BCR-ABL阴性骨髓增殖性肿瘤患者JAK2、CALR、MPL基因突变的临床分析

目的:研究BCR-ABL阴性骨髓增殖性肿瘤(MPN)患者JAK2V617F、CALR及MPL基因突变情况,并比较不同类型基因突变及突变均阴性患者部分临床参数的差异。方法:收集227例BCR-ABL阴性MPN患者各类型基因突变情况、相关检验结果及影像学检查。结果:JAK2V617F突变总检出率、CALR基因突变总检出率、MPL基因突变总检出率分别为71.4%,11.0%,2.6%。真性红细胞增多症(PV)患者中未检测到CALR、MPL基因突变。有1例原发性血小板增多症(ET)患者有两种基因突变共存(JAK2V617F+,CALR+)。在PV患者中,JAK2V617F突变阳性组年龄、白细胞计数、血小板计数、脾肿大发生率均高于JAK2V617F突变阴性组(均P0.05)。在ET患者中,JAK2V617F突变阳性组年龄、白细胞计数、血红蛋白浓度均高于CALR突变阳性组及3种基因突变均阴性组,脾肿大及血栓发生率高于CALR突变阳性组;而与MPL突变阳性组相比,仅表现为血红蛋白浓度增高(P0.05)。在原发性骨髓纤维化(PMF)患者中,仅发现JAK2V617F突变阳性组白细胞计数高于CALR突变阳性组。结论:不同MPN亚型基因突变的发生频率、分布、血栓形成风险不同,导致独特的临床表征。CALR阳性患者较JAK2V617F阳性患者更年轻、骨髓增殖水平更低、血栓事件风险更低,这提示危险分层更低、预后较好。这为MPN的诊断及个体化治疗提供证据。     

慢性淋巴细胞白血病合并骨髓增殖性肿瘤临床特征分析

目的 探究慢性淋巴细胞白血病(CLL)和骨髓增殖性肿瘤(MPN)共患病可能的发病机制并分析其临床特征。方法 回顾性纳入2000年～2020年世界范围内报道的CLL合并MPN患者112例及本院CLL合并MPN患者1例。根据MPN类型将所有患者分为真性红细胞增多症(PV)合并CLL组(35例)、原发性血小板增多症(ET)合并CLL组(53例)及原发性骨髓纤维化(PMF)合并CLL组(25例);根据JAK2V617F突变情况将已知突变情况患者95例,分为JAK2V617F阳性组(66例)和JAK2V617F阴性组(29例);根据疾病诊断顺序将所有患者分为先MPN后CLL组(60例)、同时诊断组(27例)及先CLL后MPN组(26例)。收集所有患者性别、诊断年龄、两种疾病诊断间隔时间、疾病诊断类型、JAK2V617F突变情况及其他类型突变情况、治疗及转归情况、存活状态、生存时间、MPN类型、疾病诊断顺序等临床特征资料并分组进行比较。结果 3个MPN合并CLL组中共患病以ET合并CLL最为常见(46.9%)。PV合并CLL组JAK2V617F阳性患者比例明显高于其他两组(P<0.05)。P...     

慢性骨髓增殖性疾病131例JAK2V617F突变的定量分析及临床意义

目的 研究JAK2V617F突变在慢性骨髓增殖性疾病(CMPD)中的发生率、突变类型、转录本水平及其临床意义.方法 采用突变特异性扩增系统(ARMS)-PCR方法 检测JAK2V617F的发生率及其突变类型;毛细管电泳方法 定量分析JAK2V617突变转录本水平.结果 纯合型JAK2V617F突变特发性血小板增多症(ET)患者和杂合型JAK2v617F突变ET患者其发病年龄均较野生型患者为高(P<0.05);CMPD患者发病年龄与JAK2V617F突变的发生问存在相关性(P<0.05),≥60岁患者的JAK2V617F突变转录本水平高于<60岁患者(P<0.05);JAK2V617F突变型CMPD患者中真性红细胞增多症(PV)和ET患者白细胞和血红蛋白水平均高于野生型患者(P<0.05),ET患者中纯合型突变者白细胞水平高于杂合型突变者(P<0.05);JAK2V617F突变在PV中的发生率和JAK2V617F纯合型突变在PV中的发生率均高于ET患者(P<0.05);JAK2V617F转录本水平均值在PV、ET和慢性特发性骨髓纤维化患者中无差别.结论 ARMS-PCR结合毛细管电泳定性和定量分析JAK2V617F突变可用于CMPD的诊断和预后判定.     

