We are interested in genetic models of leukemogenesis, particularly the highly conserved anti-apoptotic transcriptional pathway downstream of E2A-HLF, a chimeric oncoprotein activated by chromosomal translocation in childhood leukemia. Sequence homology between the HLF transcription factor and CES-2, a cell death specification protein in the nematode Caenorhabditis elegans, suggests that this pathway is not unique to developing B-lymphocytes, but has been evolutionarily conserved in diverse organisms. Recent evidence suggests that this pathway in mammalian cells involves Slug, a zinc-finger transcription factor that is highly related to CES-1, the gene located downstream of CES-2 in the worm. We are now focusing on the role of Slug in apoptosis, using mice with targeted disruption of the Slug gene. Our goal is to understand pathways of cell death specification in mammals --how they connect to the common machinery of programmed cell death and how they can be disrupted in malignant transformation.

Zebrafish Models of Cancer

Our group is working on cancer genetics, using the zebrafish genetic system to clarify developmental pathways subverted in human leukemias and solid tumors. The zebrafish animal model provides a powerful system for genetic analysis of vertebrate embryogenesis, organ development and disease. This model is unique within vertebrates in its capacity for "forward" genetic analysis, through use of phenotype-driven mutational screens and readily accessible transparent embryos. These properties make the zebrafish an ideal system for gene discovery based on gene function, an advantage that is very useful in dissecting pathways of gene action and identifying genes that are either activated (oncogenes) or inactivated (tumor suppressors) during malignant transformation.

We are currently conducting a genome-wide mutagenesis screen to identify genes required for normal myeloid cell development in the hematopoietic system. We have shown that zebrafish myelopoiesis is very similar to that in humans and other mammals, indicating that this screen should reveal conserved signal transduction pathways involved in normal vertebrate myeloid cell development. A subset of the mutations identified by this approach should correspond to genes whose human counterparts contribute to the differentiation arrest of cells in the myeloid lineage characteristic of two important human diseases - myelodysplastic syndrome and acute myeloid leukemia.

A second screen is underway to isolate genes that are essential for normal embryologic development of the peripheral sympathetic nervous system. A subset of the genes discovered in this screen should function as tumor suppressors in neuroblastoma, an enigmatic tumor whose spontaneous regression in infants and relentless progression in older children pose daunting problems to therapists and experimental oncologists alike. Despite major modifications of therapy over the past two decades, the long-term cure rate in children with advanced disease is still far from satisfactory.

A third area involves both transgenic and mutagenesis approaches in zebrafish to dissect pathways that lead to T-cell leukemia. We have shown that human T-cell leukemias can be divided into five major subtypes based on the expression of oncogenes that initiate malignant transformation in thymocytes. We are overexpressing these oncogenes in zebrafish T-cell progenitors using the Rag2 gene promoter to generate T-cell leukemia and thymic lymphoma in zebrafish. Complementary genetic screens are being employed to identify tumor suppressor genes whose mutational inactivation contributes to malignant transformation in each of the five oncogenic pathways.

Chemical and genetic modifier screens using tumor-prone zebrafish lines may ultimately reveal mutant genes or drugs that can suppress or modify disease progression. For example, we hope to identify mutated genes that promote specific aspects of the malignant phenotype, such as genomic instability, metastasis or invasiveness. We also hope to discover mutations or drugs that delay or totally suppress the onset tumors in transgenic zebrafish lines, thus providing candidate targets for the development of new therapies.

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Models of Conserved Cell Death Specification Pathways
and the Role of E2A-HLF in Leukemogenesis