Dissecting the Complexity of Philadelphia-Positive Mutated Populations by Ultra-Deep Sequencing of the Bcr-Abl Kinase Domain: Biological and Clinical Implications

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 692-692 ◽  
Author(s):  
Simona Soverini ◽  
Caterina De Benedittis ◽  
Katerina Machova Polakova ◽  
Adela Brouckova ◽  
Fausto Castagnetti ◽  
...  
Keyword(s):  
Deep Sequencing ◽  
Research Funding ◽  
Kinase Inhibitor ◽  
Lymphoblastic Leukemia ◽  
Kinase Domain ◽  
Mutation Status ◽  
Abl Kinase ◽  
Selection Of ◽  
Higher Sensitivity

Abstract Abstract 692 Background and Aims: In chronic myeloid leukemia (CML) and Philadelphia-positive acute lymphoblastic leukemia (ALL), tyrosine kinase inhibitor (TKI) therapy may select for drug-resistant Bcr-Abl mutants. Mutation status of resistant patients is usually investigated by Sanger sequencing (SS) of the Bcr-Abl kinase domain (KD). Novel ultra-deep sequencing (UDS) technologies allow to conjugate higher sensitivity with the unprecedented possibility to perform instant cloning of thousands of DNA molecules. We thus decided to take advantage of an UDS-based approach in order to: Methods: We retrospectively performed a longitudinal analysis of a total of 111 samples from 35 CML or Ph+ ALL patients who had received sequential treatment with multiple TKIs (two to four TKIs among imatinib, dasatinib, nilotinib, ponatinib) and had experienced sequential relapses accompanied by selection of TKI-resistant mutations. All samples had already been scored by SS; 74/111 (67%) were positive for one (n=33) or multiple (n=41) mutations. UDS of the Bcr-Abl KD was done using Roche 454 technology. UDS allowed to achieve a lower detection limit of at least 0.1% – as compared to 20% of SS. Results: Bcr-Abl KD mutation status was found to be more complex than SS had previously shown in 85/111 (77%) samples (representative examples are detailed in Table 1). In 33/74 (44%) samples known to harbour one or more mutations by SS, UDS revealed that up to four ‘minor’ mutations with 1–20% abundance were present in addition to the ‘dominant’ one(s). The type of mutations could easily be accounted for by TKI exposure history, since the majority were known to be poorly sensitive either to the current or to the previous TKI received. The higher degree of complexity was evident also when the clonal relationships of multiple mutations were reconstructed (Table 1). This revealed that identical mutations may be acquired in parallel by independent populations (e.g., one wild-type and one already harboring a mutation), via the same or different nucleotide changes leading to the same amino acid substitution (convergent evolution). In addition, longitudinal quantitative follow-up of mutated populations revealed that: Conclusions: Disclosures: Soverini: ARIAD: Consultancy; Bristol-Myers Squibb: Consultancy; Novartis: Consultancy. Castagnetti:Novartis: Honoraria; Bristol Myers Squibb: Honoraria. Luppi:CELGENE CORPORATION: Research Funding. Rosti:Bristol Myers Squibb: Consultancy, Honoraria; Novartis: Consultancy, Honoraria, Research Funding. Baccarani:ARIAD, Novartis, Bristol Myers-Squibb, and Pfizer: Consultancy, Honoraria, Speakers Bureau. Martinelli:NOVARTIS: Consultancy, Honoraria, Speakers Bureau; BMS: Consultancy, Honoraria, Speakers Bureau; PFIZER: Consultancy; ARIAD: Consultancy.

scholarly journals The 'Next-in-Cml' Study: A Prospective Multicenter Study of Deep Sequencing of the BCR-ABL1 Kinase Domain in Philadelphia Chromosome-Positive Patients with Non-Optimal Responses to Tyrosine Kinase Inhibitor Therapy