JAK2V617F突变分型和相对定量研究——附123例慢性骨髓增殖性疾病检测结果分析

目的 研究JAK2V617F突变在慢性骨髓增殖性疾病(CMPD)中的发生率和突变类型,并对突变转录本水平进行定量分析.方法 对2003年1月至2008年5月苏州大学附属第一医院就诊的CMPD患者采用突变特异性扩增系统(ARMS)PCR方法检测JAK2V617F的发生率及其突变类型;采用毛细管电泳方法定量分析JAK2V617F突变转录本水平.结果 123例CMPD患者共检出90例JAK2V617F阳性,其中35例真性红细胞增多症(PV)患者中JAK2V617F阳性率100%,85例原发性血小板增多症(ET)患者中JAK2V617F阳性率62.4%,低于PV患者,差异有统计学意义(P<0.05);3例慢性骨髓纤维化(IMF)患者中JAK2V617F阳性率66.7%.90例JAK2V617F突变患者中共检出纯合突变35例,其中PV患者17例(17/35),占48.6%,ET患者17例(17/85),占20.0%,低于PV患者,差异有统计学意义(P<0.05),IMF患者1例;毛细管电泳定量分析显示,纯合型突变患者JAK2V617F突变转录本水平较杂合型突变患者低,差异有统计学意义(P<0.05);杂合型PV患者JAK2V617F突变转录本水平高于杂和型ET患者,差异有统计学意义(P<0.05).93例患者进行了染色体检查,6例有核型异常,但未发现特异性染色体改变.结论 ARMS-PCR可作为JAK2V617F突变较准确的检测方法,结合毛细管电泳可用于此突变的定量分析以及临床CMPD的诊断.     

JAK2V617F点突变在BCR-ABL阴性骨髓增殖性疾病中的意义

目的研究JAK2V617F点突变在诊断BCR—ABL融合基因阴性的骨髓增殖性疾病（MPD）患者中的意义。方法选择51例BCR—ABL阴性MPD患者,采用等位基因特异PCR法检测各组患者JAK2V617F的突变情况。结果51例BCR—ABL融合基因阴性的MPD患者中,34例JAK2V617F突变阳性,其中原发性血小板增多症（ET）18例（69．23％）,真性红细胞增多症（PV）16例（66．67％）,特发性骨髓纤维化（IMF）1例为阴性;PV与ET患者相比更容易发生肝脾肿大、脑梗死、静脉血栓形成、高尿酸血症等并发症（P〈0．05）。ET患者中,JAK2V617F突变阳性组白细胞计数较阴性组高（P〈0．05）。ET患者中,JAK2V617F突变阳性组白细胞计数较阴性组高（P〈0．05）,ET和PV患者中JAK2V617F突变阳性组都比阴性组更容易发生上述并发症等（P均〈0．05）。结论JAK2V617F点突变在BCR—ABL融合基因阴性MPD中有较高的发生率,具有明确的诊断学意义;ET及PV患者中此突变阳性者更易发生血栓形成等并发症。     

