A single nucleotide polymorphism in the coding region of ABL and its effects on sensitivity to imatinib.

We have identified a gene polymorphism (K247R) within or close to the P-loop of BCR-ABL, which leads to the substitution of arginine for lysine. We investigated the allelic frequency of K247R by screening 157 CML patients and 213 healthy blood donors with conventional sequencing, restriction enzyme digest and single strand conformational polymorphism analysis, and found the arginine allele to be rare. Three out of five CML patients with the arginine allele of K247R failed to achieve a major cytogenetic response to imatinib, suggesting that the arginine allele may have reduced sensitivity. However, despite K247R's position in or near to the P-loop, biochemical and cellular assays of imatinib and dasatinib sensitivity showed no alteration compared to wild type. Clinicians should be aware that possession of the arginine allele of K247R does not reflect a mutation that necessitates a change in the therapeutic strategy, unless there are other signs of inadequate response to drug.

Antisense BCR-ABL oligomers cause non-specific inhibition of chronic myeloid leukemia cell lines.

We have examined the effects of antisense oligomers (AOs) of various lengths, sequences and chemistry on the proliferation of eight different cell lines, five derived from patients with chronic myelogenous leukemia (CML) and three from other sources. In general, phosphodiester AOs were inactive, presumably due to degradation by nucleases present in fetal calf serum. Both BA2 and B3A2 phosphorothiolate AOs (but not corresponding sense oligomers) significantly inhibited the proliferation of three CML cell lines (BV173, LAMA84, and KYO1), but the effect was independent of the type of breakpoint expressed by each cell line, suggesting that the inhibition was sequence dependent but not sequence specific. The CML cell lines tested showed different sensitivities to inhibition of proliferation by AOs--lines with defective expression of the normal ABL protooncogene (e.g. BV173) were more readily inhibited than lines with a normal ABL message (e.g. K562). We conclude that further studies are necessary to delineate the precise mechanism(s) by which CML cell proliferation is inhibited by AOs.

Prognostic factors for patients with chronic myeloid leukaemia in chronic phase treated with imatinib mesylate after failure of interferon alfa.

We assessed clinical results in 145 patients with chronic myeloid leukaemia in chronic phase who satisfied criteria for interferon-alpha failure and were thus eligible for treatment with imatinib at the Hammersmith Hospital. We used univariate and multivariate analyses to develop a risk score based on features defined after treatment for 3 months. We identified a low neutrophil count and poor cytogenetic response (<35% Ph-negative marrow metaphases) at 3 months as principal independent predictive factors and incorporated them into a three-tier prognostic scoring system for individual patients. For patients in the low-, intermediate- and high-risk groups, the probabilities of survival at 24 months were 100, 82 and 40% (P<0.0001) and progression-free survival 100, 66 and 15% (P<0.0001), respectively. This Hammersmith prognostic scoring system was validated with an independent cohort of patients treated at another UK centre.

Autografting in chronic myeloid leukaemia.

Autografting in chronic myeloid leukaemia (CML) has been performed at various stages in the disease using different regimens for the past 20 years. Early experience suggested that autografting in blastic transformation could prolong survival and, as with allogeneic transplantation, the use of autografting was then gradually extended to the chronic phase of the disease. Autografting techniques have been modified in recent years to exploit the presence of putative normal residual haemopoietic cells in the marrow of patients with CML. These include in vitro manipulations of marrow cells such as 10 day culture, purging with chemotherapeutic and other agents, and in vivo approaches such as collecting peripheral blood progenitor cells for autografting in the early recovery phase following high dose chemotherapy. Despite these advances, no large clinical trials have been carried out to address the question of whether autografting in CML extends the duration of chronic phase or prolongs survival. Therefore for the present autografting for CML must still be regarded as experimental, but interest in this approach continues since for the majority of patients allogeneic transplantation is not currently possible.

CD34+ cell selection in chronic phase chronic myeloid leukaemia: a comparison of laboratory grade columns.

