Mutations of the β₂‐microglobulin gene result in a lack of HLA class I molecules on melanoma cells of two patients immunized with MAGE peptides

Abstract

Mutations have been identified in the beta2-microglobulin gene of tumor cells of two metastatic melanoma patients who received immunizations with MAGE peptides. One mutation abolishes the start codon whereas the other introduces a premature stop codon. The second beta2-microglobulin allele of both tumors appears to be lost on the basis of sequence data and loss of microsatellite heterozygosity. The lack of beta2-microglobulin gene product results in the absence of HLA class I antigens on the surface of the tumor cells. This may explain why the tumors of both patients progressed despite the immunization treatment and shows the necessity of analyzing in depth the antigen presentation capability of the tumor cells for the interpretation of clinical trials involving anti-tumor vaccination.

Complete loss of HLA class I antigen expression on melanoma cells: A result of successive mutational events

“…9,13 This patient, however, demonstrates that evasion strategies can develop before the clinical application of any therapeutic regimen. Therefore, the possibility of early development of HLAloss tumor variants should be taken into consideration when immunologic treatment strategies are designed.…”

“…Paraffin-embedded tumor tissue was analyzed for b2m, HLA class I and HLA-DR expression using the mouse MAbs GRH1 (recognizing free and HLA class I heavy chain-associated b2m), 9,15 HC-10 (binding to free HLA-A, -B and -C heavy chains) 9 and GRB1 (against HLA-DR), 9,15 respectively. Staining was performed as described previously.…”

Immune Selection of Hot-Spot β2-Microglobulin Gene Mutations, HLA-A2 Allospecificity Loss, and Antigen-Processing Machinery Component Down-Regulation in Melanoma Cells Derived from Recurrent Metastases following Immunotherapy

“…The mutation harbored by the cell line 1074MEL (ATG to ATA) is novel, because it is different from two other initiation codon mutations identified in the Burkitt lymphoma cell line Daudi (ATG to ATC) (34) and in the melanoma cell line LB1622-MEL (ATG to AAG) (35). The nonsense mutation identified in the cell line 1174MEL (TCA to TGA), which introduces a premature stop in codon 31 of exon 2 of the ␤ 2 m gene (31 Ser3 Stop ), is identical with the recently described mutation in the melanoma cell line BB74-MEL (35). It is noteworthy that both LB1622-MEL and BB74-MEL cell lines were derived from lesions in patients who had undergone T cellbased immunotherapy, suggesting that these cells have been exposed to strong T cell selective pressure.…”

“…To determine the functionality of APM in melanoma cells analyzed in this study, we assessed HLA-A*0201-MART1 [27][28][29][30][31][32][33][34][35] …”

“…The monomeric HLA-A*0201-MART1 27-35 -specific scFv 8.3 was isolated from the human synthetic V H ϩ V L scFv library (Griffin.1 library) by panning with HLA-A*0201-MART1 [27][28][29][30][31][32][33][34][35] peptide complexes (M. Campoli, unpublished results). To construct tetrameric scFv 8.3, monomeric scFv 8.3 were engineered with a C-terminal BirA biotinylation tag (scFv 8.3-BirA) and expressed from ampicillin-resistant bacterial colonies, as previously described (24 -26).…”