Incidence of the BCR-ABL Kinase Domain Mutations in Imatinib Treated CML Patients: Russian Experience
Authors: Misyurin Andrey V , Misyurina Elena N , Krutov Alexey , Aksenova Elena , Soldatova Irina , Kesaeva Laura , Turkina Anna , Chelysheva Ekaterina , Savchenko Valeri G , Parovichnikova Elena 
Published: October 2019

Abstract 4420

In the period from 18.01.06 to 25.05.11 we have performed mutation analysis in CML pts with the obvious manifestation of resistance against imatinib. Chronic phase CML patients have been treated by imatinib and monitored by means of RQ-PCR using IS units. At first signs of the evident BCR-ABL level increase (more than 5–10% IS), the mutation analysis has been performed by means of direct PCR fragment sequencing. We have used original primers that enable to search for mutations in the area spanning from a3 to a11 ABL exons of BCR-ABL. Sequencing has been performed for 274 CML pts that were suspected to acquire imatinib resistance. Among them there were 47,45% (130/274) of females (median age 51 years, from 15 to 73), 52,55% (144/274) of males (median age 50 years, from 15 to 74). In the group of mutation negative pts (80/274 – 29,2%) the BCR-ABL level was more than 9% (IS). Interestingly, in this group there were a little bit more females than in the total cohort of pts (58,7% against 47,45%, p=0,034). The median age of the mutation negative females (51 years) was quite the same as in the total cohort, whereas the median age of mutation negative males significantly differed from the one in the total cohort (58 vs. 51 year, p=0,041). 70,8% (194/274) pts tested occurred to be mutation positive (42,8% females (83/194), 57,2% males (111/194)). 214 point mutations of 38 different types have been detected, among them T315I (26/194 – 12%), G250E (26/194 – 12%), T317L (16/194 – 7,4%), F359V (15/194 – 7,0%), H396R (15/194 – 7,0%), M244V (15/194 – 7,0%), E255K (12/194 – 5,6%), Y253H (12/194 – 5,6%), E255V (9/194 – 4,2%), L248V (8/194 – 3,7%), E355G (6/194 – 2,8%), M315T (6/194 – 2,8%), Q252H (5/194 – 2,3%), L387F (4/194 – 1,8%), S348L (4/194 – 1,8%), F311I (3/194 – 1,4%), F359C (3/194 – 1,4%), E255D (2/194 – 0,9%), E275K (2/194 – 0,9%), E279A (2/194 – 0,9%), K247R (2/194 – 0,9%), E292V (1/194 – 0,46%), E334G (1/194 – 0,46%), E450K (1/194 – 0,46%), E459A (1/194 – 0,46%), E459K (1/194 – 0,46%), F359I (1/194 – 0,46%), F486S (1/194 – 0,46%), L383F (1/194 – 0,46%), P441L (1/194 – 0,46%), Q252M (1/194 – 0,46%), Q491L (1/194 – 0,46%), T305I (1/194 – 0,46%), T345I (1/194 – 0,46%), V299A (1/194 – 0,46%), Y312C (1/194 – 0,46%), T520S (1/194 – 0,46%). As rare events we have observed some other types of mutations. In one CML patient with primary imatinib resistance there was a G425Stop (1/194 – 0,46%) mutation which caused a significant truncation of the C-end of the BCR-ABL protein. There were also 2 highly imatinib resistant pts with deletions of exon a7 ABL (2/194 – 0,9%). One more unusual mutation was an insertion of 72 bp between the a8 and a9 exons of ABL. The origin of this insertion was a middle part of the 8th intron of the ABL gene. This sequence is arranged as a typical human exon because it is flanked by donor and acceptor splicing signals. We found this mutation in 3 imatinib resistant CML pts (3/194 – 1,4%). This mutation gives rise to the abnormal stop-codon within the exon a9 due to a CD’s frameshift. In all pts with point L248V mutation (8/194 – 3,7%) we have also observed additional abnormal variant of BCR-ABL gene as a splicing isoform lacking 81 bp of exon a4. The point mutation L248V (742C>G) exchange the motif CAAGCT for CAAGGT and the latter is a strong splice acceptor site. It seems that this cryptic splice acceptor site in the middle of 4a exon carrying 742C>G point mutation may compete with natural splice acceptor site from 4th intron. It gives rise to two BCR-ABL transcripts: one carrying a point mutation and the other one with 81 bp’s deletion of the 3’end of exon 4a, without frameshift. These findings suggest that in the case of mutational analysis the considerable attention should be paid not only to BCR-ABL point mutations but also to the possible emergence of the other changes of the BCR-ABL mRNA structure which may occur due to arbitrary activity of splicing machinery. Our findings also suggest that among imatinib resistant CML pts some of them already acquired mutations that prevented adequate response for the 2nd generation of TKI. In our study 12% of BCR-ABL mutation positive CML pts resistant to imatinib (n=26/194) had T315I which also render nilotinib and dasatinib resistance. 23,8% (n=51/194) were positive for mutations that decreases nilotinib response (E255K/V, F359C/V, Y253H) and 7,5% (n=16/194) were positive for T317L that hinders dasatinib response.

Disclosures:

No relevant conflicts of interest to declare.