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Find video protocols related to scientific articles indexed in Pubmed.
Ras pathway mutations are highly prevalent in relapsed childhood acute lymphoblastic leukaemia, may act as relapse-drivers and confer sensitivity to MEK inhibition.
Blood
PUBLISHED: 09-26-2014
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For most children who relapse with acute lymphoblastic leukaemia, the prognosis is poor and there is a need for novel therapies to improve outcome. We screened samples from children with B lineage ALL entered into the ALL-REZ BFM 2002 clinical trial (ClinicalTrials.gov identifier: NCT00114348) for somatic mutations activating the Ras pathway (KRAS, NRAS, FLT3 and PTPN11) and showed mutation to be highly prevalent (76 from 206). Clinically, they were associated with high risk features including early relapse, CNS involvement and specifically for NRAS/KRAS mutations, chemoresistance. KRAS mutations were associated with a reduced overall survival. Mutation screening of the matched diagnostic samples found many to be wildtype but using more sensitive allelic specific assays, low level mutated subpopulations were found in many cases, suggesting that they survived up front therapy and subsequently emerged at relapse. Preclinical evaluation of the MEK1/2 inhibitor, selumetinib (AZD6244, ARRY-142886) showed significant differential sensitivity in Ras pathway mutated ALL compared to wild type cells both in vitro and in a orthotopic xenograft model engrafted with primary ALL and in the latter, reduced RAS mutated CNS leukaemia. Given these data, clinical evaluation of selumetinib may be warranted for Ras pathway mutated relapsed ALL.
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Recurrent intestinal mucinous borderline tumors of the ovary: a report of 5 cases causing problems in diagnosis, including distinction from mucinous carcinoma.
Int. J. Gynecol. Pathol.
PUBLISHED: 02-04-2014
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Intestinal mucinous borderline tumors (IMBTs) of the ovary are generally associated with a highly favorable outcome and rarely recur. We describe 5 cases of IMBT initially treated by cystectomy or by salpingo-oophorectomy that was likely incomplete, with subsequent recurrences. Three cases were received in consultation, and in each of these, the clinical and intraoperative findings were worrisome for mucinous carcinoma, and diagnostic difficulty was encountered by the referring pathologist. The patient age ranged from 28 to 69 (median 53) yrs. All tumors were clinically Stage I at presentation; in at least 3 cases, extensive adhesiolysis was required during their removal. A pathologic diagnosis of IMBT was made in 4 cases; the remaining tumor was inadequately sampled (3 blocks from a 7.5-cm tumor showed predominantly benign to focally borderline mucinous epithelium). A total of 8 recurrences, all as IMBT, developed at mean follow-up of 26 (range, 6-102) mo; 6 of these occurred within ?2 yr. In 4 cases, removal of recurrent tumor required an extensive operation because of bowel and/or vaginal involvement. Residual ovarian stroma was identified in all recurrences. There was no evidence of invasive mucinous carcinoma, pseudomyxoma peritonei, or a primary tumor elsewhere (including appendix) in any of the cases. Our findings indicate that patients with IMBTs who undergo cystectomy or oophorectomy requiring adhesiolysis are at increased risk of recurrence, which may occur early, be multiple, and potentially require extensive resection if sites such as bowel or vagina are involved. Recurrences of IMBT that develop in this setting likely represent regrowth of incompletely resected IMBT, or arise within residual ovarian tissue. This is the first detailed clinicopathologic study of such cases.
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Appendiceal or cecal endometriosis with intestinal metaplasia: a potential mimic of appendiceal mucinous neoplasms.
Am. J. Surg. Pathol.
PUBLISHED: 01-24-2014
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Appendiceal or cecal endometriosis uncommonly presents diagnostic challenges, but rare cases with intestinal metaplasia can mimic mucinous tumors of the appendix. We describe 6 cases of appendiceal or cecal endometriosis with intestinal metaplasia that grossly and microscopically mimicked appendiceal mucinous tumors. The cases were characterized by marked mural hypertrophy and elastosis, with distortion of the appendix, and virtual obliteration of the lumen of the appendix. All cases had conventional endometriosis, together with varying amounts of endometriosis with intestinal-type epithelium. In 2 cases, scant or virtually no stroma around some mucin-filled glands made their recognition as endometriosis more difficult. None of the cases had dysplasia of the mucinous epithelium. All patients had benign follow-up (mean 3.6 y). Endometriosis with intestinal metaplasia is a rare phenomenon that, when it occurs in the appendix or cecum, may lead to misdiagnosis as a low-grade appendiceal mucinous neoplasm or mucinous carcinoma.
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Ras/Raf/MEK/ERK Pathway Activation in Childhood Acute Lymphoblastic Leukemia and Its Therapeutic Targeting.