MPN患者JAK2基因V617F点突变的检测及意义

目的探讨JAK2基因V617F点突变在骨髓增殖性肿瘤（MPN）中的发生情况及其意义。方法本组受检者共100例。MPN患者82例,其中真性红细胞增多症（PV）20例,有红细胞增多但未能诊断为PV者10例,原发性血小板增多症（ET）14例,有血小板增多但未能诊断为ET者13例,原发性骨髓纤维化（PMF）9例,慢性髓性白血病（CML）2例,其他MPN 6例,其他不明原因白细胞升高患者8例;另有急性白血病（AML）1例,骨髓增生异常综合征（MDS）10例、腔隙性梗死3例,慢性肾炎1例,大B细胞性淋巴瘤1例,异体胎肝移植患者1例,正常体检者1例。提取100例受检者外周血液及骨髓样本中有核细胞DNA,采用直接测序法进行JAK2第12、14外显子突变热点的检测。结果 100例样本中,有52例检测了第12、14两个外显子,48例仅检测第14外显子。未发现第12外显子突变。V617F及其他突变总检出率为38%（38/100）。PV、ET、PMF、CML、其他MPN、其他不明原因白细胞升高患者中均检测到不同比例的JAK2基因V617F点突变;而18例非MPN患者样本中均未发现V617F点突变,两组相比,P〈0.01。结论 JAK2基因突变在MPN的发生率高于其他血液疾病,可作为诊断MPN的一项重要参考指标。     

特发性骨髓纤维化患者JAK2V617F和MPLW515L点突变及JAK2外显子12突变的研究

特发性骨髓纤维化(IMF)为慢性骨髓增殖性疾病(CMPD)的一种,缺乏特异性诊治.现在研究表明近一半的IMF患者存在JAK2V617F点突变,另外,MPLW515突变及JAK2外显子12突变的发现提示MPD除了JAK2V617F点突变以外存在其他的发病机制[1].本研究应用等位基因特异性-聚合酶链反应(AS-PCR)及基因测序检测30例IMF患者的JAK2V617F、MPLW515L点突变及JAK2外显子12突变,了解其在IMF患者中的发生情况.     

骨髓增殖性肿瘤128例JAK2V617F基因突变与血栓栓塞关系研究

目的研究骨髓增殖性肿瘤(MPN)患者JAK2V617F突变与发生血栓栓塞的关系。方法回顾性分析2008年2月至2011年7月新疆地区128例不同民族MPN患者临床特征、实验室检查、JAK2V617F基因突变及血栓栓塞事件发生情况等资料。结果 MPN患者中93例(72.6%)存在JAK2V617F突变,66例(51.6%)发生血栓事件,其中突变阳性93例患者中58例有血栓形成,35例突变阴性患者中8例有血栓形成,突变阳性组与阴性组血栓发生率比较差异有统计学意义(χ2=15.893,P<0.05)。76例汉族MPN中58例为突变阳性,42例发生血栓,52例少数民族中35例突变阳性,24例发生血栓,汉族与少数民族患者JAK2基因突变总阳性率及血栓发生率比较均无统计学意义(χ2=1.261,1.026,P>0.05)。结论真性红细胞增多症、原发性血小板增多症及特发性骨髓纤维化等经典MPN患者中均有较高的JAK2V617F基因突变率,并突变阳性者血栓发生率明显高于阴性者;新疆地区汉族与维、哈、回等少数民族MPN患者JAK2V617F基因突变阳性率及血栓形成率之间差异无统计学意义。该突变阳性、年龄≥60岁、WBC≥15×109/L、合并有血管危险因素的患者血栓发生风险可能会高,应尽早予以干预治疗。     

芦可替尼时代骨髓增殖性肿瘤的治疗策略

不同类型的费城染色体阴性的骨髓增殖性肿瘤(MPN) 具有相同的JAK2V617F 突变;JAK2 抑制剂芦可替尼 的应用,仅在一部分MPN 有效,均提示JAK2 靶点非MPN 分子通路“惟一”。其中,包括TET2 、ASXL1 等表观遗传 学基因突变参与了MPN 进展和转化。此外,真性红细胞增多症(PV) 和原发性血小板增多症(ET) 患者的治疗目标 仍然是避免血栓栓塞,降低急性白血病(AL) 和PV 或ET 后骨髓纤维化(MF) 的风险。而原发性骨髓纤维化(PMF) 的治疗目标则是提高患者生存质量,延长患者生存期。对PMF 患者而言,依据芦可替尼疗效和深度基因检测度进 行分层是对现有的临床危险分层治疗的合理补充。     

