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Background: Treating the octogenarian and nonagenarian patients who have acute myeloid leukemia (AML) with intensive chemotherapy is controversial. Several models to predict outcome were proposed, including the use of a comorbidity index. However, it is unclear whether the Charlson comorbidity index (CCI) or the hematopoietic cell transplant comorbidity index (HCTCI) is more sensitive.Methods: The authors analyzed their experience with 92 patients aged >or=80 years who had AML. Patients' pretreatment characteristics and their treatment outcomes were recorded.Results: All patients were offered intensive treatment; 59 patients (64%) were treated intensively with a variety of regimens, whereas 33 patients (36%) elected to receive supportive care. The CCI and the HCTCI had similar predictive ability for outcome in both groups. A multivariate analyses of prognostic factors identified near-normal albumin (48% of patients; 1-year survival rate, >27%) as a favorable factor for the whole cohort, age <83 years (47% of patients; 1-year survival rate, >25%) and nonmonocytic morphology (75% of patients; 1-year survival rate, >26%) as favorable factors for the intensively treated cohort, and bone marrow blasts <46% (50% of patients; 1-year survival rate, >19%) as a favorable factor for patients who received supportive care.Conclusions: This retrospective analysis was developed to assist in treatment decisions for octogenarian and nonagenarian patients with AML. The findings will need validation in a prospective study. [ABSTRACT FROM AUTHOR]

Patients with synchronous malignancies can be problematic to diagnose and manage because workup and therapeutic targeting for each individual malignancy must be coordinated carefully. This report presents a patient with concurrent chronic myeloid leukemia (CML) and chronic lymphocytic leukemia (CLL) managed with concomitant venetoclax and imatinib. Because imatinib is a moderate cytochrome P450 3A4 inhibitor, close monitoring is required when using with a substrate of 3A4 such as venetoclax. Although the target dose of venetoclax is 400 mg, it was capped at 100 mg due to the interaction. Despite the interaction and possible enhancement of toxicities, the patient has tolerated therapy well, and both diseases have responded to this novel approach. In addition, because aberrant BCL-2 activity has been implicated in CML, the use of venetoclax may contribute to success in the management of this patient's CML. This case report represents the safe concomitant use of venetoclax and imatinib in a patient with synchronous CML and CLL.

The use of immunotherapy to treat patients with myelodysplastic syndromes (MDS) shows promise but is limited by our incomplete understanding of the immunologic milieu. In solid tumors, CD141 Hi conventional dendritic cells (CD141 Hi cDCs) are necessary for antitumor immunosurveillance and the response to immunotherapy. Here, we found that CD141 Hi cDCs are reduced in MDS bone marrow and based on the premise established in solid tumors, we hypothesized that reduced numbers of CD141 Hi cDCs are associated with inferior overall survival in MDS patients. We found that MDS patients with reduced numbers of CD141 Hi cDCs, but not other DC populations, showed reduced overall survival. To examine the basis for reduction in CD141 Hi cDCs, we found fewer numbers of progenitors committed to DC differentiation in the MDS bone marrow and these progenitors expressed lower levels of interferon regulatory factor-8 (IRF8), a master regulator of CD141 Hi cDC differentiation. To rescue impaired CD141 Hi cDC differentiation, we used pharmacologic inhibition of lysine-specific demethylase 1A (LSD1) to promote CD141 Hi cDC differentiation by MDS progenitors. These data reveal a previously unrecognized element of the MDS immunologic milieu. Epigenetic regulation of CD141 Hi cDC differentiation offers an intriguing opportunity for intervention and a potential adjunct to immunotherapy for patients with MDS.

In contrast to FLT3 ITD mutations, in-frame deletions in the FLT3 gene have rarely been described in adult acute leukemia. We report two cases of AML with uncommon in-frame mutations in the juxtamembrane domain of the FLT3 gene: a 3-bp (c.1770&#95;1774delCTACGinsGT; p.F590&#95;V592delinsLF) deletion/insertion and a 12-bp (c.1780&#95;1791delTTCAGAGAATAT; p.F594&#95;Y597del) deletion. We verified by sequencing that the reading frame of the FLT3 gene was preserved and by cDNA analysis that the mRNA of the mutant allele was expressed in both cases. Given the recent development of FLT3 inhibitors, our findings may be of therapeutic value for AML patients harboring similar FLT3 mutations.

Background: Several studies have suggested that low 25(OH) vitamin D3 levels may be prognostic in some malignancies, but no studies have evaluated their impact on treatment outcome in patients with acute myeloid leukemia (AML)., Methods: Vitamin D levels were evaluated in 97 consecutive, newly diagnosed, intensively treated patients with AML. MicroRNA expression profiles and single nucleotide polymorphisms (SNPs) in the 25(OH) vitamin D3 pathway genes were evaluated and correlated with 25(OH) vitamin D3 levels and treatment outcome., Results: Thirty-four patients (35%) had normal 25(OH) vitamin D3 levels (32-100 ng/mL), 34 patients (35%) had insufficient levels (20-31.9 ng/mL), and 29 patients (30%) had deficient levels (<20 ng/mL). Insufficient/deficient 25(OH) vitamin D3 levels were associated with worse relapse-free survival (RFS) compared with normal vitamin D3 levels. In multivariate analyses, deficient 25(OH) vitamin D3 , smoking, European Leukemia Network genetic group, and white blood cell count retained their statistical significance for RFS. Several microRNAs and SNPs were associated with 25(OH) vitamin D3 levels, although none remained significant after multiple test corrections; one 25(OH) vitamin D3 receptor SNP, rs10783219, was associated with a lower complete remission rate (P = .0442) and with shorter RFS (P = .0058) and overall survival (P = .0011)., Conclusions: It remains to be determined what role microRNA and SNP profiles play in contributing to low 25(OH) vitamin D3 level and/or outcome and whether supplementation will improve outcomes for patients with AML., (© 2013 American Cancer Society.)

