Clonality analysis of pulmonary tumors by genome-wide copy number profiling

Abstract

Multiple tumors in patients are frequently diagnosed, either synchronous or metachronous. The distinction between a second primary and a metastasis is important for treatment. Chromosomal DNA copy number aberrations (CNA) patterns are highly unique to specific tumors. The aim of this study was to assess genome-wide CNA-patterns as method to identify clonally related tumors in a prospective cohort of patients with synchronous or metachronous tumors, with at least one intrapulmonary tumor. In total, 139 tumor pairs from 90 patients were examined: 35 synchronous and 104 metachronous pairs. Results of CNA were compared to histological type, clinicopathological methods (Martini-Melamed-classification (MM) and ACCP-2013-criteria), and, if available, EGFR- and KRAS-mutation analysis. CNA-results were clonal in 74 pairs (53%), non-clonal in 33 pairs (24%), and inconclusive in 32 pairs (23%). Histological similarity was found in 130 pairs (94%). Concordance between histology and conclusive CNA-results was 69% (74 of 107 pairs: 72 clonal and two non-clonal). In 31 of 103 pairs with similar histology, genetics revealed non-clonality. In two out of four pairs with non-matching histology, genetics revealed clonality. The subgroups of synchronous and metachronous pairs showed similar outcome for the comparison of histological versus CNA-results. MM-classification and ACCP-2013-criteria, applicable on 34 pairs, and CNA-results were concordant in 50% and 62% respectively. Concordance between mutation matching and conclusive CNA-results was 89% (8 of 9 pairs: six clonal and two non-clonal). Interestingly, in one patient both tumors had the same KRAS mutation, but the CNA result was non-clonal. In conclusion, although some concordance between histological comparison and CNA profiling is present, arguments exist to prefer extensive molecular testing to determine whether a second tumor is a metastasis or a second primary.

Fig 1. Example of a matching histologic pair with a non-clonal CNA-result.

A1 & A2: squamous cell carcinomas from a laryngeal tumor biopsy and a resected tumor in the left lower lobe, respectively (20x objective). B1 & B2: the corresponding CNA profiles. On the y axis is the log2 tumor to normal ratio and on the x axis the chromosomal position. MAD = median absolute deviation.

Fig 2. Example of a non-matching histologic pair with a clonal CNA-result.

A1: adenocarcinoma from a breast resection. A2: non-small cell carcinoma, favoring squamous cell carcinoma (IHC: p63 positive; TTF-1/PAS-D/Alcian blue negative) from a lung biopsy (20x objective). B1 & B2: the corresponding CNA profiles. On the y axis is the log2 tumor to normal ratio and on the x axis the chromosomal position. Gain and loss are positive and negative log2 ratio, respectively. MAD = median absolute deviation. All array data are available in the Gene Expression Omnibus database, under accession number GSE87058. Mutation analysis: not performed on both samples. Martini-Melamed and ACCP-2013: not applicable. Follow-up: no sign of metastasis after 79 months.