherwise expressly provided herein, as used in this application, each of the following terms shall have the meaning indicated below. Additional definitions are set forth throughout this application.

[0051] It should be noted that an entity designated with the term "a" or "an" refers to one or more of that entity, e.g., "a nucleotide sequence" is understood to represent one or more nucleotide sequences. Thus, the terms "a" (or "an"), "one or more," and "at least one" may be used interchangeably herein.

[0052] Furthermore, "and / or", when used herein, should be construed as a specific disclosure of each of the two specified features or components with or without the other. Thus, the term "and / or" as used herein in phrases such as "A and / or B" is intended to include "A and B", "A or B", "A" (single), and "B" (single). Similarly, the term "and / or" when used in phrases such as "A, B, and / or C" is intended to encompass each of the following aspects: A, B, and C, A, B, or C, A or C, A or B, B or C, A and C, A and B, B and C, A (single), B (single), and C (single).

[0053] Whenever an embodiment is described herein in conjunction with the phrase "comprising," it is understood that other similar embodiments described in terms of "consisting of" and / or "consisting essentially of" are also provided.

[0054] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the art to which this disclosure pertains. For example, Concise Dictionary of Biomedicine and Molecular Biology, Juo, Pei-Show, 2nd ed., 2002, CRC Press; The Dictionary of Cell and Molecular Biology, 3rd ed., 1999, Academic Press, and Oxford Dictionary Of Biochemistry And Molecular Biology, Revised, 2000, Oxford University Press provide those of ordinary skill in the art with a general dictionary of many of the terms used in this disclosure.

[0055] Units, prefixes, and symbols are shown in their Systeme International de Unites (SI) accepted form. Numeric ranges are inclusive of the numbers defining the range. Unless otherwise indicated, nucleotide sequences are written left to right in 5' to 3' orientation. Amino acid sequences are written left to right in amino to carboxy orientation. The headings provided herein are not intended to limit the various aspects of the disclosure, which can be had by reference to the specification in its entirety. Accordingly, the terms defined immediately below are more fully defined by reference to the specification in its entirety.

[0056] "Administering" refers to the physical introduction of an agent into a subject using any of a variety of methods and delivery systems known to those skilled in the art. Examples of routes of administration of the formulations disclosed herein include intravenous, intramuscular, subcutaneous, intraperitoneal, spinal or other parenteral routes of administration, for example by injection or infusion. The phrase "parenteral administration" as used herein means a method of administration other than enteral and topical administration, usually by injection, including, but not limited to, intravenous, intramuscular, intraarterial, intrathecal, intralymphatic, intralesional, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intrathecal, epidural and intrasternal injection and infusion, and in vivo electroporation. In some embodiments, the formulation is administered by a non-parenteral route, for example orally. Other non-parenteral routes include topical, epithelial, or mucosal routes of administration, such as intranasally, intravaginally, rectally, sublingually, or topically, and administration can be, for example, once, multiple times, and / or over one or more extended periods of time.

[0057] The term "T cell receptor" (TCR) as used herein refers to a heteromeric cell surface receptor that can specifically interact with a target antigen. As used herein, the term "TCR" includes, but is not limited to, naturally occurring and non-naturally occurring TCRs, full-length TCRs and their antigen-binding portions, chimeric TCRs, TCR fusion constructs, and synthetic TCRs. In humans, TCRs are expressed on the surface of T cells and are responsible for T cell recognition and targeting of antigen-presenting cells. Antigen-presenting cells (APCs) present fragments of foreign proteins (antigens) complexed with major histocompatibility complexes (MHC, also referred to herein as those complexed with HLA molecules, e.g., HLA class 1 molecules). TCRs recognize and bind antigen:HLA complexes and recruit CD3 (expressed by T cells) to activate the TCR. Activated TCRs initiate downstream signaling and immune responses, including the destruction of EPCs.

[0058] Generally, TCRs can comprise two chains, alpha and beta (or less frequently gamma and delta), interconnected by disulfide bonds. Each chain comprises a variable domain (alpha and beta chain variable domains) and a constant region (alpha and beta chain constant region). The variable domain is located distal to the cell membrane, and the variable domain interacts with the antigen. The constant region is located proximal to the cell membrane. TCRs can further comprise a transmembrane region and a short cytoplasmic tail. As used herein, the term "constant region" encompasses the transmembrane region and cytoplasmic tail, if present, as well as the conventional "constant region."

[0059] Variable domains can be further subdivided into regions of hypervariability, termed complementarity determining regions (CDRs), flanked by more conserved regions, termed framework regions (FRs). Each alpha and beta chain variable domain contains three CDRs and four FRs (FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4). Each variable domain contains a binding domain that interacts with antigen. All three CDRs of each chain are involved in antigen binding, although CDR3 is thought to be the major antigen binding region. CDR1 also interacts with antigen, while CD2 is thought to primarily recognize the HLA complex.

[0060] Unless expressly stated and unless the context dictates otherwise, the term "TCR" also includes antigen-binding fragments or portions of any TCR disclosed herein, including monovalent and bivalent fragments or portions, and single chain TCRs. The term "TCR" is not limited to naturally occurring TCRs bound to the surface of T cells. As used herein, the term "TCR" further refers to a TCR described herein expressed on the surface of a cell other than a T cell (e.g., a cell that naturally expresses or is modified to express CD3 as described herein), or a TCR described herein that does not include a cell membrane (e.g., an isolated TCR or a soluble TCR).

[0061] An "antigen binding molecule," "part of a TCR," or "TCR fragment" refers to any portion of a TCR that is less than the entire TCR. An antigen binding molecule can include the antigenic complementarity determining regions (CDRs).

[0062] "Antigen" refers to any molecule, e.g., a peptide, that can induce an immune response or that can be bound by a TCR. "Epitope" as used herein refers to a portion of a polypeptide that can induce an immune response or that can be bound by a TCR. The immune response can include antibody production, or activation of specific immunocompetent cells, or both. Those skilled in the art will readily appreciate that virtually any macromolecule, including any protein or peptide, can function as an antigen. Antigens and / or epitopes can be endogenously expressed, i.e., expressed by genomic DNA, or recombinantly expressed. Antigens and / or epitopes can be specific to a particular tissue, such as cancer cells, or they can be broadly expressed. In addition, fragments of larger molecules can function as antigens. In one aspect, the antigen is a tumor antigen. An epitope can be present in a longer polypeptide (e.g., in a protein), or an epitope can be present as a fragment of a longer polypeptide. In some embodiments, the epitope is complexed with a major histocompatibility complex (MHC; also referred to herein as a complex with an HLA molecule, eg, an HLA class I molecule).

[0063] The term "HLA" as used herein refers to human leukocyte antigens. HLA genes code for human major histocompatibility complex (MHC) proteins. MHC proteins are expressed on the surface of cells and are involved in activating immune responses. HLA class I genes code for MHC class I molecules that are expressed on the surface of cells in complexes with peptide fragments of self or non-self proteins (antigens). T cells expressing TCR and CD3 recognize the antigen:MHC class I complex and initiate an immune response to target and destroy antigen-presenting cells that present non-self proteins.

[0064] As used herein, "HLA class I molecule" or "HLA class I molecule" refers to the protein product of a wild-type or mutant HLA class I gene that encodes an MHC class I molecule. Thus, "HLA class I molecule" and "MHC class I molecule" are used interchangeably herein.

[0065] MHC class I molecules contain two protein chains: an alpha chain and a β2-microglobulin (β2m) chain. Human β2m is encoded by the B2M gene. The alpha chain of MHC class I molecules is encoded by the HLA gene complex. The HLA complex is located in the 6p21.3 region of the short arm of human chromosome 6 and contains more than 220 genes of diverse functions. HLA genes are highly mutated, with more than 20,000 HLA alleles and associated alleles, including more than 15,000 HLA class I alleles known in the art, encoding thousands of HLA proteins, including more than 10,000 HLA class I proteins (see, e.g., hla.alleles.org, last accessed February 27, 2019). The HLA complex contains at least three genes, HLA-A, HLA-B, and HLA-C, that code for MHC class I alpha chain proteins. In addition, HLA-E, HLA-F, and HLA-G encode proteins that associate with MHC class I molecules.

[0066] The term "autologous" refers to any material derived from the same individual that is subsequently reintroduced. For example, autologous T cell therapy involves administering to a subject T cells isolated from the same subject. The term "allogeneic" refers to any material derived from one individual that is then introduced into another individual of the same species. For example, allogeneic T cell transplantation involves administering to a subject T cells obtained from a donor other than the subject.

[0067] "Cancer" refers to a broad group of different diseases characterized by the uncontrolled growth of abnormal cells in the body. Uncontrolled cell division and growth leads to the formation of malignant tumors that infiltrate adjacent tissues and may metastasize to distant parts of the body via the lymphatic system or bloodstream. "Cancer" or "cancerous tissue" may include tumors. Examples of cancers that may be treated by the methods of the present invention include, but are not limited to, cancers of the immune system, including lymphomas, leukemias, and other white blood cell malignancies. In some embodiments, the methods of the present invention are directed to the treatment of cancers including, for example, bone cancer, kidney cancer, prostate cancer, breast cancer, colon cancer, lung cancer, cutaneous or intraocular malignant melanoma, pancreatic cancer, skin cancer, head and neck cancer, cutaneous or intraocular malignant melanoma, uterine cancer, ovarian cancer, rectal cancer, anal cancer, gastric cancer, testicular cancer, uterine cancer, fallopian tube cancer, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, Hodgkin's disease, non-Hodgkin's lymphoma (NHL), primary mediastinal large B-cell lymphoma (PMBC), diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), transformed follicular lymphoma, splenic marginal zone lymphoma (SMZL), esophageal cancer, small intestine cancer, endocrine system cancer, thyroid cancer, parathyroid cancer, adrenal cancer, sarcoma of soft tissue, urinary It can be used to reduce tumor size in tumors from uterine cancer, penile cancer, chronic or acute leukemia, acute myeloid leukemia (AML), chronic myeloid leukemia, acute lymphoblastic leukemia (ALL) (including non-T-cell ALL), chronic lymphocytic leukemia (CLL), childhood solid tumors, lymphocytic lymphoma, bladder cancer, kidney or ureter cancer, renal pelvis cancer, central nervous system (CNS) neoplasms, primary CNS lymphoma, tumor angiogenesis, spinal axis tumor, brain stem glioma, pituitary adenoma, Kaposi's sarcoma, epidermoid carcinoma, squamous cell carcinoma, T-cell lymphoma, environmentally induced cancers including those caused by asbestos, other B-cell malignancies, and combinations of the above cancers. Certain cancers may respond to chemotherapy or radiation therapy or the cancer may be refractory. Refractory cancer refers to cancer that cannot be corrected by surgical intervention and which either does not respond to chemotherapy or radiation therapy initially or becomes unresponsive over time.

[0068] As used herein, "anti-tumor effect" refers to a biological effect that can be manifested as a reduction in tumor volume, a reduction in the number of tumor cells, a reduction in tumor cell proliferation, a reduction in the number of metastases, an increase in overall survival or progression-free survival, an increase in life expectancy, or an improvement in various physiological symptoms associated with tumors.Anti-tumor effect can also refer to the prevention of tumor development, for example, vaccination.

[0069] As used herein, the term "progression-free survival", which may be abbreviated as PFS, refers to the time from the date of treatment to the date of disease progression or death from any cause according to the revised IWG response criteria for malignant lymphomas.

[0070] As used herein, "disease progression" or "progressive disease", which may be abbreviated as PD, refers to the worsening of one or more symptoms associated with a particular disease.For example, disease progression in a subject suffering from cancer can include the increase in the number or size of one or more malignant lesions, tumor metastasis, and death.

[0071] As used herein, "duration of response," which may be abbreviated as DOR, refers to the time from a subject's first objective response according to the revised IWG response criteria for malignant lymphoma to the date of confirmed disease progression or death.

[0072] The term "overall survival," sometimes abbreviated as OS, is defined as the time from the date of treatment to the date of death.

[0073] As used herein, a "cytokine" is a non-antibody protein released by one cell in response to contact with a specific antigen, which interacts with a second cell and mediates a response in the second cell. Cytokines may be endogenously expressed in cells or may be administered to a subject. Cytokines may be released from immune cells, including macrophages, B cells, T cells, and mast cells, to convey an immune response. Cytokines may induce various responses in recipient cells. Cytokines may include homeostatic cytokines, chemokines, proinflammatory cytokines, effectors, and acute phase proteins. For example, homeostatic cytokines, including interleukin (IL) 7 and IL-15, can promote survival and proliferation of immune cells, and proinflammatory cytokines can promote inflammatory responses. Examples of homeostatic cytokines include, but are not limited to, IL-2, IL-4, IL-5, IL-7, IL-10, IL-12p40, IL-12p70, IL-15, and interferon (IFN) gamma. Examples of proinflammatory cytokines include, but are not limited to, IL-1a, IL-1b, IL-6, IL-13, IL-17a, tumor necrosis factor (TNF)-alpha, TNF-beta, fibroblast growth factor (FGF)2, granulocyte-macrophage colony-stimulating factor (GM-CSF), soluble intercellular adhesion molecule 1 (sICAM-1), soluble vascular adhesion molecule 1 (sVCAM-1), vascular endothelial growth factor (VEGF), VEGF-C, VEGF-D, and placenta growth factor (PLGF). Examples of effectors include, but are not limited to, granzyme A, granzyme B, soluble Fas ligand (sFasL), and perforin. Examples of acute phase proteins include, but are not limited to, C-reactive protein (CRP) and serum amyloid A (SAA).

[0074] A "chemokine" is a type of cytokine that mediates cell chemotaxis or directional movement. Examples of chemokines include, but are not limited to, IL-8, IL-16, eotaxin, eotaxin-3, macrophage-derived chemokine (MDC or CCL22), monocyte chemoattractant protein 1 (MCP-1 or CCL2), MCP-4, macrophage inflammatory protein 1 alpha (MIP-1α, MIP-1a), MIP-1β (MIP-1b), gamma-inducible protein 10 (IP-10), and thymus and activation-regulated chemokine (TARC or CCL17).

[0075] Other examples of analytes and cytokines of the present invention include, but are not limited to, chemokine (CC motif) ligand (CCL) 1, CCL5, monocyte specific chemokine 3 (MCP3 or CCL7), monocyte chemoattractant protein 2 (MCP-2 or CCL8), CCL13, IL-1, IL-3, IL-9, IL-11, IL-12, IL-14, IL-17, IL-20, IL-21, granulocyte colony stimulating factor (G-CSF), leukemia inhibitory factor (LIF), oncolytic factor (CTAF), leukemia inhibitory factor (LEF ... These include tatin M (OSM), CD154, lymphotoxin (LT) beta, 4-1BB ligand (4-1BBL), proliferation-inducing ligand (APRIL), CD70, CD153, CD178, glucocorticoid-induced TNFR-related ligand (GITRL), tumor necrosis factor superfamily member 14 (TNFSF14), OX40L, TNF- and ApoL-related leukocyte-expressed ligand 1 (TALL-1), or TNF-related apoptosis-inducing ligand (TRAIL).

[0076] A "therapeutically effective amount," "effective dose," "effective amount," or "therapeutically effective dosage" of a drug or therapeutic agent is any amount of drug that, when used alone or in combination with another therapeutic agent, protects a subject against the onset of disease or promotes disease regression as evidenced by a decrease in the severity of disease symptoms, an increase in the frequency and duration of symptom-free periods of the disease, or prevention of impairment or disability due to the affliction of the disease. The ability of a therapeutic agent to promote disease regression can be evaluated using a variety of methods known to the skilled physician, for example, by assaying the activity of the agent in human subjects in clinical trials, in animal model systems predictive of efficacy in humans, or in in vitro assays.

[0077] The term "lymphocyte" as used herein includes natural killer (NK) cells, T cells, or B cells. NK cells are a type of cytotoxic (cell-toxic) lymphocyte and are a major component of the innate immune system. NK cells reject tumor and virus-infected cells. It functions through the process of apoptosis or programmed cell death. They were called "natural killers" because they do not require activation to kill cells. T cells play a major role in cell-mediated immunity (without the involvement of antibodies). The T cell receptor (TCR) distinguishes T cells from other lymphocyte types. The thymus, a specialized organ of the immune system, is primarily responsible for the maturation of T cells. There are six types of T cells, namely, helper T cells (e.g., CD4+ cells), cytotoxic T cells (TCs, also known as cytotoxic T lymphocytes, CTLs, T killer cells, cytolytic T cells, CD8+ T cells, or killer T cells), memory T cells ((i) stem memory T cells such as naive cells, SCM The cells are CD45RO-, CCR7+, CD45RA+, CD62L+ (L-selectin), CD27+, CD28+, and IL-7Rα+, but they express large amounts of CD95, IL-2Rβ, CXCR3, and LFA-1, and display numerous functional attributes characteristic of memory cells; (ii) central memory T CMThe cells express L-selectin and CCR7, they secrete IL-2 but not IFNγ or IL-4, and (iii) effector memory T EM These include T cells (which do not express L-selectin or CCR7 but produce effector cytokines such as IFNγ and IL-4), regulatory T cells (Treg, suppressor T cells, or CD4+CD25+ regulatory T cells), natural killer T cells (NKT), and gamma delta T cells. On the other hand, B cells play a major role in humoral immunity (involving the participation of antibodies). B cells make antibodies and antigens, act as antigen-presenting cells (APCs), and turn into memory B cells after activation by antigen interaction. In mammals, immature B cells are formed in the bone marrow, from which they get their name.

[0078] The term "genetically engineered" or "engineered" refers to a method of modifying the genome of a cell, including, but not limited to, deleting a coding or non-coding region or a portion thereof, or inserting a coding region or a portion thereof. In some embodiments, the modified cell is a lymphocyte, such as a T cell, or a modified cell expressing CD3, which can be obtained from a patient or a donor. The cell can be modified to express an exogenous construct, such as, for example, a T cell receptor (TCR) as disclosed herein, which is integrated into the genome of the cell. In some embodiments, the cell is modified to express CD3.

[0079] The term "modified" as used herein refers to the modification or manipulation of a target, e.g., a polypeptide, a polynucleotide, or a cell, to change one or more aspects of the target. For example, a "modified" HLA binding pocket refers to an HLA binding pocket that is altered compared to a wild-type HLA binding pocket. In some embodiments, the modification is made to the structure of the target, e.g., the structure of the HLA binding pocket. In some embodiments, the modification is to the amino acid sequence of the target, e.g., the modification comprises amino acid mutation (e.g., amino acid substitution). The term "modified" is not intended to be limited to a particular means of modifying the target. In some embodiments, the modification is achieved using a gene editing tool. In some embodiments, the gene editing tool is a CRISPR / Cas system, a transcription activator-like effector nuclease (TALEN), a meganuclease, or a zinc finger nuclease (ZFN) system.

[0080] The term "engineered" as used herein in the context of a cell refers to a cell that has been modified to contain a heterologous polynucleotide or that has been genetically engineered in a manner that alters the sequence, function, and / or expression of an endogenous polynucleotide. In some embodiments, the genome of the cell is genetically modified, for example, using gene editing tools. In some embodiments, the cell is engineered to express a heterologous polypeptide, for example, a heterologous HLA molecule, a chimeric antigen receptor (CAR), a heterologous T cell receptor (TCR), or any combination thereof.

[0081] The term "HLA binding pocket" or variations thereof refers to the peptide (e.g., antigen) binding site of an HLA molecule. The HLA binding pocket is formed by a β-sheet floor containing eight antiparallel β-sheets packed into two antiparallel α-helices to form a channel. In HLA class I molecules, the binding groove is divided into six pockets, A-F, which are defined by specific polymorphic amino acid residues that determine their tissue distribution and functionality. The B and F pockets are where the major anchor residues, the second and last positions of the peptide (P2 and PΩ, respectively), bind to HLA class I. These class I HLA molecules typically bind peptides that are 8-11 amino acids in length. Structures of peptide / HLA complexes show that conserved hydrogen bonds form between HLA side chains and the peptide backbone of nine core amino acids in the bound peptide. Additional HLA allele-specific interactions form between peptide side chains and structural pockets within the antigen binding pocket.

[0082] "Immune response" refers to the actions of cells of the immune system (e.g., T lymphocytes, B lymphocytes, natural killer (NK) cells, macrophages, eosinophils, mast cells, dendritic cells, and neutrophils) and soluble macromolecules produced either by such cells or the liver (including antibodies (Abs), cytokines, and complement) that result in the selective targeting of, binding to, damaging, destroying, and / or elimination from the vertebrate body of invading pathogens, pathogen-infected cells or tissues, cancerous cells or other abnormal cells, or, in the case of autoimmunity or pathological inflammation, normal human cells or tissues.

[0083] The term "immunotherapy" refers to the treatment of a subject suffering from a disease or at risk of suffering from or relapsing from a disease by a method that involves inducing, enhancing, suppressing, or otherwise modifying immune response. Examples of immunotherapy include, but are not limited to, T cell therapy. T cell therapy includes adoptive T cell therapy, tumor infiltrating lymphocyte (TIL) immunotherapy, autologous cell therapy, engineered autologous cell therapy (eACT), and allogeneic T cell transplantation.

[0084] The cells used in the immunotherapy described herein can be derived from sources known in the art. For example, T cells can be differentiated in vitro from a hematopoietic stem cell population, or the T cells can be obtained from a subject. T cells can be obtained, for example, from peripheral blood mononuclear cells, bone marrow, lymph node tissue, umbilical cord blood, thymus tissue, tissue from a site of infection, ascites, pleural effusion, spleen tissue, and tumors. Additionally, T cells can be obtained from one or more T cell lines available in the art. T cells can also be obtained from a unit of blood drawn from a subject using any number of techniques known to those skilled in the art, such as FICOLL™ separation and / or apheresis. Additional methods of isolating T cells for T cell therapy are disclosed in U.S. Patent Publication No. 2013 / 0287748, which is incorporated herein by reference in its entirety. Immunotherapy can also include administering modified cells to a subject, where the modified cells express CD3 and TCR as disclosed herein. In some embodiments, the modified cells are not T cells.

[0085] As used herein, a "patient" includes any human suffering from cancer (e.g., lymphoma or leukemia). The terms "subject" and "patient" are used interchangeably herein.

[0086] The terms "peptide", "polypeptide" and "protein" are used interchangeably and refer to compounds composed of amino acid residues covalently linked by peptide bonds. A protein or peptide must contain at least two amino acids, and there is no limit to the maximum number of amino acids that may comprise a protein or peptide sequence. A polypeptide includes any peptide or protein that contains two or more amino acids linked together by peptide bonds. As used herein, the term refers to both short chains, also commonly referred to in the art as peptides, oligopeptides and oligomers, for example, as well as longer chains, of which there are many varieties, commonly referred to in the art as proteins. "Polypeptides" include, for example, biologically active fragments, substantially homologous polypeptides, oligopeptides, homodimers, heterodimers, variants of polypeptides, modified polypeptides, derivatives, analogs, fusion proteins, among others. The polypeptides include natural peptides, recombinant peptides, synthetic peptides, or combinations thereof.

[0087] As used herein, "stimulation" refers to a primary response induced by the binding of a stimulatory molecule to its cognate ligand, which mediates signal transduction events. A "stimulatory molecule" is a molecule on a T cell, e.g., a T cell receptor (TCR) / CD3 complex, that specifically binds to a cognate stimulatory ligand present on an antigen-presenting cell. A "stimulatory ligand" is a ligand that, when present on an antigen-presenting cell (e.g., aAPC, dendritic cell, B cell, etc.), specifically binds to a stimulatory molecule on a T cell, thereby mediating a primary response by the T cell, including, but not limited to, activation, initiation of an immune response, proliferation, etc. Stimulatory ligands include, but are not limited to, MHC class I molecules loaded with peptides, anti-CD3 antibodies, superagonist anti-CD28 antibodies, and superagonist anti-CD2 antibodies.

