Relapsed multiple myeloma: who benefits from salvage autografts?

BACKGROUND: Multiple myeloma is incurable despite the advance of autologous stem cell transplant (ASCT) and novel agents (thalidomide, bortezomib, lenalidomide). The role of ASCT as salvage therapy in relapsed myeloma remains unclear. AIM: To identify and refine the predictors of survival following salvage ASCT for relapsed multiple myeloma, so that they can be applied clinically for patient selection. METHODS: Retrospective review of patients treated salvage ASCT for relapsed myeloma at our centre from 1992 to 2011. RESULTS: Following an initial ASCT at diagnosis, 30 patients underwent salvage ASCT for subsequent relapse, with the median time to first relapse/progression being 30.2 months. All patients received reinduction, then melphalan-based conditioning with salvage ASCT. Non-relapse mortality at 100 days following salvage ASCT was 3%. The median overall survival and progression-free survival following salvage ASCT were 45 and 22 months respectively. The progression-free interval (PFI) after initial ASCT predicted survival outcomes in a time-dependent manner. With PFI following initial ASCT of <18, 18-36 and ≥36 months, the median progression-free survival following salvage ASCT was 4.2, 13.8 and 49.1 months respectively (P < 0.0001). The median overall survival was 10.7, 30.9 and 86.1 months respectively (P < 0.0001). CONCLUSIONS: Salvage ASCT is an effective and safe treatment option in selected patients and should be considered in patients relapsing ≥36 months after their initial ASCT. The time-dependent relationship between PFI and salvage ASCT outcome is important when stratifying patient groups who may benefit from this procedure.

When should iron chelation therapy be considered in patients with myelodysplasia and other bone marrow failure syndromes with iron overload?

Despite the absence of a robust evidence base, there is growing consensus that effective treatment of iron overload leads to decreased morbidity and premature mortality in patients with good prognosis myelodysplastic syndromes (MDSs). Furthermore, new treatment modalities, including disease-modifying therapies (lenalidamide and azacytidine) and reduced intensity conditioning therapies for allogeneic blood stem cell transplants, are offering the prospect of longer survival for patients with traditionally less favourable prognosis MDS, who might also benefit from iron chelation. This article proposes assessment of patients with MDS and related bone marrow failure syndromes to determine suitability for iron chelation. Iron chelation therapy options and monitoring are discussed.

Increased T regulatory cells and decreased Th1 pro-inflammatory cytokines correlate with culture-positive infection in febrile neutropenia childhood oncology patients.

BACKGROUND: Paediatric oncology patients with febrile neutropenia are usually hospitalised and treated with empirical broad-spectrum antibiotic therapy to counter the risk of infection. However, there is currently no method available to rapidly identify bacteremia in these patients. T-helper-type-1 (Th1) cytokines are required for effective immune response to many pathogenic organisms and T regulatory cells are known suppressors of Th1 cells. We hypothesized that characterization of reduced intracellular Th1 cytokines and increased T regulatory cells (Tregs) may prove useful in identifying infection in childhood oncology patients with febrile neutropenia. METHODS: Intracellular Th 1 cytokines and Tregs were enumerated in peripheral blood from a group of childhood oncology patients with febrile neutropenia using multiparameter flow cytometry. RESULTS: There was a significant increase in the percentage of CD25(+) CD127(-) CD8(-) CD3(+) Tregs and a significant decrease in Th1 intracellular cytokines IFNγ, IL-2 and TNFα in the blood of culture positive patients compared with culture negative patients. CONCLUSIONS: Enumeration of Tregs and intracellular Th1 cytokines may provide a sensitive, specific test for determining infection in childhood oncology patients before blood culture results become available.

How we mobilize haemopoietic stem cells.

Mobilization and collection of haemopoietic stem and progenitor cells (HSPC) is the cornerstone of autologous and allogeneic stem cell transplantation for a wide variety of haematological and some non-haematological malignancies. Centres providing this service face the challenge of optimizing the likelihood of successful collection of transplantable doses of cells, while maximizing the efficiency of the apheresis unit and minimizing the risk of toxicity as well as mobilization failure. Recent developments in the understanding of the molecular mechanisms of mobilization have led to the emergence of novel strategies for HSPC mobilization, which may assist in meeting these imperatives. The task for clinicians is how to incorporate the use of these strategies into practice, in the light of emerging evidence for efficacy and safety of these agents. Herein, the literature is reviewed, and a proposed algorithm for HSPC mobilization is presented.

