The Study Of Correlation Between The Driver Gene Mutations And Cytokines In Ph-negative Myeloproliferative Neoplasms

Objective:Analysis of the correlation between cytokine expression and driver mutations and hematological parameters and clinical symptoms in MPN patients by comparing serum cytokine [tumor necrosis factors(TNF),interferon-γ(IFN-γ),interleukin(IL)-10,IL-6,IL-4 and IL-2]expression profiles of different subtypes and driver mutation status in real-world patients with classical myeloproliferative neoplasms(MPN),to investigate the state of chronic inflammatory response caused by cytokines,to provide new assays for the diagnosis of MPN and to explore possible new targets for targeting cytokine therapy.Methods:80 patients with primary Philadelphia chromosome-negative(Ph-)MPN diagnosed in our hospital were selected for the study [22 cases of polycythemia vera(PV),29 cases of essential thrombocythemia(ET),29 cases of primary myelofibrosis(PMF)],and clinical data were collected on gender,age,WBC count,Hb,PLT count,splenomegaly,and embolism,by quantitative PCR to detect JAK2,MPL and CALR,flow cytometry for serum TNF,IFN-γ,IL-10,IL-6,IL-4,and IL-2 expression levels.Results:1.Total mutations in the JAK2,CALR,or MPL genes were present in 95%(76/80)of the MPN patients,with 85%(68/80)of total JAK2 mutations,7.5%(6/80)of total CALR mutations,and 2.5%(2/80)of total MPL mutations;5.0%(4/80)of triple-negative MPN patients.Among the patients with PV,the detection rate of the JAK2 mutation was 95.5%(21/22);triple-negative patients accounted for 4.5%(1/22);no CALR or MPL mutation was found.The rate of detection of JAK2V617 F mutation in ET patients was 69%(20/29);CALR mutation was 17.2%(5/29)and MPL mutation was 6.9%(2/29);the triple-negative patients accounted for 6.9%(2/29).In PMF patients,the rate of detection of JAK2V617 F mutation was 93.1%(27/29);MPL mutation was 3.45%(1/29);and triple-negative patients accounted for 3.45%(1/29);and no CALR mutation was found.2.When comparing the cytokine levels of the three subtypes of MPN,the PV group had significantly higher levels of IFN-γ and IL-10 than the ET group(P<0.01)and higher levels of IL-4 than the ET group(P<0.05),and the PMF group had significantly higher levels of IL-10 and IL-6 than the ET group(P<0.01),and no significant differences were found in the levels of TNF and IL-2 between the three groups(P>0.05).Cytokine levels in MPN patients were compared between different driver mutation groups,but the differences were not statistically significant(P>0.05).When further comparing the cytokine levels of each subtype in JAK2V617 F mutation positive patients,PV group had significantly higher IL-10 levels than ET group(P<0.01),and higher IL-4 levels than ET group(P<0.05),PMF group had significantly higher IL-10 and IL-6 levels than ET group(P<0.01),and higher IL-4 levels than ET group(P<0.05)),no significant differences were found in levels of IFN-γ,TNF,and IL-2 between the three groups(P>0.05).3.Different subtypes of MPN showed significant differences for RBC,HCT,Hb,PLT,LDH,PT,APTT(P<0.05)and not for WBC,FIB and D-Dimer(P>0.05).In which the levels of RBC,HCT,Hb,and APTT were higher in the PV group than in the PMF and ET groups(P<0.05);PLT levels were higher in the ET group than in the PV and PMF groups(P<0.05),and RBC,HCT,and Hb levels were higher than in the PMF group(P<0.05);LDH and PT levels were higher in the PMF group than in the PV and ET groups(P<0.05).In PV,IFN-γ was negatively correlated with HCT(r=-0.481),TNF was negatively correlated with WBC(r=-0.463),IL-6 was negatively correlated with Hb(r=-0.523),IL-10 and IL-4 were negatively correlated with FIB(r=-0.507 and r=-0.606);in PMF,IFN-γ was positively correlated with PLT relationship(r=0.373),TNF was positively correlated with WBC,RBC,and PLT(r=0.407,r=0.422,r=0.533),IL-4 was positively correlated with WBC and PLT(r=0.424,r=0.559),and IL-2 was positively correlated with WBC and PLT(r=0.440,r=0.517);There was no correlation between the measured cytokines and laboratory indicators in ET patients.4.The three clinical features of MPN with different subtypes of thrombosis,hepatomegaly,and splenomegaly were compared,and the results were significantly different(P<0.01).Compared with ET and PMF patients,PV patients were more likely to develop thrombosis(P<0.01),and compared with PV and ET patients,PMF patients were more likely to develop hepatomegaly(P<0.05)and splenomegaly(P<0.01).Among patients with MPN,IFN-γ levels were significantly higher in patients with thrombosis than in patients without thrombosis(P<0.01);IL-10 and IL-6 levels were higher in patients with hepatomegaly and splenomegaly than in patients without hepatomegaly and splenomegaly(P<0.05).5.According to the IPSS scoring system,PMF patients were classified into low-risk,intermediate-risk,and high-risk groups,the three groups showed no significant difference between them for IFN-γ,TNF,IL-10,IL-2,and IL-4(P>0.05),showing a significant difference for IL-6(P<0.05);a two-by-two comparison showed that the IL-6 level was higher in the high-risk group than in the intermediate-risk group(P<0.05).Conclusion:1.Cytokine expression in MPN patients differed among subtypes,and cytokine levels also differed among JAK2 mutation-positive patient groups;some cytokine levels were higher in patients with PV and PMF than in patients with ET.It suggests that the changes of cytokine expression in MPN patients can be monitored dynamically to help determine the disease progression.2.Cytokine levels are somewhat correlated with the clinical characteristics of MPN.3.IL-6 levels were higher in the high-risk IPSS score group of PMF patients,suggesting that IL-6 may be a potential indicator related to disease risk stratification and prognosis assessment.