[0088] The terms "conditioning" and "preconditioning" are used interchangeably herein to refer to preparing a patient in need of T cell therapy. Conditioning as used herein includes, but is not limited to, reducing the number of endogenous lymphocytes, removing cytokine sinks, increasing serum levels of one or more homeostatic cytokines or proinflammatory factors, enhancing effector function of T cells administered after conditioning, enhancing antigen presenting cell activation and / or availability, or any combination thereof prior to T cell therapy. In one embodiment, "adjusting to the appropriate state" includes increasing the serum levels of one or more cytokines, such as interleukin 7 (IL-7), interleukin 15 (IL-15), interleukin 10 (IL-10), interleukin 5 (IL-5), gamma-inducible protein 10 (IP-10), interleukin 8 (IL-8), monocyte chemoattractant protein 1 (MCP-1), placenta growth factor (PLGF), C-reactive protein (CRP), soluble intercellular adhesion molecule 1 (sICAM-1), soluble vascular adhesion molecule 1 (sVCAM-1), or any combination thereof. In another embodiment, "adjusting to the appropriate state" includes increasing the serum levels of IL-7, IL-15, IP-10, MCP-1, PLGF, CRP, or any combination thereof.

[0089] "Treatment" or "treating" of a subject refers to any type of intervention or process performed on a subject, or the administration of an active agent to a subject, for the purpose of reversing, mitigating, ameliorating, inhibiting, slowing, or preventing the onset, progression, development, severity, or recurrence of a symptom, complication, or condition, or biochemical manifestations associated with a disease. In one aspect, "treatment" or "treating" includes partial remission. In another aspect, "treatment" or "treating" includes complete remission.

[0090] The use of the alternative (e.g., "or") should be understood to mean either one, both, or any combination of the alternatives. As used herein, the indefinite article "a" or "an" should be understood to refer to "one or more" of the listed or shown components.

[0091] The term "about" or "essentially comprising" refers to a value or composition that is within an acceptable error range of a particular value or composition as determined by one of ordinary skill in the art, which depends in part on how the value or composition is measured or determined, i.e., the limitations of the measurement system. For example, "about" or "essentially comprising" can mean within or more than one standard deviation per the practice in the art. Alternatively, "about" or "essentially comprising" can mean a range of up to 10% (i.e., ±10%). For example, about 3 mg can include any number between 2.7 mg and 3.3 mg (at 10%). Furthermore, particularly with respect to biological systems or processes, the term can mean up to an order of magnitude or up to 5 times the value. When a particular value or composition is provided in this application and claims, unless otherwise indicated, the meaning of "about" or "essentially comprising" should be assumed to be within an acceptable error range of that particular value or composition.

[0092] As described herein, any concentration range, percentage range, ratio range, or integer range, unless otherwise specified, should be understood to include any integer value within the recited range, and, where appropriate, fractions thereof (such as 1 / 10 and 1 / 100 of an integer).

[0093] Various aspects of the invention are described in further detail in the following subsections.

[0094] II. Methods of the Disclosure Some embodiments of the present disclosure are directed to a method of adjusting a subject in need of treatment to a suitable state, comprising modifying the HLA binding pocket of an HLA molecule expressed on a cell of the subject. Some embodiments of the present disclosure are directed to a method of enhancing an immune response in a subject in need thereof, comprising modifying the HLA binding pocket of an HLA molecule expressed on a cell of the subject. Other embodiments of the present disclosure are directed to a method of increasing the binding affinity of a peptide to an HLA molecule on a cell, comprising modifying the binding pocket of HLA on the cell. In some embodiments, the cell is present in a subject in need of treatment.

[0095] Some embodiments of the present disclosure are directed to a method for enhancing the immunogenicity of an antigen in a subject in need of treatment, comprising modifying the HLA binding pocket of an HLA molecule on a cell in the subject, wherein the HLA molecule can bind to the antigen.Some embodiments of the present disclosure are directed to a method for enhancing the immune response to a treatment in a subject in need thereof, comprising modifying the HLA binding pocket of an HLA molecule on a cell of the subject.In some embodiments, the treatment comprises immunotherapy.Other embodiments of the present disclosure are directed to a method for enhancing the immune response to an antigen in a subject in need thereof, comprising modifying the binding pocket of HLA on a cell of the subject.

[0096] In some embodiments, the modified HLA binding pocket has high binding affinity for the antigen, hi some embodiments, the binding affinity is increased by at least about 10%, at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 100%, at least about 125%, at least about 150%, at least about 175%, at least about 200%, at least about 250%, at least about 300%, at least about 350%, at least about 400%, at least about 450%, or at least about 500% compared to the binding affinity of the unmodified HLA molecule.

[0097] In some embodiments, cells comprising an HLA molecule comprising a modified HLA binding pocket have increased surface display of antigen-HLA complexes compared to cells comprising an unmodified HLA molecule, hi some embodiments, the surface display of antigen-HLA complexes is increased by at least about 10%, at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 100%, at least about 125%, at least about 150%, at least about 175%, at least about 200%, at least about 250%, at least about 300%, at least about 350%, at least about 400%, at least about 450%, or at least about 500% compared to the surface display of antigen-HLA complexes of an unmodified HLA molecule.

[0098] In some embodiments, the cells are antigen-presenting cells. In some embodiments, the cells are dendritic cells. In some embodiments, the cells are artificial antigen-presenting cells. In some embodiments, the artificial antigen-presenting cells comprise beads (e.g., silicate beads, glass beads, metal beads, or combinations thereof), nanovesicles, microvesicles, exosomes, endosomes, or any combinations thereof. In some embodiments, the cells are in vivo. In some embodiments, the cells are ex vivo. In some embodiments, the cells are allogeneic cells. In some embodiments, the cells are donor cells, i.e., cells obtained from a subject other than the subject who may ultimately receive the cells.

[0099] In some embodiments, an antigen-HLA complex comprising an HLA molecule having a modified HLA binding pocket as disclosed herein elicits a greater antigen-specific T cell response when contacted with a T cell, hi some embodiments, the antigen-specific T cell response is increased by at least about 10%, at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 100%, at least about 125%, at least about 150%, at least about 175%, at least about 200%, at least about 250%, at least about 300%, at least about 350%, at least about 400%, at least about 450%, or at least about 500% compared to the antigen-specific T cell response when the T cell is contacted with an antigen-HLA complex comprising an unmodified HLA molecule.

[0100] In some embodiments, antigen-HLA complexes comprising HLA molecules having modified HLA binding pockets as disclosed herein increase proliferation of tumor antigen-specific T cells. In some embodiments, the antigen-specific T cell response is increased by at least about 10%, at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 100%, at least about 125%, at least about 150%, at least about 175%, at least about 200%, at least about 250%, at least about 300%, at least about 350%, at least about 400%, at least about 450%, or at least about 500% compared to proliferation of tumor antigen-specific T cells when the T cells are contacted with antigen-HLA complexes comprising unmodified HLA molecules.

[0101] In some embodiments, the antigen is a tumor antigen. In some embodiments, the antigen is an antigen expressed by a pathogen. In some embodiments, the antigen is a viral antigen. In some embodiments, the antigen is a bacterial antigen. In some embodiments, the antigen is a fungal antigen.

[0102] II.A. HLA Class I Molecules Described herein are modified HLA molecules with enhanced antigen binding affinity and methods of use thereof. Any HLA molecule can be used in the methods and compositions of the disclosure. In some embodiments, the HLA molecule is an HLA class I molecule. The HLA class I molecule can be any HLA class I molecule. In some embodiments, the HLA class I molecule is selected from HLA-A, HLA-B, HLA-C, HLA-E, HLA-F, HLA-G, HLA-K, or HLA-L alleles, or any combination thereof. In some embodiments, the HLA class I molecule is selected from HLA-A, HLA-B, and HLA-C alleles. In some embodiments, the HLA class I molecule is selected from HLA-E, HLA-F, and HLA-G alleles. In some embodiments, the HLA class I molecule is an HLA-A allele. In certain embodiments, the HLA class I molecule is an HLA-B allele. In a particular embodiment, the HLA class I molecule is an HLA-C allele.

[0103] Many HLA class I alleles are known, including many HLA-A, HLA-B, and HLA-C alleles, and any of the known alleles can be used in the present disclosure. An updated list of HLA alleles is available at hla.alleles.org / (last visited October 7, 2021).

[0104] II.A.1. HLA-A alleles In some embodiments, the HLA class I molecule is an HLA-A allele. Any HLA-A allele can be used in the methods and compositions of the present disclosure. In some embodiments, the HLA molecule is an allele selected from HLA-A*01, HLA-A*02, HLA-A*03, HLA-A*11, HLA-A*23, HLA-A*24, HLA-A*25, HLA-A*26, HLA-A*29, HLA-A*30, HLA-A*31, HLA-A*32, HLA-A*33, HLA-A*34, HLA-A*36, HLA-A*43, HLA-A*66, HLA-A*68, HLA-A*69, HLA-A*74, and HLA-A*80. In some embodiments, the HLA molecule is an allele of the HLA-A*24 superfamily of alleles. In some embodiments, the HLA molecule is an HLA-A*24 allele. In some embodiments, the HLA molecule is an HLA-A*23 allele. In some embodiments, the HLA molecule is an HLA-A*32 allele.

[0105] The details are HLA-A*23:01:01:01, HLA-A*23:01:01:02, HLA-A*23: 01:01:03、SONG-A*23:01:01:04、SONG-A*23:01:01:05、SONG-A*23:01:01:0 、SONG-A*23:01:01:07、SONG-A*23:01:01:08、SONG-A*23:01:01:09、SONG-A*2 3:01:01:10、SONG-A*23:01:01:11、SONG-A*23:01:01:12、SONG-A*23:01:01: 13、SONGS-A*23:01:01:14、SONGS-A*23:01:01:15、SONGS-A*23:01:01:16、SONGS-A *23:01:01:17、SONG-A*23:01:01:18、SONG-A*23:01:01:19、SONG-A*23:01:0 1:20, SONG-A*23:01:01:21, SONG-A*23:01:01:22, SONG-A*23:01:01:23, SONG -A*23:01:01:24、SONG-A*23:01:01:25、SONG-A*23:01:01:26、SONG-A*23:0 :02、SONG-A*23:01:03、SONG-A*23:01:04、SONG-A*23:01:05、SONG-A*23:01:0 6、SONG-A*23:01:07、SONG-A*23:01:08、SONG-A*23:01:09、SONG-A*23:01:10、 SONG-A*23:01:11, SONG-A*23:01:12, SONG-A*23:01:13, SONG-A*23:01:14, HL A-A*23:01:15, SONG-A*23:01:16, SONG-A*23:01:17, SONG-A*23:01:18, SONG- A*23:01:19, SONG-A*23:01:20, SONG-A*23:01:21, SONG-A*23:01:22, SONG-A* 23:01:23、SONGS-A*23:01:24、SONGS-A*23:01:25、SONGS-A*23:01:26、SONGS-A*2 :01:27、SONG-A*23:01:28、SONG-A*23:01:29、SONG-A*23:01:30、SONG-A*23:0 1:31、SONG-A*23:01:32、SONG-A*23:01:33、SONG-A*23:01:34、SONG-A*23:02、HLA-A*23:03:01、HLA-A*23:03:02:01、HLA-A*23:03:02:02、HLA-A*23:04、HLA-A*23:05、HLA-A*23:06、HLA-A*23:07、HLA-A*23:08、HLA-A*23:09、HLA-A*23:10、HLA-A*23:11、HLA-A*23:12、HLA-A*23:13、HLA-A*23:14:01、HLA-A*23:14:02、HLA-A*23:15、HLA-A*23:16、HLA-A*23:17:01:01、HLA-A* 23:17:01:02、HLA-A*23:17:01:03、HLA-A*23:17:02、HLA-A*23:17:03、HLA-A*23:18、HLA-A*23:19、HLA-A*23:20、HLA-A*23:21、HLA-A*23:22、HLA-A*23:23、HLA-A*23:24、HLA-A*23:25、HLA-A*23:26、HLA-A*23:27、HLA-A*23:28、HLA-A*23:29、HLA-A*23:30、HLA-A*23:31、HLA-A*23:32、HLA-A*23:3 3、HLA-A*23:34、HLA-A*23:35、HLA-A*23:36、HLA-A*23:37:01、HLA-A*23:37:02、HLA-A*23:38、HLA-A*23:39、HLA-A*23:40、HLA-A*23:41、HLA-A*23:42、HLA-A*23:43、HLA-A*23:44、HLA-A*23:45、HLA-A*23:46、HLA-A*23:47、HLA-A*23:48、HLA-A*23:49、HLA-A*23:50、HLA-A*23:51、HLA-A*23:52、HL A-A*23:53、HLA-A*23:54、HLA-A*23:55、HLA-A*23:56、HLA-A*23:57、HLA-A*23:58、HLA-A*23:59、HLA-A*23:60、HLA-A*23:61、HLA-A*23:62、HLA-A*23:63、HLA-A*23:64、HLA-A*23:65、HLA-A*23:66、HLA-A*23:67、HLA-A*23:68、HLA-A*23:70、HLA-A*23:71、HLA-A*23:72、HLA-A*23:73、HLA-A*23:74、SONG-A*23:75, SONG-A*23:76, SONG-A*23:77, SONG-A*23:78, SONG-A*23:79, SONG-A*23: 80、SONGS-A*23:81、SONGS-A*23:82、SONGS-A*23:83、SONGS-A*23:84、SONGS-A*23:85、SONGS-A*2 3:86, SONG-A*23:87, SONG-A*23:88, SONG-A*23:89, SONG-A*23:90, SONG-A*23:91, SONG- A*23:92, SONG-A*23:93, SONG-A*23:94, SONG-A*23:95, SONG-A*23:96, SONG-A*23:97, HL A-A*23:98、SONGS-A*23:99、SONGS-A*23:100、SONGS-A*23:101、SONGS-A*23:102、SONGS-A*23 :103、SONG-A*23:104、SONG-A*23:105、SONG-A*23:106、SONG-A*23:107、SONG-A*23:108、 SONG-A*23:109, SONG-A*23:110, SONG-A*23:111, SONG-A*23:112, SONG-A*23:113, SONG-A *23:114, SONG-A*23:115, SONG-A*23:116, and SONG-A*23:1

