[Show abstract][Hide abstract] ABSTRACT: BACKGROUND: The monitoring of minimal residual disease (MRD) through low sensitivity real-time (RT) PCR analysis of BCR-ABL transcripts allows early detection of chronic myeloid leukemia(CML) relapse after allogeneic hematopoietic stem cell transplantation (HSCT). The introduction of more sensitive techniques, such as RT quantitative (Q)-PCR, may lead to overestimate the risk of CML relapse. PATIENTS AND METHODS: In this study we reviewed the results of peripheral blood (PB) RT Q-PCR in CML patients who underwent allogeneic HSCT from 1983 to 2007. In our laboratory, RT Q-PCR analysis was routinely performed since 2002. Eighty-seven out of 189 patients had available RT Q-PCR data; 63 patients had at least three RT Q-PCR analyses evaluable. RESULTS: Fifty-two out of 63 patients (83%) had, at least once, detectable transcript levels, but with BCR-ABL/ABL ratio <0.1% defined as <MR3, whereas 11 (17%) had persistent undetectable BCR-ABL transcripts. Six out of 52 patients with <MR3 relapsed, defined as BCR-ABL transcript numbers > 0.1% confirmed by the finding of Ph+ positive cells in bone marrow. No patients with persistent undetectable transcripts relapsed (p=0.19). Relapse did not correlate with the number of occurrences of <MR3, or with the time to the first <MR3 result. Finally, out of 46 patients with detectable transcripts who did not relapse, 35 had undetectable transcripts at last contact. RT Q-PCR analysis had low specificity (19%) and low positive predictive value (12%) in predicting relapse of CML patients after allogeneic HSCT. CONCLUSION: Our data suggest that detectionof low BCR-ABL transcript levelsby RT Q-PCR analysis has a poor accuracy in defining the risk of CML relapse, and it should not be considered as the unique indication to treatment. Fluctuation of BCR-ABL transcripts levels is common as late as ≥10 years after transplant, possibly suggesting the long-term persistence of CML stem cells.
Biology of blood and marrow transplantation: journal of the American Society for Blood and Marrow Transplantation 01/2013; · 3.15 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Nilotinib is a potent and selective BCR-ABL inhibitor. The phase 3 ENESTnd trial demonstrated superior efficacy of nilotinib vs imatinib, with higher and faster molecular responses. After a median of 24 months, the rates of progression to accelerated-blastic phase (ABP) were 0.7% and 1.1% with nilotinib 300mg and 400mg BID, respectively, significantly lower compared to imatinib (4.2%). Nilotinib has been approved for the frontline treatment of Ph+ CML. With imatinib 400mg (IRIS trial), the rate of any event and of progression to ABP were higher during the first 3 years. Consequently, a confirmation of the durability of responses to nilotinib beyond 3 years is extremely important. Aims. To evaluate the response and the outcome of patients
treated for 3 years with nilotinib 400mg BID as frontline therapy.
[Show abstract][Hide abstract] ABSTRACT: The median age of chronic myeloid leukemia (CML) patients is ~60 years, and age is still considered an important prognostic factor, included in Sokal and EURO risk scores. However, few data are available about the long-term outcome of older patients treated with imatinib (IM) frontline. We analyzed the relationship between age and outcome in 559 early chronic-phase CML patients enrolled in 3 prospective clinical trials of Gruppo Italiano Malattie Ematologiche dell'Adulto CML Working Party, treated frontline with IM, with a median follow-up of 60 months. There were 115 older patients (≥ 65 years; 21%). The complete cytogenetic and major molecular response rates were similar in the 2 age groups. In older patients, event-free survival (55% vs 67%), failure-free survival (78% vs 92%), progression-free survival (62% vs 78%), and overall survival (75% vs 89%) were significantly inferior (all P < .01) because of a higher proportion of deaths that occurred in complete hematologic response, therefore unrelated to CML progression (15% vs 3%, P < .0001). The outcome was similar once those deaths were censored. These data show that response to IM was not affected by age and that the mortality rate linked to CML is similar in both age groups. This trial was registered at www.clinicaltrials.gov as #NCT00514488 and #NCT00510926.
