The mutational oncoprint of recurrent cytogenetic abnormalities in adult patients with de novo acute myeloid leukemia.

Recurrent chromosomal abnormalities and gene mutations detected at the time of diagnosis of acute myeloid leukemia (AML) are associated with particular disease features, treatment response and survival of AML patients, and are used to denote specific disease entities in the World Health Organization classification of myeloid neoplasms and acute leukemia. However, large studies that integrate cytogenetic and comprehensive mutational information are scarce. We created a comprehensive oncoprint of mutations associated with recurrent cytogenetic findings by combining the information on mutational patterns of 80 cancer- and leukemia-associated genes with cytogenetic findings in 1603 adult patients with de novo AML. We show unique differences in the mutational profiles among major cytogenetic subsets, identify novel associations between recurrent cytogenetic abnormalities and both specific gene mutations and gene functional groups, and reveal differences in cytogenetic and mutational features between patients younger than 60 years and those aged 60 years or older. The identified associations between cytogenetic and molecular genetic data may help guide mutation testing in AML, and result in more focused application of targeted therapy in patients with de novo AML.

Maintenance therapy with decitabine in younger adults with acute myeloid leukemia in first remission: a phase 2 Cancer and Leukemia Group B Study (CALGB 10503).

In this prospective phase 2 clinical trial conducted by Cancer and Leukemia Group B (CALGB, now the Alliance), we studied decitabine as maintenance therapy for younger adults with acute myeloid leukemia (AML) who remained in first complete remission (CR1) following intensive induction and consolidation. Given that decitabine is clinically active in AML and with hypomethylating activity distinct from cytotoxic chemotherapy, we hypothesized that 1 year of maintenance therapy would improve disease-free survival (DFS) for AML patients <60 years, who did not receive allogeneic stem cell transplantation in CR1. After blood count recovery from final consolidation, patients received decitabine at 20 mg/m2 intravenously daily for 4-5 days, every 6 weeks for eight cycles. One hundred and thirty-four patients received decitabine and 85 (63%) had favorable risk AML. The median number of cycles received was 7 (range: 1-8) and the primary reason for discontinuation was relapse. DFS at 1 year and 3 years was 79% and 54%, respectively. These results are similar to the outcomes in the historical control comprising similar patients treated on recent CALGB trials. Thus, maintenance with decitabine provided no benefit overall. Standard use of decitabine maintenance in younger AML patients in CR1 is not warranted. This trial was registered at www.clinicaltrials.gov as NCT00416598.

Vascular ring presenting as dysphagia in an adult woman: a case report.

A 48-year-old woman was seen in a surgical outpatient clinic with a 2 year history of progressive dysphagia with occasional regurgitation, partially controlled with a proton pump inhibitor. Primary investigations of pH testing and gastroscopy were normal, although a barium swallow study revealed significant hold-up at the aortic arch impression and a posterior right-sided oesophageal impression suggestive of a right-sided aortic arch. A follow-up computed tomography angiogram discovered a vascular ring encircling the trachea and oesophagus, formed by a right-sided aortic arch with aberrant aortic branches, and a Kommerell's diverticulum. It was deemed that the patient's symptoms were related to this vascular ring. The patient underwent stage-one surgery - an extra-anatomic bypass of the double aortic arch and right subclavian artery - and 4 months later a stent graft insertion over the origin of the diverticulum with the aim of complete symptomatic relief. This case presents a common symptom familiar to any clinician (dysphagia), which has been caused by a rare pathology. It is even more unusual that this should present itself in adulthood.

Mutations in the CCND1 and CCND2 genes are frequent events in adult patients with t(8;21)(q22;q22) acute myeloid leukemia.

