Detection of BCR-ABL Point Mutations in Patients with Chronic Myeloid Leukemia (CML) Resistant to Imatinib and Prognosis

Reviewer: Eric Shinohara MD, MSCI
Abramson Cancer Center of the University of Pennsylvania
Ultima Vez Modificado: 21 de marzo del 2007

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Presenter: Silveira, RA
Presenter's Affiliation: UNICAMP, Brazil
Type of Session: Scientific

Background

  • Imatanib (Gleevec) has been established as first line treatment in Chronic Myeloid Leukemia (CML).
  • Resistance to Imatinib frequently occurs in Acute Blast Crisis (ABC).
  • More than 70 mutations in BRC/ABL have been identified and mutations in BRC/ABL are the most frequent mechanism associated with Imatinib resistance. 
  • Mutations in BRC/ABL have been linked with a poor prognosis.
  • This study was undertaken to detect mutations of the kinase domain of BCR/ABL in CML patients who have primary or secondary resistance to Imatinib and to describe the clinical outcome of these patients after the detection of BCR/ABL mutations.

Materials and Methods

  • 20 patients with CML that was primarily or secondarily resistant to Imatinib were evaluated. 
  • BCR/ABL mutation identification:
    • RNA was extracted from erythroblasts extracted from peripheral blood samples. 
    • Amplification of the BCR/ABL binding domain, specifically amino acids 244-486, was performed using a semi-nested real time PCR (RT-PCR).
    • PCR samples were then submitted to Genbank for sequencing and comparison of these sequences with normal BCR/ABL sequences was performed.   

Results
13 of 20 patients with primary or secondary resistance to Imatinib were found to have mutations in the BCR/ABL binding domain.

  • Of the 13 patients with resistance, five were in Blast Crisis (BC), four in Accelerated Phase (AP), and four in Chronic Phase (CP).
  • There were 11 men and nine women in the study.
  • Median age of the patients was 29.6 years
  • The majority of patients were previously treated with hydroxyurea or a combination of hydorxyuria and interferon therapy. 
  • 15% of patients had a Major Cytologic Response (MCR) to Imatinib and the median time to Imatinib resistance was 8.7 months.
Seven different types of mutations were found in these 13 patients.  The seven mutations identified are as follows, with the number of patients with that specific mutation in parenthesis:
  • T315I (5), L248V (1), G250E (1), F359V (2), M244V (1), E255K (2) and E279K (1)
  • In the remaining seven patients no mutations were found, however clononal evolution was found in two of the seven patients.
Of the 13 patients with mutations, seven were treated with Dasatinib. 
  • PCR analysis was repeated after treatment with Dasatinib and the continued presence of the mutations was demonstrated.
  • Zero out of five patients with the T315I mutation responded to treatment with Dasatanib.
T315I mutations are associated with the Imatinib binding site.

Two patients are dead, two are on treatment with hydroxyuria, one is in AP and one is in CP.  One was in AP and is awaiting bone marrow transplant.
  • One patient with the M244V mutation is still on Dasatinib with a MCR.
  • One patient with the F359V mutation in AP underwent non-myeloblative stem cell transplant from an identical sibling donor, however there was no response.  Repeat PCR found that the mutation was still present and the patient was placed on a phase II clinical trial with Dasatinib.
  • The remaining patients with Phosphate binding loop (P-loop) mutations progressed to blast crisis and died.
Kaplan Meier curves demonstrated the worst overall survival in patients with P-loop mutations, followed by those with T315I mutations, followed by patients with the remaining types of mutations grouped together. 

Author's Conclusions

  • Mutations of the BCR/ABL binding domain are frequent in patients resistant to Imatinib.
  • Patients with mutations of P-loop carry the worst prognosis, with rapid progression to blast crisis.
  • Dasatinib is effective in many patients with mutations in the BCR/ABL binding domain, however it is ineffective in patients with the T135I mutation.
  • Evaluation for mutations in BCR/ABL is important in determining how patients respond to treatment.

Clinical/Scientific Implications
The role of molecular markers continues to expand in the treatment of cancer.  Though preliminary, this study suggests that P-loop mutations may represent a prognostic marker for progression to blast crisis and that the T135I mutation may indicate Dasatinib resistance in patients with CML.   The results of this study support the need for further studies and raises several interesting questions.

  • What accounted for the Imatinib resistance in the seven patients who were resistant but for whom no mutation was found?  Other sites besides the BCR/ABL binding domain may play a role in response to Imatinib.
  • The T135I mutation involving the Imatanib binding domain showed resistance to Damatinib, other alternative targets, perhaps other than those involving the BCR/ABL binding domain, may need to be established for these patients.   
  • Previous studies have suggested that while P-loop mutations are commonly present in CML cells, they did not play a role in prognosis.  Further study of the P-loop mutation may explain this discrepancy in findings. 

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