骨髓增殖性肿瘤分子学研究进展及其临床价值

<正>William Dameshek(1951)提出骨髓增殖性肿瘤(myeloproliferative neoplasms,MPNs)的概念,认为真性红细胞增多症(PV)、原发性血小板增多症(ET)、原发性骨髓纤维化(PMF)属于同一组疾病。Kralovics等(2005)对JAK2 V617F突变在MPNs患者中的发现,更加印证了上述概念提出的重要意义。随后在少数ET和PMF患者中发现MPL突变。2013年,有研究在JAK2、MPL基因突变阴性的ET和MF患者中,首次报道CALR基因     

Detection of MPL exon10 mutations in 103 Chinese patients with JAK2V617F-negative myeloproliferative neoplasms

JAK2V617F mutation has been reported in 90% of patients with polycythemia vera (PV) and about 50% of patients with essential thromobocythemia (ET) and primary myelofibrosis (PMF). Recently, acquired mutations in the transmembrane-juxtamembrane region of MPL (MPLW515 mutations) have been reported in approximately 5% of JAK2V617F-negative PMF and about 1% of all cases of ET. MPL is the receptor for thrombopoietin that regulates the production of platelets by bone marrow. It is likely that some mutations more closely related to ET in MPL exon10 may have been missed by current assays. We inferred that there might be other mutations in MPL exon10 for MPN patients in addition to MPLW515 mutations. To investigate its mutation types and prevalence in Chinese patients with myeloproliferative neoplasms (MPN), we performed mutation detection on MPL exon10 in 103 JAK2V617F-negative MPN patients by single strand conformation polymorphism (SSCP) and allele-specific PCR (AS-PCR) combined with sequencing. As a result, one previously unrecognized MPL mutation (12-bp in-frame insertion) was identified in one patient with ET in addition to an MPLW515K mutation identified in one PMF patient. This confirms our hypothesis that BCR/ABL negative and JAK2V617F-negative MPN patients have other mutations besides W515 mutation in MPL exon10 and mutations other than single nucleotide exchange also exist. In addition, MPL mutation was associated with Chinese MPN patients.     

The analysis of JAK2 and MPL mutations and JAK2 single nucleotide polymorphisms in MPN patients by MassARRAY assay

Recent studies have shown that JAK2 V617F, MPL W515L/K and JAK2 exon 12 mutations underlie the major molecular pathogenesis of myeloproliferative disorders (MPN). Allele-Specific Polymerase Chain Reaction (AS-PCR), direct sequencing and MassARRAY assay were used to ascertain the real prevalence of these mutations and the influence of genetic susceptibility in Chinese MPN patients. The positive rate of JAK2 V617F in polycythaemia vera (PV), essential thrombocythaemia (ET) and primary myelofibrosis (PMF) was 82.0%, 36.6% and 51.1% respectively. One ET patient and two PMF patients harboured the MPL W515L mutation and three PV patients harboured JAK2 exon 12 mutations. All of these patients were confirmed as JAK2 V617F negative. Clinical data demonstrated that PV patients with JAK2 exon 12 mutations were younger, had higher haemoglobin levels and white blood cell counts than PV patients with JAK2 V617F. In addition, through analysis of 4 polymorphic loci of JAK2 gene, no significant difference of distribution frequency was found among PV, ET and PMF patients. Distribution frequency of haplotype also was not significantly different among PV, ET and PMF patients. We conclude that JAK2 V617F is a major molecular pathogenesis in Chinese MPN patients. MPL W515L mutation and JAK2 exon 12 mutations can also be found in JAK2 V617F negative MPN patients.     

