传统聚合酶链反应与新一代测序技术在急性髓系白血病中诊断FLT3、异柠檬酸脱氢酶和核仁磷酸蛋白1突变的比较：PETHEMA聚合酶链反应-线性微阵列分析研究结果

Conventional PCR Versus Next Generation Sequencing for Diagnosis of FLT3, IDH and NPM1 Mutations in Acute Myeloid Leukemia: Results of the PETHEMA PCR-LMA Study.

作者信息

Boluda Blanca, Rodriguez-Veiga Rebeca, Sargas Claudia, Ayala Rosa, Larráyoz María J, Chillón María Carmen, Soria-Saldise Elena, Bilbao Cristina, Prados de la Torre Esther, Navarro Irene, Martinez-Cuadron David, Gil Cristina, Bernal Teresa, Bergua Juan, Algarra Lorenzo, Tormo Mar, Martínez-Sanchez Pilar, Carrillo-Cruz Estrella, Serrano Josefina, Alonso-Domínguez Juan M, García Raimundo, Amigo Maria Luz, Herrera-Puente Pilar, Sayas María J, Lavilla-Rubira Esperanza, García-Pérez María José, Morán Julia, Pérez-Santaolalla Esther, Alonso-Vence Natalia, Oliva Ana, López Juan Antonio, Barrios Manuel, García-Fortes María, Olave María Teresa, Labrador Jorge, Martínez-López Joaquín, Calasanz María J, García-Sanz Ramón, Pérez-Simón José A, Gómez-Casares María T, Sánchez-Garcia Joaquín, Mendizabal Yolanda, Barragán Eva, Montesinos Pau

机构信息

Hematology Department, Hospital Universitari i Politécnic-IIS La Fe, 46026 Valencia, Spain.

Molecular Biology Unit, Hospital Universitari i Politécnic-IIS La Fe, 46026 Valencia, Spain.

出版信息

Cancers (Basel). 2025 Mar 1;17(5):854. doi: 10.3390/cancers17050854.

BACKGROUND/OBJECTIVES: This PETHEMA PCR-LMA study aimed to evaluate whether mutations detected by NGS (VAF cut-off of ≥5%) correlate with NPM1, FLT3-ITD, FLT3-TKD, IDH1, and IDH2 mutations detected using conventional PCR (analytical sensitivity 3%) in a nationwide network of seven reference laboratories.

METHODS

Between 2019 and 2021, 1685 adult AML patients with at least one centralized sample (NGS or PCR) at primary diagnosis or relapse/refractory episode were included.

RESULTS

During this period, 1288 paired NGS/PCR samples (1094 at diagnosis, 103 at relapse and 88 at refractoriness) were analyzed. Considering PCR the gold-standard, for NPM1 NGS sensitivity was 98.5% and specificity 98.9%, for FLT3-ITD 73.8% and 99.6%, for FLT3-TKD 84.5% and 99.3%, for IDH1 98.7% and 98.7%, and for IDH2 99.1% and 97.7%, respectively. Overall concordance rate of positive results between NGS (and PCR was 95% (262/276) for NPM1, 72% (149/206) for FLT3-ITD, 74% (49/66) for FLT3-TKD, 87% (77/89) for IDH1 and 84% (107/127) for IDH2. Overall, median days from sample reception until report were 7 for PCR and 28 for NGS.

CONCLUSIONS

This study shows high concordance between NPM1 and IDH results using PCR and NGS. However, sensible important discrepancies are observed for FLT3 mutations. In our context, rapid screening for these druggable mutations should be performed by conventional PCR.

背景/目的：这项PETHEMA PCR-LMA研究旨在评估在一个由七个参考实验室组成的全国性网络中，通过二代测序（VAF截止值≥5%）检测到的突变是否与使用传统PCR（分析灵敏度3%）检测到的NPM1、FLT3-ITD、FLT3-TKD、IDH1和IDH2突变相关。

方法

在2019年至2021年期间，纳入了1685例成年急性髓系白血病患者，这些患者在初次诊断或复发/难治性发作时至少有一份集中检测样本（二代测序或PCR）。

结果

在此期间，分析了1288对二代测序/PCR样本（诊断时1094对，复发时103对，难治时88对）。将PCR视为金标准，对于NPM1，二代测序的灵敏度为98.5%，特异性为98.9%；对于FLT3-ITD，灵敏度为73.8%，特异性为99.6%；对于FLT3-TKD，灵敏度为84.5%，特异性为99.3%；对于IDH1，灵敏度为98.7%，特异性为98.7%；对于IDH2，灵敏度为99.1%，特异性为97.7%。二代测序（和PCR）之间阳性结果的总体一致率，NPM1为95%（262/276），FLT3-ITD为72%（149/206），FLT3-TKD为74%（49/66），IDH1为87%（77/89），IDH2为84%（107/127）。总体而言，从样本接收至报告的中位天数，PCR为7天，二代测序为28天。