[Show abstract][Hide abstract] ABSTRACT: Variant Philadelphia (Ph) chromosome translocations have been reported in 5%-10% of patients with newly diagnosed chronic myeloid leukemia (CML). Variant translocations may involve one or more chromosomes in addition to 9 and 22, and can be generated by 2 different mechanisms, 1-step and 2-step rearrangements, as revealed by fluorescence in situ hybridization. The prognostic significance of the occurrence of variant translocations has been discussed in previous studies. The European LeukemiaNet recommendations do not provide a "warning" for patients with variant translocations, but there is limited information about their outcome after therapy with tyrosine kinase inhibitors. To identify the role of variant translocations in early chronic phase (CP) CML patients treated with imatinib mesylate, we performed an analysis in a large series of 559 patients enrolled in 3 prospective imatinib trials of the Gruppo Italiano Malattie EMatologiche dell'Adulto (GIMEMA) Working Party on CML. Variant translocations occurred in 30 patients (5%). Our data show that the presence of variant translocations has no impact on the cytogenetic and molecular response or on outcome, regardless of the involvement of different mechanisms, the number of involved chromosomes, or the presence of deletions. Therefore, we suggest that patients with variant translocations do not constitute a "warning" category in the imatinib era. This study is registered at www.clinicaltrials.gov as NCT00514488 and NCT00510926.
[Show abstract][Hide abstract] ABSTRACT: In patients with Philadelphia-positive acute lymphoblastic leukemia, resistance to treatment with tyrosine kinase inhibitors is frequent and most often associated with the development of point mutations in the BCR-ABL kinase domain. We aimed to assess: (i) in how many patients BCR-ABL kinase domain mutations are already detectable at relatively low levels at the time of diagnosis, and (ii) whether mutation detection correlates with subsequent response to therapy.
We retrospectively analyzed samples collected at diagnosis from 15 patients with Philadelphia-positive acute lymphoblastic leukemia who subsequently received tyrosine kinase inhibitor therapy (dasatinib) by cloning the BCR-ABL kinase domain in a bacterial vector and sequencing 200 independent clones per sample.
Mutations at relatively low levels (2-4 clones out of 200) could be detected in all patients--eight who relapsed and seven who achieved persistent remission. Each patient had evidence of two to eight different mutations, the majority of which have never been reported in association with resistance to tyrosine kinase inhibitors. In two patients out of six who relapsed because of a mutation, the mutation (a T315I) was already detectable in a few clones at the time of diagnosis. On the other hand, a patient who was found to harbor an F317L mutation is in persistent remission on dasatinib.
Our results suggest that the BCR-ABL kinase domain is prone to randomly accumulate point mutations in Philadelphia-positive acute lymphoblastic leukemia, although the presence of these mutations in a relatively small leukemic subclone does not always preclude a primary response to tyrosine kinase inhibitors.
[Show abstract][Hide abstract] ABSTRACT: Deletions of the derivative chromosome 9 [der(9)] have been associated with a poor prognosis in chronic myeloid leukemia (CML) across different treatment modalities. In the imatinib era, the prognostic impact of der(9) deletions has been evaluated mainly in patients with late chronic-phase (CP) CML, giving partially conflicting results. Few data are available in the early CP setting. For this reason, in 2006, the European LeukemiaNet recommendations still considered der(9) deletions as a candidate adverse prognostic factor and required a careful monitoring of the patient.
To investigate the prognostic value of der(9) deletions in early CP CML, we performed an analysis of three prospective imatinib trials of the Italian Group for Hematological Malignancies of the Adult (GIMEMA) CML Working Party.
