Initial Identification of a Blood-Based Chromosome Conformation Signature for Aiding in the Diagnosis of Amyotrophic Lateral Sclerosis.

BACKGROUND: The identification of blood-based biomarkers specific to the diagnosis of amyotrophic lateral sclerosis (ALS) is an active field of academic and clinical research. While inheritance studies have advanced the field, a majority of patients do not have a known genetic link to the disease, making direct sequence-based genetic testing for ALS difficult. The ability to detect biofluid-based epigenetic changes in ALS would expand the relevance of using genomic information for disease diagnosis. METHODS: Assessing differences in chromosomal conformations (i.e. how they are positioned in 3-dimensions) represents one approach for assessing epigenetic changes. In this study, we used an industrial platform, EpiSwitch™, to compare the genomic architecture of healthy and diseased patient samples (blood and tissue) to discover a chromosomal conformation signature (CCS) with diagnostic potential in ALS. A three-step biomarker selection process yielded a distinct CCS for ALS, comprised of conformation changes in eight genomic loci and detectable in blood. FINDINGS: We applied the ALS CCS to determine a diagnosis for 74 unblinded patient samples and subsequently conducted a blinded diagnostic study of 16 samples. Sensitivity and specificity for ALS detection in the 74 unblinded patient samples were 83∙33% (CI 51∙59 to 97∙91%) and 76∙92% (46∙19 to 94∙96%), respectively. In the blinded cohort, sensitivity reached 87∙50% (CI 47∙35 to 99∙68%) and specificity was 75∙0% (34∙91 to 96∙81%). INTERPRETATIONS: The sensitivity and specificity values achieved using the ALS CCS identified and validated in this study provide an indication that the detection of chromosome conformation signatures is a promising approach to disease diagnosis and can potentially augment current strategies for diagnosing ALS. FUND: This research was funded by Oxford BioDynamics and Innovate UK. Work in the Oxford MND Care and Research Centre is supported by grants from the Motor Neurone Disease Association and the Medical Research Council. Additional support was provided by the Northeast ALS Consortium (NEALS).

Complexity of nursing care in acute care hospital patients: results of a pilot study with a newly developed questionnaire.

AIM: The aim of this pilot study was to develop an instrument for measuring complexity of nursing care in hospitalised acute care patients as well as to examine its comprehensibility, its feasibility, the effort required for data collection, and its inter-rater reliability as well as its face validity. METHODS: This pilot study was designed as a descriptive, explorative cross-sectional survey with multiple measurements of the patient-related complexity of nursing care and a supplemental qualitative questionnaire conducted on six units of a Swiss university hospital. The instrument to assess complexity of nursing care was developed on the framework of Perrow and encompasses on three subscales a total of 15 items with a 5-point Likert scale. ETHICAL CONSIDERATIONS: The study was reviewed and approved by the Cantonal Ethics Committee. RESULTS: In total, 866 assessments of complexity of nursing care were carried out on 234 patients. The variability of the results of the six units, from three different specialties, suggests that the sampling was suitable for capturing a wide spectrum of complexity. The results of the three subscales are consistent and the discussion of them with the participating units shows that they are also plausible. The verification of the inter-rater reliability has satisfactory to high intersubjective correlation of the values. There were also a few suggestions for improving comprehensibility as well as on how to support user application. The time expenditure for the assessment between 2 to 5 minutes per patient was accurately. CONCLUSION: With the newly developed questionnaire to measure the complexity of nursing care in acute care hospitals it seems to be possible to assess and to quantify the complexity of nursing care in various acute care hospital settings. Based on the findings and the feedback of the participating users, the questionnaire needs to be improved for large-scale application.

Identification of a Titin-derived HLA-A1-presented peptide as a cross-reactive target for engineered MAGE A3-directed T cells.

MAGE A3, which belongs to the family of cancer-testis antigens, is an attractive target for adoptive therapy given its reactivation in various tumors and limited expression in normal tissues. We developed an affinity-enhanced T cell receptor (TCR) directed to a human leukocyte antigen (HLA)-A*01-restricted MAGE A3 antigen (EVDPIGHLY) for use in adoptive therapy. Extensive preclinical investigations revealed no off-target antigen recognition concerns; nonetheless, administration to patients of T cells expressing the affinity-enhanced MAGE A3 TCR resulted in a serious adverse event (SAE) and fatal toxicity against cardiac tissue. We present a description of the preclinical in vitro functional analysis of the MAGE A3 TCR, which failed to reveal any evidence of off-target activity, and a full analysis of the post-SAE in vitro investigations, which reveal cross-recognition of an off-target peptide. Using an amino acid scanning approach, a peptide from the muscle protein Titin (ESDPIVAQY) was identified as an alternative target for the MAGE A3 TCR and the most likely cause of in vivo toxicity. These results demonstrate that affinity-enhanced TCRs have considerable effector functions in vivo and highlight the potential safety concerns for TCR-engineered T cells. Strategies such as peptide scanning and the use of more complex cell cultures are recommended in preclinical studies to mitigate the risk of off-target toxicity in future clinical investigations.

