Identifying the Epitopes of Bacillus thuringiensis Cry2Aa Toxin Involved in Cadherin Interaction by a Monoclonal Antibody.

Antibodies are a useful tool for assistance to map the binding epitopes in Bacillus thuringiensis Cry toxins and their receptors, and even determine how receptors promote toxicity. In this work, a monoclonal antibody (mAb-1D2) was produced by the hybridoma cell line raised against Cry2Aa toxins, with a half inhibition concentration (IC50) of 9.16 µg/mL. The affinity constant of two recombinant toxin-binding fragments derived from Helicoverpa armigera and Plutella xylostella cadherin-like protein (HaCad-TBR or PxCad-TBR) to Cry2Aa toxin was measured to be 1.21 µM and 1.24 µM, respectively. Competitive ELISA showed that mAb-1D2 competed with HaCad-TBR or PxCad-TBR binding to Cry2Aa. Meanwhile, the toxicity of the Cry2Aa toxin to the H. armigera and P. xylostella larvae were greatly reduced when the toxin was mixed with mAb-1D2, which indicated that cadherin may play an important functional role in the toxicity of Cry2Aa. After transforming mAb-1D2 to a single-chain variable fragment (scFv), the hot spot residues of Cry2Aa with 1D2-scFv, PxCad-TBR, and HaCad-TBR were analyzed by molecular docking. It was demonstrated that the hot spot residues of Cry2Aa involving with 1D2-scFv interaction were mainly in Domain II, and some residues in Domain I. Moreover, mAb-1D2 and the two cadherin fragments shared the common hot spot residues on Cry2Aa, which could explain mAb-1D2 inhibited Cry2Aa binding with cadherin fragments. This monoclonal antibody could be a useful tool for identifying the binding epitopes between Cry2Aa and cadherin, and even assist to analyze the roles of cadherin in Cry2Aa toxicity.

[Influence of Serum TGF-ß1 and EGFR Levels on the Therapeutic Effect of High-Dose AraC in Patients with Acute Myeloid Leukemia Based on the Decision Curve].

OBJECTIVE: To analyze the influence of serum levels of transforming growth factor-ß1 (TGF-ß1) and epidermal growth factor receptor (EGFR) on the therapeutic effect of high-dose cytarabine (HD-AraC) in patients with acute myeloid leukemia (AML). METHODS: 98 patients with AML treated in our hospital from January 2019 to June 2020 were selected as the research subjects, all patients were treated with HD-AraC for 1 course of treatment every week. The effect of 2 groups were evaluated during after one course of treatment and divided into effective group and ineffective group, statistical table of baseline data was designed, the baseline data of 2 groups were counted in detail, the baseline data and serum levels of TGF-ß1 and EGFR of 2 groups were compared, Logistic regression analysis was used to examine the relationship between the levels of serum TGF-ß1, EGFR and the therapeutic effect of HD-AraC in patients with AML, the value of serum TGF-ß1 and EGFR levels in predicting the therapeutic effect of HD-AraC in AML patients was analyzed based on ROC curve and decision curve. RESULTS: After 1 course of treatment, among the 98 patients, 26 cases had complete remission, 38 cases had partially remission and 34 cases no remission, the total effective rate was 65.31% (64/98); after comparing data of 2 groups, Logistic regression analysis showed that the overexpression of serum EGFR before treatment might be a risk factor for the ineffective treatment of HD-AraC in AML patients (OR>1, P<0.05), overexpression of serum TGF-ß1 before treatment might be a protective factor for the ineffective treatment of HD-AraC in AML patients (OR<1, P<0.05); the ROC curve results showed that the AUC of serum EGFR and TGF-ß1 before treatment in predicting the risk of ineffective HD-AraC treatment in AML patients were >0.70, which had certain predictive value. The decision curve results showed that in the threshold range of 0.15-044, the prediction model combined with serum EGFR and TGF-ß1 levels in predicting the net benefit rate of HD-AraC treatment in AML patients was better than that of serum EGFR or serum TGF-ß1 alone. CONCLUSION: The levels of serum TGF-ß1 and EGFR affect the therapeutic effect of HD-AraC in patients with AML and increase the risk of ineffective treatment, serum TGF-ß1 and EGFR can be used to predict the risk of ineffective HD-AraC treatment in AML patients, and the combined prediction of net benefit rate is higher.

[Influence of Serum TGF-ß1 and EGFR Levels on the Therapeutic Effect of High-Dose AraC in Patients with Acute Myeloid Leukemia Based on the Decision Curve].