A fluorescent in situ hybridization (FISH) analysis of bone marrow cells was performed at diagnosis; der(9) deletions were detected in 60 (12%) of 521 evaluable patients. At 60 months, the cumulative incidence of complete cytogenetic response and major molecular response-and the probability of event-free survival, failure-free survival, progression-free survival, and overall survival-in patients with and without deletions were not statistically different.
Our data strongly support the notion that, when investigated by FISH, der(9) deletions are not a poor prognostic factor in patients with early CP CML treated with imatinib.
Journal of Clinical Oncology 06/2010; 28(16):2748-54. · 18.04 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Background / Purpose:
Age has no impact on the outcome of patients with Philadelphia-Positive Chronic Myeloid Leukemia (CML) in early chronic phase (CP) treated with imatinib. We found that response to imatinib was not affected by old age (defined as more than 65 years). Differences in survival curves between older and younger patients were determined by age-related complications and comorbidities.In the prognostic classifications (Sokal 1984; Hasford 1998) that were elaborated before the introduction of imatinib, age was a significant and important prognostic factor. However, few and not conclusive data are available for patients in early CP treated with imatinib with the consequence that imatinib therapy is still denied to several elderly patients. To assess the effects of imatinib on response and outcome, according to the age at diagnosis, we analyzed a large series of early CP CML patients.We analyzed the data of 559 previously untreated early CP patients who were assigned to receive imatinib 400 mg daily (76%) or 800 mg daily (24%) in three controlled, prospective, independent studies of GIMEMA CML WORKING PARTY. The median follow-up was 42 (extremes 1-64) months. Patients were divided in 2 groups of age, using the cut-off of 65 years old (WHO definition of “old” person): 444 (79%, group A) were less than 65 years (median age 46 years), while 115 patients (21%, group B) were more than 65 years old (median age 71 years). The proportion of patients who were treated with imatinib 800 mg daily was the same in both age groups. Patients were evaluated for hematologic, cytogenetic and molecular response; failure were defined according to ELN criteria (Baccarani 2006); events were considered failures and discontinuation of imatinib for any cause, including death. Survival curves were calculated using the Kaplan-Meier method and all analysis were performed according to the intention-to-treat principle.
No differences between younger (group A) and older (group B) patients were observed in terms of Complete Cytogenetic Response (CCyR; about 89%) and Major Molecular Response (MMR; about 87%).These survival differences were mainly due to CML-unrelated comorbidities and deaths (Table 4 in Poster). An analysis of FFS, PFS and OS without CML-unrelated deaths shows no significant difference in the 2 groups of age.This study represents an important step in demonstrating that imatinib has the same efficacy also in older age patients; this means that age should never be a contraindication to imatinib treatment. Also elderly patients should be included in clinical trials. However larger studies are needed to definitely eliminate age as a prognostic factor in CML.
51st American Society of Hematology Annual Meeting 2009; 03/2010
[Show abstract][Hide abstract] ABSTRACT: Before the introduction of imatinib, interferon alpha-based regimens were the gold standard for treatment of early chronic phase chronic myeloid leukemia patients. The combination of IFN-alpha with imatinib is currently being investigated in at least two large clinical trials, the German CML Study IV and the French SPIRIT trial. We reviewed the cytogenetic and molecular responses of 76 early chronic phase chronic myeloid leukemia patients who were treated with imatinib and interferon-alpha and of 419 early chronic phase chronic myeloid leukemia patients treated with imatinib alone front-line. The complete cytogenetic response rate was higher in the IM+IFN-alpha group than in the imatinib group at six months (60% vs. 42%; P=0.003), but not at 48 months (88% vs. 88%). The durability of the complete cytogenetic response was similar in the two groups with 94% and 91% of complete cytogenetic responders in continuous complete cytogenetic response at 48 months (P=0.56). The major molecular response rate was higher in the IM+IFN-alpha group at six months (58% vs. 34%; P=0.0001) and 12 months (67% vs. 47%; P=0.001) but not later on (65% vs. 57% at 48 months; P=0.25). Overall and progression free survival were comparable in the two groups; a significant trend to a better event free survival was observed in patients treated with PegIFNalpha (91% vs. 78%; P=0.02). These data suggest that the response to the combination treatment is more rapid. It is not yet known how much a rapid reduction will influence the longer-term overall and progression free survival, and the cure rate.
