bluuebunny/crossref_metadata_2025_split huggingface dataset:
A Phase II Study of Nilotinib A Novel Tyrosine Kinase Inhibitor Administered to Imatinib-Resistant or Intolerant Patients with Chronic Myelogenous Leukemia (CML) in Accelerated Phase (AP).
Authors: Kantarjian Hagop M. , Gattermann Norbert , Hochhaus Andreas , Larson Richard , Rafferty Teresa , Weitzman Aaron , Haque Ariful , Giles Francis , O’Brien S.G. , le Coutre Philipp 
Published: October 2019

Abstract
Nilotinib is a potent, highly selective, aminopyrimidine inhibitor of Bcr-Abl which in vitro is 30-fold more potent than imatinib. It is active against 32/33 imatinib resistant Bcr-Abl mutations. This open-label study was designed to evaluate the safety and efficacy as defined by hematologic/cytogenetic response (HR/CyR) rates of nilotinib at a dose of 400 mg bid in imatinib resistant or intolerant AP patients. Daily doses of nilotinib could be escalated to 600 mg BID for patients who did not adequately respond to treatment, and in the absence of safety concerns. Safety and efficacy data are reported for 64 patients of which 52 (81%) are resistant and 12 (19%) are intolerant to imatinib. More than half (63%) of the patients had CML for ≥ 5 years. The median age was 61 (range 24-79) years and the median time from CML diagnosis and AP diagnosis were 74 (range 2 to 298), and 2 (range 0-106) months, respectively. Of the 64 patients, 17 (27%) had extramedullary disease at baseline. The median duration of nilotinib exposure was 141 (range 2–380) days and the median average dose intensity (mg/days) for all patients, with and without dose escalations, was 797 (range 157 to 1136). Treatment is ongoing for 33 (52%) patients, and 31 (48%) have discontinued (14 for disease progression, 8 for adverse events, 1 each for an abnormal laboratory value, administrative problems, lost to follow up, 4 patients withdrew consent and there were 2 deaths listed as the primary reason for discontinuation). Overall, there were 7 deaths including 4 for disease progression, one related to progressive disease complicated by a cerebral hemorrhage, one cardiac failure and one due to sepsis. Confirmed HR occurred in 28 (44%) patients, of which 11 (17%) were complete, 5 (8%) were marrow responses/no evidence of leukemia, 12 (19%) were return to chronic phase. There were 7 (11%) patients with stable disease/no response, 6 (9%) with disease progression and 21 (33%) patients were not evaluable. Major CyR occurred in 20 (31%) patients, of which 11 (17%) were complete, 9 (14%) were partial, 11 (17%) were minor, and 15 (23%) were minimal.
There were 6 patients (9%) that did not respond. The rate of major CyR for the resistant and intolerant patients was 16 (31%) and 3 (25%), respectively. The majority of Grade 3 or 4 adverse events included thrombocytopenia in 21 (33%), neutropenia in 17 (27%), anemia in 10 (16%) patients, decreased hemoglobin in 4 (6%) patients, and increased lipase in 5 (8%). In summary, these data suggest nilotinib is clinically active and has an acceptable safety and tolerability profile when administered to patients with CML-AP. Updated information will be presented at the meeting.
BH3 Mimetic Augments the Induction of Apoptosis by INNO-406, a Novel Bcr-Abl/Lyn Inhibitor, in Bcr-Abl+ Leukemic Cells.
