Thiotepa-based high-dose therapy for autologous stem cell transplantation in lymphoma: a retrospective study from the EBMT.

Clinical information about thiotepa-based autologous stem cell transplantation (auto-SCT) outside the primary central nervous system lymphoma (PCNSL) field is sparse. In this registry-based retrospective study, we evaluated potential risks and benefits of thiotepa-based preparative regimens compared with BEAM (carmustine, etoposide, cytarabine, melphalan) in auto-SCT for diffuse large B-cell lymphoma (DLBCL, excluding PCNSL), follicular lymphoma (FL) or Hodgkin lymphoma (HL). A total of 14 544 patients (589 thiotepa and 13 955 BEAM) met the eligibility criteria, and 535 thiotepa- and 1031 BEAM-treated patients were matched in a 1:2 ratio for final comparison. No significant differences between thiotepa and BEAM groups for any survival end point were identified in the whole sample or disease entity subsets. For a more detailed analysis, 47 TEAM (thiotepa, etoposide, cytarabine, melphalan)-treated patients were compared with 75 matched BEAM patients with additional collection of toxicity data. Again, there were no significant differences between the two groups for any survival end point. In addition, the frequency of common infectious and non-infectious complications including secondary malignancies was comparable between TEAM and BEAM. These results indicate that thiotepa-based high-dose therapy might be a valuable alternative to BEAM in DLBCL, HL and FL. Further evaluation by prospective clinical trials is warranted.

BU/melphalan and auto-SCT in AML patients in first CR: a 'Gruppo Italiano Trapianto di Midollo Osseo (GITMO)' retrospective study.

AML patients (total 129; median age =50 years; range 16-72) in first CR received BU and melphalan (BU/Mel) as conditioning regimen before auto-SCT. In all, 82 patients (63.6%) received PBSCs and 47 patients (36.4%) received BM cells. The distribution of cytogenetic categories was conventionally defined as favorable (15.5%), intermediate (60.1%) and unfavorable (24.3%). With a median follow-up of 31 months, the 8-year projected OS and disease-free survival (DFS) was 62 and 56% for the whole population, respectively. The relapse rate was 46% and the non-relapse mortality was 4.65%. Although PBSC transplantation led to a faster hematological recovery than BM transplantation, in univariate analysis the stem cell source, cytogenetics and different BU formulations did not significantly affect OS and DFS, whereas age and the number of post-remission chemotherapy cycles did have a significant effect on the clinical outcome. Multivariate analysis identified age <55 years as the only important independent predictor for OS and DFS. Our data suggest that BU/Mel, being associated with a low toxicity profile (mainly mucositis) and mortality, is an effective conditioning regimen even for high-risk AML patients in first CR undergoing auto-SCT.

Fludarabine, arabinosyl cytosine and idarubicin (FLAI) for remission induction in poor-risk acute myeloid leukemia.

Progress in treatment of acute myeloid leukemia (AML) is slow and treatment intensification alone has limited effects, particularly in poor-risk cases. Poor-risk cases, that are identified mainly by prior history, leukemic cell mass and cytogenetic abnormalities, share multiple mechanisms of drug resistance that are responsible for treatment failure. Since Pgp-mediated resistance to anthracycline can be reduced with Idarubicin (IDA) and resistance to arabinosyl cytosine (AC) can be reduced with Fludarabine (FLUDA), we tested a combination of high dose AC (2000 mg/sqm, 5 doses), FLUDA (30 mg/sqm, 5 doses) and IDA (12 mg/sqm, 3 doses) for remission induction and consolidation in 45 consecutive cases of poor-risk AML. The complete remission (CR) rate was 71% after the first course and 82% overall, with a projected 2-year survival and relapse-free survival of 44% and 50% respectively. Non-hematologic toxicity was very mild, that is very important in elderly patients, but hemopoietic toxicity was substantial, with a time to hematologic recovery of 3 to 4 weeks and two cases of death in CR. Peripheral blood stem cells (PBSC) could be mobilized and collected successfully only in 11 cases. This three-drug combination is effective and has a limited non-hematologic toxicity, but FLUDA may increase the difficulty of obtaining PBSC early after remission induction.