CD34 positive (CD34+) cell selection is increasingly used for a number of important applications including gene therapy studies, ex vivo expansion and purging. However there are no data regarding the use of different technologies for CD34+ cell selection in chronic myeloid leukaemia (CML). We therefore compared the performance of three laboratory grade CD34+ selection columns (MiniMACS, Cellpro Ceprate LC and Baxter Isolex 50), using CML chronic phase peripheral blood (PB) and bone marrow (BM). With different CML samples the CD34+ purity from the three columns was equivalent, but comparing five paired samples the Ceprate purity was greater than MiniMACS, at 92.5 and 80.9%, respectively, P = 0.04. Combining results from paired and unpaired CML samples, MiniMACS (n = 7) gave a higher CD34+ yield than Ceprate LC (n = 8) or Isolex 50 (n = 4) with a mean of 51.1%, 24.3% and 13.2% respectively, (P = 0.04 and 0.01). Cell losses with all columns were similar. Attempts to improve the yield from the Ceprate LC columns by modifying the method were unsuccessful. Following MiniMACS and Ceprate LC separation the clonogenic potentials of CD34+ cells in the pre- and positive cell fractions were the same. The proportion of CD34+ 38- or CD34+ DR- cells was unchanged following column separation. These data suggest that the MiniMACS column may be the best column for CD34+ cell selection in CML but these results must be confirmed using large scale clinical columns once the MiniMACS column is licensed. It is possible that variations in CD34+ cell yields between the different columns reflect differences in antibody binding affinity to CML cells, or differences in column technologies.

An update on chronic myeloid leukaemia.

The therapy of CML is clearly 'work in progress' Although long-term follow-up data for patients treated with STI571 are not yet available, preliminary results are encouraging. STI571 is likely to be licensed in late 2001/early 2002 and will certainly find a place in the treatment of CML. Current therapeutic considerations are summarised in Fig 2. At all stages of treatment it is vital that the patient is fully aware of treatment options, their risks and benefits. In some cases there is little evidence to suggest that one treatment option is better than another; in these situations, treatment is likely to depend on a patient's individual circumstances and preferences.

Six-year follow-up of patients receiving imatinib for the first-line treatment of chronic myeloid leukemia.

Imatinib mesylate is considered standard of care for first-line treatment of chronic phase chronic myeloid leukemia (CML-CP). In the phase III, randomized, open-label International Randomized Study of Interferon vs STI571 (IRIS) trial, previously untreated CML-CP patients were randomized to imatinib (n=553) or interferon-alpha (IFN) plus cytarabine (n=553). This 6-year update focuses on patients randomized to receive imatinib as first-line therapy for newly diagnosed CML-CP. During the sixth year of study treatment, there were no reports of disease progression to accelerated phase (AP) or blast crisis (BC). The toxicity profile was unchanged. The cumulative best complete cytogenetic response (CCyR) rate was 82%; 63% of all patients randomized to receive imatinib and still on study treatment showed CCyR at last assessment. The estimated event-free survival at 6 years was 83%, and the estimated rate of freedom from progression to AP and BC was 93%. The estimated overall survival was 88% -- or 95% when only CML-related deaths were considered. This 6-year update of IRIS underscores the efficacy and safety of imatinib as first-line therapy for patients with CML.

On the use of lonafarnib in myelodysplastic syndrome and chronic myelomonocytic leukemia.