We designed a phase II study evaluating the upfront combination of clofarabine and daunorubicin in acute myeloid leukemia (AML) patients≥60 years old. The median age of the 21 patients was 69 (range 60-85) years. Fourteen patients (67%) had unfavorable risk features. The principal toxicities were grade ≥3 infections and prolonged myelosuppression. Three (14%) deaths occurred from infectious complications. Six (28.6%) patients achieved complete remission including three (21.4%) of 14 patients with unfavorable AML. The median disease-free survival was 6.8 months and the median overall survival was 11.2 months., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Signal transducer and activator of transcription (STAT) 3 inhibits dendritic cell (DC) differentiation and is constitutively activated in blasts of approximately half of AML patients. We investigated the correlation between STAT3 activity, DC maturation and the ability to stimulate T-cells in primary acute myeloid leukemia (AML)-derived DCs. STAT3 knock-down by shRNAmir increased the ability of AML-DCs to stimulate T-cells. Treatment of AML-DC with arsenic trioxide, but not AG490, JSI-124 or NSC-74859, led to a more mature phenotype and enhanced T-cell stimulation, while having minimal effect on normal DC. We conclude that AML-DCs have improved immunogenicity after reducing STAT3., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Aberrant expression of the B lymphoid marker, CD19, in acute myeloid leukemia (AML) has frequently been associated with t(8;21)(q22;q22). However, AML cases lacking t(8;21) may occasionally express CD19. We asked whether CD19 expression is restricted to the karyotypically abnormal leukemic cells in primary leukemia samples. We compared, by fluorescence in situ hybridization, CD19-positive and CD19-negative cells from nine patients with acute leukemia: three non-t(8;21) AML, three t(8;21) AML and three cases of acute lymphoblastic leukemia. There were no significant differences in karyotypic pattern between the CD19-positive and CD19-negative leukemic cells, raising the concern that therapeutically targeting CD19 for acute leukemia may not eradicate all malignant clones.

Understanding the biology of Waldenström macroglobulinemia is hindered by a lack of preclinical models. We report a novel cell line, RPCI-WM1, from a patient treated for WM. The cell line secretes human immunoglobulin M (h-IgM) with κ-light chain restriction identical to the primary tumor. The cell line has a modal chromosomal number of 46 and harbors chromosomal changes such as deletion of 6q21, monoallelic deletion of 9p21 (CDKN2A), 13q14 (RB1) and 18q21 (BCL-2), with a consistent amplification of 14q32 (immunoglobulin heavy chain; IgH) identical to its founding tumor sample. The clonal relationship is confirmed by identical CDR3 length and single nucleotide polymorphisms as well as a matching IgH sequence of the cell line and founding tumor. Both also harbor a heterozygous, non-synonymous mutation at amino acid 265 in the MYD88 gene (L265P). The cell line expresses most of the cell surface markers present on the parent cells. Overall, RPCI-WM1 represents a valuable model to study Waldenström macroglobulinemia.

Cytogenetic abnormalities are important diagnostic and prognostic criteria for hematologic malignancies. Karyotyping and fluorescence in situ hybridization (FISH) are the conventional methods by which these abnormalities are detected. The sensitivity of these microscopy-based methods is limited by the abundance of the abnormal cells in the samples and therefore these analyses are commonly not applicable to minimal residual disease (MRD) stages. A flow cytometry-based imaging approach was developed to detect chromosomal abnormalities following FISH in suspension (FISH-IS), which enables the automated analysis of several log-magnitude higher number of cells compared with the microscopy-based approaches. This study demonstrates the applicability of FISH-IS for detecting numerical chromosome aberrations, establishes accuracy, and sensitivity of detection compared with conventional FISH, and feasibility to study procured clinical samples of acute myeloid leukemia (AML). Male and female healthy donor peripheral blood mononuclear cells hybridized with combinations of chromosome enumeration probes (CEP) 8, X, and Y served as models for disomy, monosomy, and trisomy. The sensitivity of detection of monosomies and trisomies amongst 20,000 analyzed cells was determined to be 1% with a high level of precision. A high correlation (R(2) = 0.99) with conventional FISH analysis was found based on the parallel analysis of diagnostic samples procured from 10 AML patients with trisomy 8 (+8). Additionally, FISH-IS analysis of samples procured at the time of clinical remission demonstrated the presence of residual +8 cells indicating that this approach may be used to detect MRD and associated chromosomal defects., (Copyright © 2012 International Society for Advancement of Cytometry.)

Smoking adversely affects hematopoietic stem cell transplantation outcome. We asked whether smoking affected outcome of newly diagnosed acute myeloid leukemia (AML) patients treated with chemotherapy. Data were collected on 280 AML patients treated with high-dose cytarabine and idarubicin-containing regimens at Roswell Park Cancer Institute who had smoking status data at diagnosis. Patients' gender, age, AML presentation (de novo vs. secondary), white blood cell (WBC) count at diagnosis, karyotype and smoking status (never vs. ever) were analyzed. Among the 161 males and 119 females with a median follow-up of 12.9 months, 101 (36.1%) had never smoked and 179 (63.9%) were ever smokers. The proportion of patients between never and ever smokers was similar to respect to age, AML presentation, WBC count at diagnosis or karyotype based on univariate analysis of these categorical variables. Never smokers had a significantly longer overall survival (OS) (60.32 months) compared to ever smokers (30.89; p = 0.005). In multivariate analysis incorporating gender, age, AML presentation, WBC count, karyotype and smoking status as covariates, age, karyotype and smoking status retained prognostic value for OS. In summary, cigarette smoking has a deleterious effect on OS in AML., (Copyright © 2011 UICC.)

Obesity adversely affects outcome in pediatric acute lymphocytic leukemia and acute myeloid leukemia (AML). We asked if obesity, measured by body mass index (BMI), affected outcome in 329 adult AML patients treated with high-dose cytarabine and idarubicin-containing regimens administered according to actual body weight. Age ≥ 60, unfavorable karyotype, secondary AML, and positive smoking status had adverse impact on overall survival in a multivariate analysis, while BMI did not. We conclude that high BMI should not be a barrier to administer high-dose cytarabine-containing regimens for AML induction.

