RAD001 (everolimus) induces dose-dependent changes to cell cycle regulation and modifies the cell cycle response to vincristine.

More than 50% of adults and ~20% of children with pre-B acute lymphoblastic leukemia (ALL) relapse following treatment. Dismal outcomes for patients with relapsed or refractory disease mandate novel approaches to therapy. We have previously shown that the combination of the mTOR inhibitor RAD001 (everolimus) and the chemotherapeutic agent vincristine increases the survival of non-obese diabetic/severe combined immuno-deficient (NOD/SCID) mice bearing human ALL xenografts. We have also shown that 16 µM RAD001 synergized with agents that cause DNA damage or microtubule disruption in pre-B ALL cells in vitro. Here, we demonstrate that RAD001 has dose-dependent effects on the cell cycle in ALL cells, with 1.5 µM RAD001 inhibiting pRb, Ki67 and PCNA expression and increasing G0/1 cell cycle arrest, whereas 16 µM RAD001 increases pRb, cyclin D1, Ki67 and PCNA, with no evidence of an accumulation of cells in G0/1. Transition from G2 into mitosis was promoted by 16 µM RAD001 with reduced phosphorylation of cdc2 in cells with 4 N DNA content. However, 16 µM RAD001 preferentially induced cell death in cells undergoing mitosis. When combined with vincristine, 16 µM RAD001 reduced the vincristine-induced accumulation of cells in mitosis, probably as a result of increased death in this population. Although 16 µM RAD001 weakly activated Chk1 and Chk2, it suppressed strong vincristine-induced activation of these cell cycle checkpoint regulators. We conclude that RAD001 enhances chemosensitivity at least in part through suppression of cell cycle checkpoint regulation in response to vincristine and increased progression from G2 into mitosis.

CXCR4 antagonists mobilize childhood acute lymphoblastic leukemia cells into the peripheral blood and inhibit engraftment.

The role of CXCL12 in the bone marrow (BM) homing and growth of B-cell progenitor acute lymphoblastic leukemia (ALL) has been established. However, the effect of modulating CXCL12/CXCR4 interactions on the retention of ALL cells within the supportive BM microenvironment and the expansion and dissemination of ALL cells in vivo has not been examined. We used mouse models of human childhood and murine leukemia and specific peptide and small molecule CXCR4 antagonists to examine the importance of CXCL12/CXCR4 in the development of leukemia in vivo. CXCR4 antagonists mobilized ALL cells into the peripheral blood (PB). Extended administration of CXCR4 antagonists to mice with leukemia resulted in a reduction in the number of leukemic cells in the PB and spleens of animals compared to control treated animals in three of the five cases tested. There was also a marked reduction in the dissemination of ALL cells to extramedullary sites including liver and kidney in all cases where this occurred. Considering the inhibitory effect of stromal layers on the activity of chemotherapeutic agents and the interactive effect of CXCL12 antagonists with chemotherapeutic agents in vitro, this raises the possibility of using these agents to potentiate the effects of current chemotherapy regimens.

The management of accidental dural puncture during labour epidural analgesia: a survey of UK practice.

The management of accidental dural puncture and postdural puncture headache in obstetric practice continues to be of great interest. This survey aims to explore the current management of this complication in the United Kingdom and compares the findings to a similar survey undertaken in 1993. A postal questionnaire was sent to all maternity units (n = 248). The return rate was 71%. Of these, 144 units (85%) now have written guidelines for the management of accidental dural puncture compared to 58% in 1993. In 47 units (28%), the epidural catheter is now routinely placed intrathecally following accidental dural puncture; in 69 units (41%) the catheter is re-sited and in the remaining 53 units (31%) either option is allowed. This is in contrast to the previous survey, which found that catheters were re-sited in 99% of units. Only 31 units (18%) now limit the second stage of labour and 19 (11%) avoid pushing and deliver by ventouse or forceps, whilst 116 units (69%) allow labour to take place without any intervention. Only 44 units (26%) now treat postdural puncture headache with an epidural blood patch as soon as it is diagnosed, whereas in 120 units (71%) the blood patch is performed only after failure of conservative measures. Due to the large increase in the use of the intrathecal catheter following this complication, a follow-up questionnaire was posted 5 months later to those units (n = 99) that reported this practice in the initial survey, with a 94% response rate. The two most commonly cited reasons for intrathecal catheterisation were to avoid further dural puncture (76%) and to allow immediate analgesia for labour (75%).