Front Oncol
PUBLISHED: 01-01-2014
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Deregulation of the Ras/Raf/MEK/extracellular signal-regulated kinase pathway is a common event in childhood acute lymphoblastic leukemia and is caused by point mutation, gene deletion, and chromosomal translocation of a vast array of gene types, highlighting its importance in leukemia biology. Pathway activation can be therapeutically exploited and may guide new therapies needed for relapsed acute lymphoblastic leukemia and other high risk subgroups.
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Histotype-genotype correlation in 36 high-grade endometrial carcinomas.
Am. J. Surg. Pathol.
PUBLISHED: 10-01-2013
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Endometrioid, serous, and clear cell carcinomas are the major types of endometrial carcinoma. Histologic distinction between these different tumor types can be difficult in high-grade cases, in which significant interobserver diagnostic disagreement exists. Endometrioid and clear cell carcinomas frequently harbor ARID1A and/or PTEN mutations. Serous carcinoma acquires TP53 mutations/inactivation at onset, with a significant subset harboring an additional mutation in PPP2R1A. This study examines the correlation between tumor histotype and genotype in 36 previously genotyped high-grade endometrial carcinomas. This included 23 endometrioid/clear cell genotype and 13 serous genotype tumors. Eight subspecialty pathologists reviewed representative online slides and rendered diagnoses before and after receiving p53, p16, and estrogen receptor immunostaining results. ? statistics for histotype-genotype concordance were calculated. The average ? values for histotype-genotype concordance was 0.55 (range, 0.30 to 0.67) on the basis of morphologic evaluation alone and it improved to 0.68 (range, 0.54 to 0.81) after immunophenotype consideration (P<0.001). Genotype-incompatible diagnoses were rendered by at least 2 pathologists in 12 of 36 cases (33%) (3 cases by 2/8 pathologists, 2 by 3/8, 2 by 4/8, 3 by 6/8, 1 by 7/8, and 1 case by 8/8 pathologists). Six of the 12 were endometrioid/clear cell genotype tumors, and the other 6 were serous genotype tumors. The histopathologic features associated with histotype-genotype-discordant cases were reviewed, and specific diagnostic recommendations were made to improve concordance. This study found that although the majority of morphologic diagnoses are genotype concordant, genotype-incompatible diagnoses are made in a significant subset of cases. Judicious use and interpretation of p53 immunohistochemistry in selected scenarios can improve histotype-genotype concordance.
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HNF-1? in ovarian carcinomas with serous and clear cell change.
Int. J. Gynecol. Pathol.
PUBLISHED: 09-28-2013
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Many ovarian tumors, including high-grade serous carcinoma (HGSC), show clear cell change. Accurate diagnosis is important, however, as ovarian clear cell carcinoma (OCCC) is known to be less responsive to traditional types of ovarian cancer chemotherapies. In a previous study, the clinical, morphologic, and immunohistochemical features of 32 ovarian carcinomas, which had been previously diagnosed as pure OCCC (n=11), pure HGSC (n=11), and mixed serous and clear cell (MSC) (n=10), were analyzed. The immunoreactivities of WT1, ER, and p53, as well as the mitotic indices and stages of presentation of the MSC, were similar to those of HGSC. It was consequently concluded that MSC represented HGSC with clear cell change. Hepatocyte nuclear factor-1? (HNF-1?) is a relatively new immunohistochemical marker that has been shown to be rather sensitive and specific for OCCC. We thus sought to evaluate this marker in this specific group of tumors. One block each of pure HGSC and pure OCCC were stained with HNF-1?. In the cases of MSC, 2 blocks were stained when the serous and clear cell components were not present on the same slide. None (0/11) of the pure HGSC showed immunoreactivity for HNF-1?, whereas all (11/11) of the pure OCCC were positive. In the cases of MSC, both the serous and clear cell components were negative for HNF-1?. HNF-1? seems to be a sensitive and specific marker for OCCC and is not expressed in HGSC with clear cell change. The pattern of immunoreactivity of HNF-1? in tumors with both serous and clear cell change supports the conclusion that MSC are HGSC with clear cells.
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Variation at 10p12.2 and 10p14 influences risk of childhood B-cell acute lymphoblastic leukemia and phenotype.
Blood
PUBLISHED: 08-30-2013
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Acute lymphoblastic leukemia (ALL) is the major pediatric cancer diagnosed in economically developed countries with B-cell precursor (BCP)-ALL, accounting for approximately 70% of ALL. Recent genome-wide association studies (GWAS) have provided the first unambiguous evidence for common inherited susceptibility to BCP-ALL, identifying susceptibility loci at 7p12.2, 9p21.3, 10q21.2, and 14q11.2. To identify additional BCP-ALL susceptibility loci, we conducted a GWAS and performed a meta-analysis with a published GWAS totaling 1658 cases and 4723 controls, with validation in 1449 cases and 1488 controls. Combined analysis identified novel loci mapping to 10p12.2 (rs10828317, odds ratio [OR] = 1.23; P = 2.30 × 10(-9)) and 10p14 marked by rs3824662 (OR = 1.31; P = 8.62 × 10(-12)). The single nucleotide polymorphism rs10828317 is responsible for the N215S polymorphism in exon 7 of PIP4K2A, and rs3824662 localizes to intron 3 of the transcription factor and putative tumor suppressor gene GATA3. The rs10828317 association was shown to be specifically associated with hyperdiploid ALL, whereas the rs3824662-associated risk was confined to nonhyperdiploid non-TEL-AML1 + ALL. The risk allele of rs3824662 was correlated with older age at diagnosis (P < .001) and significantly worse event-free survivorship (P < .0001). These findings provide further insights into the genetic and biological basis of inherited genetic susceptibility to BCP-ALL and the influence of constitutional genotype on disease development.
