ALBERT

Description: Abstract 2298 We previously reported that imatinib treatment could be safely discontinued in chronic myeloid leukemia (CML) patients who had achieved a sustained complete molecular remission (CMR) lasting for at least two years in succession. However, in the multicenter French STIM study, 60 % of patients loose CMR mainly within the first 6 months after discontinuation and the probability to remain in CMR 2 years after imatinib discontinuation is around 40 %. The molecular relapse originates in persisting leukemic cells that are undetectable by current RQ-PCR techniques. The sensitivity of these routinely used techniques is below a detection threshold corresponding to a 5-log reduction in the leukemic burden. We thus hypothesized that improving the sensitivity of leukemic cell detection would be of valuable help to better select those patients who can possibly be cured after imatinib treatment. In the present work, we have increased the sensitivity of the current RQ-PCR, based on repeated PCR and on an increased number of normal ABL copies that were analyzed, in order to augment the probability to amplify BCR-ABL. For each sample, 5 microg of RNA were used to synthesize 5 cDNA and for each cDNA, 2 PCR for BCR-ABL and one for ABL were carried out, i.e 10 BCR-ABL PCR points were performed per sample. For each microg of RNA the number of ABL copies was over 20 000, so the number of ABL copies analyzed was over 100 000. The detection threshold of BCR-ABL was calculated and considered at 40 cycles of PCR (theshold Ct). The control plasmid (pME-2) (Kindly given by Martin Müller and Andreas Hochhaus from European LeukemiaNet) used as standard curve dilution was also included to test the sensitivity and for instance 4 copies were detected in 19/20 cases, 2 copies 7/10 cases,1 copy in 2/10, 0.4 copy in 1/10. Thirty one samples from healthy donors or non CML patients (BCR-ABL negative) served as controls. Among the 310 (10 × 21) PCR for BCR-ABL we found only one positive well. 65 patients enrolled in the STIM study were analyzed at the time of imatinib discontinuation, using this new RQPCR technique. In the STIM study relapse was defined as the positivity of Bcr-Abl transcripts using classical QRT-PCR confirmed by a second analysis point indicating the increase in relation to the first analysis point performed on 2 successive assessments. Among 650 PCR for BCR-ABL, 46 wells were found positive. 9 patients were found at least 3 /10 positive PCR and among them 6 relapsed. 22 patients were found only at least 1 /10 positive and among them 15 relapsed. In 43 patients, BCR-ABL was never detected using the more sensitive RQ-PCR technique and 22 of them relapsed. To conclude, the use of a new RQPCR technique with a sensitivity of detection of BCR-ABL close to 6-log does not allow the prediction of molecular relapse following imatinib discontinuation in patients in CMR. Our results are in agreement with what was previously reported using PCR on genomic DNA. The persistence of leukemic cells in CMR patients does not automatically lead to CML relapse. Disclosures: Belanger: Novartis Pharma: Employment.

