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HLA class I-restricted MYD88 L265P-derived peptides as specific targets for lymphoma immunotherapy

View Article: PubMed Central - PubMed

ABSTRACT

Genome sequencing has uncovered an array of recurring somatic mutations in different non-Hodgkin lymphoma (NHL) subtypes. If affecting protein-coding regions, such mutations may yield mutation-derived peptides that may be presented by HLA class I proteins and recognized by cytotoxic T cells. A recurring somatic and oncogenic driver mutation of the Toll-like receptor adaptor protein MYD88, Leu265Pro (L265P) was identified in up to 90% of different NHL subtype patients. We therefore screened the potential of MYD88L265P-derived peptides to elicit cytotoxic T cell responses as tumor-specific neoantigens. Based on in silico predictions, we identified potential MYD88L265P-containing HLA ligands for several HLA class I restrictions. A set of HLA class I MYD88L265P-derived ligands elicited specific cytotoxic T cell responses for HLA-B*07 and -B*15. These data highlight the potential of MYD88L265P mutation-specific peptide-based immunotherapy as a novel personalized treatment approach for patients with MYD88L265P+ NHLs that may complement pharmacological approaches targeting oncogenic MyD88 L265P signaling.

No MeSH data available.


Efficient in vitro generation of P4B*15- and P1B*07-specific CD8+ T cells from naive T cells of CLL patients and HBDs. Representative tetramer stainings of CD8+ T cells after three cycles of aAPC-based in vitro priming using CD8+ T cells derived from HLA-matched HBDs primed with (A) the HLA-B*15-restricted peptide HQKRPIPIKY (P4B*15) and (B) the HLA-B*07-restricted peptide RPIPIKYKAM (P1B*07) as well as from HLA-matched MYD88WT CLL patient (CLL-05) primed with (C) the HLA-B*07-restricted peptide RPIPIKYKAM (P1B*07): 1st column: tetramer staining of CD8+ T cells primed with the MYD88L265P-derived peptide; 2nd column: control staining with HLA-matched tetramer containing a non-relevant control peptide on CD8+ T cells derived from the same population as T cells depicted in the 1st column; 3rd column: ex vivo tetramer staining of CD8+ T cells. In vitro primings with HBD-derived PBMCs were performed in six (P1B*07) and three (P4B*15) independent replicates, respectively. For the in vitro priming with PBMCs of CLL patients two independent replicates were conducted. Abbreviations: neg., negative.
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f0003: Efficient in vitro generation of P4B*15- and P1B*07-specific CD8+ T cells from naive T cells of CLL patients and HBDs. Representative tetramer stainings of CD8+ T cells after three cycles of aAPC-based in vitro priming using CD8+ T cells derived from HLA-matched HBDs primed with (A) the HLA-B*15-restricted peptide HQKRPIPIKY (P4B*15) and (B) the HLA-B*07-restricted peptide RPIPIKYKAM (P1B*07) as well as from HLA-matched MYD88WT CLL patient (CLL-05) primed with (C) the HLA-B*07-restricted peptide RPIPIKYKAM (P1B*07): 1st column: tetramer staining of CD8+ T cells primed with the MYD88L265P-derived peptide; 2nd column: control staining with HLA-matched tetramer containing a non-relevant control peptide on CD8+ T cells derived from the same population as T cells depicted in the 1st column; 3rd column: ex vivo tetramer staining of CD8+ T cells. In vitro primings with HBD-derived PBMCs were performed in six (P1B*07) and three (P4B*15) independent replicates, respectively. For the in vitro priming with PBMCs of CLL patients two independent replicates were conducted. Abbreviations: neg., negative.