The lung as a target organ in patients with hematologic disorders.

The lung is one of the organs most severely affected by complications during the course of hematologic disorders. In the last years an impressive amount of progress has been made in clarifying the pathogenesis of lung diseases, particularly those occurring in conditions of severe immunosuppression such as bone marrow transplantion, acquired immunodeficiency syndrome or leukemia. Peculiar anatomical characteristics render the lung parenchyma highly susceptible to infections, but the clinical outcome is due not only to the injury induced by the pathogens but also to their interactions with inflammatory cells and particularly to the effects of a wide network of secreted cytokines. Polymorphonuclear cells, macrophages, lymphocytes and structural pulmonary cells (epithelial cells, interstitial cells) generate a variety of cytokines and growth factors which, in turn, may be responsible for the majority of the clinical effects in response to infections, such as those of Pneumocystis carinii and cytomegalovirus, but also to certain drugs or to radiation. The pathogenesis of graft-versus-host disease (GVHD) is still poorly understood, but animal models seem to demonstrate the involvement of a number of cytokines and growth factors, together with toxic effects induced by conditioning regimens.

A prospective study of alpha-interferon and autologous bone marrow transplantation in chronic myeloid leukemia. The Italian Co-operative Study Group on Chronic Myeloid Leukemia.

BACKGROUND AND OBJECTIVE: Alpha-interferon (alphaIFN) can induce cytogenetic remissions in chronic myeloid leukemia (CML). Hemopoietic progenitors can be collected from the marrow in remission and utilized for autologous repopulation after high dose chemotherapy. This study was designed with the purpose of evaluating the feasibility of a combined treatment policy of alphaIFN followed by autologous bone marrow transplantation (autoBMT). DESIGN AND METHODS: A prospective study of alphaIFN and autoBMT was begun in 1989. Two hundred and seventy-two consecutive previously untreated non-blastic Ph positive chronic myeloid leukemia (CML) patients, who were less than 56 years old, were enrolled over a 3-year period (1989-1991) and were assigned to receive human recombinant alphaIFN 2a (Roferon-A) at a dose of 9 MIU daily for at least one year. If they achieved a cytogenetic response consisting in a percentage of Ph neg metaphases of more than 25%, they were eligible for marrow harvesting and subsequent autografting after high dose busulfan (16 mg/kg) and melphalan (60 mg/m(2)). RESULTS: Seventy-six patients (28%) were eligible for a marrow harvest but the marrow was harvested in only 37 cases (14%), and only twenty-three patients (8%) were actually autografted. One patient died of infection nine days after autoBMT. The other patients recovered and did not suffer any late adverse events. Five patients progressed to blastic phase, six are alive in complete hematologic remission and eleven are alive in complete hematologic and cytogenetic remission. AlphaIFN treatment was reinstituted after autoBMT in 18 of 22 cases, but four patients who are in continuous complete cytogenetic remission were not given alphaIFN anymore. The progression-free survival of the autografted patients is 65% 8 years after registration. INTERPRETATION AND CONCLUSIONS: This study shows that bone marrow hemopoietic progenitors (Ph neg and Ph pos) can be collected from patients who respond to alphaIFN and can be used to rescue hemopoietic activity after high dose chemotherapy. Though some complete and durable cytogenetic remissions were obtained, the treatment could be applied only to a small group of good risk patients, highlighting that selection is very important and results cannot be extrapolated to the average patient.

Big BU/CY is associated with a favorable long-term outcome in patients allotransplanted for chronic myelogenous leukemia in chronic phase.

