Molecular Remission to Imatinib in Patients with Chronic Myeloid Leukaemia (CML) Is Less Durable Compared to Patients after Allografting.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 276-276
Author(s):  
Thoralf Lange ◽  
Thomas Bumm ◽  
Marc Mueller ◽  
Sandra Otto ◽  
Haifa K. Al-Ali ◽  
...  
Keyword(s):  
Nested Pcr ◽  
Bone Marrow Cells ◽  
Residual Disease ◽  
Standard Dose ◽  
Healthy Individuals ◽  
Rt Pcr ◽  
Molecular Remission ◽  
Conventional Cytogenetics ◽  
Risk Of Relapse

Abstract Objectives: Patients with CML who achieve molecular remission (MR, defined as a RT-PCR negativity for BCR-ABL transcripts) after myeloablative stem cell transplantation (SCT) have a low risk of relapse, and the majority may be cured. The frequency of MR on imatinib varies greatly and the durability of these responses has not been reported. To investigate if MR after SCT and on imatinib are equally stable, we directly compared two cohorts of patients treated with imatinib or SCT, respectively, from the time of their first negative RT-PCR result. Patients and Methods: One hundred and forty-four CML patients in chronic (n=104) or accelerated phase (n=40) treated with standard dose imatinib were routinely monitored by conventional cytogenetics, quantitative RT-PCR (qPCR) and conventional nested PCR in case of negative qPCR results. Nineteen patients (13.2%) had at least 1 negative nested PCR. To assess the level of residual disease in patients with a single negative RT-PCR result, 10 replicate reactions were performed, each corresponding to > 106 white bone marrow cells. Thirty-six samples (median 3, range 1–4) from patients in MR on imatinib and 45 samples (median 2, range 1–3) from patients in MR after SCT were available. Twenty samples from healthy individuals were tested as controls. Results: The first negative result was noted after a median of 16.8 months (range 11.5–36.1) of imatinib therapy and 6.6 months (range 4.7–9.5) after SCT, respectively. The projected risk of molecular relapse at 12 months after the first negative RT-PCR result was 83% in patients on imatinib but only 20% in patients after SCT (P = 0.0001). Only two patients on imatinib remained in molecular remission at 13.8 and 16.6 months. While none of the patients with molecular relapse after allograft lost CCyR, one patient on imatinib progressed to cytogenetic relapse. The replicate assay was positive in 18/36 samples (50%) from patients on imatinib, 8/46 (17.4%) after allografting and 4/20 (20%) from healthy individuals. These differences were significant between patients on imatinib and after allografting (P = 0.003) and between patients on imatinib and healthy individuals (P = 0.005), but not between patients after allografting and healthy individuals (P = 0.9). Negativity by replicate testing was more stable in patients after allografting, although, even in these patients, positive replicate reactions continued to occur with longer follow-up. Conclusion: Imatinib-induced MR is usually not durable, in contrast to MR after transplant. Consistent with this, the level of residual disease in samples negative by single nested PCR is higher in patients on imatinib compared to patients after SCT. These results suggest that disease eradication with imatinib monotherapy may be rare. Patients on imatinib followed by PCR should be made aware of the fact that a single negative test does not have the same significance as in patients after SCT.

scholarly journals Imatinib Suspension and Validation (ISAV) Study: Final Results at 79 Months

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 461-461 ◽  
Author(s):  
Silvia Mori ◽  
Philipp le Coutre ◽  
Elisabetta Abruzzese ◽  
Bruno Martino ◽  
Ester Pungolino ◽  
...  
Keyword(s):  
Relapse Rate ◽  
Median Duration ◽  
Research Funding ◽  
Well Being ◽  
Imatinib Treatment ◽  
Rt Pcr ◽  
Molecular Remission ◽  
The Impact ◽  
Risk Of Relapse

