Dataset Information

Hotspot DNMT3A and IDH1/2 Mutations in Acute Myeloid Leukemia and Their Relevance as Targets for Immunotherapy

ABSTRACT: DNA methyltransferase 3A (DNMT3A) and isocitrate dehydrogenase 1 & 2 (IDH1/2) are genes involved in epigenetic regulation, each mutated in 7-23% of acute myeloid leukemia (AML). Here, we investigated whether hotspot mutations in these genes encode neoantigens that can be targeted by immunotherapy. Five EBV-B cell lines expressing common HLA class I alleles were transduced with a minigene construct containing mutations that often occur in DNMT3A or IDH1/2. Peptides eluted from HLA class I alleles on these EBV-B cell lines were analyzed by tandem mass spec-trometry. We identified an HLA-A*01:01-binding DNMT3AR882H peptide and HLA-B*07:02-binding IDH2R140Q peptide, for which we searched for specific T cells in healthy individuals using pep-tide-HLA tetramers. Various T-cell clones were isolated showing specific reactivity against cell lines transduced with full-length DNMT3AR882H or IDH2R140Q genes, while cell lines transduced with wildtype genes were not recognized. One T-cell clone for DNMT3AR882H also reacted against patient AML cells with the mutation, while AML cells without the mutation were not recognized, thereby validating surface presentation of a DNMT3AR882H neoantigen that can potentially be targeted on HLA-A*01:01 positive AML by immunotherapy.

INSTRUMENT(S): Orbitrap Fusion Lumos, Orbitrap Exploris 480

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

SUBMITTER: Rayman Tjokrodirijo

LAB HEAD: Peter A.

PROVIDER: PXD050560 | Pride | 2024-06-23

REPOSITORIES: Pride

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Hotspot DNA Methyltransferase 3A (DNMT3A) and Isocitrate Dehydrogenase 1 and 2 (IDH1/2) Mutations in Acute Myeloid Leukemia and Their Relevance as Targets for Immunotherapy.

Struckman Nadine E NE de Jong Rob C M RCM Honders M Willy MW Smith Sophie-Anne I SI van der Lee Dyantha I DI Koutsoumpli Georgia G de Ru Arnoud H AH Mikesch Jan-Henrik JH van Veelen Peter A PA Falkenburg J H Frederik JHF Griffioen Marieke M

Biomedicines 20240514 5

DNA methyltransferase 3A (DNMT3A) and isocitrate dehydrogenase 1 and 2 (IDH1/2) are genes involved in epigenetic regulation, each mutated in 7-23% of patients with acute myeloid leukemia. Here, we investigated whether hotspot mutations in these genes encode neoantigens that can be targeted by immunotherapy. Five human B-lymphoblastoid cell lines expressing common HLA class I alleles were transduced with a minigene construct containing mutations that often occur in DNMT3A or ...[more]

PMID: 38791049

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Project description:Human leukocyte antigen (HLA) - B*51:01 and endoplasmic reticulum aminopeptidase 1 (ERAP1) are two strong predisposing genetic factors to Behçet's disease (BD). Previous studies have focused on two subgroups of HLA-B*51 peptidome containing Proline (Pro) or Alanine (Ala) at position 2 (P2). Little is known about the unconventional non-Pro/Ala2 HLA-B*51-bound peptides. We aimed to study the features of this novel sub-peptidome, and investigate its regulation by ERAP1. CRISPR-Cas9 was used to generate an HLA-ABC-knockout HeLa cell line (HeLa.ABC-KO), which was subsequently transduced to express HLA-B*51:01 (HeLa.ABC-KO.B51). ERAP1 was silenced using lentiviral shRNA. Peptides bound to HLA-B*51:01 were eluted and analyzed by Mass Spectrometry. The characteristics of non-Pro/Ala2, Pro2 and Ala2 peptides, and their alteration by ERAP1 silencing were investigated. Effects of ERAP1 silencing on cell surface expression of HLA-B*51:01 were studied using flow cytometry. More than 20% of peptides eluted from HLA-B*51:01 lacked Proline or Alanine at P2. This unconventional group of HLA-B*51:01-bound peptides was enriched for 8-mers (with fewer 9-mers) compared to Pro2 and Ala2 sub-peptidomes, and had similar N-terminal and C-terminal residue usages to Ala2 peptides with the exception of the less abundant Leucine at position Ω. Knockdown of ERAP1 increased the percentage of non-Pro/Ala2 to approximately 40%, increased the percentage of longer (10-mer and 11-mer) peptides eluted from HLA-B*51:01 complexes, and abrogated the predominance of Leucine at P1. Interestingly knockdown of ERAP1 altered the length and N-terminal residue usage of non-Ala2&Pro2 and Ala2 but not the Pro2 peptides. Finally, ERAP1 silencing regulated the expression levels of cell surface HLA-B*51 in a cell type - dependent manner. The HLA-B*51:01 peptidome includes a surprisingly high proportion of unconventional non-Pro/Ala2 peptides which are increased by ERAP1 silencing, mimicking the loss-of-function BD risk variant.

			Action	DRS
	AdR_JYW5098_dB_Homo2020.msf	Msf
	E20202002438a.raw	Raw
	E20202002438b.raw	Raw
	E20202002439a.raw	Raw
	E20202002439b.raw	Raw

Dataset Information

Hotspot DNMT3A and IDH1/2 Mutations in Acute Myeloid Leukemia and Their Relevance as Targets for Immunotherapy

Dataset's files

Publications

Hotspot DNA Methyltransferase 3A (<i>DNMT3A</i>) and Isocitrate Dehydrogenase 1 and 2 (<i>IDH1/2</i>) Mutations in Acute Myeloid Leukemia and Their Relevance as Targets for Immunotherapy.

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