Background: Constitutive activation of signal transducer and activator of transcription-3 (STAT3) was detected in blasts from approximately 50% of patients with acute myeloid leukemia (AML) and was correlated with an adverse outcome. In vitro treatment of AML blasts with arsenic trioxide (ATO) down-regulated STAT3 activity within 6 hours associated with a reduced viability within 48 hours., Methods: A phase 1 clinical trial to evaluate the biologically effective dose and/or the maximally tolerated dose (MTD) of ATO in vivo in conjunction with high-dose cytarabine (Hidac) and idarubicin (Ida) in patients with AML aged <60 years was conducted. Data were compared with 117 historic AML patients who had received treatment with Hidac/Ida., Results: In total, 61 patients were enrolled onto 11 different dose levels (from 0.01 to 0.65 mg/kg ideal body weight). The MTD was 0.5 mg/kg. Compared with historic controls, patients who received ATO/Hidac/Ida, although they had similar pretreatment characteristics, had better overall survival (P = .039)., Conclusions: ATO priming may have improved the outcome of patients aged <60 years with AML who received Hidac/Ida. The current data suggested that ATO may enhance the effect of chemotherapy. The authors concluded that further studies of this novel combination are warranted., (Copyright © 2011 American Cancer Society.)

We examined the predictive impact of HIF-1α protein expression on clinical outcome of 84 normal karyotype acute myeloid leukemia (NK-AML) patients (median age 66.5 years) at our institute. Thirty percent of NK-AML cells expressed cytoplasmic HIF-1α. In univariate analysis, low HIF-1α (≤ 5%, n = 66) was associated with improved event-free survival (p = 0.0453, HR = 0.22). Multivariate analysis incorporating age, complete remission, FLT3-ITD mutation, and marrow blast percentage demonstrated that HIF-1α was independently associated with poorer overall and event-free survival. HIF-1α expression correlated with VEGF-C but not VEGF-A, marrow angiogenesis, FLT3 ITD or NPM1 mutations. These results support HIF-1α as an outcome marker for NK-AML., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Blastic transformation of myeloproliferative neoplasms (MPN) is still poorly understood. We describe a cohort of 23 Roswell Park Cancer Institute (RPCI) patients and 89 additional cases from the English literature for whom biologic features were described. We initially compared our 23 patients to the 89 cases from the literature. Our population had significantly less patients with prior history of polycythemia vera (PV), shorter time from MPN diagnosis to blastic transformation, <3 prior therapies, more frequent use of hydroxyurea and erythropoietin and less frequent use of alkylating agents. Interestingly, the overall survival of the two cohorts from the time of blastic transformation was similar. We therefore looked at the outcome of the entire cohort (n=112). Patients with prior history of essential thrombocythemia survived longer than patients with prior history of myelofibrosis or PV. Further, patients with <3 prior therapies, those who lacked complex karyotype and those <60 year old at MPN diagnosis had significantly longer survival. Among the PRCI population, 20/23 patients underwent induction treatment with cytarabine and an anthracycline containing regimens; 12 achieved remission and their overall survival was significantly longer than those who did not. Three patients underwent an allogeneic transplantation and their survival was significantly longer than those who did not. Patients with <3 prior therapies, those who lack complex karyotype and those <60 at MPN diagnosis have longer survival following blastic transformation. Finally, allogeneic transplantation represents the only chance for long-term survival in these patients., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Normal karyotype (NK) is the most common cytogenetic group in acute myeloid leukemia (AML) diagnosis; however, up to 50% of these patients at relapse will have aberrant karyotype (AK) AML. To determine the etiology of relapsed AK AML cells, we evaluated cytogenetic, immunophenotypic, and molecular results of 17 patients with diagnostic NK AML and relapsed AK AML at our institute. AK AML karyotype was diverse, involving no favorable and largely (8 of 17) complex cytogenetics. Despite clear cytogenetic differences, immunophenotype and NPM1/FLT3 gene mutation status did not change between presentation and relapse in 83% (10 of 12) and 94% (15 of 16) cases, respectively. High-resolution array-based comparative genomic hybridization (aCGH) performed via paired aCGH on NK AML and AK AML samples from the same patient confirmed cytogenetic aberrations only in the relapse sample. Analysis of 16 additional diagnostic NK AML samples revealed no evidence of submicroscopic aberrations undetected by conventional cytogenetics in any case. These results favor evolution of NK AML leukemia cells with acquisition of novel genetic changes as the most common etiology of AK AML relapse as opposed to secondary leukemogenesis. Additional studies are needed to confirm whether AK AML cells represent selection of rare preexisting clones below aCGH detection and to further characterize the molecular lesions found at time of AK AML relapse., (Copyright © 2010 Elsevier Inc. All rights reserved.)

The prognosis of adult normal karyotype (NK) precursor B-cell acute lymphoblastic leukemia (B-ALL) has not improved over the last decade, mainly because separation into distinct molecular subsets has been lacking and no targeted treatments are available. We screened the genome of blasts from 10 adult NK B-ALL patients for novel genomic alterations by array comparative genomic hybridization and verified our results with fluorescent in situ hybridization and gene expression profile with the same probes. The results demonstrate cryptic deletions of 9q34 involving SET, PKN3, NUP188, ABL1, and NUP214 in three of the samples. The smallest deletion resulted in the likely juxtaposition of the SET and NUP214 genes. This aberration has not been described before in adult NK B-ALL. Larger number of samples is warranted to determine the prognostic significance of this cryptic deletion., (2010 Elsevier Inc. All rights reserved.)

Smoking is associated with both acute myeloid leukemia (AML) and lung cancer. We therefore searched our database for concomitant presentation of AML and lung cancer. Among 775 AML cases and 5225 lung cancer cases presenting to Roswell Park Cancer Institute between the years January 1992 and May 2008 we found 12 (1.5% of AML cases; 0.23% of lung cancer cases) cases (seven metachronous and five synchronous) with AML and lung cancer. All but one patient were smokers. There were no unique characteristic of either AML or lung cancer in these patients. Nine patients succumbed to AML, one died from an unrelated cause while undergoing treatment for AML, one died of lung cancer and one patient is alive after allogeneic transplantation for AML. In summary, this study supports the need for effective smoking cessation programs.