Abstract Introduction. It is known that imatinib can be safely discontinued in patients (pts) with Chronic Myeloid Leukemia (CML) with minimal residual disease. Here we report an update of the Imatinib Suspension And Validation (ISAV) study at 79 months (mts) from study initiation to provide long term follow up data. Aims. The ISAV study aims to validate the capability of digital PCR (dPCR) to predict relapses after imatinib discontinuation in CML pts with negative Q-RT-PCR and to evaluate relapse rate, time to recurrence, survival and the impact of imatinib treatment on Quality of Life (QoL). Methods. This study involves 15 sites, 10 in Italy and 1 in each of the following countries: Germany, Spain, The Netherlands, Canada and Israel. CML pts (chronic or accelerated phase) treated with imatinib for more than 2 years and in complete molecular remission (CMR) were eligible. Patients had to be in CMR for at least 18 mts, with a minimum of 3 Q-RT-PCR performed at their own sites. After discontinuation of imatinib therapy, Q-RT-PCR was performed monthly (mts 1-6), bimonthly for 36 mts and then every 6 mts for additional 2 years, to assess the maintenance of the molecular remission. The loss of molecular remission was defined as two consecutive positive Q-RT-PCR tests with at least one BCR-ABL/ABL value above 0.1%. Patients losing molecular remission resumed imatinib treatment at the same dosage used before interruption. dPCR was performed at screening and at 36 mts for those pts who were still in remission. Patients' QoL during imatinib discontinuation/resumption was evaluated through the EORTC QLQ-C30 questionnaire. Results. The ISAV study enrolled 112 pts with a median follow-up time of 60.0 mts [95% CI: 59.6-60.6] for pts who do not relapsed; 66.1% of them completed the study as per protocol. The 58.9% of pts were male and 37.4% were aged 65 or older; median duration of imatinib treatment was 103.2 mts with median duration of CMR of 25.6 mts before imatinib discontinuation. At 79 mts from imatinib discontinuation, 56 pts of the 107 eligible ones relapsed and resumed imatinib with a relapse rate of 52.3% [95%CI: 20.4-32.6]; 69.6% of them relapsed in the first 9 mts. Of the 52 not-relapsed pts, 40 (76.9%) regained Q-RT-PCR positivity without losing MMR. In this latter group 2 pts experienced late relapses, at 30.6 and 45.5 mts respectively. A loss of CCyR occurred in 13 pts (23.6%): 10/13 CCyR losses were recovered, the remaining 3 were not assessed for response. No case of CML progression or resistance to imatinib was observed. After the resumption of imatinib the median time to MMR/CMR was 1.8 [95% CI: 1.0-2.0] mts. No significant correlation between relapse and previous duration of imatinib treatment, use of interferon, time to CCyR, Sokal score or duration of CMR was identified, while an inverse relationship between pts age and risk of relapse was evident. dPCR results before imatinib discontinuation showed that 23.4% of pts were positive and 76.6% negative at the time of discontinuation, with a Negative Predictive Value ratio (dPCR/Q-RT-PCR) of 1.1 [95%CI: 0.99-1.22]. At 36 mts from imatinib discontinuation 80.4% [95%CI: 30.6-50.4] of the pts tested were positive in dPCR. Moreover, the results of dPCR performed at imatinib discontinuation and age together can predict the risk of relapse: pts with less than 45 years and with a positive dPCR had the highest risk of relapse (100%) as opposed to pts ≥45 years and with negative dPCR (36.1%). The analysis of QoL evidenced a statistically significant improvement in the general well-being and symptoms scales at 1 month after imatinib discontinuation, particularly with regard to nausea, diarrhea, fatigue and insomnia (p<0.05). An inverse and transient trend toward increased pain emerged at mts 1 and 3. Conclusions. At 79 mts from the beginning of the study, 52.3% of pts relapsed, with 24% loosing CCyR. The majority of relapses occurred in the first 9 mts after discontinuation however late relapses were also observed, up to the 4th year. Therefore, pts who discontinue imatinib should be monitored for a long period of time, especially if they show positive PCR values after discontinuation. All relapsed pts including those who lost CCyR regained their original response after restarting TKI. Age <45 years and dPCR positivity are significantly associated with relapses. QoL analysis showed a significant decrease in symptoms after imatinib discontinuation. Funded by Regione Lombardia. Disclosures le Coutre: Pfizer: Honoraria; Incyte: Honoraria; BMS: Honoraria; Novartis: Honoraria. Abruzzese:BMS: Consultancy; Novartis: Consultancy; Pfizer: Consultancy; Ariad: Consultancy. Assouline:Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Roche: Honoraria, Research Funding, Speakers Bureau; Pfizer: Honoraria, Research Funding, Speakers Bureau; BMS: Honoraria, Research Funding, Speakers Bureau; Novartis: Research Funding. Kim:Pfizer: Research Funding; BMS: Research Funding; Ilyang: Research Funding; Novartis: Research Funding. Gambacorti-Passerini:Pfizer: Consultancy, Honoraria, Research Funding; BMS: Consultancy.

Can Targeted Therapy for CML Still Learn From Transplant? Using Post-transplant RQ-PCR monitoring to Clarify the Importance of the Depth of Molecular Remission On the Risk of Subsequent Relapse.

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 2789-2789
Author(s):  
Frederick Pimm ◽  
Richard Szydlo ◽  
Letizia Foroni ◽  
Francesco Dazzi ◽  
Jaspal S Kaeda ◽  
...  
Keyword(s):  
Conflicts Of Interest ◽  
Residual Disease ◽  
Unrelated Donor ◽  
Molecular Response ◽  
Molecular Remission ◽  
Post Transplant ◽  
Long Term Treatment ◽  
Risk Of Relapse