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Common variation at 3q26.2, 6p21.33, 17p11.2 and 22q13.1 influences multiple myeloma risk.
Nat. Genet.
PUBLISHED: 03-06-2013
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To identify variants for multiple myeloma risk, we conducted a genome-wide association study with validation in additional series totaling 4,692 individuals with multiple myeloma (cases) and 10,990 controls. We identified four risk loci at 3q26.2 (rs10936599, P = 8.70 × 10(-14)), 6p21.33 (rs2285803, PSORS1C2, P = 9.67 × 10(-11)), 17p11.2 (rs4273077, TNFRSF13B, P = 7.67 × 10(-9)) and 22q13.1 (rs877529, CBX7, P = 7.63 × 10(-16)). These data provide further evidence for genetic susceptibility to this B-cell hematological malignancy, as well as insight into the biological basis of predisposition.
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Papillary proliferation of the endometrium: a clinicopathologic study of 59 cases of simple and complex papillae without cytologic atypia.
Am. J. Surg. Pathol.
PUBLISHED: 03-06-2013
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Papillary proliferation of the endometrium (PPE) without cytologic atypia is uncommon and has only been studied in detail by Lehman and Hart in 2001. On histologic examination, PPE ranges from simple papillae with fibrovascular cores, often involving the surface of endometrial polyps, to complex intracystic proliferations; some consider the latter to be analogous to nonatypical complex hyperplasia. To further characterize PPE, with emphasis on the risk of and features associated with concurrent or subsequent neoplasia, the clinicopathologic features of 59 cases without cytologic atypia were studied. The cases were classified into 2 groups according to the degree of architectural complexity and extent of proliferation. Group 1 consisted of those with localized simple papillae. Simple papillae were defined as those with short, predominantly nonbranching stalks; those with occasional secondary branches and/or detached papillae were also included in this group. Localized proliferations were those with 1 or 2 foci involving the surface or the subjacent glands of polyps or nonpolypoid endometrium. Group 2 consisted of those with complex papillae and/or those with diffuse and crowded intracystic papillae. Complex papillae were those with either short or long stalks, with frequent secondary and complex branches. Diffuse proliferation was defined as presence of 3 or more foci within a specimen or involvement of >50% of the endometrial polyp by simple or complex PPE. Any coexistent or subsequent hyperplasia of conventional type (World Health Organization classification) or adenocarcinoma was recorded. The age of patients ranged from 23 to 82 years (median, 53 y); 36 (61%) were postmenopausal. The majority presented with abnormal vaginal bleeding. Sixteen patients (27%) were receiving hormonal preparations including 5 who were treated with a progestogen for preexisting endometrial hyperplasia or low-grade endometrioid adenocarcinoma. The histologic diagnosis of PPE was made in 49 biopsies and in 10 hysterectomy specimens. Thirty-six cases (61%) were classified as group 1 and 23 (39%) as group 2. In 47 cases (80%), there was a coexisting endometrial polyp, 39 (66%) of which were involved by the PPE. Fifty-three cases (90%) had coexisting epithelial metaplastic changes, 41 (77%) of which were involved by the PPE. The most common type of metaplasia was mucinous (41 of 59 cases, or 69%). Follow-up information was known for 46 patients (78%). Coexistent or subsequent nonatypical and atypical hyperplasia was found in 8 (17%) and 6 cases (13%), respectively. In 6 of the 46 cases (13%), a low-grade endometrioid adenocarcinoma was present either in the original specimen or during follow-up. In contrast to group 1 PPE, those with group 2 features were significantly associated with concurrent or subsequent premalignant lesions (nonatypical and atypical hyperplasia) or carcinoma (P<0.0001). This study indicates that localized and architecturally simple PPEs confined to a completely removed polyp are usually associated with a benign outcome and may be appropriately labeled as "benign papillary proliferation of the endometrium." Lesions with architecturally complex papillae, especially when extensive, have an increased risk of concurrent or subsequent endometrial hyperplasia and carcinoma and should probably be regarded as analogous to atypical complex hyperplasia, and the term "complex papillary hyperplasia" is appropriate. As the distinction between simple and complex PPE may be difficult in small endometrial aspirational samples, consideration for curettage should be given to ascertain whether the lesion has been completely removed.
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Post-transcriptional exon shuffling events in humans can be evolutionarily conserved and abundant.
Genome Res.
