Evaluation of CD8+ T-cell frequencies by the Elispot assay in healthy individuals and in patients with metastatic melanoma immunized with tyrosinase peptide

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Abstract
The lack of reproducible, quantitative assays for T‐cell responses has been a limitation in the development of cancer vaccines to elicit T‐cell immunity. We utilized the Elispot assay, which allows a quantitative and functional assessment of T cells directed against specific peptides after only brief in vitro incubations. CD8+ T‐cell reactivity was determined with an interferon (IFN)‐γElispot assay detecting T cells at the single cell level that secrete IFN‐γ. We studied both healthy individuals and patients with melanoma. Healthy HLA‐A*0201‐positive individuals showed a similar mean frequency of CD8+ cells recognizing a tyrosinase peptide, YMDGTMSQV, when compared with melanoma patients prior to immunization. The frequencies of CD8+ cells recognizing the tyrosinase peptide remained relatively constant over time in healthy individuals. Nine HLA‐A*0201‐positive patients with stage IV metastatic melanoma were immunized intradermally with the tyrosinase peptide together with the immune adjuvant QS‐21 in a peptide dose escalation study with 3 patients per dose group. Two patients demonstrated a significant increase in the frequency of CD8+ cells recognizing the tyrosinase peptide during the course of immunization, from approx. 1/16,000 CD8+ T cells to approx. 1/4,000 in the first patient and from approx. 1/14,000 to approx. 1/2,000 in the second patient. These results demonstrate that modest expansion of peptide‐specific CD8+ T cells can be generated in vivo by immunization with peptide plus QS‐21 in at least a subset of patients with melanoma. Int. J. Cancer 87:391–398, 2000.

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