Description: Abstract 1676 Background: Pts treated with nilotinib in the ENESTnd phase 3 trial achieved higher and faster rates of major molecular response (MMR, ≤ 0.1% BCR-ABLIS), deeper molecular responses (MR4, ≤ 0.01%IS and MR4.5, ≤ 0.0032%IS), significantly lower rates of progression to accelerated phase/blast crisis (AP/BC), and fewer CML-related deaths compared with imatinib by 1, 2, and 3 y. Here, we report data with a minimum follow-up of 3 y; efficacy and safety data based on longer follow-up of 4 y will be presented to further assess the impact of nilotinib vs imatinib in pts with newly diagnosed Ph+ CML-CP. Methods: Adult pts (N = 846) with newly-diagnosed Ph+ CML-CP were randomized to nilotinib 300 mg twice daily (BID; n = 282), nilotinib 400 mg BID (n = 281), or imatinib 400 mg once daily (QD; n = 283). MMR, MR4, MR4.5, time to progression to AP/BC, progression-free survival (PFS), and overall survival (OS) were evaluated. Results: Significantly higher rates of MMR, MR4, and MR4.5 by 3 y were achieved in nilotinib- vs imatinib-treated pts (Table). Nilotinib led to the achievement of higher rates of molecular responses regardless of Sokal risk group or age. The difference in the rates of both MR4 and MR4.5 continued to be significantly higher for nilotinib, with the difference in favor of nilotinib increasing from 1 to 3 y (MR4: 9%-14% difference by 1 y, 18%-24% difference by 3 y; MR4.5: 6%-10% difference by 1 y, 13%-17% difference by 3 y). Among patients who achieved MMR, more pts achieved MR4 or MR4.5 on nilotinib 300 mg BID (68%) and nilotinib 400 mg BID (62%) compared with imatinib (49%). No pt in any arm progressed after achieving MR4.5. Significantly fewer pts progressed to AP/BC on nilotinib vs imatinib (Table). No new progressions occurred on core treatment between the 2-y and 3-y analyses. When events occurring after treatment discontinuation were included, the rates of progression to AP/BC were also significantly lower with nilotinib vs imatinib (Table). Nearly twice as many pts had emergent mutations on imatinib (n = 21) vs either nilotinib arm (n = 11 in each arm), with 5 pts overall developing mutations between 2 and 3 y. OS remained similar in all groups at 3 y, but fewer CML-related deaths occurred in both the nilotinib 300 mg BID (n = 5) and 400 mg BID (n = 4) arms vs imatinib (n = 14). Both drugs were well tolerated. Few new adverse events (AEs) and laboratory abnormalities were observed between 2 and 3 y. Rates of discontinuation due to AEs were 10%, 14%, and 11% in the nilotinib 300 mg BID, nilotinib 400 mg BID, and imatinib arms, respectively. Conclusions: Nilotinib continues to demonstrate superiority vs imatinib, yielding faster and deeper molecular responses and a significantly decreased risk of progression. Results of ENESTnd support the use of nilotinib as a standard of care option in newly diagnosed adult pts with Ph+ CML-CP and should be considered to replace imatinib as the standard-of-care frontline therapy for patients with Ph+ CML-CP. Disclosures: Kantarjian: Novartis: Consultancy, Research Funding; BMS: Research Funding; Pfizer: Research Funding. Kim:Novartis: Consultancy, Honoraria, Research Funding, Speakers Bureau; BMS: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Pfizer: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; ARIAD: Research Funding; II-Yang: Research Funding. Clark:Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; BMS: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Honoraria, Membership on an entity's Board of Directors or advisory committees. Reiffers:BMS: Expense reimbursement for travel expenses Other; Novartis: Expense reimbursement for travel expenses, Expense reimbursement for travel expenses Other. Nicolini:Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; BMS: Honoraria, Research Funding, Speakers Bureau; Ariad: Research Funding, Speakers Bureau; Pfizer: Consultancy, Honoraria. Hughes:Novartis: Consultancy, Honoraria, Research Funding; BMS: Consultancy, Honoraria, Research Funding; Ariad: Consultancy, Honoraria; CSL: Research Funding. Hochhaus:BMS: Consultancy, Honoraria, Research Funding; Pfizer: Consultancy, Honoraria, Research Funding; Novartis: Consultancy, Honoraria, Research Funding; Ariad: Consultancy, Honoraria, Research Funding. Kemp:Novartis Pharmaceuticals Corp: Employment. Fan:Novartis Pharmaceuticals Corp: Employment. Waltzman:Novartis Pharmaceuticals Corp: Employment, Equity Ownership. Saglio:Novartis: Consultancy, Speakers Bureau; BMS: Consultancy, Speakers Bureau; Pfizer: Consultancy. Larson:Novartis: Consultancy, Honoraria, Research Funding; BMS: Consultancy, Research Funding; Pfizer: Consultancy; Ariad: Consultancy, Research Funding.