Authors: Kuroda Junya , Kimura Shinya , Strasser Andreas , Andreeff Michael , Ashihara Eishi , Tabe Yoko , Taniwaki Masafumi , Maekawa Taira 
Published: October 2019

Abstract
Aberrant tyrosine kinase activity of Bcr-Abl fusion oncoprotein by Philadelphia chromosome translocation is the causative as well as the primary molecular target in most of CML and approximately 20% of adult ALL. Though imatinib mesylate has improved the treatment of Bcr-Abl+ leukemia, resistance is often reported in patients with advanced-stage disease. To override imatinib-resistance mechanisms, we have developed INNO-406 (formerly NS-187; Kimura et al, Blood 2005), a novel Bcr-Abl/Lyn inhibitor that is 25~55-folds more potent than imatinib in suppressing Bcr-Abl+ leukemias, and is active against most of imatinib-resistant mutated Bcr-Abl except T315I. Phase I study on INNO-406 started in July 2006. To date, there is no data on the clinical-resistance profile of INNO-406. However, some patients may become resistance after the prolonged administration of INNO-406, as with other Abl kinase inhibitors such as dasatinib and nilotinib. A combination of targeted drugs with different specificities might overcome the problem of resistance (Komarova & Wodarz, PNAS 2005). Thus, we tried to find the best partner for INNO-406 based on the apoptosis-induction mechanisms of each agent. We found that apoptosis induced by INNO-406 was mediated by a Bcl-2 family-regulated intrinsic apoptosis pathway and required the activation of several BH3-only proteins such as Bim, Bad, Bmf or Bik. We investigated the combined effects of 17-AAG or bortezomib, both agents those are known to increase Bim and/or other BH3-only proteins. Neither 17-AAG nor bortezomib showed any additive effects on the accumulation of BH3-only proteins compared with INNO-406 alone. This finding suggested that INNO-406 alone activated BH3-only proteins maximally and combined effects for induction of apoptosis via the activation of BH3-only proteins could not be expected. Thus, we next examined the effects of ABT-737 (produced by Michael Andreeff), which was not an inducer of BH3-only proteins but an inhibitor for anti-apoptotic Bcl-2/Bcl-XL proteins. Intriguingly, ABT-737 augmented significantly the induction of apoptosis by INNO-406 not only in wild-type but also mutated Bcr-Abl+ leukemic cells (Figure). In conclusion, the combined therapy with a Bcr-Abl inhibitor, such as INNO-406, which induces BH3-only proteins strongly and an inhibitor for anti-apoptotic Bcl-2/Bcl-XL proteins such as ABT-737 might be the most rationale apoptosis-based molecular targeting approach against Bcr-Abl+ leukemias.
Figure Figure
Imatinib-Resistant BCR-ABL Mutant Cellular Screening Paradigm To Characterize Promising Small-Molecule Inhibitors: T315I-BCR-ABL Targeted Drug Discovery.
Authors: Azam Mohammad , Shakespeare William C. , Metcalf Chester , Wang Yihan , Sunderamoorthi Raji , Zhu Xiaotan , Huang Wei-Sheng , Zou Dong , Thomas Mathew , Romero Jan , Keats Jeff , Xu Qihang , Dalgarno David , Clackson Tim , Sawyer Tomi K. , Daley George Q. 
Published: October 2019

Abstract
In patients with chronic myeloid leukemia (CML), kinase domain mutations account for imatinib resistance in the majority of cases. Mutations cause either a direct steric hindrance to drug binding or a conformational change that favors kinase activation, which therefore precludes imatinib binding. We have previously characterized the dual Src-Abl kinase inhibitor AP23464 and found it to effectively suppress the growth of cells expressing native and essentially all imatinib-resistant variants of BCR-ABL, with the notable exception of the gatekeeper T315I mutant (Azam et al., Proc. Natl. Acad. Sci. USA, 103: 9244, 2006). Following this work, we have used mutant panel screening and integrated structural analysis to further characterize key analogs designed to overcome T315I resistance, as exemplified by AP23846 and AP24163. Both molecules effectively inhibit the tyrosine kinase activity of wild type (WT) and T315I variants of BCR-ABL, and inhibit the proliferation of BaF3-derived cell lines expressing these enzymes (see Table below). AP24163 was further characterized against a broader panel of imatinib-resistant BCR-ABL-expressing cell lines and showed a promising profile of proliferation inhibition. Comparison of these data with structural models of the mutants provides insights into the basis for the ability of AP24163 to overcome imatinib resistance.