Inactivating mutations of the histone methyltransferase gene EZH2 in myeloid disorders.

Abnormalities of chromosome 7q are common in myeloid malignancies, but no specific target genes have yet been identified. Here, we describe the finding of homozygous EZH2 mutations in 9 of 12 individuals with 7q acquired uniparental disomy. Screening of a total of 614 individuals with myeloid disorders revealed 49 monoallelic or biallelic EZH2 mutations in 42 individuals; the mutations were found most commonly in those with myelodysplastic/myeloproliferative neoplasms (27 out of 219 individuals, or 12%) and in those with myelofibrosis (4 out of 30 individuals, or 13%). EZH2 encodes the catalytic subunit of the polycomb repressive complex 2 (PRC2), a highly conserved histone H3 lysine 27 (H3K27) methyltransferase that influences stem cell renewal by epigenetic repression of genes involved in cell fate decisions. EZH2 has oncogenic activity, and its overexpression has previously been causally linked to differentiation blocks in epithelial tumors. Notably, the mutations we identified resulted in premature chain termination or direct abrogation of histone methyltransferase activity, suggesting that EZH2 acts as a tumor suppressor for myeloid malignancies.

Transcription factor mutations in myelodysplastic/myeloproliferative neoplasms.

BACKGROUND: Aberrant activation of tyrosine kinases, caused by either mutation or gene fusion, is of major importance for the development of many hematologic malignancies, particularly myeloproliferative neoplasms. We hypothesized that hitherto unrecognized, cytogenetically cryptic tyrosine kinase fusions may be common in non-classical or atypical myeloproliferative neoplasms and related myelodysplastic/myeloproliferative neoplasms. DESIGN AND METHODS: To detect genomic copy number changes associated with such fusions, we performed a systematic search in 68 patients using custom designed, targeted, high-resolution array comparative genomic hybridization. Arrays contained 44,000 oligonucleotide probes that targeted 500 genes including all 90 tyrosine kinases plus downstream tyrosine kinase signaling components, other translocation targets, transcription factors, and other factors known to be important for myelopoiesis. RESULTS: No abnormalities involving tyrosine kinases were detected; however, nine cytogenetically cryptic copy number imbalances were detected in seven patients, including hemizygous deletions of RUNX1 or CEBPA in two cases with atypical chronic myeloid leukemia. Mutation analysis of the remaining alleles revealed non-mutated RUNX1 and a frameshift insertion within CEBPA. A further mutation screen of 187 patients with myelodysplastic/myeloproliferative neoplasms identified RUNX1 mutations in 27 (14%) and CEBPA mutations in seven (4%) patients. Analysis of other transcription factors known to be frequently mutated in acute myeloid leukemia revealed NPM1 mutations in six (3%) and WT1 mutations in two (1%) patients with myelodysplastic/myeloproliferative neoplasms. Univariate analysis indicated that patients with mutations had a shorter overall survival (28 versus 44 months, P=0.019) compared with patients without mutations, with the prognosis for cases with CEBPA, NPM1 or WT1 mutations being particularly poor. CONCLUSIONS: We conclude that mutations of transcription and other nuclear factors are frequent in myelodysplastic/myeloproliferative neoplasms and are generally mutually exclusive. CEBPA, NPM1 or WT1 mutations may be associated with a poor prognosis, an observation that will need to be confirmed by detailed prospective studies.

Fusion of PDGFRB to two distinct loci at 3p21 and a third at 12q13 in imatinib-responsive myeloproliferative neoplasms.