[Show abstract][Hide abstract] ABSTRACT: In chronic myeloid leukemia, different methods are available to monitor the response to therapy: chromosome banding analysis (CBA), interphase fluorescence in situ hybridization (I-FISH), and real-time quantitative polymerase chain reaction (RT-Q-PCR). The GIMEMA CML WP (Gruppo Italiano Malattie Ematologiche Adulto Chronic Myeloid Leukemia Working Party) has performed a prospective study to compare CBA and I-FISH for the definition of complete cytogenetic response (CCgR). Samples (n = 664) were evaluated simultaneously by CBA and I-FISH. Of 537 cases in CCgR, the number of positive nuclei by I-FISH was less than 1% in 444 cases (82.7%). Of 451 cases with less than 1% positive nuclei by I-FISH, 444 (98.4%) were classified as CCgR by CBA. The major molecular response rate was significantly greater in cases with I-FISH less than 1% than in those with I-FISH 1% to 5% (66.8% vs 51.6%, P < .001) and in cases with CCgR and I-FISH less than 1% than in cases with CCgR and I-FISH 1% to 5% (66.1% vs 49.4%, P = .004). I-FISH is more sensitive than CBA and can be used to monitor CCgR. With appropriate probes, the cutoff value of I-FISH may be established at 1%. These trials are registered at http://www.clinicaltrials.gov as NCT00514488 and NCT00510926.
[Show abstract][Hide abstract] ABSTRACT: We show the molecular and functional characterization of a novel population of lineage-negative CD34-negative (Lin(-)CD34(-)) hematopoietic stem cells from chronic myelogenous leukemia (CML) patients at diagnosis. Molecular karyotyping and quantitative analysis of BCR-ABL transcript demonstrated that approximately one-third of CD34(-) cells are leukemic. CML Lin(-)CD34(-) cells showed kinetic quiescence and limited clonogenic capacity. However, stroma-dependent cultures induced CD34 expression on some cells and cell cycling, and increased clonogenic activity and expression of BCR-ABL transcript. Lin(-)CD34(-) cells showed hematopoietic cell engraftment rate in 2 immunodeficient mouse strains similar to Lin-CD34(+) cells, whereas endothelial cell engraftment was significantly higher. Gene expression profiling revealed the down-regulation of cell-cycle arrest genes and genes involved in antigen presentation and processing, while the expression of genes related to tumor progression, such as angiogenic factors, was strongly up-regulated compared with normal counterparts. Phenotypic analysis confirmed the significant down-regulation of HLA class I and II molecules in CML Lin(-)CD34(-) cells. Imatinib mesylate did not reduce fusion transcript levels, BCR-ABL kinase activity, and clonogenic efficiency of CML Lin(-)CD34(-) cells in vitro. Moreover, leukemic CD34(-) cells survived exposure to BCR-ABL inhibitors in vivo. Thus, we identified a novel CD34(-) leukemic stem cell subset in CML with peculiar molecular and functional characteristics.