Refinement of small-molecule kinase inhibitors by the integration of sequential screening of panels of mutants coupled with structural analysis is a powerful drug discovery paradigm that is applicable to an increasing number of targeted therapeutic agents.
INHIBITION OF PROLIFERATION OF BAF3 CELLS EXPRESSING BCR-ABL AND ITS VARIANTS (IC50 in nM) IMATINIB AP23464 AP23846 AP24163 WT 600 14 500 7 T315I >20000 >1000 500 480 L248R >20000 92 ND 64 G250E 5000 25 ND 63 Q252H 3000 40 ND 42 Y253H 18000 32 ND 44 E255K 12000 74 ND 24 BAF3+IL3 >20000 >1000 500 >10000 Figure Figure
In Vitro and In Vivo Inhibition of the T315I Mutant BCR/ABL Kinase.
Authors: Zhu Hong , Hanna Ehab , Lohse Dan , Stoughton Silva , Tam Betty , Barrett Kathy , Noronha Glenn , Dneprovskaia Elena , Cao Jon , Chow Chun , Hwang Linda , Mak Chiching , McPherson Andy , Palanki Moorthy , Pathak Ved P. , Renick Joe , Soll Richard , Zeng Binqi , Hood John 
Published: October 2019

Abstract
The T315I point mutation situated at the ATP-binding site of BCR/ABL tyrosine kinase remains to be the major unmet medical need in treating CML patients. None of the marketed CML drugs, including Imatinib or the recently approved Dasatinib, are able to inhibit this clinically most prevalent mutant of BCR/ABL kinase. To overcome the T315I mutation-presented hurdle, a novel small molecule BCR/ABL inhibitor, TG101223, has been synthesized and characterized in vitro and in vivo. Enzymatic activity of T315I mutant kinase and proliferation of BaF3:BCR/ABLT315I cells were inhibited with an IC50 of 50 nM and EC50 of 500 nM respectively. However, little effect of TG101223 was observed on the proliferation of the control BaF3 cells (IC50≥5,000 nM). Consistent with these observations, 24 hour treatment with 2,000 nM TG101223 induced DNA fragmentation in BCR/ABLT315I cells, but not in the control BaF3 cells. Furthermore, caspase-3 activation was detected in BCR/ABLT315I cells by an in-cell caspase assay following 4 h treatment with 2,000 nM TG101223. The above results strongly suggest that TG101223 inhibits BCR/ABLT315I-mediated cell proliferation and survival. In contrast, Imatinib did not inhibit AblT315I enzyme nor BCR/ABLT315I cell proliferation. The kinase selectivity of TG101223 was determined using a commercial enzyme assay with a panel of 49 phylogentically diverse protein kinases. Only Abl, Flt3 and the mutants of those two kinases were inhibited by more than 80% with 500 nM TG101223. Importantly, TG101223 demonstrated a desirable pharmacokinetic profile in mouse (oral bioavailability: 40%, clearance: 23 mL/min/kg and T½: 5 h). The tumorogenicity of BCR/AblT315I cells were confirmed by the observation that tail vein infusion with BCR/ABLT315I cells induced splenomegaly in SCID mice and death by day 14. To assess the in vivo activities of TG101223, mice were dosed orally once with TG101223 on day 13, followed by harvesting of their spleens 7 hours after dosing for Western blot-based protein phosphorylation analysis. A significant induction in phosphorylation of both CrkL and BCR/ABL was observed in the spleens of T315I cell recipient mice, supporting the notion that splenomegaly is a result of the infiltration of proliferating BCR/ABLT315I cells. The above induction in spleen CrkL and BCR/ABL phosphorylation was inhibited by TG101223. These data suggest that TG101223 is a promising candidate for treating the drug resistant CML patients.
Prognostic Significance of Minimal Residual Disease (MRD) in Childhood B-Precursor ALL and Its Relation to Other Risk Factors. A Children’s Oncology Group (COG) Study.
