F311l, m351t, and t315i and all patients with preexisting bcrabl mutations exhibited imatinib resistance 33. T315i mutation of bcrabl1 into human philadelphia chromosome. Aurora a kinase inhibitor aki603 induces cellular senescence. It might represent a treatment option for patients with this mutation and complement the current clinically. T315i mutation is at an elevated incidence than other aminoacid exchanges and the recorded t315i mutation incidence in cml patients with imatinib resistance ranges between 2% and 20% with an inconsistency related to methods of detection as well as type of treatment and patient cohort characteristics. In this study, we retrospectively traced small clones with the t315i mutation using dpcr at several specific time points in patients who received allogeneic hsct following dasatinibbased chemotherapy. Do i need to test for the bcr1abl transcript before ordering the abl kinase domain mutation in cml, cellbased test. Bcrabl point mutations and tki treatment in cml patients. Fine3, colleen gritzen1, john hood2, jason kang3, dan lohse2, chi ching mak1, andrew mcpherson1, glenn noronha1, ved p. Ponatinib thwarts t315i gene mutation in chronic myeloid leukemia november 29, 2012, news medical analysis of mutations in the bcrabl1 kinase domain, using direct sequencing. About 20% of patients with relapsed or resistant cml show a specific mutation on the threoninetoisoleucine gene at position 315 t315i mutation. Mar 16, 2016 allogeneic stem cell transplantation sct is currently the only curative treatment option for chronic myeloid leukemia cml patients with bcrabl t315i mutations.
Inhibitors of abl and the ablt315i mutation bentham science. Allogeneic stem cell transplantation for patients with t315i. The result was positive for the t315i gatekeeper mutation. Energies of the ligand, free protein, and the proteinlignad. Pdf prevalence of bcrabl t315i mutation in malaysian.
Pdf objective chronic myeloid leukemia cml is caused by a reciprocal translocation between. By comparison, the t315i mutations for axitinib, bosutinib, imatinib, nilotinib. In this study, we present the crystal structure of the kinase domain of the cabi t315i mutant, as well as the wildtype form, in complex with a pyrrolopyridine inhibitor, ppya. The t315i is a unique mutation because of its resistance to all approved bcrabl inhibitors, prior to ponatinib. Association of t315i mutation with resistance to tyrosine. T315i mutation in bcrabl causes resistance to therapy with tyrosine kinase inhibitors tkis in philadelphia. The highly conserved threonine 315 residue is called the gatekeeper because it is located near the abl catalytic domain controlling the access to a hydrophobic pocket of the enzymatic active site. The aim of this study was to evaluate the prevalence of this mutation in bcrablpositive cml and all patients. Detection of t315i using digital polymerase chain reaction in. More than 40 kinds of different point mutations have been classified to be related with clinical resistance to imatinib, and especially 20% of all the point mutations are identified to be a stubborn point mutation, namely the t315i point mutation. Several 3rd generation inhibitors such as ap24534, vx680 mk0457, pha 739358, ppya, xl228, sgx70393, fty720 and tg1011 are being developed to target the t315i mutation. Prevalence of bcrabl t315i mutation in malaysian patients with. Request pdf evaluation of t315i mutation frequency in chronic myeloid leukemia patients after imatinib resistance the occurrence of resistance mutations in the abl kinase domain plays a. Overcoming t315i and other tkiresistant mutations in cml alfonso quintascardama, md hematologyoncology fellow division of cancer medicine the university of texas m.
The t315i mutant form of bcrabl lacks a threonine residue, which provides a. A blood sample was sent to the hemostasis and thrombosis laboratory at jordan university hospital to look for bcrabl kd mutations. The bcrablt315i mutation confers drug resistance to. We report design and preclinical evaluation of ap24534, a potent, orally available multitargeted kinase inhibitor active against t315i and other bcrabl mutants. Pdf antileukemic activity of axitinib against cells. Jun 30, 2017 certain genetic changes in cml and all can make it more difficult to treat. A singletube allele specificpolymerase chain reaction aspcr method was developed to detect t315i mutation in a mixture of normal and mutant alleles of varying dilutions. The abl kinase domain contains a t315i mutation that predicts highlevel resistance to imatinib, dasatinib, nilotonib, and bosutinib.
