Melanoma antigen recognition by tumour-infiltrating T lymphocytes (TIL): effect of differential expression of Melan-A/MART-1

Abstract

SUMMARYWe have isolated, from an individual patient with metastatic melanoma, a series of eight TIL clones capable of lysing autologous melanoma cell targets. Six of the eight clones expressed TCRAV2S1 and lysed targets expressing HLA-A2 and the Melan-A/MART-1 peptide: AAGIGILTV. Polymerase chain reaction-single stranded conformational polymorphism (PCR-SSCP) analysis showed that the Melan-A/ MART-1-specific clones were predominant in the bulk culture prior to cloning. However, the tumour progressed in vivo even in the presence of these tumour cell-lytic clones. Using the antiMelan-A/MART-1 MoAb (A-103), we noted that Melan-A/MART-1 expression on three melanoma cell lines varied considerably during in vitro culture, in the absence of T cell immunoselection, relative to cell density. Tumour cells which spontaneously decreased Melan-A/MART-1 expression were less susceptible to specific TIL lysis. Melan-A/MART-1 expression and susceptibility to lysis increased in cells cultured at lower density. These data suggest that modulation of tumour antigen may account for tumour progression in the presence of tumour cell-lytic T lymphocytes. The observations suggest a possible explanation for the common finding of Melan-A/MART-1-specific lytic TIL in clinically progressing melanomas, as well as a possible pathway for therapeutic intervention.

A Novel Autocrine Pathway of Tumor Escape from Immune Recognition: Melanoma Cell Lines Produce a Soluble Protein That Diminishes Expression of the Gene Encoding the Melanocyte Lineage Melan-A/MART-1 Antigen Through Down-Modulation of Its Promoter

“…MU tumor cells were established at 1 ϫ 10 5 /ml in 1-ml aliquots in individual wells of 24-well culture plates, as previously described (35), which were then incubated at 37°C for between 1 and 7 days, before harvesting cells for fluorescent staining and cytotoxicity assays, as described below. Although production of MASA was constitutive, titers increased as long as tumor cells remained viable, generally proving most potent in supernatants collected after 3-4 days.…”

“…Previously, we have shown that spontaneous down-modulation of Melan-A/ MART-1 expression was observed when melanoma cells were grown at high density, whereas this Ag continued to be expressed by tumor cells cultured at low density, and could be rejuvenated by low density culture (35). The reduced expression of Melan-A/ MART-1 protein at high density correlated with decreased T cell recognition (35). These observations suggested that expression of Melan-A/MART-1 could be down-modulated in vitro during melanoma cell growth through an autocrine pathway.…”

“…These observations have led numerous investigators to use T cell-based immunotherapy in attempts to manage this highly malignant tumor (8 -11). This is illustrated by the intense ongoing efforts to vaccinate melanoma patients with the immunodominant HLA-A2-restricted Melan-A/MART-1 peptide (aa [27][28][29][30][31][32][33][34][35] (5,6,12). Although it has been demonstrated that such a peptide-based vaccination has led to enhanced cell-mediated immunity against targets expressing this peptide, the escape from T cell recognition of Melan-A/ MART-1-negative tumors has limited the efficacy of therapeutic intervention targeting this Ag (13)(14)(15)(16)(17).…”

A Screening Assay to Identify Agents That Enhance T-Cell Recognition of Human Melanomas

“…However, with each new advance comes recognition of new hurdles to therapeutic success. As our own group noted over a decade ago, 73 one of the Achilles' Heels of targeted immunotherapy is the need for the tumor to express the target antigen. Indeed, we and others have shown that tumors have a survival advantage if they express lower levels of both target antigens and the presenting HLA alleles.…”

Multiple mechanisms underlie defective recognition of melanoma cells cultured in three-dimensional architectures by antigen-specific cytotoxic T lymphocytes

“…Previous work suggests that high cell density in monolayer cultures could affect TAA expression (Ramirez-Montagut et al, 2000). In an attempt to clarify molecular mechanisms underlying our observations, we addressed antigen expression in target cells.…”