[0106] The timings are HLA-A*24:02:01:01, HLA-A*24:02:01:02, HLA-A*24: 02:01:03、SONG-A*24:02:01:04、SONG-A*24:02:01:05、SONG-A*24:02:01:0 、SONG-A*24:02:01:07、SONG-A*24:02:01:08、SONG-A*24:02:01:09、SONG-A*2 4:02:01:10、SONG-A*24:02:01:11、SONG-A*24:02:01:12、SONG-A*24:02:01: 13、SONGS-A*24:02:01:14、SONGS-A*24:02:01:15、SONGS-A*24:02:01:16、SONGS- A*24:02:01:17、SONG-A*24:02:01:18、SONG-A*24:02:01:19、SONG-A*24:02: 01:20, SONG-A*24:02:01:21, SONG-A*24:02:01:22, SONG-A*24:02:01:23, HL A-A*24:02:01:24、SONG-A*24:02:01:25、SONG-A*24:02:01:26、SONG-A*24:0 2:01:27、SONG-A*24:02:01:28、SONG-A*24:02:01:29、SONG-A*24:02:01:3 、SONG-A*24:02:01:31、SONG-A*24:02:01:32、SONG-A*24:02:01:33、SONG-A*2 4:02:01:34、SONG-A*24:02:01:35、SONG-A*24:02:01:36、SONG-A*24:02:01: 37、SONGS-A*24:02:01:38、SONGS-A*24:02:01:39、SONGS-A*24:02:01:40、SONGS-A *24:02:01:41、SONG-A*24:02:01:42、SONG-A*24:02:01:43、SONG-A*24:02: 01:44, SONG-A*24:02:01:45, SONG-A*24:02:01:46, SONG-A*24:02:01:47, HL A-A*24:02:01:48、SONG-A*24:02:01:49、SONG-A*24:02:01:50、SONG-A*24:0 2:01:51, SONG-A*24:02:01:52, SONG-A*24:02:01:53, SONG-A*24:02:01:54,HLA-A*24:02:01:55、HLA-A*24:02:01:56、HLA-A*24:02:01:57、HLA-A*24:02:01:58、HLA-A*24:02:01:59、HLA-A*24:02:01:60、HLA-A*24:02:01:61、HLA-A*24:02:01:62、HLA-A*24:02:01:63、HLA-A*24:02:01:64、HLA-A*24:02:01:65、HLA-A*24:02:01:66、HLA-A*24:02:01:67、HLA-A*24:02:01:6 8、HLA-A*24:02:01:69、HLA-A*24:02:01:70、HLA-A*24:02:01:71、HLA-A*24:02:01:72、HLA-A*24:02:01:73、HLA-A*24:02:01:74、HLA-A*24:02:01:75、HLA-A*24:02:01:76、HLA-A*24:02:01:77、HLA-A*24:02:01:78、HLA-A*24:02:01:79、HLA-A*24:02:01:80、HLA-A*24:02:01:81、HLA-A*24:02:01 :82、HLA-A*24:02:01:83、HLA-A*24:02:01:84、HLA-A*24:02:01:85、HLA-A*24:02:01:86、HLA-A*24:02:01:87、HLA-A*24:02:01:88、HLA-A*24:02:01:89、HLA-A*24:02:01:90、HLA-A*24:02:01:91、HLA-A*24:02:01:92、HLA-A*24:02:01:93、HLA-A*24:02:01:94、HLA-A*24:02:01:95、HLA-A*24:02: 01:96、HLA-A*24:02:01:97、HLA-A*24:02:01:98、HLA-A*24:02:01:99、HLA-A*24:02:01:100、HLA-A*24:02:01:101、HLA-A*24:02:01:102、HLA-A*24:02:01:103、HLA-A*24:02:01:104、HLA-A*24:02:01:105、HLA-A*24:02:01:106、HLA-A*24:02:01:107、HLA-A*24:02:01:108、HLA-A*24:02:01:109、HLA-A*24:02:01:110、HLA-A*24:02:02、HLA-A*24:02:03、HLA-A*24:02:04、HLA-A*24:02:05、HLA-A*24:02:06、HLA-A*24:02:07、HLA-A*24:02:08、HLA-A*24:02:09、HLA-A*24:02:10、HLA-A*24:02:11、HLA-A*24:02:12、HLA-A*24:02:13、HLA-A*24:02:14、HLA-A*24:02:15、HLA-A*24:02:16、HLA-A* 24:02:17、HLA-A*24:02:18、HLA-A*24:02:19、HLA-A*24:02:20、HLA-A*24:02:21、HLA-A*24:02:22、HLA-A*24:02:23、HLA-A*24:02:24、HLA-A*24:02:25、HLA-A*24:02:26、HLA-A*24:02:27、HLA-A*24:02:28、HLA-A*24:02:29、HLA-A*24:02:30、HLA-A*24:02:31、HLA-A*24:02:32、HLA-A*24:02:33、H LA-A*24:02:34、HLA-A*24:02:35、HLA-A*24:02:36、HLA-A*24:02:37、HLA-A*24:02:38、HLA-A*24:02:39、HLA-A*24:02:40:01、HLA-A*24:02:40:02、 HLA-A*24:02:41、HLA-A*24:02:42、HLA-A*24:02:43、HLA-A*24:02:44、HLA-A*24:02:45、HLA-A*24:02:46、HLA-A*24:02:47、HLA-A*24:02:48、HLA-A *24:02:49、HLA-A*24:02:50、HLA-A*24:02:51、HLA-A*24:02:52、HLA-A*24:02:53、HLA-A*24:02:54、HLA-A*24:02:55、HLA-A*24:02:56、HLA-A*24:02:57、HLA-A*24:02:58、HLA-A*24:02:59、HLA-A*24:02:60、HLA-A*24:02:61、HLA-A*24:02:62、HLA-A*24:02:63、HLA-A*24:02:64、HLA-A*24:02:65、HLA-A*24:02:66、HLA-A*24:02:67、HLA-A*24:02:68、HLA-A*24:02:69、HLA-A*24:02:70、HLA-A*24:02:71、HLA-A*24:02:72、HLA-A*24:02:73、HLA-A*24:02:74、HLA-A*24:02:75、HLA-A*24:02:76、HLA-A*24:02:77、HLA-A*24:02:78、HLA-A*24:02:79、HLA-A*24:02:80、HLA-A*24:02:81、HLA-A* 24:02:82、HLA-A*24:02:83、HLA-A*24:02:84、HLA-A*24:02:85、HLA-A*24:02:86、HLA-A*24:02:87、HLA-A*24:02:88、HLA-A*24:02:89、HLA-A*24:02:90、HLA-A*24:02:91、HLA-A*24:02:92、HLA-A*24:02:93、HLA-A*24:02:94、HLA-A*24:02:95、HLA-A*24:02:96、HLA-A*24:02:97、HLA-A*24:02:9 8、HLA-A*24:02:99、HLA-A*24:02:100、HLA-A*24:02:101、HLA-A*24:02:102:01、HLA-A*24:02:102:02、HLA-A*24:02:103、HLA-A*24:02:104、HLA-A*24:02:105、HLA-A*24:02:106、HLA-A*24:02:107、HLA-A*24:02:108、HLA-A*24:02:109、HLA-A*24:02:110、HLA-A*24:02:111、HLA-A*24:02:112 ,HLA-A*24:02:113,HLA-A*24:02:114,HLA-A*24:02:115:01,HLA-A*24:02:115:02,HLA-A*24:02:116,HLA-A*24:02:117,HLA-A*24:02:118,HLA-A*24:02:119,HLA-A*24:02:120,HLA-A*24:02:121,HLA-A*24:02:122,HLA-A*24:02:123,HLA-A*24:02:124,HLA-A*24:02:125,HLA-A*24:02:126,HLA-A*24:02:127、HLA-A*24:02:128、HLA-A*24:02:129、HLA-A*24:02:130、HLA-A*24:02:131、HLA-A*24:02:132、HLA-A*24:02:133、HLA-A*24:02:134、HLA-A*24:02:135、HLA-A*24:02:136、HLA-A*24:02:137、HLA-A*24:02:138、HLA-A*24:02:139、HLA-A*24:02:140、HLA-A*24:02:141、HLA-A*24:0 2:142、HLA-A*24:02:143、HLA-A*24:02:144、HLA-A*24:02:145、HLA-A*24:02:146、HLA-A*24:02:147、HLA-A*24:02:148、HLA-A*24:02:149、HLA-A*24:03:01:01、HLA-A*24:03:01:02、HLA-A*24:03:01:03、HLA-A*24:03:02、HLA-A*24:03:03、HLA-A*24:03:04、HLA-A*24:04、HLA-A*24:05:01、HLA-A* 24:05:02、HLA-A*24:06、HLA-A*24:07:01:01、HLA-A*24:07:01:02、HLA-A*24:07:01:03、HLA-A*24:07:02、HLA-A*24:07:03、HLA-A*24:07:04、HLA-A*24:08、HLA-A*24:09、HLA-A*24:10:01:01、HLA-A*24:10:01:02、HLA-A*24:10:02、HLA-A*24:11、HLA-A*24:13:01、HLA-A*24:13:02、HLA-A*24:14:0 1:01、HLA-A*24:14:01:02、HLA-A*24:14:01:03、HLA-A*24:14:01:04、HLA-A*24:15、HLA-A*24:17:01:01、HLA-A*24:17:01:02、HLA-A*24:18、HLA-A*24:19、HLA-A*24:20:01:01、HLA-A*24:20:01:02、HLA-A*24:20:02、HLA-A*24:21:01、HLA-A*24:21:02、HLA-A*24:22、HLA-A*24:23、HLA-A*24:24、HLA-A*24:25、HLA-A*24:26、HLA-A*24:27、HLA-A*24:28、HLA-A*24:29、HLA-A*24:30、HLA-A*24:31、HLA-A*24:32、HLA-A*24:33、HLA-A*24:34、HLA-A*24:35、H, LA-A*24:36, SONG-A*24:37, SONG-A*24:38, SONG-A*24:39, SONG-A*24:40, HL A-A*24:41, SONG-A*24:42, SONG-A*24:43, SONG-A*24:44, SONG-A*24:45, SONG- A*24:46, SONG-A*24:47, SONG-A*24:48, SONG-A*24:49, SONG-A*24:50, SONG-A* 24:51、SONGS-A*24:52、SONGS-A*24:53、SONGS-A*24:54、SONGS-A*24:55、SONGS-A*2 :56:01、SONG-A*24:56:02、SONG-A*24:57、SONG-A*24:58、SONG-A*24:59、SONG- A*24:60, SONG-A*24:61, SONG-A*24:62, SONG-A*24:63, SONG-A*24:64, SONG-A* 24:66、SONGS-A*24:67、SONGS-A*24:68、SONGS-A*24:69、SONGS-A*24:70、SONGS-A*2 :71、SONGS-A*24:72、SONGS-A*24:73、SONGS-A*24:74:01、SONGS-A*24:74:02、SONGS- A*24:75, SONG-A*24:76, SONG-A*24:77, SONG-A*24:78, SONG-A*24:79, SONG-A* 24:80、SONGS-A*24:81、SONGS-A*24:82、SONGS-A*24:83、SONGS-A*24:84、SONGS-A*2 :85、SONGS-A*24:86、SONGS-A*24:87、SONGS-A*24:88、SONGS-A*24:89、SONGS-A*24:9 0:01、SONG-A*24:90:02、SONG-A*24:91、SONG-A*24:92、SONG-A*24:93、SONG-A* 24:94、SONGS-A*24:95、SONGS-A*24:96、SONGS-A*24:97、SONGS-A*24:98、SONGS-A*2 :99、SONGS-A*24:100、SONGS-A*24:101、SONGS-A*24:102、SONGS-A*24:103、SONGS-A* 24:104, SONG-A*24:105, SONG-A*24:106, SONG-A*24:107, SONG-A*24:108, SONG -A*24:109, SONG-A*24:110, SONG-A*24:111, SONG-A*24:112, SONG-A*24:1HLA-A*24:114、HLA-A*24:115、HLA-A*24:116、HLA-A*24:117、HLA-A*24:118、HLA-A*24:119、HLA-A*24:120、HLA-A*24:121、HLA-A*24:122、HLA-A*24:123、HLA-A*24:124、HLA-A*24:125、HLA-A*24:126、HLA-A*24:127、HLA-A*24:128、HLA-A*24:129、HLA-A*24:130、HLA-A*24:131、HLA-A*24:132、H LA-A*24:133、HLA-A*24:134、HLA-A*24:135:01、HLA-A*24:135:02、HLA-A*24:136、HLA-A*24:137、HLA-A*24:138、HLA-A*24:139、HLA-A*24:140、HL A-A*24:141、HLA-A*24:142:01、HLA-A*24:142:02、HLA-A*24:143、HLA-A*24:144、HLA-A*24:145、HLA-A*24:146、HLA-A*24:147、HLA-A*24:148、HLA-A*24:148 -A*24:149, HLA-A*24:150, HLA-A*24:151, HLA-A*24:152, HLA-A*24:153, HLA-A*24:154, HLA-A*24:155, HLA-A*24:156, HLA-A*24:157, HLA-A*24:158, HLA-A*24:159, HLA-A*24:160, HLA-A*24:161, HLA-A*24:162, HLA-A*24:163, HLA-A*24:164, HLA-A*24:165, HLA-A*24:166, HLA-A*24:167, HLA- A*24:168、HLA-A*24:169、HLA-A*24:170、HLA-A*24:171、HLA-A*24:172:01、HLA-A*24:172:02、HLA-A*24:173、HLA-A*24:174、HLA-A*24:175、HLA-A*24:176、HLA-A*24:177、HLA-A*24:178、HLA-A*24:179、HLA-A*24:180、HLA-A*24:181、HLA-A*24:182、HLA-A*24:183、HLA-A*24:184、HLA-A-24:185、HLA-A*24:186、HLA-A*24:187、HLA-A*24:188、HLA-A*24:189、HLA-A*24:190、HLA-A*24:191、HLA-A*24:192、HLA-A*24:193、HLA-A*24:194、HLA-A*24:195、HLA-A*24:196、HLA-A*24:197、HLA-A*24:198、HLA-A*24:199、HLA-A*24:200、HLA-A*24:201、HLA-A*24:202、HLA-A*24:203、HLA-A*24:204、H LA-A*24:205、HLA-A*24:206、HLA-A*24:207:01、HLA-A*24:207:02、HLA-A*24:208:01、HLA-A*24:208:02:01、HLA-A*24:208:02:02、HLA-A*24:209、 HLA-A*24:210、HLA-A*24:212、HLA-A*24:213、HLA-A*24:214、HLA-A*24:215、HLA-A*24:216、HLA-A*24:217、HLA-A*24:218、HLA-A*24:219、HLA-A*24 :220、HLA-A*24:221、HLA-A*24:222、HLA-A*24:223、HLA-A*24:224、HLA-A*24:225:01、HLA-A*24:225:02、HLA-A*24:226:01、HLA-A*24:226:02、HLA-A*24:227、HLA-A*24:228、HLA-A*24:229、HLA-A*24:230、HLA-A*24:231、HLA-A*24:232、HLA-A*24:233、HLA-A*24:234、HLA-A*24:235、HLA-A*24:23 6、HLA-A*24:237、HLA-A*24:238、HLA-A*24:239、HLA-A*24:240、HLA-A*24:241、HLA-A*24:242、HLA-A*24:243、HLA-A*24:244、HLA-A*24:245、HLA-A*24:246、HLA-A*24:247、HLA-A*24:248、HLA-A*24:249、HLA-A*24:250、HLA-A*24:251、HLA-A*24:252、HLA-A*24:253、HLA-A*24:254、HLA-A*24:255、HLA-A*24:256、HLA-A*24:257、HLA-A*24:258、HLA-A*24:259、HLA-A*24:260、HLA-A*24:261、HLA-A*24:262、HLA-A*24:263、HLA-A*24:264、HLA-A*24:265、HLA-A*24:266、HLA-A*24:267、HLA-A*24:268、HLA-A*24:269、HLA-A*24:270、HLA-A*24:271、HLA-A*24:272、HLA-A*24:273、HLA-A*24:274、HLA -A*24:275, HLA-A*24:276, HLA-A*24:277, HLA-A*24:278, HLA-A*24:279, HLA-A*24:280, HLA-A*24:281, HLA-A*24:282, HLA-A*24:283, HLA-A*24:284, HLA-A*24:285, HLA-A*24:286, HLA-A*24:287, HLA-A*24:288, HLA-A*24:289, HLA-A*24:290, HLA-A*24:291, HLA-A*24:292, HLA-A*24:293, HLA-A* 24:294、HLA-A*24:295、HLA-A*24:296、HLA-A*24:297、HLA-A*24:298、HLA-A*24:299、HLA-A*24:300、HLA-A*24:301、HLA-A*24:302、HLA-A*24:303、HLA-A*24:304、HLA-A*24:305、HLA-A*24:306、HLA-A*24:307、HLA-A*24:308、HLA-A*24:309、HLA-A*24:310:01、HLA-A*24:310:02、HLA-A*24:311、HLA -A*24:312、HLA-A*24:313:01、HLA-A*24:313:02、HLA-A*24:314、HLA-A*24:315、HLA-A*24:316、HLA-A*24:317、HLA-A*24:318、HLA-A*24:319、HLA-A*24:320、HLA-A*24:321、HLA-A*24:322、HLA-A*24:323、HLA-A*24:324、HLA-A*24:325、HLA-A*24:326、HLA-A*24:327、HLA-A*24:328、HLA-A*24:329、HLA-A*24:330, HLA-A*24:331, HLA-A*24:332, HLA-A*24:333, HLA-A*24:334, HLA-A*24:335, HLA-A*24:336, HLA-A*24:337, HLA-A*24:338, HLA-A*24:339, HLA-A*24:340, HLA-A*24:341, HLA-A*24:342, HLA-A*24:343, HLA-A*24:344, HLA-A*24:345, HLA-A*24:346, HLA-A*24:347:01, HLA-A*24:34 7:02, HLA-A*24:348, HLA-A*24:349, HLA-A*24:350, HLA-A*24:351, HLA-A*24:352, HLA-A*24:353, HLA-A*24:354, HLA-A*24:355, HLA-A*24:356, HLA-A*24:357, HLA-A*24:358, HLA-A*24:359, HLA-A*24:360, HLA-A*24:361, HLA-A*24:362, HLA-A*24:363, HLA-A*24:364, HLA-A*24:365, HLA-A*24:3 66、HLA-A*24:367、HLA-A*24:368、HLA-A*24:369、HLA-A*24:370、HLA-A*24:371、HLA-A*24:372、HLA-A*24:373、HLA-A*24:374、HLA-A*24:375、HLA-A*24:376、HLA-A*24:377、HLA-A*24:378、HLA-A*24:379、HLA-A*24:380、HLA-A*24:381、HLA-A*24:382、HLA-A*24:383、HLA-A*24:384、HLA-A-24:38 5、HLA-A*24:386、HLA-A*24:387、HLA-A*24:388、HLA-A*24:389、HLA-A*24:390、HLA-A*24:391、HLA-A*24:392、HLA-A*24:393、HLA-A*24:394、HLA-A*24:395、HLA-A*24:396、HLA-A*24:397、HLA-A*24:398、HLA-A*24:399、HLA-A*24:400、HLA-A*24:401、HLA-A*24:402、HLA-A*24:403、HLA-A*24:404、HLA-A*24:405、HLA-A*24:406、HLA-A*24:407、HLA-A*24:408、HLA-A*24:409、HLA-A*24:410、HLA-A*24:411、HLA-A*24:412、HLA-A*24:413、HLA-A*24:414、HLA-A*24:415、HLA-A, *24:416、HLA-A*24:417、HLA-A*24:418、HLA-A*24:419、HLA-A*24:420、HLA-A*24:421、HLA-A*24:422、HLA-A*24:423、HLA-A*24:424、HLA-A*24:425、HLA-A*24:426、HLA-A*24:427、HLA-A*24:428、HLA-A*24:429、HLA-A*24:430、HLA-A*24:431:01、HLA-A*24:431:02、HLA-A*24:432、HLA-A*24:433、HL A-A*24:434、HLA-A*24:435、HLA-A*24:436、HLA-A*24:437、HLA-A*24:438、HLA-A*24:439、HLA-A*24:440、HLA-A*24:441、HLA-A*24:442、HLA-A*24:443、HLA-A*24:444、HLA-A*24:445、HLA-A*24:446、HLA-A*24:447、HLA-A*24:448、HLA-A*24:449、HLA-A*24:450、HLA-A*24:451、HLA-A*24:452、HLA-A *24:453、HLA-A*24:454、HLA-A*24:455、HLA-A*24:456、HLA-A*24:457、HLA-A*24:458、HLA-A*24:459、HLA-A*24:460:01:01、HLA-A*24:460:01:02、HLA-A*24:461、HLA-A*24:462、HLA-A*24:463、HLA-A*24:464、HLA-A*24:465、HLA-A*24:466、HLA-A*24:467、HLA-A*24:468、HLA-A*24:469、HLA-A*24: 470、HLA-A*24:472、HLA-A*24:473、HLA-A*24:474、HLA-A*24:475、HLA-A*24:476、HLA-A*24:477、HLA-A*24:478、HLA-A*24:479、HLA-A*24:480、HLA-A*24:481、HLA-A*24:482、HLA-A*24:483、HLA-A*24:484、HLA-A*24:485、HLA-A*24:486、HLA-A*24:487、HLA-A*24:488、HLA-A*24:489、HLA-A*24:490、HLA-A alleles selected from HLA-A*24:491, HLA-A*24:492, HLA-A*24:493, HLA-A*24:494, HLA-A*24:495, HLA-A*24:496, HLA-A*24:497, HLA-A*24:498, HLA-A*24:499, HLA-A*24:500, HLA-A*24:501, HLA-A*24:502, HLA-A*24:503, HLA-A*24:504, HLA-A*24:505, HLA-A*24:506, HLA-A*24:507, HLA-A*24:508, HLA-A*24:509, HLA-A*24:510, HLA-A*24:511, HLA-A*24:512, HLA-A*24:513, HLA-A*24:514, HLA-A*24:515, HLA-A*24:516, HLA-A*24:517, HLA-A*24:518, HLA-A*24:519, HLA-A*24:520, HLA-A*24:521, HLA-A*24:522, HLA-A*24:523, HLA-A*24:524, HLA-A*24:525, HLA-A*24:526, HLA-A*24:527, HLA-A*24:528, HLA-A*24:529, HLA-A*24:530, HLA-A*24:531, HLA-A*24:532, HLA-A*24:533, HLA-A*24:534, HLA-A*24:535, HLA-A*24:536, HLA-A*24:537, HLA-A*24:538, HLA-A*24:539, HLA-A*24:540, HLA-A*24:541, HLA-A*24:542, HLA-A*24:543, HLA-A*24:544, HLA-A*24:545, HLA-A*24:546, HLA-A*24:547, HLA-A*24:548, HLA-A*24:549, HLA-A*24:550, and HLA-A*24:551. In some embodiments, the HLA molecule is the HLA-A*24:02 allele.,

[0107] Its values ​​are HLA-A*25:01:01:01, HLA-A*25:01:01:02, HLA-A*25:0 1:01:03、SONG-A*25:01:01:04、SONG-A*25:01:01:05、SONG-A*25:01:01:06、 SONG-A*25:01:01:07、SONG-A*25:01:02、SONG-A*25:01:03、SONG-A*25:01:0 、SONG-A*25:01:05、SONG-A*25:01:06、SONG-A*25:01:07、SONG-A*25:01:08、H LA-A*25:01:09、SONGS-A*25:01:10、SONGS-A*25:01:11、SONGS-A*25:01:12、SONGS -A*25:01:13、SONG-A*25:01:14、SONG-A*25:01:15、SONG-A*25:01:16、SONG-A *25:01:17、SONGS-A*25:01:18、SONGS-A*25:01:19、SONGS-A*25:01:20、SONGS-A*2 5:01:21、SONGS-A*25:02、SONGS-A*25:03、SONGS-A*25:04、SONGS-A*25:05、SONGS-A*2 5:06、SONGS-A*25:07、SONGS-A*25:08、SONGS-A*25:09、SONGS-A*25:10、SONGS-A*25: 11. SONG-A*25:12, SONG-A*25:13, SONG-A*25:14, SONG-A*25:15, SONG-A*25:1 、SONGS-A*25:17、SONGS-A*25:18、SONGS-A*25:19:01、SONGS-A*25:19:02、SONGS-A*2 5:20、SONG-A*25:21、SONG-A*25:22、SONG-A*25:23、SONG-A*25:24、SONG-A*25: 25、SONG-A*25:26、SONG-A*25:27:01、SONG-A*25:27:02、SONG-A*25:28、SONG-A *25:29、SONGS-A*25:30、SONGS-A*25:31、SONGS-A*25:32、SONGS-A*25:33、SONGS-A*2 5:34、SONGS-A*25:35、SONGS-A*25:36、SONGS-A*25:37、SONGS-A*25:38、SONGS-A*25: 39、SONG-A*25:40、SONG-A*25:41、SONG-A*25:42、SONG-A*25:43、SONG-A*25:44、SONG-A*25:45, SONG-A*25:46, SONG-A*25:47, SONG-A*25:48, SONG-A*25:49, SONG-A*25:50, SONG-A*25:51, SONG-A*25:52, SONG-A *25:53、SONGS-A*25:54、SONGS-A*25:55、SONGS-A*25:56、SONGS-A*25:57、SONGS-A*25:58、SONGS-A*25:59、SONGS-A*25:60、SONGS-A*25:61 、SONG-A*25:62、SONG-A*25:63、SONG-A*25:64、SONG-A*25:65、SONG-A*25:66、SONG-A*25:67、SONG-A*25:68、SONG-A*25:69、SONG-A *25:70、SONGS-A*25:71、SONGS-A*25:72、SONGS-A*25:73、SONGS-A*2 :74, HLA-A*25:75, and HLA-A*25:7

[0108] Its values ​​are HLA-A*31:01:02:01, HLA-A*31:01:02:02, HLA-A*31:0 1:02:03, SONG-A*31:01:02:04, SONG-A*31:01:02:05, SONG-A*31:01:02:06, SONG-A*31:01:02:07、SONG-A*31:01:02:08、SONG-A*31:01:02:09、SONG-A*3 :01:02:10、SONG-A*31:01:02:11、SONG-A*31:01:02:12、SONG-A*31:01:02:1 3, SONG-A*31:01:02:14, SONG-A*31:01:02:15, SONG-A*31:01:02:16, SONG-A* 31:01:02:17、SONG-A*31:01:02:18、SONG-A*31:01:02:19、SONG-A*31:01:02 :20、SONG-A*31:01:02:21、SONG-A*31:01:02:22、SONG-A*31:01:02:23、SONG- A*31:01:02:24、SONG-A*31:01:02:25、SONG-A*31:01:02:26、SONG-A*31:01:0 2:27, SONG-A*31:01:02:28, SONG-A*31:01:02:29, SONG-A*31:01:02:30, SONG -A*31:01:02:31、SONG-A*31:01:02:32、SONG-A*31:01:02:33、SONG-A*31:0 :02:34、SONG-A*31:01:02:35、SONG-A*31:01:02:36、SONG-A*31:01:02:37、H LA-A*31:01:02:38, SONG-A*31:01:02:39, SONG-A*31:01:02:40, SONG-A*31: 01:02:41、SONGS-A*31:01:03、SONGS-A*31:01:04、SONGS-A*31:01:05、SONGS-A*3 :01:06、SONGS-A*31:01:07、SONGS-A*31:01:08、SONGS-A*31:01:09、SONGS-A*31:0 1:10、SONG-A*31:01:11、SONG-A*31:01:12、SONG-A*31:01:13、SONG-A*31:01: 14、SONG-A*31:01:15、SONG-A*31:01:16、SONG-A*31:01:17、SONG-A*31:01:18、HLA-A*31:01:19、HLA-A*31:01:20、HLA-A*31:01:21、HLA-A*31:01:22、HLA-A*31:01:23、HLA-A*31:01:24、HLA-A*31:01:25、HLA-A*31:01:26、HLA-A*31:01:27、HLA-A*31:01:28、HLA-A*31:01:29、HLA-A*31:01:30、HLA-A*31:01:31、HLA-A*31:01:32、HLA-A*31:01:33、HLA-A*31:01:34、HLA-A*31: 01:35、HLA-A*31:01:36、HLA-A*31:01:37、HLA-A*31:01:38、HLA-A*31:01:39、HLA-A*31:01:40、HLA-A*31:01:41、HLA-A*31:01:42、HLA-A*31:01:43、HLA-A*31:01:44、HLA-A*31:01:45、HLA-A*31:01:46、HLA-A*31:01:47、HLA-A*31:02:01、HLA-A*31:02:02、HLA-A*31:03、HLA-A*31:04:01:01、HLA -A*31:04:01:02, HLA-A*31:04:02, HLA-A*31:05, HLA-A*31:06, HLA-A*31:07, HLA-A*31:08, HLA-A*31:09, HLA-A*31:10, HLA-A*31:11, HLA-A*31:12, HLA-A*31:13, HLA-A*31:14, HLA-A*31:15, HLA-A*31:16, HLA-A*31:17, HLA-A*31:18, HLA-A*31:19, HLA-A*31:20, HLA-A*31:21, HLA-A*31:22, HLA -A*31:23, HLA-A*31:24, HLA-A*31:25, HLA-A*31:26, HLA-A*31:27, HLA-A*31:28, HLA-A*31:29, HLA-A*31:30, HLA-A*31:31, HLA-A*31:32, HLA-A*31:33, HLA-A*31:34, HLA-A*31:35, HLA-A*31:36, HLA-A*31:37, HLA-A*31:38, HLA-A*31:39, HLA-A*31:40, HLA-A*31:41, HLA-A*31:42, HLA-A*31:43,HLA-A*31:44、HLA-A*31:45、HLA-A*31:46、HLA-A*31:47、HLA-A*31:48、HLA-A*31:49、HLA-A*31:50、HLA-A*31:51、HLA-A*31:52、HLA-A*31:53、HLA-A*31:54、HLA-A*31:55、HLA-A*31:56、HLA-A*31:57、HLA-A*31:58、HLA-A*31:59、HLA-A*31:60、HLA-A*31:61、HLA-A*31:62、HLA-A*31:63、HLA-A*31: 64、HLA-A*31:65、HLA-A*31:66、HLA-A*31:67、HLA-A*31:68、HLA-A*31:69、HLA-A*31:70、HLA-A*31:71、HLA-A*31:72、HLA-A*31:73、HLA-A*31:74、HLA-A*31:75、HLA-A*31:76、HLA-A*31:77、HLA-A*31:78、HLA-A*31:79、HLA-A*31:80、HLA-A*31:81、HLA-A*31:82、HLA-A*31:83、HLA-A*31:84、HLA-A* 31:85、HLA-A*31:86、HLA-A*31:87、HLA-A*31:88、HLA-A*31:89、HLA-A*31:90、HLA-A*31:91、HLA-A*31:92、HLA-A*31:93、HLA-A*31:94、HLA-A*31:95、HLA-A*31:96、HLA-A*31:97、HLA-A*31:98、HLA-A*31:99、HLA-A*31:100、HLA-A*31:101、HLA-A*31:102、HLA-A*31:103、HLA-A*31:104、HLA-A*31:1 05、HLA-A*31:106、HLA-A*31:107、HLA-A*31:108、HLA-A*31:109、HLA-A*31:110、HLA-A*31:111、HLA-A*31:112、HLA-A*31:113、HLA-A*31:114、HLA-A*31:115、HLA-A*31:116、HLA-A*31:117、HLA-A*31:118、HLA-A*31:119、HLA-A*31:120、HLA-A*31:121、HLA-A*31:122、HLA-A*31:123、HLA-A*31:124、SONG-A*31:125, SONG-A*31:126, SONG-A*31:127, SONG-A*31:128, SONG-A*31: 129、SONGS-A*31:130、SONGS-A*31:131、SONGS-A*31:132、SONGS-A*31:133、SONGS-A* 31:134, SONG-A*31:135, SONG-A*31:136, SONG-A*31:137, SONG-A*31:138, HL A-A*31:139, SONG-A*31:140, SONG-A*31:141, SONG-A*31:142, SONG-A*31:1 、SONGS-A*31:144、SONGS-A*31:145、SONGS-A*31:146、SONGS-A*31:147、SONGS-A*3 :148、SONGS-A*31:149、SONGS-A*31:150、SONGS-A*31:151、SONGS-A*31:152、SONGS-A *31:153、SONGS-A*31:154、SONGS-A*31:155、SONGS-A*31:156、SONGS-A*31:157、H LA-A*31:158, SONG-A*31:159, SONG-A*31:160, SONG-A*31:161, SONG-A*31:1 2、SONGS-A*31:163、SONGS-A*31:164、SONGS-A*31:165、SONGS-A*31:166、SONGS-A*3 1:167, SONG-A*31:168, SONG-A*31:169, SONG-A*31:170, SONG-A*31:171, SONG- A*31:172, SONG-A*31:173, SONG-A*31:174, SONG-A*31:175, SONG-A*31:1 SONG-A*31:177, SONG-A*31:178, SONG-A*31:179, SONG-A*31:180, SONG-A*31:1 81、SONGS-A*31:182、SONGS-A*31:183、SONGS-A*31:184、SONGS-A*31:185、SONGS-A* 31:186, SONG-A*31:187, SONG-A*31:188, SONG-A*31:189, SONG-A*31:190, SONG -A*31:191, SONG-A*31:192, SONG-A*31:193, SONG-A*31:194, SONG-A*31:195 SONG-A*31:196, SONG-A*31:197, and SONG-A*31:1