Twenty-six adult patients, median age 36 years (range 21-53) with chronic myeloid leukemia in first chronic phase were allotransplanted between October 1989 and May 1995. The preparative regimen consisted of busulphan 16 mg/kg and cyclophosphamide 200 mg/kg (big BU/CY). Cyclosporin A and methotrexate were used for GVHD prophylaxis. Twenty-two donors were HLA-identical siblings and four donors were mismatched for one antigen of class I. The global incidence of acute GVHD was 50%, that of severe aGVHD (grades 3-4) was 11%; the global incidence of chronic GVHD was 30%. No patients developed veno-occlusive disease of the liver or interstitial pneumonia. Five patients died, one of relapse, four of transplant-related causes, mostly related to aGVHD; thus, the transplant-related mortality was 16%. Twenty-one patients are alive, in remission, with a median follow-up of 55 months (range 24-90); actuarial probability of survival is 78% (CI 64-96). Our study shows that this conditioning regimen is relatively easy to administer and seems to be as effective as, if not superior to, regimens containing TBI, in patients with chronic myeloid leukemia in chronic phase and the transplant-related mortality is not excessive even in older patients.

Prior treatment with alpha-interferon does not adversely affect the outcome of allogeneic BMT in chronic phase chronic myeloid leukemia.

BACKGROUND AND OBJECTIVE: Controlled clinical trials have shown that Interferon-alpha (IFN-alpha) is able to control myeloid proliferation and to suppress the Ph+ clonal hemopoiesis in early chronic phase chronic myeloid leukemia (CML): a growing number of patients are treated with this agent from diagnosis. However, if a CML patient has an HLA-identical sibling, bone marrow transplant (BMT) represents the best choice of treatment. Since IFN-alpha is known to modify the immunologic response and to increase marrow fibrosis, information is needed on the outcome of patients transplanted after IFN-alpha treatment. DESIGN AND METHODS: We analyzed retrospectively 32 Ph+ CML patients submitted to BMT in the last 6 years in Institute "Serágnoli". All the patients were in 1st chronic phase, their median age was 37 years, the donors were HLA-identical (27/32) or 1 Ag-mismatched (5/32) siblings. Big BuCy was the conditioning regimen employed for all and GVHD prophylaxis was based on CsA in 4 patients and Csa+MTX in 28 patients; all patients received homogeneous pre and post-transplant supportive care, antimicrobial and antiviral prophylaxis. These patients were divided into 2 groups according to the treatment before BMT: 16 received IFN from diagnosis to BMT (mean dose 6.9 MU/daily) for at least 6 mos (mean 23 mos, range 8-75) and 16 received chemotherapy alone (hydroxyurea [HU]). RESULTS: Hematological recovery was comparable in the two groups: time to 0.5 x 10(9)/L PMN was 20.5 days (range 11-32) in the IFN group and 20 days (range 10-32) in the HU group; time to 50 x 10(9)/L platelets was 28 days (range 20-117) in the IFN group and 27 days (range 20-112) in the HU group. The incidence of acute GVHD was not different in the two groups for any grade of the disease; in patients who survived more than 100 days, chronic GVHD occurred in the two groups with the same frequency. Seven patients died of transplant related mortality (TRM), 4 in the IFN group and 3 in the HU group. Hematological relapse was observed in only one case in the HU group; no cytogenetic relapse occurred. Disease free survivals at 7 years are 61% and 72%, respectively; the difference is not significant. INTERPRETATIONS AND CONCLUSIONS: Notwithstanding the low number of patients included in this study, the data reported here confirm that prior treatment with alpha-IFN does not adversely affect transplant outcome.

Autoimmune mediated thrombocytopenia associated with the use of interferon-alpha in chronic myeloid leukemia.

We report on two patients with Ph+ chronic myeloid leukemia (CML) in chronic phase who developed severe thrombocytopenia during treatment with interferon-alpha 2A (IFN-alpha 2A). In both cases, we detected the presence of platelet-associated antibodies (PAIg) by autoimmune flow cytometry. We postulated an immune mediated platelet destruction mechanism; corticosteroid therapy was employed and interferon therapy was withdrawn, resulting in an increase in platelet count and a reduction of PAIg. Our observation reports the detection of PAIg associated with IFN-alpha 2A therapy in CML and suggests that this immunomodulant drug could induce thrombocytopenia through a mechanism other than antiproliferation.

Cost-effectiveness of interferon-alpha and conventional chemotherapy in chronic myelogenous leukemia.

