Array CGH on Archival Fixed Paraffin Embedded Tissues on a Series of Malignant Mesothelioma
M Paciencia, M Brevet, M Figeac, V Abonnet, N Le Stang, F Galateau-Salle. CHU, Caen, France; Institute of Cancer Research, Lille, France
Background: Array based comparative genomic hybridization (aCGH) is a promising tool for molecular subclassification in malignant mesothelioma (MM). Because of the interest to perform retrospective studies applied to a specific focus on the slide, we aim to evaluate the reliability of archival FFPE tissues on a series of MM cases.
Design: Total DNA was extracted from 15 frozen and 9 FFPE tissue samples retrieved from the MESOPATH center. FFPE were selected up to 10 years, with time of fixation up to 7 days. aCGH was performed on a 180 K pangenomic Agilent array yelding a 13kb overall median probe spacing. The sensitivity of aCGH was investigated using serially diluted tumor DNA specimens. The minimum percentage of cells and amount of DNA was also evaluated.
Results: Focusing on FFPE samples we tested ULS and Klenow method for aCGH. Noticeably the Klenow one was the best. We also obtain better quality increasing the amount of recommended DNA. One of the crucial points was to co-hybridize same quality DNA. Therefore we extracted tumoral and non tumoral DNA from FFPE. Highly sensitive detection required a previous dissection from the H&E to obtain at least 70% of tumor cells. Among the FFPE tumor samples 88% yielded a high quality aCGH showing deleted and amplified fragments ranging from 3.5 Kb up to a full chromosome deletion. Among the many interesting detected aberration, we found a gain of JUN (3.5kb) and an amplification of FOS(37.5kb amplified fragment). The homozygous deletion of 9p21 was observed in 66% of the cases correlated with loss of immunoexpression of p16 in 100%.
Conclusions: Long term archival FFPE is suitable for aCGH analysis if the DNA quantity is adequate with at least 70% of tumor cells.
Tuesday, March 23, 2010 1:00 PM
Poster Session IV # 252, Tuesday Afternoon