“Human Acute Myeloid Leukemia Is Organized as a Hierarchy That Originates from a Primitive Hematopoietic Cell” (1997), by Dominique Bonnet and John E. Dick

In the 1990s, John E. Dick and Dominique Bonnet, researchers at the University of Toronto, in Toronto, Ontario, investigated how a blood cancer called acute myeloid leukemia, or AML, arises from blood-forming cells. Researchers in the field at the time did not know which cells initiate and maintain AML in the body, and there were conflicting hypotheses about which cells were responsible. Scientists hypothesized that AML might be mainly composed of more specialized and differentiated blood cells. Dick and Bonnet conducted experiments in which they grew human AML cells in mice and investigated which cells initiated and maintained the cancer to explore that hypothesis. Their study was one of the first to use sensitive mouse models to characterize and uncover which cells initiated and comprised AML, which contributed to the understanding of its cellular origin and aided in the development of targeted therapies for leukemia treatment.

1. Background and Context
2. The Experiment
3. Impacts

Background and Context

Dick and Bonnet’s study contributed to scientists’ understanding of cancer stem cells and the role they play in initiating and maintaining cancer. Cancer stem cells are cells found in tumors that can replicate and facilitate the spread of cancer throughout the body. Like all stem cells, cancer stem cells can self-renew, a process in which they produce identical copies of themselves. They can also differentiate, a process by which a cell turns into a more specialized cell. In 1994, Dick’s research team identified cancer stem cells, and they found that cancer stem cells can play a key role in initiating and maintaining cancer after treatment.

Within the context of normal blood cell production, there exists a hierarchy of blood cells. At the top of the hierarchy are hematopoietic stem cells, or HSCs, which are blood stem cells that can develop into more specialized blood cells. Next in the hierarchy are progenitor cells, which HSCs produce. Progenitor cells are more specialized than HSCs and have a decreased ability to further specialize and differentiate. Then, progenitor cells produce the last in the hierarchy, fully differentiated and functional blood cells that make up the body’s blood system. Those blood cells include red blood cells, white blood cells, and platelets. Researchers hypothesized that a cellular hierarchy also existed within the context of blood cancers like AML, which mirrored normal blood cell hierarchy. A lack of understanding of blood cancer stem cells in the field at the time made it difficult to fully understand the various cells in the blood-forming cascade that produce AML. That gap in knowledge even made it challenging to explain the differences seen in people with leukemia, including variations in responses to treatment. Dick and Bonnet’s study investigated the hierarchy in AML and what cells were the origins of AML.

Dick received his doctoral degree from the University of Manitoba in Winnipeg, Manitoba in 1984. Afterward, Dick completed a postdoctoral fellowship at the Ontario Cancer Institute in Toronto before starting his own lab in 1986 at the University of Toronto in Toronto. According to Dick, his lab was initially interested in understanding the effects of inserting genes that cause cancer into normal stem cells in the blood and human blood cancer cells into mice. In 1993, Bonnet received her doctoral degree from the Paris Diderot University in Paris, France, where she studied human genetics. That same year, Bonnet joined Dick’s lab as a postdoctoral research fellow. Together, Dick and Bonnet began studying the role of leukemic stem cells and the AML cell hierarchy in the initiation and maintenance of AML.

At the time, one theory about the blood cancer hierarchy in AML posited that there were multiple cell types, both differentiated and undifferentiated, that could give rise to leukemic cancer cells. Another theory suggested that there could be a single undifferentiated cell that forms the basis of the hierarchy and gives rise to those leukemic cancer cells. In 1994, Dick’s lab published a study that identified a leukemia-initiating cell, or L-IC, that could initiate the growth of AML in mice. That study did not investigate the AML hierarchy.