[0109] The details are HLA-A*32:01:01:01, HLA-A*32:01:01:02, HLA-A*32: 01:01:03、SONG-A*32:01:01:04、SONG-A*32:01:01:05、SONG-A*32:01:01:0 、SONG-A*32:01:01:07、SONG-A*32:01:01:08、SONG-A*32:01:01:09、SONG-A*3 2:01:01:10、SONG-A*32:01:01:11、SONG-A*32:01:01:12、SONG-A*32:01:01: 13、SONG-A*32:01:01:14、SONG-A*32:01:01:15、SONG-A*32:01:01:16、SONG-A *32:01:01:17、SONG-A*32:01:01:18、SONG-A*32:01:01:19、SONG-A*32:01:0 1:20、SONGS-A*32:01:01:21、SONGS-A*32:01:01:22、SONGS-A*32:01:01:23、SONGS -A*32:01:01:24、SONG-A*32:01:01:25、SONG-A*32:01:01:26、SONG-A*32:0 :01:27、SONG-A*32:01:01:28、SONG-A*32:01:01:29、SONG-A*32:01:01:30、H LA-A*32:01:02、SONGS-A*32:01:03、SONGS-A*32:01:04、SONGS-A*32:01:05、SONGS -A*32:01:06、SONG-A*32:01:07、SONG-A*32:01:08、SONG-A*32:01:09、SONG-A *32:01:10、SONGS-A*32:01:11、SONGS-A*32:01:12、SONGS-A*32:01:13、SONGS-A*3 2:01:14、SONGS-A*32:01:15、SONGS-A*32:01:16、SONGS-A*32:01:17、SONGS-A*32: 01:18、SONGS-A*32:01:19、SONGS-A*32:01:20、SONGS-A*32:01:21、SONGS-A*32:01 :22、SONG-A*32:01:23、SONG-A*32:01:24、SONG-A*32:01:25、SONG-A*32:01:2 6、SONG-A*32:01:27、SONG-A*32:01:28、SONG-A*32:01:29、SONG-A*32:01:30、HLA-A*32:01:31、HLA-A*32:01:32、HLA-A*32:01:33、HLA-A*32:01:34、HLA-A*32:01:35、HLA-A*32:01:36、HLA-A*32:01:37、HLA-A*32:01:38、HLA-A*32:01:39、HLA-A*32:01:40、HLA-A*32:01:41、HLA-A*32:01:42、HLA-A*32:01:43、HLA-A*32:01:44、HLA-A*32:01:45、HLA-A*32:01:46、HLA-A*32: 01:47, HLA-A*32:02, HLA-A*32:03:01:01, HLA-A*32:03:01:02, HLA-A*32:04, HLA-A*32:05, HLA-A*32:06, HLA-A*32:07, HLA-A*32:08, HLA-A*32:09, HLA-A*32:10, HLA-A*32:11, HLA-A*32:12, HLA-A*32:13, HLA-A*32:14, HLA-A*32:15, HLA-A*32:16, HLA-A*32:17, HLA-A*32:18, HLA-A*32:19, HLA -A*32:20、HLA-A*32:21、HLA-A*32:22、HLA-A*32:23、HLA-A*32:24、HLA-A*32:25、HLA-A*32:26:01、HLA-A*32:26:02、HLA-A*32:27、HLA-A*32:28、HLA-A*32:29、HLA-A*32:30:01、HLA-A*32:30:02、HLA-A*32:31、HLA-A*32:32、HLA-A*32:33:01、HLA-A*32:33:02、HLA-A*32:33:03、HLA-A*32:34、HLA -A*32:35, HLA-A*32:36, HLA-A*32:37, HLA-A*32:38, HLA-A*32:39, HLA-A*32:40, HLA-A*32:41, HLA-A*32:42, HLA-A*32:43:01, HLA-A*32:43:02, HLA-A*32:44, HLA-A*32:45, HLA-A*32:46:01, HLA-A*32:46:02, HLA-A*32:47, HLA-A*32:48, HLA-A*32:49, HLA-A*32:50, HLA-A*32:51, HLA-A*32:52,HLA-A*32:53、HLA-A*32:54、HLA-A*32:55:01、HLA-A*32:55:02、HLA-A*32:55:03、HLA-A*32:56、HLA-A*32:57、HLA-A*32:58、HLA-A*32:59、HLA-A*32:60、HLA-A*32:61、HLA-A*32:62、HLA-A*32:63、HLA-A*32:64、HLA-A*32:65、HLA-A*32:66、HLA-A*32:67、HLA-A*32:68、HLA-A*32:69、HLA-A*32:70、 HLA-A*32:71、HLA-A*32:72、HLA-A*32:73、HLA-A*32:74、HLA-A*32:75、HLA-A*32:76、HLA-A*32:77、HLA-A*32:78、HLA-A*32:79、HLA-A*32:80、HLA-A*32:81、HLA-A*32:82、HLA-A*32:83、HLA-A*32:84、HLA-A*32:85、HLA-A*32:86、HLA-A*32:87、HLA-A*32:88、HLA-A*32:89、HLA-A*32:90、HLA-A*32:9 1、HLA-A*32:92、HLA-A*32:93、HLA-A*32:94、HLA-A*32:95、HLA-A*32:96、HLA-A*32:97、HLA-A*32:98、HLA-A*32:99、HLA-A*32:100、HLA-A*32:101、HLA-A*32:102、HLA-A*32:103、HLA-A*32:104、HLA-A*32:105、HLA-A*32:106:01:01、HLA-A*32:106:01:02、HLA-A*32:107、HLA-A*32:108、HLA-A*32:1 09、HLA-A*32:110、HLA-A*32:111、HLA-A*32:112、HLA-A*32:113、HLA-A*32:114、HLA-A*32:115、HLA-A*32:116、HLA-A*32:117、HLA-A*32:118、HLA-A*32:119、HLA-A*32:120、HLA-A*32:121、HLA-A*32:122、HLA-A*32:123、HLA-A*32:124、HLA-A*32:125、HLA-A*32:126、HLA-A*32:127、HLA-A*32:128、SONG-A*32:129, SONG-A*32:130, SONG-A*32:131, SONG-A*32:132, SONG-A*32:133, SONG-A*32:134, SONG-A*32:135, SONG-A *32:136, SONG-A*32:137, SONG-A*32:138, SONG-A*32:139, SONG-A*32:140, SONG-A*32:141, SONG-A*32:142, SONG-A*32:1 3, SONG-A*32:144, SONG-A*32:145, SONG-A*32:146, SONG-A*32:147, SONG-A*32:148, SONG-A*32:149, SONG-A*32:150, SONG- A*32:151、SONGS-A*32:152、SONGS-A*32:153、SONGS-A*32:154、 HLA-A*32:155 and HLA-A*32:156.

[0110] In some embodiments, the HLA molecule of the present disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-A allele. In some embodiments, the modified HLA binding pocket is the F pocket of the HLA molecule. In some embodiments, the HLA molecule of the present disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an allele in the HLA-A*24 superfamily of alleles. In some embodiments, the HLA molecule of the present disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-A*24 allele, and the modified HLA binding pocket comprises an amino acid substitution at a position corresponding to amino acid residue 79, amino acid residue 80, amino acid residue 81, amino acid residue 82, or amino acid residue 83 of SEQ ID NO:1. In some embodiments, an HLA molecule of the disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-A*24:02 allele and the modified HLA binding pocket comprises an amino acid substitution at a position corresponding to amino acid residue 79, amino acid residue 80, amino acid residue 81, amino acid residue 82, or amino acid residue 83 of SEQ ID NO: 1. In some embodiments, an HLA molecule of the disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-A*24 allele and the modified HLA binding pocket comprises an amino acid substitution at amino acid residue 81, which corresponds to the amino acid sequence set forth in SEQ ID NO: 1. In some embodiments, an HLA molecule of the disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-A*24:02 allele and the modified HLA binding pocket comprises an amino acid substitution at amino acid residue 81, which corresponds to the amino acid sequence set forth in SEQ ID NO: 1. [Table 2]

[0111] In some embodiments, an HLA molecule of the disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-A*23 allele and the modified HLA binding pocket comprises an amino acid substitution at a position corresponding to amino acid residue 79, amino acid residue 80, amino acid residue 81, amino acid residue 82, or amino acid residue 83 of SEQ ID NO: 1. In some embodiments, an HLA molecule of the disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-A*23 allele and the modified HLA binding pocket comprises an amino acid substitution at amino acid residue 81, which corresponds to the amino acid sequence shown in SEQ ID NO: 1.

[0112] In some embodiments, an HLA molecule of the disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-A*25 allele and the modified HLA binding pocket comprises an amino acid substitution at a position corresponding to amino acid residue 79, amino acid residue 80, amino acid residue 81, amino acid residue 82, or amino acid residue 83 of SEQ ID NO: 1. In some embodiments, an HLA molecule of the disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-A*25 allele and the modified HLA binding pocket comprises an amino acid substitution at amino acid residue 81, which corresponds to the amino acid sequence shown in SEQ ID NO: 1.

[0113] In some embodiments, an HLA molecule of the disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-A*31 allele and the modified HLA binding pocket comprises an amino acid substitution at a position corresponding to amino acid residue 79, amino acid residue 80, amino acid residue 81, amino acid residue 82, or amino acid residue 83 of SEQ ID NO: 1. In some embodiments, an HLA molecule of the disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-A*31 allele and the modified HLA binding pocket comprises an amino acid substitution at amino acid residue 81, which corresponds to the amino acid sequence shown in SEQ ID NO: 1.

[0114] In some embodiments, an HLA molecule of the disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-A*32 allele and the modified HLA binding pocket comprises an amino acid substitution at a position corresponding to amino acid residue 79, amino acid residue 80, amino acid residue 81, amino acid residue 82, or amino acid residue 83 of SEQ ID NO: 1. In some embodiments, an HLA molecule of the disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-A*32 allele and the modified HLA binding pocket comprises an amino acid substitution at amino acid residue 81, which corresponds to the amino acid sequence shown in SEQ ID NO: 1.

[0115] In some embodiments, the original amino acid replaced is alanine. In some embodiments, the new amino acid replacing the original amino acid is selected from the group consisting of leucine, valine, isoleucine, methionine, phenylalanine, tyrosine, and tryptophan. In some embodiments, the new amino acid replacing the original amino acid is leucine. In some embodiments, the amino acid substitution is A81L, which corresponds to the amino acid sequence set forth in SEQ ID NO:1. In certain embodiments, the HLA molecule is an HLA-A*24:02 allele comprising a modified HLA binding pocket, and the modified HLA binding pocket comprises an A81L substitution, which corresponds to the amino acid sequence set forth in SEQ ID NO:1.

[0116] II.A.2. HLA-B alleles In some embodiments, the HLA molecule is an HLA-B allele. Any HLA-B allele can be used in the methods and compositions of the disclosure. In some embodiments, the HLA molecule is HLA-B*07, HLA-B*08, HLA-B*13, HLA-B*14, HLA-B*15, HLA-B*18, HLA-B*27, HLA-B*35, HLA-B*37, HLA-B*38, HLA-B*39, HLA-B*40, HLA-B*41, HLA-B*42, HLA-B*44, HLA-B*45, HLA-B*46, HLA-B*47, HLA-B*4 In some embodiments, the HLA molecule is HLA-B*44. In some embodiments, the HLA molecule is HLA-B*51. In some embodiments, the HLA molecule is HLA-B*58. In some embodiments, the HLA molecule is HLA-B*58.

[0117] In some embodiments, an HLA molecule of the disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-B*44 allele and the modified HLA binding pocket comprises an amino acid substitution at a position corresponding to amino acid residue 79, amino acid residue 80, amino acid residue 81, amino acid residue 82, or amino acid residue 83 of SEQ ID NO: 1. In some embodiments, an HLA molecule of the disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-B*44 allele and the modified HLA binding pocket comprises an amino acid substitution at amino acid residue 81, which corresponds to the amino acid sequence shown in SEQ ID NO: 1.

[0118] In some embodiments, an HLA molecule of the disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-B*51 allele and the modified HLA binding pocket comprises an amino acid substitution at a position corresponding to amino acid residue 79, amino acid residue 80, amino acid residue 81, amino acid residue 82, or amino acid residue 83 of SEQ ID NO: 1. In some embodiments, an HLA molecule of the disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-B*51 allele and the modified HLA binding pocket comprises an amino acid substitution at amino acid residue 81, which corresponds to the amino acid sequence shown in SEQ ID NO: 1.

[0119] In some embodiments, an HLA molecule of the disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-B*58 allele and the modified HLA binding pocket comprises an amino acid substitution at a position corresponding to amino acid residue 79, amino acid residue 80, amino acid residue 81, amino acid residue 82, or amino acid residue 83 of SEQ ID NO: 1. In some embodiments, an HLA molecule of the disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-B*58 allele and the modified HLA binding pocket comprises an amino acid substitution at amino acid residue 81, which corresponds to the amino acid sequence shown in SEQ ID NO: 1.

[0120] Its values ​​are HLA-B*44:02:01:01, HLA-B*44:02:01:02, HLA-B*44:0 2:01:03, SONG-B*44:02:01:04, SONG-B*44:02:01:05, SONG-B*44:02:01:06, SONG-B*44:02:01:07, SONG-B*44:02:01:08, SONG-B*44:02:01:09, SONG-B*4 :02:01:10, SONG-B*44:02:01:11, SONG-B*44:02:01:12, SONG-B*44:02:01:1 、SONG-B*44:02:01:14、SONG-B*44:02:01:15、SONG-B*44:02:01:16、SONG-B*4 4:02:01:17、SONGS-B*44:02:01:18、SONGS-B*44:02:01:19、SONGS-B*44:02:01: 20、SONG-B*44:02:01:21、SONG-B*44:02:01:22、SONG-B*44:02:01:23、SONG-B *44:02:01:24、SONGS-B*44:02:01:25、SONGS-B*44:02:01:26、SONGS-B*44:02:0 :27、SONG-B*44:02:01:28、SONG-B*44:02:01:29、SONG-B*44:02:01:30、SONG- B*44:02:01:31、SONG-B*44:02:01:32、SONG-B*44:02:01:33、SONG-B*44:02: 01:34, SONG-B*44:02:01:35, SONG-B*44:02:01:36, SONG-B*44:02:01:37, SONG-B*44:02:01:37, HL A-B*44:02:01:38、SONG-B*44:02:01:39、SONG-B*44:02:01:40、SONG-B*44:0 :01:41、SONG-B*44:02:01:42、SONG-B*44:02:01:43、SONG-B*44:02:01:44、H LA-B*44:02:01:45, SONG-B*44:02:01:46, SONG-B*44:02:01:47, SONG-B*44:0 2:01:48, SONG-B*44:02:01:49, SONG-B*44:02:01:50, SONG-B*44:02:01:51, SONGS-B*44:02:01:52、SONGS-B*44:02:02、SONGS-B*44:02:03、SONGS-B*44:02:04、SONG-B*44:02:05、SONG-B*44:02:06, SONG-B*44:02:07, SONG-B*44:02:08, SONG-B*44:02:08 A-B*44:02:09, SONG-B*44:02:10, SONG-B*44:02:11, SONG-B*44:02:12, SONG- B*44:02:13, SONG-B*44:02:14, SONG-B*44:02:15, SONG-B*44:02:16, SONG-B* 44:02:17、SONG-B*44:02:18、SONG-B*44:02:19、SONG-B*44:02:20、SONG-B*44: 02:21、SONG-B*44:02:22、SONG-B*44:02:23、SONG-B*44:02:24、SONG-B*44:0 :25、SONG-B*44:02:26、SONG-B*44:02:27、SONG-B*44:02:28、SONG-B*44:02:2 9, SONG-B*44:02:30, SONG-B*44:02:31, SONG-B*44:02:32, SONG-B*44:02:3 SONG-B*44:02:34, SONG-B*44:02:35, SONG-B*44:02:36, SONG-B*44:02:37, SONG -B*44:02:38、SONG-B*44:02:39、SONG-B*44:02:40、SONG-B*44:02:41、SONG-B *44:02:42、SONGS-B*44:02:43、SONGS-B*44:02:44、SONGS-B*44:02:45、SONGS-B*4 4:02:46、SONG-B*44:02:47、SONG-B*44:02:48、SONG-B*44:02:49:01、SONG-B* 44:02:49:02、SONG-B*44:02:50、SONG-B*44:02:51、SONG-B*44:02:52、SONG-B* 44:02:53、SONGS-B*44:02:54、SONGS-B*44:02:55、SONGS-B*44:02:56、SONGS-B*4 :02:57、SONGS-B*44:02:58、SONGS-B*44:02:59、SONGS-B*44:02:60、SONGS-B*44:0 2:61、SONG-B*44:02:62、SONG-B*44:02:63、SONG-B*44:02:64、SONG-B*44:02: 65、SONG-B*44:02:66、SONG-B*44:02:67、SONG-B*44:02:68、SONG-B*44:02:69、SONG-B*44:02:70, SONG-B*44:02:71, SONG-B*44:02:72, SONG-B*44:02:73, SONG-B*44:02:73, SONG-B*44:02:7 A-B*44:02:74, SONG-B*44:02:75, SONG-B*44:03:01:01, SONG-B*44:03:01:0 2、SONGS-B*44:03:01:03、SONGS-B*44:03:01:04、SONGS-B*44:03:01:05、SONGS-B* 44:03:01:06、SONGS-B*44:03:01:07、SONGS-B*44:03:01:08、SONGS-B*44:03:0 :09、SONG-B*44:03:01:10、SONG-B*44:03:01:11、SONG-B*44:03:01:12、SONG- B*44:03:01:13, SONG-B*44:03:01:14, SONG-B*44:03:01:15, SONG-B*44:03: 01:16, SONG-B*44:03:01:17, SONG-B*44:03:01:18, SONG-B*44:03:01:19, HL A-B*44:03:01:20, SONG-B*44:03:01:21, SONG-B*44:03:01:22, SONG-B*44:0 3:01:23, SONG-B*44:03:01:24, SONG-B*44:03:01:25, SONG-B*44:03:01:26, SONG-B*44:03:01:27, SONG-B*44:03:01:28, SONG-B*44:03:01:29, SONG-B*4 :03:01:30、SONGS-B*44:03:01:31、SONGS-B*44:03:01:32、SONGS-B*44:03:01:3 3, SONG-B*44:03:01:34, SONG-B*44:03:01:35, SONG-B*44:03:01:36, SONG-B* 44:03:01:37, SONG-B*44:03:01:38, SONG-B*44:03:01:39, SONG-B*44:03:0 :40、SONG-B*44:03:01:41、SONG-B*44:03:01:42、SONG-B*44:03:01:43、SONG- B*44:03:02:01、SONG-B*44:03:02:02、SONG-B*44:03:02:03、SONG-B*44:03: 03、SONG-B*44:03:04、SONG-B*44:03:05、SONG-B*44:03:06、SONG-B*44:03:07、SONG-B*44:03:08, SONG-B*44:03:09, SONG-B*44:03:10, SONG-B*44:03:11, SONG-B*44:03:11, SONG A-B*44:03:12, SONG-B*44:03:13, SONG-B*44:03:14, SONG-B*44:03:15, SONG- B*44:03:16, SONG-B*44:03:17, SONG-B*44:03:18, SONG-B*44:03:19, SONG-B* 44:03:20、SONGS-B*44:03:21、SONGS-B*44:03:22、SONGS-B*44:03:23、SONGS-B*4 :03:24、SONG-B*44:03:25、SONG-B*44:03:26、SONG-B*44:03:27、SONG-B*44:0 3:28、SONG-B*44:03:29、SONG-B*44:03:30、SONG-B*44:03:31、SONG-B*44:03: 32、SONG-B*44:03:33、SONG-B*44:03:34、SONG-B*44:03:35、SONG-B*44:03:3 、SONG-B*44:03:37、SONG-B*44:03:38、SONG-B*44:03:39、SONG-B*44:03:40、H LA-B*44:03:41、SONGS-B*44:03:42、SONGS-B*44:03:43、SONGS-B*44:03:44、SONGS -B*44:03:45、SONG-B*44:03:46、SONG-B*44:03:47、SONG-B*44:03:48、SONG-B *44:03:49、SONGS-B*44:03:50、SONGS-B*44:03:51、SONGS-B*44:03:52、SONGS-B*4 4:03:53、SONG-B*44:03:54、SONG-B*44:03:55、SONG-B*44:03:56、SONG-B*44: 03:57、SONGS-B*44:03:58、SONGS-B*44:04、SONGS-B*44:05:01:01、SONGS-B*44:05 :01:02、SONG-B*44:05:01:03、SONG-B*44:05:01:04、SONG-B*44:05:02、SONG- B*44:05:03, SONG-B*44:05:04, SONG-B*44:05:05, SONG-B*44:06, SONG-B*44: 07、SONGS-B*44:08、SONGS-B*44:09、SONGS-B*44:10、SONGS-B*44:11、SONGS-B*44:12、SONG-B*44:13, SONG-B*44:14, SONG-B*44:15:01:01, SONG-B*44:15:01:02, SONG-B *44:16、SONG-B*44:17、SONG-B*44:18:01:01、SONG-B*44:18:01:02、SONG-B*44:1 19 SONG-B*44:20, SONG-B*44:21, SONG-B*44:22, SONG-B*44:23:01:01, SONG-B*44:23:01:02, SONG-B*44:24, SONG-B*4 :25、SONG-B*44:26、SONG-B*44:27:01:01、SONG-B*44:27:01:02、SONG-B*44:27:01:03、SONG-B*44:27:02、SONG-B*4 4:27:03, SONG-B*44:27:04, SONG-B*44:28:01, SONG-B*44:28:02, SONG-B*44:29, SONG-B*44:30, SONG-B*44:31, SONG -B*44:32, SONG-B*44:33, SONG-B*44:34:01, SONG-B*44:34:02, SONG-B*44:35, SONG-B*44:36, SONG-B*44:37:01, HL A-B*44:37:02、SONGS-B*44:38、SONGS-B*44:39、SONGS-B*44:40、SONGS-B*44:41:01、SONGS-B*44:41:02、SONGS-B*44:42、H LA-B*44:43:01, SONG-B*44:43:02, SONG-B*44:44, SONG-B*44:45, SONG-B*44:46:01, SONG-B*44:46:02, SONG-B*44: 47、SONGS-B*44:48、SONGS-B*44:49、SONGS-B*44:50:01、SONGS-B*44:50:02、SONGS-B*44:50:03、SONGS-B*44:51、SONGS-B*44:51 :52、SONGS-B*44:53:01、SONGS-B*44:53:02、SONGS-B*44:54、SONGS-B*44:55、SONGS-B*44:56、SONGS-B*44:57、SONGS-B*44:5 74、SONGS-B*44:59:01、SONGS-B*44:59:02、SONGS-B*44:60、SONGS-B*44:6 SONG-B*44:62, SONG-B*44:63, SONG-B*44:64:01, SONG-B*44:64:02, SONG-B*44:65SONG-B*44:66, SONG-B*44:67, SONG-B*44:68, SONG-B*44:69:01, SONG-B*44:69:02, SONG-B *44:70、SONGS-B*44:71、SONGS-B*44:72、SONGS-B*44:73、SONGS-B*44:74、SONGS-B*44:75、SONGS-、 B*44:76, SONG-B*44:77, SONG-B*44:78, SONG-B*44:79:01, SONG-B*44:79:02 SONG-B*44:80, SONG-B*44:81, SONG-B*44:82, SONG-B*44:83, SONG-B*44:84:0 SONG-B*44:84:02, SONG-B*44:85:01, SONG-B*44:85:02, SONG-B*44:86, SONG-B *44:87、SONGS-B*44:88、SONGS-B*44:89、SONGS-B*44:90、SONGS-B*44:91、SONGS-B*4 :92、SONGS-B*44:93、SONGS-B*44:94、SONGS-B*44:95、SONGS-B*44:96、SONGS-B*44:9 7、SONGS-B*44:98、SONGS-B*44:99、SONGS-B*44:100、SONGS-B*44:101、SONGS-B*44:1 2、SONGS-B*44:103、SONGS-B*44:104、SONGS-B*44:105、SONGS-B*44:106、SONGS-B*4 :107、SONGS-B*44:108、SONGS-B*44:109、SONGS-B*44:110、SONGS-B*44:111、SONGS-B* 44:112, SONG-B*44:113, SONG-B*44:114, SONG-B*44:115, SONG-B*44:116, SONG -B*44:117, SONG-B*44:118, SONG-B*44:119, SONG-B*44:120, SONG-B*44:121, H LA-B*44:122、SONGS-B*44:123、SONGS-B*44:124、SONGS-B*44:125、SONGS-B*44:1 6:01, SONG-B*44:126:02, SONG-B*44:127, SONG-B*44:128:01, SONG-B*44:128: 02、SONGS-B*44:129、SONGS-B*44:130、SONGS-B*44:131、SONGS-B*44:132、SONGS-B*4 4:133, SONG-B*44:134, SONG-B*44:135, SONG-B*44:136, SONG-B*44:137, SONG-B *44:138, SONG-B*44:139, SONG-B*44:140, SONG-B*44:141, SONG-B*44:142, HL A-B*44:143, SONG-B*44:144, SONG-B*44:145, SONG-B*44:146, SONG-B*44:1SONG-B*44:148、SONG-B*44:1 95、SONGS-B*44:150、SONGS-B*44:151、SONGS-B*44:152、SONGS-B*44:153、SONGS-B*44:154、SONGS-B*44 :155、SONGS-B*44:156、SONGS-B*44:157、SONGS-B*44:158、SONGS-B*44:159、SONGS-B*44:160、SONGS-B*4 4:161, SONG-B*44:162, SONG-B*44:163:01, SONG-B*44:163:02, SONG-B*44:164, SONG-B*44:165 SONG-B*44:166, SONG-B*44:167, SONG-B*44:168, SONG-B*44:169, SONG-B*44:170, SONG-B*44:1 62. SONG-B*44:172, SONG-B*44:173, SONG-B*44:174, SONG-B*44:175, SONG-B*44:176, SONG-B*44:177, H LA-B*44:178, SONGS-B*44:179, SONGS-B*44:180, SONGS-B*44:181, SONGS-B*44:182, SONGS-B*44:183, SONGS-B *44:184, SONG-B*44:185, SONG-B*44:186:01, SONG-B*44:186:02, SONG-B*44:187, SONG-B*44:188, SONG -B*44:189, SONG-B*44:190, SONG-B*44:191, SONG-B*44:192:01, SONG-B*44:192:02, SONG-B*44:192:0 、SONGS-B*44:192:04、SONGS-B*44:193、SONGS-B*44:194:01、SONGS-B*44:194:02、SONGS-B*44:195、SONGS-B*4 4:196, SONG-B*44:197, SONG-B*44:198, SONG-B*44:199, SONG-B*44:200, SONG-B*44:201, SONG-B*44:2 、SONGS-B*44:203:01、SONGS-B*44:203:02、SONGS-B*44:204:01、SONGS-B*44:204:02、SONGS-B*44:205:01、H LA-B*44:205:02, SONGS-B*44:206, SONGS-B*44:207, SONGS-B*44:208, SONGS-B*44:209, SONGS-B*44:210:01SONG-B*44:210:02, SONG-B*44:211, SONG-B*44:212, SONG-B*44:213, SONG-B*4 4:214, SONG-B*44:215, SONG-B*44:216, SONG-B*44:217, SONG-B*44:218, SONG- B*44:219, SONG-B*44:220, SONG-B*44:221, SONG-B*44:222, SONG-B*44:223, H LA-B*44:224、SONGS-B*44:225、SONGS-B*44:226、SONGS-B*44:227、SONGS-B*44:2 、SONGS-B*44:229、SONGS-B*44:230、SONGS-B*44:231、SONGS-B*44:232、SONGS-B*44: 233、SONGS-B*44:234、SONGS-B*44:235、SONGS-B*44:236、SONGS-B*44:237、SONGS-B* 44:238, SONG-B*44:239, SONG-B*44:240, SONG-B*44:241, SONG-B*44:242, SONG -B*44:243, SONG-B*44:244, SONG-B*44:245, SONG-B*44:247, SONG-B*44:248, H LA-B*44:249, SONG-B*44:250, SONG-B*44:251, SONG-B*44:252, SONG-B*44:2 3, SONG-B*44:254, SONG-B*44:255, SONG-B*44:256, SONG-B*44:257, SONG-B*4 :258、SONGS-B*44:259、SONGS-B*44:260、SONGS-B*44:261、SONGS-B*44:262、SONGS-B *44:263、SONGS-B*44:264、SONGS-B*44:265、SONGS-B*44:266:01、SONGS-B*44:266: 02、SONGS-B*44:267、SONGS-B*44:268、SONGS-B*44:269、SONGS-B*44:270:01、SONGS- B*44:270:02, SONG-B*44:271, SONG-B*44:272, SONG-B*44:273, SONG-B*44:2 4. SONG-B*44:275, SONG-B*44:276, SONG-B*44:277, SONG-B*44:278, SONG-B*4 :279、SONGS-B*44:280、SONGS-B*44:281:01、SONGS-B*44:281:02、SONGS-B*44:282、SONG-B*44:283, SONG-B*44:284, SONG-B*44:285, SONG-B*44:286, SONG-B*44:287, SONG-B*44:2 、SONGS-B*44:289、SONGS-B*44:290、SONGS-B*44:291、SONGS-B*44:292、SONGS-B*44:293、SONGS-B*44:2 、SONGS-B*44:295、SONGS-B*44:296、SONGS-B*44:297、SONGS-B*44:298、SONGS-B*44:299、SONGS-B*44:3 、SONGS-B*44:301、SONGS-B*44:302、SONGS-B*44:303、SONGS-B*44:304、SONGS-B*44:305、SONGS-B*44:3 7, SONG-B*44:307, SONG-B*44:308, SONG-B*44:309, SONG-B*44:310, SONG-B*44:311, SONG- B*44:312, SONG-B*44:313, SONG-B*44:314, SONG-B*44:315, SONG-B*44:316, SONG-B*44:3 7, SONG-B*44:318, SONG-B*44:319, SONG-B*44:320, SONG-B*44:321, SONG-B*44:322, SONG-B *44:323, SONG-B*44:324, SONG-B*44:325, SONG-B*44:326, SONG-B*44:327, SONG-B*44:3 79. SONG-B*44:329, SONG-B*44:330, SONG-B*44:331, SONG-B*44:332, SONG-B*44:333, SONG-B*44:334, SONG-B*44:335, SONG- B*44:336, SONG-B*44:337, SONG-B*44:338, SONG-B*44:339, SONG-B*44:340, SONG-B*44:341, SONG-B*44:342, SONG-B*44:34 3, SONG-B*44:344, SONG-B*44:345, SONG-B*44:346, SONG-B*44:437, SONG-B*44:438, SONG-B*44:439, SONG-B*44:440, SONG-B *44:441, SONG-B*44:442, SONG-B*44:443, SONG-B*44:444, SONG-B*44:445, SONG-B*44:446, SONG-B*44:447, SONG-B*44:4SONGS-B*44:4 SONG-B*44:450, SONG-B*44:451, SONG-B*44:452, SONG-B*44:453, SONG-B*4 :454、SONGS-B*44:455、SONGS-B*44:456、SONGS-B*44:457、SONGS-B*44:458、SONGS- B*44:459, SONG-B*44:460, SONG-B*44:461, SONG-B*44:462, SONG-B*44:4 SONG-B*44:464:01:01, SONG-B*44:464:01:02, SONG-B*44:465, SONG-B*44:4 67、SONGS-B*44:468、SONGS-B*44:469、SONGS-B*44:470、SONGS-B*44:471、SONGS-B* 44:472, SONG-B*44:473, SONG-B*44:474, SONG-B*44:475, SONG-B*44:476, HL A-B*44:477, SONG-B*44:478, SONG-B*44:479, SONG-B*44:480, SONG-B*44:4 1, SONG-B*44:482, SONG-B*44:483, SONG-B*44:484, SONG-B*44:485, SONG-B*4 4:486, SONG-B*44:487, SONG-B*44:488, SONG-B*44:489, SONG-B*44:490, HL A-B*44:491, SONG-B*44:492, SONG-B*44:493, SONG-B*44:494, SONG-B*44:4 5、SONGS-B*44:496、SONGS-B*44:497、SONGS-B*44:498、SONGS-B*44:499、SONGS-B*4 4:500、SONGS-B*44:501、SONGS-B*44:502、SONGS-B*44:503、SONGS-B*44:504、SONGS -B*44:505、SONGS-B*44:506、SONGS-B*44:507、SONGS-B*44:508、SONGS-B*44:509 、SONGS-B*44:510、SONGS-B*44:511、SONGS-B*44:512、SONGS-B*44:513、SONGS-B*4 :514、SONG-B*44:515、SONG-B*44:516、SONG-B*44:517、SONG-B*44:518、SONG- B*44:519, SONG-B*44:520, SONG-B*44:521, SONG-B*44:522, SONG-B*44:523and HLA-B*44:524.