Anaemia characterises patients with myelofibrosis harbouring Mpl mutation

The clinical and haematological phenotype of patients with myelofibrosis harbouring MPL(W515L/K) mutation has not been thoroughly investigated. Of 217 myelofibrosis subjects, 18 (8.2%) had an MPL mutation, four of which (22%) co-existed with JAK2(V617F) mutation. When compared with MPL wild-type patients, irrespective of JAK2(V617F) status, those with MPL(W515L/K), were more frequently female, were older (61 years vs. 57 years; P = 0.02), presented with more severe anaemia (haemoglobin, 101 g/l vs. 121 g/l; P = 0.002) and were more likely to require regular transfusional support (P = 0.012). These data indicate that MPL mutation in myelofibrosis characterises patients with more severe anaemic phenotype.     

Prevalence of overt myeloproliferative neoplasms and JAK2 V617F mutation in Korean patients with splanchnic vein thrombosis

Introduction: Myeloproliferative neoplasm (MPN) is known to be a major risk factor of splanchnic vein thrombosis (SVT). Recent studies revealed that a significant proportion of patients with SVT harbor a gain‐of‐function mutation in the JAK2 gene (V617F) with or without MPN. In this study, the authors investigated the prevalence of MPN and JAK2 V617F mutation in Korean patients with SVT. Methods: The study subjects were 26 patients diagnosed as having SVT based on Doppler ultrasound and/or computed tomography from January 2008 to January 2010 (16 men and 10 women; mean age 44 years, range 15–75 years). The clinical and laboratory data were reviewed. The JAK2 V617F mutation was detected by allele‐specific polymerase chain reaction and direct sequencing analyses using DNA from peripheral blood leukocytes. Results: Among 26 study patients, 12 had portal vein thrombosis, five had hepatic vein thrombosis, three had mesenteric, and two had splenic vein thrombosis. Four patients had thrombosis involving more than one splanchnic vein. Two patients (7.7%; 2/26) had overt MPN (essential thrombocythemia). JAK2 V617F was detected in three patients (11.5%) including the two patients with overt MPN. Thus, the prevalence of the JAK2 V617F mutation in patients with SVT but without overt MPN was 4.2% (1/24). Conclusion: The prevalence of overt MPN and that of JAK2 V617F were lower in Korean patients with SVT than in previous reports. Data from a larger number of patients with long‐term follow‐up are needed to reveal the clinical relevancy of JAK2 V617F in Korean patients with SVT.     

JAK2 GGCC haplotype in MPL mutated myeloproliferative neoplasms

JAK2 (V617F) is associated with a genetic predisposition to its acquisition,as it is preferentially found in subjects with a common constitutional JAK2 haplotype known as 46/1 or GGCC. A recent study suggests that a genetic predisposition to acquisition of MPL mutation may exist in sporadic patients, since an association was found with the JAK2 46/1 haplotype. We genotyped 509 patients with myeloproliferative neoplasms (MPN), 7% of which carrying a somatic mutation of MPL Exon 10. We found that the JAK2 GGCC haplotype was closely associated with JAK2 (V617F) (OR 1.84, P < 0.001) but not with MPL mutations (OR 0.98), suggesting a different genetic background for these molecular lesions.     

Progenitor genotyping reveals a complex clonal architecture in a subset of CALR‐mutated myeloproliferative neoplasms

The identification of acquired CALR mutations in patients with essential thrombocythaemia (ET ) or myelofibrosis (MF ) has meant that disease‐initiating mutations can now be detected in about 90% of all patients with a myeloproliferative neoplasm (MPN ). Here, we show that only those CALR mutations that cause a +1 frameshift, thereby altering the carboxy‐terminus of calreticulin, promote cytokine independence in vitro; in‐frame deletions were not functional, and are unlikely to be the pathogenetic mutation underlying some MPN cases. Expression of the thrombopoietin receptor, MPL , was also necessary for factor‐independence. Although the CALR mutations are considered to occur only in JAK 2 V617F‐negative cases and in a heterozygous state, progenitor genotyping revealed that this is not always true. Notably, CALR mutation‐positive MPN s can be polyclonal: in one case, two distinct CALR mutation‐positive subpopulations could be identified; in another, separate populations of JAK 2 V617F‐positive and CALR ‐mutated cells were present. Mitotic recombination involving chromosome 19 in a third instance resulted in the emergence of a CALR mutation‐homozygous subclone. Collectively, our studies demonstrate that occasional patients with CALR mutation‐positive ET or MF carry other MPN ‐initiating genetic mutations (including JAK 2 V617F), acquire “secondary mutations” before or after the CALR mutation, or evolve over time to being CALR mutation‐homozygous.     