Abstract Abstract 2789 The use of tyrosine kinase inhibitors (TKI) in the management of chronic myeloid leukemia (CML) has dramatically improved survival, with some 80% of patients achieving a deep and durable molecular remission (MR). The current focus for these patients is the ability to withdraw long-term treatment and a number of ‘stopping’ studies have been initiated worldwide. Many of these approaches are derived from the French STIM study which showed that 40% of patients who had been real-time quantitative PCR (RT-qPCR) negative for BCR-ABL1 for two years could cease treatment without experiencing disease relapse. However, the RT-qPCR assay used in this study was particularly stringent with a sensitivity of 10−5, compatible with a five log reduction in BCR-ABL1 transcripts (MR5), and it is not clear that the same level of success will result from studies using MR4 and MR4.5 as the indication for treatment cessation. Furthermore, because of the lack of accuracy in RT-qPCR assays when the number of BCR-ABL1 transcripts approach zero, some laboratories report as undetectable, transcript numbers <6 or even <11. In order to investigate the importance of the depth of molecular response on the risk of subsequent disease recurrence, we studied the long-term follow-up of, and RT-qPCR results from, patients who received allogeneic stem cell transplantation as treatment for CML at a time when minimal residual disease detection was performed by RT-qPCR using ABL1 as the control gene. We analysed data from 180 patients transplanted from January 1998 onwards who received an allo-SCT from an HLA-identical sibling or a matched unrelated donor and who had survived for at least 6 months post-transplant with a consistent sequence of 5 or more RT-qPCR results from the time of transplant to the end of follow-up. Patients were assessed on the depth of their MR; 9 categories of ‘complete’ MR were defined based on BCR-ABL1 transcript threshold for negativity (BCR-ABL1=0, BCR-ABL1>0 but <6, BCR-ABL1>5 but <11) and control transcript number (CTN) (CTN>104 but <104.5, CTN>104.5 but <105, but CTN>105). We ranked these categories, firstly by BCR-ABL1 transcript threshold, defining negativity at a lower threshold as a deeper response, and then sub-ranked by CTN, defining a larger CTN as a deeper response. Of the 180 patients, 49 (27%) did not achieve ‘complete’ MR by any definition and for the 131 (73%) patients who did reach some degree of ‘complete’ MR, the median time from transplant to best molecular response was 8.7 months (range, 1.0–103 months). We defined relapse as progression to an RT-qPCR level that triggered the use of donor lymphocyte infusions i.e. BCR-ABL1/ABL ratio exceeded 0.02% in 3 samples, or exceeded 0.05% in 2 samples, or showed rising levels with the last 2 samples higher than 0.02%, or worse (loss of cytogenetic or haematological remission). The 2 year relapse incidence post SCT was 94% in the group who did not achieve any degree of ‘complete’ MR, 94% in the group who achieved MR with BCR-ABL1<11 and >0, CTN>104 (n=32, 17.8%), 55% in the group BCR-ABL1=0, CTN>104 and <104.5 (n=19, 11%), 26% in the group BCR-ABL1=0, CTN>104.5 and <105 (n=47, 26%), and 6% in the group BCR-ABL1=0, CTN>105 (n=33, 18%) (p<0.0001). In multivariate analysis with adjustment for donor type, classifying the 33 patients who achieved BCR-ABL1=0, CTN>105 as the optimal molecular responders the relative risk of relapse was 90.1 in 49 patients who never achieved MR by any definition, (p<0.0001), 21.7 in the group BCR-ABL1<11 and >0, CTN>104 (n=32) (p<0.0001), 8.1 in the group BCR-ABL1=0, CTN>104 and<104.5 (n=19) (p<0.0001), and 2.11 in the group BCR-ABL1=0, CTN>104.5 and <105 (n=47) (p=0.002). In conclusion, fewer detectable BCR-ABL1 transcripts with larger numbers of control transcripts, i.e. a deeper response, predict a lower risk of relapse in post-transplant survivors and may have important implications for the ability to stop long-term TKI therapy. Disclosures: No relevant conflicts of interest to declare.

Qualitative and Quantitative Molecular Follow up of Minimal Residual Disease after Non Myeloablative Allografting for Multiple Myeloma.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 5128-5128 ◽  
Author(s):  
B. Bruno ◽  
M. Ladetto ◽  
M. Astolfi ◽  
L. Veneziano ◽  
L. Cimolin ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Minimal Residual Disease ◽  
Nested Pcr ◽  
Genomic Dna ◽  
Residual Disease ◽  
Patient Specific ◽  
Molecular Remission ◽  
Post Transplant ◽  
Minimal Residual

Abstract New allogeneic transplant protocols with non myeloablative conditioning regimens for treatment of multiple myeloma (MM) have been developed in the attempt to reduce the transplant related toxicity associated with myeloablation. Preliminary data have been encouraging with remarkable clinical response rates (Maloney et al, Blood 2003). However, data on the achievement of molecular remission, prerequisite for eventual cure, are still lacking. We implemented a tandem transplant approach consisting of high dose melphalan (200 mg/sqm) with autografting followed by non myeloablative low dose (2.0 Gy) total body irradiation and G-CSF mobilized PBSC infusion from HLA-identical siblings. The curative potential relies exclusively upon a potent graft versus myeloma (GVM) effect through donor T cells. At diagnosis, patient specific clonal markers were generated based upon the rearrangement of the immunoglobulin heavy chain (IgH) genes and used for nested polymerase chain reaction (PCR) detection of minimal residual disease after transplant. Molecular remission was defined as the disappearance of the molecular marker post transplant in both bone marrow and blood. The sensitivity of the nested PCR-based assay was 1 in 100000 cells. A patient specific marker was generated in 11/15 (73%) patients who entered the study. After a median follow up of 16 months (range 5–50), molecular follow up post transplant showed that 3/11 (27%) reached molecular remission at 1, 3 and 7 months post allografting, respectively. Of the remaining 8 patients, 3/8 and 5/8 reached clinical complete remission, defined as the disappearance of the monoclonal paraprotein by immunofixation, and partial remission, respectively. However, minimal residual disease by nested PCR could be detected at all timepoints. The molecular remissions have been durable at 7, 30, and 48 months post transplant, respectively. In 1 case the remission was achieved and sustained in the absence of graft versus host disease (GVHD) which is consistent with the notion that GVHD is not essential for GVM. Furthermore, in 4/11 patients real-time quantification of IgH rearrangements was performed on genomic DNA samples using tumor specific primers and consensus probes. All patients showed a considerable tumor burden reduction post autografting. Samples from two patients became negative by real time PCR at 3 months post allografting, but became PCR-negative by nested PCR at 3 and 7 months, respectively. This discrepancy is explained by the greater sensitivity of nested PCR and the larger amount of IgH copies which are expected in cDNA compared to genomic DNA. The remaining two patients only obtained a clinical partial response throughout the study period. This report indicates that the tandem auto-allo transplant approach can lead to molecular remission in MM. Prospective quantitative monitoring of disease response may be helpful to design individual additional immunotherapeutic manoeuvres, such as donor lymphocyte infusions, to enhance GVM. Longer follow up on a larger series of patients is needed to determine the frequency and durability of molecular remissions.