OBJECTIVE: To compare the cost-effectiveness of interferon-alpha with that of hydroxyurea as initial therapy for patients with chronic myelogenous leukemia (CML) in the chronic phase. DESIGN: A decision analysis and Markov model that described the natural history of the therapeutic process. The Markov model contained two treatment arms (interferon-alpha and hydroxyurea) and eight states of health (complete hematologic remission with cytogenetic response, complete hematologic remission without cytogenetic response, partial hematologic remission, chronic phase without hematologic remission, accelerated phase, blast crisis, bone marrow transplantation, and death). Probabilities, costs, and utilities were obtained from published clinical studies and clinical investigators. MEASUREMENT: Quality-adjusted years of life saved and costs and qualities discounted at 5% per year. SETTING: University medical centers in North America and Europe. PATIENTS: Meta-analysis of results from patients studied in clinical trials. RESULTS: The model's predictions of median survival (69 months with interferon-alpha therapy and 58 months with hydroxyurea therapy) were derived from data in the recent literature. In patients 50 years of age, interferon-alpha improved life expectancy over hydroxyurea by approximately 18 months. The marginal cost-effectiveness of interferon-alpha (incremental discounted cost of interferon-alpha compared with that of conventional therapy) was $34800 per quality-adjusted year of life saved. The model was sensitive to the monthly cost of interferon-alpha therapy (if the cost of interferon-alpha is reduced by one third, the cost-effectiveness becomes $19300 per quality-adjusted year of life saved) but was not particularly sensitive to the costs associated with blast crisis or bone marrow transplantation. The other significant variable was quality of life during therapy with interferon-alpha; when this measure was varied from 70% to 100% of the quality of life during hydroxyurea therapy, cost-effectiveness changed from $123200 to $25620 per quality-adjusted year of life saved. When the quality of life associated with interferon-alpha was less than 62% of the quality of life associated with hydroxyurea, the discounted quality-adjusted life expectancy with interferon-alpha was less than that with hydroxyurea. CONCLUSION: Compared with hydroxyurea, interferon-alpha is, in most clinical scenarios, a cost-effective initial therapy for patients with chronic-phase CML who can tolerate the drug.

Neutralizing anti-interferon-alpha antibodies and response to treatment in patients with Ph+ chronic myeloid leukaemia sequentially treated with recombinant (alpha 2a) and lymphoblastoid interferon-alpha.

Neutralizing anti-IFN alpha antibodies (nIFN alpha Abs) occur in a significant proportion of patients with hairy cell leukaemia, hepatitis or solid tumours treated with recombinant IFN alpha (IFN alpha 2a or IFN alpha 2b), but information on their incidence in chronic myeloid leukaemia (CML) is scanty and their clinical relevance is not yet completely defined. By using an IFN alpha antiviral neutralization bioassay, the frequency of nIFN alpha 2a Abs was evaluated in 67Ph+ CML patients during IFN alpha 2a therapy at doses ranging from 6 to 9 MU/d. 15 patients (22%) developed nIFN alpha 2a Abs (titre ranging from 1:40 to 1:20480) and 11/15 (73%) were haematologically and/or karyotypically unresponsive to treatment. 52 patients did not develop antibodies and 11 of them (21%) were unresponsive. The negative relationship between the positivity for nIFN alpha 2a Abs and the response to treatment was highly significant (P = 0.0001). In nine nIFN alpha 2a Abs positive patients, treatment was changed from recombinant IFN alpha 2a to lymphoblastoid IFN alpha (IFN alpha-ly), at the same dose and schedule. After 9 months of IFN alpha-ly treatment a haematological response was achieved in 4/7 cases who were non-responsive to prior IFN alpha 2a therapy and was maintained in the other two patients previously responsive to IFN alpha 2a. However, no karyotypic response was observed. This data shows that a significant proportion of Ph+ CML patients receiving treatment with IFN alpha 2a can develop neutralizing antibodies and that these antibodies are associated with a loss of IFN alpha 2a efficacy. Changing the patients to treatment with lymphoblastoid IFN alpha may restore haematological response but it is not likely to induce a karyotypic response.

FLANG (fludarabine + cytosine arabinoside + novantrone + G-CSF) induces partial remission in lymphoid blast transformation of Ph+chronic myelogenous leukaemia.