[Show abstract][Hide abstract] ABSTRACT: Nilotinib has a higher binding affinity and selectivity for BCR-ABL with respect to imatinib and is an effective treatment of chronic myeloid leukemia (CML) after imatinib failure. In a phase 2 study, 73 early chronic-phase, untreated, Ph(+) CML patients, received nilotinib at a dose of 400 mg twice daily. The primary endpoint was the complete cytogenetic response (CCgR) rate at 1 year. With a median follow-up of 15 months, the CCgR rate at 1 year was 96%, and the major molecular response rate 85%. Responses were rapid, with 78% CCgR and 52% major molecular response at 3 months. During the first year, the treatment was interrupted at least once in 38 patients (52%). The mean daily dose ranged between 600 and 800 mg in 74% of patients, 400 and 599 mg in 18% of patients, and was less than 400 mg in 8% of patients. Dose interruptions were mainly due to nonhematologic and biochemical side effects. Myelosuppression was irrelevant. One patient progressed to blastic crisis after 6 months; one went off-treatment for lipase increase grade 4 (no pancreatitis). Nilotinib is safe and very active in early chronic-phase CML. These data support a role for nilotinib for the frontline treatment of CML. This study was registered at ClinicalTrials.gov as NCT00481052.
[Show abstract][Hide abstract] ABSTRACT: Dasatinib and nilotinib are tyrosine kinase inhibitors (TKIs) developed to overcome imatinib resistance in Philadelphia-positive leukemias. To assess how Bcr-Abl kinase domain mutation status evolves during sequential therapy with these TKIs and which mutations may further develop and impair their efficacy, we monitored the mutation status of 95 imatinib-resistant patients before and during treatment with dasatinib and/or nilotinib as second or third TKI. We found that 83% of cases of relapse after an initial response are associated with emergence of newly acquired mutations. However, the spectra of mutants conferring resistance to dasatinib or nilotinib are small and nonoverlapping, except for T315I. Patients already harboring mutations had higher likelihood of relapse associated with development of further mutations compared with patients who did not harbor mutations (23 of 51 vs 8 of 44, respectively, for patients who relapsed on second TKI; 13 of 20 vs 1 of 6, respectively, for patients who relapsed on third TKI).
[Show abstract][Hide abstract] ABSTRACT: An increase in the serum concentration of pancreatic enzymes (amylase and lipase) was reported in a proportion of imatinib-resistant and/or intolerant Philadelphia-positive chronic myeloid leukemia patients treated with nilotinib. Acute pancreatitis was very rare, and the relevance of these laboratory alterations remains unknown. We report on 8 chronic myeloid leukemia patients who developed serum lipase/amylase elevation during treatment with nilotinib. After a median follow-up of 26 months, none of these patients developed an acute pancreatitis or clinical signs of pancreatic disease. Pancreatic hyperenzymemia never led to permanent drug discontinuation and required nilotinib temporary interruption in one case only. The median cumulative duration of dose interruptions and response to treatment were comparable in patients with or without pancreatic enzyme elevation. The mechanisms of action of nilotinib on pancreatic enzymes deserves to be investigated: however, in our experience, the relevance of pancreatic hyperenzymemia was clinically very limited.
[Show abstract][Hide abstract] ABSTRACT: Imatinib mesylate has become the treatment of choice for chronic myeloid leukemia (CML): the standard dose for chronic- phase (CP) CML is 400 mg daily. Response rates are different according to Sokal score, being significantly lower in intermediate and high Sokal risk patients. Phase 1 and 2 trials have shown a dose-response effect and high-dose imatinib trials in early CP CML showed better results compared with standard dose. Our study is the first prospective trial planned to evaluate the efficacy and tolerability of high-dose imatinib in previously untreated intermediate Sokal risk CML patients. Seventy-eight patients were treated with 400 mg imatinib twice daily: complete cytogenetic response (CCgR) rates at 12 and 24 months were 88% and 91%; moreover, at 12 and 24 months 56% and 73% of CCgR patients achieved a major molecular response. The incidence of adverse events was slightly higher than reported by the most important standard-dose trials. With a median follow-up of 24 months, 3 patients progressed to advanced phase. In intermediate Sokal risk newly diagnosed CML patients, high-dose imatinib induced rapid and high response rates, apparently faster than those documented in the International Randomized Study of IFN and Imatinib for the same risk category. These clinical trials are registered at www.clinicaltrials.gov as no. NCT00510926.