Lonafarnib is an orally bio-available farnesyltransferase inhibitor that prevents farnesylation of specific target proteins including Ras. In a multicenter study, 67 patients with advanced myelodysplastic syndrome (MDS) and chronic myelomonocytic leukemia (CMML) were treated with a continuous oral dose of 200-300 mg of lonafarnib and were evaluated for hematologic, pathologic and pharmacodynamic response. The median age of patients was 70 years (range 44-86). There were 32 patients with MDS (RAEB-20 and RAEB-t-12) and 35 with CMML. Overall 16 (24%) of the patients responded with two patients achieving a complete remission and one a partial response. Responses were seen in 6/32 and 10/35 patients with MDS and CMML, respectively. Of the 19 patients who were platelet transfusion-dependent prior to treatment, 5 (26%) became transfusion-free for a median duration of 185 days. A decrease in the farnesylation of the HDJ-2 protein measured in patient-derived cells was observed in the majority of patients during treatment with lonafarnib, but no clear correlation between changes in farnesylation and clinical effect could be made. Gastrointestinal toxicity was significant with 19% of patients discontinuing therapy due to diarrhea, nausea and/or anorexia. Lonafarnib has demonstrable activity in patients with advanced MDS and CMML.

Autografting for chronic myeloid leukaemia.

For most chronic myeloid leukaemia (CML) patients the option of a potentially 'curative' allogeneic stem cell transplant is not available because of age or lack of donor. Interferon alpha appears to extend survival when used in the chronic phase of the disease but probably does not produce long-term disease-free survivors. Autografting is being actively explored as a therapeutic option which may improve on the survival data seen with interferon and numerous different autografting methodologies are being investigated. While it seems reasonable to hope that a suitably robust and safe approach to autografting may improve survival it is unlikely with current technology that long-term disease-free survival will be achieved. To date no compelling trial data are available to confirm the efficacy of autografting but large prospective randomized studies are underway to investigate whether autografting can indeed extend survival for CML patients who do not have the option of an allograft.

Scattering by irregular inhomogeneous particles via the digitized Green's function algorithm.

The digitized Green's function (DGF) algorithm and the underlying theory are described. This finite element algorithm models dielectric particles of arbitrary shape and arbitrary optical structure. DGF predictions of differential and total cross sections are compared with predictions of Mie and EBCM algorithms for several homogeneous spheres and spheroids. Results of tests of convergence of the DGF calculation as the number of elements are increased are presented. Computer time and storage requirements as functions of wavelength and particle size, shape, and optical structure are discussed.

Scattering of millimeter waves by snow crystals and equivalent homogeneous symmetric particles.

The digitized Green's function code was used to compute differential and total cross sections of several model snow crystals and of several homogeneous highly symmetric equivalent particles of the same general shape and size as the snow crystals at a wavelength of 10 mm. Optical constants of equivalent particles were derived using the Biot-Arago, Lorentz-Lorenz, and Bruggemann mixing rules. Reasonable agreement was found for equivalent particles whose mass distributions were most similar to those of the snow crystals. Overall, the Bruggemann mixing rule produced the best match.

Residual Ph-negative stem cells in chronic myeloid leukemia--sometimes or always?

Some patients with Philadelphia (Ph) chromosome-positive chronic myeloid leukemia (CML) may achieve partially or completely Ph-negative hemopoiesis after treatment with high doses of cytotoxic drugs or interferon-alpha at standard dosage. This observation leads to three important questions: 1) are Ph-negative myeloid cells in such patients strictly normal? 2) can such Ph-negative cells be identified in all newly diagnosed patients or only in a minority? and 3) what is the basis for the proliferative advantage manifested by CML cells and why might it be temporarily lost as a result of treatment? The mechanisms that might prevent the proliferation of normal cells in an environment of Ph-positive cells and the relevance of these questions to the design of a strategy aimed at obtaining complete remission in the majority of patients are considered. Such a strategy might incorporate autografting with Ph-negative stem cells harvested during the recovery phase of high dose chemotherapy.

Physics-Based Visualization of Dense Natural Clouds. II. Cloud-Rendering Algorithm.

We discuss the representation of aerosol-scattering properties, boundary information, and the use of these results in line-of-sight rendering applications for visualization of a modeled atmosphere based on a discrete ordinates three-dimensional radiative-transport method. The outputs of the radiative-transfer model provide spatial and angular distributions of limiting path radiance, given an input density distribution and external illumination conditions. We discuss the determination of the direct attenuated radiance, integrated path radiance, and background radiance for each pixel in the rendered scene. Orthographic and perspective projection approaches for displaying these results are described, and sample images are shown.