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 3097-3097
Author(s):  
Simona Soverini ◽  
Caterina De Benedittis ◽  
Stefania Stella ◽  
Anna Serra ◽  
Francesca Carnuccio ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Tyrosine Kinase Inhibitor ◽  
Board Of Directors ◽  
Deep Sequencing ◽  
Kinase Inhibitor ◽  
Kinase Domain ◽  
Second Phase ◽  
Advisory Committees ◽  
Mutation Status

Abstract Introduction Some retrospective studies in tyrosine kinase inhibitor (TKI)-resistant Philadephia-positive (Ph+) leukemia patients (pts) have suggested that deep sequencing (DS) may provide a more accurate picture of BCR-ABL1 kinase domain (KD) mutation status as compared to conventional sequencing (CS). However, the frequency and clinical relevance of low burden mutations remains to be explored prospectively in large series of unselected pts. In addition, the implementation of routine BCR-ABL1 DS in multiple molecular diagnostic laboratories has never been attempted. These open issues led us to design a multi-center, multi-laboratory prospective study ('NEXT-IN-CML') aimed to assess the feasibility, performance and informativity of DS for BCR-ABL1 KD mutation screening. Aims The first phase of the study was aimed to establish a network of 5 reference labs sharing a standardized DS workflow, a joint database for clinical and mutational data storage and a common pipeline of data analysis, interpretation and clinical reporting. The second phase of the study, involving 54 Italian Hematology Units, is aimed to assess the frequency and clinical significance of low burden mutations detectable by DS by prospective collection and analysis of samples from chronic myeloid leukemia (CML) pts who exhibit failure (F) or warning (W) responses and relapsed Ph+ acute lymphoblastic leukemia (ALL) pts. Methods A PCR and an amplicon DS protocol already set up and optimized for the Roche GS Junior in the framework of the IRON II international consortium was adopted. In the first phase, 5 batches of blinded cDNA samples were prepared and shipped to evaluate individual lab performances. The batches included archival samples with known BCR-ABL1 mutation status as assessed by CS and serial dilutions of BaF3 T315I+ cells in BaF3 unmutated cells, simulating mutation loads of 20% down to 1%. In the ongoing second phase prospectively, consecutively collected CML and Ph+ ALL samples are being analyzed in parallel by CS and DS. Clinical history and follow-up data are used for correlations. Results In the first phase of the study, 312/320 amplicons were successfully generated and sequenced. A median of 124,686 (range, 48,181-170,687) high quality reads were obtained across the 5 labs. Median number of forward and reverse reads was 1,757 (range 884-7,838), with no coverage dropouts for any amplicon or index. Comparison of observed vs expected mutations showed that 76/78 evaluable samples were accurately scored. In the remaining two, the analysis software failed to detect the 35bp insertion ('35INS') commonly detectable between exons 8 and 9. Quantitation of point mutation burden was highly reproducible across the entire range of frequencies, from 100% to 1%. The second phase of the