The predisposition of monosomy 7 to diabetes insipidus (DI) in acute myeloid leukemia (AML) led us to ask whether AML associated with monosomy 7 and DI will differ from AML associated with other karyotype aberrations and DI and whether the outcome of patients with AML and DI will differ from those without DI. We describe 2 patients from Roswell Park Cancer Institute and discuss 29 additional cases from the literature. AML with monosomy 7 and DI (n = 25) had a trend towards a lower complete remission (p = 0.0936) and worse survival (p = 0.0480) than AML with other karyotype changes and DI (n = 6). Further, AML with monosomy 7 and DI had worse complete remission rate and overall survival than AML with monosomy 7 but without DI. In conclusion, it appears that AML with monosomy 7 and DI is a disease entity with specifically poor outcome.

Background: We have shown that colon and breast cancer contains large amounts of urokinase (uPA), and that these cells are the actual sites of its synthesis. We isolated a large complex molecule consisting of the beta-chain of uPA, both chains of haptoglobin (Hp), and part or all of an NCAM-like molecule. The question arose whether it would be possible to show the presence of Hp in the same cells where uPA was found., Materials and Methods: Human colon and breast adenocarcinomas were investigated for expression of Hp and uPA by immunohistochemistry. Fluorescence in situ hybridization was used to identify the cells of origin of these antigens., Results: Hp was expressed in 8 of 11 colon adenocarcinomas, and in 10 of 12 breast tumors. uPA was demonstrable on the cell membrane and in the cytoplasm of all 11 colon adenocarcinomas studied, and cytoplasmic uPA-mRNA was found in all cases. While uPA was also detected in some stromal and inflammatory cells, Hp was present abundantly in such cells, as well as in capillary endothelial cells. Hp-mRNA was also found in both colon and breast tumors wherever the antigens were expressed., Conclusions: uPA and Hp are produced by the cancer cells and are not taken up by stromal elements. While the role of uPA in the malignant process is well documented, that of Hp is largely unexplored. Its ubiquity, shown here, suggests that it is also involved in some aspects of that process.

Adjuvant chemotherapy and radiation therapy for breast cancer are associated with therapy-related acute myeloid leukemia (AML)/myelodysplastic syndromes (MDS), but little is known about additional risk factors. Thirty-four patients with AML (n=26)/MDS (n=8) following breast cancer (cases) were compared with 2029 breast cancer patients without AML/MDS (controls). Cases were older at breast cancer diagnosis (mean 60.2 years versus 54.5 years; p=0.01) and more commonly had additional cancers (29% versus 4.9%; p<0.0001) and >or=4 first-degree relatives with any type of cancer (OR: 5.37, CI: 1.44-19.9). Thus risk factors for AML/MDS following breast cancer include older age, other cancers and multiple first-degree relatives with cancer.

Therapy-related myelodysplastic syndrome and acute myeloid leukemia (t-MDS/t-AML) have been reported only rarely following treatment of AML. We report five patients treated for de novo AML who developed t-MDS/t-AML, all with chromosome 7 abnormalities, including -7, del(7)(q22q36) and del(7)(p11.22p22). All had been treated with cytarabine, topoisomerase 2 inhibitors and granulocyte or granulocyte-monocyte colony-stimulating factor and three with alkylating agents as part of autologous transplant regimens. These cases further document t-MDS/t-AML as a complication of therapy for AML. Presence of chromosome 7 abnormalities in patients with and without prior alkylating agent therapy suggests possible association with the antimetabolite cytarabine.

Mitoxantrone is a DNA-topoisomerase 2 inhibitor used as a single agent for treatment of relapsing-remitting or progressive multiple sclerosis (MS). We present here two patients treated with mitoxantrone for MS who subsequently developed acute promyelocytic leukemia (APL). These constitute, to our knowledge, the eighth and ninth reports of APL in patients treated with mitoxantrone for MS. Topoisomerase 2 inhibitors are associated with therapy-related acute myeloid leukemia (t-AML) with 11q23 abnormalities, but therapy-related APL (t-APL) is less common, and documentation of nine cases of t-APL after mitoxantrone therapy for MS suggests a specific association.

The BCR/ABL tyrosine kinase inhibitor imatinib mesylate produces a high rate of cytogenetic responses in patients with Philadelphia (Ph)-positive chronic myeloid leukemia (CML), but secondary clonal chromosome abnormalities may develop in Ph-negative cells, and acute myeloid leukemia (AML) has been reported in patients with secondary chromosome abnormalities. We report a patient who developed AML during imatinib treatment of Ph-positive CML despite a cytogenetic response and absence of secondary chromosome abnormalities. Thus, development of AML as a rare event in CML patients with cytogenetic responses to imatinib therapy does not depend on the development of secondary cytogenetic abnormalities.

Reciprocal chromosomal translocations involving the MLL gene at chromosome region 11q23 are recurring cytogenetic abnormalities in both de novo and therapy-related acute myeloid leukemia (AML) and in acute lymphoblastic leukemia. We report a t(4;11)(p12;q23) with rearrangement of MLL and FRYL (also known as AF4p12), a human homolog to the furry gene of Drosophila, in an adult patient with therapy-related AML after fludarabine and rituximab therapy for small lymphocytic lymphoma and radiation therapy for breast carcinoma. To our knowledge, t(4;11)(p12;q23) has been reported in two previous patients, and MLL and FRYL rearrangement was demonstrated in one of them. Both of the previous patients had therapy-related leukemias after exposure to topoisomerase II inhibitors, whereas our patient had received cytotoxic therapy that did not include a topoisomerase II inhibitor. Thus, t(4;11)(p12;q23) with MLL and FRYL involvement represents a new recurring 11q23 translocation, to date seen only in therapy-related acute leukemias.

