Zebrafish have emerged as a powerful model of development and cancer. Human, mouse, and zebrafish malignancies exhibit striking histopathologic and molecular similarities, underscoring the remarkable conservation of genetic pathways required to induce cancer. Zebrafish are uniquely suited for large-scale studies in which hundreds of animals can be used to investigate cancer processes. Moreover, zebrafish are small in size, optically clear during development, and amenable to genetic manipulation. Facile transgenic approaches and new technologies in gene inactivation have provided much needed genomic resources to interrogate the function of specific oncogenic and tumor suppressor pathways in cancer. This manuscript focuses on the unique attribute of labeling leukemia cells with fluorescent proteins and directly visualizing cancer processes in vivo including tumor growth, dissemination, and intravasation into the vasculature. We will also discuss the use of fluorescent transgenic approaches and cell transplantation to assess leukemia-propagating cell frequency and response to chemotherapy. Zebrafish Models of Leukemia Zebrafish models of hematological malignancies exhibit striking similarities with human and mouse disease Although characterized by increased circulating white blood counts, chronic leukemias are often much slower growing and take months or years to progress. Leukemias can be further subdivided based on the blood lineage in which cells have become transformed Zebrafish first emerged as a powerful genetic model of leukemia with the description of transgenic approaches in which cMYC was overexpressed in developing thymocytes Advances in Hematology Moreover, GFP+ thymocytes exhibited stereotypical homing to the nasal placode, periocular space, and kidney marrow when assessed by serial fluorescent imaging over days Many exciting new models of hematopoietic malignancy have been created including B-cell acute lymphoblastic leukemia (B-ALL), acute myeloid leukemia (AML), and myeloproliferative neoplasm Fluorescent Transgenic Approaches to Label Leukemia Cells Cell Transplantation Approaches to Visualize Tumor Cell Engraftment Investigators have utilized cell transplantation of fluorescently labeled cancer cells into sublethally irradiated adult zebrafish to assess tumorigenicity Cell Transplantation Approaches to Examine Tumor Cell Homing and Intravasation into Vessels Intravasation of cancer cells into the vasculature is a critical step in cancer progression, allowing the spread of tumor cells beyond the site of origin Fluorescence Imaging to Visualize Leukemia Responses to Drug Treatment and Gamma-Irradiation Fluorescence imaging of transplanted cancer cells can also be used to visualize response to chemotherapy and radiation. For example, the Revskoy group recently showed that GFP-labeled T-ALL cells could be serially transplanted into syngeneic strain larvae Cell Transplantation Approaches to Quantify Leukemia Propagating Cell Frequency and Aggression Leukemia-propagating cells (LPCs) have the capacity to produce all the other cell types contained within the leukemia, are responsible for continued tumor growth, and ultimately drive relapse. Investigators have used fluorescence-activated cell sorting (FACS) to identify unique cell populations and limiting dilution cell transplantation to assess if molecularly defined leukemia cells retain LPC activity in human disease. For example, in AML a rare CD34+, CD38− cell enriches for leukemia-propagating potential Conclusion and Challenges for the Future Zebrafish has fast emerged as a powerful model of leukemia. When coupled with fluorescent transgenic approaches and powerful imaging techniques, these models are uniquely positioned to uncover mechanisms driving tumor dissemination, progression, and relapse. Moreover, the use of multifluorescent transgenic animals will allow for labeling of tumor cell compartments similar to those defined in RASinduced rhabdomyosarcoma model