study has started in Jan 2016. As of Jul 31st, a total of 106 consecutive pts (CML, n=96; Ph+ ALL, n=10) have been enrolled. The present analysis focuses on the first 75 CML pts (60 F and 15 W), for whom sequencing results are currently available (analysis of the entire population of patients enrolled up to Nov 2016 will be presented at the meeting). Clinically actionable mutations have been detected in 10/75 (14%) pts by CS and in 20/75 pts (27%) by DS. Notably, among the 10 pts positive for clinically actionable mutations by DS but not by CS, 3 had a low burden T315I (2 F [dasatinib, imatinib] and 1 W [dasatinib]). In 5 additional pts negative for mutations by CS (3 F and 2 W), DS identified multiple low burden mutations with unknown IC50, suggesting that the cooperation of individually 'weak' mutants may be a new mechanism underlying reduced TKI efficacy. Longitudinal analysis and follow-up of pts are shaping the clinical significance of different types of low burden mutations and will be presented. Conclusions The 'NEXT-in-CML' study is demonstrating that DS of BCR-ABL1 can successfully be implemented in national lab networks and is an important step forward towards routine use of this technology. We have now adapted the protocol for both the Ion Torrent PGM and the Illumina Miseq platforms. For a minimum of 15 samples per sequencing run, DS costs are estimated to equal those of CS (cost per sample, reagents only: ≈100€ for PGM (314 chip) and Miseq (nano kit v2) vs ≈95€ for CS) with comparable turnaround times for delivery of results. Our study is also contributing useful data for the clinical interpretation of DS findings. Disclosures Soverini: Bristol-Myers Squibb: Consultancy; Ariad: Consultancy; Novartis: Consultancy. Castagnetti:ARIAD Pharmaceuticals: Consultancy, Honoraria; Novartis: Consultancy, Honoraria; Bristol-Myers Squibb: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria. Ciceri:MolMed SpA: Consultancy. Breccia:Novartis: Consultancy, Honoraria; Bristol Myers Squibb: Honoraria; Celgene: Honoraria; Ariad: Honoraria; Pfizer: Honoraria. Di Raimondo:Janssen-Cilag: Honoraria. Bassan:Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Pfizer: Honoraria; Ariad: Honoraria, Membership on an entity's Board of Directors or advisory committees. Cavo:Millennium: Consultancy, Honoraria; Amgen: Consultancy, Honoraria; Janssen-Cilag: Consultancy, Honoraria; Bristol-Myers Squibb: Consultancy, Honoraria; Celgene: Consultancy, Honoraria. Rosti:Novartis: Consultancy, Honoraria; Bristol-Myers Squibb: Consultancy, Honoraria; Ariad: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria. Baccarani:Novartis: Consultancy, Honoraria; Bristol-Myers Squibb: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria. Saglio:Roche: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; ARIAD: Consultancy, Honoraria; BMS: Consultancy, Honoraria; Novartis: Consultancy, Honoraria. Martinelli:Ariad: Consultancy, Speakers Bureau; Pfizer: Consultancy, Speakers Bureau; Roche: Consultancy, Speakers Bureau; Novartis: Speakers Bureau; BMS: Speakers Bureau; Celgene: Consultancy, Speakers Bureau; Amgen: Consultancy, Speakers Bureau; MSD: Consultancy; Genentech: Consultancy.