We identified four patients who presented with BCR-ABL1 negative myeloproliferative neoplasms and cytogenetically visible abnormalities of chromosome band 5q31-35. Fluorescence in situ hybridization indicated that the platelet-derived growth factor receptor beta gene (PDGFRB) was disrupted in all four cases and 5' rapid amplification of cDNA ends identified in-frame mRNA fusions between PDGFRB and WDR48 (3p21), GOLGA4 (3p21) and BIN2 (12q13). Strikingly, all three genes encode proteins involving intracellular trafficking. Imatinib, a known inhibitor of PDGFRbeta, selectively blocked the growth of t(3;5) myeloid colonies and produced clinically significant responses in all patients. We conclude that PDGFRB fuses to diverse partner genes in atypical myeloproliferative neoplasms (MPNs). Although very rare, identification of these fusions is critical for proper management of affected individuals.

TFG, a target of chromosome translocations in lymphoma and soft tissue tumors, fuses to GPR128 in healthy individuals.

BACKGROUND: The formation of fusion genes plays roles in both oncogenesis and evolution by facilitating the acquisition of novel functions. Here we describe the first example of a human polymorphic in-frame fusion of two unrelated genes associated with a copy number variant. DESIGN AND METHODS: Array comparative genomic hybridization was used to identify cryptic oncogenic fusion genes. Fusion gene structure and origin was examined using molecular biological and computational methods. Phenotype associations were examined using PopGen cohorts. RESULTS: Targeted array comparative genomic hybridization to identify cryptic oncogenic fusion genes in patients with atypical myeloproliferative neoplasms identified a 111 kb amplification with breakpoints within the TRK-fused gene (TFG, a target of translocations in lymphoma and thyroid tumors) and G-protein-coupled receptor 128 (GPR128) resulting in an expressed in-frame TFG-GPR128 fusion transcript. The fusion gene was also identified in healthy individuals at a frequency of 0.02 (3/120). Normally both genes are in identical orientations with TFG immediately downstream of GPR128. In individuals with a copy number variant amplification, one or two copies of the TFG-GPR128 fusion are found between the two parental genes. The breakpoints share a region of microhomology, and haplotype and microsatellite analysis indicate a single ancestral origin. Analysis of PopGen cohorts showed no obvious phenotype association. An in silico search of EST databases found no other copy number variant amplification-associated fusion transcripts, suggesting that this is an uncommon event. Conclusions The finding of a polymorphic gene fusion in healthy individuals adds another layer to the complexity of human genome variation and emphasizes the importance of careful discrimination of oncogenic changes found in tumor samples from non-pathogenic normal variation.

Frequent CBL mutations associated with 11q acquired uniparental disomy in myeloproliferative neoplasms.

Recent evidence has demonstrated that acquired uniparental disomy (aUPD) is a novel mechanism by which pathogenetic mutations in cancer may be reduced to homozygosity. To help identify novel mutations in myeloproliferative neoplasms (MPNs), we performed a genome-wide single nucleotide polymorphism (SNP) screen to identify aUPD in 58 patients with atypical chronic myeloid leukemia (aCML; n = 30), JAK2 mutation-negative myelofibrosis (MF; n = 18), or JAK2 mutation-negative polycythemia vera (PV; n = 10). Stretches of homozygous, copy neutral SNP calls greater than 20Mb were seen in 10 (33%) aCML and 1 (6%) MF, but were absent in PV. In total, 7 different chromosomes were involved with 7q and 11q each affected in 10% of aCML cases. CBL mutations were identified in all 3 cases with 11q aUPD and analysis of 574 additional MPNs revealed a total of 27 CBL variants in 26 patients with aCML, myelofibrosis or chronic myelomonocytic leukemia. Most variants were missense substitutions in the RING or linker domains that abrogated CBL ubiquitin ligase activity and conferred a proliferative advantage to 32D cells overexpressing FLT3. We conclude that acquired, transforming CBL mutations are a novel and widespread pathogenetic abnormality in morphologically related, clinically aggressive MPNs.

JAK2 haplotype is a major risk factor for the development of myeloproliferative neoplasms.

Chronic myeloproliferative neoplasms (MPNs) are a group of related conditions characterized by the overproduction of cells from one or more myeloid lineages. More than 95% of cases of polycythemia vera, and roughly half of essential thrombocythemia and primary myelofibrosis acquire a unique somatic 1849G>T JAK2 mutation (encoding V617F) that is believed to be a critical driver of excess proliferation. We report here that JAK2(V617F)-associated disease is strongly associated with a specific constitutional JAK2 haplotype, designated 46/1, in all three disease entities compared to healthy controls (polycythemia vera, n = 192, P = 2.9 x 10(-16); essential thrombocythemia, n = 78, P = 8.2 x 10(-9) and myelofibrosis, n = 41, P = 8.0 x 10(-5)). Furthermore, JAK2(V617F) specifically arises on the 46/1 allele in most cases. The 46/1 JAK2 haplotype thus predisposes to the development of JAK2(V617F)-associated MPNs (OR = 3.7; 95% CI = 3.1-4.3) and provides a model whereby a constitutional genetic factor is associated with an increased risk of acquiring a specific somatic mutation.

