Expression Validation Of CD123 Related Long Non-coding RNA In Acute Myeloid Leukemia

Background:Acute myelogenous leukemia（AML）is a hematological disease characterized by the malignant clonal proliferation of myeloid primitive cells.Hematopoietic function is disturbed and inhibited.To date,many studies have shown that gene alterations and epigenetic regulation are involved in the occurrence and development of AML.Although there has been gradual progress in the accurate diagnosis and reasonable treatment of AML in recent years,and patients can receive traditional chemotherapy,targeted therapy and allogeneic stem cell transplantation（allo-HSCT）,many patients still suffer from complete remission and relapse or refractory problems,the overall survival period is short,and the prognosis is poor.Leukemia stem cells（LSCs）are considered to be resistant to chemotherapy drugs and are the main reason for relapse and refractory leukemia.Therefore,finding potential therapeutic targets for AML and finding new methods to interfere with AML-related genes and epigenetic changes in order to improve the prognosis and survival conditions of AML patients is a very worthy research area.Compared with normal hematopoietic stem cells（HSCs）,LSCs show specific mutation characteristics,specific metabolic characteristics and epigenetic modification.LSCs maintained many properties of HSCs,including multi-differentiation ability,self-repair ability and strong growth and proliferation ability,and had some same surface markers as normal hematopoietic stem cells:CD34⁺CD38^-,HLA-DR,CD71^-.At present,many studies are focused on the identification of specific membrane biomarkers on LSCs to accurately identify LSCs and distinguish LSCs from HSCs.One of the most promising therapeutic markers is CD123/IL-3Rα（Interleukin-3 alpha receptor）.CD123,theαsubunit of the IL-3 receptor,a type 1 transmembrane glycoprotein,is a member of the cytokine receptor superfamily.CD123 is a highly specific immunophenotype that is highly expressed in leukemia stem cells but almost not in hematopoietic stem cells.So far,CD123has been shown to increase expression in a variety of hematological malignancies.Existing literature also reported that patients with high expression of CD123 had lower clinical remission rate,higher recurrence rate,poor treatment response,and shorter disease-free survival than patients without expression of CD123,suggesting that CD123 is associated with poor prognosis of AML patients.Long non-coding RNA（lnc RNA）is a nucleotide molecule longer than other types of regulatory RNA,with a length of more than 200bp,lacking instructions to encode proteins.lnc RNA can regulate gene expression in a variety of ways and regulate life activities through pre-transcriptional,transcriptional and post-transcriptional pathways.With the wide application of high-throughput sequencing technology and chip technology,lnc RNA research has gradually become a hot field of scientific research.At present,abundant lnc RNA have been found in a variety of malignant tumor tissues.They play a key role in the occurrence and progression of malignant tumor diseases,either promoting cancer genes or inhibiting cancer genes,which also provides a certain research basis for the discovery,prevention and treatment of clinically refractory cancers,as well as the evaluation of disease progression and prognosis.According to existing research reports,CD123 may play an important role in the occurrence and development of AML,and is considered to be related to the poor prognosis of AML patients.At present,there are few reports about the role of CD123-related long non-coding RNA in the occurrence,development and prognosis of AML.In the previous stage,high-throughput sequencing technology was applied to screen out differentially expressed non-coding RNA in bone marrow samples of CD123^-and CD123⁺newly treated AML patients.In this study,real-time fluorescence quantitative PCR（q RT-PCR）was used to verify whether there were differences in expression levels of CD123-related long non-coding RNA in AML.Differential expression of CD123-related long non-coding RNA may play a role in the initiation and development of AML and affect the survival and prognosis of patients.This study provides a theoretical basis for further understanding the pathogenesis of AML and targeted therapy of AML.Objective:1.To analyze the correlation between the expression of CD123 and the general clinical characteristics and efficacy of newly treated AML patients.2.To verify the differential expression of CD123/IL-3Rα-related long non-coding RNA in patients with primary AML CD123^-and CD123⁺by real-time fluorescence quantitative PCR（q RT-PCR）.Methods:1.The target lnc RNA to be verified were identified from nc RNA differentially expressed in bone marrow samples of CD123^-and CD123⁺newly treated AML patients screened by high-throughput sequencing in the previous research group;2.Clinical bone marrow specimens of newly diagnosed AML patients were collected.There were 10 and 8 newly treated AML patients in the CD123^-and CD123⁺groups,respectively.3.Clinical and laboratory data of newly treated AML patients were collected,including age,sex,white blood cell count at initial diagnosis,hemoglobin concentration,platelet count,myeloid leukemia cell ratio,chromosome karyotype,gene mutation,etc.,and the efficacy of the first chemotherapy and two courses of chemotherapy were observed.4.The target lnc RNA of bone marrow samples from AML patients was verified by real-time fluorescence quantitative PCR.SPSS 26.0 and Graph Pad Prism 9.0 statistical software were used for data analysis and graph processing.According to the type of data distribution,if the data were in line with normal distribution,mean±standard deviation was used for statistical description,and unpaired T test was used for statistical analysis of differences between groups.If the data did not conform to the normal distribution,the median（interquartile）was used for statistical description,and the Mann-Whitney U test was used for statistical analysis.Fisher’s exact probability method was used to compare the two classification variables.Bilateral P<0.05 indicated that the difference between the two groups was statistically significant.Results:1.The proportion of myeloid leukemia cells in patients with CD123⁺AML at initial diagnosis was higher than that in patients with CD123^-AML.2.Compared with the CD123^-group,JHDM1D-AS1 and FBXL19-AS1 were i ncreased in the CD123⁺group.3.Compared with the CD123^-group,the differential expressions of BOLA3-A S1,HSPC324,LINC01355,FAM13A-AS1,LOC339803,MAGI2-AS3,BAALC-AS2,LINC00877 in the CD123⁺group were not statistically significant.Conclusion:1.Compared with the CD123^-group,patients in the CD123⁺group had a higher percentage of myeloid leukemia cells at initial diagnosis.2.Real-time fluorescence quantitative PCR（q RT-PCR）verified that differentially expressed lnc RNA in bone marrow of newly treated CD123^-AML patients and CD123⁺AML patients included JHDM1D-AS1,FBXL19-AS1.Compared with the CD123^-group,JHDM1D-AS1 and FBXL19-AS1 were increased in the CD123⁺group.