Ultra-Deep Sequencing of the Bcr-Abl Kinase Domain Allows Earlier Detection and More Accurate Characterization of Resistant Subclones in Philadelphia-Positive Acute Lymphoblastic Leukemia Patients Receiving Tyrosine Kinase Inhibitor-Based Therapies

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 284-284
Author(s):  
Simona Soverini ◽  
Caterina De Benedittis ◽  
Katerina Machova Polakova ◽  
Adela Brouckova ◽  
Cristina Papayannidis ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Tyrosine Kinase ◽  
Tyrosine Kinase Inhibitor ◽  
Deep Sequencing ◽  
Sanger Sequencing ◽  
Kinase Inhibitor ◽  
Lymphoblastic Leukemia ◽  
Mutation Screening ◽  
Kinase Domain ◽  
Abl Kinase

Abstract Abstract 284 Background and Aims: Selection of drug-resistant mutations in the Bcr-Abl kinase domain (KD) is a critical problem undermining the long-term efficacy of tyrosine kinase inhibitor (TKI)-based therapies in Philadelphia-positive (Ph+) acute lymphoblastic leukemia (ALL) patients. Bcr-Abl KD mutation screening is routinely performed by Sanger sequencing (SS). Before the advent of ultra-deep sequencing (UDS) technologies, no method was available that could conjugate the possibility to scan the KD for the so many mutations known to be associated with TKI resistance with a sensitivity higher than that of SS. UDS technologies also allow high throughputness and accurate quantitation of mutated clones and their application in a diagnostic setting is not far to come. We used an UDS strategy for Bcr-Abl KD mutation screening in order to study the dynamics of expansion of mutated clones in Ph+ ALL patients receiving TKI-based therapies and to test the ability of UDS to highlight emerging clones harboring critical mutations. Methods: 72 samples from 25 Ph+ ALL patients who had developed resistance to one or multiple lines of TKI (imatinib, dasatinib, nilotinib, bosutinib, ponatinib) therapy were selected for this retrospective analysis. All the patients had previously been analyzed by Sanger sequencing (SS) and were known to have developed one or more TKI-resistant Bcr-Abl KD mutations on treatment. In order to reconstruct the dynamics of mutation emergence, longitudinal re-analysis of monthly collected samples was perfomed with UDS on a Roche GS Junior. UDS allowed to achieve a lower detection limit of at least 0.1% (by generating a minimum of 5,000 sequence reads/patient), as compared to 20% of SS. Results: 39 samples were known to harbor one (n=27 samples) or more (n=12 samples) TKI-resistant mutations with >20% abundance, as assessed by SS. UDS could successfully detect all the 54 mutations previously identified by SS. In addition, UDS detected one or multiple lower-level (<20%) mutations in 42/72 (58%) samples, demonstrating that in more than half of the cases SS may misclassify Bcr-Abl KD mutation status or underestimate its complexity. Lower-level mutations were indeed found both in samples that had been scored as wild-type by SS and in samples already harboring mutations with >20% abundance. The type of lower-level mutations detected by UDS could easily be accounted for by TKI exposure history, since the majority were known to be poorly sensitive either to the TKI being administered or to the previous TKI received. Overall, 44 samples turned out to carry multiple (two to five) mutations at any level, distributed in the same and/or in different subpopulations with a complex clonal architecture that UDS allowed to reconstruct. Of note, in 14/25 (56%) patients with molecularly detectable disease but not yet evidence of cytogenetic or hematologic relapse, UDS could identify emerging TKI-resistant mutations 1 to 2 months before they became detectable by SS. These outgrowing mutations were detected at 1–19% abundance in 12 patients and at 0.1–1% abundance in 2 patients. In the remaining 11 patients, dynamics of outgrow of the TKI-resistant mutations (five T315I, two Y253H, two E255K, one E255V and one F317L) was so rapid that not even strict monthly monitoring could allow to pick them up before they became dominant. Conclusions: Now that multiple options are available, Bcr-Abl KD mutation monitoring has become a precious tool for rational decision-making in order to maximize the efficacy of TKI-based regimens as induction or salvage therapy for Ph+ ALL patients. UDS proved as reliable as SS for the detection of mutations with >20% abundance and to have comparable costs. As a key advantage, UDS added precious quantitative and qualitative information on the full repertoire of mutated populations, that SS failed to appreciate in more than half of the samples analyzed. TKI-resistant mutations leading to patient relapse were not necessarily preexisting at low levels at diagnosis or at the time of switchover to another TKI, underlining the importance of regular monitoring of patients. Although TKI-resistant populations may arise and take over very rapidly, in approximately half of the patients monthly monitoring with UDS would have allowed to identify them earlier than SS and well in advance of clinical relapse, thus allowing a more timely therapeutic intervention. Disclosures: Soverini: Novartis: Consultancy; Bristol-Myers Squibb: Consultancy; ARIAD: Consultancy. Luppi:CELGENE CORPORATION: Research Funding. Baccarani:ARIAD, Novartis, Bristol Myers-Squibb, and Pfizer: Consultancy, Honoraria, Speakers Bureau. Martinelli:NOVARTIS: Consultancy, Honoraria, Speakers Bureau; BMS: Consultancy, Honoraria, Speakers Bureau; PFIZER: Consultancy; ARIAD: Consultancy.

Treating Ph+ Acute Lymphoblastic Leukemia (ALL) in the Elderly: The Sequence of Two Tyrosine Kinase Inhibitors (TKI) (Nilotinib and Imatinib) Does Not Prevent Mutations and Relapse.

