TP53 and Decitabine in Acute Myeloid Leukemia and Myelodysplastic Syndromes.

BACKGROUND: The molecular determinants of clinical responses to decitabine therapy in patients with acute myeloid leukemia (AML) or myelodysplastic syndromes (MDS) are unclear. METHODS: We enrolled 84 adult patients with AML or MDS in a single-institution trial of decitabine to identify somatic mutations and their relationships to clinical responses. Decitabine was administered at a dose of 20 mg per square meter of body-surface area per day for 10 consecutive days in monthly cycles. We performed enhanced exome or gene-panel sequencing in 67 of these patients and serial sequencing at multiple time points to evaluate patterns of mutation clearance in 54 patients. An extension cohort included 32 additional patients who received decitabine in different protocols. RESULTS: Of the 116 patients, 53 (46%) had bone marrow blast clearance (<5% blasts). Response rates were higher among patients with an unfavorable-risk cytogenetic profile than among patients with an intermediate-risk or favorable-risk cytogenetic profile (29 of 43 patients [67%] vs. 24 of 71 patients [34%], P<0.001) and among patients with TP53 mutations than among patients with wild-type TP53 (21 of 21 [100%] vs. 32 of 78 [41%], P<0.001). Previous studies have consistently shown that patients with an unfavorable-risk cytogenetic profile and TP53 mutations who receive conventional chemotherapy have poor outcomes. However, in this study of 10-day courses of decitabine, neither of these risk factors was associated with a lower rate of overall survival than the rate of survival among study patients with intermediate-risk cytogenetic profiles. CONCLUSIONS: Patients with AML and MDS who had cytogenetic abnormalities associated with unfavorable risk, TP53 mutations, or both had favorable clinical responses and robust (but incomplete) mutation clearance after receiving serial 10-day courses of decitabine. Although these responses were not durable, they resulted in rates of overall survival that were similar to those among patients with AML who had an intermediate-risk cytogenetic profile and who also received serial 10-day courses of decitabine. (Funded by the National Cancer Institute and others; ClinicalTrials.gov number, NCT01687400 .).

RBF and Rno promote photoreceptor differentiation onset through modulating EGFR signaling in the Drosophila developing eye.

The retinoblastoma gene Rb is the prototype tumor suppressor and is conserved in Drosophila. We use the developing fly retina as a model system to investigate the role of Drosophila Rb (rbf) during differentiation. This report shows that mutation of rbf and rhinoceros (rno), which encodes a PHD domain protein, leads to a synergistic delay in photoreceptor cell differentiation in the developing eye disc. We show that this differentiation delay phenotype is caused by decreased levels of different components of the Epidermal Growth Factor Receptor (EGFR) signaling pathway in the absence of rbf and rno. We show that rbf is required for normal expression of Rhomboid proteins and activation of MAP kinase in the morphogenetic furrow (MF), while rno is required for the expression of Pointed (Pnt) and Ebi proteins, which are key factors that mediate EGFR signaling output in the nucleus. Interestingly, while removing the transcription activation function of dE2F1 is sufficient to suppress the synergistic differentiation delay, a mutant form of de2f1 that disrupts the binding with RBF but retains the transcription activation function does not mimic the effect of rbf loss. These observations suggest that RBF has additional functions besides dE2F1 binding that regulates EGFR signaling and photoreceptor differentiation.

Retinoblastoma family protein promotes normal R8-photoreceptor differentiation in the absence of rhinoceros by inhibiting dE2F1 activity.

The retinoblastoma gene Rb is a prototype tumor suppressor which is conserved in Drosophila. Although much is known about the roles of Rb in cell proliferation and apoptosis, much less is known about how Rb regulates cell differentiation. Inactivation of Drosophila Rb (rbf) exhibited subtle differentiation defects similar to inactivation of Rb in mice, suggesting the existence of redundant mechanisms in the control of cell differentiation. To test this possibility and to characterize the role of Rbf in cell differentiation during retinal development, we carried out a genetic screen and identified a mutation in rhinoceros (rno), which leads to synergistic differentiation defects in conjunction with rbf inactivation. Characterization of an early differentiation defect, the multiple-R8 phenotype, revealed that this phenotype was caused by limiting amounts of Notch signaling due to reduced expression of the Notch ligand, Delta (Dl). Decreasing the gene dosage of Dl enhanced the multiple-R8 phenotype, while increasing the level of Dl suppressed this phenotype. Interestingly, removal of the transcriptional activation of dE2F1 partially restores Dl expression in rbf,rno mutant clones and suppresses the associated differentiation defects, indicating that this differentiation function of RBF is mediated by its regulation of dE2F1 activity.

Proneural basic helix-loop-helix proteins and epidermal growth factor receptor signaling coordinately regulate cell type specification and cdk inhibitor expression during development.

Cell differentiation and cell cycle exit are coordinately regulated during development; however, the molecular logic underlying this regulation is not known. The Drosophila cdk inhibitor Dacapo (Dap) is one of the key cell cycle regulators that exhibit dynamic expression during development and contribute to the developmental regulation of the cell cycle. In this study, regulation of Dap expression during cell type specification was investigated. The expression of Dap in the R2 and R5 precursors of the developing eye and in the newly recruited leg disc femoral sense organ precursors was found to be controlled by the epidermal growth factor receptor signaling-regulated transcription factor Pointed (Pnt) and the proneural basic helix-loop-helix proteins Atonal (Ato) and Daughterless (Da). We show that Pnt, Ato, and Da regulate Dap expression directly through their respective binding sites precisely at the time when these transcription factors function to specify neural fates. These results show that Dap expression is directly regulated by developmental mechanisms that simultaneously control cell type specification. This is potentially a general mechanism by which the expression of key cell cycle regulators is coordinated with differentiation during normal development. The direct regulation of key cell cycle regulators by the differentiation factors ensures coordinated regulation of cell cycle and differentiation.

Control of cell cycle entry and exiting from the second mitotic wave in the Drosophila developing eye.

BACKGROUND: In the morphogenetic furrow (MF) of the Drosophila developing eye, all cells arrest in G1 and photoreceptor cell differentiation initiates. As the cells exit the MF, Notch signaling is required for the uncommitted cells to enter a synchronous round of cell division referred to as the "second mitotic wave" (SMW). How cell cycle entry and exit in SMW is regulated remains unclear. Recent studies have suggested that Notch signaling controls S phase in the SMW by regulating Cyclin A and the E2F transcription factor independent of Cyclin E. In this manuscript, we investigate the developmental regulation of cell cycle entry into and exit from SMW. RESULTS: We demonstrate here that Cyclin E-dependent kinase activity is required for S phase entry in SMW. We show that removal of Su(H), a key transcription factor downstream of Notch signaling, blocks G1/S transition in SMW with strong upregulation of the Cyclin E/Cdk2 inhibitor Dacapo (Dap). We further show that the upregulation of Dap, which is mediated by bHLH protein Daughterless (Da), is important for cell cycle arrest of Su(H) mutant cells in SMW. Finally we show that removal of Dap leads to additional cell proliferation and an accumulation of the non-photoreceptor cells in the Drosophila developing eye. CONCLUSION: Our data demonstrate that Cyclin E/Cdk2 kinase activity is absolutely required for S phase in SMW, and that Dap is required for the proper cell cycle arrest of cells exiting the SMW. In addition, our results suggest that the G1 arrest of notch and Su(H) mutant cells in the SMW are regulated by distinct mechanisms, and that the upregulation of Dap contributes the G1 arrest of Su(H) mutant cells.