High Rate of BCR-ABL Transcript Undetectability Achieved by Treating with Imatinib Mesylate Very Late CML Patients in Stable CCR after IFN.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2938-2938
Author(s):  
Alimena Giuliana ◽  
Diverio Daniela ◽  
Breccia Massimo ◽  
Mancini Marco ◽  
Giuliani Nicoletta ◽  
...  
Keyword(s):  
Imatinib Mesylate ◽  
Internal Control ◽  
Molecular Level ◽  
Residual Disease ◽  
Standard Dose ◽  
Transcript Level ◽  
High Rate ◽  
Molecular Response ◽  
Rt Pcr

Abstract Background. Interferon alfa (IFN a) induces complete cytogenetic response (CCR) in small proportion of CML patients, with almost all of these patients still presenting BCR-ABL transcript at the molecular level, even after prolonged period of CCR. Imatinib mesylate induces CCR in a 60–80% of patients but a high percentage of these still show residual disease as detectable by RT-PCR, and can be at risk of disease relapse. Thus combining synergistic drugs might be required to eliminate the disease effectively. Methods. We treated with imatinib 16 CML patients (8 M and 8 F) who were in very late CP and in stable CCR achieved with IFNa, but still were persistently positive at molecular level by RT-PCR. According to Sokal’s score, 11 patients were low risk (LR) and 5 were intermediate risk (IR). Median time from diagnosis was 125 mos.(r. 52–202), median duration of stable CCR was 79 mo.s (r.9–148). Imatinib was administered at the standard dose of 400 mg/die, after stopping IFN for 1 week. The level of residual disease was then monitored on BM cells at planned intervals ( baseline, 3, 6, 12 mo.s), by assaying BCR-ABL transcript at nested PCR and real-time quantitative RT-PCR; ABL was used as an internal control and results expressed as a ratio of BCR-ABL/ABL transcripts on a log scale. Results. The median transcript levels, as measured at various time points, appeared to progressively and consistently decrease in all patients with respect to the baseline values. In particular, BCR-ABL transcript undectectability was observed in 6/15 patients with evaluable analyses at 3 mo.s, in 9/16 patients analysed at 6 mo.s, and in 5/8 with available results also at 12 mo.s; of these latter patients, 4 were already negative in previous analyses and one become negative at 12 mo.s. Thus, altogether, 10/16 (62.5%) patients with at least two examinations within 12 mo.s had at least one negative molecular result. Correlations between degree of transcript reduction and clinical/biological factors detected at diagnosis and during follow-up, evidenced not significant results for age, type of transcript (either b2a3 or b3a3), baseline transcript level pre-imatinib, duration of disease and of stable CCR prior imatinib, while a trend was apparent for a higher rate of LR vs IR score among the 10 patients reaching transcript undetectability (8/10 LR vs 2/10 IR) with respect to the 6 persistently positive subjects(3 LR vs 3 IR). In present cases, imatinib was well tolerated and no side effects required drug dose reduction or discontinuation. At a median follow-up of 13 months (8–16) from start imatinib, all 16 patients are alive in CCR with progressively improving molecular response, 10 of whom with persistent transcript undetectability. Conclusion. Albeit obtained in a series of very selected patients, present results represent a further evidence stressing on the efficacy of combining imatinib and IFN a; through different, possibly complementary, mode of action, these two drugs might mutually potentiate their effect while reducing the emergence of drug resistance. The additive toxicity caused by their concurrent use might be overcome by a sequential combination; this could also be applied to patients reaching stable CCR on imatinib but still maintaining detectable residual disease.

Molecular Long-Term Surveillance of CML Patients on Imatinib Therapy. Follow-Up of German Patients Treated within the IRIS Trial.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 1003-1003
Author(s):  
Martin C. Mueller ◽  
P. Paschka ◽  
T. Lahaye ◽  
Ch. Lorentz ◽  
N. Gattermann ◽  
...  
Keyword(s):  
Median Time ◽  
Nested Pcr ◽  
Residual Disease ◽  
Response To Therapy ◽  
Imatinib Therapy ◽  
Interferon Α ◽  
Tyrosine Kinase Domain ◽  
Molecular Response