Description: Despite the excellent efficacy of imatinib in chronic myeloid leukemia (CML), the response in patients is heterogeneous, which may in part be caused by pharmacogenetic variability. Imatinib has been reported to be a substrate of the P-glycoprotein pump. In the current study, we focused on the ABCB1 (MDR1) genotype. We analyzed the 3 most relevant single nucleotide polymorphisms of MDR1 in 90 CML patients treated with imatinib. Among the patients homozygous for allele 1236T, 85% achieved a major molecular response versus 47.7% for the other genotypes (P = .003). For the 2677G〉T/A polymorphism, the presence of G allele was associated with worse response (77.8%, TT/TA; vs 47.1%, GG/GA/GT; P = .018). Patients with 1236TT genotype had higher imatinib concentrations. One of the haplotypes (1236C-2677G-3435C) was statistically linked to less frequent major molecular response (70% vs 44.6%; P = .021). Hence, we demonstrated the usefulness of these single nucleotide polymorphisms in the identification of CML who may or may not respond optimally to imatinib.

Description: Using high-performance liquid chromatography–tandem mass spectrometry, we assessed trough imatinib plasma levels in 68 patients with chronic myeloid leukemia (CML) who responded or not to standard-dose imatinib, after at least 12 months' treatment. Mean trough imatinib plasma levels were significantly higher in the group with complete cytogenetic response (56 patients) than in the group without (12 patients; P = .03) and higher in the group with major molecular response (MMR) than in the group without (34 patients [1452 ± 649 ng/mL] versus 34 patients [869 ± 427 ng/mL]; P 〈 .001). Regarding trough imatinib plasma levels and their discrimination potential for MMR, the area under receiver operating characteristic curve was 0.775, with best sensitivity (77%) and specificity (71%) at a plasma threshold of 1002 ng/mL. Therefore, monitoring of imatinib plasma levels could be very useful for the management of patients with CML or should at least be checked in the case of treatment failure or suboptimal response.

Description: Abstract LBA-1 Background: Nilotinib is a highly potent and the most selective inhibitor of BCR-ABL, the only proven molecular target for CML therapy. ENESTnd (Evaluating Nilotinib Efficacy and Safety in Clinical Trials-Newly Diagnosed Patients) is a phase 3, randomized, open-label, multicenter study comparing the efficacy and safety of 300 or 400 mg bid nilotinib with 400 mg qd imatinib in patients (pts) with newly diagnosed Ph+ CML in chronic phase (CML-CP). Methods: 846 pts with newly diagnosed Ph+ CML-CP, diagnosed within 6 mos, and stratified by Sokal risk score, were randomized 1:1:1 to nilotinib 300 mg bid (n=282), nilotinib 400 mg bid (n=281), and imatinib 400 mg qd (n=283) arms. The primary endpoint was rate of major molecular response (MMR) at 12 months (mos). All pts had a minimum of 12 mos of treatment or discontinued early; median follow-up was 14 mos. MMR was defined as a value of ≤ 0.1% of BCR-ABL/ABL ratio on the International Scale. Molecular response was assessed by RQ-PCR at baseline, monthly for 3 mos and every 3 mos thereafter. Samples were analyzed at a central PCR laboratory. The major secondary endpoint was rate of complete cytogenetic response (CCyR) by 12 mos based on bone marrow cytogenetics. Results: Baseline demographics, disease characteristics, and Sokal scores were well balanced among the 3 arms; pts with high-risk Sokal scores were 28% in all arms. Median dose intensities of nilotinib delivered were 592 mg/day for 300 mg bid and 779 mg/day for 400 mg bid; imatinib dose intensity was 400 mg/day. Overall, 84%, 82%, and 79% of pts remained on the study for 300 mg bid nilotinib, 400 mg bid nilotinib, and 400 mg qd imatinib, respectively. Rates of MMR at 12 mos (Table) were superior for nilotinib 300 mg bid compared with imatinib 400 mg qd (44% vs. 22%,P 〈 .0001) and also for nilotinib 400 mg bid compared with imatinib 