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 2601-2601 ◽  
Author(s):  
Cristina Papayannidis ◽  
Paola Fazi ◽  
Alfonso Piciocchi ◽  
Francesco Di Raimondo ◽  
Giovanni Pizzolo ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Tyrosine Kinase ◽  
Research Funding ◽  
Kinase Inhibitors ◽  
Lymphoblastic Leukemia ◽  
The Elderly ◽  
Kinase Domain ◽  
Intensive Chemotherapy ◽  
Molecular Responses ◽  
Abl Kinase

Abstract Abstract 2601 Background: Tyrosine Kinase Inhibitors (TKI) have been shown to be very effective for the treatment of Acute Lymphoblastic Leukemia (ALL), with a Complete Hematologic Remission (CHR) rate close to 100%, and a high rate of Complete Cytogenetic and Molecular responses (CCgR and CMR). However, when they are used alone, as single agents, most patients relapse, so that they are currently used in combination with chemotherapy and as a preparation to allogeneic stem cell transplantation (SCT). Since Ph+ ALL is more frequent in the elderly, many patients cannot tolerate intensive chemotherapy and are not eligible for SCT. We have explored if the administration of two TKIs, Nilotinib (NIL) and Imatinib (IM) can improve the results without increasing the toxicity. Aims: To evaluate the response and the outcome of Ph+ ALL patients treated with the sequential administration of NIL and IM, to investigate the type and number of BCR-ABL kinase domain mutations developing during and after the study. Methods: We have designed a study (ClinicalTrials.gov. NCT01025505) in which patients more than 60 years old or unfit for intensive chemotherapy and SCT where treated with two TKIs, NIL 400 mg twice daily, and IM 300 mg twice daily, alternating for 6 weeks for a minimum of 24 weeks (study core) and indefinitely in case of response. The 6-weeks rotation schedule was respected, irrespectively of temporary discontinuations. The primary end-point was the rate of Disease Free Survival (DFS) at 24 weeks (4 courses of treatment); the secondary end points included the evaluation of CHR, CCgR and CMR rates. Mutation analysis was performed by nested RT-PCR amplification of the ABL kinase domain of the BCR-ABL transcript (codons 206 through 421). Amplified products were screened by denaturing-high performance liquid chromatography (D-HPLC). Samples scored positive for the presence of sequence variations were then subjected to direct automatic sequencing to characterize the mutation. Results: 39 patients have been enrolled in 15 Italian hematologic Centers (median age 66 years, range 28–84). Among these, 8 patients were unfit for standard chemotherapy or SCT (median age 50 years, range 28–59). 27 patients were p190, 5 were p210 and 7 were p190/p210. After 6 weeks of treatment, 36 patients were evaluable for response: 34 were in CHR (94%) and 2 in PHR (6%). 23 patients have already completed the study core (24 weeks), 87% were in CHR and 17 are currently continuing therapy in the protocol extension phase. Thus, the OS at 1 year is 79%, and 64% at 2 years. Overall, 1 patient was primarily resistant and 13 patients have relapsed, with a median time to relapse of 7.6 months (range 0.8–16.1 months), for a DFS of 51.3% at 12 months (Figure 1). Mutations detected were T315I in 2 cases, Y253H in 3 cases, T315I and Y253H in 1 case, E255K in 1 case, T315I and E255K in 1 case, E255V and Y253H in 1 case. Two patients were WT. A detailed kinetics of Molecular responses is shown in Table 1. Data on mutational analysis are reported in Table 2. Further details about Cytogenetic and Molecular responses, and about Adverse Events will be provided on site. Conclusions: In this small cohort of Ph+ ALL elderly/unfit patients, the rates of relapse and progression were not likely to be different from the rates observed with Imatinib alone (Vignetti et al, Blood 2007, May 1;109(9):3676-8) and Dasatinib alone (Foà, Blood 2011, Dec 15;118(25):6521-8). It's important to notice that the mutations that occurred at the time of relapse were sensitive to other TKIs (Dasatinib and Ponatinib). Acknowledgments: COFIN, Bologna University, BolognAIL, PRIN, Fondazione del Monte di Bologna e Ravenna, INPDAP. Disclosures: Pizzolo: Hoffmann-La Roche: Consultancy, Honoraria. Luppi:CELGENE CORPORATION: Research Funding. Vallisa:CELGENE CORPORATION: Research Funding. Martinelli:NOVARTIS: Consultancy, Honoraria, Speakers Bureau; BMS: Consultancy, Honoraria, Speakers Bureau; PFIZER: Consultancy; ARIAD: Consultancy. Baccarani:ARIAD, Novartis, Bristol Myers-Squibb, and Pfizer: Consultancy, Honoraria, Speakers Bureau.

