Development of a real-time reverse transcription polymerase chain reaction assay for c-myc expression that allows the identification of a subset of c-myc+ diffuse large B-cell lymphoma.
Absence of a reliable method for determining the level of c-myc expression has impeded the analysis of its biological and clinical relevance in tumors. We have standardized the conditions for a real-time reverse transcription polymerase chain reaction analysis for c-myc expression, including the selection of an endogenous reference (18S rRNA), the adequate number of measurements for each sample (2 cDNA in triplicate), and suitable controls for determining inter- and intrarun variability (standard curve and calibrator). Subsequently, in a series of 56 non-Hodgkin's lymphomas, we analyzed the expression of c-myc mRNA, using real-time reverse transcription polymerase chain reaction, and of other functionally related proteins (bcl-6, p27, cyclin D3, and p53). As expected, all eight Burkitt's lymphoma cases analyzed had high levels of c-myc mRNA expression compared with that observed in reactive lymphoid tissue. There was a wider range of expression in diffuse large B-cell lymphoma, with 30% (15 of 48) of cases overexpressing c-myc. This overexpression was largely independent of c-myc translocations (4 of 5), as demonstrated by fluorescence in situ hybridization. In this large B-cell lymphoma series, a high level of c-myc expression predicted lower survival probability, irrespectively of the International Prognostic Index risk group classification. A slightly increased frequency of p53 inactivation was observed in the cases with c-myc overexpression, which suggests a growth advantage in lymphomas with concurrent deregulation of c-myc and p53. In addition, a moderate increase in bcl-6 protein expression was observed in the c-myc-positive cases, suggesting the existence of a complex interrelationship between these two genes. These findings suggest that c-myc may play a relevant role in the pathogenesis of a subset of large B-cell lymphoma and suggest the existence of additional regulatory mechanisms of c-myc expression to c-myc rearrangements.
Sáez
Ana-Isabel
AI
Centro Nacional de Investigaciones Oncológicas, Toledo, Spain.
Artiga
María-Jesús
MJ
Romero
Cristina
C
Rodríguez
Sandra
S
Cigudosa
Juan-Cruz
JC
Pérez-Rosado
Alberto
A
Fernández
Isabel
I
Sánchez-Beato
Margarita
M
Sánchez
Esther
E
Mollejo
Manuela
M
Piris
Miguel A
MA
D001402:B-Lymphocytes / Q000473:pathology*; D014408:Biomarkers, Tumor / Q000032:analysis; D002051:Burkitt Lymphoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D018076:DNA, Complementary / Q000032:analysis; D004273:DNA, Neoplasm / Q000032:analysis*; D016259:Genes, myc / Q000235:genetics*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D016403:Lymphoma, Large B-Cell, Diffuse / Q000235:genetics* / Q000378:metabolism / Q000401:mortality / Q000473:pathology; D009363:Neoplasm Proteins / Q000032:analysis; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics* / Q000378:metabolism; D019310:Pseudolymphoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D012334:RNA, Neoplasm / Q000032:analysis; D012335:RNA, Ribosomal / Q000032:analysis; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D015996:Survival Rate
D014408:Biomarkers, Tumor; D018076:DNA, Complementary; D004273:DNA, Neoplasm; D009363:Neoplasm Proteins; D016271:Proto-Oncogene Proteins c-myc; D012334:RNA, Neoplasm; D012335:RNA, Ribosomal