Outcome of patients with myelodysplastic syndromes: experience from a single institution in South Australia.

AIMS: To study disease characteristics of adult patients with myelodysplastic syndromes (MDS) in South Australia and to analyse their outcome and survival. METHODS: One hundred and eight adult patients with confirmed MDS from marrow biopsies in the 76-month period before April 2006 were retrospectively included in an MDS database. RESULTS: The median age at diagnosis of this cohort was 70 years, with skewing of refractory anaemia with excess blasts and refractory cytopenia with multilineage dysplasia in the younger patients. Clonal cytogenetic abnormalities were present in 42% of patients. Median survival was 48 months, and secondary transformation to acute myeloid leukaemia was seen in 27%. Survival, according to the World Health Organization subtypes in ascending order, was refractory anaemia with excess blasts, refractory anaemia, refractory anaemia with ringed sideroblast, refractory cytopenia with multilineage dysplasia and del(5q). The International Prognostic Scoring System score stratified MDS patients into different risk groups and effectively discriminated significantly different survivals, ranging from a median 4 months for high-risk patients to 72 months for low-risk patients. CONCLUSION: An MDS database provides useful information regarding the disease characteristics and survival of MDS patients in South Australia and confirms the prognostic usefulness of the International Prognostic Scoring System. The future prospective collection of results will be invaluable in evaluating the effect of novel therapies on patient prognosis.

A four-gene LincRNA expression signature predicts risk in multiple cohorts of acute myeloid leukemia patients.

Prognostic gene expression signatures have been proposed as clinical tools to clarify therapeutic options in acute myeloid leukemia (AML). However, these signatures rely on measuring large numbers of genes and often perform poorly when applied to independent cohorts or those with older patients. Long intergenic non-coding RNAs (lincRNAs) are emerging as important regulators of cell identity and oncogenesis, but knowledge of their utility as prognostic markers in AML is limited. Here we analyze transcriptomic data from multiple cohorts of clinically annotated AML patients and report that (i) microarrays designed for coding gene expression can be repurposed to yield robust lincRNA expression data, (ii) some lincRNA genes are located in close proximity to hematopoietic coding genes and show strong expression correlations in AML, (iii) lincRNA gene expression patterns distinguish cytogenetic and molecular subtypes of AML, (iv) lincRNA signatures composed of three or four genes are independent predictors of clinical outcome and further dichotomize survival in European Leukemia Net (ELN) risk groups and (v) an analytical tool based on logistic regression analysis of quantitative PCR measurement of four lincRNA genes (LINC4) can be used to determine risk in AML.

Multicycle high-dose chemotherapy and filgrastim-mobilized peripheral-blood progenitor cells in women with high-risk stage II or III breast cancer: five-year follow-up.

PURPOSE: To determine the safety and efficacy of multiple cycles of dose-intensive, nonablative chemotherapy in women with poor-prognosis breast cancer. PATIENTS AND METHODS: Women with stage II breast cancer and 10 or more involved nodes or four or more involved nodes and estrogen receptor-negative tumors and women with stage III disease received three cycles of epirubicin 200 mg/m2 and cyclophosphamide 4 g/m2, with progenitor cell and filgrastim support every 28 days (n = 79) or 21 days (n = 20). Patients were reviewed at least twice yearly thereafter. Twenty-six patients had bone marrow and apheresis collections assessed for the presence of micrometastatic tumor cells. RESULTS: Ninety-nine women (median age, 43 years; range, 24 to 60 years) were treated. Ninety-two completed all three cycles of chemotherapy. The major toxicity was severe, reversible myelosuppression that was more prolonged with successive cycles, and this did not differ between patients given treatment every 28 days and those treated every 21 days. Febrile neutropenia occurred in 176 (61%) of 287 cycles. Severe mucositis (grade 3 or 4) occurred in 23% of cycles but tended to be short-lived and was reversible. The cardiac ejection fraction fell by a median of 4% during treatment, and three patients developed evidence of cardiac failure after chemotherapy. Two patients (2%) died of acute toxicity. Three of 26 patients had evidence of circulating micrometastatic tumor cells. The actuarial distant disease-free and overall survival rates at 60-month follow-up were 64% (95% confidence interval [CI], 53% to 75%) and 67% (95% CI, 56% to 78%), respectively. CONCLUSION: Multiple cycles of dose-intensive, nonablative chemotherapy is a feasible and safe approach. Disease control and survival are similar to those in other studies of myeloablative chemotherapy in poor-prognosis breast cancer. The regimen is being evaluated in a randomized trial of the International Breast Cancer Study Group.

