Different pattern of allelic loss in Epstein-Barr virus-positive gastric cancer with emphasis on the p53 tumor suppressor pathway

Abstract

Both Helicobacter pylori (HP) and Epstein-Barr virus (EBV) have been implicated in carcinogenesis of the stomach. Fifty-seven gastric carcinomas were tested for microsatellite instability and allelic loss at several tumor suppressor loci using 21 polymorphic microsatellite markers. Furthermore, immunohistochemistry for p53 and DPC4/SMAD4 was performed. Results were analyzed according to HP and EBV status of the tumors, as assessed by immunohistochemistry and RNA in situ hybridization, respectively. Fractional allelic loss was lower in EBV-positive carcinomas (n = 15) when compared to EBV-negative carcinomas (P < 0.001). EBV positivity was inversely associated with allelic loss at specific markers on chromosomal arms 5q (APC), 17p (TP53), and 18q (DPC4/SMAD4). Allelic loss at the TP53 locus was not encountered in EBV-positive carcinomas, but occurred in 51% of EBV-negative carcinomas (P < 0.005). Moreover, none of the EBV-positive carcinomas showed unequivocal p53 immunopositivity in contrast to 39% of the EBV-negative carcinomas (P < 0.01). EBV-status was not related to microsatellite instability. There was no correlation between HP-status and any of the molecular alterations tested. In conclusion, EBV-positive gastric carcinomas follow a distinct pathogenesis at the molecular level, in which p53 is not, or differently inactivated.

Figure 1.

Representative examples of LOH. Electropherograms of labeled PCR products of paired tumor (T) and normal (N) DNA are shown. The tumor number and the allelic imbalance factor are shown at the top of each frame and the lost alleles are marked by asterisks (see Materials and Methods for calculation of the allelic imbalance factor and scoring of LOH). The tested marker is indicated at the bottom of each frame and individual alleles are marked by arrows.

Figure 2.

Frequency distribution of allelic loss of EBV-positive and EBV-negative gastric carcinomas at the chromosomal arms tested. Asterisks indicate statistically significant differences at specific chromosomal arms.

Figure 3.

Relation of EBV and p53 alterations, illustrated by two representative tumors (PA57 and SE8). A: Epstein-Barr virus was detected by in situ hybridization for EBER1 nuclear RNA transcripts. PA57 is negative whereas SE8 shows strong nuclear positivity in the vast majority of tumor cells. Immunohistochemistry for p53 is strongly positive in PA57 but negative in SE8. B: Electropherograms of labeled PCR products of paired tumor (T) and normal (N) DNA. Two microsatellite markers at the TP53 locus are shown (p53 Alu and TP53). Both PA57 and SE8 are informative for each marker. The individual alleles are indicated by arrows. PA57 (EBV-negative, p53-immunopositive) shows LOH at both markers. The lost alleles are marked by asterisks. SE8 (EBV-positive, p53-immunonegative) has retained both alleles in the tumor.