Authors: Borowitz Michael J. , Devidas Meenakshi , Bowman W. Paul , Carroll William L. , Chen I.-Ming , Harvey Richard , Hunger Stephen , Martin Paul L. , Pullen Jeanette , Viswanatha David , Willman Cheryl L. , Winick Naomi , Camitta Bruce 
Published: October 2019

Abstract
Early response to therapy, including assessment of MRD is a powerful prognostic factor in childhood ALL. However, there are many other known prognostic factors, and the precise relationship among these and MRD has been incompletely explored. In COG protocol P9900, patients were assigned to one of 3 risk-directed therapies based on NCI risk group, as well as TEL-AML1 translocations (TEL) and simultaneous trisomies of chromosomes 4 and 10 (DT). Infants and Ph+ patients were excluded. Good risk patients (9904) comprised NCI SR patients with either TEL or DT without CNS disease. Poor risk patients (9906) included a subset of NCI HR patients with particularly unfavorable risk factors based largely on age and white count, while 9905 had both NCI SR and HR patients not eligible for 9904 or 9906, including NCI HR patients with TEL or DT. MRD was satisfactorily measured with at least .01% sensitivity by 4-color flow cytometry in the marrow at end of induction (D29) or in blood at Day 8 (D8). D29 MRD was associated with poor outcome in all studies, and in both HR and SR patients on 9905 as shown:
Study 4 y EFS ± SE (n) p value MRD <.01% MRD >.01% 9904 Overall 94±1%(668) 75±6%(105) <.0001 TEL-AML1 94±2%(367) 68±12%(38) <.0001 DT 95±1%(316) 80±7%(69) <.0001 9905 NCI-Standard Risk 89±2%(462) 63±6%(119) <.0001 9905 NCI-High Risk 83±4%(292) 49±9%(73) <.0001 9906 74±6%(156) 39±8%(84) <.0001
The proportion of D29 MRD positive DT(17.9%) patients was higher than that of TEL patients (9.4%) but MRD was also prognostic when each group was considered separately; similar findings were seen in HR TEL and DT patients. TEL or DT was associated with a significantly better prognosis in both HR and SR patients who were MRD negative, but were not favorable factors in MRD positive patients. D8 blood MRD was also prognostic in all studies, with higher MRD levels generally associated with poorer outcome. Patients who were both D29 and D8 MRD negative had a good outcome overall (93% 4 y EFS); combining all favorable risk factors, including TEL, DT, NCI risk group, D8 and D29 MRD identified a group of patients (about 13% of B-precursor ALL patients) with 98± 1% 4 y EFS. In multivariate analysis, D29 MRD was the most important prognostic factor. NCI Risk group, DT and D8 blood were also significant, but TEL and day 8 morphologic assessment of marrow were not. We conclude that MRD is a major prognostic factor in childhood ALL even accounting for other risk factors. The prognostic significance of TEL and DT depends on MRD status. The combination of good risk clinical and genetic factors, and absence of MRD in both D8 blood and D29 marrow identifies a group of patients with outstanding outcome on limited therapy.
Mutations in the BCR/ABL Kinase Encoding the Resistance to Imatinib Mesylate Do Not Modify the Sensitivity to Genotoxic Treatment.