The adenine nucleoside analogue, fludarabine phosphate, in combination with cytosine-arabinoside (Ara-C) and granulocyte-colony stimulating factor (G-CSF) (the so called FLAG regimen) has recently been shown to be effective in the treatment of poor-prognosis acute non-lymphoid leukaemia. We used this combination plus novantrone (FLANG regimen) in a case of Ph1+ chronic myeloid leukaemia (CML) unresponsive to interferon alpha that had progressed to an acute phase, after 3 months of treatment with 6-mercaptopurine and hydroxyurea. The patient was treated with two courses of fludarabine 30 mg/m2 (days 1-5) + Ara-C 2 g/m2 (days 1-5) + novantrone 5 mg/m2 (days 1-3) and G-CSF from day 0 to neutrophil recovery. After the first cycle of chemotherapy, bone marrow blasts decreased from 100% to less than 5% (clinical complete remission), with a progressive clearance of Ph1+ metaphases (from 100% to 12%). At the end of the second course, a progressive increase of blasts was observed again and karyotypic detection of Ph+ cells was also documented (from 12% to 42.9%). During this partial remission, the patient underwent an allogeneic bone marrow transplantation from an HLA matched identical brother. At the time of this report, he is still alive and well and in complete karyotypic remission. This partial therapeutic success was compared with the result obtained in another previously reported CML case: differences in the therapeutic efficacy of protocols employing fludarabine nucleosides and the type of blastic cells involved are discussed.

Interferon-alpha and hydroxyurea in early chronic myeloid leukemia: a comparative analysis of the Italian and German chronic myeloid leukemia trials with interferon-alpha.

In 1994, the Italian and the German Chronic myeloid leukemia (CML) trials comparing interferon-alpha (IFN-alpha) with conventional chemotherapy were published. The survival advantage in favor of IFN-alpha compared with hydroxyurea (HU; 72 v 52 months) was significant in the Italian (P < .002), but not in the German trial (66 v 56 months, P < .44). We set up a collaborative study to identify the reasons for the different outcomes. There are major differences in the trial protocols concerning admission criteria, treatment strategy, and definitions. The German patients were older and more seriously sick. Fifty-two of the 327 patients in the German IFN and HU arms did not fulfil Italian admission criteria, and 41 of the 322 Italian patients did not fulfil German admission criteria. Using mutually uniform admission criteria, the median survival times of the IFN patients are 76 (Italian) and 72 (German) months (P = .56). The Italian group administered IFN combined with HU as needed, whereas the German group strictly used IFN as monotherapy with rerandomization to busulfan (BU) or HU after IFN resistance or intolerability. The differences seen between the Italian and the German trial results can be accounted for by objective differences in study design, especially the admission criteria, treatment strategy, and bias due to intention to treat analysis. The detailed analysis of the data suggests that the combination of IFN with HU as needed is more effective than either agent alone.

Interferon-alpha in the treatment of chronic myeloid leukemia. A summary and an update of the Italian studies.

Six-hundred patients were recruited between 1986 and 1991 for studies of the treatment of Ph positive chronic myeloid leukemia (CML) with interferon-alpha (IFN-alpha). The median survival of the patients who were assigned to treatment with IFN-alpha was 6 years or longer than 6 years, and was more than the survival of the patients who were assigned to conventional chemotherapy. Survival prolongation was significantly related with the achievement of a cytogenetic response. IFN-alpha treatment was not harmful for subsequent allogeneic or autologous bone marrow transplantation.

A new method of "in-cell reverse transcriptase-polymerase chain reaction" for the detection of BCR/ABL transcript in chronic myeloid leukemia patients.