[0121] The numbers are HLA-B*51:01:01:01, HLA-B*51:01:01:02, HLA-B*51: 01:01:03, SONG-B*51:01:01:04, SONG-B*51:01:01:05, SONG-B*51:01:01:0 、SONG-B*51:01:01:07、SONG-B*51:01:01:08、SONG-B*51:01:01:09、SONG-B*5 1:01:01:10, SONG-B*51:01:01:11, SONG-B*51:01:01:12, SONG-B*51:01:01: 13、SONG-B*51:01:01:14、SONG-B*51:01:01:15、SONG-B*51:01:01:16、SONG- B*51:01:01:17, SONG-B*51:01:01:18, SONG-B*51:01:01:19, SONG-B*51:01: 01:20, SONG-B*51:01:01:21, SONG-B*51:01:01:22, SONG-B*51:01:01:23, SONG-B*51:01:01:23, HL A-B*51:01:01:24, SONG-B*51:01:01:25, SONG-B*51:01:01:26, SONG-B*51:0 1:01:27, SONG-B*51:01:01:28, SONG-B*51:01:01:29, SONG-B*51:01:01:3 、SONG-B*51:01:01:31、SONG-B*51:01:01:32、SONG-B*51:01:01:33、SONG-B*5 1:01:01:34, SONG-B*51:01:01:35, SONG-B*51:01:01:36, SONG-B*51:01:01: 37、SONGS-B*51:01:01:38、SONGS-B*51:01:01:39、SONGS-B*51:01:01:40、SONGS-B *51:01:01:41、SONGS-B*51:01:01:42、SONGS-B*51:01:01:43、SONGS-B*51:01: 01:44, SONG-B*51:01:01:45, SONG-B*51:01:01:46, SONG-B*51:01:01:47, SONG-B*51:01:01:47, HL A-B*51:01:01:48, SONG-B*51:01:01:49, SONG-B*51:01:01:50, SONG-B*51:0 1:01:51, SONG-B*51:01:01:52, SONG-B*51:01:01:53, SONG-B*51:01:01:54,SONG-B*51:01:01:55, SONG-B*51:01:01:56, SONG-B*51:01:01:57, SONG-B*5 :01:01:58, SONG-B*51:01:01:59, SONG-B*51:01:01:60, SONG-B*51:01:01:6 1, SONG-B*51:01:01:62, SONG-B*51:01:01:63, SONG-B*51:01:01:64, SONG-B* 51:01:01:65、SON-B*51:01:01:66、SON-B*51:01:01:67、SON-B*51:01:0 :68、SONG-B*51:01:01:69、SONG-B*51:01:01:70、SONG-B*51:01:01:71、SONG- B*51:01:01:72、HLA-B*51:01:01:73、HLA-B*51:01:01:74、HLA-B*51:01: 01:75, SONG-B*51:01:01:76, SONG-B*51:01:01:77, SONG-B*51:01:01:78, HL A-B*51:01:02:01, SONG-B*51:01:02:02, SONG-B*51:01:03, SONG-B*51:01:0 4、SONGS-B*51:01:05、SONGS-B*51:01:06、SONGS-B*51:01:07、SONGS-B*51:01:08、 SONG-B*51:01:09, SONG-B*51:01:10, SONG-B*51:01:11, SONG-B*51:01:12, SONG-B*51:01:12, SONG-B*51:01:1 A-B*51:01:13, SONG-B*51:01:14, SONG-B*51:01:15, SONG-B*51:01:16, SONG- B*51:01:17, SONG-B*51:01:18, SONG-B*51:01:19, SONG-B*51:01:20, SONG-B* 51:01:21、SONGS-B*51:01:22、SONGS-B*51:01:23、SONGS-B*51:01:24、SONGS-B*5 :01:25、SONG-B*51:01:26、SONG-B*51:01:27、SONG-B*51:01:28、SONG-B*51:0 1:29, SONG-B*51:01:30, SONG-B*51:01:31, SONG-B*51:01:32, SONG-B*51:01: 33, SONG-B*51:01:34, SONG-B*51:01:35, SONG-B*51:01:36, SONG-B*51:01:3SONG-B*51:01:38, SONG-B*51:01:39, SONG-B*51:01:40, SONG-B*51:01:41, SONG-B*51:01:41, SONG-B*51:01:4 A-B*51:01:42, SONG-B*51:01:43, SONG-B*51:01:44, SONG-B*51:01:45, SONG- B*51:01:46, SONG-B*51:01:47, SONG-B*51:01:48, SONG-B*51:01:49, SONG-B* 51:01:50、SONGS-B*51:01:51、SONGS-B*51:01:52、SONGS-B*51:01:53、SONGS-B*51: 01:54、SONGS-B*51:01:55、SONGS-B*51:01:56、SONGS-B*51:01:57、SONGS-B*51:01 :58、SONGS-B*51:01:59、SONGS-B*51:01:60、SONGS-B*51:01:61、SONGS-B*51:01:6 2、SONGS-B*51:01:63、SONGS-B*51:01:64、SONGS-B*51:01:65、SONGS-B*51:01:66、 SONG-B*51:01:67, SONG-B*51:01:68, SONG-B*51:01:69, SONG-B*51:01:70, SONG -B*51:01:71, SONG-B*51:01:72, SONG-B*51:01:73, SONG-B*51:01:74, SONG-B *51:01:75、SONGS-B*51:01:76、SONGS-B*51:01:77、SONGS-B*51:01:78、SONGS-B*5 1:01:79, SONG-B*51:01:80, SONG-B*51:01:81, SONG-B*51:01:82, SONG-B*51: 01:83、SONG-B*51:01:84、SONG-B*51:01:85、SONG-B*51:01:86、SONG-B*51:01: 87、SONGS-B*51:01:88、SONGS-B*51:01:89、SONGS-B*51:01:90、SONGS-B*51:01:9 、SONG-B*51:01:92、SONG-B*51:02:01:01、SONG-B*51:02:01:02、SONG-B*51:0 2:01:03、SONG-B*51:02:02、SONG-B*51:02:03、SONG-B*51:02:04、SONG-B*51: 02:05、SONGS-B*51:02:06、SONGS-B*51:02:07、SONGS-B*51:03、SONGS-B*51:04:01、SONG-B*51:04:02, SONG-B*51:05, SONG-B*51:06:01:01, SONG-B*51:06:01:02, SONG-B*51:06:01:03, SONG-B*51:06:02 A-B*51:06:03, SONG-B*51:06:04, SONG-B*51:07:01, SONG-B*51:07:02, SONG-B*51:08:01:01, SONG-B*51:08:01:02, SONG- B*51:08:01:03, SONG-B*51:08:01:04, SONG-B*51:08:02, SONG-B*51:08:03, SONG-B*51:08:04, SONG-B* 51:09:01、SONGS-B*51:09:02、SONGS-B*51:09:03、SONGS-B*51:10、SONGS-B*51:11、SONGS-B*51:12、SONGS-B*51:13:01、SONGS-B*51 :13:02、SONGS-B*51:14、SONGS-B*51:15、SONGS-B*51:16、SONGS-B*51:17、SONGS-B*51:18、SONGS-B*51:19、SONGS-B*51:20、SONGS-B*5 1:21, SONG-B*51:22, SONG-B*51:23, SONG-B*51:24:01, SONG-B*51:24:02, SONG-B*51:24:03, SONG-B*51:24:04, SONG-B*51: 24:05、SONGS-B*51:26、SONGS-B*51:27、SONGS-B*51:28、SONGS-B*51:29、SONGS-B*51:30、SONGS-B*51:31、SONGS-B*51:32、SONGS-B*51:32 :33、SONGS-B*51:34、SONGS-B*51:35、SONGS-B*51:36、SONGS-B*51:37、SONGS-B*51:38、SONGS-B*51:39、SONGS-B*51:40、SONGS-B*51:41 74、SONGS-B*51:42、SONGS-B*51:43、SONGS-B*51:4 64, SONG-B*51:45, SONG-B*51:46, SONG-B*51:48, SONG-B*51:49, SONG-B*51:50, SONG-B*51:51, SONG-B*51: 52, SONG-B*51:53, SONG-B*51:54, SONG-B*51:55, SONG-B*51:56:01, SONG-B*51:56:02, SONG-B*51:56:03SONG-B*51:57, SONG-B*51:58, SONG-B*51:59, SONG-B*51:60, SONG-B*51:61:01, SONG-B*5 :61:02、SONGS-B*51:62、SONGS-B*51:63:01、SONGS-B*51:63:02、SONGS-B*51:64、SONGS-B*51:65 、SONGS-B*51:66、SONGS-B*51:67、SONGS-B*51:68、SONGS-B*51:69、SONGS-B*51:70、SONGS-B*51:7 、SONGS-B*51:72、SONGS-B*51:73、SONGS-B*51:74、SONGS-B*51:75、SONGS-B*51:76、SONGS-B*51:7 、SONGS-B*51:78:01、SONGS-B*51:78:02、SONGS-B*51:79、SONGS-B*51:80、SONGS-B*51:81、SONGS-B*51:81、SONGS-B*51:8 *51:82、SONGS-B*51:83、SONGS-B*51:84、SONGS-B*51:85、SONGS-B*51:86、SONGS-B*51:87、SONGS-B *51:88、SONGS-B*51:89、SONGS-B*51:90、SONGS-B*51:91、SONGS-B*51:92:01、SONGS-B*51:92:02 、SONGS-B*51:93、SONGS-B*51:94、SONGS-B*51:95、SONGS-B*51:96、SONGS-B*51:97、SONGS-B*51:9 18 SONG-B*51:99, SONG-B*51:100, SONG-B*51:101, SONG-B*51:102, SONG-B*51:103, SONG-B*51:104:01, SONG-B*51:104:02 SONG-B*51:105, SONG-B*51:106:01, SONG-B*51:106:02, SONG-B*51:107, SONG-B*51:108, SONG-B*51:109, SONG-B*51:110 84 SONG-B*51:111, SONG-B*51:112, SONG-B*51:113, SONG-B*51:114, SONG -B*51:115, SONG-B*51:116, SONG-B*51:117, SONG-B*51:118, SONG-B* 51:119, SONG-B*51:120, SONG-B*51:121, SONG-B*51:122, SONG-B*51: 123. SONG-B*51:124, SONG-B*51:125, SONG-B*51:126, SONG-B*51:127.SONG-B*51:128, SONG-B*51:129, SONG-B*51:130, SONG-B*51:131, SONG-B*51:132, SONG-B*51:1 、SONGS-B*51:134、SONGS-B*51:135、SONGS-B*51:136、SONGS-B*51:137、SONGS-B*51:138、SONGS-B*51:13、 9, SONG-B*51:140, SONG-B*51:141, SONG-B*51:142, SONG-B*51:143, SONG-B*51:144, SONG -B*51:145, SONG-B*51:146, SONG-B*51:147, SONG-B*51:148, SONG-B*51:149, SONG-B*51: 150. SONG-B*51:151, SONG-B*51:152, SONG-B*51:153, SONG-B*51:154, SONG-B*51:155, H LA-B*51:156, SONG-B*51:157, SONG-B*51:158:01, SONG-B*51:158:02, SONG-B*51:159, H LA-B*51:160, SONG-B*51:161, SONG-B*51:162:01, SONG-B*51:162:02, SONG-B*51:163 SONG-B*51:164, SONG-B*51:165, SONG-B*51:166, SONG-B*51:167, SONG-B*51:168, SONG-B* 51:169, SONG-B*51:170, SONG-B*51:171, SONG-B*51:172, SONG-B*51:173, SONG-B*51:1 :01、SONGS-B*51:174:02、SONGS-B*51:175、SONGS-B*51:176、SONGS-B*51:177、SONGS-B*51:1 57. SONG-B*51:179, SONG-B*51:180, SONG-B*51:181, SONG-B*51:182, SONG-B*51:183, SONG-B*51:1 31. SONG-B*51:185, SONG-B*51:186, SONG-B*51:187, SONG-B*51:188, SONG-B*51:189, SONG-B*51:190, SONG-B *51:191, SONG-B*51:192, SONG-B*51:193, SONG-B*51:194, SONG-B*51:195, SONG-B*51:196, SONG-B*51:197, H LA-B*51:198, SONGS-B*51:199, SONGS-B*51:200, SONGS-B*51:201, SONGS-B*51:202, SONGS-B*51:203, SONGS-B*51:2 04:01, SONGS-B*51:204:02, SONGS-B*51:205, SONGS-B*51:206, SONGS-B*51:207, SONGS-B*51:208, SONGS-B*51:209SONG-B*51:210, SONG-B*51:211, SONG-B*51:212, SONG-B*51:213, SONG-B*51:214, SONG -B*51:215, SONG-B*51:216, SONG-B*51:217, SONG-B*51:218, SONG-B*51:219, SONG-B*5 1:220, SONG-B*51:221, SONG-B*51:222, SONG-B*51:223, SONG-B*51:224, SONG-B*51:2 25. SONG-B*51:226, SONG-B*51:227, SONG-B*51:228, SONG-B*51:229, SONG-B*51:230:0 1:01、SONGS-B*51:230:01:02、SONGS-B*51:230:01:03、SONGS-B*51:231、SONGS-B*51:2 :01、SONGS-B*51:232:02、SONGS-B*51:233、SONGS-B*51:234、SONGS-B*51:235、SONGS-B*51:2 36、SONGS-B*51:237:01、SONGS-B*51:237:02、SONGS-B*51:238、SONGS-B*51:239、SONGS-B*51 :240、SONGS-B*51:241、SONGS-B*51:242、SONGS-B*51:243、SONGS-B*51:244、SONGS-B*51:2 64 SONG-B*51:246, SONG-B*51:247, SONG-B*51:248, SONG-B*51:249, SONG -B*51:250, SONG-B*51:251, SONG-B*51:252, SONG-B*51:253, SONG-B* 51:254, SONG-B*51:255, SONG-B*51:256, SONG-B*51:257, SONG-B*51: 258. SONG-B*51:259, SONG-B*51:260, SONG-B*51:261, SONG-B*51:262. SONG-B*51:263, SONG-B*51:264, SONG-B*51:265, SONG-B*51:266, SONG -B*51:267, SONG-B*51:268, SONG-B*51:269, SONG-B*51:270, SONG-B* 51:271, SONG-B*51:272, SONG-B*51:273, SONG-B*51:274, SONG-B*51: 275, SONG-B*51:276, SONG-B*51:277, SONG-B*51:278, SONG-B*51:2SONG-B*51:280, SONG-B*51:281, SONG-B*51:282, SONG-B*51:283, SONG-B*51:284, SONG-B*51:285, SONG-B*51:286, SONG-B*51:287, H LA-B*51:288, SONG-B*51:289, SONG-B*51:290, SONG-B*51:291, SONG-B*51:292, SONG-B*51:293, SONG-B*51:294, SONG-B*51:295, SONG -B*51:296, HLA-B*51:297, HLA-B*51:298, HLA-B*51:299, HLA-B*51:300, HLA-B*51:301, HLA-B*51:302, HLA-B*51:303, HLA- B*51:304, HLA-B*51:305, HLA-B*51:306, HLA-B*51:307, HLA-B*51:308, HLA-B*51:309, HLA-B*51:310, HLA-B*51:311, HLA-B* 51:312, SONG-B*51:313, SONG-B*51:314, SONG-B*51:315, SONG-B*51:316, SONG-B*51:317, SONG-B*51:318, SONG-B*51:319, SONG-B*5 1:320, SONG-B*51:321, SONG-B*51:322, SONG-B*51:323, SONG-B*51:324, SONG-B*51:325, SONG-B*51:326, SONG-B*51:327, SONG-B*51: 328. SONG-B*51:329, SONG-B*51:330, SONG-B*51:331, SONG-B*51:332, SONG-B*51:333, SONG-B*51:334, SONG-B*51:335, SONG-B*51:3 36. SONG-B*51:337, SONG-B*51:338, SONG-B*51:339, SONG-B*51:340, SONG-B*51:341, SONG-B*51:342, SONG-B*51:343, SONG-B*51:344 31. The HLA-B*51:345 range is also available.