Thus, free energy decreases more in case of binding during dfgin phase, which is therefore preferentially inhibited by dasatinib, because it is thermodynamically favored. However, axitinib, a vascular endothelial growth factor receptor inhibitor, is effective against this mutation. Mutation t315i chronic myeloid leukemia cml chronic. Mutation testing is not recommended in newly diagnosed chronic phase patients, during routine monitoring, or when there is no evidence of tki resistance.
It is evident that there is substantial increase of t315i mutation and reduction of m244v mutation in all cml profile, suggesting t315i is more favored and m244v is less favo red towards the advanced phases. Noticeably, m351t, instead of t315i, mutation is the most frequent mutation in chroni c cml. Although the t315i mutation does not disturb the overall structure of the bcrabl protein, it affects the topology of the atp binding region. Characteristics and outcomes of patients with chronic. Clinical features and outcomes in chronic myeloid leukemia. Therapeutic options against bcrabl1 t315ipositive chronic. Ponatinib, a third generation tki, is the drug of choice in such cases. Inductions of bcrabl mutations by imatinib treatment journal of. The availability of a simple, sensitive and quick assay, allowing a rapid detection of the t315i mutation is therefore crucial, as the detection of this mutation represents an important element in clinical decision for cml patients. The kit provides all necessary reagents, optimized for rapid and sensitive detection of a low percentage of mutant dna in a background of wildtype genomic dna. Chronic myeloid leukaemia with extreme thrombocytosis and. We show here the first crystal structure of the kinase domain of abl t315i in complex with pha739358, an aurora kinase inhibitor currently in clinical development for solid and hematologic malignancies. Bad prognosis and resistance to tkis in such cases boosted development of novel drugs. On the other hand, the above results indicated no significant relationship between t315i mutation, duration of response to treatment, ccyr to treatment, blood parameters and patients clinical signs.
Ponatinib is the only known tki which showed clinical efficiency in t315i positive cml patients. T315i bcrabl mutation analysis quantitative indication. Iclusig ponatinib, previously known as ap24534 is an orally active multityrosine kinase inhibitor and is currently approved by the us food and drug administration for patients with chronic myeloid leukemia and philadelphia chromosomepositive acute lymphoblastic leukemia, specifically targeting the bcrabl gene mutation, t315i. Moreover, mln4924 inhibited the survival and selfrenewal of primary human cml cd34. Pdf on may 1, 2010, michele cea and others published a t315i mutation in e19a2 bcrabl1 chronic myeloid leukemia responding to dasatinib find, read and cite all the research you need on. All of the bcrabl mutants are inhibited by the 2nd generation inhibitors with the exception of the t315i mutant. Some of these mutations have clinical significance, but none as much as ploop and t315i mutations. The chromosomal defect in the philadelphia chromosome is a reciprocal translocation, in which parts of two chromosomes, 9 and 22, swap places. Significance of ploop mutations in cml 89 imatinib resistant pts in 5 french centers. Discovery of asciminib abl001, an allosteric inhibitor. T315i mutation in phpositive acute lymphoblastic leukemia is. Incidence of t315i mutation in bcrablpositive cml and. Short report open access pharmacokinetics of dasatinib. Pdf chronic myelogenous leukemia cml is a hematological stem cell disorder caused by increased.
T315i mutation of bcrabl1 into human philadelphia chromosomepositive leukemia cell lines by homologous recombination using the crisprcas9 system. The majority of relapses 3554 occurred in the first 3 months, and only one patient relapsed after 7 months. The bcrablt315i mutation confers drug resistance to fdaapproved targeted therapeutics imatinib mesylate, dasatinib, and nilotinib. This compound inhibits in vitro the kinase activity of wildtype abl and of several mutants, including t315i. A kbm5 t315i and 32d t315i cells were treated with indicated concentration of aki603 for 96 h. The results obtained from the application of the molecular mechanicspoissonboltzmann surface area method to the evaluation of the binding free energy of wt and t315i mutant ablcore domains with imatinib led us to the conclusion that the analyzed mutation can be claimed to be one of the molecular causes of resistance when identified in patients. This mutation has been reported to confer a particularly poor prognosis compared to other bcrabl1 mutations. A singletube allele specificpolymerase chain reaction to. Abl kinase domain kd mutations remains the most documented one in vivo, identified in at least 50% of tyrosine kinase inhibitors tkis failures and disease progression. A costeffectiveness method for detection of abl mutations in. Design, syntheses and sar of inhibitors targeting the. T315imutated bcrabl in chronic myeloid leukemia and.