Rapid hematopoietic recovery after multicycle high-dose chemotherapy: enhancement of filgrastim-induced progenitor-cell mobilization by recombinant human stem-cell factor.

PURPOSE: To assess the mobilization potential and safety of recombinant human stem-cell factor (SCF) when coadministered with filgrastim to untreated women with poor-prognosis breast cancer. PATIENTS AND METHODS: Eligible women had breast cancer with 10 or more positive axillary nodes, or estrogen receptor-negative tumor with 4 positive nodes, or stage III disease. Patients were randomized to receive SCF plus filgrastim or filgrastim alone. Filgrastim 12 microg/kg daily was administered for 6 days by continuous subcutaneous infusion. SCF was administered by daily subcutaneous injection at 5, 10, or 15 microg/kg concurrent with filgrastim for 7 days, or 10 microg/kg daily starting 3 days before filgrastim for a total of 10 days (SCF pretreatment). Apheresis was performed on days 5, 6, and 7 of filgrastim administration. Patients then had three cycles of epirubicin 200 mg/m2 and cyclophosphamide 4 g/m2 every 28 days, each supported by one third of the apheresis product. RESULTS: Sixty-two women were treated. Greater yields occurred in patients who received SCF 10 microg/kg daily plus filgastim than those who received filgrastim alone (P=.013 for CD34+ cells; P=.07 for granulocyte-macrophage colony-forming cells [GM-CFCs]). The difference was more marked with SCF-pretreatment than concurrent SCF. Fewer aphereses were required to reach the predetermined target of peripheral-blood progenitor/stem cells (PBPCs) in women who received SCF. SCF was generally well tolerated. Hematologic recovery was rapid after each of the three cycles of chemotherapy. There was no difference in recovery between the different treatment groups. CONCLUSION: Mobilization of PBPCs by filgrastim is significantly enhanced by coadministration of SCF, and commencing SCF before filgrastim can optimize this effect. SCF has the potential to reduce the number of aphereses required to collect a target number of PBPCs.

The detection of Philadelphia chromosome negative metaphases in long-term bone marrow cultures of the peripheral blood from patients with chronic myeloid leukemia predicts response to interferon-alpha 2a.

The cytogenetic response of 10 patients with chronic myeloid leukaemia (CML) to human recombinant interferon-alpha 2a (rhIFN alpha 2a) was compared to the Philadelphia chromosome (Ph) status of the pre-treatment peripheral blood cells after in vitro culture under long-term bone marrow culture (LTBMC) conditions. Pre-treatment light density peripheral blood cells were cultured in LTBMC on sex-mismatched irradiated allogeneic stromal layers with weekly cytogenic examination of metaphases in the non-adherent cell fraction. This was correlated with the patients' response to rhIFN alpha. Two groups of patients, five showing a cytogenetic response (responsive) and five who failed to achieve a cytogenetic response (nonresponsive) were studied. At the initiation of the LTBMCs the Ph' was found to be present in 100% of the cells analysed for nine patients and 97% for one patient. Pretreatment peripheral blood from four responsive patients demonstrated a decline in the proportion of Ph'-positive cells (Ph+) after 1 to 2 weeks in LTBMC. In contrast, peripheral blood from all the non-responsive subjects showed persistence of the Ph+ clone in 100% of the cells analysed out to a maximum of 3 to 5 weeks in LTBMC. A significant difference was observed (Fisher exact test, p = 0.023) between the two patient groups in respect to the appearance of normal clones in the nonadherent population. The presence of Ph- metaphases in LTBMC of peripheral blood cells of CML patients may predict their cytogenetic response to rhIFN alpha 2a.

Peripheral blood is a source of BCR-ABL-negative pre-progenitors in early chronic phase chronic myeloid leukemia.

