Detection of circulating tumor lysate–reactive CD4⁺ T cells in melanoma patients

Purpose: We wanted to study whether an allogeneic melanoma lysate would be a feasible stimulatory antigen source for detection of a peripheral CD4⁺ T-cell immune response in patients with medically untreated malignant melanoma. The lysate was produced from a melanoma cell line (FM3.29) which expresses high amounts of melanoma antigens. Methods: Fresh peripheral blood was incubated with and without lysate for 6 h in the presence of anti-CD28/anti-CD49d MoAb (for costimulation). After flow cytometric estimation of the frequency of CD69⁺/IFN-γ⁺ cells in the CD4⁺ population, the response to lysate was calculated as the difference between the number of activated IFN-γ-producing CD4⁺ cells in the lysate-stimulated and the nonstimulated sample. Results: An immune response to lysate was observed in blood samples from 11 of 15 patients (73%) with metastatic melanoma. A weak response was found in 1 of 4 patients radically operated for localized disease, whereas no responders were seen among 7 healthy donors. The fraction of circulating lysate-activated T cells ranged from 0.0037% to 0.080% of the CD4⁺ population. A negative result of the assay was found occasionally, especially in donors with high background levels of spontaneous IFN-γ production, indicating an inhibitory effect of the lysate. Conclusions: This method for detection of a peripheral T-cell immune response in melanoma patients has several advantages for clinical use. The tumor lysate preparations may contain large numbers of stimulating antigens (known, as well as unknown) and are easily prepared and handled. Potentially, the assay might be useful as a diagnostic tool, a marker of residual or recurrent disease, a prognostic factor, or a predictor or monitor of the effect of antineoplastic therapy including immune-modulating therapy.