Abstract High rates of complete cytogenetic response (CCR), the availability of sensitive methods to detect residual disease, and direct therapeutic consequences are leading motives to integrate regular molecular monitoring into the standards for the management of patients (pts) with chronic myeloid leukemia (CML). We sought to determine long-term dynamics of BCR-ABL mRNA expression levels in 132 CML pts (75 m, 57 f, median age 51, range 20–71 yrs) recruited into the IRIS study in 17 German centers. Pts were randomized to receive imatinib (n=69) or interferon α+Ara-C (IFN, n=63). Due to intolerance or lack of response 41 pts crossed over from IFN to imatinib. Response to therapy was sequentially monitored by conventional cytogenetics from bone marrow metaphases (n=806). BCR-ABL transcripts were determined in 1414 peripheral blood samples by quantitative real time RT-PCR (RQ-PCR) using the LightCycler technology. In case of low level (&lt;10 transcripts/2μl cDNA) or neg RQ-PCR, nested PCR was performed. Total ABL transcripts were quantified as internal controls. A single series of BCR-ABL plasmid dilutions served as standard for both BCR-ABL and ABL transcripts. In pts on 1st-line imatinib therapy median ratios BCR-ABL/ABL gradually decreased: 4.8% at mo 3, 0.88% at mo 6, 0.22% at mo 12, 0.17% at mo 18, 0.058% at mo 24, 0.066% at mo 30, and 0.023% at mo 36. After crossover to imatinib results were not significantly different: 15.5% at mo 3, 1.6% at mo 6, 0.28% at mo 12, 0.068% at mo 18, 0.045% at mo 24, and 0.041% at mo 30. After a median follow-up of 40 mo (1–47) 31/69 pts (45%) on 1st-line imatinib were still RQ-PCR pos, 20 pts (29%) were RQ-PCR neg and nested PCR pos, and in 4 pts (5.8%) BCR-ABL became undetectable by RQ- and nested PCR. After a median time of 25 mo (3–43) on 2nd-line imatinib therapy 19/41 pts (46%) were RQ-PCR pos, 9 pts (22%) were RQ-PCR neg and nested PCR pos, and in 5 pts (12%) BCR-ABL was undetectable by RQ- and nested PCR. Considering adequate RNA quality BCR-ABL became repeatedly undetectable in 4 pts after 18–33 mo of 1st-line imatinib therapy and in 5 pts 9–33 mo after crossover from IFN to imatinib. In one patient, BCR-ABL remained undetectable after a treatment free interval of 4 weeks. After achieving CCR, 5 pts (7.2%) on 1st-line and 2 pts (4.9%) on 2nd-line imatinib therapy experienced cytogenetic relapse after a median time of 10 mo (4–21). In none of these pts mutations of the tyrosine kinase domain of BCR-ABL were detected. BCR-ABL/ABL ratios after 12 mo of imatinib therapy were significantly lower in pts in continuous CCR vs pts with subsequent relapse (0.18 vs 0.60%, respectively, p=0.04). None of the relapsing patients had achieved a ratio BCR-ABL/ABL &lt;0.12% after 12 mo, which represents a 3-log reduction from baseline. During total follow-up ratios BCR-ABL/ABL &lt;0.12% have been achieved in 51 pts (74%) on 1st-line and in 21 pts (51%) on 2nd-line imatinib therapy. We conclude that (i) treatment with imatinib in newly diagnosed CML pts is associated with a rapid and steady decrease of BCR-ABL transcript levels, (ii) a short trial of IFN does not jeopardize molecular response to subsequent imatinib therapy, (iii) an increasing minority of pts achieve complete molecular remission, and (iv) ratios of BCR-ABL/ABL &lt;0.12% after 12 mo of therapy predict for long-term response.

Use of 5-Fold Staining for Multiparameter Flow Cytometry-Based Quantification of Minimal Residual Disease in Acute Myeloid Leukemia Results in Improved Sensitivity.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2025-2025
Author(s):  
Wolfgang Kern ◽  
Daniela Voskova ◽  
Claudia Schoch ◽  
Wolfgang Hiddemann ◽  
Susanne Schnittger ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Bone Marrow Cells ◽  
Residual Disease ◽  
Multiparameter Flow Cytometry ◽  
Normal Bone Marrow ◽  
Prognostic Information ◽  
Normal Bone ◽  
Triple Staining ◽  
Marrow Cells