scholarly journals Unraveling the complexity of tyrosine kinase inhibitor–resistant populations by ultra-deep sequencing of the BCR-ABL kinase domain

Blood ◽  
2013 ◽  
Vol 122 (9) ◽  
pp. 1634-1648 ◽  
Author(s):  
Simona Soverini ◽  
Caterina De Benedittis ◽  
K. Machova Polakova ◽  
Adela Brouckova ◽  
David Horner ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Tyrosine Kinase Inhibitor ◽  
Deep Sequencing ◽  
Sanger Sequencing ◽  
Kinase Inhibitor ◽  
Kinase Domain ◽  
Abl Kinase ◽  
Key Points ◽  
Conventional Methods

Key Points UDS demonstrated that BCR-ABL KD mutations detectable with conventional methods may just be the tip of the iceberg. The information provided by conventional Sanger sequencing may not always be sufficient to predict responsiveness to a given TKI.

A Novel BCR-ABL Mutation Predicts Resistance to Tyrosine Kinase Inhibitor (TKI) Therapy by a Unique Mechanism in Patients (pts) with Philadelphia-Positive (Ph+) Leukemia.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 4773-4773
Author(s):  
Alfonso Quintas-Cardama ◽  
Jorge Cortes ◽  
Hagop M. Kantarjian ◽  
Moshe Talpaz ◽  
Ji Wu ◽  
...  
Keyword(s):  
Kinase Activity ◽  
Kinase Inhibitor ◽  
Lymphoblastic Leukemia ◽  
Chronic Phase ◽  
Kinase Domain ◽  
Abl Kinase ◽  
High Level ◽  
A Minor ◽  
Unique Mechanism