[0122] The numbers are HLA-B*58:01:01:01, HLA-B*58:01:01:02, HLA-B*58: 01:01:03, SONG-B*58:01:01:04, SONG-B*58:01:01:05, SONG-B*58:01:01:0 、SONG-B*58:01:01:07、SONG-B*58:01:01:08、SONG-B*58:01:01:09、SONG-B*5 8:01:01:10, SONG-B*58:01:01:11, SONG-B*58:01:01:12, SONG-B*58:01:01: 13、SONGS-B*58:01:02、SONGS-B*58:01:03、SONGS-B*58:01:04、SONGS-B*58:01:0 、SONG-B*58:01:06、SONG-B*58:01:07、SONG-B*58:01:08、SONG-B*58:01:09、H LA-B*58:01:10、SONGS-B*58:01:11、SONGS-B*58:01:12、SONGS-B*58:01:13、SONGS -B*58:01:14, SONG-B*58:01:15, SONG-B*58:01:16, SONG-B*58:01:17, SONG-B *58:01:18、SONGS-B*58:01:19、SONGS-B*58:01:20、SONGS-B*58:01:21、SONGS-B*5 8:01:22、SONGS-B*58:01:23、SONGS-B*58:01:24、SONGS-B*58:01:25、SONGS-B*58: 01:26, SONG-B*58:01:27, SONG-B*58:01:28, SONG-B*58:01:29, SONG-B*58:0 :30、SONGS-B*58:01:31、SONGS-B*58:01:32、SONGS-B*58:01:33、SONGS-B*58:01:3 4, SONG-B*58:01:35, SONG-B*58:01:36, SONG-B*58:01:37, SONG-B*58:01:38, SONG-B*58:01:39, SONG-B*58:02:01:01, SONG-B*58:02:01:02, SONG-B*58:0 :01:03、SONG-B*58:02:02、SONG-B*58:04、SONG-B*58:05、SONG-B*58:06、SONG- B*58:07, SONG-B*58:08:01, SONG-B*58:08:02, SONG-B*58:09, SONG-B*58:10SONGS-B*58:11, SONGS-B*58:12, SONGS-B*58:13, SONGS-B*58:14, SONGS-B*58:15, SONGS-B*58:16:01, SONGS-B*58:16:02, SONGS A-B*58:17, SONG-B*58:18, SONG-B*58:19, SONG-B*58:20, SONG-B*58:21, SONG-B*58:22, SONG-B*58:23, SONG-B*58:24 、SONGS-B*58:25、SONGS-B*58:26、SONGS-B*58:27、SONGS-B*58:28:01、SONGS-B*58:28:02、SONGS-B*58:29、SONGS-B*58:31、SONGS-B*58:31、SONGS A-B*58:32, SONG-B*58:33, SONG-B*58:34, SONG-B*58:35, SONG-B*58:36, SONG-B*58:37, SONG-B*58:38, SONG-B*58:39 45, SONG-B*58:40, SONG-B*58:41, SONG-B*58:42, SONG-B*58:43, SONG-B*58:44, SONG-B*58:45:01 SONG-B*58:45:02, SONG-B*58:46, SONG-B*58:47, SONG-B*58:48, SONG-B*58:49, SONG-B*58:50, SONG -B*58:51, SONG-B*58:52, SONG-B*58:53, SONG-B*58:54, SONG-B*58:55, SONG-B*58:56, SONG-B*58: 57. SONG-B*58:58, SONG-B*58:59:01, SONG-B*58:59:02, SONG-B*58:60, SONG-B*58:61, SONG-B*58:6 2, SONG-B*58:63, SONG-B*58:64, SONG-B*58:65, SONG-B*58:66, SONG-B*58:67, SONG-B*58:68, SONG- B*58:69, SONG-B*58:70, SONG-B*58:71, SONG-B*58:72, SONG-B*58:73, SONG-B*58:74, SONG-B*58:75 、SONGS-B*58:76、SONGS-B*58:77、SONGS-B*58:78、SONGS-B*58:79、SONGS-B*58:80、SONGS-B*58:81、SONGS-B *58:82, SONG-B*58:83, SONG-B*58:84, SONG-B*58:85, SONG-B*58:86, SONG-B*58:87, SONG-B*58:8SONGS-B*58:89, SONGS-B*58:90, SONGS-B*58:91, SONGS-B*58:92, SONGS-B*58:93, SONGS-B*58:94, HL A-B*58:95, SONG-B*58:96, SONG-B*58:97, SONG-B*58:98, SONG-B*58:99, SONG-B*58:100, SONG -B*58:101, SONG-B*58:102, SONG-B*58:103, SONG-B*58:104, SONG-B*58:105, SONG-B*58:1 、SONGS-B*58:107、SONGS-B*58:108、SONGS-B*58:109、SONGS-B*58:110、SONGS-B*58:111、SONGS-B*58: 112. SONG-B*58:113, SONG-B*58:114, SONG-B*58:115, SONG-B*58:116, SONG-B*58:117, SONG-B *58:118, SONG-B*58:119, SONG-B*58:120, SONG-B*58:121, SONG-B*58:122, SONG-B*58:123, HL A-B*58:124, SONG-B*58:125, SONG-B*58:126, SONG-B*58:127, SONG-B*58:128, SONG-B*58:1 9. HLA-B*58:130, HLA-B*58:131, HLA-B*58:132, HLA-B*58:1

[0123] In some embodiments, the HLA molecule of the present disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-B allele. In some embodiments, the modified HLA binding pocket is the F pocket of the HLA molecule. In some embodiments, the HLA molecule of the present disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-B allele, and the modified HLA binding pocket comprises an amino acid substitution at a position corresponding to amino acid residue 79, amino acid residue 80, amino acid residue 81, amino acid residue 82, or amino acid residue 83 of SEQ ID NO: 1. In some embodiments, the HLA molecule of the present disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-B allele, and the modified HLA binding pocket comprises an amino acid substitution at amino acid residue 81, which corresponds to the amino acid sequence shown in SEQ ID NO: 1.

[0124] In some embodiments, the original amino acid replaced is alanine. In some embodiments, the new amino acid replacing the original amino acid is selected from the group consisting of leucine, valine, isoleucine, methionine, phenylalanine, tyrosine, and tryptophan. In some embodiments, the new amino acid replacing the original amino acid is leucine. In some embodiments, the amino acid substitution is A81L, which corresponds to the amino acid sequence set forth in SEQ ID NO:1.

[0125] II.A.3. HLA-C alleles In some embodiments, the HLA class I molecule is an HLA-C allele. Any HLA-C allele can be used in the disclosed methods and compositions. In some aspects, the HLA-C allele is selected from the HLA-C*05:01 allele, the HLA-C*05:03 allele, the HLA-C*05:04 allele, the HLA-C*05:05 allele, and the HLA-C*05:06 allele. In certain aspects, the HLA-C allele is the HLA-C*05:01 allele. In certain aspects, the HLA-C allele is the HLA-C*05:03 allele. In certain aspects, the HLA-C allele is the HLA-C*05:04 allele. In certain aspects, the HLA-C allele is the HLA-C*05:05 allele. In certain aspects, the HLA-C allele is the HLA-C*05:06 allele.

[0126] In some embodiments, the HLA molecule of the present disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-C allele. In some embodiments, the modified HLA binding pocket is the F pocket of the HLA molecule. In some embodiments, the HLA molecule of the present disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-C allele, and the modified HLA binding pocket comprises an amino acid substitution at a position corresponding to amino acid residue 79, amino acid residue 80, amino acid residue 81, amino acid residue 82, or amino acid residue 83 of SEQ ID NO: 1. In some embodiments, the HLA molecule of the present disclosure comprises a modified HLA binding pocket, wherein the HLA molecule is an HLA-C allele, and the modified HLA binding pocket comprises an amino acid substitution at amino acid residue 81, which corresponds to the amino acid sequence shown in SEQ ID NO: 1.

[0127] In some embodiments, the original amino acid replaced is alanine. In some embodiments, the new amino acid replacing the original amino acid is selected from the group consisting of leucine, valine, isoleucine, methionine, phenylalanine, tyrosine, and tryptophan. In some embodiments, the new amino acid replacing the original amino acid is leucine. In some embodiments, the amino acid substitution is A81L, which corresponds to the amino acid sequence set forth in SEQ ID NO:1.

[0128] II.A.3. HLA Class I Binding Pocket Various aspects of the present disclosure are directed to HLA molecules that contain modified HLA binding pockets. HLA class I molecules contain six binding pockets: A, B, C, D, E, and F. The data presented herein demonstrate that modifying the amino acid sequence of one or more residues in one or more of the six binding pockets can enhance the binding affinity of HLA molecules to target antigens. In doing so, the present disclosure provides a novel method for increasing the antigen binding affinity of HLA molecules that can be applied to a large number of different HLA alleles.

[0129] In some embodiments, the HLA molecule comprises a modified F pocket. In some embodiments, the HLA molecule comprises a modified A pocket. In some embodiments, the HLA molecule comprises a modified B pocket. In some embodiments, the HLA molecule comprises a modified C pocket. In some embodiments, the HLA molecule comprises a modified D pocket. In some embodiments, the HLA molecule comprises a modified E pocket.

[0130] In some embodiments, more than one HLA binding pocket of an HLA molecule is modified. In some embodiments, the HLA molecule comprises (i) a modified F pocket and (ii) a modified A pocket. In some embodiments, the HLA molecule comprises (i) a modified F pocket and (ii) a modified B pocket. In some embodiments, the HLA molecule comprises (i) a modified F pocket and (ii) a modified C pocket. In some embodiments, the HLA molecule comprises (i) a modified F pocket and (ii) a modified D pocket. In some embodiments, the HLA molecule comprises (i) a modified F pocket and (ii) a modified E pocket.

[0131] In some embodiments, the HLA molecule comprises (i) a modified F pocket, (ii) a modified B pocket, and (iii) a modified A pocket. In some embodiments, the HLA molecule comprises (i) a modified F pocket, (ii) a modified B pocket, and (iii) a modified C pocket. In some embodiments, the HLA molecule comprises (i) a modified F pocket, (ii) a modified B pocket, and (iii) a modified D pocket. In some embodiments, the HLA molecule comprises (i) a modified F pocket, (ii) a modified B pocket, and (iii) a modified E pocket.

[0132] In some embodiments, the HLA molecule comprises (i) a modified F pocket, (ii) a modified A pocket, and (iii) a modified C pocket. In some embodiments, the HLA molecule comprises (i) a modified F pocket, (ii) a modified A pocket, and (iii) a modified D pocket. In some embodiments, the HLA molecule comprises (i) a modified F pocket, (ii) a modified A pocket, and (iii) a modified E pocket.

[0133] In some embodiments, the HLA molecule comprises (i) a modified F pocket, (ii) a modified C pocket, and (iii) a modified D pocket. In some embodiments, the HLA molecule comprises (i) a modified F pocket, (ii) a modified C pocket, and (iii) a modified E pocket.

[0134] In some embodiments, the HLA molecule comprises (i) a modified F pocket, (ii) a modified D pocket, and (iii) a modified E pocket.

[0135] II.B. Cancer Treatment Methods Certain aspects of the present disclosure are directed to a method of treating cancer in a subject in need thereof comprising administering to the subject a nucleic acid molecule disclosed herein, a recombinant TCR disclosed herein, a bispecific TCR disclosed herein, an epitope disclosed herein, or an HLA class I molecule disclosed herein, or a vector or cell comprising any of the above.

[0136] In some embodiments, the cancer is melanoma, bone cancer, kidney cancer, prostate cancer, breast cancer, colon cancer, lung cancer, cutaneous or intraocular malignant melanoma, pancreatic cancer, skin cancer, head and neck cancer, uterine cancer, ovarian cancer, rectal cancer, anal cancer, gastric cancer, testicular cancer, uterine cancer, fallopian tube cancer, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, Hodgkin's disease, non-Hodgkin's lymphoma (NHL), primary mediastinal large B-cell lymphoma (PMBC), diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), transformed follicular lymphoma, splenic marginal zone lymphoma (SMZL), esophageal cancer, small intestine cancer, endocrine system cancer, thyroid cancer, parathyroid cancer, adrenal gland cancer, soft tissue carcinoma, In some embodiments, the cancer is selected from the group consisting of tumors, urethral cancer, penile cancer, chronic or acute leukemia, acute myeloid leukemia (AML), chronic myeloid leukemia, acute lymphoblastic leukemia (ALL) (including non-T cell ALL), chronic lymphocytic leukemia (CLL), solid tumors of childhood, lymphocytic lymphoma, bladder cancer, kidney or ureter cancer, renal pelvis cancer, neoplasms of the central nervous system (CNS), primary CNS lymphoma, tumor angiogenesis, spinal axis tumor, brain stem glioma, pituitary adenoma, Kaposi's sarcoma, epidermoid carcinoma, squamous cell carcinoma, T cell lymphoma, environmentally induced cancers including those due to asbestos, other B cell malignancies, and combinations of the above cancers. In some embodiments, the cancer is melanoma.

[0137] In some embodiments, the cancer is recurrent. In some embodiments, the cancer is refractory. In some embodiments, the cancer is progressive. In some embodiments, the cancer is metastatic.

[0138] In some embodiments, the methods disclosed herein treat cancer in a subject. In some embodiments, the methods disclosed herein reduce the severity of one or more symptoms of cancer. In some embodiments, the methods disclosed herein reduce the size or number of tumors resulting from cancer. In some embodiments, the methods disclosed herein increase the overall survival of a subject compared to a subject not provided with the methods disclosed herein. In some embodiments, the methods disclosed herein increase the progression-free survival of a subject compared to a subject not provided with the methods disclosed herein. In some embodiments, the methods disclosed herein result in a partial response in a subject. In some embodiments, the methods disclosed herein result in a complete response in a subject.

[0139] In some aspects, the methods disclosed herein include treating cancer in a subject in need thereof, comprising administering to the subject a cell described herein, wherein the cell comprises a nucleic acid molecule disclosed herein, a vector disclosed herein, a recombinant TCR disclosed herein, and / or a bispecific antibody disclosed herein. In some aspects, the cell is a T cell. In some embodiments, the cell is a cell that has been modified to express CD3.

[0140] In some embodiments, the cells, e.g., T cells, are obtained from the subject. In some embodiments, the cells, e.g., T cells, are obtained from a donor other than the subject.

[0141] In some embodiments, the subject is preconditioned prior to administering the cells. Preconditioning can include any substance that promotes T cell function and / or survival. In some embodiments, preconditioning includes administering to the subject chemotherapy, cytokines, proteins, small molecules, or any combination thereof. In some embodiments, preconditioning includes administering an interleukin. In some embodiments, preconditioning includes administering IL-2, IL-4, IL-7, IL-9, IL-15, IL-21, or any combination thereof. In some embodiments, preconditioning includes administering cyclophosphamide, fludarabine, or both. In some embodiments, preconditioning includes administering vitamin C, AKT inhibitors, ATRA (vesanoid, tretinoin), rapamycin, or any combination thereof.

[0142] In some embodiments, the compositions disclosed herein (e.g., antigen-HLA complexes, cells expressing modified HLA, or vaccines disclosed herein) are administered to subjects in combination with immunotherapy. Without being bound to a particular mechanism, according to the present disclosure, modification of HLA binding pocket increases the affinity of HLA to antigen, thereby increasing the surface display of antigen on cells expressing modified HLA. This increased surface display enhances the immune response to antigen. Thus, the modified HLA molecules disclosed herein can act to enhance the immune response to immunotherapy.

[0143] Any immunotherapy can benefit from combined administration with the modified HLA molecules disclosed herein. As used herein, "combined administration" refers to the administration of at least two therapies within a set period of time. In some embodiments, at least two therapies are administered simultaneously (e.g., at the same time). In some embodiments, at least two therapies are administered consecutively (e.g., one after the other). In some embodiments, at least two therapies are administered on the same day. In some embodiments, at least two therapies are administered on consecutive days. In some embodiments, at least two therapies are administered during the same administration cycle (e.g., according to the prescribed administration schedule of the immunotherapy).

[0144] In some embodiments, the immune cell therapy comprises administering a plurality of immune cells to the subject (i.e., immune cell therapy or cell-based therapy). Immune cell therapy has emerged as a promising means of treating various diseases, including cancer. In some embodiments, the immune cell therapy comprises administering a plurality of T cells, NK cells, tumor infiltrating lymphocytes (TILs), or any combination thereof. In some embodiments, the immune cells are modified. In some embodiments, the immune cells are modified to express a chimeric antigen receptor (CAR), a heterologous T cell receptor (TCR), an engineered TCR, or any combination thereof. In some embodiments, the immune cell therapy comprises administering an engineered T cell, where the engineered T cell comprises a nucleic acid molecule encoding a CAR, a heterologous TCR, an engineered TCR, or any combination thereof. In some embodiments, the immune cell therapy comprises administering an engineered NK cell, where the engineered NK cell comprises a nucleic acid molecule encoding a CAR, a heterologous TCR, an engineered TCR, or any combination thereof.

[0145] In some embodiments, the immunotherapy comprises an antagonist (inhibitor or blocker) of a protein that inhibits T cell activation (e.g., an immune checkpoint inhibitor), such as CTLA-4, PD-1, PD-L1, PD-L2, GITR, LAG-3, Galectin 9, CEACAM-1, BTLA, CD69, Galectin-1, TIGIT, CD113, GPR56, VISTA, B7-H3, B7-H4, 2B4, CD48, GARP, PD1H, LAIR1, mesothelin, CD27, CD96, TIM-1, TIM-3, and TIM-4. In some embodiments, the immunotherapy comprises a PD-1 inhibitor, a PD-L1 inhibitor, a CTLA-4 inhibitor, a LAG3 inhibitor, a TIM3 inhibitor, or any combination thereof.

[0146] In some embodiments, the immunotherapy comprises a cancer vaccine.

[0147] In some embodiments, the immunotherapy comprises an antibody or antigen-binding portion thereof that specifically binds to PD-1 or PD-L1. In some embodiments, the immunotherapy comprises an anti-PD-1 antibody selected from nivolumab (OPDIVO®) and pembrolizumab (KEYTRUDA®). In some embodiments, the immunotherapy comprises an anti-PD-1 antibody selected from YERVOY® (ipilimumab) or tremelimumab (against CTLA-4), galiximab (against B7.1), BMS-936558 (against PD-1), MK-3475 (against PD-1), atezolizumab (TECENTRIQ®), AMP224 (against B7DC), BMS-936559 (against B7-H1). (against B7-H1), MPDL3280A (against B7-H1), MEDI-570 (against ICOS), AMG557 (against B7H2), MGA271 (against B7H3), IMP321 (against LAG-3), BMS-663513 (against CD137), PF-05082566 (against CD137), CDX-1127 (against CD27), anti-OX40 (Providence Health Services), huMAbOX40L (against OX40L), atacicept (against TACI), CP-870893 (against CD40), lucatumumab (against CD40), dacetuzumab (against CD40), muromonab-CD3 (against CD3), anti-GITR antibody MK4166, TRX518, Medi1873, INBRX-110, LK2-145, GWN-323, GITRL-Fc, and any combination thereof.

[0148] In some embodiments, the immunotherapy includes agents that target (or specifically bind to) members of the B7 family of membrane-bound ligands, including B7-1, B7-2, B7-H1 (PD-L1), B7-DC (PD-L2), B7-H2 (ICOS-L), B7-H3, B7-H4, B7-H5 (VISTA), and B7-H6, or costimulatory or co-inhibitory receptors or ligands that specifically bind to members of the B7 family. In some embodiments, the immunotherapy includes agonists of proteins that stimulate T cell activation, such as B7-1, B7-2, CD28, 4-1BB (CD137), 4-1BBL, GITR, ICOS, ICOS-L, OX40, OX40L, CD70, CD27, CD40, DR3, and CD28H. In some embodiments, the immunotherapy includes antagonists of inhibitory receptors on NK cells or agonists of activating receptors on NK cells, such as antagonists of KIR (e.g., lirilumab).

[0149] In some embodiments, a composition disclosed herein (e.g., an antigen-HLA complex, a cell expressing a modified HLA, or a vaccine disclosed herein) is administered to a subject in combination with another anti-cancer agent, e.g., chemotherapy, a cytokine, radiation therapy, surgery, or any combination thereof.In some embodiments, the additional anti-cancer agent is radiation therapy and / or chemotherapy using, for example, camptothecin (CPT-11), 5-fluorouracil (5-FU), cisplatin, doxorubicin, irinotecan, paclitaxel, gemcitabine, cisplatin, paclitaxel, carboplatin-paclitaxel (taxol), doxorubicin, or camptothecin + apo21 / TRAIL (6-fold combo), one or more proteasome inhibitors (e.g., bortezomib or MG132), one or more Bcl-2 inhibitors (e.g., BH3 I-2' (bcl-xl inhibitors), indoleamine dioxygenase-1 inhibitors (e.g., INCB24360, indoximod, NLG-919, or F001287), AT-101 (R-(-)-gossypol derivatives), ABT-263 (small molecule), GX-15-070 (obatoclax), or MCL-1 (myeloid leukemia cell differentiation protein-1) antagonists), iAP (inhibitor of apoptosis proteins) antagonists (e.g., smac7, smac4, small molecule smac mimetics, synthetic smac peptides (Fulda et al., Nat. Med. 2002;8:808-15), ISIS 23722 (LY2181308), or AEG-35156 (GEM-640)), HDAC (histone deacetylase) inhibitors, anti-CD20 antibodies (e.g., rituximab), angiogenesis inhibitors (e.g., bevacizumab), antiangiogenic agents targeting VEGF and VEGFR (e.g., Avastin), synthetic triterpenoids (Hyer et al., Cancer Research, 2005;65:4799-808), c-FLIP (cellular FLICE inhibitory protein) modulators (e.g., natural and synthetic ligands of PPARγ (peroxisome proliferator activated receptor gamma), 5809354 or 5569100), kinase inhibitors (e.g., sorafenib), trastuzumab, cetuximab, temsirolimus, mTOR inhibitors such as rapamycin and temsirolimus, bortezomib, JAK2 inhibitors, HSP90 inhibitors, PI3K-AKT inhibitors, lenalidomide, GSK3P inhibitors, IAP inhibitors and / or genotoxic drugs.

[0150] In some embodiments, the anti-cancer agent comprises one or more anti-proliferative cytotoxic agents. In some embodiments, the anti-cancer agent comprises an alkylating agent (including but not limited to nitrogen mustards, ethylenimine derivatives, alkyl sulfonates, nitrosoureas, and triazenes). In some embodiments, the anti-cancer agent comprises uracil mustard, chlormethine, cyclophosphamide (CYTOXAN®), phosphamide, melphalan, chlorambucil, pipobroman, triethylenemelamine, triethylenethiophosphoramine, busulfan, carmustine, lomustine, streptozocin, dacarbazine, temozolomide, and any combination thereof.