Abstract The quantification of minimal residual disease (MRD) by multiparameter flow cytometry (MFC) using triple staining has been shown to yield prognostic information independent of other parameters in patients with acute myeloid leukemia (AML). Due to the immunophenotypic heterogeneity of AML the application of 5-fold staining may result in a better characterization of the leukemia-associated aberrant immunophenotype (LAIP) and thus in an improved sensitivity of the method as compared to triple staining. We analyzed bone marrow samples from 114 patients with newly diagnosed and untreated AML by MFC using a comprehensive antibody panel with 5-fold combinations. Sensitivity was estimated by quantification of LAIP-positive cells for each LAIP in 18 normal bone marrow samples. In each patient at least one LAIP was identified (total, 203 LAIPs). The LAIPs were present on a median of 15.88% of the bone marrow cells at diagnosis (range, 2.11% to 79.64%). The median number of normal bone marrow cells displaying the LAIPs ranged from 0.001% to 0.065% (median, 0.010%). As a result, the logarithmic difference (LD) in LAIP-positive cells between leukemic and normal bone marrow amounted to a median of 3.33 (range, 1.96 to 4.88). Similarly, if only the most sensitive LAIP was considered for each patient the median frequencies of LAIP-positive cells were 14.07% (range, 2.11% to 77.57%) in leukemic bone marrow and 0.010% (range, 0.001% to 0.065%) in normal bone marrow. Importantly, however, in this setting the resulting LD amounted to a median of 3.45 (range, 1.96 to 4.88). In order to estimate the impact of applying 5-fold staining on the sensitivity the information of each of the applied colors was skipped once while the results of the other four colors, respectively, were used. Skipping one color resulted in an increase of LAIP-positive normal bone marrow cells (median, 0.050%; range, 0.001% to 3.6%) while the percentages of LAIP-positive leukemic cells changed only marginally (median, 22.65%; range, 2.25% to 90.06%). The gain in LD by applying 5-fold staining in comparison to 4-fold staining amounted to a median of 0.58 (maximum gain, 3.14). In 32 patients a total of 120 follow-up samples have been analyzed appyling the combination of antibodies that allowed the best LAIP definition. The LD from diagnosis to follow-up amounted to a median of 2.82 (range, 0.77 to 4.82). Clinical follow-up data is available in 26 of these 32 patients. MRD assessment after completion of consolidation therapy has been performed in 15 patients. The median LD between diagnosis and follow-up assessment is 2.84 (range, 1.07 to 4.33). Separating patients according to this median LD identified a group of patients with no relapses yet (LD >2.84) while patients with an LD <2.84 had an event-free survival of only 50% at one year (p=0.075). These data confirm that flow cytometrically-based assessment of MRD is feasible in AML and results in prognostic information. It is suggested that the application of 5-fold staining significantly improves the sensitivity and thereby the overall accuracy of the method.

Molecular Remission Can Be Attained in Relapsed or Refractory Chronic Lymphocytic Leukemia (CLL) and Follicular Lymphomas after Reduced Intensity Conditioning (RIC) and Allogeneic Stem Cell Transplantation (ALLO-SCT).

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2317-2317
Author(s):  
Lucia Farina ◽  
Matteo Carrabba ◽  
Anna Dodero ◽  
Elena Rizzo ◽  
Elisabetta Zorzan ◽  
...  
Keyword(s):  
Nested Pcr ◽  
Chronic Gvhd ◽  
Conditioning Regimen ◽  
Lymphocytic Leukemia ◽  
Clinical Relapse ◽  
Molecular Remission ◽  
Continuous Increase ◽  
Taqman Pcr ◽  
Follicular Lymphomas

Abstract RIC followed by allo-SCT is an effective salvage treatment for some relapsed hematologic malignancies due to the postulated graft versus tumour (GVT) effect. In order to evaluate the quality of the clinical response, we have investigated the molecular status of patients receiving allo-SCT for relapsed disease. Forty-four patients (19 chronic lymphocytic leukemias (CLL), 21 follicular lymphomas (FCL) and 4 small lymphocytic lymphomas (SLL)) were enrolled in a prospective phase II study. The median age was 54 years (range: 32–69 years). The median number of previous chemotherapy regimens was 2 (range: 1–5) and 23% of patients had already failed an auto-SCT. Before transplant 34% of patients were chemorefractory and 34% of the chemosensitive patients were in complete remission (CR). The conditioning regimen consisted of thiotepa 10mg/kg, fludarabine 60mg/ms and cyclophosphamide 60mg/kg; short course of methotrexate and cyclosporin were used as GVHD prophylaxis. Minimal residual disease (MRD) was monitored by nested PCR for IgH or Bcl-2 genes; in PCR-positive patients a TaqMan based quantitative monitoring was also employed. All patients engrafted. On day +30 after transplant 39% of patients achieved CR. Acute GVHD (aGVHD) was observed in 57% of patients and 52% of 42 evaluable patients developed chronic GVHD; no difference in the incidence of GVHD between FCL and CLL/SLL was observed. In 30 of 44 patients (68%) a PCR marker for MRD monitoring was found. Twenty-five patients (10 CLL, 2 SLL, 13 FCL) of 37 patients in CR after allo-SCT were monitored by nested PCR and 4 PCR-positive patients were monitored by TaqMan PCR. At a median molecular follow up of 15 months (range: 3–62) 15 of 25 patients (60%) were alive and in molecular remission; one CLL patient died of TRM in molecular remission (MR); five of these patients were chemorefractory. Nine patients (3 FCL, 5 CLL, 1 SLL) never achieved PCR negativity and 3 of them relapsed (2 CLL; 1 SLL) after a median time of 270 days. In one of these patients the TaqMan PCR system could detect a continuous increase of tumour genomes in the marrow prior to the clinical relapse. The SLL patient achieved MR after chemotherapy and DLI, developing limited cGVHD; the other two patients never developed GVHD, even after DLI. Eighty percent of PCR-negative patients developed GVHD and it preceded or was concomitant with the achievement of MR. The better molecular outcome of FCL seems to be due to a longer follow up (19 months vs 12 months) if compared to CLL/SLL, in which a slow clearance of MRD has been observed. In conclusion, MR can be achieved in relapsed and chemorefractory patients affected by indolent lymphoproliferative disorders; quantitative PCR monitoring can be used to modulate post-transplant immunotherapy; a longer follow up is warranted to evaluate if the GVT effect can sustain MR in the long-term.

Minimal Residual Disease (MRD) Analysis in Acute Myeloid Leukemia (AML) with Favorable Cytogenetics [t(8;21) and inv(16)] by Real Time PCR(RT-PCR) and Flow Cytometry (FC).