Methods of detecting minimal residual disease (MRD) in chronic myeloid leukemia (CML) include chromosome analysis, Southern blotting, polymerase chain reaction (PCR) and fluorescent in situ hybridization (FISH) techniques. We report a novel method to detect intracellular messenger RNA (mRNA) by combining the techniques of reverse transcription (RT) and PCR performed directly inside the cells, without extraction of the nucleic acid. We applied this method, which we call "in-cell RT-PCR", to detect hybrid BCR/ABL transcript within single cells. After cellular permeabilization and fixation of single cells in suspension, the neoplastic mRNA was reverse transcribed into cDNA, and the cDNA was amplified by PCR with fluorescent primers, specific for bcr/abl. Flow cytometry was used to detect cells positive for the amplified DNA within the cell cytoplasm. After transferring the amplified cells onto slides by cytospin, the positive cells for BCR/ABL cDNA were observed by fluorescent microscopy. The technique was capable of detecting low abundancy signals and distinguishing different levels of gene expression. The amplification products were found in the cells and supernatants. The distribution was critically affected by the protease digestion condition. The specificity of amplification was confirmed by a nested RT-PCR of BCR/ABL performed on extracted mRNA from the same sample, and by reamplification of supernatants. We have used the technique to study 10 Ph+ CML patients and three normal subjects as controls. Four patients were 100% Ph+ at diagnosis time and RT-PCR+ at cytogenetic and molecular analysis, respectively. In-cell RT-PCR showed that the residual non-neoplastic cells could be observed in all cases. In two patients undergoing interferon-alpha (IFN-alpha) therapy and in four bone-marrow transplanted patients, the in-cell RT-PCR was used to compare the level of Ph+ positivity detected by cytogenetic analysis with the number of cells expressing BCR/ABL transcript. In this manner, we could estimate the MRD. Our preliminary application of the technique suggests that it is capable of accurately identifying cells transcribing bcr/abl, and that it may have significant clinical applications in the detection of MRD.

Does the type of BCR/ABL junction predict the survival of patients with Ph1-positive chronic myeloid leukemia?

The prognostic value of the site of DNA rearrangement within the M-BCR on chromosome 22 or of the type of transcript has been debated in the last years. The majority of the studies do not support the hypothesis of a predictive value of such molecular parameters. Results coming from a multicentric, prospective trial, based on alpha-IFN therapy, seem to indicate a better karyotypic response in 3' rearranged patients. The possibility of evoking a cytotoxic immune response directed towards peptides originating from each of the different BCR/ABL junctions constitute an important challenge for the future.

Apoptosis induction with fludarabine on freshly isolated chronic myeloid leukemia cells.

BACKGROUND: Fludarabine (FLU) is a fluorinated purine analogue with antineoplastic activity in lymphoproliferative malignancies. Recently, some in vitro reports have showed the effective role of FLU on the activation of apoptosis. MATERIALS AND METHODS: The induction of programmed cell death (apoptosis) by fludarabine (FLU), an adenine nucleoside analogue, alpha-interferon (alpha-IFN), and FLU plus alpha-IFN was evaluated in vitro against freshly isolated, chronic-phase Ph1+ chronic myeloid leukemia (CML). Apoptosis was detected by electrophoresis gel of DNA oligonucleosomal fragments in 8 CML samples. RESULTS: Only FLU and FLU plus alpha-IFN significantly activated apoptosis in all the samples, suggesting selective activity on CML cells. On the other hand, alpha-IFN alone did not activate programmed cell death. CONCLUSIONS: Our data show apoptotic activity for FLU on CML cells. Programmed cell death may be suppressed in cells carrying the bcr-abl transcript, and FLU might remove this resistance in the neoplastic cell cycle. This preliminary report justifies using FLU in pilot clinical trials for chronic phase Ph1+ CML patients.

The type of BCR/ABL junction does not predict the survival of patients with Ph1-positive chronic myeloid leukaemia.

The prognostic value of the location of the breakpoint on chromosome 22 in patients with Ph1+ chronic myeloid leukaemia (CML) is still controversial. We analysed both DNA rearrangement and transcript type in a new continuous series of CML patients. By Southern blotting analysis, we found that, out of 72 patients, 43 had a 5' rearrangement and 29 a 3' one, of the 43 5'-rearranged patients, 35 carried an a2b2 transcript and eight an a2b3 one, while, of the 29 patients rearranged in the 3' part of the M-BCR area, 26 had an a2b3 transcript, one had an a2b2 transcript and two carried both types of transcript. Thus, mRNA studies allow to detect an a2b3 transcript in 17.7% of 5' rearranged patients. However, no correlation was observed between type of transcript and survival, as after DNA studies.