Imatinib is effective for the treatment of chronic myeloid leukemia (CML). However even undetectable BCR-ABL1 by Q-RT-PCR does not equate to eradication of the disease. Digital-PCR (dPCR), able to detect 1 BCR-ABL1 positive cell out of 10 7 , has been recently developed. The ISAV study is a multicentre trial aimed at validating dPCR to predict relapses after imatinib discontinuation in CML patients with undetectable Q-RT-PCR. CML patients under imatinib therapy since more than 2 years and with undetectable PCR for at least 18 months were eligible. Patients were monitored by standard Q-RT-PCR for 36 months. Patients losing molecular remission (two consecutive positive Q-RT-PCR with at least 1 BCR-ABL1/ABL1 value above 0.1%) resumed imatinib. The study enrolled 112 patients, with a median follow-up of 21.6 months. Fifty-two of the 108 evaluable patients (48.1%), relapsed; 73.1% relapsed in the first 9 months but 14 late relapses were observed between 10 and 22 months. Among the 56 not-relapsed patients, 40 (37.0% of total) regained Q-RT-PCR positivity but never lost MMR. dPCR results showed a significant negative predictive value ratio of 1.115 [95% CI: 1.013-1.227]. An inverse relationship between patients age and risk of relapse was evident: 95% of patients <45 years relapsed versus 42% in the class 45 to <65 years and 33% of patients 65 years [P(v 2 ) < 0.0001]. Relapse rates ranged between 100% (<45 years, dPCR1) and 36% (>45 years, dPCR-). Imatinib can be safely discontinued in the setting of continued PCR negativity; age and dPCR results can predict relapse.

Dasatinib and nilotinib are tyrosine kinase inhibitors (TKIs) developed to overcome imatinib resistance in Philadelphiapositive leukemias. To assess how BcrAbl kinase domain mutation status evolves during sequential therapy with these TKIs and which mutations may further develop and impair their efficacy, we monitored the mutation status of 95 imatinib-resistant patients before and during treatment with dasatinib and/or nilotinib as second or third TKI. We found that 83% of cases of relapse after an initial response are associated with emergence of newly acquired mutations. However, the spectra of mutants conferring resistance to dasatinib or nilotinib are small and nonoverlapping, except for IntroductionResistance to the tyrosine kinase inhibitor (TKI) imatinib mesylate (IM) in chronic myeloid leukemia (CML) and Philadelphiapositive (Ph ϩ ) acute lymphoblastic leukemia (ALL) is often caused by selection of mutations in the Bcr-Abl kinase domain (KD), [1][2][3][4][5][6][7][8][9] altering residues that are directly or indirectly critical for IM binding. To overcome this problem, novel TKIs have been developed. Dasatinib and nilotinib [10][11][12][13] have been the first ones to enter clinical evaluation and to receive marketing approval in IMresistant patients. Preclinical experience with these agents showed that they are active against several IM-resistant Bcr-Abl mutants, with the exception of T315I. 10,11,14 However, both TKIs have been hypothesized to retain their own "Achilles heels." Recent studies have tried to profile mutations that will probably emerge under dasatinib and nilotinib, inducing random mutagenesis and then selecting for cell clones retaining viability when cultured in the presence of the inhibitors. [15][16][17][18] Results suggested that, besides T315I, other mutants might be critical. To assess how Bcr-Abl KD mutation status evolves under the selective pressure of sequential therapy with novel TKIs and which mutations among those predicted by in vitro studies may indeed develop in patients who relapse on dasatinib or nilotinib, we have monitored the mutation status of 95 IM-resistant patients before and during sequential treatment with one or both of these agents. Methods Patients and definitionsThis report focuses on 95 patients (Table 1) who were referred to our laboratory for mutation analysis at the time of IM failure and who received up to 2 subsequent TKIs (dasatinib and/or nilotinib). Thirty-eight patients had chronic-phase (CP) CML, 46 patients had advanced-phase CML (accelerated-phase [AP], n ϭ 11; myeloid blast crisis [BC], n ϭ 18; lymphoid BC, n ϭ 17), and 11 patients had Ph ϩ ALL. For CML patients, IM failure was defined according to European LeukemiaNet recommendations. 19 Similar criteria were applied to define IM failure in Ph ϩ ALL. All 95 patients received a second TKI (dasatinib, n ϭ 55; nilotinib, n ϭ 40). Twenty-six of 95 patients received a third TKI after relapse on second TKI (dasatinib, n ϭ 16; nilotinib, n ϭ 10). For the purpose of this analysis, response to dasat...

Several papers authored by international experts have proposed recommendations on the management of BCR-ABL1+ chronic myeloid leukemia (CML). Following these recommendations, survival of CML patients has become very close to normal. The next, ambitious, step is to bring as many patients as possible into a condition of treatment-free remission (TFR). The Gruppo Italiano Malattie EMatologiche dell’Adulto (GIMEMA; Italian Group for Hematologic Diseases of the Adult) CML Working Party (WP) has developed a project aimed at selecting the treatment policies that may increase the probability of TFR, taking into account 4 variables: the need for TFR, the tyrosine kinase inhibitors (TKIs), the characteristics of leukemia, and the patient. A Delphi-like method was used to reach a consensus among the representatives of 50 centers of the CML WP. A consensus was reached on the assessment of disease risk (EUTOS Long Term Survival [ELTS] score), on the definition of the most appropriate age boundaries for the choice of first-line treatment, on the choice of the TKI for first-line treatment, and on the definition of the responses that do not require a change of the TKI (BCR-ABL1 ≤10% at 3 months, ≤1% at 6 months, ≤0.1% at 12 months, ≤0.01% at 24 months), and of the responses that require a change of the TKI, when the goal is TFR (BCR-ABL1 &gt;10% at 3 and 6 months, &gt;1% at 12 months, and &gt;0.1% at 24 months). These suggestions may help optimize the treatment strategy for TFR.

BACKGROUND:Patients with Philadelphia chromosome-positive (Ph1) acute lymphoblastic leukemia (ALL) frequently relapse on imatinib with acquisition of BCR-ABL kinase domain (KD) mutations. To analyze the changes that second-generation tyrosine kinase inhibitors (TKIs) have brought in mutation frequency and type, a database review was undertaken of the results of all the BCR-ABL KD mutation analyses performed in the authors' laboratory from January 2004 to January 2013. METHODS: Interrogation of the database retrieved 450 mutation analyses in 272 patients with Ph1 ALL. Prescreening of samples was performed with denaturing highperformance liquid chromatography (D-HPLC), followed by direct sequencing of D-HPLC-positive cases. RESULTS: BCR-ABL KD mutations were detected in 70% of imatinib-resistant patients, with T315I, E255K, and Y253H mutations accounting for 75% of cases. Seventy-eight percent of the patients reported to be resistant to second-generation TKIs after imatinib failure were positive for mutations, and 58% of them had multiple mutations. Analysis of patients relapsing on dasatinib revealed a newly acquired T315I mutation in almost two-thirds of the cases. Direct sequencing detected no mutations at diagnosis, even in patients who relapsed after a few months. CONCLUSIONS: Second-generation TKIs ensure a more rapid debulking of the leukemic clone and have much fewer insensitive mutations, but long-term disease control remains a problem, and the T315I mutation is revealed to be an even more frequent enemy. BCR-ABL KD mutation screening of patients with Ph1 ALL who are receiving imatinib or second-generation TKIs would be a precious ally for timely treatment optimization. In contrast, the clinical usefulness of conventional direct sequencing at diagnosis seems to be very low. Cancer 2014;120:1002-9.