The Experiment

Dick and Bonnet conducted their experiment to confirm which blood-forming cells give rise to AML and investigate how cells that makeup AML are organized in a hierarchy. Their goal was to use sensitive mice to inject AML cells and understand which cells grew the cancer. The first step was to determine the best type of mice to use for their experiments. They tested both severe combined immunodeficient, or SCID, mice and non-obese diabetic/severe combined immunodeficiency, or NOD/SCID, mice. Both mice have weakened immune systems, which allows cancer cells to grow in them. Dick and Bonnet tested the ability of those two types of mice to accept human AML cells and maintain the cancer. They found that they needed to inject fewer AML cancer cells in NOD/SCID mice compared to SCID mice to achieve the same number of AML cells that were growing in the mice. As a result, they decided to use NOD/SCID mice in future experiments.

Dick and Bonnet then isolated stem cells from AML patient samples using cell surface proteins as markers of the leukemic stem cells. They looked at cell surface protein markers CD34 and CD38. CD34 is typically found in blood stem cells, and CD38 is typically absent in blood stem cells. Dick and Bonnet used those markers because Dick’s lab previously showed that cells with CD34 but lacking CD38 were capable of initiating AML in mice.

After isolating AML stem cells, Dick and Bonnet injected them into mice to investigate if those cells were able to self-renew and differentiate in the cancer. They found that some of the injected cells were able to differentiate into more specialized blood cells like progenitor cells or leukemic blasts, which can sustain the cancer. They also found that about 2 percent of the originally injected AML cells kept the same CD34-positive and CD38-negative characteristics, which suggested that those had the ability to self-renew, meaning they could make more stem cells. That self-renewal ensures that a small proportion of leukemic stem cells is always present, which can then generate new cancer cells over time.

Dick and Bonnet also injected cells that had both CD34 and CD38 on their surface. Those cells would be more specialized due to the presence of CD38, which is not found on stem cells and is usually found on more differentiated cells. They found that cells with that combination of cell surface proteins did not grow in mice, suggesting leukemia could not likely start from those more specialized cells.

The team used serial transplantation of those cells in mice to further investigate the self-renewal capabilities of the cells with CD34 but lacking CD38. Serial transplantation is a process by which the team grows cells in one mouse and then transplants them into another mouse. The goal was to understand whether the leukemic stem cells were able to continuously produce new leukemia cells over multiple generations, which would indicate their self-renewal capacity. They found that after serial transplantation, there was a 30- to 100-fold expansion of the initial leukemic stem cells, further suggesting that they have the ability to self-renew.

Dick and Bonnet’s results demonstrated that cells with CD34 but lacking CD38 were likely leukemic stem cells. Their results also suggested that those leukemic stem cells that had the ability to self-renew and differentiate existed at the top of the AML hierarchy. They demonstrated that the leukemic stem cell is the cell responsible for maintaining AML growth and can produce leukemic progenitor cells and fully functional cancer cells, such as red blood cells, white blood cells, and platelets.

Impacts

Dick and Bonnet’s study showed that leukemic stem cells sit at the top of the AML hierarchy because of their ability to initiate and maintain leukemia in mice through self-renewal and differentiation. The results of the experiments challenged other theories that AML might originate from those more specialized and differentiated blood cells. According to Dick and Bonnet, through the characterization of those leukemic stem cells, there can be further investigation of genes that cause normal cells to turn into cancer cells, known as oncogenes. They assert that the investigation of those oncogenes could lead to therapies that focus on specifically targeting leukemic stem cells.

The study laid the framework for further understanding how AML originates and is maintained in the body. It further explained the AML blood cell hierarchy and what cell drives AML growth in the body. That result is crucial for understanding differences in responses to certain treatments. Additionally, the study provided further evidence for cancer stem cells existing in leukemia, which laid the grounds for research on cancer stem cells in other cancers, including in solid tumors. In one study published in 2003, researchers at the University of Michigan Medical School in Ann Arbor, Michigan expanded on Dick and Bonnet’s study and identified one of the first cancer stem cells in breast cancer. Dick and Bonnet’s study is highly cited in the field of cancer research and therapy, with more than 9,700 citations as of 2025.