PUBLISHED: 09-23-2011
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In silico analyses have established that transcripts from some genes can be processed into RNAs with rearranged exon order relative to genomic structure (post-transcriptional exon shuffling, or PTES). Although known to contribute to transcriptome diversity in some species, to date the structure, distribution, abundance, and functional significance of human PTES transcripts remains largely unknown. Here, using high-throughput transcriptome sequencing, we identify 205 putative human PTES products from 176 genes. We validate 72 out of 112 products analyzed using RT-PCR, and identify additional PTES products structurally related to 61% of validated targets. Sequencing of these additional products reveals GT-AG dinucleotides at >95% of the splice junctions, confirming that they are processed by the spliceosome. We show that most PTES transcripts are expressed in a wide variety of human tissues, that they can be polyadenylated, and that some are conserved in mouse. We also show that they can extend into 5 and 3 UTRs, consistent with formation via trans-splicing of independent pre-mRNA molecules. Finally, we use real-time PCR to compare the abundance of PTES exon junctions relative to canonical exon junctions within the transcripts from seven genes. PTES exon junctions are present at <0.01% to >90% of the levels of canonical junctions, with transcripts from MAN1A2, PHC3, TLE4, and CDK13 exhibiting the highest levels. This is the first systematic experimental analysis of PTES in human, and it suggests both that the phenomenon is much more widespread than previously thought and that some PTES transcripts could be functional.
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Detection and monitoring of normal and leukemic cell populations with hierarchical clustering of flow cytometry data.
Cytometry A
PUBLISHED: 09-05-2011
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Flow cytometry is a valuable tool in research and diagnostics including minimal residual disease (MRD) monitoring of hematologic malignancies. However, its gradual advancement toward increasing numbers of fluorescent parameters leads to information rich datasets, which are challenging to analyze by standard gating and do not reflect the multidimensionality of the data. We have developed a novel method to analyze complex flow cytometry data, based on hierarchical clustering analysis (HCA) but with a new underlying algorithm, using Mahalanobis distance measure. HCA is scalable to analyze complex multiparameter datasets (here demonstrated on up to 12 color flow cytometry and on a 20-parameter synthetic dataset). We have validated this method by comparison with standard gating approaches when performed independently by expert cytometrists. Acute lymphoblastic leukemia blast populations were analyzed in diagnostic and follow-up datasets (n = 123) from three centers. HCA results correlated very well (Passing-Bablok correlation coefficient = 0.992, slope = 1, intercept = -0.01) with standard gating data obtained by the I-BFM FLOW-MRD study group. To further improve the performance in follow-up samples with low MRD levels and to automate MRD detection, we combined HCA with support vector machine (SVM) learning. HCA in combination with SVM provides a novel diagnostic tool that not only allows analysis of increasingly complex flow cytometry data but also is less observer-dependent compared with classical gating and has potential for automation.
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Potent, selective inhibitors of fibroblast growth factor receptor define fibroblast growth factor dependence in preclinical cancer models.
Mol. Cancer Ther.
PUBLISHED: 07-15-2011
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We describe here the identification and characterization of 2 novel inhibitors of the fibroblast growth factor receptor (FGFR) family of receptor tyrosine kinases. The compounds exhibit selective inhibition of FGFR over the closely related VEGFR2 receptor in cell lines and in vivo. The pharmacologic profile of these inhibitors was defined using a panel of human tumor cell lines characterized for specific mutations, amplifications, or translocations known to activate one of the four FGFR receptor isoforms. This pharmacology defines a profile for inhibitors that are likely to be of use in clinical settings in disease types where FGFR is shown to play an important role.
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Casitas B lymphoma mutations in childhood acute lymphoblastic leukemia.
Genes Chromosomes Cancer
PUBLISHED: 06-03-2011
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Casitas B-lineage lymphoma (CBL) proteins are RING finger ubiquitin E3 ligases that attenuate the signaling of receptor tyrosine kinases and are mutated in a number of myeloid disorders. In this study, mutational screening of the linker-RING domains of CBL and CBLB was performed by denaturing high performance liquid chromatography in a cohort of diagnostic (n = 180) or relapse (n = 46) samples from children with acute lymphoblastic leukemia. Somatic mutations were identified in three children, giving an overall incidence of 1.7% and involved small deletions affecting the intron/exon boundaries of exon 8, leading to skipping of exon 8 and abolishing E3 ligase function. Mutated primary samples were associated with constitutive activation of the RAS pathway and sensitivity to MEK inhibitors was shown. Thus, mutation of CBL is an alternative route to activate the RAS pathway and may identify children who are candidates for MEK inhibitor clinical trials.
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Müllerian adenosarcomas with unusual growth patterns: staging issues.
Int. J. Gynecol. Pathol.
