Allogeneic stem cell transplantation for patients with chronic myeloid leukemia and acute lymphocytic leukemia after Bcr-Abl kinase mutation–related imatinib failure

Imatinib mesylate is a potent and selective tyrosine kinase inhibitor that has become standard therapy for patients with chronic myeloid leukemia (CML).^1,2  Most newly diagnosed CML patients with chronic-phase (CP) disease treated with imatinib achieve durable responses. However, a small percentage of CP patients and most advanced-phase subjects relapse on therapy.^3-5 

Resistance to imatinib occurs at a rate of approximately 4% per year among patients treated in early CP and at higher rates in those treated in advanced stages. Resistance is most frequently mediated by mutations within the kinase domain of Bcr-Abl and, to a lesser extent, by amplification of the Bcr-Abl genomic locus and other mechanisms.^6,7  Point mutations have been reported in 30% to 90% of resistant patients.

Allogeneic stem cell transplantation (Allo-SCT) is a potentially curative treatment for CML.⁸  However, there is little data in patients with imatinib resistance–associated mutations. In order to address this problem, we analyzed the outcome of patients with Bcr-Abl kinase mutations treated with related or unrelated donor hematopoietic stem cell transplantation (HSCT) at our institution.

Between July 2002 and October 2005, 10 patients with imatinib-refractory CML (n = 9) or Philadelphia-positive (Ph-positive) acute lymphocytic leukemia (ALL; n = 1) received allo-SC transplants from human leukocyte antigen (HLA)–matched related (n = 5) or unrelated (n = 5) donors. Failure to imatinib was defined as (1) loss of a cytogenetic or complete hematologic response (CHR) while on therapy; (2) failure to achieve a CHR (for patients in CP) or any hematologic response (for patients in accelerated phase [AP] or blast phase [BP]) after 3 months of imatinib therapy; or (3) persistence of 100% Ph-positive metaphases after 6 months or greater than 35% after 12 months of therapy.

Preparative regimens are described in Table 1. Graft-versus-host disease (GVHD) prophylaxis consisted of tacrolimus and methotrexate 5 mg/m² intravenously on days 1, 3, 6, and 11 after transplantation. All patients received granulocyte colony-stimulating factor 5 μg/kg daily from day 7 until neutrophil engraftment. Antithymocyte globulin was given to recipients of unrelated donor transplants. Informed consent was obtained in all cases. The M. D. Anderson Cancer Center Institutional Review Board approved this retrospective study.

Standard criteria were used to score CML response.⁹  A complete molecular response (CMR) was defined as an undetectable Bcr-Abl transcript level. A major molecular response (MMR) was defined as Bcr-Abl/Abl ratio less than 0.05%.

Bcr-Abl transcripts were quantitated in a single-tube multiplex real-time reverse transcription–polymerase chain reaction (RT-PCR) assay using TaqMan chemistry (Applied Biosystems, Foster City, CA).¹⁰  Total RNA was isolated from peripheral blood or bone marrow samples by Trizol solubilization (Gibco-BRL, Gaithersburg, MD) and cDNA was synthesized by reverse transcriptase (Superscript II; Invitrogen, Carlsbad, CA). The resulting cDNA was subjected to PCR to amplify b2a2, b3a2, and e1a2 Bcr-Abl fusion transcripts, and quantitative Bcr-Abl levels were normalized to total abl transcript levels as described previously.¹⁰ 

For mutational analysis, the entire kinase domain of the Bcr-Abl fusion transcript was sequenced using a nested PCR method. The Bcr-Abl fusion transcript was first amplified followed by 2 separate PCR reactions that cover exons 221 to 390 and codons 350 to 500 of the Abl kinase domain, respectively, essentially as described.¹¹  Standard dideoxy chain termination cycle sequencing was done using a 3100 genetic analyzer (Applied Biosystems) with analysis using Seqscape v2.0 software (Applied Biosystems). All mutations were confirmed by sequencing of forward and reverse strands, with approximate sensitivity of 15% mutation-bearing cells.¹¹ 

Hematopoietic chimerism was evaluated on peripheral blood or bone marrow by multiplex PCR-based DNA microsatellite polymorphism analysis using PCR D6S264, D3S1282, D18S62, and D3S1300 fluorescence-labeled primer sets, followed by separation by capillary electrophoresis and analysis using GeneScan software (Applied Biosystems). Mixed chimerism was defined as the presence of any detectable (1%) recipient DNA.