[Show abstract][Hide abstract] ABSTRACT: The outcome for adult with Ph + leukemias (ALL and CML) has improved dra- matically with current therapy including use of Tyrosine Kinase Inhibitors (TKIs), such as imatinib, nilotinib or dasatinib. The complete hematological remission is obtained in about 98% of early chronic phase CML patients treat- ed with TKIs. But the emergence of resistance to imatinib has become a signif- icant problem: the most common cause of imatinib resistance is the selection of leukemic clones with point mutations in the Abl kinase domain. These muta- tions lead to amino acid substitutions and prevent the appropriate binding of imatinib. These is an unexpected event in about 4% of CML patients, during first year of TKI therapy. Current approaches to risk classification based not only on well-established clinical parameters such as Sokal's Score for CML, but including genetic lesions of acute Ph + leukemia cell at diagnosis, as well as early response parameters are proposed. Several novel agents have been developed showing efficacy in overcoming imatinib resistance: new therapeutic approach- es that interfere specifically with mutated forms of Ph + leukemias and activate specifically the apoptotic pathway on leukemic blast cells are now available, such as Dasatinib, Nilotinib, Bosutinib and we highlights the latter as that may be applicable to the treatment of adult Ph + leukemias.
[Show abstract][Hide abstract] ABSTRACT: Ikaros plays an important role in the control of differentiation and proliferation of all lymphoid lineages. The expression of short isoforms lacking DNA-binding motifs alters the differentiation capacities of hematopoietic progenitors, arresting lineage commitment. We sought to determine whether molecular abnormalities involving the IKZF1 gene were associated with resistance to tyrosine kinase inhibitors (TKIs) in Ph+ acute lymphoblastic leukemia (ALL) patients. Using reverse-transcribed polymerase chain reaction, cloning, and nucleotide sequencing, only the non-DNA-binding Ik6 isoform was detected in 49% of Ph+ ALL patients. Ik6 was predominantly localized to the cytoplasm versus DNA-binding Ik1 or Ik2 isoforms, which showed nuclear localization. There was a strong correlation between nonfunctional Ikaros isoforms and BCR-ABL transcript level. Furthermore, patient-derived leukemia cells expressed oncogenic Ikaros isoforms before TKI treatment, but not during response to TKIs, and predominantly at the time of relapse. In vitro overexpression of Ik6 strongly increased DNA synthesis and inhibited apoptosis in TKI-sensitive cells. Genomic sequence and computational analyses of exon splice junction regions of IKZF1 in Ph+ ALL patients predicted several mutations that may alter alternative splicing. These results establish a previously unknown link between specific molecular defects that involve alternative splicing of the IKZF1 gene and the resistance to TKIs in Ph+ ALL patients.
[Show abstract][Hide abstract] ABSTRACT: Very recently the JAK2V617F mutation has been described in BCR⁄ABL+ patients for the first time. Even though characterized by very different clinical patterns, these cases raise some questions
about the biological meaning of the coexistence between these disease-specific molecular markers.
The present report confirms that the unexpected coexistence between these two disease-specific molecular markers - the
JAK2V617F mutation and the BCR⁄ABL translocation - is
European Journal Of Haematology 08/2008; 81(1):75-6. · 2.55 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In 2004, we reported the short-term results of a multicentric, phase 2 study of imatinib 400 mg daily and pegylated interferon-alpha in the treatment of 76 early chronic phase Philadelphia-positive chronic myeloid leukemia patients. In this report, we update the results with an observation time of five years. After two years of treatment, all but 10 patients (13%) had discontinued pegylated interferon-alpha. The complete cytogenetic response rate at five years was 87%, and 94% of complete cytogenetic responders maintained the complete cytogenetic response after five years. All but one complete cytogenetic response also achieved a major molecular response. These data confirm the excellent response to imatinib front-line and the stability of the complete cytogenetic response. Any possible additional benefit of the combination with interferon-alpha remains uncertain, due to low patient compliance.