The t(1;9)(p34;q34) and t(8;12)(p11;q15) fuse pre-mRNA processing proteins SFPQ (PSF) and CPSF6 to ABL and FGFR1.

We have investigated two patients with acquired chromosomal rearrangements, a male presenting with a t(1;9)(p34;q34) and B cell progenitor acute lymphoid leukemia and a female presenting with a t(8;12)(p11;q15) and the 8p11 myeloproliferative syndrome. We determined that the t(1;9) fused ABL to SFPQ (also known as PSF), a gene mapping to 1p34 that encodes a polypyrimidine tract-binding protein-associated splicing factor. The t(8;12) fused CPSF6, a cleavage and polyadenylation specificity factor, to FGFR1. The fusions were confirmed by amplification of the genomic breakpoints and RT-PCR. The predicted oncogenic products of these fusions, SFPQ-ABL and CPSF6-FGFR1, are in-frame and encode the N-terminal domain of the partner protein and the entire tyrosine kinase domain and C-terminal sequences of ABL and FGFR1. SFPQ interacts with two FGFR1 fusion partners, ZNF198 and CPSF6, that are functionally related to the recurrent PDGFRalpha partner FIP1L1. Our findings thus identify a group of proteins that are important for pre-mRNA processing as fusion partners for tyrosine kinases in hematological malignancies.

A constitutively active SPTBN1-FLT3 fusion in atypical chronic myeloid leukemia is sensitive to tyrosine kinase inhibitors and immunotherapy.

OBJECTIVES: To determine the consequences and significance of an acquired 46XX,t(2;13;2;21)(p13;q12;q33;q11.2) in atypical chronic myeloid leukemia (aCML). METHODS: Translocation breakpoints were identified by fluorescence in situ hybridization and a novel fusion gene identified by rapid amplification of cDNA ends polymerase chain reaction. Functional analysis of the fusion was performed using the Ba/F3 transformation assay and specific inhibition demonstrated using small molecule inhibitors. RESULTS: Fluorescence in situ hybridization indicated that FLT3 at 13q12 was disrupted and 5'-rapid amplification of cDNA ends polymerase chain reaction identified a novel in-frame mRNA fusion between exon 3 of SPTBN1 (spectrin, beta, nonerythrocytic 1) at chromosome 2p16 and exon 13 of FLT3. Expression of SPTBN1-FLT3 transformed Ba/F3 cells to growth factor independence and was accompanied by constitutive phosphorylation of the fusion protein and the downstream substrate extracellular signal-regulated kinase 1/2. The growth of transformed cells was inhibited in a dose-dependent fashion by SU11657, PKC412, and TKI258 (CHIR-258), but not by imatinib. To determine if FLT3 might be involved more widely in BCR-ABL-negative aCML, we analyzed 40 cases and found two were internal tandem duplication-positive, but D835 mutations were not observed. The t(2;13;2;21) patient was initially treated with hydroxyurea and subsequently underwent an unrelated donor bone marrow transplantation. She relapsed cytogenetically at 4 years, but responded to donor lymphocyte infusion, achieving sustained cytogenetic and molecular (nested reverse transcription polymerase chain reaction) remission. CONCLUSION: Although FLT3 abnormalities are uncommon in aCML, SPTBN1-FLT3 is a novel constitutively active tyrosine kinase that appears to responsive to both targeted signal transduction therapy and immunotherapy.

Activity of TKI258 against primary cells and cell lines with FGFR1 fusion genes associated with the 8p11 myeloproliferative syndrome.

