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Understanding CD8 + T-cell responses toward the native and alternate HLA-A * 02:01-restricted WT1 epitope

View Article: PubMed Central - PubMed

ABSTRACT

Ry: The Wilms' tumor 1 (WT1) antigen is expressed in solid and hematological malignancies, but not healthy tissues, making it a promising target for cancer immunotherapies. Immunodominant WT1 epitopes, the native HLA-A2/WT1126-134 (MFPNAPYL) (HLA-A2/RMFPNAPYL epitope (WT1A)) and its modified variant MFPNAPYL (HLA-A2/YMFPNAPYL epitope (WT1B)), can induce WT1-specific CD8+ T cells, although WT1B is more stably bound to HLA-A*02:01. Here, to further determine the benefits of those two targets, we assessed the naive precursor frequencies; immunogenicity and cross-reactivity of CD8+ T cells directed toward these two WT1 epitopes. Ex vivo naive WT1A- and WT1B-specific CD8+ T cells were detected in healthy HLA-A*02:01+ individuals with comparable precursor frequencies (1 in 105–106) to other naive CD8+ T-cell pools (for example, A2/HIV-Gag77-85), but as expected, ~100 × lower than those found in memory populations (influenza, A2/M158-66; EBV, A2/BMLF1280-288). Importantly, only WT1A-specific naive precursors were detected in HLA-A2.1 mice. To further assess the immunogenicity and recruitment of CD8+ T cells responding to WT1A and WT1B, we immunized HLA-A2.1 mice with either peptide. WT1A immunization elicited numerically higher CD8+ T-cell responses to the native tumor epitope following re-stimulation, although both regimens produced functionally similar responses toward WT1A via cytokine analysis and CD107a expression. Interestingly, however, WT1B immunization generated cross-reactive CD8+ T-cell responses to WT1A and could be further expanded by WT1A peptide revealing two distinct populations of single- and cross-reactive WT1A+CD8+ T cells with unique T-cell receptor-αβ gene signatures. Therefore, although both epitopes are immunogenic, the clinical benefits of WT1B vaccination remains debatable and perhaps both peptides may have separate clinical benefits as treatment targets.

No MeSH data available.


Expansion of cross-reactive WT1A-specific CD8+ T cells. In one experiment, splenocytes from mice immunized with WT1A (n=4), WT1B (n=4) or an irrelevant HPV peptide (n=3) were cultured with WT1A, WT1B or SV4 peptides in the presence of IL-2. After 6 days of culture, cells were costained with WT1A-tetramer-PE and WT1B-tetramer APC (co-stain 3). Representative tetramer co-staining profiles (a) of individual mice cultured with WT1A and WT1B peptides from both WT1A/WT1B-vaccinated groups. Bar graph (b) showing absolute numbers of expanded single-tetramer+ and dual-tetramer+ CD8+ T cells (mean±s.d. are shown).
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fig5: Expansion of cross-reactive WT1A-specific CD8+ T cells. In one experiment, splenocytes from mice immunized with WT1A (n=4), WT1B (n=4) or an irrelevant HPV peptide (n=3) were cultured with WT1A, WT1B or SV4 peptides in the presence of IL-2. After 6 days of culture, cells were costained with WT1A-tetramer-PE and WT1B-tetramer APC (co-stain 3). Representative tetramer co-staining profiles (a) of individual mice cultured with WT1A and WT1B peptides from both WT1A/WT1B-vaccinated groups. Bar graph (b) showing absolute numbers of expanded single-tetramer+ and dual-tetramer+ CD8+ T cells (mean±s.d. are shown).

Mentions: WT1A peptide vaccination trials in AML and MDS patients have found that WT1A-specific CD8+ T-cell responses are short-lived and expand poorly following in vitro culture or repeated vaccination.19, 20 In addition, it is very difficult to directly compare the de novo efficacy of different vaccine formulations within the same individual. To compare the proliferative capacity of CD8+ T cells following WT1A versus WT1B vaccination in our mouse system, 2–3 × 107 splenocytes per mouse from each group were expanded for 6 days in the presence of WT1A, WT1B or the control SV4 peptide, before determining tetramer specificity based on co-stain 3 (WT1A-tetramer PE versus WT1B-tetramer APC) (Figure 5a). The cultures generated with the SV4 peptide served as control cultures from the vaccination groups. In addition, cells from the HPV-E7/CpG vaccination group were stimulated with the WT1A and WT1B peptide and as expected, no expansion was detected confirming the absence of nonspecific expansion by these peptides.


