Construction of a Novel CD34⁺ Hematopoietic Stem Cell Derived Xenograft Model of Human Hyperleukocytic Acute Myeloid Leukemia

Acute myeloid leukemia (AML) is an abnormal clonal proliferative disease of hematopoietic progenitors in the bone marrow, which is the most common malignant disease in the blood system. Hyperleukocytic leukemia (HLL) is one of the high-risk types and commonly defined as peripheral blood leukocyte counts exceeding 50-100×10⁹/l. The patients with hyperleukocytic acute myeloid leukemia (HL-AML) usually suffer from poor prognosis due to low responsiveness to initial chemotherapeutic agents, so these patients have higher mortality with life-threatening complication than those without hyperleukocytosis. However, there is no available patient derived xenograft model (PDX) in HL-AML, especially the CD34+ hematopoietic stem cell derived xenograft model of human HL-AML.

In this study, firstly, the primary HL-AML cells were collected and separated from the patients who underwent leukapheresis by using Fresenius COM.TEC machine, and the CD34⁺ hematopoietic stem cells were isolated from primary HL-AML cells using magnetic beads. Then, the NOD-SCID-IL2rγ^null (B-NSG) mice were treated with 1.5 Gy χ-ray for total body irradiation at day 0, and intravenously injected with isolated 1.5 × 10⁷ HLL cells or 1.5 × 10⁶ CD34⁺ cells mixed with G-CSF within 24 h of irradiation. After inoculation, the blood smear, bone marrow smear, immunophenotyping and histopathology were used to test whether the model was successfully constructed.

For construction of hematopoietic stem cell derived xenograft model in HL-AML, at day 9, the immunophenotypes of hCD45+ and hCD34+ cells were found and had higher ratios in CD34+ group than HLL group in peripheral blood, which suggested the model was successfully established. The body weight of mice in CD34+ group were decreased faster than the HLL group due to severe tumor load and the leukocyte counts were increased higher than the correspondent HLL group after cell inoculation. In CD34+ group, the blood routine examination had changed, especially the WBC counts and PLT counts. On the 25th day, leukemic cells were found by the blood smears analysis and detected by flow cytometry, and which were more obviously found in bone marrow, and the immunophenotypes ratio of hCD45⁺ and hCD34⁺ were increased in CD34⁺ group. A lot of CD34+ cells were infiltrated into bone marrow, liver and spleen tissues on the 25th day that detected by histopathological analysis, which indicated that the more obviously model construction using hematopoietic stem cell derived leukemic cell.

In a conclusion, we successfully constructed a novel CD34+ hematopoietic stem cells derived animal model of human hyperleukocytic AML, which is available for mechanism research and drug treatment.