Manipulation of autologous bone marrow cells (BM) for transplantation in chronic myeloid leukemia (CML) to enrich for normal cells is a novel approach that may improve survival for patients not suitable for allogeneic transplantation. Limitations of this technique include the reported low frequency of normal stem cells in CML and the difficulties in obtaining sufficient BM for manipulation. To address these problems we compared the apheresis product with the diagnostic bone marrow at diagnosis as a source of primitive BCR/ABL-negative progenitors. We analyzed the CD34+ HLA-DR- and CD34+CD38(-) populations in five CML patients to evaluate the frequency of BCR-ABL-negative progenitors and pre-progenitors in these populations. Progenitor analysis was performed by RT-PCR of individual hemopoietic colonies from a standard CFU-GM assay. Analysis of pre-progenitors involved RT-PCR of secondary colonies derived from a stroma-free pre-CFU assay. Our results show variable levels of BCR-ABL-negative progenitors in the 34+DR- population but very low levels of BCR-ABL-negative progenitors in the 34+38- population in blood. Analysis of pre-progenitors from the 34+DR- fraction of peripheral blood (PB) and BM showed 80-100% and 85-100% of colonies were BCR-ABL negative at days 14 and 28, respectively. Analysis of pre-progenitors from the 34+38- fraction of PB and BM showed 23-100% and 42-100% of colonies were BCR-ABL negative at days 14 and 28, respectively. In summary, pre-progenitors from the 34+DR- and 34+38- populations are predominantly BCR-ABL negative in both marrow and blood at diagnosis. Apheresis product collected at diagnosis is a more abundant sources of BCR-ABL-negative pre-progenitors than BM. Thus, apheresis product could potentially be utilized as a source of BCR-ABL-negative stem cells in CML.

Successful peripheral blood stem cell mobilisation with filgrastim in patients with chronic myeloid leukaemia achieving complete cytogenetic response with imatinib, without increasing disease burden as measured by quantitative real-time PCR.

Imatinib mesylate (Glivec) is a selective inhibitor of bcr-abl tyrosine kinase, the product of the Philadelphia chromosome, which is the hallmark of chronic myeloid leukaemia (CML). With imatinib, complete cytogenetic response (CCR) can be achieved in over 70% of newly diagnosed patients with CML. However, the optimal long-term management of patients who achieve CCR after imatinib is unknown. With longer follow-up, it is anticipated that some patients are likely to progress and become candidates for autologous transplantation. We studied filgrastim (r-metHuG-CSF) mobilisation of peripheral blood stem cells (PBSC) in 32 patients who have achieved CCR with imatinib. Our data demonstrate that (1) the target CD34(+) cell yields of >/=2.0 x 10(6)/kg were attained with filgrastim 10 microg/kg/day, in 9/18 (50%) of patients during uninterrupted imatinib therapy, and in 10/14 (70%) when imatinib was temporarily withheld. The median CD34(+) cell yield per aphaeresis was 0.70 x 10(6)/kg (range 0.14-2.18) and 2.90 x 10(6)/kg (range 0.15-8.71) in the two groups, respectively (P&<0.005). (2) The cell yields did not correlate with the duration of imatinib administration. (3) There was no impact of the mobilisation procedure on the level of leukaemia as measured by serial blood bcr-abl levels using real-time quantitative PCR with either protocol. (4) bcr-abl remained detectable at low levels in the harvests in most but not all patients. In conclusion, filgrastim can safely be used to mobilise PBSC in patients who have achieved CCR with imatinib, but CD34(+) cell yields are significantly improved when imatinib is temporarily withheld.

Adjuvant treatment of high-risk breast cancer using multicycle high-dose chemotherapy and filgrastim-mobilized peripheral blood progenitor cells.