PUBLISHED: 05-31-2011
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Uterine adenosarcomas are uncommon mixed Müllerian neoplasms, most commonly arising in the uterine corpus. A new Federation of International Gynecologic Organization staging system for these tumors has recently been implemented. This staging system is an improvement on the earlier generic application of the 1988 Federation of International Gynecologic Organization staging system for endometrial cancer to adenosarcoma. Herein, we report 3 uterine adenosarcomas with unusual features. For 2 of these, no specific staging guidelines are provided by either the earlier or, more importantly, the new staging system. The first case is of an adenosarcoma arising in the eutopic endometrium with involvement of underlying adenomyosis without myometrial invasion; the second originated in a mural adenomyoma in the absence of eutopic endometrial involvement; and the third case encompassed synchronous endometrial and extrauterine (peritoneal) neoplasms. Such cases are rare, and there is insufficient evidence to be definitive about staging. Thus, we suggest a descriptive reporting strategy for adenosarcomas with these unusual features. We also propose a reporting nomenclature for such cases to ensure standardization such that they can be adequately recorded in synoptic reporting protocols. This will facilitate reliable data collection such that an evidence-based staging system for these scenarios may be derived.
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Genomic characterization implicates iAMP21 as a likely primary genetic event in childhood B-cell precursor acute lymphoblastic leukemia.
Blood
PUBLISHED: 04-28-2011
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Intrachromosomal amplification of chromosome 21 (iAMP21) defines a distinct subgroup of childhood B-cell precursor acute lymphoblastic leukemia (BCP-ALL) that has a dismal outcome when treated with standard therapy. For improved diagnosis and risk stratification, the initiating genetic events need to be elucidated. To investigate the genetic basis of BCP-ALL, genomes of 94 iAMP21 patients were interrogated by arrays, FISH, and multiplex ligation-dependent probe amplification. Most copy number alterations targeted chromosome 21, reinforcing the complexity of this chromosome. The common region of amplification on chromosome 21 was refined to a 5.1-mb region that included RUNX1, miR-802, and genes mapping to the Down syndrome critical region. Recurrent abnormalities affecting genes in key pathways were identified: IKZF1 (22%), CDKN2A/B (17%), PAX5 (8%), ETV6 (19%), and RB1 (37%). Investigation of clonal architecture provided evidence that these abnormalities, and P2RY8-CRLF2, were secondary to chromosome 21 rearrangements. Patient outcome was uniformly poor with standard therapy irrespective of the presence or absence of these changes. This study has provided evidence that chromosome 21 instability is the only anomaly among those so far investigated that is common to all iAMP21 patients, and therefore the initiating event is likely to be found among the complex structural rearrangements of this abnormal chromosome.
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Comparative pre-clinical evaluation of receptor tyrosine kinase inhibitors for the treatment of multiple myeloma.
Leuk. Res.
PUBLISHED: 01-14-2011
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Fibroblast growth factor receptor 3 (FGFR3) is up-regulated as a result of the t(4;14)(p16;q32) translocation that occurs in up to 20% of multiple myeloma (MM) patients. Recent studies have demonstrated that up-regulation of FGFR3 promotes cell survival, growth and drug resistance in malignant plasma cells, both in vitro and in vivo. Therefore, inhibition of FGFR3 signalling is potential target for the chemotherapeutic intervention in t(4;14) MM.
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MHC variation and risk of childhood B-cell precursor acute lymphoblastic leukemia.
Blood
PUBLISHED: 11-08-2010
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A role for specific human leukocyte antigen (HLA) variants in the etiology of childhood acute lymphoblastic leukemia (ALL) has been extensively studied over the last 30 years, but no unambiguous association has been identified. To comprehensively study the relationship between genetic variation within the 4.5 Mb major histocompatibility complex genomic region and precursor B-cell (BCP) ALL risk, we analyzed 1075 observed and 8176 imputed single nucleotide polymorphisms and their related haplotypes in 824 BCP-ALL cases and 4737 controls. Using these genotypes we also imputed both common and rare alleles at class I (HLA-A, HLA-B, and HLA-C) and class II (HLA-DRB1, HLA-DQA1, and HLA-DQB1) HLA loci. Overall, we found no statistically significant association between variants and BCP-ALL risk. We conclude that major histocompatibility complex-defined variation in immune-mediated response is unlikely to be a major risk factor for BCP-ALL.
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Genome-wide homozygosity signatures and childhood acute lymphoblastic leukemia risk.
Blood
PUBLISHED: 03-15-2010
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Recent studies have reported that regions of homozygosity (ROH) in the genome are detectable in outbred populations and can be associated with an increased risk of malignancy. To examine whether homozygosity is associated with an increased risk of developing childhood B-cell precursor acute lymphoblastic leukemia (BCP-ALL), we analyzed 824 ALL cases and 2398 controls genotyped for 292 200 tagging SNPs. Across the genome, cumulative distribution of ROH was not significantly different between cases and controls. Four common ROH at 10p11.2-10q11.21, 1p31.1, 19p13.2-3, and 20q11.1-23 were, however, associated with ALL risk at P less than .01 (including 1 ROH to which the erythropoietin receptor [EPOR] gene maps, P = .005) but were nonsignificant after adjusting for multiple testing. Our findings make it unlikely that levels of measured homozygosity, caused by autozygosity, uniparental isodisomy, or hemizygosity, play a major role in defining BCP-ALL risk in predominantly outbred populations.