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2989-2989
Author(s):  
Granada Perea ◽  
Adriana Lasa ◽  
Anna Aventin ◽  
Alicia Domingo ◽  
Neus Villamor ◽  
...  
Keyword(s):  
Myeloid Leukemia ◽  
Residual Disease ◽  
Fusion Transcript ◽  
Good Prognosis ◽  
Rt Pcr ◽  
Free Survival ◽  
End Of Treatment ◽  
The Mean ◽  
Minimal Residual

Abstract Objectives: To analyze MRD in 65 patients (pts) with good prognosis AML: 30 t(8;21) and 35 inv(16), using both FC and RT-PCR, and to investigate the prognostic value of MRD in the pts outcome. Methods: MRD was monitored in CR pts (n=55) by FC in 101 follow-up samples obtained after various cycles of treatment, as follows: 40 post-induction (ind), 30 post-intensification (int) and 31 at the end of treatment (ttm), and by RT-PCR in 76 samples: 31, 23 and 22, respectively. In 35 pts the two techniques were applied at the same time of the ttm. MRD by FC was assessed using fixed combinations of three monoclonal antibodies. AML1/ETO and CBFb/MYH11 were analyzed following the BIOMED protocol. Results: Twenty-seven percent (n=15) of CR pts relapsed: 6 with t(8;21) and 9 with inv(16). The mean MRD by FC was 1.1% after ind, 0.2% after int and 0.1% at the end of ttm. At the end of ttm, the MRD detected by FC in relapsed and not relapsed pts were significativaly different: 0.3% vs 0.08% (p=0.002). By RT-PCR, the mean of fusion transcript copies/ablx104 differed between relapsed and nonrelapsed pts: 2385 vs 122 (p=0.001) after ind, 56 vs 7.6 after int (p=0.0001) and 75 vs 3.3 (p=0.0001) at the end of ttm. Relapses were more commonly observed in those pts with FC MRD level >0.1% at the end of ttm than in pts with ≤0.1%: 50% vs 12% (p=ns); likewise, using RT-PCR, a cutoff level of >10 copies at the end of ttm correlated with high risk of relapse: 80% of pts with RT-PCR >10 relapsed compared to 12% of pts with levels <10 (p=0.009). The overall survival (OS) probability was 86% for pts with CF MRD ≤0.1 at the end of ttm and 0% for pts with MRD >0.1 (p=0.1) and the leukemia free survival (LFS) was 78% and 44%, respectively (p=0.05). For pts with RT-PCR ≤10 at the end of ttm, the OS was 100% and for pts with RT-PCR >10 it was 30% (p=0.007) and the LFS was 87% and 20%, respectively (p=0.001). MRD was identified after ind in 55% of relapsed pts and at the end of ttm in 83% of relapsed pts. Only 1 pt (1/13) with FC MRD <0.1 and RT-PCR <10 at the end of ttm relapsed. For patients in complete remission, the mean copy level of chimeric transcript was higher for pts with t(8;21) than for those with inv(16): 30.2 vs 17.4 (p=0.0001). Comments: In tandem analysis of MRD by FC and RT-PCR could improve MRD detection in AML pts.

Determination of Minimal Residual Disease in Patients with Acute Myeloid Leukemia by Multiparameter Flow Cytometry: Prognostic Impact at Different Checkpoints.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 400-400 ◽  
Author(s):  
Wolfgang Kern ◽  
Daniela Voskova ◽  
Claudia Schoch ◽  
Wolfgang Hiddemann ◽  
Susanne Schnittger ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Myeloid Leukemia ◽  
Bone Marrow Cells ◽  
Residual Disease ◽  
Multiparameter Flow Cytometry ◽  
Prognostic Impact ◽  
Minimal Residual ◽  
Marrow Cells

Abstract Guiding antileukemic treatment in patients with acute myeloid leukemia (AML) is increasingly based on levels of minimal residual disease (MRD) which can be quantified with high sensitivity by multiparameter flow cytometry (MFC). The optimum checkpoint for determination of MRD during the course of therapy, however, has not yet been determined. We applied MFC using a comprehensive panel of antibodies to identify leukemia-associated aberrant immunophenotypes (LAIPs) at diagnosis and to quantify MRD by individually selected antibody combinations. The prognostic impact of MRD levels was assessed in comparison to cytogenetics and age. Patients received double induction, consolidation, and maintenance therapies and underwent allogeneic stem cell transplantation if they were younger than 60 years and had a matched related donor. In 286 patients with newly diagnosed and untreated AML MFC-based assessment for the presence of LAIP has been performed. The median percentage of LAIP-positive bone marrow cells at diagnosis was 16.04% (range, 2.54%–76.14%). All individual LAIPs were applied to 26 normal bone marrow samples to estimate sensitivity based on the median percentages of LAIP-positive normal bone marrow cells which ranged from 0.00% to 1.01% (median, 0.02%). A total of 550 follow-up samples has been analyzed in these patients at different checkpoints (CP1, up to day 21 after start of therapy, n=85; CP2, day 22–60, n=122; CP3, day 61–120, n=158; CP4, day 121–365, n=137; CP5, after day 365, n=48). In order to adjust for differences in the percentages of LAIP-positive bone marrow cells at diagnosis the logarithmic difference (LD) between diagnosis and follow-up was calculated for each follow-up sample. The median LDs at the respective checkpoints were: CP1, 2.02; CP2, 2.29; CP3, 2.39; CP4, 2.53; and CP5, 2.81. Separation of patients according to the respective median LDs resulted in differences in event-free survival (EFS; CP1: 21.1 vs. 9.1 months, p=0.0711; CP2: 14.2 vs. 9.3 months, p=0.0095; CP3: 30.9 vs. 13.5 months, p=0.0055; CP4: median not reached vs. 14.1 months, p<0.0001; CP5: median not reached vs. 22.5 months, p=0.0001) and overall survival (OS; CP3: median not reached vs. 21.6 months, p=0.0332; CP4: 90% vs. 53% at 2 years, p=0.0058). Cox analysis using the LDs at the different checkpoints as continuous variables confirmed the prognostic impact on EFS (CP2, p=0.002; CP3, p=0.0003; CP4, p<0.0001; CP5, p<0.0001) and revealed an impact also on OS (CP3, p=0.003; CP4, p=0.001; CP5, p=0.029). Cox regression analysis taking into consideration cytogenetics and age as covariates proved the independent prognostic impact of LD at checkpoints 2 to 5 on both EFS and OS with the exception of LD at checkpoint 2 and OS. In fact, LD at checkpoint 5 was the only parameter independently related to EFS and OS. These data suggest that quantification of MRD by MFC in AML results in powerful and independent prognostic parameters. In particular during the first year of treatment MRD levels provide important prognostic information. Clincal trials should use MRD-based stratification in order to assess the efficacy of early treatment intensification in high-risk AML patients.