[0151] In some embodiments, the anti-cancer agent comprises an antimetabolite (including but not limited to folate antagonists, pyrimidine analogs, purine analogs, and adenosine deaminase inhibitors). In some embodiments, the anti-cancer agent comprises methotrexate, 5-fluorouracil, floxuridine, cytarabine, 6-mercaptopurine, 6-thioguanine, fludarabine phosphate, pentostatin, gemcitabine, and any combination thereof.

[0152] In some embodiments, the anticancer agent is a taxane, paclitaxel (e.g., TAXOL™), docetaxel, discodermolide (DDM), dictyostatin (DCT), peloruside A, epothilone, epothilone A, epothilone B, epothilone C, epothilone D, epothilone E, epothilone F, furanoepothilone D, desoxyepothilone Bl,

[17] -dehydrodesoxyepothilone B,

[18] dehydro Desoxyepothilone B, C12,13-cyclopropyl-epothilone A, C6-C8 bridged epothilone A, trans-9,10-dehydroepothilone D, cis-9,10-dehydroepothilone D, 16-desmethylepothilone B, epothilone BIO, discodermolide, patupilone (EPO-906), KOS-862, KOS-1584, ZK-EPO, ABJ-789, XAA296A (discodermolide ), TZT-1027 (Sobridotin), ILX-651 (Tacidotin hydrochloride), Halichondrin B, Eribulin mesylate (E-7389), Hemiasterin (HTI-286), E-7974, Shiptophycin, LY-355703, Maytansinoid immunoconjugate (DM-1), MKC-1, ABT-751, Tl-38067, T-900607, SB-715992 (Ispinesib), SB-743921 , MK-0731, STA-5312, eleutherobin, 17β-acetoxy-2-ethoxy-6-oxo-B-homo-estra-1,3,5(10)-trien-3-ol, cyclostreptin, isolaulimalide, laulimalide, 4-epi-7-dehydroxy-14,16-didemethyl-(+)-discodermolide, and the microtubule stabilizing agent cryptotyron 1, and any combination thereof.

[0153] In some embodiments, the anti-cancer agent comprises lymphodepleting chemotherapy. In some embodiments, the lymphodepleting chemotherapy is administered prior to the modified immune cells. In some embodiments, the lymphodepleting therapy comprises cyclophosphamide. In some embodiments, the lymphodepleting therapy comprises fludarabine. In some embodiments, the lymphodepleting therapy comprises cyclophosphamide and fludarabine.

[0154] In some embodiments, the anti-cancer agent comprises a cytokine. In some embodiments, the cytokine comprises an interleukin. In some embodiments, the cytokine is selected from IL2, IL7, IL12, IL15, IL17, IL21, granulocyte-macrophage colony-stimulating factor (GM-CSF), and interferon (IFN)-α. In some embodiments, the cytokine comprises IL-2.

[0155] II.C. Methods for Engineering Modified HLA In some embodiments of the present disclosure, the HLA binding pocket of the HLA molecule is modified using a gene editing tool. In some embodiments, the modification occurs in a cell expressing the HLA molecule ex vivo. In some embodiments, the modification occurs in a cell expressing the HLA molecule in a test tube. In some embodiments, the modification occurs in a cell expressing the HLA molecule in vivo. In some embodiments, the methods disclosed herein include genetically modifying an HLA molecule of a human subject using an in vivo gene editing tool.

[0156] Any gene editing tool can be used in the method of the present disclosure. In some embodiments, the gene editing tool comprises CRISPR / Cas9. In some embodiments, the gene editing tool comprises TALEN. In some embodiments, the gene editing tool comprises zinc finger nuclease. In some embodiments, the gene editing tool comprises meganuclease.

[0157] II.C.1.CRISPR / Cas9 In some embodiments, gene editing tools that can be used in the present disclosure include CRISPR / Cas systems. Such systems can employ, for example, Cas9 nuclease, which is optionally codon-optimized for the desired cell type (e.g., antigen-presenting cell) in which it is to be expressed. CRISPR / Cas systems use Cas nuclease, for example, Cas9 nuclease, which targets genomic sites by complexing with guide RNA (e.g., synthetic guide RNA) (gRNA) that hybridizes to the target DNA sequence immediately preceding the NGG motif recognized by Cas nuclease, for example, Cas9. A double-stranded break is generated three bases upstream of the NGG motif. Additional fusions with other enzymes can result in site-specific base editing in the absence of double-stranded breaks. The unique capability of the CRISPR / Cas9 system is its ability to simultaneously target multiple separate genomic loci by co-expressing a single Cas9 protein with two or more gRNAs (e.g., at least 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 gRNAs).

[0158] The CRISPR system used herein can use a fusion crRNA-tracrRNA construct (i.e., a single transcript) that functions with a codon-optimized Cas9. This single RNA is often referred to as a guide RNA or gRNA or single guide RNA or sgRNA. Within the gRNA, the crRNA portion is identified as the "target sequence" for a given recognition site, and the tracrRNA is often referred to as the "scaffold." Briefly, a short DNA fragment containing the target sequence is inserted into a guide RNA expression plasmid. The gRNA expression plasmid contains the target sequence (in some embodiments, about 20 nucleotides), some form of tracrRNA sequence (scaffold), as well as a suitable promoter that is active in the cell and the elements necessary for proper processing in eukaryotic cells. Many of the systems rely on custom-made complementary oligos that are annealed to form double-stranded DNA and then cloned into the gRNA expression plasmid.

[0159] The gRNA expression cassette and the Cas9 expression cassette are then introduced into the cells. See, e.g., Mali P et al., (2013), Science, 2013, Feb. 15; 339(6121):823-6; Jinek M. et al., Science, 2012, Aug. 17; 337(6096):816-21; Hwang WYet al., Nat. Biotechnol. 2013, March; 31(3):227-9; Jiang W. et al., Nat. Biotechnol. 2013, March; 31(3):233-9; Cronican et al., ACS. Chem. Biol. 5(8):747-52(2010); and Cong Let al., Science, 2013, Feb. 15; 339(6121):819-23, each of which is incorporated by reference herein in its entirety.

[0160] In some embodiments, the HLA binding pocket of the HLA molecule is modified using CRISPR / Cas9.

[0161] II.C.2.TALEN In some embodiments, the gene editing tool that can be used in the present disclosure comprises nuclease agents such as transcription activator-like effector nuclease (TALEN).TAL effector nuclease is a class of sequence-specific nuclease that can be used to make double-strand breaks at specific target sequences in the genome of prokaryotes or eukaryotes.TAL effector nuclease is created by fusing natural or engineered transcription activator-like (TAL) effector, or its functional part, to the catalytic domain of endonuclease, for example, FokI.

[0162] The unique modular DNA-binding domain of TAL effectors allows the design of proteins with potentially any given DNA recognition specificity, and thus the DNA-binding domain of a TAL effector nuclease can be engineered to recognize a specific DNA target site and thus used to effect a double-strand break at the desired target sequence. See, WO2010 / 079430; Morbitzer et al., (2010), PNAS, 10.1073 / pnas.1013133107; Scholze & Boch, (2010), Virulence, 1:428-432; Christian et al., Genetics, (2010) 186:757-761; Li et al., (2010), Nuc. Acids Res. (2010), doi:10.1093 / nar / gkq704; and Miller et al., (2011), Nature Biotechnology, 29:143-148, all of which are incorporated by reference in their entireties.

[0163] In some embodiments, the TAL effector nuclease is engineered to cleave at or near a target nucleic acid sequence, e.g., in a genomic locus of interest, where the target nucleic acid sequence is at or near a sequence that is modified by a targeting vector. TAL nucleases suitable for use in the various methods and compositions provided herein include those that are specifically designed to bind at or near a target nucleic acid sequence that is modified by a targeting vector described herein.

[0164] II.C.3. Zinc finger nucleases In some embodiments, gene editing tools that can be used in the present disclosure include nuclease agents, such as zinc finger nuclease (ZFN) systems. Zinc finger-based systems include fusion proteins that include two protein domains: a zinc finger DNA binding domain and an enzymatic domain. A "zinc finger DNA binding domain", "zinc finger protein", or "ZFP" is a protein or a domain within a larger protein that binds to DNA in a sequence-specific manner via one or more zinc fingers, which are regions of amino acid sequence within the binding domain whose structure is stabilized by the coordination of a zinc ion. The zinc finger domain binds to a target DNA sequence, thereby directing the activity of the enzymatic domain to the vicinity of the sequence, thus inducing modification of an endogenous target gene in the vicinity of the target sequence. Zinc finger domains can be engineered to bind to virtually any desired sequence. Thus, after identifying a target locus that contains a target DNA sequence for which cleavage or recombination is desired, one or more zinc finger binding domains can be engineered to bind to one or more target DNA sequences in the target locus. Modification is achieved at the target locus by expression in a cell of a fusion protein containing the zinc finger binding domain and the enzyme domain.

[0165] Typically, a single zinc finger domain is about 30 amino acids long. An individual zinc finger binds to a three nucleotide (i.e., triplet) sequence (or a four nucleotide sequence that may overlap by one nucleotide with the four nucleotide binding site of an adjacent zinc finger). Thus, the length of the sequence (e.g., target sequence) to which the zinc finger binding domain is engineered to bind determines the number of zinc fingers of the engineered zinc finger binding domain. For example, for ZFPs whose finger motifs do not bind to overlapping subsites, a six nucleotide target sequence is bound by two finger binding domains, a nine nucleotide target sequence is bound by three finger binding domains, and so on. The binding sites (i.e., subsites) of the individual zinc fingers of a target site need not be contiguous, but can be separated by one or several nucleotides, depending on the length and nature of the amino acid sequence between the zinc fingers in the multi-finger binding domain (i.e., the inter-finger linker). In some embodiments, the DNA binding domain of an individual ZFN contains between three and six individual zinc finger repeats, each capable of recognizing between nine and eighteen base pairs.

[0166] Zinc finger binding domains can be engineered to bind to a sequence of choice. See, e.g., Beerli et al., (2002), Nature Biotechnol. 20:135-141; Pabo et al., (2001), Ann. Rev. Biochem. 70:313-340; Isalan et al., (2001), Nature Biotechnol. 19:656-660; Segal et al., (2001), Curr. Opin. Biotechnol. 12:632-637, Choo et al., (2000), Curr. Opin. Struct. Biol. 10:411-416; 2002-2003, Catalogue, New England Biolabs, Beverly, Mass.; and Belfort et al., (1997), Nucleic Acids Res. 25:3379-3388, each of which is incorporated by reference in its entirety. Engineered zinc finger binding domains can have novel binding specificities relative to naturally occurring zinc finger proteins. Engineering methods include, but are not limited to, rational design and various types of selection.

[0167] Any means for target DNA sequence selection can be used in the methods described herein. Target sites generally have a length of at least 9 nucleotides, and thus zinc finger binding domains containing at least three zinc fingers bind. However, for example, four finger binding domains can bind to 12 nucleotide target sites, five finger binding domains can bind to 15 nucleotide target sites, or six finger binding domains can bind to 18 nucleotide target sites. Obviously, larger binding domains (e.g., seven, eight, nine fingers and larger) can also bind to even longer target sites.

[0168] The enzyme domain portion of the zinc finger fusion protein can be obtained from any endo- or exonuclease. Exemplary endonucleases from which the enzyme domain can be derived include, but are not limited to, restriction endonucleases and homing endonucleases. See, e.g., 2002-2003, Catalogue, New England Biolabs, Beverly, Mass.; and Belfort et al., (1997), Nucleic Acids Res. 25:3379-3388. Additional enzymes that cleave DNA are known (e.g., 51 nuclease; mung bean nuclease; pancreatic DNase I; micrococcal nuclease; yeast HO endonuclease; see also Linn et al. (eds.), Nucleases, Cold Spring Harbor Laboratory Press, 1993). One or more of these enzymes (or functional fragments thereof) can be used as the source of the cleavage domain.

[0169] II.C.4. Meganucleases In some embodiments, the gene editing tool that can be used is a meganuclease system. The domains, structures and functions of meganucleases are known, see, for example, Guhan and Muniyappa, (2003), Crit. Rev. Biochem. Mol. Biol. 38: 199-248; Lucas et al., (2001), Nucleic Acids Res. 29: 960-9; Jurica and Stoddard, (1999), Cell Mol. Life Sci. 55: 1304-26; Stoddard, (2006), Q Rev. Biophys. 38: 49-95; and Moure et al., (2002), Nat. Struct. Biol. 9: 764.

[0170] In some instances, naturally occurring mutant and / or engineered derivative meganucleases are used. Methods for modifying kinetics, cofactor interactions, expression, optimum conditions, and / or recognition site specificity, as well as screening for activity, are known, e.g., see Epinat et al., (2003), Nucleic Acids Res. 31:2952-62; Chevalier et al., (2002), Mol. Cell, 10:895-905; Gimble et al., (2003), Mol. Biol. 334:993-1008; Seligman et al., (2002), Nucleic Acids Res. 30:3870-9; Sussman et al., (2004), J. Mol. Biol. 342:31-41; Rosen et al., (2006), Nucleic Acids Res. 34:4791-800; Chames et al., (2006), Nucleic Acids Res. 34:4791-800; each of which is incorporated herein by reference in its entirety. See, e178, Smith et al., (2006), Nucleic Acids Res. 34: e149, Gruen et al., (2002), Nucleic Acids Res. 30: e29, Chen and Zhao, (2005), Nucleic Acids Res. 33: e154, WO2005105989, WO2003078619, WO2006097854, WO2006097853, WO2006097784, and WO2004031346.

[0171] II. Methods for enriching a target population of DT cells Without being bound to a particular mechanism, the modified HLA molecules described herein have increased affinity for antigens, thereby allowing surface display of antigens that are recognized by low affinity TCRs, and allowing the identification of novel TCRs and the enrichment of T cells expressing such novel TCRs.

[0172] Some embodiments of the present disclosure are further directed to a method for identifying a novel T cell receptor (TCR) capable of binding to a target antigen-HLA complex, comprising: (i) contacting a target antigen with an antigen-HLA complex comprising a binding pocket that has been modified to increase the affinity of the HLA for the antigen; and (ii) contacting a plurality of TCRs with the target antigen-HLA complex. Some embodiments of the present disclosure are further directed to a method for identifying a novel T cell receptor (TCR) capable of binding to a target antigen-HLA complex, comprising: (i) contacting a target antigen with an engineered antigen-presenting cell, wherein the engineered antigen-presenting cell comprises a binding pocket that has been modified to increase the affinity of the HLA for the antigen; and (ii) contacting a plurality of TCRs with the target antigen-HLA complex.

[0173] In some embodiments, after contacting, the enriched population of T cells comprises a greater number of T cells capable of binding to the antigen-HLA complex compared to the number of T cells capable of binding to the antigen-HLA complex prior to contacting.

[0174] In some embodiments, T cells associated with antigen-HLA complexes are isolated. Conventional methods can be used to identify and sequence the TCR that binds to antigen-HLA complexes. The identified TCR is recombinantly expressed in immune cells, conferring the immune cells the ability to target antigens.

[0175] Some embodiments of the present disclosure are directed to a method for selecting T cells that can target tumor cells.In some embodiments, the method comprises contacting a population of isolated T cells with an antigen-HLA complex in vitro, wherein the HLA comprises a modified HLA binding pocket as described herein.In some embodiments, the T cells are obtained from a human subject.

[0176] The T cells obtained from the human subject can be any T cells disclosed herein. In some embodiments, the T cells obtained from the human subject are tumor infiltrating lymphocytes (TILs). In some embodiments, the method further comprises administering the enriched T cells to the human subject. In some embodiments, the subject is preconditioned prior to receiving the T cells as described herein.

[0177] III. Compositions of the Present Disclosure Some embodiments of the present disclosure are directed to HLA molecules that include modified HLA binding pockets. Modified HLA molecules can include any modified HLA molecules disclosed herein, for example, in section II.A. above. The modified HLA molecules described herein have improved affinity for antigens. Thus, some embodiments of the present disclosure are directed to antigen-HLA complexes that include HLA molecules that include modified binding pockets, as disclosed herein.

[0178] In some embodiments, the antigen is a tumor antigen. In some embodiments, the antigen is an antigen expressed by a pathogen. In some embodiments, the antigen is a viral antigen. In some embodiments, the antigen is a bacterial antigen. In some embodiments, the antigen is a fungal antigen. In some embodiments, the antigen is a polypeptide that is less than about 30 amino acids, less than about 29 amino acids, less than about 28 amino acids, less than about 27 amino acids, less than about 26 amino acids, less than about 25 amino acids, less than about 24 amino acids, less than about 23 amino acids, less than about 22 amino acids, less than about 21 amino acids, less than about 20 amino acids, less than about 19 amino acids, less than about 18 amino acids, less than about 17 amino acids, less than about 16 amino acids, less than about 15 amino acids, less than about 14 amino acids, less than about 13 amino acids, less than about 12 amino acids, less than about 11 amino acids, less than about 10 amino acids in length. In some embodiments, the antigen is an antigen recognized by a low affinity TCR (e.g., a low affinity antigen).

[0179] III.A. Cells Containing Modified HLA Some embodiments of the present disclosure are directed to cells comprising modified HLA molecules disclosed herein or nucleic acid molecules encoding modified HLA molecules. In some embodiments, the cells are mammalian cells. In some embodiments, the cells are human cells. In some embodiments, the cells are antigen-presenting cells. In some embodiments, the cells are dendritic cells. In some embodiments, the cells are artificial antigen-presenting cells. In some embodiments, the artificial antigen-presenting cells comprise beads (e.g., silicate beads, glass beads, metal beads, or combinations thereof), nanovesicles, microvesicles, exosomes, endosomes, or any combinations thereof. In some embodiments, the cells are in vivo. In some embodiments, the cells are ex vivo.

[0180] In some embodiments, the T cells are isolated from a human subject. In some embodiments, the cells are allogeneic cells. In some embodiments, the human subject is the same subject who will eventually receive T cell therapy. In some embodiments, the cells are donor cells, i.e., cells obtained from a subject other than the subject who may eventually receive the cells.

[0181] In some embodiments, the cell is a cell that does not naturally express CD3, and the cell is modified to express CD3. In some embodiments, the cell comprises a transgene that encodes CD3, and the transgene is expressed by the cell. In some embodiments, the cell comprises a transgene that encodes a protein that activates the expression of endogenous CD3 by the cell. In some embodiments, the cell comprises a transgene that encodes a protein or siRNA that inhibits the inhibitor of CD3 expression in the cell. In some embodiments, the transgene is integrated into the genome of the cell. In some embodiments, the transgene is not integrated into the genome of the cell.

[0182] In some embodiments, the cells are derived from pluripotent stem cells, such as embryonic stem cells (ESCs), hematopoietic stem cells (HSCs), or induced pluripotent stem cells (iPSCs). In some embodiments, the cells are isolated from peripheral blood mononuclear cells (PBMCs). In some embodiments, the cells are isolated from tumor biopsies, such as tumor infiltrating lymphocytes (TILs).

[0183] III.B. Nucleic Acid Molecules and Vectors Certain aspects of the present disclosure are directed to nucleic acid molecules encoding modified HLA molecules as disclosed herein. Some aspects of the present disclosure are directed to vectors comprising nucleic acid molecules encoding modified HLA molecules as disclosed herein. In some aspects, the vector is a viral vector. In some aspects, the vector is a viral particle or virus. In some aspects, the vector is a mammalian vector. In some aspects, the vector is a bacterial vector.

[0184] In certain embodiments, the vector is a retroviral vector. In some embodiments, the vector is selected from the group consisting of adenoviral vector, lentivirus, Sendai virus, baculovirus vector, Epstein-Barr virus vector, papovavirus vector, vaccinia virus vector, herpes simplex virus vector, and adeno-associated virus (AAV) vector. In certain embodiments, the vector is an AAV vector. In some embodiments, the vector is a lentivirus. In certain embodiments, the vector is an AAV vector. In some embodiments, the vector is a Sendai virus. In some embodiments, the vector is a hybrid vector. Examples of hybrid vectors that can be used in the present disclosure can be found in Huang and Kamihira, Biotechnol. Adv. 31(2):208-23(2103), which is incorporated herein by reference in its entirety.

[0185] III.C. Vaccines A particular embodiment of the present disclosure is a cancer vaccine comprising an antigen-HLA complex, wherein the HLA comprises a modified HLA binding pocket as disclosed herein. In some embodiments, the antigen is a tumor antigen. In some embodiments, the vaccine further comprises one or more excipients. In some embodiments, the vaccine further comprises one or more additional peptides. In some embodiments, the one or more additional peptides comprise one or more additional epitopes.

[0186] Any of the various aspects, embodiments, and options described herein may be combined in any and all variations.

[0187] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference.

[0188] Having generally described the present disclosure, a further understanding can be obtained by reference to the examples provided herein, which are for illustrative purposes only and are not intended to be limiting. EXAMPLES

[0189] Example 1: Methods

[0190] Cell samples Peripheral blood samples were obtained from healthy donors after Institutional Review Board approval. Mononuclear cells were obtained by density gradient centrifugation (Ficoll-Paque PLUS; GE Healthcare, Chicago, IL). K562 is an erythroleukemia cell line deficient in HLA expression. T2 is a T-cell leukemia / B-LCL hybrid cell line. Jurkat76 is a T-cell leukemia cell line lacking expression of TCR and CD8. K562, T2, and Jurkat76 cell lines were grown in RPMI1640 supplemented with 10% FBS and 50 μg / ml gentamicin (Thermo Fisher Scientific). HEK293T cell line was grown in DMEM supplemented with 10% FBS and 50 μg / ml gentamicin. K562, T2, and HEK293T cells were obtained from the American Type Culture Collection (ATCC, Manassas, VA). TILs isolated from metastatic melanoma patients were expanded in vitro. High-resolution HLA DNA typing was performed on TIL samples.

[0191] peptide

[0192] The synthetic peptides were dissolved in DMSO at 50 mg / ml. The peptide used was the A2-restricted heteroclitic NY-ESO-1 157-165 (SLLMWITQV), gp100 154-162 (KTWGQYWQV), and HIV pol 476-484 (ILKEPVHGV), and HLA-A24:02-restricted gp100-intron 4 170-178 (VYFFLPDHL), gp100-intron 4 161-180 (PSQPIIHTCVYFFLPDHLSF), gp100-intron 4 166-185 (IHTCVYFFLPDHLSFGRPFH), wild-type WT1 235-243 (CMTWNQMNL), heteroclitic WT1 235-243 (CYTWNQMNL), HTLV-1tax 301-309 (SFHSLHLLF), and HIVenv 584-592 The peptide was (RYLRDQQLL). 476-484HTLV-1 tax 301-309 , and HIV env 584-592 Peptides were used as negative controls. Examples of peptide sequences examined for peptide-HLA binding assays and for measuring peptide exchange efficiency are listed in Table 1.

[0193] gene

[0194] All HLA-A*24:02 genes were linked to a truncated NGFR (ΔNGFR) gene using the Furin-SGSG-F2A sequence and cloned into a pMX retroviral plasmid. The full-length gp100 gene was purchased from Dharmacon (Lafayette, CO). Genomic DNA of gp100 was isolated using the PureLink Genomic DNA Mini Kit (Thermo Fisher Scientific, Waltham, MA). All genes were cloned into a pMX retroviral vector and transduced using the 293GPG cell-based retroviral system.