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Mismatch repair and the downstream target genes, PAX5 and Ikaros, in childhood acute lymphoblastic leukemia.
Leuk. Res.
PUBLISHED: 02-08-2010
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The mismatch repair (MMR) pathway is a post-replicative DNA repair process and MMR deficiency is a common feature of ALL cell lines. In this study we have investigated MMR deficiency in a large cohort of primary relapsed ALL (n=40) and investigated coding microsatellites (MS) of the lymphoid transcription factors, PAX5 and IKZF1 as downstream target genes. Only one patient showed MMR deficiency, as evidenced by microsatellite instability, which was acquired at relapse and was associated with reduced expression of both MLH1 and MSH2. Coding MS in candidate target genes including PAX5, IKZF1, BAX and TGFBRII were all wild type in this patient but the MMR-deficient cell line REH, was confirmed to have a coding MS in both PAX5 and TGFBRII. Whilst MMR deficiency is not highly prevalent in primary ALL, optimisation of the drug regimen to omit/replace thioguanines should be considered for children with MMR deficiency and/or reduced expression of key pathway components.
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NFkappaB modulators in a model of glucocorticoid resistant, childhood acute lymphoblastic leukemia.
Leuk. Res.
PUBLISHED: 01-27-2010
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Glucocorticoids (GCs) are pivotal agents in the treatment of childhood acute lymphoblastic leukaemia (ALL) but the molecular basis of GC-resistance remains unclear. Expression-array studies have shown that commonly upregulated genes associated with GC-sensitivity include GR, glucocorticoid-induced leucine zipper (GILZ) and IkappaBalpha, which all negatively interact with components of the pro-survival NFkappaB pathway and therefore may be critical determinants of GC-sensitivity. We have investigated these regulators and their effect on NFkappaB activity in GC-resistant descendents of the B-lineage ALL cell line, PreB 697. We show that while differential up regulation of the modulators (GILZ, GR and IkappaBalpha) was demonstrated in GC-sensitive compared to GC-resistant sub-lines, this was not coupled with altered nuclear translocation or functionality of the RelA, p50 or c-Rel subunits of NFkappaB. Thus, GC-resistance in the PreB 697 cell line model is not mediated by NFkappaB, however further investigation of the impact of these GC-sensitive associated proteins on other survival pathways, such as the RAS-RAF-MEK-ERK pathway, is warranted.
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Variation in CDKN2A at 9p21.3 influences childhood acute lymphoblastic leukemia risk.
Nat. Genet.
PUBLISHED: 01-25-2010
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Using data from a genome-wide association study of 907 individuals with childhood acute lymphoblastic leukemia (cases) and 2,398 controls and with validation in samples totaling 2,386 cases and 2,419 controls, we have shown that common variation at 9p21.3 (rs3731217, intron 1 of CDKN2A) influences acute lymphoblastic leukemia risk (odds ratio = 0.71, P = 3.01 x 10(-11)), irrespective of cell lineage.
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Flow minimal residual disease monitoring of candidate leukemic stem cells defined by the immunophenotype, CD34+CD38lowCD19+ in B-lineage childhood acute lymphoblastic leukemia.
Haematologica
PUBLISHED: 11-30-2009
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Flow cytometric minimal residual disease (MRD) monitoring could become more powerful if directed towards the disease-maintaining leukemic stem cell (LSC) compartment. Using a cohort of 48 children with B-lineage acute lymphoblastic leukemia (ALL), we sought the newly proposed candidate-LSC population, CD34(+)CD38(low)CD19(+), at presentation and in end of induction bone marrow samples. We identified the candidate LSC population in 60% of diagnostic samples and its presence correlated with expression of CD38, relative to that of normal B-cell progenitors. In addition, the candidate LSC was not detectable in all MRD positive samples. The absence of the population in 40% of diagnostic and 40% of MRD positive samples does not support the use of this phenotype as a generic biomarker to track LSCs and suggests that this phenotype may be an artifact of CD38 underexpression rather than a biologically distinct LSC population. ClinicalTrials.gov Identifier: NCT00222612.
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Deregulated expression of cytokine receptor gene, CRLF2, is involved in lymphoid transformation in B-cell precursor acute lymphoblastic leukemia.
