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Primary acute myelogenous leukaemia (AML) samples are heterogeneous in clonogenicity, both among patients and within the leukaemic cell population of a single patient. To explain this heterogeneity the leukaemia stem cell model postulates that leukaemic hematopoiesis is organized in a hierarchy, sustained by leukaemia stem cells that may either self-renew or differentiate aberrantly to give rise to blasts that can no longer proliferate. This process is akin to the irreversible growth arrest entered by terminally differentiating normal blood cells. We wished to identify genes associated with clonogenicity in AML. To obtain pure populations of cells of defined growth abilities, we analyzed low passage cultures of the cell line OCI-AML4. This cell line resembles primary AML cells in several important respects; it is growth factor-dependent, contains a low proportion of clonogenic cells, and has a relatively simple karyotype. Clones consisting of four cells were micromanipulated so that a single cell was sampled for global RT-PCR while its three clonal siblings served as reporters of clonogenicity. By microarray analysis we found the orphan nuclear receptor EAR-2 to be expressed four-fold lower in leukemia single cells that spontaneously lose proliferative ability, compared to single cells with greater proliferative capacity. EAR-2 is a member of the COUP transcription factor family, which play roles in various developmental processes through interactions with nuclear receptors and other transcription factors. We assessed expression of EAR-2 in monoblastic leukaemia U937 cells induced to differentiate with a variety of induction agents. Treatment with dimethylsulfoxide, phorbol ester, vitamin D3, and all trans retinoic acid (ATRA) all induced significant decreases in EAR-2 expression. This phenomenon was also seen in a mouse model of acute promyelocytic leukaemia (APL). When primary bone marrow cultures of hCG-NuMA-RAR transgenic mice were induced to differentiate with ATRA, an average decrement in EAR-2 expression of 5.58 fold was observed (p To characterize the effect of forced expression of EAR-2 on clonogenicity we transduced U937 cells with a retrovirus encoding either EAR-2 (U937-EAR2) or EGFP (U937-GFP). Analysis of FACS-purified U937-EAR2 and U937-GFP cultures showed that forced expression of EAR-2 reduces the doubling time of these populations (U937-EAR2 = 24h; U937-GFP = 34h; p<< 0.001), while no significant difference was observed in cell cycle profile. The decrease in doubling time of U937-EAR2 cells may reflect a decrease in the rate of cell loss in the population, consistent with the hypothesis that EAR-2 functions as a repressor of terminal differentiation. We have observed that expression of the orphan nuclear receptor EAR-2 is positively associated with maintenance of proliferative capacity and negatively associated with differentiation. These observations establish the importance of EAR-2 in the regulation of clonogenicity and terminal differention.

8277 Background: Older patients with AML often have a poor prognosis and often do not tolerate intensive chemotherapy. When weighing alternate management strategies, impact upon QOL is an important consideration. However, there is limited published research in this area. Methods: Patients aged 60 or older with newly diagnosed AML were enrolled at diagnosis regardless of treatment intent. Patients were excluded if life expectancy was less than one month or they did not speak English. QOL was measured at baseline and at 1, 4, and 6 months. Questionnaires included the European Organization for the Research and Treatment of Cancer QLQ-C30 (global health, 5 QOL domains, 3 symptom domains) and the Functional Assessment of Cancer Therapy Anemia and Fatigue subscale. Demographic and clinical information was also recorded. Patients were stratified by intensive versus non-intensive treatment. Results: From June to December 2003, 14 patients (mean age 73) were enrolled. At baseline, global health, role function, and...

8273 Background: Physical and psychological function in older patients with AML has not been studied previously. It is important to understand whether intensive treatment of leukemia adversely affects functional status in older patients. We evaluated the functional, mood, and cognitive changes over time in older patients with AML. Methods: All patients age 60 or greater with a new diagnosis of de novo or secondary AML were approached. Exclusion criteria were another actively treated malignancy, lack of English fluency and life expectancy less than 1 month. Baseline patient data collected included personal and medical information. Activities of daily living (ADL) were assessed by the Barthel Questionnaire and instrumental activities of daily living (IADL) by the Lawton-Brody tool. The Geriatric Depression Scale (GDS) was given to assess mood. Cognition was assessed via the Folstein Mini Mental Status Examination (MMSE). Follow up assessments were obtained 1, 4, and 6 months after entry into the study. Resu...

8183 Background: QOL is becoming increasingly important in oncology and is usually measured using general cancer scales and symptom-specific scales. AML is an aggressive hematological tumor that affects primarily older adults and often responds poorly to standard treatment. Understanding QOL will assist in treatment decision-making, symptom palliation, and evaluation of new biologic agents. Yet there is no validated English-language symptom module for use in AML. Methods: In this pilot study, we set out to translate and perform English-language validation of an existing German symptom-specific module for AML designed by Schumacher and Kuchler for use with the European Organization for the Research and Treatment of Cancer (EORTC) QLQ-C30 (Leukemia 1998; 12:586). Forward-backward translation of the 20-item module (7 subscales) was performed. Face and content validation of the English module were assessed by a panel of hematologists and QOL researchers. The module was then administered along with the QLQ-C30...

Mouse embryos lacking the receptor tyrosine kinase, Flk1, die without mature endothelial and hematopoietic cells. To investigate the role of Flk1 during vasculogenesis and hematopoiesis, we examined the developmental potential of Flk1 −/− embryonic stem cells in chimeras. We show that Flk1 is required cell autonomously for endothelial development. Furthermore, Flk1 −/− cells do not contribute to primitive hematopoiesis in chimeric yolk sacs or definitive hematopoiesis in adult chimeras and chimeric fetal livers. We also demonstrate that cells lacking Flk1 are unable to reach the correct location to form blood islands, suggesting that Flk1 is involved in the movement of cells from the posterior primitive streak to the yolk sac and, possibly, to the intraembryonic sites of early hematopoiesis.