Preliminary Study On The Expression And Clinical Significance Of PD-L1 In Multiple Myeloma Cells And PD-1 In T And NK Cells

Background and Objective:Multiple myeloma（MM）is a malignant tumor with clonal proliferation of plasma cells originating from the blood system,which has a variety of clinical symptoms,complex pathogenesis and poor prognosis.It is one of the common hematologic malignancies.In recent years,the incidence of MM has been increasing year by year.At present,most patients with MM appear disease recurrence and progression,primary drug resistance after conventional treatment,leading to final death.Programmed cell death protein 1（PD-1）and programmed cell death 1 Ligand1（PD-L1）activate T cells by negatively regulating signals.It is one of the most promising immunotherapeutic strategies,mainly by changing the state of the immune surveillance of tumor cells.In this research,the expression of programmed cell death ligand 1（PD-L1）on myeloma cells and the expression level of programmed cell death protein 1（PD-1）on T cells in patients with Multiple Myeloma（MM）,were detected by Flow Cytometry（FCM）.The aims were to analysis the relationship between PD-1/PD-L1 and the MM patients’clinical significance,and further explore the mechanism of PD-1/PD-L1expression in Multiple Myeloma.The studies lays a theoretical foundation for a new treatment plan for patients with multiple myeloma.Methods:（1）Collection:a total of 97 cases who were diagnosed with multiple myeloma were collected from July 2018 to November 2020 in the Oncology Department of the First Affiliated Hospital of Zhengzhou University,including 68 newly diagnosed patients,12 refractory patients with recurrence and 17 patients with complete remission.Anticoagulant bone marrow of the subjects were collected,corresponding clinical data and laboratory examinations were reviewed and statistically analyzed using the electronic records system.And the therapeutic effect of the patients was tracked for at least half a year.（2）Flow cytometry was used to detect the expression of T cell subsets,NK cells and the expression of surface PD-1 in the bone marrow of patients with multiple myeloma,and the expression of PD-L1 on the surface of plasma cells.（3）Relationship between the expression of PD-L1 in the bone marrow of patients with multiple myeloma and its clinical laboratory characteristics was analyzed.（4）Means and quantles[M（P25～P75）]were used to represent the non-normal distribution of experimental data.Comparison between groups was performed by Kruskal-Wallis H test,pairwise comparison was performed by Mann-Whitney U test,Rank sum test was used to compare two groups’data.Statistical software SPSS 22.0was used to analyze acquired data,and Graph Pad Prism 8 was used to carry out relevant data mapping.P<0.05 means that the difference is statistically significant.Results:（1）A total of 97 patients with multiple myeloma were diagnosed,including55 males（57%）and 42 females（43%）.The onset age ranged from 40 to 84 years old（median age was 60 years old）.Among them,there were 67 cases of Ig G type,6 cases of Ig M type,21 cases of Ig A type,2 cases of non-secretory type,and 1 case of light chain type.According to the Internationalized Staging System（ISS）:10 cases were stage I,31 cases were stage II,and 56 cases were stage III.（2）Compared with the normal control group,the proportion of CD4+T cells,CD8+T cells and the ratio of CD4/CD8 in the bone marrow of patients with multiple myeloma in the newly diagnosed group and the remission group were not statistically significant（P>0.05）.（3）Respectively compared with the control group and the newly diagnosed group,the proportion of CD4+T cells in the bone marrow of patients with multiple myeloma in the relapse and refractory group were reduced（P<0.05）,the proportion of CD8+T cells was increased（P<0.05）,and the ratio of CD4/CD8 was reduced（P<0.05）,the differences were statistically significant.Compared with the control group,the expression level of PD-1 increased in CD3+CD4+T cells on the bone marrow of patients with multiple bone marrow tumors in the newly diagnosed group and the relapsed and refractory group,and the expression level in CD3+CD8+T cells also increased.The difference has statistical significance（P<0.05）.（4）Compared with the control group,the proportion of NK cells in the newly diagnosed group and the refractory relapse group was increased,but the increase of NK cells in the newly diagnosed group was not statistically significant（P>0.05）,and the difference of NK cells in the refractory relapse group was statistically significant（P<0.05）.Compared with the newly diagnosed group,the NK cells in the bone marrow of the relapse and refractory group were increased,and the difference was statistically significant（P<0.05）.Compared with the control group,the expression of PD-1 on NK cells in the bone marrow of patients with multiple myeloma in the relapsed and refractory group was increased,and the difference was statistically significant（P<0.05）.The expression of PD-1 on NK cells（CD3-CD56+）in the bone marrow of patients with multiple myeloma in the relapse and refractory group was higher than that in the newly diagnosed group,and the difference was statistically significant（P<0.05）.（5）Flow cytometry was used to analyze the expression of abnormal plasma cells and PD-L1 on their surface in patients with multiple myeloma.The results showed that PD-L1 was expressed in multiple myeloma cells,and the positive rate Abstractwas 16.25%（13/80）.The positive rate of PD-L1 expression in the initial treatment group was 14.7%（10/68）,and the positive rate of PD-L1 expression in the relapse and refractory group was 25%（3/12）.Compared with the initial treatment group,the positive rate of PD-L1 expression in multiple myeloma was significantly higher in the relapse and refractory group（P<0.05）,and the difference was statistically significant.（6）There was no correlation between the expression of PD-L1 in multiple myeloma cells and the patients’hemogram（white blood cells,hemoglobin）,albumin,globulin,bone marrow plasma cell proportion,lactate dehydrogenase,ferritin andβ₂-microglobulin,and the difference was not statistically significant.Conclusion:（1）PD-L1 was expressed in multiple myeloma,and the expression level in the refractory group was higher than that in the newly diagnosed group.There was no association between PD-L1 expression and relevant clinical examination features in patients with multiple myeloma.（2）The expression of PD-1 and PD-L1 in the bone marrow of patients with multiple myeloma is related to the progression of the disease.PD-1/PD-L1 may be involved in the immune escape of multiple myeloma.