Panel a and b show t315i positive cells from a patient with 60 % of mutated cells positive cells, panel c and d show t315i positive cells from a patient with 45 % of mutated cells. The role of cytokines in governing the expansion of the. The result is that a fusion gene is created by juxtaposing the abl1 gene on chromosome 9 region q34 to a part of the bcr breakpoint cluster region gene on chromosome 22 region q11. Our patient achieved a major cytogenetic response and major. This article cites 19 articles, 9 of which you can access for free at. Optimizing the selection of kinase inhibitors for chronic myeloid leukemia patients. Pdf the bcrabl mutations t315i and y253h do not confer. A total of 38 18% patients harbored the t315i mutation in this period. The most important mutations are the ploop mutations and the t315i mutation. Characteristics and outcomes of patients with chronic myeloid. Pdf prevalence of bcrabl t315i mutation in malaysian patients. Nov 29, 2014 however, t315i mutation also was observed in 10 cases post im failure 4 cp, 5 ap, and 1 blast phase and 2 cases post nilotinib treatment in the study by cortes et al.
The philadelphia chromosome or philadelphia translocation ph is a specific genetic abnormality in chromosome 22 of leukemia cancer cells particularly chronic myeloid leukemia cml cells. Overcoming t315i and other tkiresistant mutations in cml. Pdf inhibitors of abl and the ablt315i mutation glenn. Design, syntheses and sar of inhibitors targeting the t315i abl mutation jianguo cao1, kathy barrett2, richard m. The prevalence of the t315i mutation was found to be 7% 460. Despite three approved therapeutic options, the crossresistant bcrabl t315i mutation and compound mutants selected on sequential inhibitor therapy remain major clinical challenges. Mutations on other sites of the kinase have also been reported, for example on the chelix, sh2 domain, substrate binding site, activation loop and cterminal lobe.
Denaturing high performance liquid chromatography dhplc and direct sequencing were performed as a comparison to aspcr. Pcrbased pyrosequencing of rna is performed to examine mutational status of codons 315 of the bcrabl fusion transcript. In this study, we investigated axitinib activity against ponatinibresistant cells and found that axitinib inhibited cellular growth and apoptosis in baf3 t315i mutant. Mutations identified by massively parallel sequencing test to consider bcrabl1 mutation analysis for tyrosine kinase inhibitor resistance by next generation sequencing 2008420 order only for patients with an established diagnosis of a bcrabl1positive leukemia use to determine if a mutation is present that would. Therefore, inhibitors that target the t315i mutation are needed to address this.
Mutationspecific control of bcrabl t315i positive leukemia. Targeted kinase inhibitors are a major therapeutic class in the treatment of cancer. Antitumoreffectsofblockingproteinneddylation in t315ibcrabl. The cml patients with t315i mutation have the median of pfs only 11. Furthermore, the t315i mutation induces a conformational change in several amino acid residues, which are important for the binding of imatinib and bcrabl the allosteric effect. T315i is one of the most common acquired mutations in this domain, which occurs in atp binding site and inhibits the formation of hydrogen bond with im. The side chain of ile315 is accommodated in the abi t315i mutant structure without large conformational changes proximal to the site of mutation. Hyperdiploidy associated with t315i mutation in bcrabl. Despite the remarkable success of imatinib against bcrabl, development of secondary resistance, most often due to point mutations in the bcrabl tyrosine kinase tk domain, is quite common.