Molecular Response of CML Patients to INFα Based Treatment or to STI Based Therapy.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 4872-4872
Author(s):  
Debora Luzi ◽  
Rosanna Capozzi ◽  
Annamaria Rauco ◽  
Roberta Pace ◽  
Emilio Donti ◽  
...  
Keyword(s):  
Myeloid Leukemia ◽  
Response Duration ◽  
Cytogenetic Response ◽  
The Other ◽  
Entire Group ◽  
Molecular Response ◽  
Rt Pcr ◽  
Molecular Remission ◽  
First Time

Abstract Introduction: Continous improving results have been obtained during last two decades in the control of Ph’positive chronic myeloid leukemia(CML). However the final goal of molecular remission remains difficult to obtain even in the STI age. Aims : Evaluation of the rate of molecular response to IFNα,IFNα based treatment,to STI or to STI-INFα combination was analized in 100 consecutive Ph+ CML patients observed in a single Institution over a period of 20 years. Patients, Methods and Results All patients were treated at the time of diagnosis (87) or late (13) during the course of their disease. Distribution according to treatment was: INFα,63pts (late or early:13,50);INFα-ARA-C combination,20pts;STI,14 pts;STI-INFα association, 3 pts. Two pts, both initially assigned to INFα-ARA-C combination, were crossed-over to STI, one because relapsing off-therapy after a long lasting continous (25 mths) molecular remission and the other in cytogenetic response because intolerant to the initial treatment. In addition, other 3 pts patients, with persistent complete cytogenetic, but not molecular remission to INFα or INFα-ARA-C combination were subsequentially trated with the STI-IFNα association. At present,99/100 pts are evaluable. The median times of follow-up for the entire group and form the different treatment subgroups are: late IFNα 154 months(42–263); early IFNα, 71 months(1–197); IFNα-ARA-C, 61 months(5–203); STI- IFNα,78 mths(11–47), STI,31 mths(3–41). A complete kariotypic remission(CKR) was observed in 15/63 IFNα treated pts, in 10/20 IFNα-ARA-C pts group, in 10/13 cases of STI group and in 3 /3 pts who received STI-IFNα. A molecular response(RT-nested PCR, JQ Guo, Leukemia: 2002,15,2447–53) was observed in 4/15,2/10,5/10 and in 2/3 CKR pts initially trated with the different modalities listed above. Response was confirmed from 2 to 7 consecutive or not consecutive times in the 2/4 cases responsive to INFα, in the 2 cases responsive to INFα-ARA-C combination,4/5cases responsive to STI and in 2/3 cases responsive to STI-IFNα association. The 2nd and the 3rd molecular remission to STI were obtained in the patient molecularly and cytogenetically relapsed off-therapy and, for the first time from the diagnosis, in the other patient in CKR to IFNα-ARA-C combination and crossed to STI treatment. Furthermore, all 3 cases, in CKR, but not molecular response to other treatments at the time of cross-over to STI-IFNα combination, achieved a persistent (in 2 to 3 tests over a period ranging from 6+ to 12+ mths) molecular remission. The first interval between the start of the treatment and the first molecular response varied from 12 to 52, from 3 to 22, from 11 to 24, from 5 to 11 mths in the groups initially treated with IFNα, IFNα-ARA-C, STI or STI-IFNα respectively. The 2 pts, crossed-over to STI alone, both, obtained a response after 29 mths of therapy. In addition in the 3 pts crossed-over to STI-IFNα therapy, the molecular response was obtained after 14,23 and 25 mths from the start of last treatment. Conclusion It is not possible to achieve any conclusion regarding the treatment effect on molecular response duration because of the different length of follow-up of various groups of patients. However in responsive patients to IFN alone or combined to ARA-C or STI, consecutive negative RT-PCR tests were observed more frequently than in patients receving STI alone.

Sign in / Sign up

Export Citation Format

Share Document