Mentions: To assess whether MYD88L265P-derived peptide-specific T cell responses can be induced from naive T cells in vitro, we isolated CD8+ T cells from six HLA-B*07+ and three HLA-B*15+ HBDs as well as from two HLA-B*07+MYD88WT CLL patients. We performed artificial antigen-presenting cell (aAPC)-based in vitro priming using the three HLA-B*07-restricted MYD88L265P-derived peptides RPIPIKYKAM (P1B*07), RPIPIKYKA (P2B*07) and SPGAHQKRPI (P3B*07) as well as the HLA-B*15-restricted peptide HQKRPIPIKY (P4B*15). Using HBD-derived CD8+ T cells, P4B*15-tetramer-positive CD8+ populations with frequencies of 1.50–10.68% of viable cells were detected in 2/3 HBDs after priming (Fig. 3A). For the HLA-B*07-restricted peptide P1B*07, we observed tetramer-positive CD8+ populations with frequencies of 0.47–14.04% of viable cells in 6/6 HBDs (Fig. 3B). For P2B*07, 1/3 (33%) HBDs showed a tetramer-positive CD8+ population with a frequency of 0.15% of viable cells (Fig. S1A). P3B*07-tetramer positive CD8+ populations with frequencies of 0.20–1.56% of viable cells were detected in 3/4 HBDs (Fig. S1B). Notably, after aAPC-based in vitro priming of CD8+ T cells from MYD88WT B*07+ CLL patients without previous T cell reactivity for P1B*07 (as detected by 12-day recall IFNγ ELISPOT assay and ex vivo tetramer staining), we observed in 1/2 patients a population of 0.40% P1B*07-specific CD8+ T cells within the viable cells (Fig. 3C). No tetramer-positive T cell populations >0.10% (>0.50%) were detectable in control stainings with an HLA-B*07 (HLA-B*15)-tetramer containing a control peptide. In ex vivo control stainings no tetramer-positive T cell populations >0.01% were detectable. Furthermore, Peper et al.31 demonstrated that T cell responses observed after three rounds of aAPC-based stimulations were mediated by in vitro primed naive T cells rather than by pre-existing memory T cells, as short-time stimulation of the same PBMC did not result in the detection of specific T cell populations. Collectively, all 4/4 (100%) refolded MYD88L265P-derived peptides thus are able to efficiently prime mutation-specific T cells in certain HLA contexts in HBDs as well as in CLL patients.Figure 3.