Physics-based visualization of dense natural clouds. I. Three-dimensional discrete ordinates radiative transfer.

A technique is developed to model radiative transfer in three-dimensional natural clouds with a standard discrete ordinates finite-element method modified to evaluate cell-surface-averaged radiances. A log-least-squares-based scale transformation is used to improve the discrete phase-function model. We handle dense media by assuming constant diffuse radiances over input faces to cubic cells, allowing analytical forms for transmittance factors. Transmission equations are combined with diffuse volumetric single-scattering calculations to support evaluations of cell energy balance. Energy not accounted for volumetrically is treated with surface-based effects. Results produced show accurate flux computations at over 30 optical depths per modeled cell. Comparisons with nonuniform cloud Monte Carlo calculations show less than 1% rms error and correlations greater than 0.999 for cases in which cloud-density fluctuations are resolved.

Orthotopic cardiac transplantation for dilated cardiomyopathy after allogeneic bone marrow transplantation.

We report the case of a 26-year-old man with acute myeloid leukemia (AML) who underwent bone marrow transplantation (BMT) from an HLA-identical sibling-donor when in first complete remission. He developed a dilated cardiomyopathy 9 months post-BMT and subsequently underwent orthotopic heart transplantation. He remains in complete hematological remission more than 12 months after surgery with normal cardiac function and an excellent performance status. Although cardiac transplantation has been carried out for chemotherapy-induced cardiomyopathy in cancer patients, to our knowledge cardiac transplantation following BMT has not been reported previously.

Chronic intrahepatic cholestasis in sickle cell disease requiring exchange transfusion.

A 15 year old boy with sickle cell disease developed intrahepatic cholestasis. A course of exchange transfusion successfully corrected the extreme hyperbilirubinaemia over one year. Upon stopping the exchange transfusion programme the hyperbilirubinaemia relapsed but transfusion was effective when reinstituted.

Effects of imatinib mesylate (STI571, Glivec) on the pharmacokinetics of simvastatin, a cytochrome p450 3A4 substrate, in patients with chronic myeloid leukaemia.

The inhibition by imatinib of the cytochrome p450 3A4 isoenzyme may reduce the CYP3A4-mediated metabolic clearance of clinically important coadministered drugs. The main purpose of this study was to evaluate the effect of the coadministration of imatinib on the pharmacokinetics of simvastatin, a probe CYP3A4 substrate. In total, 20 patients with chronic myeloid leukaemia received an oral dose of 40 mg of simvastatin on study day 1. On study days 2-7, each patient received 400 mg of imatinib once daily orally and on study day 8, 400 mg imatinib together with 40 mg of simvastatin was given. Blood levels of simvastatin were measured predose and for 24 h postdose on study days 1 and 8. Two additional blood samples were taken for imatinib pharmacokinetic (PK) assessment on day 8 before, and 24 h after, imatinib administration. Imatinib increased the mean maximum concentration (C(max)) value of simvastatin two-fold and the area under concentration-time curve (AUC ((0-inf))) value 3.5-fold (P<0.001) compared with simvastatin alone. There was a statistically significant decrease in total-body clearance of drug from the plasma (CL/F) with a mean reduction of 70% for simvastatin (P<0.001): the mean half-life of simvastatin was prolonged from 1.4-2.7 h when given together with imatinib. No changes in imatinib PK parameters were found when given concomitantly with simvastatin. In conclusion, the coadministration of imatinib at steady state with 40 mg simvastatin increases the exposure (C(max) and AUCs) of simvastatin significantly (P<0.001) by two-three-fold. Caution is therefore required when administering imatinib with CYP3A4 substrates with a narrow therapeutic window. The coadministration of simvastatin with imatinib (400 mg) was well tolerated and no major safety findings were reported in this study.