Toxicity was graded according to National Cancer Institute criteria. GVHD was graded according to consensus criteria.¹² 

A total of 10 patients (3 in CP, 4 in AP, 2 in BP, and 1 with Ph-positive ALL) received a transplant. The median age was 44 years (range, 26-63 years). Nine different protein kinase mutations were detected. Five patients harbored a P-loop mutation. At the time of Allo-SCT, 1 patient (patient unique identifier [UPIN] 5) was in MMR (Ph-positive ALL, mutation Q252H), 1 (UPIN 7) was in major cytogenetic response, and 2 (UPIN 4 and 10) were in complete hematologic response. The 6 remaining patients were in CP (n = 2) and AP (n = 4) with no hematologic response. Patients' characteristics are summarized in Table 1.

All patients engrafted at a median of 13 days (range, 10-17 days). There were no major toxicities and no treatment-related deaths. Chimerism studies at days 30 and 100 after HSCT were 100% of donor type in all but 1 patient. Nine patients achieved a complete cytogenetic response 1 month after transplantation. All patients were assessed for molecular response more than once: 7 achieved a CMR at a median of 2 months (range, 1-6 months) after transplantation; 2 achieved an MMR (mutations F359V and T315I) by 3 and 4 months after transplantation, with transcript levels of 0.012% and 0.04%, respectively (the first one [UPIN 9] is still responding with the last transcript level of 0.004%, while the second relapsed [UPIN 6] after 6 months). Three patients (ALL with Q252H [UPIN 5], AP with T315I [UPIN 6], and BP with E255K [UPIN 10]) progressed and died of their disease, respectively, 7, 11, and 4 months after Allo-SCT. Patients UPIN 5 and 6 had achieved a CMR and an MMR, whereas patient UPIN 10 did not respond to Allo-SCT. Seven patients are alive (6 in CMR and 1 in MMR) after a median follow-up of 19 months (range, 13-24 months). Outcomes of treatment are summarized in Table 2.

This is the first report on the outcome of imatinib mesylate–refractory patients with kinase mutations undergoing Allo-SCT. Specifically, we showed that in such cases, Allo-SCT is feasible, safe, and frequently effective. A high rate of complete clinical and molecular responses was achieved, despite the fact that most patients received a transplant with imatinib-resistant advanced disease. In this small series, we could detect no difference in outcome between P-loop versus non–P-loop mutations.⁹  Interestingly, 1 of the 2 patients with the T315I mutation responded and remains in molecular remission after 17 months of follow-up, whereas the other relapsed 6 months after transplantation and died shortly after that. This mutation confers resistance to imatinib and to the novel tyrosine kinase inhibitors (NTKIs).^13,14 

The NTKIs dasatinib (BMS-354825) and AMN-107, currently being evaluated in clinical trials, have shown promising results in patients with CML resistant to imatinib.^15-17  Both drugs are active in patients with mutant Bcr-Abl kinase except when the disease is driven by the T315I mutation. In advanced-stage patients such as ours, consideration should be given to maintenance use of NTKI or other agents after transplantation.

These preliminary results support the use of Allo-SCT as a salvage option for patients developing resistance to imatinib through Bcr-Abl mutations. This subset of patients is likely to benefit from early transplantation, should a suitable related or unrelated donor be identified. Long-term follow-up of a larger number of patients will be necessary to fully document the efficacy and benefit of allogeneic transplantation in this setting.