Objectives: To assess the forms and extent of genomic instability in thyroid cancers and colorectal neoplasms and to determine if such measurements could explain the generally excellent prognosis of thyroid malignant neoplasms compared with colon carcinoma., Design: Tumor genome analyses. Genomic instability was measured by the following 4 methods, listed in ascending order based on the size of events detected: inter-simple sequence repeat polymerase chain reaction (ISSR-PCR), fractional allelic loss (FAL) analysis, array-based comparative genomic hybridization (aCGH), and spectral karyotyping (SKY)., Results: The genomic instability index of 32 thyroid carcinomas, 59 colon carcinomas, and 11 colon polyps was determined by ISSR-PCR; no difference was seen among the 3 groups by this method. Fractional allelic loss rates were comparable in thyroid cancers and colon polyps and lower than FAL rates in colorectal cancers. Indolent papillary thyroid carcinomas were essentially diploid with no large-scale alterations in chromosome number or structure when evaluated by aCGH or SKY. In anaplastic thyroid cancers, aCGH revealed abundant chromosome alterations. Colorectal carcinomas showed extensive copy number changes and chromosomal rearrangements when analyzed by aCGH and SKY., Conclusions: Genomic alterations in papillary thyroid carcinoma, such as in benign colon polyps, are principally smaller events detected by ISSR-PCR. With the more aggressive tumor types (ie, anaplastic thyroid and colorectal carcinomas), larger events detected by FAL analysis, aCGH, and SKY were revealed. We hypothesize that mutations caused by smaller genomic alterations enable thyroid cells to achieve a minimal malignant phenotype. Mutations for aggressive biological behavior appear with larger genomic events.

Diagnostic evaluation of acute leukemia at Roswell Park Cancer Institute has routinely included immunophenotyping by multiparameter flow cytometry. In a retrospective analysis of 646 cases, morphology and cytochemistry established lineage in 612, but not in 34 (5%), of which 26, 5, and 3 were myeloid, undifferentiated, and lymphoid, respectively, based on immunophenotyping. In addition, immunophenotyping changed the lineage assigned based on morphology and cytochemistry in 11 cases (2%); 8 changed from lymphoid to myeloid, and 3 from myeloid to lymphoid. The data support routine inclusion of at least limited immunophenotyping in the diagnostic evaluation of acute leukemia., (Copyright 2006 International Society for Analytical Cytology.)

A growing body of literature reports therapy-related myelodysplastic syndrome (t-MDS) and acute myeloid leukemia (t-AML) in patients treated successfully for acute promyelocytic leukemia (APL). We report a t-MDS with an isochromosome 1q as a sole abnormality, 47,XY,+1,i(1)(q10), in a 46-year-old man with APL 14 years after he was treated with cytosine arabinosine and daunorubicin. The literature on t-MDS/t-AML after APL therapy and on isochromosome 1q is reviewed.

Recent improved treatments for lymphoid malignancies produce more long-term survivors, yet increase the risk for secondary malignancies. Therapy-related myelodysplasia and acute myeloid leukemia are well described, but secondary chronic myeloid leukemia (CML) has only rarely been reported. We report three patients with CML diagnosed 8, 10 and 2.5 years following Hodgkin's disease, non-Hodgkin's lymphoma and chronic lymphocytic leukemia therapy, respectively. BCR-ABL transcripts were not detected after completion of primary therapy in two cases. All three patients received imatinib therapy, with one patient subsequently undergoing allogeneic hematopoietic stem cell transplantation. All three patients have ongoing favorable responses to CML therapy.

Dendritic-like cells from t(9;22) acute lymphoblastic leukemia (ALL) blasts can activate T cells, while the original unmodified leukemic blasts cannot. To determine whether these functional differences were associated with differences in antigen-processing machinery (APM) component expression, we have measured the level of APM component expression in unmodified blasts and ALL-derived dendritic-like cells. Seven t(9;22) ALL patient samples and one cell line were studied for APM component expression utilizing a unique panel of recently developed monoclonal antibodies and a recently developed intracellular staining technique. In addition, the HLA class I antigen cell surface expression was measured. HLA class I antigens were similarly expressed on the unmodified blasts and on the autologous dendritic-like cells. Intracellular HLA class I antigen and tapasin expression (P=0.03 for both) were upregulated in all t(9;22) ALL-derived dendritic-like cells, in comparison to the unmodified blasts. These results provide a potential mechanism for the ability of t(9;22) ALL-derived dendritic-like cells to induce T-cell activation and, suggest that tapasin upregulation may serve as a marker to standardize and monitor the quality of the dendritic-like cells used in immunotherapy.

Myeloid sarcoma (MS) is a tumor mass of myeloblasts or immature myeloid cells occurring in an extramedullary site. In this study, seven cases of MS [stomach (1), testis (1), skin (2), and lymph node (3)] and 3 synchronous and 1 follow-up bone marrow (BM) samples were studied for genomic abnormalities using array comparative genomic hybridization (array-CGH). Array-CGH construction used approximately 5,400 bacterial artificial chromosome clones from the RPCI-11 library, spanning the human genome. Data were analyzed using the DNAcopy software and custom heuristics. All MS cases had genomic abnormalities detected by array-CGH. Unbalanced genomic abnormalities in five MS cases were confirmed by conventional cytogenetics (CC) and/or fluorescence in situ hybridization (FISH); these abnormalities included loss of 4q32.1-q35.2, 6q16.1-q21, and 12p12.2-p13.2 and gain of 8q21.2-q24.3, 8, 11q21-q25, 13q21.32-q34, 19, and 21. Array-CGH was also invaluable in identifying possible deletions, partner translocations, and breakpoints that were questionable by CC. The remaining two MS cases had genomic aberrations detected by array-CGH, but were not studied further by CC/FISH. Chromosome 8 was most commonly abnormal (3/7 cases). Identical genomic abnormalities were demonstrated in MS and in synchronous BM in two cases. These results demonstrate that array-CGH is a powerful tool to screen MS tissue for unbalanced genomic abnormalities, allowing identification of chromosome abnormalities when concurrent BM is nonanalyzable or nonleukemic., ((c) 2005 Wiley-Liss, Inc.)

Objectives: Pediatric embryonal heterologous rhabdomyosarcoma of the cervix is a rare tumor., Methods: We present an interesting clinico-pathologic situation of two sisters presenting with pediatric embryonal heterologous rhabdomyosarcoma of the cervix., Results: Pediatric embryonal heterologous rhabdomyosarcomas of the cervix are relatively uncommon. After a Pubmed search from 1952 to present, to our knowledge, this is the only report involving sisters presenting with this disease. Treatment has been extrapolated from collaborative groups such as Intergroup Rhabdomyosarcoma Study (IRS) Group so that optimal management may be achieved., Conclusions: We urge continued reporting of these rare tumors to enhance understanding if there may be a genetic component associated with them.