[0195] Transformants

[0196] Jurkat76 / CD8 cells were transduced with individual TCRα and TCRβ genes as previously reported (see, e.g., T. Ochi et al., Optimization of T-cell reactivity by exploiting TCR chain centricity for the purpose of safe and effective antitumor TCR gene therapy. Cancer Immunol. Res. 3, 1070-1081 (2015), which is incorporated by reference in its entirety). Jurkat76 / CD8-derived TCR transfectants were purified (purity >95%) using CD3 microbeads (Miltenyi Biotec, Bergisch Gladbach, Germany). Retroviral supernatant from PG13 was used to transduce TCR genes into human primary T cells. T2 cells were retrovirally transduced with HLA-A*24:02 (wild type) or A*24:02(A81L, L82R, R83G) to generate T2-A*24:02 or T2-A*24:02(A81L, L82R, R83G), respectively. β2m sgRNA plasmid (Origene, Rockville, MD) was electroporated into T2 cells using a Gene Pulser Xcell (Bio-Rad, Hercules, CA). Cells were stained with biotin-conjugated anti-β2m antibody (clone 2M2, BioLegend, San Diego, CA) and β2m-negative cells were isolated using anti-biotin microbeads (Miltenyi Biotec, Bergisch Gladbach, German) to obtain β2m knockout T2 (T2 / β2mKO) cells. T2 / β2mKO cells were retrovirally transduced with β2m-binding HLA-A*02:01 (wild type or L81A) to generate T2 / β2mKO / β2m-A*02:01 (wild type or L81A). All HLA-A*24:02 genes were tagged with the ΔNGFR gene as described above, and ΔNGFR+ cells were purified (>95% purity) and used in subsequent experiments.

[0197] Flow cytometry

[0198] Cell surface molecules were stained with PC5-conjugated anti-CD8 mAb (clone B9.11; Beckman Coulter, Brea, CA), FITC-conjugated anti-NGFR (clone ME20.4; BioLegend, San Diego, CA), and APC / Cy7-conjugated anti-CD3 (clone UCHT1; BioLegend, San Diego, CA). Dead cells were identified with the LIVE / DEAD Fixable Aqua dead cell staining kit (Life Technologies, Carlsbad, CA). For intracellular staining, cells were fixed and permeabilized using the Cytofix / Cytoperm kit (BD Biosciences Franklin Lakes, NJ). Stained cells were analyzed by flow cytometry (Biosciences Franklin Lakes, NJ), and data analysis was performed using FlowJo (BD Franklin Lakes, NJ).

[0199] Peptide-HLA binding assay

[0200] T2-A24 (wild-type) cells or T2-A24 cells carrying single amino acid substitutions at positions 81, 82, or 83 were pulsed with 50 μg / ml of biotinylated peptides overnight at 37° C. After extensive washing, cells were stained with PE-conjugated streptavidin (SA-PE), washed, and fluorescence intensity was measured by flow cytometry analysis.

[0201] Cytokine ELISPOT analysis

[0202] IL-2 and IFN-γ ELISPOT analysis was performed as described previously. For IL-2 ELISPOT assays, PVDF plates (Millipore, Bedford, MA) were coated with capture mAb (SEL002; R&D Systems, Minneapolis, MN). T cells were cultured at 2 × 10 per well in the presence or absence of peptide. 4The plates were then washed and incubated with biotin-conjugated detection mAb (SEL002, R&D Systems, Brandywine, MD). After washing, alkaline phosphatase-conjugated streptavidin (Jackson ImmunoResearch, West Grove, PA) was added. The plates were washed and incubated with NBT / BCIP (nitroblue tetrazolium / 5-bromo-4-chloro-3-indolyl phosphate, Promega, Madison, WI) to develop IL-2 spots. For IFN-γ ELISPOT analysis, PVDF plates (Millipore, Bedford, MA) were coated with capture mAb (1-D1K; MABTECH, Mariemont, OH) and T cells were plated at 2 × 10 per well. 4 The plates were incubated with 1000 target cells at 37°C for 20–24 h. The plates were then washed and incubated with biotin-conjugated detection mAb (7-B6-1; MABTECH). HRP-conjugated SA (Jackson ImmunoResearch, West Grove, PA) was then added to develop the IFN-γ spots. The reaction was stopped by rinsing thoroughly with cold tap water. ELISPOT plates were scanned and counted using an ImmunoSpot plate reader and ImmunoSpot version 5.0 software (Cellular Technology Limited, Shaker Heights, OH).

[0203] CD8 in an HLA-A24-restricted, peptide-specific manner + TIL expansion

[0204] CD8 + CD8 T cells were isolated by negative magnetic selection using a T cell isolation kit (Miltenyi Biotec, Bergisch Gladbach, Germany). +TILs were purified. HLA-A*24:02 aAPCs were pulsed with 10 μg / ml of the class I-restricted peptide of interest for 6 h. The aAPCs were then irradiated at 200 Gy, washed, and added to the TILs at an effector to target (E:T) ratio of 20:1. After 48 h, 10 IU / ml IL-2 (Novartis, Basel, Switzerland), 10 ng / ml IL-15 (Peprotech, East Windsor, NJ), and 30 ng / ml IL-21 (Peprotech, East Windsor, NJ) were added to the cultures every 3 days.

[0205] Primary CD8 transduced with TCR + T cell proliferation

[0206] CD3 T cells were isolated via negative magnetic selection using a pan-T cell isolation kit (Miltenyi Biotec, Bergisch Gladbach, Germany). + T cells were purified. Purified T cells were stimulated with 200 Gy irradiated aAPC / mOKT3 at an E:T ratio of 20:1. After overnight incubation, the cloned TCR genes were retrovirally transduced into activated T cells via centrifugation at 1,000 g for 1 h at 32°C for three consecutive days. After 48 h, 100 IU / ml IL-2 and 10 ng / ml IL-15 were added to the TCR-transduced T cells. Culture medium was replenished every 2 to 3 days.

[0207] Human cell-based production of pHLA multimers

[0208] The affinity-matured HLA-A*24:02 gene was engineered to carry a Glu (E) residue instead of a Gln (Q) residue at position 115 of the α2 domain and an α3 domain derived from the mouse Kb gene instead of the α3 domain of HLA class I. The soluble A*24:02 was produced by fusing the extracellular domain of the affinity-matured HLA-A*24:02 gene with a Gly-Ser (GS) flexible linker followed by a 6×His tag. Q115E -K bWe generated soluble A*24:02 using the 293GPG cell-based retroviral system. Q115E -K b The genes were individually transfected into HEK293T cells. Soluble affinity matured A*24:02 Q115E -K b Stable HEK293T cells expressing A*24:02 were grown to confluence and the medium was replaced. After 48 hours, conditioned medium was collected and used immediately or frozen at -80°C for later use. Soluble A*24:02 produced by HEK293T transfectants Q115E -K b The containing supernatant was mixed with 100 μg / ml of the desired A*24:02-restricted peptide for in vitro peptide exchange and incubated overnight at 37° C. Anti-His mAB (clone AD1.1.10; Abcam) conjugated to a fluorescent dye such as phycoerythrin (PE) was used at a molar ratio of 2:1 to obtain soluble monomeric A*24:02 peptide-loaded peptides. Q115E -K b was dimerized at room temperature for 2 hours or at 4°C overnight. Q115E -K b The concentration of the molecules was measured by specific ELISA using anti-HLA class I mAb (clone W6 / 32) and anti-His tag biotinylated mAb (clone AD1.1.10, R&D Systems) as capture and detection antibodies, respectively.

[0209] Staining of pHLA multimers

[0210] T cells (2×10 5) in the presence of 50 nM dasatinib (LC Laboratories, Woburn, MA) for 30 min at 37 °C. Cells were then washed and incubated with 5–10 μg / ml pHLA multimer for 30 min at room temperature, followed by AffiniPure Fab fragment goat anti-mouse IgG1 conjugated to R-phycoerythrin (Jackson ImmunoResearch, West Grove, PA) for 15 min at 4 °C. Cells were then washed three times and co-stained with anti-CD8 mAb for 15 min at 4 °C. Dead cells were identified using the LIVE / DEAD Fixable Aqua Dead Cell Stain Kit.

[0211] Measurement of peptide exchange efficiency using ELISA

[0212] The efficiency of peptide exchange in the monomer was assessed using a competitive binding assay and an enzyme-linked immunosorbent assay (ELISA). 37 TYFSLNNK(-biotin)F 45The monomers were loaded with 100 μg / ml of biotinylated peptides bearing the nucleotide sequence 101-101-101 and incubated overnight at 37°C. The biotinylated peptide-HLA was purified and exchanged into phosphate-buffered saline (PBS) using an Amicon Ultra filter (molecular weight cut-off (MWCO) 10 kDa) (MilliporeSigma, Burlington, MA), mixed with 1 mg / ml of the peptide of interest, and incubated overnight at 37°C. ELISA plates were coated with 10 μg / ml of anti-HLA class I mAb (clone W6 / 32) in PBS overnight at 4°C. The plates were washed and blocked with 10% nonfat dry milk in PBS for 30 min at room temperature. pHLA monomer was added and incubated for 2 h at room temperature. After washing, the plates were incubated with streptavidin-conjugated alkaline phosphatase for 30 min at room temperature. Finally, the plates were washed and incubated with p-nitrophenyl phosphate (PNPP) substrate (Pierce, Rockford, IL) at room temperature. The reaction was stopped by adding 1 mol / L NaOH. Optical density (OD) (405 nm) was read (Spectramax 190 Microplate Reader; Molecular Devices, Sunnyvale, CA). The OD values ​​of control wells containing non-biotinylated peptide were subtracted from the OD values ​​of test wells containing biotinylated peptide. The peptide exchange efficiency for each monomer was calculated as follows: peptide exchange efficiency = [1 - (OD value with peptide / OD value with DMSO)] x 100. All samples were assayed in triplicate wells.

[0213] statistical analysis

[0214] Statistical analysis was performed using GraphPad Prism (GraphPad Software, San Diego, CA). To determine whether two groups were significantly different for a given variable, analysis was performed using Welch's t-test (two-tailed). P values ​​less than 0.05 were considered significant.

[0215] Example 2: Leucine substitution at position 81 of HLA-A*24:02 (A81L) During the evaluation of peptide immunogenicity, allelic variation among HLA class I alleles was examined. Among the key residues forming the peptide-binding pocket of HLA class I, two polymorphisms were found at position 81 within the F pocket: HLA-A and HLA-B alleles expressed either alanine (Ala) or leucine (Leu) residues, whereas HLA-C alleles expressed only the Leu residue. Furthermore, Leu was the most frequently found residue at position 81 among all HLA class I alleles, except for most members of the A24 supertype. Furthermore, as shown in Figure 1, when only the most frequently found HLA class I alleles within the general population are aligned, the α1 domain of HLA-A*24:02 differs from that of other HLA alleles at positions 81–83, suggesting that these residues may uniquely define HLA-A*24:02 and may affect its peptide binding.

[0216] While the A24 supertype is present in all ethnicities and is the second most common HLA-A allele worldwide, it is most prevalent in Asian ethnicities, with HLA-A*24:02 being the most common allele among the Japanese population. Therefore, in an attempt to understand how HLA-A*24:02 presents antigens, we investigated the biological effects of amino acid substitutions within the HLA-A*24:02 peptide-binding pocket.

[0217] T2 cells were used to transduce HLA-A*24:02 constructs expressing single amino acid substitutions at positions 81–83, substituting each position with the most common residue found in other HLA class I alleles (Figure 1). T2 lines expressing HLA-A*24:02 with an Ala to Leu substitution at position 81 (A81L), a Leu to arginine (Arg) substitution at position 82 (L82R), or an Arg to glycine substitution at position 83 (R83G) were all successfully transduced. However, significant differences in anti-HLA antibody binding were observed (Figure 2A). Whereas a pan-anti-HLA class I antibody (clone W6 / 32) was able to bind similarly to all HLA-A*24:02-transduced T2 cells with a higher mean fluorescence intensity (MFI) than to untransduced T2 cells, we found that anti-HLA-A*24 (clone 22E1) lost the ability to detect mutant HLA-A*24:02(L82R).

[0218] We investigated whether T2 transformants expressing HLA-A*24:02 constructs could present antigens to generate superagonist HLA, allowing one engineered HLA molecule to accommodate many native peptides, as opposed to producing many heteroclitic peptides. We pulsed T2 cells with a number of known HLA-A*24:02-restricted peptides derived from virus- and tumor-associated antigens, and analyzed cell surface presentation of pHLA complexes by flow cytometry. Consistent with the increased pan-HLA class I expression observed in T2 transformants, we found that all HLA-A*24:02 constructs, including the HLA-A*24:02(L82R) construct, were able to present peptides (Figure 2B), suggesting that the substitutions at positions 81–83 did not alter the surface expression or folding of HLA-A*24:02, but rather affected epitope recognition by anti-HLA-A*24 (clone 22E1) antibodies. However, assays showed that T2-HLA-A*24:02(A81L) had a significantly increased ability to present HLA-A*24:02-restricted peptides compared to wild-type or other HLA-A*24:02 variants. These results highlight the importance of this substitution in peptide anchoring and suggest that the HLA-A*24:02(A81L) superagonist construct may better sustain the presentation of HLA-A*24:02 antigens on the cell surface of antigen-presenting cells (APCs).

[0219] Example 3: Surface expression of HLA-A*02:01 Due to the expression of the Leu residue at position 81, HLA-A*02:01 is more similar to other common HLA-B and HLA-C alleles than HLA-A*24:02 (Figure 1). Because the A81L substitution in HLA-A*24:02 has a profound effect on surface expression of pMHC complexes, we investigated whether such a substitution alters antigen presentation involving HLA-A*02:01. To control for endogenous expression of HLA-A*02:01 in T2 cells, we generated β2m-binding HLA-A*02:01 with the L81A substitution expressed as a single chain (β2m-HLA-A*02:01) and transduced it into β2m knockout T2 cells (T2 / β2mKO). Consistent with previous observations, endogenous HLA expression was detected on the surface of wild-type T2 cells but was absent in T2 / β2mKO when detected using anti-β2m or pan-anti-HLA antibodies (Figure 3A). Unlike β2m-HLA-A*02:01 (wild-type), β2m-HLA-A*02:01 containing the L81A substitution showed significantly reduced expression, with slightly higher staining than in T2 / β2mKO (Figure 3A). To examine whether the L81A substitution only disrupted surface expression of β2m-HLA-A*02:01, we stained fixed and permeabilized T2 cells and found that although intracellular β2m expression was intact, intracellular staining using the conformation-dependent pan-HLA class I antibodies W6 / 32 or B9.12.1 did not detect β2m-HLA-A*02:01(L81A) at levels higher than surface staining (Figure 3B).

[0220] To confirm that reduced surface expression of β2m-HLA-A*02:01(L81A) translates into biological activity, we performed a 10-mL IgG antibody assay using HLA-A*02:01-restricted NY-ESO-1 157-165 Specific TCR (clone 1G4LY) or gp100 154-162 Specific TCR (clone gp100 154Functional assays were performed using J76 / CD8 cells expressing HLA-A*02:01 (wild type). Whereas T2-β2m-HLA-A*02:01 (wild type) pulsed with the cognate peptide was able to induce IFNγ from TCR-J76 / CD8 cells in ELISPOT assays (Fig. 3C), coculture with T2-β2m-HLA-A*02:01(L81A) significantly reduced IFNγ production (Fig. 3C). These data suggest that the reduced surface expression of β2m-HLA-A*02:01(L81A) was not due to altered epitope recognition by pan-anti-HLA antibodies, but rather was likely due to disruption of the formation of stable HLA-A*02:01pMHC complexes by the L81A substitution.

[0221] Example 4: HLA-A*24:02(A81L) enhances peptide-specific T cell responses We investigated whether increased surface expression of the superagonist HLA leads to increased T cell activation. HLA-A*24:02 / WT1 235-243 Jurkat76 / CD8 cells transduced with TCRs of various avidity against the epitope were used to detect WT1 235-243 We found that T2-HLA-A*24:02(A81L) was more potent at inducing T cell secretion of IL-2 compared to T2-HLA-A*24:02(wild type) when either was continuously present in the culture medium (Fig. 4A, top) or pulsed onto the cells prior to the T cell assay (Fig. 4A, bottom). All WT1 235-243 These results suggest that the A81L substitution may improve the ability to retain surface expression of HLA-A*24:02 when observed for specific TCR. Since short peptides have the ability to bind to cell surface HLA, thereby bypassing potential endosomal processing, we investigated whether the A81L substitution alters the ability to cross-present internalized HLA-A*24:02-restricted peptides. Using T2 transfectants, we expressed HLA-A*24:02-restricted gp100int4 170-178Consistent with the ability to enhance T cell activation to short peptides, T2-HLA-A*24:02(A81L)-activated TCR-transduced T cells pulsed with long peptides showed a more than four-fold increase in the number of IFNγ-secreting spots compared to HLA-A*24:02(wild type) (Figure 4B).

[0222] Example 5: HLA-A*24:02(A81L) multimers strongly stain low affinity antigen-specific TCRs A major limitation of TCR-based cancer immunotherapy is the ability to identify TCRs with low affinity antigens. Designing multimers that can reliably stain such TCRs could lead to more promising cancer treatment options. Having found that the A81L substitution enhances antigen presentation of naturally occurring HLA-A*24:02 peptides to T cells, we investigated whether superagonist HLA constructs could be used as reagents to improve testing of HLA-A*24:02-restricted T cells. Novel peptide-exchangeable affinity-matured HLA class I multimers were previously designed to be used to detect, select, and clone tumor antigen-specific TCRs from tumor infiltrates. Using this platform, we generated affinity-matured HLA-A*24:02 multimers expressing the A81L substitution to determine whether this substitution would further enhance the utility of these multimers in detecting low affinity TCRs. The efficiency of peptide exchange with HLA-A*24:02-restricted peptides derived from a number of virus-associated and tumor-associated antigens was examined using a cell-free assay. Compared to wild-type HLA-A*24:02 monomer, the A81L substitution significantly improved the peptide exchange efficiency for all peptides that had low exchange efficiency in wild-type HLA-A*24:02 affinity matured monomer, increasing the exchange efficiency by at least 2-fold (Figures 5A-5B). 76-84For peptides, affinity matured HLA-A*24:02(A81L) monomers, but not wild-type monomers, resulted in detectable peptide exchange (Figures 5A-5B, peptide 32). The A81L substitution did not compromise peptide specificity, and the HLA-B*18 restricted peptide MAGE-A3 167-176 , HLA-B*27 restricted peptide VEGF(UTR), and HLA-C*06 restricted peptide GAGE1 / 2 / 8 9-16 did not result in obvious exchange to affinity matured HLA-A*24:02(A81L) monomers (Figures 5A-5B, peptides 65-67). Furthermore, we found HLA-A*24:02(A81L) monomers capable of efficient peptide exchange, and therefore wild-type multimers and A81LHLA-A*24:02WT1 235-243 We stained Jurkat76 / CD8 cells expressing cognate TCRs of various avidities with the multimers. Consistent with the exchange efficiency, the HLA-A*24:02(A81L) superagonist multimers showed a higher ability to stain TCRs of low avidity compared to the HLA-A*24:02 wild-type multimers. For TCRs A133 and A186, the HLA-A*24:02(A81L) multimers produced staining of T cells that would have been undetectable under the experimental conditions used (Figure 5C).

[0223] Example 6: HLA-A*24:02(A81L)aAPC enhances proliferation of tumor antigen-specific T cells In addition to enhancing the detection of low avidity T cells, we investigated whether the A81L substitution would be useful for the expansion of T cells derived from tumor infiltrates. Using a previously developed platform of artificial antigen-presenting cells (aAPCs), we identified T cells expressing HLA-A*24:02(A81L) or wild-type and expressing gp100int4. 170-178 or control HTLV-1tax 301-309 Short-term expansion of tumor-infiltrating lymphocytes (TILs) from melanoma patients was performed using peptide-pulsed aAPCs. After 2 weeks of coculture, gp100int4 170-178 aAPC-HLA-A*24:02(A81L) pulsed with HLA-A*24:02(A81L) upregulated antigen-specific CD8+ This resulted in a greater than two-fold expansion of T cells (Figure 6A). This ability to enhance antigen-specific T cell proliferation was consistent across three replicate experiments (Figure 6B), resulting in a 6-fold expansion from baseline TILs (Figure 6C).

[0224] The data presented herein show that generation of HLA-A*24:02-derived superagonist HLA constructs expressing the A81L substitution enhances T cell activation in coculture assays, increases cytokine secretion and T cell proliferation, and does not interfere with the processing and presentation of HLA-A*24:02-restricted antigens. In cell-free assays, the superagonist HLA significantly increased the efficiency of peptide exchange for HLA-A*24:02-restricted peptides, and importantly, did not alter the restriction of peptides to HLA-A*24:02, as no increase in peptide presentation by T2 cells or for HLA-A*24:02-nonrestricted peptides was observed in cell-free peptide exchange assays.

[0225] Given the importance of residue 81 in forming part of the F-binding pocket, it was surprising that the A81L substitution did not adversely affect binding of HLA-A*24:02-restricted peptides and in fact increased the surface availability of peptide-HLA-A*24:02 complexes and enhanced T cell responses. This was especially true for HLA-A*24:02-restricted peptides, which barely bound wild-type HLA-A*24:02. As with other peptide-HLA I complexes, the overall structure of HLA-A*24:02 is similar to other HLA I alleles, adopting the well-described α1 and α2 domains formed by an antiparallel β-sheet and two long α-helices that constitute the peptide-binding interface presented to the TCR. Unique to HLA-A*24:02, however, are the unusually deep B and F peptide-binding pockets that can accommodate bulky aromatic and large hydrophobic side chains such as the anchor residues Y or F (at position 2) and F, L, I, or W (at the C-terminus) of peptide ligands. It is unclear whether the A81L substitution (see Figures 7A-7B for in silico modeling) would alter this preference; however, no bias was observed for the presentation of specific HLA-A*24:02-restricted peptides, but the ability of all peptides examined to enhance peptide presentation and T cell activation. This observation likely highlights the importance of secondary peptide anchors, which are known to strongly influence the binding capacity of peptide ligands to HLA-A*24:02. Indeed, the A81L substitution may affect how the secondary peptide anchors affect the unique conformation that peptides adopt within HLA-A*24:02. Several crystal structures have shown that peptide ligands in HLA-A*24:02 adopt a moderately sized “A” or “M” shape at the central residue (anchored at position 5) compared to other HLA I alleles, providing a unique peptide “bulge” for TCR docking and T cell recognition.Interestingly, similar to the unique effects of secondary anchors on various HLAI alleles, the Leu modification at position 81 may be unique to HLA-A*24:02, as a substitution at position 81 within HLA-A*02:01 renders this allele unable to activate T cells, highlighting the importance of this residue in surface expression of the pHLA complex.

[0226] Since peptide / HLA affinity, and thus surface expression, are key factors determining the immunogenicity of pMHC complexes, superagonist modifications may increase the binding affinity of peptides to HLA-A*24:02 or alter the conformation of peptide binding to enhance T cell activation. When this substitution is applied to HLA multimer staining, superagonist multimers can be used to detect TCRs that are likely low affinity but not detected with wild-type monomers. Consistent with this observation, aAPCs expressing HLA-A*24:02(A81L) increased the proliferation of antigen-specific T cells derived from TILs. The ability of superagonist multimers to detect low affinity TCRs, together with the ability of aAPCs to expand antigen-specific T cells, may allow for a more powerful probing of both high and low affinity T cell repertoires for any given epitope derived from tumor antigens.

[0227] Overall, these data present a novel method to enhance pHLA immunogenicity and show that modification of the HLA binding pocket can be used to enhance T cell activation and proliferation to a broad range of peptides while maintaining antigen restriction. The specific A81L substitution can be directly applied to HLA class I alleles containing Ala at position 81. Namely, members of the A24 supertype (HLA-A*23 and HLA-A*24 alleles), as well as the A01- (HLA-A*25, HLA-A*31, HLA-A*32 alleles), B07- (HLA-B*51 alleles), B44- (HLA-B*44 alleles), and B58- (HLA-B*58 alleles) supertypes share F-pocket peptide specificity with HLA-A*24:02, but most of the various members of the alleles do not fall into a particular supertype (HLA-B*13, HLA-B*=38, HLA-B*49, HLA-B*52, HLA-B*53, HLA-B*57) (2). For other alleles, applying crystal structure-guided modifications from position 81 onwards may reduce the hassle of developing heteroclitic peptides for each peptide epitope individually.

Claims

[Claim 1] The invention as described herein and in the drawings.

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