Women with primary breast cancer associated with extensive axillary node involvement or large primary tumors have a very poor prognosis despite treatment with standard-dose adjuvant chemotherapy. In an attempt to improve the outlook of these patients, we investigated the safety and feasibility of delivering three cycles of high-dose epirubicin and cyclophosphamide supported with filgrastim-mobilized peripheral blood progenitor cells (PBPC). Fifteen previously untreated women, median age 50 (range, 30-58) years, with poor prognosis early stage breast cancer received filgrastim (12 microgram/kg daily for 6 days) prior to chemotherapy to mobilize progenitor cells. Patients were then given three cycles of epirubicin (200 mg/m2) and cyclophosphamide (4 g/m2) at planned 28-day intervals, each followed by infusion of one third of the PBPC collected and daily administration of filgrastim (5 microgram/kg s.c.). Three leukaphereses collected a median of 114.9 (range, 22.7-273.5) x 10(4) granulocyte-macrophage-colony-forming cells/kg body weight. Hemopoietic recovery was rapid after each cycle, and there was no correlation between the rate of recovery and the number of granulocyte-macrophage-colony-forming cells infused. There was a small but significant progressive delay in recovery from hematological and nonhematological toxicities across the three cycles. Left ventricular ejection fraction fell to below 50% in eight (53%) patients, but none developed congestive cardiac failure. Two patients did not complete three cycles because of insufficient PBPC for a third cycle (n = 1) and 2-mercaptoethane sodium sulfonate- related drug reaction during the second cycle (n = 1). There were no deaths during the study or during the follow-up period (median, 70 weeks; range, 50-85 weeks), and no late toxicities occurred. Therefore, we concluded that the delivery of multiple cycles of nonmyeloablative, dose-intensive chemotherapy supported by PBPC and filgrastim is safe, and may be widely applicable to a variety of common chemosensitive cancers with a poor prognosis. The efficacy of three cycles of high-dose epirubicin and cyclophosphamide is to be compared with standard-dose chemotherapy in a randomized trial in patients with high-risk, operable stage II and III breast cancer.

CFU-mix are no better than CFU-GM in predicting hemopoietic reconstitutive capacity of peripheral blood stem cells collected in the very early remission phase of acute nonlymphoblastic leukemia.

High levels of circulating myeloid progenitor cells (CFU-GM) occur during the very early remission phase of acute nonlymphoblastic leukemia (ANLL). Autologous stem cell rescue using blood cells collected during this phase has shown that successful hemopoietic reconstitution can be achieved, but a higher CFU-GM dose appears to be required than when bone marrow cells are used. This suggests that during very early remission, the level of marrow repopulating pluripotent stem cells (PSC) in blood does not undergo the same amount of increase as does the CFU-GM. This study set out to determine whether the levels of the multilineage progenitor cell (CFU-Mix) would be better indicators of the PSC in these cells than the CFU-GM. Serial peripheral blood CFU-Mix and CFU-GM measurements were carried out in six ANLL patients during very early remission. The levels of peripheral blood CFU-Mix showed a mean 12-fold increase, as compared to a mean 20-fold increase in the CFU-GM. The timing of the increase in the CFU-Mix paralleled that of the CFU-GM. These findings suggest that the CFU-Mix is no better than the CFU-GM in predicting PSC levels during very early remission of ANLL, and is closer to the CFU-GM than to the PSC in ontogeny.

c-kit is expressed by primitive human hematopoietic cells that give rise to colony-forming cells in stroma-dependent or cytokine-supplemented culture.

Using monoclonal antibody (MAB) YB5.B8, we have examined the expression of the c-kit protein, the receptor for the hematopoietic cytokine stem cell factor (SCF), on primitive hematopoietic cells. Bone marrow mononuclear cells (BMMNC) enriched for immature cells by differential agglutination using the lectin soybean agglutinin (SBA) were subjected to multiparameter fluorescence activated cell sorting (FACS) based on light-scattering properties, the expression of the c-kit protein and the CD34 antigen, and the retention of the vital fluorescent dye, Rhodamine 123 (Rh123). Sorted populations were assayed for their content of directly clonogenic progenitor cells (colony-forming units-granulocyte/macrophage [CFU-GM], burst-forming units-erythroid [BFU-E], and multipotential colony-forming units [CFU-Mix]) and for the presence of more primitive progenitor cells ("pre-CFU"). The latter were assayed by (1) their ability to initiate and sustain hematopoiesis in a standard stromal cell-dependent culture system and (2) their capacity for de novo generation of clonogenic progenitors in response to a combination of six recombinant hematopoietic cytokines in a stroma-independent suspension culture assay. A mean of 76% of CD34+ cells were found to coexpress c-kit. The majority of directly clonogenic cells (98% of CFU-GM, 98% of CFU-Mix, and 85% of BFU-E) were found in the CD34+c-kit+ fraction. Similarly, all pre-CFU were recovered in the CD34+c-kit+Rh123dull fraction, irrespective of whether the cells were maintained on marrow stromal cells or in cytokine-supplemented liquid culture. A mean of 87% (range 70-100%) of the CD34+Rh123dull cells also expressed c-kit. Since SCF has been reported to act as a growth factor for early lymphoid cells as well as myeloid cells, we looked for coexpression of c-kit and early lymphoid markers in the CD34+ population by multiparameter flow cytometry. Coexpression of c-kit on a minority of cells with markers of B or T lineages was observed. The majority of early lymphoid cells, however, appeared to lack c-kit expression. This was confirmed by the finding that only 4% of c-kit+CD34+ cells showed terminal deoxynucleotidyl transferase (TdT) activity, compared with 25% of the c-kit-CD34+ cells.

