Zhisheng Her1, Kylie Su Mei Yong1, Kathirvel Paramasivam1, Wilson Wei Sheng Tan1, Xue Ying Chan1, Sue Yee Tan1, Min Liu1,2, Yong Fan3, Yeh Ching Linn4, Kam Man Hui1,5, Uttam Surana1,6,7*, and Qingfeng Chen1,3,5,8*.
1 Institute of Molecular and Cell Biology, Agency for Science, Technology and Research (A*STAR), Singapore;
2 Department of Medicine, Yong Loo Lin School of Medicine, National University of Singapore, Singapore;
3 Key Laboratory for Major Obstetric Diseases of Guangdong Province, The Third Affiliated Hospital of Guangzhou Medical University, Guangzhou, 510150, China;
4 Department of Haematology, Singapore General Hospital, Singapore;
5 Division of Cellular and Molecular Research, National Cancer Centre, Singapore;
6 Department of Pharmacology, National University of Singapore, Singapore
7 Bioprocessing Technology Institute, Agency for Science, Technology and Research, Singapore;
8 Department of Microbiology and Immunology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.
* Corresponding author: Qingfeng Chen.
E-mail: firstname.lastname@example.org. Uttam Surana, email: email@example.com;
Published in Journal of Hematology and Oncology on 30 August 2017.
Background: Xenotransplantation of patient-derived AML (acute myeloid leukemia) cells in NOD-scid Il2rγnull (NGS) mice is the method of choice for evaluating this human hematologic malignancy. However, existing models constructed using intravenous injection in adult or newborn NSG mice have inferior engraftment efficiency, poor peripheral blood engraftment or are difficult to construct.
Methods: Here, we describe an improved AML xenograft model where primary human AML cells were injected into NSG newborn pups intrahepatically.
Results: Introduction of primary cells from AML patients resulted in high levels of engraftment in peripheral blood, spleen and bone marrow (BM) of recipient mice. The phenotype of engrafted AML cells remained unaltered during serial transplantation. The mice developed features that are consistent with human AML including spleen enlargement and infiltration of AML cells into multiple organs. Importantly, we demonstrated that although leukemic stem cell activity is enriched and mediated by CD34+CD117+ subpopulation, CD34+CD117- subpopulation can acquire CD34+CD117+ phenotype through de-differentiation. Lastly, we evaluated the therapeutic potential of Sorafenib and Regorafenib in this AML model and found that periphery and spleen AML cells are sensitive to these treatments, whereas BM provides a protective environment to AML.
Conclusions: Collectively, our improved model is robust, easy-to-construct and reliable for pre-clinical AML studies.
Figure legend:In vivo Sorafenib and Regorafenib treatment can efficiently eliminate AML in peripheral but not in bone marrow. Magnetically sorted CD34+ pooled BM cells and splenocytes from secondary engrafted NSG mice were injected intrahepatically in newborn NSG pups after sublethal irradiation (1 × 105 cells per pup). Successfully engrafted mice with more than 30 human CD45+ cells per μl of blood (between week 12 to 16 post-engraftment) were randomly assigned to either untreated (n = 3), Regorafenib (n = 6; 5 mg/kg body weight; gavage-fed once daily) or Sorafenib (n = 6; 10 mg/kg body weight; gavage-fed once daily) treatment groups and monitored for one month. a Longitudinal effect of Regorafenib and Sorafenib treatment on peripheral blood engraftment at week 0, 2 and 4 post-drug treatment. Change in AML engraftment for each group at each time point is expressed as fold change relative to the absolute human CD45+ count per μl of blood at week 0 post-drug treatment. Data are presented as mean fold change ± SEM. Two-tailed Mann Whitney U test; *; p < 0.05. b Soft tissue sarcoma and reduced spleen size were observed in Regorafenib- or Sorafenib-treated mice. Representative images of abdomen and spleen was shown; scale bar: 1 cm. Weight of spleen are presented as mean ± SEM. Two-tailed Mann Whitney U test; *; p < 0.05. c Comparison of the absolute count of human CD45+ cells (c, above) and CD34+ cells (c, below) in peripheral blood, spleen and BM between different treatment groups after 4 weeks post-drug treatment. Data are presented as mean absolute count ± SEM. Two-tailed Mann Whitney U test; *; p < 0.05.
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