| JAK2?CALR and MPL gene mutation detection and clinical observation of 1648 Philadelphia chromosome negative myeloproliferative neoplasms patients of a single center(1995-2016)Objective To analyse systematically the clinical and laboratory features of 2724 Ph-MPNs patients in our laboratory from the following parts: MPNs driven gene distribution,clinical parameters,cytogenetics and progression of the disease,and reveal the clinical characteristics of Ph-MPNs in our area.Methods The clinical information of 2724 Ph-MPNs patients followed the WHO(2016)criterion from 2009 to 2016 was collected to analyse the mutations of driver genes of MPNs patients,and the clinical characteristics.Results 1.General features of 2724 patients with Ph-MPNs: disease distribution: there are 1754 cases of ET patients(64.39%),784 cases of PV patients(28.78%),186 cases of PMF patients(6.83%).Age distribution and sex ratio: In our patient cohort,first onset age of MPNs patient can at any age,the median age of onset was 59(3-95 years old),predominantly at the age of 40-79 years old.Male to female ratio: 1.015:1.2.The mutations of driver genes of 2724 patients with Ph-MPNs: The JAK2V617 F was found in 70.2% MPNs(1904/2711),in 62.03% of ET patients(1088/1754),in 90.1% of PV patients(706/784),in 63.58% of PMF patients(110/173);JAK2 exon12 mutation was found in 0.77% MPNs(16/2077),0.1% of ET patients(2/1754),4.3% of PV patients(14/323),CALR exon9 gene mutation was found in 32.7% of MPNs(230/703),in 32.9% of ET patients(206/625),in 30.77% of PMF patients(24/78);MPL exon10 gene mutation was found in 3% of MPNs(17/568),in 2.8% of ET patients(14/500),in 4.4% of PMF patients(3/68).At the same time,we found a case of ET with co-occurrenc of JAK2V617 F and CALR exon9 gene mutation.3.The relevance of driver gene mutations and clinical characteristics of 2724 patients with Ph-MPNs: In our study,the median onset age and white blood cell count of patients with JAK2V617 F was significant higher than patients with JAK2 exon12 mutation or without mutations(P<0.001).In ET group,patients with JAK2V617 F had higher white blood cell count and hemoglobin level than patients with CALR and MPL mutation(differences are statistically significant).The platelet count of patients with CALR mutations was significantly higher than patients with JAK2V617F(P <0.001).The patients without mutation were younger,and had lower white blood cell count and hemoglobin level compared with the other groups.4.The chromosome abnormalities and prognosis of 2724 patients with PhMPNs: The incidence of clonal chromosomal abnormality in ET patients was 5%(66/1339).And the main type of chromosome aberration was chromosome number abnormality,of which –Y was and the most common abnormality(18.18%).In PV patients,the incidence of clonal chromosomal abnormality was 5.2%(31/593),and the most common abnormal chromosome was trisomy 8 and trisomy 9 in each 6 cases(19.35%).In patients with PMF,the incidence of clonal chromosomal abnormality was 24.6%(34/138),dominantly in complex abnormal clonal chromosomal(47.06%).The incidence of abnormal chromosome in PMF was significantly higher than that of ET and PV(P<0.001).0.51% of the ET patients transformed into AML(9/1754),0.2% of which transformed into MDS(3/1754),0.4% of which transformed into MF(7/1754),and 0.2% of which transformed into PV(3/1754).Meanwhile 0.12% of the PV patients transformed into MDS(1/784),0.64% of which transformed into MF(5/784).However 4.8% of the PMF patients transformed to AML(9/186),1.1% of which transformed into MDS(2/186).The incidence of PMF transformed into AML was significantly higher than that of PV and ET(P<0.001).Conclusions1.In our patient cohort,first onset age of MPNs patient is predominantly at the age of 40-79 years old.At the same time,there was more patients of ET than PV and PMF.2 The driver gene mutation spectrum of our center Ph-MPNs patients was consistent with previous reports,different subtypes of MPNs had different types and frequencies of driver gene mutations,and different driver gene mutations lead to unique