Core-binding factor acute myeloid leukemia (CBF-AML) is defined by the presence of either t(8;21)(q22;q22)/RUNX1-RUNX1T1 or inv(16)(p13.1q22)/t(16;16)(p13.1;q22)/CBFB-MYH11. The resulting fusion genes require a 'second hit' to initiate leukemogenesis. Mutation assessment of 177 adults with CBF-AML, including 68 with t(8;21) and 109 with inv(16)/t(16;16), identified not only mutations well known in CBF-AML but also mutations in the CCND1 and CCND2 genes, which represent novel frequent molecular alterations in AML with t(8;21). Altogether, CCND1 (n=2) and CCND2 (n=8) mutations were detected in 10 (15%) patients with t(8;21) in our cohort. A single CCND2 mutation was also found in 1 (0.9%) patient with inv(16). In contrast, CCND1 and CCND2 mutations were detected in only 11 (0.77%) of 1426 non-CBF-AML patients. All CCND2 mutations cluster around the highly conserved amino-acid residue threonine 280 (Thr280). We show that Thr280Ala-mutated CCND2 leads to increased phosphorylation of the retinoblastoma protein, thereby causing significant cell cycle changes and increased proliferation of AML cell lines. The identification of CCND1 and CCND2 mutations as frequent mutational events in t(8;21) AML may provide further justification for cell cycle-directed therapy in this disease.

Comprehensive mutational analysis of primary and relapse acute promyelocytic leukemia.

Acute promyelocytic leukemia (APL) is a subtype of myeloid leukemia characterized by differentiation block at the promyelocyte stage. Besides the presence of chromosomal rearrangement t(15;17), leading to the formation of PML-RARA (promyelocytic leukemia-retinoic acid receptor alpha) fusion, other genetic alterations have also been implicated in APL. Here, we performed comprehensive mutational analysis of primary and relapse APL to identify somatic alterations, which cooperate with PML-RARA in the pathogenesis of APL. We explored the mutational landscape using whole-exome (n=12) and subsequent targeted sequencing of 398 genes in 153 primary and 69 relapse APL. Both primary and relapse APL harbored an average of eight non-silent somatic mutations per exome. We observed recurrent alterations of FLT3, WT1, NRAS and KRAS in the newly diagnosed APL, whereas mutations in other genes commonly mutated in myeloid leukemia were rarely detected. The molecular signature of APL relapse was characterized by emergence of frequent mutations in PML and RARA genes. Our sequencing data also demonstrates incidence of loss-of-function mutations in previously unidentified genes, ARID1B and ARID1A, both of which encode for key components of the SWI/SNF complex. We show that knockdown of ARID1B in APL cell line, NB4, results in large-scale activation of gene expression and reduced in vitro differentiation potential.

Prognostic and biologic significance of DNMT3B expression in older patients with cytogenetically normal primary acute myeloid leukemia.

DNMT3B encodes a DNA methyltransferase implicated in aberrant epigenetic changes contributing to leukemogenesis. We tested whether DNMT3B expression, measured by NanoString nCounter assay, associates with outcome, gene and microRNA expression and DNA methylation profiles in 210 older (⩾60 years) adults with primary, cytogenetically normal acute myeloid leukemia (CN-AML). Patients were dichotomized into high versus low expressers using median cut. Outcomes were assessed in the context of known CN-AML prognosticators. Gene and microRNA expression, and DNA methylation profiles were analyzed using microarrays and MethylCap-sequencing, respectively. High DNMT3B expressers had fewer complete remissions (CR; P=0.002) and shorter disease-free (DFS; P=0.02) and overall (OS; P<0.001) survival. In multivariable analyses, high DNMT3B expression remained an independent predictor of lower CR rates (P=0.04) and shorter DFS (P=0.04) and OS (P=0.001). High DNMT3B expression associated with a gene expression profile comprising 363 genes involved in differentiation, proliferation and survival pathways, but with only four differentially expressed microRNAs (miR-133b, miR-148a, miR-122, miR-409-3p) and no differential DNA methylation regions. We conclude that high DNMT3B expression independently associates with adverse outcome in older CN-AML patients. Gene expression analyses suggest that DNMT3B is involved in the modulation of several genes, although the regulatory mechanisms remain to be investigated to devise therapeutic approaches specific for these patients.

