Most methods for estimation of rates of RNA production are not applicable in human in vivo clinical studies. We describe here an approach for measuring ribosomal RNA turnover in vivo using [6,6-(2)H(2)]-glucose as a precursor for de novo RNA synthesis. Because this method involves neither radioactivity nor toxic metabolites, it is suitable for human studies.For method development in vitro, a lymphocyte cell line (PM1) was cultured in the presence of [6,6-(2)H(2)]-glucose. RNA was extracted, hydrolyzed enzymatically to ribonucleosides, and derivatized to either the aldonitrile tetra-acetate or the pentafluoro triacetate derivative of the pentose before GC-MS. We identified optimum derivatization and analysis conditions and demonstrated quantitative incorporation of deuterium from glucose into RNA of dividing cells.Pilot clinical studies demonstrated the applicability of this approach to blood leukocytes and solid tissues. A patient with chronic lymphocytic leukemia received [6,6-(2)H(2)]-glucose (1 g/kg) orally in aliquots administered every 30 min for a period of 10 h. When we analyzed CD3(-) B cells that had been purified by gradient centrifugation and magnetic-bead adhesion, we observed deuterium enrichment, a finding consistent with a ribosomal RNA production rate of about 7%/day, despite the slow division rates observed in concurrent DNA-labeling analysis. Similarly, in 2 patients with malignant infiltration of lymph nodes, administration of [6,6-(2)H(2)]-glucose (by intravenous infusion for 24 h) before excision biopsy allowed estimation of DNA and RNA turnover in lymph node samples.Our study results demonstrate the proof-of-principle that deuterium-labeled glucose may be used to analyze RNA turnover, in addition to DNA production/cell proliferation, in clinical samples.
Since their discovery just under a century ago, growth factors (GFs) have been used almost ubiquitously in haematology. Many haematological cancers are associated with bone marrow failure, either as a direct consequence of the disease or its treatment. Colony stimulating factors (CSFs) have been used to address the problems associated with the resulting cytopenias, however, concerns about the potential leukaemogenic effects of some of these CSFs led to a degree of initial hesitancy in usage, particularly in the management of acute myeloid leukaemia (AML). This has now been largely overcome. Other limitations have included cost and side effect profiles (the latter particularly with the multilineage factors). There has been wide variation locally, nationally and internationally in the usage of GFs. The American Society of Clinical Oncologists (ASCO) attempted to rationalise the usage of GFs by producing a consensus document enumerating the evidence-based indications for use of GFs. There is little information on cost effectiveness, this remains an important issue for the future. Peripheral blood stem cell transplantation (PBSCT) has revolutionised the management of many malignant conditions and has contributed to the increased use of growth factors. Many other indications are emerging for GFs used singly or in combination. Current clinical applications of GFs include: i) amelioration of cytopenias following chemotherapy and stem cell transplantation, ii) chemotherapy dose maintenance and escalation, iii) chemosensitisation and modification of disease states, iv) optimisation of methods for mobilisation of progenitor stem cells, v) immunotherapy, and vi) as therapeutic targets for treatment of haematolgical malignancies.
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