Authors: Bargaoanu Milene , Skorski Tomasz 
Published: October 2019

Abstract
Imatinib mesylate (IM), a selective inhibitor of ABL kinase activity, revolutionized the treatment of BCR/ABL-positive leukemias. Unfortunately, clinical and experimental observations reveal that resistance to IM is a rising problem, which obscures an otherwise very successful oncogene-targeted therapy. IM resistance can be achieved by point mutations in the kinase domain of BCR/ABL and have been detected in 50–90% of patients with acquired resistance to IM, including ~23% of the IM-naive patients. Strategies to enhance the effect of IM and eventually overcome the resistance are dose escalation, addition of a growth factor, and combinations with novel tyrosine kinase inhibitors like AMN107 and dasatinib, or with inhibitors targeting downstream BCR/ABL effectors, e.g. PI-3k. Unfortunately, resistance to other small molecule inhibitors is likely to appear, as well. Therefore, we tested the sensitivity of leukemia cells expressing IM-resistant BCR/ABL mutants to genotoxic agents. Baf3 cells expressing similar levels of p210BCR/ABL wild-type (WT) and Y253F, Y253H, E255K, E255V, T315I, M351T, and H396P mutants were established. These mutants were selected since they represent several functionally distinct ABL kinase domain regions, including P-loop (Y253F/H and E255K/V), the site of a hydrogen bond with IM (T315I), the activation loop hinge (M351T), and the activation loop (H396P). In addition, they exhibit altered transformation potency, kinase activity, and substrate utilization, irrespective of sensitivity to IM. Western analysis demonstrated similarity, but also differences in the patterns of tyrosine phosphorylated proteins in total cell lysates. As expected, these clones displayed different level of sensitivity to IM: T315I>Y253H>E255V>E255K=Y253F>M251T=H396P>WT. However, leukemia cells expressing the WT and IM-resistant BCR/ABL kinase mutants demonstrated similar sensitivity to cytotoxic drugs such as hydroxyurea (HU), mitomycin C (MMC), and N-Methyl N’-nitro-N-nitrosoguanidine (MNNG). Surprisingly, clones expressing IM-resistant BCR/ABL kinase mutants were more sensitive to γ-irradiation than their WT counterparts. γ-H2AX (histone H2AX phosphorylated on S139) nuclear foci, which serve as marker of DNA double-strand breaks (DSBs), the most lethal DNA lesions, were detected by immunofluorescence. Leukemia cells expressing WT and IM-resistant p210BCR/ABL proteins accumulated similar numbers of DSBs after MMC treatment and γ-irradiation. This observation suggests that leukemia cells expressing IM-resistant p210BCR/ABL mutants may be eliminated by genotoxic treatment, especially γ-irradiation. In addition, IM-resistant BCR/ABL kinase mutants, in contrast to the WT kinase may not be able to modulate the mechanisms protecting from apoptosis induced by γ-irradiation.
Clinical BCR-ABL Peptide Vaccination in Chronic Myeloid Leukaemia: Results of the EPIC Study.
Authors: Rojas Jose M. , Knight Katy , Wang Li-Hui , Clark Richard E. 
Published: October 2019

Abstract
Chronic myeloid leukaemia (CML) is characterised by the BCR-ABL oncoprotein. The amino-acid sequences spanning the junctional region are completely leukaemia-specific. In vitro pulsing of antigen presenting cells elicits immune response to CML cells. Vaccination of CML patients with peptides from this junctional region could therefore elicit/augment immune responses to CML cells. In our Evaluation of Peptide Immunisation in CML (EPIC) study, the patient’s entry requirements were as follows:
first chronic phase of CML, expression of e14a2 (b3a2) BCR-ABL transcript, and prior treatment with imatinib daily (at least 400mg) at a stable dose for at least 6 months.
Each patient received intradermally a cocktail of 3 BCR-ABL peptides:
a 9-mer spanning the e14a2 region, the same 9-mer linked to PADRE (a 15-mer non-natural peptide shown to activate CD4+ T cells), and a 13-mer consensus e14a2 junctional peptide linked to PADRE.