Single high doses of cyclophosphamide enable the collection of high numbers of hemopoietic stem cells from the peripheral blood.

We used single high doses of cyclophosphamide (4 g/m2) to produce rebound increases in peripheral blood (PB) stem cells (PBSC) during recovery from myelosuppression, enabling their collection by apheresis for later autotransplantation. Thirty-three courses of cyclophosphamide were given to 30 patients with malignant lymphoma, multiple myeloma, or solid tumors. The neutrophil count was less than 0.5 x 10(9)/liter for a mean of 6.9 days (median 7 days), and fever occurred in 17 of 33 courses. Positive blood cultures occurred in two patients, one of whom died. The mean peak level of PB granulocyte-macrophage colony-forming units (CFU-GM) was 1517 x 10(3)/liter (median 2447 x 10(3)/liter), a 14-fold increase above the mean in normal subjects. The peak occurred at a mean of 16.6 days (median 16 days) after cyclophosphamide, generally coinciding with the time to reach 1.0 x 10(9) neutrophils per liter. Normal or minimally involved bone marrow and a rapid rise in leukocyte count during recovery were independent variables correlated to the peak of the rebound increase in PB CFU-GM levels. Previous chemotherapy and the duration of neutropenia were additional independent variables in the group with peak PB CFU-GM levels of greater than 1000 x 10(3)/liter. The mean total CFU-GM collected after a mean of five aphereses was 43.8 x 10(4)/kg body weight (BW) (median 35.5 x 10(4)/kg BW), significantly correlated with the mononuclear cell yield. We conclude that single 4 g/m2 doses of cyclophosphamide effectively produce high levels of PBSC, particularly but not exclusively in patients with normal or minimally involved bone marrow and who have not had intensive recent chemotherapy.

Contribution of chemotherapy mobilization to disease control in multiple myeloma treated with autologous hematopoietic cell transplantation.

In patients with multiple myeloma (MM) undergoing autologous hematopoietic cell transplantation (auto-HCT), peripheral blood progenitor cells may be collected following mobilization with growth factor alone (GF) or cytotoxic chemotherapy plus GF (CC+GF). It is uncertain whether the method of mobilization affects post-transplant outcomes. We compared these mobilization strategies in a retrospective analysis of 968 patients with MM from the Center for International Blood and Marrow Transplant Research database who received an auto-HCT in the US and Canada between 2007 and 2012. The kinetics of neutrophil engraftment (⩾0.5 × 10(9)/L) was similar between groups (13 vs 13 days, P=0.69) while platelet engraftment (⩾20 × 10(9)/L) was slightly faster with CC+GF (19 vs 18 days, P=0.006). Adjusted 3-year PFS was 43% (95% confidence interval (CI) 38-48) in GF and 40% (95% CI 35-45) in CC+GF, P=0.33. Adjusted 3-year OS was 82% (95% CI 78-86) vs 80% (95% CI 75-84), P=0.43 and adjusted 5-year OS was 62% (95% CI 54-68) vs 60% (95% CI 52-67), P=0.76, for GF and CC+GF, respectively. We conclude that MM patients undergoing auto-HCT have similar outcomes irrespective of the method of mobilization and found no evidence that the addition of chemotherapy to mobilization contributes to disease control.

Residual leukemia cannot be detected in very early remission peripheral blood stem cell collections in acute non-lymphoblastic leukemia.

