Steenaard Rebecca V, Feelders Richard A, Dogan Fadime, van Koetsveld Peter M, Creemers Sara G, Ettaieb Madeleine H T, van Kemenade Folkert J, Haak Harm R, Hofland Leo J
Department of Internal Medicine, Division of Endocrinology, Erasmus Medical Center, 3015 CN Rotterdam, The Netherlands.
Department of Internal Medicine, Máxima MC, 5504 DB Veldhoven, The Netherlands.
Biomedicines. 2023 Jul 17;11(7):2013. doi: 10.3390/biomedicines11072013.
The differentiation between benign and malignant adrenocortical tumors based on pathological assessment can be difficult. We present a series of 17 patients with unclear malignant tumors, of whom six had recurrent or metastatic disease. The assessment of the methylation pattern of insulin-like growth factor 2 () regulatory regions in fresh frozen material has shown to be valuable in determining the malignancy of adrenocortical tumors, although this has not been elaborately tested in unclear malignant tumors. Since fresh frozen tissue was only available in six of the patients, we determined the feasibility of using formalin-fixed paraffin-embedded (FFPE) tissue for this method. We isolated DNA from FFPE tissue and matched the fresh frozen tissue of three patients with adrenocortical carcinoma. Methylation patterns of regulatory regions were determined by pyrosequencing using different amounts of bisulfite-converted DNA (5 ng, 20 ng, 40 ng). Compared to fresh frozen tissue, FFPE tissue had a higher failure rate (fresh frozen 0%; FFPE 18.5%) and poor-to-moderate replicability (fresh frozen rho = 0.89-0.99, median variation 1.6%; FFPE rho = -0.09-0.85, median variation 7.7%). There was only a poor-to-moderate correlation between results from fresh frozen and FFPE tissue (rho = -0.28-0.70, median variation 13.2%). In conclusion, FFPE tissue is not suitable for determining the methylation score in patients with an unclear malignant adrenocortical tumor using the currently used method. We, therefore, recommend fresh frozen storage of resection material for diagnostic and biobank purposes.
基于病理评估来区分肾上腺皮质良性和恶性肿瘤可能具有挑战性。我们报告了17例恶性肿瘤情况不明的患者，其中6例出现复发或转移。对新鲜冷冻材料中胰岛素样生长因子2（IGF2）调控区域甲基化模式的评估已证明在确定肾上腺皮质肿瘤的恶性程度方面具有价值，尽管在恶性情况不明的肿瘤中尚未对此进行详尽测试。由于仅6例患者有新鲜冷冻组织，我们确定了使用福尔马林固定石蜡包埋（FFPE）组织进行该方法的可行性。我们从FFPE组织中分离DNA，并与3例肾上腺皮质癌患者的新鲜冷冻组织进行匹配。使用不同量的亚硫酸氢盐转化DNA（5 ng、20 ng、40 ng）通过焦磷酸测序确定IGF2调控区域的甲基化模式。与新鲜冷冻组织相比，FFPE组织的失败率更高（新鲜冷冻组织为0%；FFPE组织为18.5%），且重复性较差至中等（新鲜冷冻组织rho = 0.89 - 0.99，中位变异为1.6%；FFPE组织rho = -0.09 - 0.85，中位变异为7.7%）。新鲜冷冻组织和FFPE组织的结果之间仅存在较差至中等的相关性（rho = -0.28 - 0.70，中位变异为13.2%）。总之，使用当前方法时，FFPE组织不适用于确定恶性肾上腺皮质肿瘤情况不明患者的IGF2甲基化评分。因此，我们建议为诊断和生物样本库目的将切除材料新鲜冷冻保存。
