Mutation Analysis of BCR-ABL Tyrosine Kinase Domain In New Chronic Phase-Chronic Myeloid Leukemia Patients with Suboptimal Response or Treatment Failure From Imatinib Treatment.
Authors: Kim Dong-Wook , Kim Dongho , Kim Soo-Hyun , Goh Hyun-Gyung , Pane Fabrizio , Hughes Timothy , Radich Jerald P. , Lin Peter , Saglio Giuseppe , Branford Susan , Martinelli Giovanni , Soverini Simona , Hochhaus Andreas 
Published: October 2019

Abstract
Abstract 3441
Resistance to imatinib can occur in chronic myeloid leukemia (CML). Mutations in BCR-ABL kinase domain have been known as the clinically most relevant mechanisms of imatinib resistance. Several studies have shown that 40~80% of the imatinib resistant patients have BCR-ABL kinase domain mutations. However, there has not been much information about mutation status in CML patients with suboptimal response to imatinib. With samples from Tyrosine Kinase Inhibitor Optimization and Selectivity (TOPS) study which investigates efficacy of imatinib 400 mg daily and 800 mg daily with Philadelphia positive CML CP patients, and relationship between various responses and emergence of mutation was investigated by both standard and highly sensitive mutation assays.
As a clinical correlative study (CCS) program with samples from TOPS study, we analyzed BCR-ABL mutation from 51 patients with optimal response, suboptimal response or treatment failure to imatinib in order to investigate their mutation status at different time points including diagnosis, 6 months, 12 months and 18 months. In addition, 53 non-clinical trial patients from Seoul St. Mary's hospital were involved in this analysis. All patients were in CP.
Suboptimal responders and treatment failures were selected based on European Leukemia Net (ELN) 2009 guideline. CML patients' samples from the TOPS study and Seoul St. Mary's hospital were collected at particular time points after initiation of imatinib treatment and stored as cryopreserved cells or isolated RNAs. Mutations in BCR-ABL kinase domain were analyzed using direct sequencing and allele-specific oligonucleotide (ASO)-PCR (for Y253H, Y253F, G250E, E255K, E255V, T315I, F359V, and M351T).
We performed mutation analysis with total 164 samples collected from 104 patients at different time points including diagnosis, 6 months, 12 months and 18 months after initiation of imatinib treatment. Serially collected samples at all time points were not available for all the patients; samples from 31 patients were available at diagnosis, 41 patients at 6 months, 58 patients at 12 months and 34 patients at 18 months (Table 1). We found ten BCR-ABL kinase domain point mutations including G250E, Q252H, Y253H, T315I, F317L, E355G, F359V, F359I and D444Y in 13 patients. In addition, we also found other mutations including 35 base pair insertion between exon 8 and exon 9 of ABL, and deletion of exon 7 of ABL in other 10 patients.
No mutation was found from the patients' samples collected at diagnosis. At 6 months, mutation was found 5% (1 of 21), 18% (2 of 11) and 22% (2 of 9) patients in optimal response, suboptimal response and treatment failure group, respectively. ASO-PCR revealed that one patient in optimal response group had T315I. The same mutation status of the patient maintained at 12 months and the patients showed treatment failure at 12 months. At 12 months, mutation portion was 0% (0 of 15), 13% (2 of 15) and 25% (7 of 28) in optimal response, suboptimal response and treatment failure group, respectively. At 18 months, 36% (5 of 14) of suboptimal molecular responders who achieved CCyR, but no MMR showed mutation, and 62% (8 of 13) of failure group who showed less than CCyR had mutations. No particular difference in mutation frequency was found between 400mg group and 800mg group.
Patients with suboptimal response or treatment failure showed much higher chance of BCR-ABL point mutation, 35 base pair insertion or exon 7 deletion in comparison with optimal responders, suggesting that mutation screening is important for patients with suboptimal response as well as treatment failure on the basis of ELN guideline. Treatment failure who achieved less than CCyR at 18 months and suboptimal responders who achieved CCyR, but no MMR at 18 months were highly recommended for mutation screening based on these data. Highly sensitive ASO-PCR provided early detection of point mutation in BCR-ABL kinase domain. However, clinical relevance of low level mutant clone, 35 base pair insertion and exon 7 deletion require long-term follow up for better understanding.
Table 1. Response Diagnosis 6 months 12 months 18 months Pt with mutation Total patients Pt with mutation Total patients Pt with mutation Total patients Pt with mutation Total patients Optimal response 0 31 1 (5%) 21 0 (0%) 15 0 (0%) 7 Suboptimal response 2 (18%) 11 2 (13%) 15 5 (36%) 14 Failure 2 (22%) 9 7 (25%) 28 8 (62%) 13 Total Patients 31 41 58 34

Disclosures:
Pane: Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees. Saglio:Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Hochhaus:Ariad: Consultancy, Membership on an entity's Board of Directors or advisory committees.