A stem cell-like gene expression signature associates with inferior outcomes and a distinct microRNA expression profile in adults with primary cytogenetically normal acute myeloid leukemia.

Acute myeloid leukemia (AML) is hypothesized to be sustained by self-renewing leukemia stem cells (LSCs). Recently, gene expression signatures (GES) from functionally defined AML LSC populations were reported, and expression of a 'core enriched' (CE) GES, representing 44 genes activated in LCSs, conferred shorter survival in cytogenetically normal (CN) AML. The prognostic impact of the CE GES in the context of other molecular markers, including gene mutations and microRNA (miR) expression alterations, is unknown and its clinical utility is unclear. We studied associations of the CE GES with known molecular prognosticators, miR expression profiles, and outcomes in 364 well-characterized CN-AML patients. A high CE score (CE(high)) associated with FLT3-internal tandem duplication, WT1 and RUNX1 mutations, wild-type CEBPA and TET2, and high ERG, BAALC and miR-155 expression. CE(high) patients had a lower complete remission (CR) rate (P=0.003) and shorter disease-free (DFS, P<0.001) and overall survival (OS, P<0.001) than CE(low) patients. These associations persisted in multivariable analyses adjusting for other prognosticators (CR, P=0.02; DFS, P<0.001; and OS, P<0.001). CE(high) status was accompanied by a characteristic miR expression signature. Fifteen miRs were upregulated in both younger and older CE(high) patients, including miRs relevant for stem cell function. Our results support the clinical relevance of LSCs and improve risk stratification in AML.

Escalation of daunorubicin and addition of etoposide in the ADE regimen in acute myeloid leukemia patients aged 60 years and older: Cancer and Leukemia Group B Study 9720.

Untreated de novo (n=421) and secondary (n=189) acute myeloid leukemia (AML) patients ≥60 years received intensified chemotherapy, including daunorubicin 60 mg/m(2) and etoposide 100 mg/m(2) during days 1, 2, 3 with cytarabine 100 mg/m(2) during days 1-7, with a second induction if needed and one consolidation course with these drugs and doses for 2, 2 and 5 days, respectively. In all, 287 (47%) achieved complete remission (CR), 136 (22%) died and 187 (31%) were non-responders. CR rates were 27, 44 and 52% for complex karyotypes, rare aberrations and neither (P<0.001), 52 and 37% for de novo and secondary AML (P=0.003), and 53 and 42% for age 60-69 and ≥70 years (P=0.015). In multivariable analysis, CR predictors included non-complex/non-rare karyotypes (P<0.001), de novo AML (P<0.001), better performance status (PS) (P<0.001) and younger age (P=0.001). Disease-free (DFS) and overall (OS) survival medians were 6.8 (95% CI: 6.2, 7.8) and 7.2 (95% CI: 6.4, 8.6) months. In multivariable analysis, DFS was shorter for complex karyotypes (P<0.001) and increasing white blood count (WBC) (P<0.001) and age (P=0.038), and OS for complex karyotypes (P<0.001), increasing WBC (P=0.001) and age (P<0.001), poorer PS (P<0.001) and secondary AML (P=0.010). Outcomes and prognostic factors were similar to those in previous Cancer and Leukemia Group B studies.

Receiving mixed signals: uncoupling oligodendrocyte differentiation and myelination.

The development and maturation of an oligodendroglial cell is comprised of three intimately related processes that include proliferation, differentiation, and myelination. Here we review how proliferation and differentiation are controlled by distinct molecular mechanisms and discuss whether differentiation is merely a default of inhibited proliferation. We then address whether differentiation and myelination can be uncoupled in a similar manner. This task is particularly challenging because an oligodendrocyte cannot myelinate without first differentiating, and these processes are therefore not mutually exclusive. Is it solely the presence of the axon that distinguishes a differentiated oligodendrocyte from a myelinating one? Uncoupling these two processes requires identifying specific signals that regulate myelination without affecting the differentiation process. We will review current understanding of the relationship between differentiation and myelination and discuss whether these two processes can truly be uncoupled.

