Division of Hematology-Oncology, Chang Gung Memorial Hospital at Linkou, Taoyuan, Taiwan.
College of Medicine, Chang Gung University, Taoyuan, Taiwan.
J Clin Pathol. 2018 Jun;71(6):514-521. doi: 10.1136/jclinpath-2017-204829. Epub 2017 Dec 4.
We investigated the clinical and prognostic relevance of the mutational status of driver genes with allele burden and endogenous erythroid colony (EEC) growth in 203 Taiwanese patients with primary myelofibrosis (PMF).
Pyrosequencing was used to detect mutational status and measure allele burden, while (exon 10) mutations were analysed by PCR assay and then by direct sequencing. exon 9 mutations were first screened for length changes by GeneScan followed by sequencing. The allele burden of the mutated gene was measured by pyrosequencing. The EEC assay was conducted using a serum-free culture system.
The frequencies of the three driver mutations and triple-negative status were similarly distributed between pre-PMF and overt PMF patients, except that pre-PMF patients had a higher incidence of type 2/type-2 like mutations and a lower allele burden. EEC growth and mutations conferred favourable overall survival (OS). A lower allele burden and grade 3 bone marrow fibrosis were associated with shorter OS and decreased leukaemia-free survival (LFS). Type 2/type 2-like mutations were associated with better LFS compared with type1/type 1-like mutations. Patients with triple-negative mutation status had significantly worse OS and LFS. The allele burden of mutations remained unchanged, while some mutations showed clonal expansion in patients during secondary acute myeloid leukaemia transformation.
Our study showed that EEC growth, a higher allele burden and mutations, especially type 2, were independent predictors for better outcomes in PMF. The allele burden of mutations remained stable, but the allele burden of mutations was variable during leukaemia transformation.
我们研究了在 203 例台湾原发性骨髓纤维化（PMF）患者中，等位基因负担和内源性红细胞集落（EEC）生长与驱动基因突变的临床和预后相关性。
采用焦磷酸测序法检测突变状态和等位基因负担，采用 PCR 法和直接测序法分析 （exon 10）突变，exon 9 突变先通过 GeneScan 检测长度变化，然后进行测序。通过焦磷酸测序测量突变 基因的等位基因负担。采用无血清培养系统进行 EEC 检测。
三种驱动基因突变和三阴性状态的频率在前期 PMF 和显性 PMF 患者之间分布相似，除前期 PMF 患者 2/2 样突变和 等位基因负担较高，而 突变发生率较低。EEC 生长和 突变与总生存（OS）较好相关。较低的 等位基因负担和 3 级骨髓纤维化与较短的 OS 和降低的无白血病生存（LFS）相关。2/2 样 突变与 1/1 样突变相比，与更好的 LFS 相关。三阴性突变状态患者的 OS 和 LFS 显著较差。 突变的等位基因负担保持不变，而在继发性急性髓系白血病转化中，一些 突变显示出克隆性扩张。
我们的研究表明，EEC 生长、较高的 等位基因负担和 突变，特别是 2 型，是 PMF 患者预后较好的独立预测因素。 突变的等位基因负担保持稳定，但在白血病转化过程中 突变的等位基因负担是可变的。