400 mg qd (43% vs. 22%,P 〈 .0001). Median time to MMR among pts who achieved MMR was faster for nilotinib 300 mg bid (5.7 mos) and nilotinib 400 mg bid (5.8 mos) compared with imatinib 400 mg qd (8.3 mos). Rates of CCyR by 12 mos were significantly higher for both nilotinib at either 300 mg bid compared with imatinib 400 mg qd (80% vs. 65%,P 〈 .0001) and for nilotinib 400 mg bid compared with imatinib 400 mg qd (78% vs. 65%,P = .0005). Overall, progression to advanced disease was lower for nilotinib 300 mg bid (2 pts) and nilotinib 400 mg bid (1 pt) compared with imatinib 400 mg qd (11 pts). Overall, both drugs were well-tolerated. Rates of discontinuation due to adverse events or laboratory abnormalities were 7% for nilotinib 300 mg bid, 11% for nilotinib 400 mg bid, and 9% for imatinib 400 mg qd. Pts were monitored for QT prolongation and LVEF. No patients in any treatment arm showed a QTcF interval 〉 500 msec. There was no decrease from baseline in mean LVEF anytime during treatment in any arm. The study is ongoing. Conclusions: Nilotinib at both 300 mg bid and 400 mg bid induced significantly higher and faster rates of MMR and CCyR compared with imatinib 400 mg qd, the current standard of care in pts with newly diagnosed CML. Nilotinib was effective across all Sokal scores. After only one year of treatment, both nilotinib arms resulted in a meaningful clinical benefit compared to imatinib, with reduction of transformation to AP/BC. Nilotinib exhibited a favorable safety and tolerability profile. The superior efficacy and favorable tolerability profile of nilotinib compared with imatinib suggests that nilotinib may become the standard of care in newly diagnosed CML. Disclosures: Saglio: Novartis: Consultancy, Speakers Bureau; BMS: Consultancy, Speakers Bureau. Off Label Use: Nilotinib is not currently approved for first-line treatment of CML. The presentation will report the results from a randomized study of imatinib versus nilotinib in patients with newly diagnosed Ph+ CML-CP. Kim:Novartis: Honoraria, Membership on an entity’s Board of Directors or advisory committees, Research Funding; BMS: Honoraria, Membership on an entity’s Board of Directors or advisory committees, Research Funding; Wyeth: Research Funding. le Coutre:Novartis: Honoraria, Research Funding; BMS: Honoraria. Reiffers:Novartis: Research Funding. Pasquini:Novartis: Consultancy, Membership on an entity’s Board of Directors or advisory committees; BMS: Membership on an entity’s Board of Directors or advisory committees; Schering: Membership on an entity’s Board of Directors or advisory committees. Clark:Novartis: Honoraria, Research Funding, Speakers Bureau. Hughes:Novartis: Honoraria, Research Funding; BMS: Honoraria, Research Funding. Hochhaus:Novartis: Research Funding. Gallagher:Novartis: Employment, Equity Ownership. Hoenekopp:Novartis: Employment. Dong:Novartis: Employment, Equity Ownership. Haque:Novartis: Employment. Larson:Novartis:

Description: Stem cell factor (SCF) has been shown to synergize with filgrastim to mobilize CD34+ cells into the peripheral blood. To determine if addition of SCF to chemotherapy and filgrastim reduces the number of leukaphereses required to achieve a target yield of 5 × 106 CD34+ cells/kg, 102 patients with multiple myeloma were randomized to receive mobilization chemotherapy with cyclophosphamide (4 g/m2) and either SCF (20 μg/kg/d) combined with filgrastim (5 μg/kg/d) or filgrastim alone (5 μg/kg/d), administered daily until leukaphereses were completed. After collection, patients were treated with myeloablative therapy supported by autologous peripheral blood progenitor cell (PBPC) infusion and filgrastim (5 μg/kg/d). There was a significant difference between the treatment groups in the number of leukaphereses required to collect 5 × 106 CD34+ cells/kg (median of 1 v 2 for SCF + filgrastim and filgrastim alone, respectively, P = .008). Patients receiving the combination of SCF plus filgrastim had a 3-fold greater chance of