JAK2 V617F‐positive acute myeloid leukaemia (AML): a comparison between de novo AML and secondary AML transformed from an underlying myeloproliferative neoplasm. A study from the Bone Marrow Pathology Group

The JAK2 V617F mutation is characteristic of most Philadelphia chromosome‐negative myeloproliferative neoplasms (MPNs) and occurs rarely in de novo acute myeloid leukaemia (AML). We sought to characterize AMLs that harbour this mutation and distinguish those that arise de novo (AML‐DN) from those that reflect transformation of an underlying MPN (AML‐MPN). Forty‐five patients with JAK2 V617F‐mutated AML were identified; 15 were AML‐DN and 30 were AML‐MPN. AML‐MPN cases were more likely to have splenomegaly (P = 0·02), MPN‐like megakaryocytes and higher mean JAK2 V617F VAF at diagnosis (P = 0·04). Mutations involving TET2 were exclusively identified in AML‐DN patients. Mutations of genes affecting DNA methylation were more common in AML‐DN (P < 0·01). A complex karyotype was more frequent in AML‐MPN cases than in AML‐DN (P < 0·01), with AML‐DN more likely to display a normal karyotype (P = 0·02). Bone marrow histology after recovery from induction chemotherapy in AML‐DN cases revealed no morphological evidence of any previously occult MPNs, while this was evident in most of the AML‐MPN specimens (P < 0·01). These findings in this largest study of JAK2 V617F‐mutated AMLs indicate that AML‐DN is distinct from AML‐MPN.     

Phenotypic variations and new mutations in JAK2 V617F-negative polycythemia vera, erythrocytosis, and idiopathic myelofibrosis

OBJECTIVE: The chronic myeloproliferative disorders (MPD), polycythemia vera (PV), essential thrombocytosis, and idiopathic myelofibrosis (IMF), are characterized by a spectrum of clinical features and linked by common genetic lesions in JAK2 and MPL. However, the clinical phenotypes in genetically undefined MPD patients are similar to those patients with JAK2 and MPL lesions. We, therefore, sought to determine whether there were JAK2 or MPL lesions in a well-defined, JAK2 V617F-negative MPD cohort, and to determine if clinical associations could be identified based on variations identified in these genes. METHODS: We examined the JAK2 and MPL genes in JAK2 V617F-negative PV, IMF, and idiopathic erythrocytosis patients for sequence variations. RESULTS: We identified two previously unrecognized JAK2 mutations and three previously unrecognized MPL mutations in JAK2 V617F-negative PV, erythrocytosis, and IMF patients. We identified JAK2 exon 12 lesions in 30% of JAK2 V617F-negative PV patients, and either JAK2 V617F or JAK2 exon 12 lesions in 9% of erythrocytosis patients. In IMF, in addition to the MPL gene mutation, W515K, we identified three additional mutations: 204P and two intervening sequence transitions, IVS 11/12 and 10/11. CONCLUSIONS: While the clinical phenotype of JAK2 exon 12 lesions in the MPD was predominantly erythroid, there was significant disease spectrum overlap between JAK2 V617F and JAK2 exon 12 mutations. By contrast, MPL gene mutations were not associated with erythrocytosis, but segregated primarily with the phenotypes of thrombocytosis, extramedullary disease, myelofibrosis, and osteosclerosis.