Cd200 induces apoptosis and inhibits bcrabl signaling in. T315i mutation exerts a dismal prognosis on adult bcrabl1. For monitoring of the levels of the thr to ile mutation at codon 315 t315i of the bcrabl kinase seen in cml patients who have developed resistance to imatinib or other kinase inhibitors. The existence of drug resistance caused by mutations in the breakpoint cluster regionabelson tyrosine kinase bcrabl kinase domain remains a clinical challenge due to limited effective treatment options for chronic myeloid leukemia cml. The role of cytokines in governing the expansion of the t315i mutation in chronic myeloid leukaemia oilin lee the melissa white laboratory cancer theme, south australian heath and medical research institute sahmri. Accordingly, t315i yields imatinib resistance more strongly than other point mutations figure 425. This chromosome is defective and unusually short because of reciprocal translocation, t9. Although the t315i mutation does not disturb the overall structure of the bcrabl protein. The proportion of patients with t315i mutation were higher than that of control. The substitution of the threonine residue at position 315 of the bcrabl1 protein by isoleucine t315i places the bulky isoleucine side chain in the. Cleavage of bcrabl transcripts at the t315i point mutation by. A t315i mutation in e19a2 bcrabl1 chronic myeloid leukemia responding to dasatinib. The t315i rgq pcr kit enables detection of the t315i mutation in the abl gene exon 6 using realtime pcr on the rotorgene q 5plex hrm instrument.
We analyzed 27 patients with t315i, including 20 who developed t315i after imatinib failure representing 11% of 186 patients with imatinib failure, and 7 of 23 who developed new mutations after second tyrosine kinase inhibitor tki. Chronic myeloid leukemia cml with t315i mutation has been reported to have poor prognosis. T315imutated bcrabl in chronic myeloid leukemia and imatinib. The results of the study confirmed that this method is low cost and easy tool to operate for t315i mutation screening and direct sequencing should be performed in positive cases for. Thus, the usefulness of crisprcas9 system for functional analysis of somatic mutations in cancers was demonstrated. Aki603 and imatinib synergistically inhibit proliferation of cells harboring t315i mutation. Ap24534 is an orally active multitargeted kinase inhibitor that potently inhibits bcrabl t315i.
The bcrabl fusion transcript encodes the bcrabl tyrosine kinase tk, which causes chronic myelogenous leukemia cml. T315i affects a contact residue for most available tyrosine kinase inhibitors tkis, thus patients with chronic myeloid leukemia cml harboring this mutation are resistant to these agents. Detection of bcrabl t315i mutation by peptide nucleic. Among these mutations, the t315i gatekeeper mutation confers resistance to both imatinib, and secondgeneration tkis such as nilotinib and. As was mentioned in the section about targeted therapy, in some patients on tki treatment, the cancer cells develop a gene change called the t315i mutation that keeps most of the tkis from working.
Optimizing kinase inhibitor selection for cml patients. Cml and all patients with the t315i mutation often become resistant to standard treatment. If your cml stops responding to treatment with a tki, another one may be tried. Ponatinib avoids the steric hindrance imposed by the threonine. Prevalence of bcrabl t315i mutation in malaysian patients with imatinibresistant chronic myeloid leukemia. Herein we report a novel series of benzothiazolebased inhibitors that are effective against wildtype and t315i mutant bcrabl kinases. Pdf on jun, 20, ivana horvat and others published dynamics of t315i mutation in philadelphia positive patients on tyrosine kinase inhibitor therapy. Treating t315ipositive chronic myeloid leukemia cml.
Notably, p27kip1 silencing attenuated the suppressive effect of mln4924 on the maintenance of lscs in cml. Pdf a t315i mutation in e19a2 bcrabl1 chronic myeloid. The bcrabl mutations t315i and y253h do not confer a growth advantage in the absence of imatinib. Evaluation of t315i mutation frequency in chronic myeloid. All four patients with mutation were in advance phases and had previously lost all their responses. Therefore, detection of t315i mutation in cml patients are clinically useful in the selection of appropriate. For monitoring of the levels of the thr to ile mutation at codon 315 t315i of the bcrabl kinase seen in cml patients who have developed resistance to imatinib or other kinase inhibitors methodology. Predicting resistance of clinical abl mutations to. Bcrabl t315i, a mutant resistant to all approved abl kinase inhibitors, is emerging as a common pathway of failure on all three inhibitors. Several published studies have demonstrated the effectiveness of sct for cml patients with t315i mutation 4,11,12. Association of t315i mutation with resistance to tyrosine kinase. Consequently, several conformationaltering mutations, except for t315i, are susceptible to dasatinib action, anyway.
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