This paper reports a 73-year old woman with simultaneous presentation of acute monoblastic leukemia (acute myeloid leukemia (AML), French-American-British (FAB) type M5a) and mantle cell lymphoma. The patient presented with wasting, generalized lymphadenopathy, an extensive infiltrative rash and pancytopenia. Bone marrow and lymph node histopatholology showed extensive infiltration by leukemic monoblasts. Marrow cytogenetics revealed a complex karyotype, including t(8;16)(p11;p13). Flow cytometric immunophenotyping of peripheral blood, lymph node and bone marrow demonstrated two populations, expressing CD5, CD19, CD20 and CD22 and CD45, HLA-DR, CD13, CD33, CD14 and CD38, respectively. A focus of abnormal lymphocytes in the lymph node biopsy demonstrated BCL1 expression and t(11;14)(p11;p13) by fluorescence in situ hybridization and immunoglobulin heavy chain gene rearrangement by the polymerase chain reaction. The patient received infusional cytarabine, daunorubicin and etoposide chemotherapy, with complete remission of both the AML and the mantle cell leukemia. To the authors' knowledge, this is the first report of simultaneous presentations of AML, FAB M5a and mantle cell lymphoma. The case is discussed and the literature is reviewed.

Background: Mutations in the imprinted gene CDKN1C account for approximately 10% of Beckwith-Wiedemann syndrome (BWS) cases. Fibroblasts from BWS patients with loss of methylation (LOM) at the imprinting control region (ICR) KvDMR1 have reduced CDKN1C expression. Another group of BWS patients with downregulated CDKN1C expression but with normal methylation at KvDMR1 has been identified., Objective: To investigate the mechanism of CDKN1C silencing in BWS in these two classes of patients., Methods: The CDKN1C promoter region was analysed for changes in DNA methylation using bisulphite sequencing, and for alterations in chromatin structure using the chromatin immunoprecipitation (ChIP) assay., Results: There was only spurious CpG methylation of the CDKN1C promoter in fibroblast DNA from both normal individuals and patients with BWS, irrespective of the methylation status of KvDMR1. There was no detectable change in chromatin structure at the CDKN1C promoter in patients with LOM at KvDMR1. BWS patients with downregulated CDKN1C and normal methylation at KvDMR1 had depletion of dimethylated H3-K4 and enrichment of dimethylated H3-K9 and HP1gamma at the CDKN1C promoter, suggesting that in these cases gene silencing is associated with repressive chromatin changes., Conclusions: CDKN1C may be downregulated by multiple mechanisms including some that do not involve promoter methylation. In BWS patients with normal methylation at KvDMR1 and reduced expression of CDKN1C, repressive chromatin may play a role, but the absence of methylation and repressive chromatin structure at the CDKN1C promoter in BWS patients with LOM at KvDMR1 argues for a direct role of this epimutation in silencing CDKN1C.

Klinefelter syndrome is an underdiagnosed chromosomal disorder that has significant implications for health and for medical management. This report presents 5 adult male patients diagnosed with previously unsuspected Klinefelter syndrome as a result of cytogenetic testing for suspected hematologic malignancies. This case series highlights the importance of maintaining a comprehensive and holistic approach to medical care. The medical, genetic, and psychosocial implications of the Klinefelter diagnosis are discussed.

The purpose of this study was to evaluate fluorescence in situ hybridization abnormalities of the 2p23 anaplastic lymphoma kinase (ALK) gene loci in lymphomas with anaplastic morphology. We studied 24 anaplastic large cell lymphomas (ALCL) classified by World Health Organization criteria [17 primary nodal/systemic (10 ALK+, 7 ALK-), seven primary cutaneous], and 17 additional non-Hodgkin's lymphomas [one ALK+ B-lineage lymphoma, 14 ALK- diffuse large B-cell lymphomas (seven anaplastic variants, five nonanaplastic, two secondary CD30+), two follicular lymphomas]. ALK- lymphomas with anaplastic morphology showed extra nonrearranged anaplastic lymphoma kinase gene loci (P=0.004) due to trisomy 2 irrespective of the following factors: B or T/null phenotype (P=0.315), diagnostic categories of systemic or cutaneous ALCL or the above-mentioned B-cell lymphomas (P=0.131), and CD30 positivity by immunohistochemistry (P=1.000). Trisomy 2 was absent in all ALK+ lymphomas (P=0.009), which showed rearranged ALK gene loci (P<0.001). Whether trisomy 2 is a primary or secondary event that leads to ALK- lymphomas cannot be determined from this study. Its presence in secondary B-cell lymphomas suggests that trisomy 2 may be a secondary cytogenetic aberration in lymphomas in general. Further investigation of this finding is necessary to further our understanding of the heterogeneous group of ALK- lymphomas.

Genomic instability in cancer is frequently described as being either chromosomal instability or microsatellite instability, although when events within chromosomes are monitored, extensive intrachromosomal instability is also found. Spectral karyotyping was used to visualize how extensively genomic instability gives rise to intratumor genomic heterogeneity in sporadic colorectal carcinomas. Two factors were then examined which might relate to intrachromosomal instability in colorectal cancers: the presence of the glutathione transferase-Ml gene to detoxify potential carcinogens, and the presence of activated ras which has been associated with chromosomal instability when first expressed. Intrachromosomal genomic instability was previously determined by inter-(simple sequence repeat) PCR (inter-SSR PCR) and by fractional allelic loss rate for 348 markers. GSTM1 status was determined for each of 49 tumors through use of specific PCR, and 28 of the tumors showed the GSTM1 null genotype. A significant association was found between GSTMl-null status and elevated inter-(simple sequence repeat) PCR instability. In contrast, no association was found with fractional allelic loss rate. The first exons of the K-ras and H-ras oncogenes were sequenced in 72 colorectal cancers; 19 of the tumors had a mutation in codon 12 of the K-ras gene (24.5%), but no H-ras mutations were found. A weak correlation (p=0.10) was observed between mutant K-ras and inter-(simple sequence repeat) PCR genomic instability, and no association existed with fractional allelic loss rate.