reaching 5 × 106 CD34+ cells/kg in a single leukapheresis compared with patients mobilized with filgrastim alone. The median CD34+ cell yield was significantly increased for the SCF group in the first leukapheresis (11.3 v 4.0 × 106/kg, P = .003) and all leukaphereses (12.4v 8.2 × 106/kg, P = .007). Total colony-forming unit–granulocyte-macrophage (CFU-GM) and mononuclear cell counts were also significantly higher in the SCF group in the first leukapheresis and in all leukaphereses. As expected for patients mobilized to an optimal CD34+ cell yield, the time to engraftment was similar between the 2 treatment groups. Cells mobilized with the combination of SCF plus filgrastim were thus considered effective and safe for achieving rapid engraftment. Treatment with SCF plus filgrastim was well tolerated, with mild to moderate injection site reactions being the most frequently reported adverse events. There were no serious allergic-like reactions to SCF. The addition of SCF to filgrastim after cyclophosphamide for PBPC mobilization resulted in a significant increase in CD34+cell yield and a concomitant reduction in the number of leukaphereses required to collect an optimal harvest of 5 × 106CD34+ cells/kg.

Description: Targeting the tyrosine kinase activity of Bcr-Abl with STI571 is an attractive therapeutic strategy in chronic myelogenous leukemia (CML). A few CML cell lines and primary progenitors are, however, resistant to this compound. We investigated the mechanism of this resistance in clones of the murine BaF/3 cells transfected with BCR-ABL and in 4 human cell lines from which sensitive (s) and resistant (r) clones were generated by various methods. Although the resistant cells were able to survive in the presence of STI571, their proliferation was approximately 30% lower than that of their sensitive counterparts in the absence of the compound. The concentration of STI571 needed for a 50% reduction in viable cells after a 3-day exposure was on average 10 times higher in the resistant (2-3 μmol/L) than in the sensitive (0.2-0.25 μmol/L) clones. The mechanism of resistance to STI571 varied among the cell lines. Thus, in Baf/BCR-ABL-r, LAMA84-r, and AR230-r, there was up-regulation of the Bcr-Abl protein associated with amplification of the BCR-ABL gene. In K562-r, there was no Bcr-Abl overexpression, but the IC50 for the inhibition of Bcr-Abl autophosphorylation was increased in the resistant clones. Sequencing of the Abl kinase domain revealed no mutations. The multidrug resistance P-glycoprotein (Pgp) was overexpressed in LAMA84-r, indicating that at least 2 mechanisms of resistance operate in this cell line. KCL22-r showed neither Bcr-Abl up-regulation nor a higher threshold for tyrosine kinase inhibition by STI571. We conclude that BCR-ABL–positive cells can evade the inhibitory effect of STI571 by different mechanisms, such as Bcr-Abl overexpression, reduced intake mediated by Pgp, and, possibly, acquisition of compensatory mutations in genes other than BCR-ABL.

Description: Pharmacokinetic monitoring is widely used in different medical specialities, but it has been rarely applied in clinical oncology practice. The current gold standard treatment of chronic myelogenous leukemia (CML) is imatinib, a tyrosine kinase inhibitor. We have previously shown the necessity to obtain a trough plasma threshold of 1000 ng/mL for efficient treatment with imatinib. We routinely perform centralized quantification for patients in France and this has allowed the assessment of imatinib therapeutic monitoring and its use in a real-life setting. After 16 months of data collection, we had gathered 1607 samples for 1044 CML patients (mean age 55 years, F/M sex ratio 0.67) treated with imatinib 400 mg (median) range (100–800mg). We received only one sample for 739 patients and more than one sample for 305 patients. The mean trough plasma concentration of imatinib (Cmin) was 1043 ng/mL (median: 876 ng/mL) and 596 of the 1044 CML patients (57%) had a Cmin