The 8p11 myeloproliferative syndrome (EMS) is an aggressive, atypical stem cell myeloproliferative disorder associated with chromosome translocations that disrupt and constitutively activate FGFR1 by fusion to diverse partner genes. To explore the possibility of targeted therapy for EMS, we have investigated the use of TKI258, a multitargeted receptor tyrosine kinase inhibitor with activity against FGFR, VEGFR, PDGFR, FLT3, and KIT that is currently being assessed for the treatment of a variety of malignancies in phase 1 clinical studies. The viability of Ba/F3 cells transformed to IL3 independence by ZNF198-FGFR1 or BCR-FGFR1 was specifically inhibited by TKI258 with IC(50) values of 150 nM and 90 nM, respectively. Inhibition was accompanied by dose-dependent inhibition of phosphorylation of each fusion gene, ERK, and STAT5. TKI258 also specifically inhibited proliferation and survival of the FGFR1OP2-FGFR1-positive KG1 and KG1A cell lines, resulting in increased levels of apoptosis. Primary cells from EMS patients showed significant, dose-dependent responses in liquid culture and in methylcellulose colony assays compared with controls. This work provides evidence that targeted therapy may be beneficial for patients with EMS.

Two novel imatinib-responsive PDGFRA fusion genes in chronic eosinophilic leukaemia.

We identified two patients with a t(2;4)(p24;q12) and a t(4;12)(q2?3;p1?2), respectively, in association with BCR-ABL and FIP1L1-PDGFRA negative chronic eosinophilic leukaemia. Molecular analysis revealed a novel STRN-PDGFRA fusion for the t(2;4) and ETV6-PDGFRA for the t(4;12). The fusions were confirmed by specific amplification of the genomic breakpoints, reverse transcription polymerase chain reaction and fluorescence in situ hybridisation. Both patients were treated with imatinib and, following a rapid haematological response, achieved cytogenetic remission and a major molecular response. In conclusion, PDGFRA fuses to diverse partner genes in myeloid disorders. Identification of these fusions is important as they are particularly sensitive to imatinib.

Haemopoietic growth factors significantly improve the mitotic index and chromosome quality in cytogenetic cultures of myeloid neoplasia.

Haemopoietic growth factors stimulate bone marrow cell division, differentiation, and survival in vivo. We have investigated the use of recombinant human haemopoietic growth factors in vitro to improve cytogenetic cultures. Using a combination of granulocyte colony stimulating factor, granulocyte macrophage colony stimulating factor, stem cell factor, and interleukin-3, we developed an additive for bone marrow cultures intended to stimulate myeloid cell growth. Sixty-seven paired parallel cultures were analyzed, of which 50 were abnormal. The growth factor (GF) cultures showed a median four- to five-fold increase in mitotic index (MI) (P < 0.0001). In addition, the chromosome morphology was significantly improved in the GF cultures with a median increase in United Kingdom National External Quality Assessment Scheme quality score of 1.25 points (P < 0.0001). There was no statistically significant difference in the number of abnormal cells between the two culture methods. The combination of higher MI and improved chromosome quality substantially reduces the time required to process a case; furthermore, the GF medium is cheaper than the medium with which it was compared. This method is suitable for both diagnostic and follow-up cytogenetic analysis of acute and chronic myeloid neoplasia and is particularly useful for poorly cellular marrow samples or blood samples that would be expected to fail on standard culture. The use of this method has enabled substantial improvements in work efficiency in our oncology cytogenetic laboratory and reduced average reporting times from 9.0 days (2004/5) to 7.1 days (2005/6), despite a 6% increase in sample numbers.

Durable responses to imatinib in patients with PDGFRB fusion gene-positive and BCR-ABL-negative chronic myeloproliferative disorders.

Fusion genes derived from the platelet-derived growth factor receptor beta (PDGFRB) or alpha (PDGFRA) play an important role in the pathogenesis of BCR-ABL-negative chronic myeloproliferative disorders (CMPDs). These fusion genes encode constitutively activated receptor tyrosine kinases that can be inhibited by imatinib. Twelve patients with BCR-ABL-negative CMPDs and reciprocal translocations involving PDGFRB received imatinib for a median of 47 months (range, 0.1-60 months). Eleven had prompt responses with normalization of peripheral-blood cell counts and disappearance of eosinophilia; 10 had complete resolution of cytogenetic abnormalities and decrease or disappearance of fusion transcripts as measured by reverse transcriptase-polymerase chain reaction (RT-PCR). Updates were sought from 8 further patients previously described in the literature; prompt responses were described in 7 and persist in 6. Our data show that durable hematologic and cytogenetic responses are achieved with imatinib in patients with PDGFRB fusion-positive, BCR-ABL-negative CMPDs.

Der(6)t(1;6)(q21-23;p21.3): a specific cytogenetic abnormality in myelofibrosis with myeloid metaplasia.