Sources

1. Bonnet, Dominique, and John E. Dick. “Human Acute Myeloid Leukemia Is Organized as a Hierarchy That Originates from a Primitive Hematopoietic Cell.” Nature Medicine 3 (1997): 730–7.
2. Canadian Medical Hall of Fame. “John E. Dick, PhD” Canadian Medical Hall of Fame. https://www.cdnmedhall.ca/laureates/johndick (Accessed May 26, 2025).
3. Chu, Xianjing, Wentao Tian, Jiaoyang Ning, Gang Xiao, Yunqi Zhou, Ziqi Wang, Zhuofan Zhai, Guilong Tanzhu, Jie Yang, and Rongrong Zhou. “Cancer Stem Cells: Advances in Knowledge and Implications for Cancer Therapy.” Signal Transduction and Targeted Therapy 9 (2024): 170. https://pmc.ncbi.nlm.nih.gov/articles/PMC11224386/ (Accessed May 26, 2025).
4. Lapidot, Tsvee, Christian Sirard, Josef Vormoor, Barbara Murdoch, Trang Hoang, Julio Caceres-Cortes, Mark Minden, Bruce Paterson, Michael A. Caligiuri, and John E. Dick. “A Cell Initiating Human Acute Myeloid Leukemia After Transplantation Into SCID Mice.” Nature 367 (1994): 645–8. https://d1wqtxts1xzle7.cloudfront.net/62875691/AML_stem_cell__-_first_ever_paper_on_CSC20200408-51194-1h5uwug-libre.pdf?1586363327=&response-content-disposition=inline%3B+filename%3DAML_stem_cell_first_ever_paper_on_CSC.pdf&Expires=1744677061&Signature=eZVUocpkwueMUyAx8nN8Qtgw3KhX1pCWybM91mme3z4G2JCH4Ai0OtpWLkXdX-W~fu-MwXwnKikwTDe5XMBnE8Qw2MvJBuj4NLfp4itC7HrueRBAHNLYf-n8apJxeEN6tY4WW~BfYiBGslwinItA4Qpiu-fIgIE8gp4vf4rGGxAzvJBtFvGUajlsV8OAmToDYrAEgE~VsQKm0fvcti5BeHc57RPpu0U8frnhunQwJ5QjCLDZ8wRcsRYYVOfcG1GVCfttT9Z5ZEqw~ZS~uAJnP4xcuvgtwBUHXBe-i21jzW0T~1JbHic8ZnMjD80rwlmZbuQPfcAIoDDlpO67WXaEWw__&Key-Pair-Id=APKAJLOHF5GGSLRBV4ZA (Accessed May 26, 2025).
5. The Francis Crick Institute. “Dominique Bonnet.” The Francis Crick Institute. https://www.crick.ac.uk/research/find-a-researcher/dominique-bonnet (Accessed May 26, 2025).
6. University Health Network. “Dr. John Dick on His Famous Discovery of Cancer Stem Cells, and the Puzzle of Research.” University of Toronto. https://www.uhn.ca/corporate/News/UHN_Podcasts/Behind_the_Breakthrough/Pages/Season2_Episode10_John_Dick.aspx (Accessed May 26, 2025).
7. Zeng, Andy G. X., Suraj Bansal, Liqing Jin, Amanda Mitchell, Weihsu Claire Chen, Hussein A. Abbas, Michelle Chan-Seng-Yue, Veronique Voisin, Peter van Galen, Anne Tierens, Meyling Cheok, Claude Preudhomme, Hervé Dombret, Naval Daver, P. Andrew Futreal, Mark D. Minden, James A. Kennedy, Jean C. Y. Wang, and John E. Dick. “A Cellular Hierarchy Framework for Understanding Heterogeneity and Predicting Drug Response in Acute Myeloid Leukemia.” Nature Medicine 28 (2022): 1212–23.