HLA class I-restricted MYD88 L265P-derived peptides as specific targets for lymphoma immunotherapy
Efficient in vitro generation of P4B*15- and P1B*07-specific CD8+ T cells from naive T cells of CLL patients and HBDs. Representative tetramer stainings of CD8+ T cells after three cycles of aAPC-based in vitro priming using CD8+ T cells derived from HLA-matched HBDs primed with (A) the HLA-B*15-restricted peptide HQKRPIPIKY (P4B*15) and (B) the HLA-B*07-restricted peptide RPIPIKYKAM (P1B*07) as well as from HLA-matched MYD88WT CLL patient (CLL-05) primed with (C) the HLA-B*07-restricted peptide RPIPIKYKAM (P1B*07): 1st column: tetramer staining of CD8+ T cells primed with the MYD88L265P-derived peptide; 2nd column: control staining with HLA-matched tetramer containing a non-relevant control peptide on CD8+ T cells derived from the same population as T cells depicted in the 1st column; 3rd column: ex vivo tetramer staining of CD8+ T cells. In vitro primings with HBD-derived PBMCs were performed in six (P1B*07) and three (P4B*15) independent replicates, respectively. For the in vitro priming with PBMCs of CLL patients two independent replicates were conducted. Abbreviations: neg., negative.
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f0003: Efficient in vitro generation of P4B*15- and P1B*07-specific CD8+ T cells from naive T cells of CLL patients and HBDs. Representative tetramer stainings of CD8+ T cells after three cycles of aAPC-based in vitro priming using CD8+ T cells derived from HLA-matched HBDs primed with (A) the HLA-B*15-restricted peptide HQKRPIPIKY (P4B*15) and (B) the HLA-B*07-restricted peptide RPIPIKYKAM (P1B*07) as well as from HLA-matched MYD88WT CLL patient (CLL-05) primed with (C) the HLA-B*07-restricted peptide RPIPIKYKAM (P1B*07): 1st column: tetramer staining of CD8+ T cells primed with the MYD88L265P-derived peptide; 2nd column: control staining with HLA-matched tetramer containing a non-relevant control peptide on CD8+ T cells derived from the same population as T cells depicted in the 1st column; 3rd column: ex vivo tetramer staining of CD8+ T cells. In vitro primings with HBD-derived PBMCs were performed in six (P1B*07) and three (P4B*15) independent replicates, respectively. For the in vitro priming with PBMCs of CLL patients two independent replicates were conducted. Abbreviations: neg., negative.
Mentions: To assess whether MYD88L265P-derived peptide-specific T cell responses can be induced from naive T cells in vitro, we isolated CD8+ T cells from six HLA-B*07+ and three HLA-B*15+ HBDs as well as from two HLA-B*07+MYD88WT CLL patients. We performed artificial antigen-presenting cell (aAPC)-based in vitro priming using the three HLA-B*07-restricted MYD88L265P-derived peptides RPIPIKYKAM (P1B*07), RPIPIKYKA (P2B*07) and SPGAHQKRPI (P3B*07) as well as the HLA-B*15-restricted peptide HQKRPIPIKY (P4B*15). Using HBD-derived CD8+ T cells, P4B*15-tetramer-positive CD8+ populations with frequencies of 1.50–10.68% of viable cells were detected in 2/3 HBDs after priming (Fig. 3A). For the HLA-B*07-restricted peptide P1B*07, we observed tetramer-positive CD8+ populations with frequencies of 0.47–14.04% of viable cells in 6/6 HBDs (Fig. 3B). For P2B*07, 1/3 (33%) HBDs showed a tetramer-positive CD8+ population with a frequency of 0.15% of viable cells (Fig. S1A). P3B*07-tetramer positive CD8+ populations with frequencies of 0.20–1.56% of viable cells were detected in 3/4 HBDs (Fig. S1B). Notably, after aAPC-based in vitro priming of CD8+ T cells from MYD88WT B*07+ CLL patients without previous T cell reactivity for P1B*07 (as detected by 12-day recall IFNγ ELISPOT assay and ex vivo tetramer staining), we observed in 1/2 patients a population of 0.40% P1B*07-specific CD8+ T cells within the viable cells (Fig. 3C). No tetramer-positive T cell populations >0.10% (>0.50%) were detectable in control stainings with an HLA-B*07 (HLA-B*15)-tetramer containing a control peptide. In ex vivo control stainings no tetramer-positive T cell populations >0.01% were detectable. Furthermore, Peper et al.31 demonstrated that T cell responses observed after three rounds of aAPC-based stimulations were mediated by in vitro primed naive T cells rather than by pre-existing memory T cells, as short-time stimulation of the same PBMC did not result in the detection of specific T cell populations. Collectively, all 4/4 (100%) refolded MYD88L265P-derived peptides thus are able to efficiently prime mutation-specific T cells in certain HLA contexts in HBDs as well as in CLL patients.Figure 3.

View Article: PubMed Central - PubMed

ABSTRACT

Genome sequencing has uncovered an array of recurring somatic mutations in different non-Hodgkin lymphoma (NHL) subtypes. If affecting protein-coding regions, such mutations may yield mutation-derived peptides that may be presented by HLA class I proteins and recognized by cytotoxic T cells. A recurring somatic and oncogenic driver mutation of the Toll-like receptor adaptor protein MYD88, Leu265Pro (L265P) was identified in up to 90% of different NHL subtype patients. We therefore screened the potential of MYD88L265P-derived peptides to elicit cytotoxic T cell responses as tumor-specific neoantigens. Based on in silico predictions, we identified potential MYD88L265P-containing HLA ligands for several HLA class I restrictions. A set of HLA class I MYD88L265P-derived ligands elicited specific cytotoxic T cell responses for HLA-B*07 and -B*15. These data highlight the potential of MYD88L265P mutation-specific peptide-based immunotherapy as a novel personalized treatment approach for patients with MYD88L265P+ NHLs that may complement pharmacological approaches targeting oncogenic MyD88 L265P signaling.

No MeSH data available.