Understanding CD8 + T-cell responses toward the native and alternate HLA-A * 02:01-restricted WT1 epitope
Expansion of cross-reactive WT1A-specific CD8+ T cells. In one experiment, splenocytes from mice immunized with WT1A (n=4), WT1B (n=4) or an irrelevant HPV peptide (n=3) were cultured with WT1A, WT1B or SV4 peptides in the presence of IL-2. After 6 days of culture, cells were costained with WT1A-tetramer-PE and WT1B-tetramer APC (co-stain 3). Representative tetramer co-staining profiles (a) of individual mice cultured with WT1A and WT1B peptides from both WT1A/WT1B-vaccinated groups. Bar graph (b) showing absolute numbers of expanded single-tetramer+ and dual-tetramer+ CD8+ T cells (mean±s.d. are shown).
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC5382434&req=5

fig5: Expansion of cross-reactive WT1A-specific CD8+ T cells. In one experiment, splenocytes from mice immunized with WT1A (n=4), WT1B (n=4) or an irrelevant HPV peptide (n=3) were cultured with WT1A, WT1B or SV4 peptides in the presence of IL-2. After 6 days of culture, cells were costained with WT1A-tetramer-PE and WT1B-tetramer APC (co-stain 3). Representative tetramer co-staining profiles (a) of individual mice cultured with WT1A and WT1B peptides from both WT1A/WT1B-vaccinated groups. Bar graph (b) showing absolute numbers of expanded single-tetramer+ and dual-tetramer+ CD8+ T cells (mean±s.d. are shown).
Mentions: WT1A peptide vaccination trials in AML and MDS patients have found that WT1A-specific CD8+ T-cell responses are short-lived and expand poorly following in vitro culture or repeated vaccination.19, 20 In addition, it is very difficult to directly compare the de novo efficacy of different vaccine formulations within the same individual. To compare the proliferative capacity of CD8+ T cells following WT1A versus WT1B vaccination in our mouse system, 2–3 × 107 splenocytes per mouse from each group were expanded for 6 days in the presence of WT1A, WT1B or the control SV4 peptide, before determining tetramer specificity based on co-stain 3 (WT1A-tetramer PE versus WT1B-tetramer APC) (Figure 5a). The cultures generated with the SV4 peptide served as control cultures from the vaccination groups. In addition, cells from the HPV-E7/CpG vaccination group were stimulated with the WT1A and WT1B peptide and as expected, no expansion was detected confirming the absence of nonspecific expansion by these peptides.

View Article: PubMed Central - PubMed

ABSTRACT

Ry: The Wilms' tumor 1 (WT1) antigen is expressed in solid and hematological malignancies, but not healthy tissues, making it a promising target for cancer immunotherapies. Immunodominant WT1 epitopes, the native HLA-A2/WT1126-134 (MFPNAPYL) (HLA-A2/RMFPNAPYL epitope (WT1A)) and its modified variant MFPNAPYL (HLA-A2/YMFPNAPYL epitope (WT1B)), can induce WT1-specific CD8+ T cells, although WT1B is more stably bound to HLA-A*02:01. Here, to further determine the benefits of those two targets, we assessed the naive precursor frequencies; immunogenicity and cross-reactivity of CD8+ T cells directed toward these two WT1 epitopes. Ex vivo naive WT1A- and WT1B-specific CD8+ T cells were detected in healthy HLA-A*02:01+ individuals with comparable precursor frequencies (1 in 105–106) to other naive CD8+ T-cell pools (for example, A2/HIV-Gag77-85), but as expected, ~100 × lower than those found in memory populations (influenza, A2/M158-66; EBV, A2/BMLF1280-288). Importantly, only WT1A-specific naive precursors were detected in HLA-A2.1 mice. To further assess the immunogenicity and recruitment of CD8+ T cells responding to WT1A and WT1B, we immunized HLA-A2.1 mice with either peptide. WT1A immunization elicited numerically higher CD8+ T-cell responses to the native tumor epitope following re-stimulation, although both regimens produced functionally similar responses toward WT1A via cytokine analysis and CD107a expression. Interestingly, however, WT1B immunization generated cross-reactive CD8+ T-cell responses to WT1A and could be further expanded by WT1A peptide revealing two distinct populations of single- and cross-reactive WT1A+CD8+ T cells with unique T-cell receptor-αβ gene signatures. Therefore, although both epitopes are immunogenic, the clinical benefits of WT1B vaccination remains debatable and perhaps both peptides may have separate clinical benefits as treatment targets.

No MeSH data available.