Clonality Analysis of Multifocal Lung Cancers by Loss of Heterozygosity, P53 Gene Sequencing and X Chromosome Inactivation Studies
X Wang, M Wang, FW Abdul-Karim, GT MacLennan, JN Eble, TD Jones, F Olobatuyi, R Eisenberg, OW Cummings, A Lopez-Beltran, R Montironi, S Zheng, D Bratman, DD Davidson, L Cheng. Indiana University, Indianapolis; Case Western Reserve University, Cleveland; Cordoba University, Cordoba, Spain; Polytechnic University of Marche Region, Ancona, Italy
Background: The incidence of multifocality of lung cancer ranges from 0.2% to 2%. The knowledge of clonality of multiple anatomically separate, but histologically similar lung tumors is limited. Detailed characterization of genetic changes may provide insights into the mechanisms of multifocality of this malignancy.
Design: We analyzed 70 lung tumors from 30 patients (23 females, 7 males) (26 patients with non-small cell carcinomas and 4 patients with carcinoid/atypical carcinoid tumors) who underwent surgical resection for lung epithelial tumors. All patients had multiple tumors (two to five) involving one or both lungs. Genomic DNA was prepared from paraffin-embedded tissue sections using laser capture microdissection. Loss of heterozygosity (LOH) study was performed using a panel of microsatellite markers at 3p14-21 (D3S1766), 4q33-34 (D4S408), 9p21 (IFNA, D9S171), 11q13 (D11S970) and 17p13.1 (TP53). Exons 5, 7 and 8 of p53 gene were screened for mutations by direct DNA sequencing. X-chromosome inactivation was methylation and PCR based analysis.
Results: All thirty cases showed loss of heterozygosity in at least one of the six polymorphic microsatellite markers (ranging from one to four markers). Completely concordant LOH patterns between synchronous or metachronous cancers in individual patients were seen in 26 of 30 informative cases (87%). The identical p53 point mutations were present in 8 of 10 cases, who showed p53 mutation by sequencing. Nineteen of 23 (83%) informative cases showed identical X chromosome inactivation pattern. With the combination of LOH studies, p53 gene mutation screening and X chromosome inactivation analyses, 23 of 30 (77%) cases demonstrated identical genetic changes, consistent with monoclonal origin among separate tumors.
Conclusions: Our data indicate that the great majority of multifocal lung cancers have a common clonal origin, and that multifocality in lung cancer represents local and regional intrapulmonary metastasis.
Monday, March 9, 2009 1:00 PM
Poster Session II # 226, Monday Afternoon