Luteinizing hormone signaling and breast cancer: polymorphisms and age of onset.

Estrogen exposure has repeatedly been shown to associate with the risk of developing breast cancer. Estrogen synthesis is under the control of LH and FSH, where LH, through its receptor (LHR), stimulates production of ovarian androgens; and FSH, their aromatization to estrogens. Here, we investigated whether functional polymorphic variants in the LH signaling pathway are associated with the risk of breast cancer or its clinical phenotype. A PCR-restriction fragment length polymorphism genotyping approach was used to investigate this in 266 breast cancers. The LHR18insLQ allele does not seem to influence breast cancer risk. However, women who were homozygous for the LHR18insLQ allele were, on average, 8.3 yr younger at diagnosis, compared with those homozygous for the wild-type LHR allele (mean age, 51.9 yr vs. 60.2 yr; P = 0.03). Trends were observed for associations between LHR18insLQ carriers and nodal involvement or larger tumor size. Patients who were LHR18insLQ carriers revealed a significantly worse overall survival, compared with those who were homozygous for LHR [hazard ratio = 2.4; 95% CI (1.3-4.3); P = 0.006]. In contrast, no associations between the LH genotype and any of the clinical parameters were observed. Our findings suggest that the LHR18insLQ gene polymorphism determines an earlier age of disease onset and is prognostic for poor outcome of breast cancer.

Efficacy and safety of gemtuzumab ozogamicin in patients with poor-prognosis acute myeloid leukemia.

The objective of this work was to determine the safety and efficacy of gemtuzumab ozogamicin in patients with poor prognosis acute myeloid leukemia (AML). Patients with the following diagnoses/characteristics were treated with 1-3 infusions of gemtuzumab ozogamicin at a dose of 9 mg/m2: (1) relapse of AML < or = 6 months of first complete remission (CR); (2) AML refractory to chemotherapy at initial induction or at first relapse; (3) AML in second or greater relapse; (4) myeloid blast crisis of chronic myeloid leukemia (CML); (5) untreated patients > or = 70 years or > or = 55 years with abnormal cytogenetics (excluding inv 16, t(15;17) and t(8;21)) and/or an antecedent hematologic disorder; (6) refractory anemia with excess blasts in transformation (RAEBT). Forty-three patients, ages 19-84 (mean 62), were treated, including 7 patients with untreated AML age > 70 years, 2 with untreated RAEBT, 14 with AML first salvage (first remission 0-6 months), 15 with AML > or = second salvage and 14 with myeloid blast phase of CML. The overall response rate was 14%, with 4/43 (9%) patients achieving CR and 2/43 (5%) achieving CR without platelet recovery. The most significant toxicity was neutropenic fever, which occurred in 84% of patients. In conclusion, in patients with relapsed/refractory AML, gemtuzumab ozogamicin has a comparable response rate to single-agent chemotherapy and may offer a more favorable toxicity profile.

Chimeric fusion proteins--diphtheria toxin-based.

Most cancer patients receive chemotherapy drugs that target DNA or the cell division apparatus. Many of these patients develop multidrug-resistant tumor cells, thus, novel methods to overcome drug resistance are needed. One approach is to target tumor cell protein synthesis. Peptide toxins, which catalytically inactivate protein synthesis, have been re-engineered to selectively bind and intoxicate tumor cells. Diphtheria toxin (DT), a member of the class of peptide toxins, has been subjected to structural and genetic analysis and protein engineering for several decades. In this review, we will examine the structure, function, synthesis and pharmacology of anticancer DT conjugates.

Postremission therapy in older patients with de novo acute myeloid leukemia: a randomized trial comparing mitoxantrone and intermediate-dose cytarabine with standard-dose cytarabine.