Chromosome anomalies are detected in approximately half of patients with myelofibrosis with myeloid metaplasia (MMM) although none of the most prevalent lesions are specific to the disease. In a prospective cytogenetic study of 81 patients with MMM, we encountered three with an unbalanced translocation between chromosomes 1 and 6 with specific breakpoints; der(6)t(1;6)(q21-23;p21.3). A subsequent Mayo Clinic cytogenetic database search identified 12 patients with this chromosome anomaly among 17 791 consecutive patients. A similar database search from Royal Hallamshire Hospital in Sheffield, UK revealed two additional patients among 8000 cases. The clinical phenotype and survival for each of these 14 patients was typical of MMM. These findings suggested that der(6)t(1;6)(q21-23;p21.3) is a highly specific cytogenetic anomaly that may harbour gene(s) specifically associated with MMM. In a preliminary fluorescence in situ hybridization study, the breakpoints on chromosome 6 in two additional cases were found to be telomeric to the gene for 51 kDa FK506-binding protein (FKBP51).

NUP98-LEDGF fusion and t(9;11) in transformed chronic myeloid leukemia.

The molecular basis for disease progression in chronic myeloid leukaemia (CML) is poorly understood, but is believed to be a consequence of additional acquired genetic lesions. We describe here a case of CML who presented de novo in transformation with a t(9;11)(p21;p15) and NUP98-LEDGF fusion in addition to the t(9;22). The t(9;11) was present in only 2/45 (4%) of bone marrow metaphases, but 17/20 (85%) of metaphases from peripheral blood, suggesting an extramedullary or focal origin. This is the first description of NUP98-LEDGF in CML and strengthens the association between disease progression in and NUP98 abnormalities.

Widespread occurrence of the JAK2 V617F mutation in chronic myeloproliferative disorders.

The analysis of rare chromosomal translocations in myeloproliferative disorders has highlighted the importance of aberrant tyrosine kinase signaling in the pathogenesis of these diseases. Here we have investigated samples from 679 patients and controls for the nonreceptor tyrosine kinase JAK2 V617F mutation. Of the 480 myeloproliferative disorder (MPD) samples, the proportion of positive cases per disease subtype was 30 (20%) of 152 for atypical or unclassified MPD, 2 of 134 (2%) for idiopathic hypereosinophilic syndrome, 58 of 72 (81%) for polycythemia vera, 24 of 59 (41%) essential thrombocythemia (ET), and 15 of 35 (43%) for idiopathic myelofibrosis. V617F was not identified in patients with systemic mastocytosis (n = 28), chronic or acute myeloid leukemia (n = 35), secondary erythrocytosis (n = 4), or healthy controls (n = 160). Homozygosity for V617F was seen in 43% of mutant samples and was closely correlated with chromosome 9p uniparental disomy. Homozygosity was significantly less common in ET compared with other MPD subtypes. In 53 cases analyzed, the median level of PRV1 expression was significantly higher in V617F-positive cases compared with cases without the mutation. We conclude that V617F is widespread in MPDs. Detection of this acquired mutation is likely to have a major impact on the way patients with MPD are diagnosed, as well as serving as an obvious target for signal transduction therapy.

p53-Binding protein 1 is fused to the platelet-derived growth factor receptor beta in a patient with a t(5;15)(q33;q22) and an imatinib-responsive eosinophilic myeloproliferative disorder.

We describe the fusion of TP53BP1 to PDGFRB in a patient with a chronic myeloid leukemia-like disorder associated with eosinophilia and a t(5;15)(q33;q22). TP53BP1 encodes 53BP1, a p53-binding protein that plays a role in cellular responses to DNA damage. The 53BP1-PDGFRbeta fusion protein is predicted to retain the kinetochore-binding domain of 53BP1 fused to the transmembrane and intracellular tyrosine kinase domain of PDGFRbeta. The presence of the fusion was confirmed by two-color fluorescence in situ hybridization, reverse transcription-PCR, and by characterizing the genomic breakpoints. The reciprocal fusion, which would contain the p53-binding 53BP1 BRCA1 COOH-terminal domains, was not detectable by fluorescence in situ hybridization or nested PCR. Imatinib, a known inhibitor of PDGFRbeta, blocked the growth of patient colony-forming unit, granulocyte-macrophage in vitro and produced a clinically significant response before relapse and subsequent death with imatinib-resistant disease. We conclude that TP53BP1-PDGFRB is a novel imatinib target in atypical chronic myeloid leukemia.