The Degree of Segmental Aneuploidy Measured by Total Copy Number Abnormalities Predicts Survival and Recurrence in Superficial Gastroesophageal Adenocarcinoma

Abstract Background: Prognostic biomarkers are needed for superficial gastroesophageal adenocarcinoma (EAC) to predict clinical outcomes and select therapy. Although recurrent mutations have been characterized in EAC, little is known about their clinical and prognostic significance. Aneuploidy is predictive of clinical outcome in many malignancies but has not been evaluated in superficial EAC

The degree of segmental aneuploidy measured by total copy number abnormalities predicts survival and recurrence in superficial gastroesophageal adenocarcinoma

Background: Prognostic biomarkers are needed for superficial gastroesophageal adenocarcinoma (EAC) to predict clinical outcomes and select therapy. Although recurrent mutations have been characterized in EAC, little is known about their clinical and prognostic significance. Aneuploidy is predictive of clinical outcome in many malignancies but has not been evaluated in superficial EAC. Methods: We quantified copy number changes in 41 superficial EAC using Affymetrix SNP 6.0 arrays. We identified recurrent chromosomal gains and losses and calculated the total copy number abnormality (CNA) count for each tumor as a measure of aneuploidy. We correlated CNA count with overall survival and time to first recurrence in univariate and multivariate analyses. Results: Recurrent segmental gains and losses involved multiple genes, including: HER2, EGFR, MET, CDK6, KRAS (recurrent gains); and FHIT, WWOX, CDKN2A/B, SMAD4, RUNX1 (recurrent losses). There was a 40-fold variation in CNA count across all cases. Tumors with the lowest and highest quartile CNA count had significantly better overall survival (p = 0.032) and time to first recurrence (p = 0.010) compared to those with intermediate CNA counts. These associations persisted when controlling for other prognostic variables. Significance: SNP arrays facilitate the assessment of recurrent chromosomal gain and loss and allow high resolution, quantitative assessment of segmental aneuploidy (total CNA count). The non-monotonic association of segmental aneuploidy with survival has been described in other tumors. The degree of aneuploidy is a promising prognostic biomarker in a potentially curable form of EAC. © 2014 Davison et al

Schematic Illustration of Copy Number Abnormalities in Two Tumors Representing High and Low CNA Count and a Spectrum of Genomic Complexity.

<p>For both A and B, the horizontal axis represents position of segments along chromosome 7 (in Mb) and the vertical axis represents estimated copy number (truncated at 16). (A) Tumor 2515 (dark grey) has a large number of independent CNAs on chromosome 7 (N = 56), including a complex copy number gains at ∼55 Mb (region of EGFR) as well as other gains and losses throughout the chromosome. (B) By contrast, tumor 2634 (light grey) has few independent copy number changes (N = 2, gain at ∼39 Mb and loss at ∼97.4 Mb). For this illustration, copy numbers in the normal range of 1.7–2.3 were assigned a value of 2.</p

Representative EGFR and MYC FISH Results.

<p>FISH to determine EGFR and MYC copy number. Gene specific probes are labeled red (EGFR and MYC) while corresponding centromere probes (CEP7 and CEP8, respectively) are labeled green. (A) Tumor cells with a near normal 1∶1 ratio of EGFR/CEP7 and approximately 2 signals from each probe per cell. (B) High level EGFR amplification. EGFR amplification was detected as clusters of numerous red fluorescent signals which is a characteristic pattern caused by high EGFR copy number (see reference <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0079079#pone.0079079-Marx1" target="_blank">[57]</a>) with variable chromosome 7 centromere copy number. (C) Tumor cells with a near normal MYC and centromere 8 copy number. (D) High level MYC amplification characterized by numerous MYC signals per cell and a high MYC/CEP8 ratio.</p

Total CNA Count in Relation to Potential Pathologic Prognostic Variables.

<p>For categorical variables, none of the differences between groups was statistically significant when comparing across the three groups nor in pairwise comparisons with low CNA count (p-value>0.05, Fisher's exact test).</p><p>The difference in tumor size and mean number of resected lymph nodes was not significant across the three CNA count groups (p-value>0.05, Kruskal-Wallis one way ANOVA), nor in pairwise comparisons (p-value>0.05, Mann-Whitney U Test).</p