OBJECTIVE: To analyze the influence of serum levels of transforming growth factor-ß1 (TGF-ß1) and epidermal growth factor receptor (EGFR) on the therapeutic effect of high-dose cytarabine (HD-AraC) in patients with acute myeloid leukemia (AML). METHODS: 98 patients with AML treated in our hospital from January 2019 to June 2020 were selected as the research subjects, all patients were treated with HD-AraC for 1 course of treatment every week. The effect of 2 groups were evaluated during after one course of treatment and divided into effective group and ineffective group, statistical table of baseline data was designed, the baseline data of 2 groups were counted in detail, the baseline data and serum levels of TGF-ß1 and EGFR of 2 groups were compared, Logistic regression analysis was used to examine the relationship between the levels of serum TGF-ß1, EGFR and the therapeutic effect of HD-AraC in patients with AML, the value of serum TGF-ß1 and EGFR levels in predicting the therapeutic effect of HD-AraC in AML patients was analyzed based on ROC curve and decision curve. RESULTS: After 1 course of treatment, among the 98 patients, 26 cases had complete remission, 38 cases had partially remission and 34 cases no remission, the total effective rate was 65.31% (64/98); after comparing data of 2 groups, Logistic regression analysis showed that the overexpression of serum EGFR before treatment might be a risk factor for the ineffective treatment of HD-AraC in AML patients (OR>1, P<0.05), overexpression of serum TGF-ß1 before treatment might be a protective factor for the ineffective treatment of HD-AraC in AML patients (OR<1, P<0.05); the ROC curve results showed that the AUC of serum EGFR and TGF-ß1 before treatment in predicting the risk of ineffective HD-AraC treatment in AML patients were >0.70, which had certain predictive value. The decision curve results showed that in the threshold range of 0.15-044, the prediction model combined with serum EGFR and TGF-ß1 levels in predicting the net benefit rate of HD-AraC treatment in AML patients was better than that of serum EGFR or serum TGF-ß1 alone. CONCLUSION: The levels of serum TGF-ß1 and EGFR affect the therapeutic effect of HD-AraC in patients with AML and increase the risk of ineffective treatment, serum TGF-ß1 and EGFR can be used to predict the risk of ineffective HD-AraC treatment in AML patients, and the combined prediction of net benefit rate is higher.

hsa­miR­429 targets CBX8 to promote cell apoptosis in diffuse large B­cell lymphoma.

Diffuse large B­cell lymphoma (DLBCL) is the most common type of non­Hodgkin lymphoma worldwide. Several studies have indicated that Homo sapiens (hsa)­microRNA (miR)­429 exerts a tumor­suppressive effect on a variety of malignant tumors. To the best of our knowledge, the molecular function and mechanism of action of hsa­miR­429 in DLBCL have not been evaluated to date. The present study demonstrated that the expression of hsa­miR­429 in DLBCL cells was significantly reduced. hsa­miR­429 inhibited the proliferation of the DLBCL cell lines, SUDHL­4 and DB, and promoted apoptosis. A dual luciferase reporter assay was used to demonstrate that chromobox 8 (CBX8) was the target gene of hsa­miR­429. Overexpression of CBX8 promoted the proliferation of SUDHL­4 and DB cells and inhibited apoptosis, thereby playing a cancer­promoting role. Transfection of hsa­miR­429 mimic into DB cells overexpressing CBX8 antagonized the effect of CBX8 on the proliferation of DB cells. Moreover, the apoptotic rate was increased in DB cells overexpressing CBX8 and transfected with hsa­miR­429 mimic, while the proportion of cells in the G2/M phase was significantly reduced. These results demonstrated the antagonistic effect of hsa­miR­429 on the oncogenic function of CBX8. Therefore, in DLBCL, the tumor suppressor effect of hsa­miR­429 may be achieved by targeted downregulation of CBX8, suggesting that hsa­miR­429 may be used as a diagnostic marker and a potential nucleic acid drug for DLBCL. CBX8 may also represent an effective therapeutic target for DLBCL.

[Expression and Clinical Significance of CD73 in Acute Myeloid Leukemia Patients with NPM1 Mutation].

OBJECTIVE: To investigate the expression of CD73 in acute myeloid leukemia (AML) patients with NPM1 mutant and wild-type, and to evaluate the therapeutic efficacy and prognosis of CD73 to the AML patients. METHODS: 160 patients with AML treated in our hospital from June 2015 to June 2019 were enrolled, and 40 non-AML bone marrow samples from healthy people were selected as controls during the same period. The expression of CD73 in healthy people, NPM1 mutation and NPM1 wild-type AML patients were compared, and the relationship between the expression of CD73 and its clinicopathological characteristics, as while as efficacy in AML patients were analyzed. The patients were followed up, and the influence of CD73 to the prognosis of different AML patients was analyzed. RESULTS: The positive expression rate of CD73 in AML patients (23.75%) was significantly higher than that in the healthy control group (0.62%), and the positive expression rate of CD73 in AML patients with NPM1 mutation (74.75%) was significantly higher than that with NPM1 wild-type (25.51%) (both P<0.001). AML patients with CD73 positive expression was associated with age, FAB typing, disease risk classification, and NPM1 gene mutation (both P<0.05). The overall survival rate of AML patients with NPM1 gene mutation was 75.98%, which was significantly higher than the patients with NPM1 wild-type (34.68%)(P<0.001), the median survival time of AML patients with NPM1 gene mutation in the CD73+ group was 21 months, which was significantly longer than the patients in the CD73- group (11 months)(P<0.001), the median survival time of AML patients with NPM1 wild-type in the CD73+ group was 13 months, which was significantly shorter than the patients in the CD73- group (18 months) (P<0.001). CONCLUSION: The expression of CD73 was increased in AML patients with NPM1 gene mutation, and CD73 showed different prognostic significance in AML patients with different NPM1 gene mutation. The combination of clinicopathologic features, CD73 expression and NPM1 gene in AML patients is helpful to determine their prognosis and guide the formulation of relevant treatment plans.