Blood
PUBLISHED: 07-29-2009
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We report 2 novel, cryptic chromosomal abnormalities in precursor B-cell acute lymphoblastic leukemia (BCP-ALL): a translocation, either t(X;14)(p22;q32) or t(Y;14)(p11;q32), in 33 patients and an interstitial deletion, either del(X)(p22.33p22.33) or del(Y)(p11.32p11.32), in 64 patients, involving the pseudoautosomal region (PAR1) of the sex chromosomes. The incidence of these abnormalities was 5% in childhood ALL (0.8% with the translocation, 4.2% with the deletion). Patients with the translocation were older (median age, 16 years), whereas the patients with the deletion were younger (median age, 4 years). The 2 abnormalities result in deregulated expression of the cytokine receptor, cytokine receptor-like factor 2, CRLF2 (also known as thymic stromal-derived lymphopoietin receptor, TSLPR). Overexpression of CRLF2 was associated with activation of the JAK-STAT pathway in cell lines and transduced primary B-cell progenitors, sustaining their proliferation and indicating a causal role of CRLF2 overexpression in lymphoid transformation. In Down syndrome (DS) ALL and 2 non-DS BCP-ALL cell lines, CRLF2 deregulation was associated with mutations of the JAK2 pseudokinase domain, suggesting oncogenic cooperation as well as highlighting a link between non-DS ALL and JAK2 mutations.
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Establishment and validation of a standard protocol for the detection of minimal residual disease in B lineage childhood acute lymphoblastic leukemia by flow cytometry in a multi-center setting.
Haematologica
PUBLISHED: 04-18-2009
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Minimal residual disease detection, used for clinical management of children with acute lymphoblastic leukemia, can be performed by molecular analysis of antigen-receptor gene rearrangements or by flow cytometric analysis of aberrant immunophenotypes. For flow minimal residual disease to be incorporated into larger national and international trials, a quality assured, standardized method is needed which can be performed in a multi-center setting. We report a four color, flow cytometric protocol established and validated by the UK acute lymphoblastic leukemia Flow minimal residual disease group. Quality assurance testing gave high inter-laboratory agreement with no values differing from a median consensus value by more than one point on a logarithmic scale. Prospective screening of B-ALL patients (n=206) showed the method was applicable to 88.3% of patients. The minimal residual disease in bone marrow aspirates was quantified and compared to molecular data. The combined risk category concordance (minimal residual disease levels above or below 0.01%) was 86% (n=134). Thus, this standardized protocol is highly reproducible between laboratories, sensitive, applicable, and shows good concordance with molecular-based analysis.
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Alternative splicing of hMLH1 in childhood acute lymphoblastic leukaemia and characterisation of the variably expressed Delta9/10 isoform as a dominant negative species.
Leuk. Res.
PUBLISHED: 04-15-2009
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Mismatch repair (MMR) deficiency is a common feature of acute lymphoblastic leukaemia (ALL) cell lines and in some cases is due to the mutations of hMLH1 which affect mRNA splicing. Therefore, we have analysed alternative splicing of hMLH1 in a cohort of children with ALL. We show that alternative splicing of hMLH1 is highly variable in normal and leukaemic cells and can occur by exon skipping or by the use of an alternative splice site, both serving to down-regulate the amount of full-length hMLH1 mRNA/protein produced. Aberrant splicing was found in one child with an aggressive leukaemia in which there was a predominant hMLH1Delta6 form and an associated loss of wild-type hMLH1 protein but this was not accompanied by microsatellite instability. Functional analysis of one of the most abundant spliced forms, hMLH1Delta9/10, was shown to have a significant dominant negative effect on the functionality of the MMR pathway but again was similarly expressed in ALL and normal cells.
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Loci on 7p12.2, 10q21.2 and 14q11.2 are associated with risk of childhood acute lymphoblastic leukemia.
Nat. Genet.
PUBLISHED: 04-08-2009
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To identify risk variants for childhood acute lymphoblastic leukemia (ALL), we conducted a genome-wide association study of two case-control series, analyzing the genotypes with respect to 291,423 tagging SNPs in a total of 907 ALL cases and 2,398 controls. We identified risk loci for ALL at 7p12.2 (IKZF1, rs4132601, odds ratio (OR) = 1.69, P = 1.20 x 10(-19)), 10q21.2 (ARID5B, rs7089424, OR = 1.65, P = 6.69 x 10(-19)) and 14q11.2 (CEBPE, rs2239633, OR = 1.34, P = 2.88 x 10(-7)). The 10q21.2 (ARID5B) risk association appears to be selective for the subset of B-cell precursor ALL with hyperdiploidy. These data show that common low-penetrance susceptibility alleles contribute to the risk of developing childhood ALL and provide new insight into disease causation of this specific hematological cancer. Notably, all three risk variants map to genes involved in transcriptional regulation and differentiation of B-cell progenitors.
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Microcystic stromal tumor of the ovary: report of 16 cases of a hitherto uncharacterized distinctive ovarian neoplasm.
Am. J. Surg. Pathol.