We asked herein whether functional dendritic-like cells could be generated from t(9;22) acute lymphoblastic leukemia (ALL) blasts. We first determined that the combination of interleukin (IL)-1beta, IL-3, IL-7, tumor necrosis factor-alpha, stem cell factor and CD40 ligand was optimal for generating dendritic-like cells from t(9;22) ALL cell lines. Following 6 days in culture, four of five cell lines demonstrated dendrite-like morphology, upregulation of CCR7, CD54, CD80 and CD86, uptake of extracellular proteins and activation of T cells, and similar results were obtained with blasts from three t(9;22) ALL patients. The dendritic-like cells appeared to be composed of populations resembling both immature and mature dendritic cells (DCs), distinguished by CD80 expression. CD80-CD86+ cells were classified as immature DCs, demonstrating high endocytic capability and inducing minimal allogeneic T cell proliferation, while CD80+CD86+ cells were classified as mature DCs, exhibiting negligible endocytic capability and inducing robust allogeneic T cell proliferation. These mature dendritic-like cells induced autologous cytotoxic T cell responses against the unmodified blasts in a patient who achieved prolonged remission. In summary, CD80+CD86+ cells generated from t(9;22) ALL blasts may be useful in adoptive immunotherapy for t(9;22) ALL.

Acute myeloid leukemia (AML) with rearrangement of the core-binding factor (CBF) alpha or beta subunit gene has a favorable prognosis, but CD56 expression in CBFalpha-AML is associated with short disease-free survival. A proposed mechanism is overexpression of the multidrug resistance (MDR) protein P-glycoprotein (Pgp). CD56 expression, Pgp expression and function, and expression of the additional MDR proteins multidrug resistance protein-1 (MRP-1), lung resistance protein (LRP) and breast cancer resistance protein (BCRP) were studied in pretreatment blasts from 25 CBF-AML patients. CD56 expression was frequent in CBFalpha but rare in CBFbeta, and Pgp expression and function were frequent in both subtypes. CD56 expression did not correlate with Pgp expression or function, nor with expression of the other MDR proteins. Treatment failure associated with CD56 expression in CBFalpha-AML is not likely attributable to Pgp.

Massive hyperdiploidy (>50 chromosomes) and tetraploidy (4n) are rare cytogenetic abnormalities in myelocytic malignancies, and their significance is unknown. We report on 11 patients with acute myelocytic leukemia (AML) and two patients with a myelodysplastic syndrome (MDS) with massive hyperdiploidy (10 patients) or tetraploidy (3 patients) seen at our institution over a 12-year period. Eleven patients were male and two were female. Age range was 44-84 years (median, 70 years). Only one AML patient had a previous MDS, and no patient had therapy-related disease. One or more copies of chromosomes 8 and 19 were gained in eight patients each; other frequently gained chromosomes included 13, 15, and 21. Eight patients had structural abnormalities in addition to chromosome gain; del(5q) was most common (five patients). Eleven patients received induction chemotherapy, but only four achieved complete remission. Survival ranged from 1 to 22 months, with a median of 6 months. We conclude that massive hyperdiploidy and tetraploidy are infrequent abnormalities in AML and MDS, are seen primarily in de novo disease in older male patients and are associated with a low remission rate and short survival. Massive hyperdiploidy and tetraploidy define a prognostically unfavorable cytogenetic group in de novo AML.

Purpose: Pancreatic cancer remains a devastating problem with the majority of patients succumbing to death from this disease. A hallmark of pancreatic cancer is the loss of basement membrane that may be attributed to the action of urinary plasminogen activator (uPA) and matrix metalloproteinase-9 (MMP-9). These enzymes are also implicated in angiogenesis. uPA and microvessel density have been shown to be good prognostic indicators for breast and colon cancer. MMP-9 and microvessel density have not been investigated in pancreatic cancer. We have therefore investigated by immunohistochemistry: (a) frequency of uPA expression and its receptor uPAR and the site of synthesis of uPA by in situ hybridization (ISH); (b) MMP-9 and its coexpression with uPA; (c) microvessel density as determined by von Willebrand factor staining and its relationship to uPA and MMP-9 expression; and (d) correlation of these parameters with survival., Experimental Design: Archival paraffin sections of 27 pancreatic tumors were semiquantitatively investigated by immunohistochemistry using the following antibodies: (a) monoclonal antibodies (MAbs) uPA(1) and uPA(2) (3689 and 394, respectively); (b) MAb uPAR, (no. 3932); (c) MAb MMP-9 (no. 936); and (d) rabbit anti-F8RA/vWF. ISH was performed using a uPA cDNA., Results: Both uPA antibodies revealed overexpression of uPA (93%) often with uniform staining of tumor cells. uPAR and MMP-9 showed focal staining in only 52 and 37% of tumors, respectively. Morphologically normal appearing ductal cells in close proximity to tumors overexpressed uPA in contrast to distally located normal cells (P = <0.001). uPA staining was also investigated in pancreatic intraepithelial neoplasia (PanIN) lesions. PanIN 1A/B staining for uPA was seen in 8 cases (30%), that for PanIN 2 in 19 cases (70%), and for PanIN3 in 12 cases (44%). Lumen of microvessels in the tumor stroma also revealed staining of uPA in 10 cases (37%). ISH experiments revealed the presence of uPA mRNA not only in the cytoplasm of tumor cells but also in adjacent normal appearing ducts as well as in PanIN lesions. Patients with overexpression of uPA, uPAR, or MMP-9 had a trend toward poorer survival than those who did not express it. Microvessel density did not show any significant relationship with uPA, uPAR, and MMP-9 expression and survival., Conclusions: We conclude that uPA and MMP-9 are potential prognostic indicators in pancreatic cancer, whereas microvessel density may not be one. This study confirms our previous observation that uPA is made by the tumor cells themselves. Presence of uPA in vessels of tumor stroma suggests that uPA is in circulation, and its measurement and that of MMP-9 in the blood of these patients may aid in prognosis. Patients showing overexpression of uPA and MMP-9 have a trend toward shorter survival time.