Abstract Albeit most pts with chronic myeloid leukemia (CML) treated with imatinib (IM) have a favorable outcome, some will acquire resistance, mainly due to the development of Abl kinase domain mutations, which confer varying levels of TKI resistance. We describe a novel V304D mutation in pts with Ph+ leukemia who failed TKI therapy. Expression of V304D mutation in BCR-ABL failed to induce cytokine-independence in Ba/F3 cells. Studies in Cos-7 cells demonstrated that this mutant did not induce autophosphorylation and was deficient in kinase activity. We detected V304D mutation in 13 (18%) of 70 IM-resistant pts screened (12 CML, 1 Ph+ acute lymphoblastic leukemia [ALL]), and it was present in a median of 37% (range, 20% to 80%) resistant clones. Median age was 60 years (range, 30 to 81) and median time from diagnosis to IM therapy was 39 months (range, 1 to 91). Eleven (92%) of 12 pts with CML were in chronic phase (CP) at IM start and 1 was in blast phase (BP). Pts received IM for a median of 35 months (range, 2 to 66). Nine pts with CML had failed interferon and 2 (1 CML, 1 Ph+ALL) allogeneic stem cell transplantation prior to IM. Ten (83%) of 12 pts started IM at 400 mg/d but all eventually received ≥600 mg/d. Six pts with CML achieved a complete hematologic response (CHR), 1 BP returned to chronic phase (RCP), and 6 (5 CML, 1 Ph+ALL) had primary hematologic resistance (HR). No cytogenetic (CG) responses were observed and 7 pts with CML CP progressed (4 to AP and 3 to BP) after IM discontinuation. Four pts with CML (1 CP, 2 AP, 1 BP) received nilotinib after IM failure for a median of 2 months (range, 1 to 3.5). Two pts (1 CP, 1 AP) showed primary HR, 1 AP progressed to BP, and 1 BP (on 600 mg twice daily) had a transient (6 weeks) CHR before showing secondary HR. Twelve pts (11 CML, 1 Ph+ALL) received dasatinib: 7 at 70 mg twice daily, 1 at 90 mg daily, 1 at 140 mg daily, 1 at 180 mg daily, 1 at 90 mg twice daily, and 1 at 120 mg twice daily. Dasatinib was administered for a median of 8 months (range, 1 to 23). Two pts achieved CHR and a minor CG response in 1 analysis (75% and 65% Ph+ cells, respectively), 1 RCP, 1 no evidence of leukemia, and 8 (67%) primary HR. One of 4 pts who started dasatinib in CP progressed to AP. Responders to dasatinib had V304D mutation in 20%, 20%, and 25% of clones, respectively. Four pts exhibited concomitant Abl kinase mutations developed prior to dasatinib therapy: 3 with F317L and 1 with G250E. One pt had a 6 base pair in-frame insertion in the TK domain. T315I mutation evolved in 1 pt after dasatinib discontinuation. Eight pts discontinued dasatinib due to disease progression (7 died), 2 were lost to follow-up, and 2 remain on CHR after 17+ and 23+ months on dasatinib. In vitro studies of cells from one pt in CP with V304D mutation (50% of clones) failed to detect CrkL phosphorylation despite detectable expression of the Bcr-Abl protein. In summary, the V304D mutation in the Abl kinase domain results in kinase inactivation and is associated with high-level resistance to TKI therapy, transformation to AP/BP in CML and a particularly poor prognosis. Loss of kinase activity by mutation represents a very unique mechanism of kinase inhibitor resistance and predicts acquisition of other transforming events that support CML cell survival.

scholarly journals A Comprehensive Cancer Center Experience with Hyper-CVAD Plus Ponatinib As Frontline Therapy for Adult Philadelphia Chromosome-Positive Acute Lymphoblastic Leukemia

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 4394-4394
Author(s):  
Tamer Othman ◽  
Benjamin Moskoff ◽  
Matthew Tenold ◽  
Tali Azenkot ◽  
Margaret Krackeler ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Bacterial Infection ◽  
Research Funding ◽  
Kinase Inhibitor ◽  
Lymphoblastic Leukemia ◽  
Blast Crisis ◽  
Philadelphia Chromosome ◽  
Mixed Phenotype ◽  
Philadelphia Chromosome Positive