clinical phenotype.The incidence of clonal chromosomal abnormality and transformation into AML of the PMF patients is higher than that of ET and PV.WT1 gene expression in primary myelofibrosis and its relationship with DIPSS-plusObjective: To reveal WT1 gene expression in PMF and its relationship with DIPSS-plus,we measured the level of WT1 gene expression in 57 newly diagnosed PMF.Methods: 1.We measured the transcript levels of the WT1 gene and ABL gene in 57 newly diagnosed PMF patients by real-time quantitative RT-PCR.The WT1 transcripts were normalized with respect to the number of ABL transcripts and expressed as WT1 or copy numbers every 104 copies of ABL.In this study,a value of 300copies/10000 abl copies was set as the cut-off value for WT1 m RNA expression.2.We analyzed whether the levels of WT1 m RNA expression were associated with clinical characteristics(DIPSS-plus)by statistical methods.Thereby discuss its relationship with disease progression and clinical significance.Results: 1.According to DIPSS-plus,the PMF patients were divided into four groups: low risk,intermediate 1 risk,intermediate 1 risk,high risk groups,in which the WT1 gene expression increased gradually.The expression of WT1 in the high-risk group was significantly higher than that of the other three groups(differences are statistically significant).However,there was an increase trend in the low risk,intermediate 1 risk,intermediate 2 risk groups,but no statistically significant reached.2.The WT1 expression of PMF was significantly higher after transformation into AML.than that of the total of PMF patiens and also the high risk group.(P=0.004,0.013)3.Patients with high WT1 expression were more likely to have low platelets P=0.023).Accordingly,high WT1 expression were associated with more frequent constitutional symptoms(P=0.026)and palpable splenomegaly(P=0.013).Conclusion: There are different levels of WT1 gene expression in PMF patients,and they tended to increase with disease progression.It can be used for risk assessment and monitor of disease progression in PMF patients.Application of an NGS in 62 triple negative myeloproliferative neoplasmsObjective: To identify the disease-causing mutation in triple-negative cases of MPN by applying the next generation sequencing technique for the purpose of identifying somatic mutations.Methods: 1.62 of triple negative MPN patients were enrolled(29 ET,25 PV,8 PMF),whose DNA samples were processed according to the manufacturer's instructions and Sequencing was performed using the Illumina Hiseq 4000 platform.2 Gene mutations were divided into active signaling,chromatin modifer,DNA repair,epigenetic modifier,RNA splicing,transcription factor,and the others 7groups according to gene function.Mutation frequency and patterns of the respective MPN subgroups were studied.Results: 1.82.3%(51/62)of the triple negative MPN were identified gene mutations,the left 17.7% parts were not founded,of which 6 were ET,4 were PV,1 were PMF.126 gene mutations were detected in all of the cases,including 51 gene mutations in ET group,46 gene mutations in PV group,29 gene mutations in PMF group.There were 1.76 gene mutations in each ET,1.84 mutations in each PV,while 3.63 gene mutations in each PMF.2.There were significantly more active signaling mutation(including KIT,NRAS,KRAS,MPL,CBL,MPL,PV)in PMF group than in ET and PV group(P=0.016,P=0.029),and there was no difference between the ET and PV groups(P=0.967).3.We found one case of ET and one case of PV haborned.R443 X and T810 I mutation respectively in exon 11 and 18 of JAK2 gene.Meanwhile Y591 D and G446 E were founded in one ET and PMF which located in exon 12 and exon 9 of MPLgene.Conclusion: The mutational landscape in triple negative MPN is much more complex than initially thought.Per capita number of mutations in PMF is more than ET and PV.The mutation patterns of PMF was significantly different from ET and PV.There are other mutations outside of most frequently region of JAK2 and MPL gene which can also cause disease. |
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