Role of ITGA5 in Malignant Proliferation and Infiltration of Acute Myeloid Leukemia

Abstract Introduction: Acute myeloid leukemia is a clonal hematopoietic stem cell malignancy, and excessive leukemic cell migration into the peripheral blood can lead to fatal complications when leukemic cells in the bone marrow proliferate and infiltrate malignantly. Integrins are a class of adhesion molecules expressed on hematopoietic cells and leukemia cells. At present, it is believed that the interaction between integrins and their ligands also plays an important role in the migration and adhesion of leukemia cells. We performed proteomic sequencing analysis of AML bone marrow samples in our previous work and found that integrin ITGA5 expression was significantly increased in AML leukemia cells compared with normal subjects, and combined with previous work basis and literature reports we speculated that integrin molecule ITGA5 may be involved in the process of malignant proliferation and infiltration of AML.

Methods: We collected bone marrow cells from AML patients and healthy controls for single-cell transcriptome and proteomics sequencing.Bone marrow CD34 + cells and AML cell lines from normal subjects and AML patients were collected, and integrin gene level expression was verified by PCR.We knocked down the ITGA5 gene in AML cell lines and analyzed ITGA5 expression by flow cytometry.AML cell lines were treated with anti-human ITGA5 monoclonal antibody and ITGA5 inhibitor ATN161, respectively, to determine the effects on apoptosis, migration, invasion, and adhesion, in addition, the surface molecular force of the cell lines was determined by a novel molecular fluorescence tension probe based on DNA structure.Using the MLL-AF9 mouse model, we tested the effect of blocking ITGA5 in combination with cytarabine chemotherapy.The doses of mice in each group were as follows: ATN161 (50 mg/kg) + cytarabine (50 mg/kg), ATN161 (50 mg/kg), cytarabine (50 mg/kg), and normal saline.

Results: Proteomic sequencing analysis of bone marrow samples revealed that integrin ITGA5 expression was significantly increased in AML leukemia cells compared with normal subjects.RNA were extracted from AML cell lines Kasumi-1, MV411, MOLM-13, and bone marrow CD34 + cells from AML patients and detected by QPCR, and ITGA5 expression was significantly increased in AML cell lines and AML patient cells compared with normal controls.ITGA5 expression in AML cells was inhibited by either ITGA5 monoclonal antibody or ITGA5 inhibitor ATN161, and migration and invasion of AML cell lines were significantly decreased, but the effect on apoptosis was not very prominent.Using a novel molecular fluorescent tension probe based on DNA structure to visualize the mechanical signal of viable cells, AML cell lines, AML patients CD34 + can produce a strong signal on the base compared with controls, and the force signal generated on the base is significantly inhibited after the use of ATN161.We constructed a MLL-AF9 leukemia mouse model and found that ATN161 could significantly reduce the proportion of leukemic cells in the liver, spleen, and bone marrow of leukemia mice and effectively alleviate the malignant proliferation and infiltration of leukemia cells by ITGA5 inhibitor ATN161 combined with cytarabine treatment, and the therapeutic effect was more obvious after combination therapy.

Conclusion: ITGA5 plays an important role in the migration, invasion and adhesion of leukemia cells. Inhibition of ITGA5 by ATN16 combined with chemotherapy can significantly play a better therapeutic role, and inhibit extramedullary infiltration of leukemia cells. Our findings will contribute to a better understanding of the role of ITGA5 in malignant proliferative infiltration of AML and the role of ITGA5 as a preclinical target in AML.

Disclosures No relevant conflicts of interest to declare.