Abstract Background Ponatinib, a third-generation BCR-ABL1 tyrosine kinase inhibitor (TKI), + hyper-CVAD showed remarkable activity against Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph+ ALL), and may be superior to chemotherapy + earlier generation TKIs in terms of depth of remission, event-free survival (EFS), and overall survival (OS). However, this regimen's efficacy and tolerability have yet to be externally validated. Here, we summarize our real-world experience with ponatinib + hyper-CVAD for untreated Ph+ ALL and other Ph+ acute or blast phase leukemias. Methods We retrospectively analyzed all adults treated at the University of California, Davis (UCD) from March 2012 to May 2021 with ponatinib + hyper-CVAD upfront. The primary endpoints were 3-year OS and EFS. Secondary endpoints were complete molecular response (CMR), measurable residual disease (MRD) negativity by multiparameter flow cytometry (MFC), complete cytogenetic response (CCyR) rates, and adverse events (AEs). Time to event analyses were done via the Kaplan-Meier method. Patients alive were censored at their last follow-up date. Patients undergoing allogeneic hematopoietic cell transplant (HCT) after 6 months of achieving complete remission (CR) were censored at the time of HCT for the landmark analysis. Patients with missing data were excluded from the response analyses. Results We identified 13 Ph+ ALL patients who received ponatinib + hyper-CVAD for initial induction. The baseline characteristics for the Ph+ ALL patients are summarized in Table 1. The median follow-up was 16 months. The median number of hyper-CVAD cycles completed was 8 (range, 1-8) with ponatinib. Two patients proceeded to HCT in CR1, one at 3.5 months after starting induction, and due to difficulty controlling the patient's concurrent multiple myeloma prior to HCT and recovery from anti-neoplastic therapy, the second was delayed to 46 months after starting induction. The 3-year OS and EFS with ponatinib + hyper-CVAD were each 92% (95% confidence interval, 78.9-100) (Figure 1). Landmark analysis completed 6 months following CR showed a 3-year OS of 100% in patients treated with ponatinib + hyper-CVAD without HCT in first CR (CR1). The CMR, CCyR, and MRD-negativity by MFC rates with ponatinib were all 92.3% (12/13). The median time to CMR, CCyR, and MRD-negativity by MFC were 51 days, 22 days, and 53 days, respectively. Notable AEs with ponatinib include neutropenic fever (92%), bacterial infection (69%), transaminitis (38%), venous thromboembolism (31%), invasive fungal infection (15%), hemorrhage (15%), cerebrovascular accident (CVA) (15%), and tumor lysis syndrome (8%). One patient died during induction with ponatinib due to a bacterial infection. Two patients switched to a different TKI due to a CVA after 4 and 24 months. Only 2 patients did not complete 8 cycles of hyper-CVAD, due to death during induction (n=1) and proceeding to HCT after 3 cycles (n=1). As for similar Ph+ leukemias, 3 chronic myeloid leukemia with lymphoid blast crisis (CML-LBC), 1 CML with mixed phenotype blast crisis (CML-MPBC), and 1 with mixed phenotype acute leukemia (MPAL) were treated with ponatinib + hyper-CVAD. The MPAL patient achieved CMR within 55 days, while the CML-MPBC and 2 CML-LBC patients achieved CMR after HCT. The third CML-LBC patient is in CR with ongoing treatment. After median follow-up of 25 months, all 5 were alive, and only the MPAL patient relapsed 28 months after starting treatment and 1 year after HCT. Conclusion To our knowledge, this is the first report externally validating the efficacy and tolerability of ponatinib + hyper-CVAD for Ph+ ALL. We also show the feasibility of using this regimen in patients with Ph+ CML-LBC, CML-MPBC and MPAL. Despite the small sample size and retrospective nature, our study supports existing data demonstrating that this regimen challenges both the designation of Ph+ ALL as a high-risk disease and the trend to transplant in CR1. Our findings support that ponatinib + hyper-CVAD should be considered a standard of care for Ph+ ALL. Figure 1 Figure 1. Disclosures Kaesberg: Incyte: Speakers Bureau. Rosenberg: Takeda, Janssen: Speakers Bureau. Abedi: BMS/Celgene: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Abbvie: Speakers Bureau; Seattle Genetics: Speakers Bureau. Tuscano: Genentech, Pharamcyclics, Abbvie, BMS, Acrotech, Seattle Genetics, Takeda: Research Funding. Jonas: AbbVie, BMS, Genentech, GlycoMimetics, Jazz, Pfizer, Takeda, Treadwell: Consultancy; 47, AbbVie, Accelerated Medical Diagnostics, Amgen, AROG, Celgene, Daiichi Sankyo, F. Hoffmann-La Roche, Forma, Genentech/Roche, Gilead, GlycoMimetics, Hanmi, Immune-Onc, Incyte, Jazz, Loxo Oncology, Pfizer, Pharmacyclics, Sigma Tau, Treadwell: Research Funding; AbbVie: Other: Travel reimbursement. OffLabel Disclosure: Ponatinib is approved for Philadelphia chromosome positive acute lymphoblastic leukemia (Ph+ALL) that is resistant or intolerant to prior tyrosine kinase inhibitor therapy. In this study, we described outcomes with ponatinib in combination with hyperCVAD in the frontline setting, which is off-label.

Sign in / Sign up

Export Citation Format

Share Document