We have used a combined cell culture and cytogenetic approach to study the level of residual leukemia during the very early remission (VER) phase of acute non-lymphoblastic leukemia. Clonogenic leukemic cells were induced to proliferate by phytohemagglutinin-stimulated leucocyte conditioned medium and identified by a leukemia-associated karyotype t(8;21) and a morphological marker (Auer rod). When leukemic blasts were cultured, the leukemic karyotype and Auer rods were most readily detected after 3-9 days. When VER blood cells were cultured, no leukemia-associated karyotype or Auer rods could be detected. Based on the number of VER blood cell derived metaphases analysed, the incidence of leukemic blasts among dividing cells is less than 2%.

A differential sensitivity to recombinant human interferon-alpha 2a between normal and chronic myeloid leukaemic peripheral blood granulocyte-macrophage colony-forming units.

The sensitivity to recombinant human interferon-alpha 2a (IFN) of peripheral blood granulocyte-macrophage colony-forming units (PB CFU-GM) from patients with chronic myeloid leukaemia (CML) was studied in a semi-solid clonogenic assay, and compared with normal PB CFU-GM. Like normal PB CFU-GM, the growth of CML PB CFU-GM in vitro was found to be dependent on the plating concentration used. The optimal CFU-GM growth occurred when CML PB mononuclear cells (MNC) were plated at low concentrations in the range of 0.01-0.1 x 10(5)/ml, compared to the range of 0.3-3.0 x 10(5)/ml optimal for CFU-GM growth in normal subjects. The optimal plating concentration for CML PB CFU-GM was similar to that observed in PB collected from patients with ovarian carcinoma during haematological recovery following chemotherapy-induced myelosuppression (recovery phase). The recovery phase PB was used as a source of non-leukaemic cells with a higher incidence of CFU-GM similar to that of CML. IFN produced a dose-related inhibition of CFU-GM growth in normal, recovery phase ovarian carcinoma and CML, PB MNC. The IFN concentration required to inhibit 50% of the CFU-GM in culture (LD50) was found to be significantly influenced by the plating concentration. When cells were cultured at 1.0 x 10(5) MNC/ml the mean LD50 for 7 CML patients was similar to that in normal (n = 5) or recovery phase (n = 5) peripheral blood, 273 i.u./ml, 1047 i.u./ml and 795 i.u./ml, respectively. In contrast when CML cells were cultured at 0.03 x 10(5) MNC/ml the concentration for optimal CML CFU-GM growth, the mean LD50 was significantly lower than that in normal PB and recovery phase PB, 4 i.u./ml, 251 i.u./ml and 78 i.u./ml, respectively (p less than 0.05). This is the first report of a differential sensitivity to IFN between CML and non-CML progenitors using an optimized PB CFU-GM assay system and proposes that further study of the in vitro culture of CML progenitors may increase our understanding of the clinical effects of IFN.

Bone marrow biopsy during induction chemotherapy for acute myeloid leukaemia identifies only 50% of patients with resistant disease.

A study was carried out to determine whether bone marrow biopsy performed on day 6 of induction therapy for acute myeloid leukaemia (AML) can identify those patients with resistant disease who would need an intensification of the first course of induction. Bone marrow biopsies were performed on day 6 of induction chemotherapy in 44 patients with AML treated with daunorubicin, cytosine arabinoside and thioguanine. Biopsies were assessed for blast count, trephine cellularity and leukaemic index. Discrimination between patients who went on to achieve remission and those with resistant disease was best achieved using the reduction in bone marrow cellularity from pretreatment marrow to day-6 marrow. However, this discriminator identified only 50% of the patients with resistant disease and included 13% of patients who achieved remission with the first course of chemotherapy. The other parameters of response were even less effective at discriminating between chemotherapy-resistant and chemotherapy-responsive disease.

Spontaneous regression of relapsed non-Hodgkin's lymphoma in a patient who received an autologous transplant for primary resistant disease.

Poor prognosis malignant lymphoma is often treated with autologous bone marrow transplantation. Relapse after transplantation is usually associated with disease progression and resistance to further therapy. We present a case in which a 50-year-old patient relapsed with multiple pulmonary metastases shortly after peripheral blood stem cell transplantation. Remarkably, this relapse remitted spontaneously. This may provide valuable insights into tumour cell biology and mechanisms by which stem cell mobilisation may alter this.