Peptides were administered at either 100 (5 patients), 300 (5 patients), 600 (5 patients), or 1000μg (4 patients) with sargramostim on 6 occasions over 2 months. Immune responses to the vaccine were monitored by IFN-γ and IL-5 ELISPOT assays on peripheral blood mononuclear cells. Molecular responses were assessed by quantitative real-time PCR of BCR-ABL mRNA. At entry no patient showed a detectable immune response to PADRE, but all 19 patients had detectable CD4+ T cells responses within 3 months of commencing vaccination. This indicated that the vaccination protocol was capable of stimulating T cell responses in all 19 patients. Immune responses to the 9-mer BCR-ABL junctional peptide used in the vaccine were detected in 11/19 patients, and demonstrated to be CD8+ T cells by cytokine analysis in flow cytometry. BCR-ABL immune responses were also assessed against a longer 18-mer peptide spanning the whole e14a2 junctional region. CD4+ T cells specific for this 18-mer peptide were detected in 14/19 patients. Interestingly, immunophenotyping indicated that these BCR-ABL-specific T cells were of a memory phenotype (CD45RO+). Serial molecular responses were available for at least 12 months on all cases. Of the 6 patients not in major cytogenic response (MCR) at baseline, molecular improvement was only observed in one case. However 12/13 patients in at least MCR at baseline had at least a 1 log fall in BCR–ABL transcripts, though this occurred several months after completing vaccination. Moreover, vaccination improved the fall in BCR–ABL transcripts in patients who had received imatinib for more than 12 months. These data show that peptide immunisation in CML can elicit anti-BCR-ABL peptide responses in CD4+ and CD8+ T cells. It also demonstrates that BCR-ABL peptide vaccination may improve control of CML, especially in patients responding well to imatinib.
The Immunogenicity of BCR/ABL+ Cells Is Dominated by BCR/ABL-Regulated Antigens and Is Impaired by the Inhibition of BCR/ABL Kinase Activity.
Authors: Scheich Florian , Peschel Christian , Bernhard Helga 
Published: October 2019

Abstract
The inhibition of BCR/ABL kinase activity by imatinib mesylate (IM, STI571, Gleevec®) is the standard therapy for patients with Philadelphia chromosome+ (Ph+) chronic myeloid leukemia (CML). However, the long term treatment with IM or other BCR/ABL kinase inhibitors may be limited due to the development of resistant disease and accumulating side effects. Immunotherapeutical approaches directed against Ph+ CML may overcome these problems. So far, the research to develop an immunotherapy against Ph+ leukemia focused on the BCR-ABL fusion protein, giving the promising opportunity to stimulate cytotoxic T lymphocytes against the joining region segment of p210 BCR/ABL. However, only a limited number of peptides spanning the fusion region is endogenously processed and presented in context with HLA class I molecules. In contrast, the constitutively active BCR/ABL kinase leads to the upregulation and activation of multiple proteins, which may subsequently result in the expression of different, probably leukemia-associated antigens on the cell surface of BCR/ABL-positive malignant cells. In this study, we investigated the immunogenicity of antigens upregulated by BCR-ABL kinase activity, and verified the capacity of these antigens to induce an anti-leukemia T cell response in vitro. We performed CD8+ T-cell stimulations with dendritic cells transfected with mRNA coding BCR/ABL wild-type (WT). Following two stimulations, the proliferating T-cell populations were analyzed for cytokine secretion (IFN-γ) in response to target cells expressing either BCR/ABL WT or a kinase deficient (KD, K1172R) variant of BCR/ABL. With this experimental setting it was possible to compare the immunogenic potential of antigens upregulated by BCR/ABL with the immunogenicity of the BCR/ABL protein itself. We were not able to activate T-cell populations directed against either the breakpoint-region of BCR/ABL or the BCR- or ABL part of the fusion protein. In contrast, we show here that the constitutively active kinase domain of BCR/ABL has a key role in enhancing the immunogenicity of BCR/ABL+ cells. A broad, HLA-dependent T-cell immune response specifically directed against BCR/ABL regulated antigens was detected. The inhibition of BCR/ABL kinase activity by IM significantly impaired the immunogenicity of BCR/ABL+ cells. We found the same T-cell reactivity pattern in several healthy donors and a CML patient. This is the first study demonstrating the major contribution of the BCR/ABL kinase domain to the immunogenicity of BCR/ABL+ cells as T-cell responses against these cells are dominated by BCR/ABL regulated antigens, and not by BCR/ABL itself. These results may contribute to the design of clinical vaccination trials for the treatment CML patients with minimal residual disease, e.g. following successful induction therapy with BCR/ABL inhibitors.