Induction of primary NY-ESO-1 immunity: CD8+ T lymphocyte and antibody responses in peptide-vaccinated patients with NY-ESO-1+ cancers

Abstract

Cancer-testis antigen NY-ESO-1 is one of the most immunogenic tumor antigens defined to date. Spontaneous humoral and CD8+ T-cell responses to NY-ESO-1 are detected in 40-50% of patients with advanced NY-ESO-1-expressing tumors. A clinical trial was initiated to study the immunological effects of intradermal vaccination with 3 HLA-A2-binding NY-ESO-1 peptides in 12 patients with metastatic NY-ESO-1-expressing cancers. Seven patients were NY-ESO-1 serum antibody negative, and five patients were NY-ESO-1 serum antibody positive at the outset of the study. Primary peptide-specific CD8+ T-cell reactions and delayed-type hypersensitivity responses were generated in four of seven NY-ESO-1 antibody-negative patients. Induction of a specific CD8+ T-cell response to NY-ESO-1 in immunized antibody-negative patients was associated with disease stabilization and objective regression of single metastases. NY-ESO-1 antibody-positive patients did not develop significant changes in baseline NY-ESO-1-specific T-cell reactivity. However, stabilization of disease and regression of individual metastases were observed in three of five immunized patients. These results demonstrate that primary NY-ESO-1-specific CD8+ T-cell responses can be induced by intradermal immunization with NY-ESO-1 peptides, and that immunization with NY-ESO-1 may have the potential to alter the natural course of NY-ESO-1-expressing tumors.

Figure 1

Immunological and clinical effects of vaccination in NY-ESO-1 seronegative (A) and seropositive (B) patients. ELISPOT assays: number of IFN-γ-releasing CD8+ T lymphocytes per 2.5 × 10⁴ CD8+ T cells after presensitization with two NY-ESO-1 peptides (11 mer, SLLMWITQCFL; p157–167; 9 mer, SLLMWITQC; p157–165). No measurable CD8+ T-cell responses were found against the third NY-ESO-1 peptide used for immunization (QLSLLMWIT; p155–163) or against an irrelevant peptide (GILTVILGV; p29–37) used as a negative control in the assays (data not shown). ne, not evaluable because of high background reactivity. Cytotoxicity against peptide-pulsed T2 cells: +, >20% specific lysis above background. DTH reactions classified according to the area of redness and induration: −, no reaction; −/+, weak or questionable reaction; white circle, positive reaction >8 mm; red circle, inflammatory reaction >2 cm. ns, immediate nonspecific skin reaction after injection of 100% DMSO/peptide solution. The development of single metastases is documented according to the World Health Organization classification of response as described. Abbreviations: nd, not done; LU, lung; CNS, brain; Pl, pleura; SC, s.c.; LN, lymph node; LI, liver; P, peritoneum. *, The designation “progressive disease” (PD) was given because of an enlargement of the lesion. However, this enlargement was attributable to increasing necrosis of the s.c. mass. Additional cycles of vaccination were carried out in patients NW415 and NW886 (total number of cycles, five) and in patient NW924 (total number of cycles, four).

Figure 2

NY-ESO-1-specific cytotoxicity of CD8+ T lymphocytes of patient NW745 (lymphocytes obtained on day 127 of immunization) and patient NW903 (lymphocytes obtained on day 78 of immunization) after presensitization with the NY-ESO-1 nonamer (p157–165) (a) and the NY-ESO-1 11-mer (p157–167) (b) peptides. Target cells were T2 cells alone (white bars); T2 cells pulsed with the NY-ESO-1 9 mer (p157–165; gray bars); T2 cells pulsed with the NY-ESO-1 11 mer (p157–167; black bars); NY-ESO-1+, HLA-A2+ melanoma cell lines NW-MEL-38 (striped bars); SK-MEL-37 (checkered bars); and NY-ESO-1−, HLA-A2+ melanoma cell line NW-MEL-145 (crossed bars). The chromium-51 release assay is described in Materials and Methods. nd, not done.

Figure 3

Classification of DTH reactions induced by the NY-ESO-1 11-mer peptide (p165–167). Reaction patterns: −, no detectable redness or induration; −/+, weak or questionable skin reaction with redness <8 mm in diameter and without palpable induration; +, redness and induration >8 mm in diameter; ++, inflammatory reaction >2 cm in diameter.

Figure 4

(Upper) Development of NY-ESO-1 serum antibody during immunization of patient NW745 with NY-ESO-1 peptides as demonstrated by Western blot. NY-ESO-1 antibody was first detected on day 113 and showed an increasing titer with continued immunization. Serum dilution was 1:250. Co, blot with 0,25 μg of NY-ESO-1 recombinant “short” protein (14 kDa) and serum from a melanoma patient with NY-ESO-1 antibody. (Lower) Development of NY-ESO-1 CD8+ T cells during immunization of patient NW745 with NY-ESO-1 peptides as demonstrated by ELISPOT analysis. CD8+ T-cell reactivity was first detected on day 64 and increased in frequency with continued immunization. The CD8+ T-cell response preceded the development of NY-ESO-1 antibody in patient NW745.