PMID- 34266882 OWN - NLM STAT- MEDLINE DCOM- 20220111 LR - 20220111 IS - 2051-1426 (Electronic) IS - 2051-1426 (Linking) VI - 9 IP - 7 DP - 2021 Jul TI - Influence of DM-sensitivity on immunogenicity of MHC class II restricted antigens. LID - 10.1136/jitc-2021-002401 [doi] LID - e002401 AB - BACKGROUND: Graft-versus-host-disease (GvHD) is a major problem in allogeneic stem cell transplantation. We previously described two types of endogenous human leukocyte antigen (HLA)-II restricted antigens depending on their behavior towards HLA-DM. While DM-resistant antigens are presented in the presence of HLA-DM, DM-sensitive antigens rely on the expression of HLA-DO-the natural inhibitor of HLA-DM. Since expression of HLA-DO is not upregulated by inflammatory cytokines, DM-sensitive antigens cannot be presented on non-hematopoietic tissues even under inflammatory conditions. Therefore, usage of CD4+ T cells directed against DM-sensitive antigens might allow induction of graft-versus-leukemia effect without GvHD. As DM-sensitivity is likely linked to low affinity peptides, it remains elusive whether DM-sensitive antigens are inferior in their immunogenicity. METHODS: We created an in vivo system using a DM-sensitive and a DM-resistant variant of the same antigen. First, we generated murine cell lines overexpressing either H2-M or H2-O (murine HLA-DM and HLA-DO) to assign the two model antigens ovalbumin (OVA) and DBY to their category. Further, we introduced mutations within the two T-cell epitopes and tested the effect on DM-sensitivity or DM-resistance. Furthermore, we vaccinated C57BL/6 mice with either variant of the epitope and measured expansion and reactivity of OVA-specific and DBY-specific CD4+ T cells. RESULTS: By testing T-cell recognition of OVA and DBY on a murine B-cell line overexpressing H2-M and H2-O, respectively, we showed that OVA leads to a stronger T-cell activation in the presence of H2-O demonstrating its DM-sensitivity. In contrast, the DBY epitope does not rely on H2-O for T-cell activation indicating DM-resistance. By introducing mutations within the T-cell epitopes we could generate one further DM-sensitive variant of OVA and two DM-resistant counterparts. Likewise, we designed DM-resistant and DM-sensitive variants of DBY. On vaccination of C57BL/6 mice with either epitope variant we measured comparable expansion and reactivity of OVA-specific and DBY-specific T-cells both in vivo and ex vivo. By generating T-cell lines and clones of healthy human donors we showed that DM-sensitive antigens are targeted by the natural T-cell repertoire. CONCLUSION: We successfully generated DM-sensitive and DM-resistant variants for two model antigens. Thereby, we demonstrated that DM-sensitive antigens are not inferior to their DM-resistant counterpart and are therefore interesting tools for immunotherapy after allogeneic stem cell transplantation. CI - (c) Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY. Published by BMJ. FAU - Bernhardt, Anna Luise AU - Bernhardt AL AD - Department of Internal Medicine 5 - Hematology and Internal Oncology, Friedrich-Alexander University Erlangen-Nuremberg, Erlangen, Bayern, Germany. FAU - Zeun, Julia AU - Zeun J AD - Department of Internal Medicine 5 - Hematology and Internal Oncology, Friedrich-Alexander University Erlangen-Nuremberg, Erlangen, Bayern, Germany. FAU - Marecek, Miriam AU - Marecek M AD - Department of Internal Medicine 5 - Hematology and Internal Oncology, Friedrich-Alexander University Erlangen-Nuremberg, Erlangen, Bayern, Germany. FAU - Reimann, Hannah AU - Reimann H AD - Department of Internal Medicine 5 - Hematology and Internal Oncology, Friedrich-Alexander University Erlangen-Nuremberg, Erlangen, Bayern, Germany. FAU - Kretschmann, Sascha AU - Kretschmann S AD - Department of Internal Medicine 5 - Hematology and Internal Oncology, Friedrich-Alexander University Erlangen-Nuremberg, Erlangen, Bayern, Germany. FAU - Bausenwein, Judith AU - Bausenwein J AD - Department of Internal Medicine 5 - Hematology and Internal Oncology, Friedrich-Alexander University Erlangen-Nuremberg, Erlangen, Bayern, Germany. FAU - van der Meijden, Edith D AU - van der Meijden ED AD - Department of Internal Medicine 5 - Hematology and Internal Oncology, Friedrich-Alexander University Erlangen-Nuremberg, Erlangen, Bayern, Germany. FAU - Karg, Margarete M AU - Karg MM AD - Department of Internal Medicine 5 - Hematology and Internal Oncology, Friedrich-Alexander University Erlangen-Nuremberg, Erlangen, Bayern, Germany. AD - Schepens Eye Research Institute, Harvard Medical School, Boston, Massachusetts, USA. FAU - Haug, Tabea AU - Haug T AD - Department of Internal Medicine 5 - Hematology and Internal Oncology, Friedrich-Alexander University Erlangen-Nuremberg, Erlangen, Bayern, Germany. FAU - Meintker, Lisa AU - Meintker L AD - Department of Internal Medicine 5 - Hematology and Internal Oncology, Friedrich-Alexander University Erlangen-Nuremberg, Erlangen, Bayern, Germany. FAU - Lutzny-Geier, Gloria AU - Lutzny-Geier G AD - Department of Internal Medicine 5 - Hematology and Internal Oncology, Friedrich-Alexander University Erlangen-Nuremberg, Erlangen, Bayern, Germany. FAU - Mackensen, Andreas AU - Mackensen A AD - Department of Internal Medicine 5 - Hematology and Internal Oncology, Friedrich-Alexander University Erlangen-Nuremberg, Erlangen, Bayern, Germany. FAU - Kremer, Anita N AU - Kremer AN AUID- ORCID: 0000-0001-9309-8972 AD - Department of Internal Medicine 5 - Hematology and Internal Oncology, Friedrich-Alexander University Erlangen-Nuremberg, Erlangen, Bayern, Germany anita.kremer@uk-erlangen.de. LA - eng PT - Journal Article PT - Research Support, Non-U.S. Gov't PL - England TA - J Immunother Cancer JT - Journal for immunotherapy of cancer JID - 101620585 RN - 0 (DNA-Binding Proteins) RN - 0 (Transcription Factors) RN - 148770-78-9 (HIVEP2 protein, human) SB - IM MH - Animals MH - Antigen Presentation/*immunology MH - DNA-Binding Proteins/*metabolism MH - Humans MH - Immunotherapy/*methods MH - Mice MH - Mice, Transgenic MH - Transcription Factors/*metabolism PMC - PMC8286791 OTO - NOTNLM OT - CD4-positive T-lymphocytes OT - adoptive OT - antigen presentation OT - antigens OT - immunotherapy COIS- Competing interests: None declared. EDAT- 2021/07/17 06:00 MHDA- 2022/01/12 06:00 PMCR- 2021/07/15 CRDT- 2021/07/16 05:51 PHST- 2021/06/18 00:00 [accepted] PHST- 2021/07/16 05:51 [entrez] PHST- 2021/07/17 06:00 [pubmed] PHST- 2022/01/12 06:00 [medline] PHST- 2021/07/15 00:00 [pmc-release] AID - jitc-2021-002401 [pii] AID - 10.1136/jitc-2021-002401 [doi] PST - ppublish SO - J Immunother Cancer. 2021 Jul;9(7):e002401. doi: 10.1136/jitc-2021-002401.