MLL gene rearrangements leading to production of MLL fusion proteins are commonly detected in infant leukemia patients; the most common MLL fusion associated with infant leukemia is the MLL-AF4 fusion. A single case of chromosomal rearrangement leading to production of an MLL fusion with AF5Q31, a gene structurally similar to AF4, has been detected recently in the malignant cells of an infant leukemia patient. We have identified a second case of MLL-AF5Q31 fusion, arising from an insertion of MLL sequences into chromosome 5, also in an infant leukemia patient. Because MLL and AF5Q31 are transcribed in opposite orientations, a simple balanced chromosomal translocation cannot produce a fusion protein, and complex chromosomal rearrangements such as insertions and inversions are required to produce an MLL-AF5Q31 fusion protein. This report demonstrates that chromosomal insertion of MLL sequences is a rare but recurrent abnormality associated with infant leukemia., (Copyright 2003 Wiley-Liss, Inc.)

Double minute chromosomes (dmin) are small chromatin bodies consisting of genes amplified in an extrachromosomal location. dmins are uncommon in hematologic malignancies; they are seen primarily in acute myeloid leukemia, with amplification of the MYC oncogene or, less frequently, the MLL transcription factor. Nine patients with hematologic malignancies with dmin were seen at the Roswell Park Cancer Institute between 1985 and 2000; eight had acute myeloid leukemia and one a myelodysplastic syndrome. Fluorescence in situ hybridization (FISH) demonstrated MYC amplification on dmin in four patients, but MLL amplification was not seen. Spectral karyotyping showed that the dmin derived from chromosome 11 in one patient and from chromosome 19 in two others without MYC or MLL amplification; derivation from these chromosomes was confirmed by FISH with chromosome paint probes. The dmin of chromosome 11 origin hybridized to a bacterial artificial chromosome (BAC) RP11-112M22 that maps to 11q24.3 and is predicted to contain ETS1 and other markers, including D11S11351 and D11S4091. The dmin of chromosome 19 origin in one patient hybridized to BACs RP11-46I12 and RP11-110J19; in the other patient, these clones did not hybridize with the dmin, but were found to be amplified on a marker chromosome that was derived from chromosome 19 in that patient's cells. These BACs have been mapped to 19q12-19q13.1 and 19q11-19q13.1, respectively, and are predicted to contain the markers D19S409 and D19S919 and the gene for ubiquinol-cytochrome C reductase, Rieske iron-sulfur polypeptide1 (UQCRFS1). dmin originating from chromosome 19 have not been reported previously in hematologic malignancies., (Copyright 2002 Wiley-Liss, Inc.)

The MLL gene at chromosome band 11q23 is specifically cleaved at a unique site within its breakpoint cluster region (bcr) during the higher order chromatin fragmentation associated with apoptosis. We now show that the same specific DNA cleavage event can be detected in an exogenous MLL bcr fragment that is integrated into the genome outside of its normal chromosomal context, as well as in an extrachromosomal episome containing an MLL bcr fragment. We also show that episomal or randomly integrated copies of the MLL bcr behave similar to the endogenous MLL bcr when tested in a scaffold-associated region (SAR) assay. Furthermore, an episomal murine MLL bcr introduced into human cells is cleaved at the same site as the endogenous murine MLL bcr; this episomal murine MLL bcr also functions as a SAR in human cells. We conclude that both nuclear DNA scaffold attachment as well as site-specific DNA cleavage can be directed by sequences contained within the MLL bcr, and that it is feasible to study these events using episomal shuttle vectors.

Methotrexate is in widespread use as second-line therapy for rheumatoid arthritis. Treatment with methotrexate in this and other settings has not been associated with the development of therapy-related leukemias. Four patients with rheumatoid arthritis are reported who developed acute myeloid leukemia (AML) while receiving low dose weekly methotrexate therapy in the absence of previous or concomitant treatment with known leukemogenic agents. AML in these four patients was of different morphologic subtypes and was associated with heterogeneous cytogenetic abnormalities, cell surface marker expression and multidrug resistance protein expression. None of the recognized features of therapy-related leukemia were present in these four nor in five previously-reported patients. It is likely that the occurrence of AML in patients with rheumatoid arthritis in the setting of methotrexate therapy represents the coincidence of these two diseases, and does not reflect a causal relationship.

Chronic myeloid leukemia in myeloid blast crisis (CML-MBC) is highly resistant to standard induction chemotherapy regimens. Anecdotal results from previous clinical trials support the concept of dose escalation in patients with CML-MBC. Eight patients with CML-MBC were treated with cytosine arabinoside (Ara-C) 1.5-3.0 g/m2 intravenously over 1 hr every 12 hr for 12 doses and idarubicin 12 mg/m2 intravenously daily for 3 days. Sixteen previous reports describing the use of Ara-C-based chemotherapy regimens in patients with CML-MBC were also reviewed. Our patients' median age was 62 years (range, 42-69 years). One patient achieved complete hematologic remission (95% confidence interval, 0.3%, 53%). The median survival for our patients was 7.3 months. These results were not different from previous published reports using Ara-C-based chemotherapy regimens to treat CML-MBC. In summary, the combination of high-dose Ara-C and idarubicin did not improve the overall prognosis of patients with CML-MBC. Innovative approaches need to be explored for this patient population., (Copyright 2001 Wiley-Liss, Inc.)

Signal transducer and activator of transcription (STAT) proteins are implicated in the control of cell survival, proliferation and differentiation in response to hematopoietic cytokines. C-terminally truncated STAT isoforms (STATbeta), as opposed to the full length form (STATalpha), have a competitive or even transdominant negative effect on gene induction mediated by the STAT pathway. We have previously demonstrated that while constitutively active STAT proteins were detected in ten of 36 (28%) for STAT3 and eight of 36 (22%) for STAT5 in pretreatment samples from newly diagnosed acute myeloid leukemia (AML) patients, a significantly larger fraction of samples [21 of 27 (78%)] expressed STATbeta proteins. To determine whether STATbeta expression was maintained or increased after relapse in AML, we compared STAT activity and isoform expression at diagnosis and at relapse in 17 patients. In this selected group, constitutively active STAT3 was detected in 13 of 17 (76%) AML samples at diagnosis but was detected in only four of these patients at relapse. Constitutively active STAT5 was detected in three of 17 (18%) AML samples at diagnosis; but only two at relapse. In contrast, STATbeta protein expression was observed in 12 of the 17 pretreatment samples (71%) and in 16 of 17 samples at relapse. Only one patient did not express STATbeta at relapse. Our results suggest that STATbeta isoform expression, rather than level of constitutive activity, may be involved in disease progression in AML.