The treatment of older patients with acute myeloid leukemia (AML) remains unsatisfactory, with complete remission (CR) achieved in only approximately 50% and long-term disease-free survival in 10% to 20%. Three hundred eighty-eight patients (60 years of age and older) with newly diagnosed de novo AML were randomly assigned to receive placebo (P) or granulocyte-macrophage colony-stimulating factor (GM-CSF) or GM in a double-blind manner, beginning 1 day after the completion of 3 days of daunorubicin and 7 days of cytarabine therapy. No differences were found in the rates of leukemic regrowth, CR, or infectious complications in either arm. Of 205 patients who achieved CR, 169 were medically well and were randomized to receive cytarabine alone or a combination of cytarabine and mitoxantrone. With a median follow-up of 7.7 years, the median disease-free survival times were 11 months and 10 months for those randomized to cytarabine or cytarabine/mitoxantrone, respectively. Rates of relapse, excluding deaths in CR, were 77% for cytarabine and 82% for cytarabine/mitoxantrone. Induction randomization had no effect on leukemic relapse rate or remission duration in either postremission arm. Because cytarabine/mitoxantrone was more toxic and no more effective than cytarabine, it was concluded that this higher-dose therapy had no benefit in the postremission management of older patients with de novo AML. These results suggest the need to develop novel therapeutic strategies for these patients. (Blood. 2001;98:548-553)

High frequency of immunophenotype changes in acute myeloid leukemia at relapse: implications for residual disease detection (Cancer and Leukemia Group B Study 8361).

Multiparameter flow cytometry (MFC) has the potential to allow for sensitive and specific monitoring of residual disease (RD) in acute myeloid leukemia (AML). The use of MFC for RD monitoring assumes that AML cells identified by their immunophenotype at diagnosis can be detected during remission and at relapse. AML cells from 136 patients were immunophenotyped by MFC at diagnosis and at first relapse using 9 panels of 3 monoclonal antibodies. Immunophenotype changes occurred in 124 patients (91%); they consisted of gains or losses of discrete leukemia cell populations resolved by MFC (42 patients) and gains or losses of antigens on leukemia cell populations present at both time points (108 patients). Antigen expression defining unusual phenotypes changed frequently: CD13, CD33, and CD34, absent at diagnosis in 3, 33, and 47 cases, respectively, were gained at relapse in 2 (67%), 15 (45%), and 17 (36%); CD56, CD19, and CD14, present at diagnosis in 5, 16, and 20 cases, were lost at relapse in 2 (40%), 6 (38%), and 8 (40%). Leukemia cell gates created in pretreatment samples using each 3-antibody panel allowed identification of relapse AML cells in only 68% to 91% of cases, but use of 8 3-antibody panels, which included antibodies to a total of 16 antigens, allowed identification of relapse AML cells in all cases. Thus, the immunophenotype of AML cells is markedly unstable; nevertheless, despite this instability, MFC has the potential to identify RD in AML if multiple antibody panels are used at all time points. (Blood. 2001;97:3574-3580)

Bone marrow cytogenetic abnormalities of aplastic anemia.

Cytogenetic abnormalities in association with aplastic anemia have been reported fairly infrequently. Clonal cytogenetic abnormalities at initial diagnosis are uncommon. A retrospective study was performed of the cytogenetic findings in patients with typical morphological and clinical features of severe aplastic anemia from a single institution for the years 1988 through 1998. A total of 30 cases of aplastic anemia, 16 men and 14 women, were identified. The median age was 60 with females being significantly older (67.5 years) in comparison to males (44 years). Bone marrow specimens failed to yield metaphases in 16 cases and normal karyotypes were detected in 11 cases. Cytogenetic abnormalities were detected in 3 cases. Clonal abnormalities, as defined, occurred in only 2 cases (6.7%). A review of the literature identified a total of 24 cases of aplastic anemia with abnormal cytogenetic findings. Overall, the most common chromosome abnormalities are trisomies of 6 and 8 and loss of chromosome 7. Trisomy 6 is more common at diagnosis while loss of chromosome 7 is more common after therapy.