PUBLISHED: 03-26-2009
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We have encountered 16 ovarian neoplasms of probable stromal origin whose most distinctive feature is microcystic change, which is usually conspicuous. On the basis of our extensive experience with ovarian tumors, the neoplasm is unique and warrants separate categorization; we have elected to designate it "microcystic stromal tumor" because of its most striking feature. The patients ranged from 26 to 63 (mean 45) years of age and typically presented with a pelvic mass. Hormonal manifestations were possibly present in only 2. All tumors were unilateral with a mean size of 8.7 (range: 2 to 27) cm and none had evidence of extraovarian spread. The tumors were solid-cystic (11 cases), solid (3 cases), or predominantly cystic (2 cases). The solid component was usually firm and tan or white-tan, but in 1 case was yellowish; soft foci were present in 3 cases and small foci of hemorrhage, necrosis, or both, in 3. On microscopic examination the appearance of the tumors varied according to the relative prominence of their 3 fundamental components: microcysts, solid cellular regions, and fibrous stroma. Microcysts dominated in 9 cases, were roughly equal to noncystic morphology in 5 cases and were minor in 2. The microcystic pattern was characterized by small rounded to oval cystic spaces, in areas coalescing to larger irregular channels; intracytoplasmic vacuoles were also frequently present. The solid cellular areas were usually focally intersected by fibrous bands and hyaline plaques reminiscent of thecoma. The cells contained moderately conspicuous finely granular, lightly eosinophilic cytoplasm, with generally bland, round to oval or spindle-shaped nuclei with fine chromatin and small indistinct nucleoli. Foci of bizarre nuclei were, however, present in 10 cases. Mitotic rate was low in all cases, ranging from 0 to 2 mitoses/10 high-power fields. Immunohistochemical results were as follows: CD10, 16/16 cases positive; vimentin, 16/16 cases positive; inhibin, 1/16 cases weakly positive; calretinin, 1/16 cases positive; cytokeratin, 4/16 cases focally positive; and epithelial membrane antigen, 0/16 cases positive. Microcystic change can be observed in a wide variety of ovarian tumors and the broad potential differential diagnosis is discussed in the text. For tumors which have been well sampled and exhibit (1) a microcystic pattern and regions with lobulated cellular masses with intervening, sometimes hyalinized fibrous stroma, (2) an absence of morphologic features enabling any other specific diagnosis in the sex cord-stromal category, (3) an absence of epithelial elements, and (4) an absence of teratomatous or other germ cell elements, we propose the designation "microcystic stromal tumor." The characteristic immunophenotype is CD10/vimentin+/epithelial membrane antigen-, with focal cytokeratin-positivity in one-quarter of cases; inhibin and/or calretinin are usually negative. Seven patients with available follow-up are without evidence of disease at a mean of 4.25 years (range: 1.5 to 12.5 y) from the time of initial diagnosis. These tumors, to date, have occurred over a wide age range in postpubertal females, are characteristically unilateral, and confined to the ovary at presentation. They represent, in addition to the sclerosing stromal tumor (segregated out 3 decades ago), a distinctive subtype of ovarian tumor, likely also belonging to the stromal category based on current evidence.
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MMSET deregulation affects cell cycle progression and adhesion regulons in t(4;14) myeloma plasma cells.
Haematologica
PUBLISHED: 02-13-2009
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The recurrent immunoglobulin translocation, t(4;14)(p16;q32) occurs in 15% of multiple myeloma patients and is associated with poor prognosis, through an unknown mechanism. The t(4;14) up-regulates fibroblast growth factor receptor 3 (FGFR3) and multiple myeloma SET domain (MMSET) genes. The involvement of MMSET in the pathogenesis of t(4;14) multiple myeloma and the mechanism or genes deregulated by MMSET upregulation are still unclear.
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A comprehensive analysis of the CDKN2A gene in childhood acute lymphoblastic leukemia reveals genomic deletion, copy number neutral loss of heterozygosity, and association with specific cytogenetic subgroups.
Blood
PUBLISHED: 02-07-2009
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Inactivation of the tumor suppressor gene, CDKN2A, can occur by deletion, methylation, or mutation. We assessed the principal mode of inactivation in childhood acute lymphoblastic leukemia (ALL) and frequency in biologically relevant subgroups. Mutation or methylation was rare, whereas genomic deletion occurred in 21% of B-cell precursor ALL and 50% of T-ALL patients. Single nucleotide polymorphism arrays revealed copy number neutral (CNN) loss of heterozygosity (LOH) in 8% of patients. Array-based comparative genomic hybridization demonstrated that the mean size of deletions was 14.8 Mb and biallelic deletions composed a large and small deletion (mean sizes, 23.3 Mb and 1.4 Mb). Among 86 patients, only 2 small deletions were below the resolution of detection by fluorescence in situ hybridization. Patients with high hyperdiploidy, ETV6-RUNX1, or 11q23/MLL rearrangements had low rates of deletion (11%, 15%, 13%), whereas patients with t(9;22), t(1;19), TLX3, or TLX1 rearrangements had higher frequencies (61%, 42%, 78%, and 89%). In conclusion, CDKN2A deletion is a significant secondary abnormality in childhood ALL strongly correlated with phenotype and genotype. The variation in the incidence of CDKN2A deletions by cytogenetic subgroup may explain its inconsistent association with outcome. CNN LOH without apparent CDKN2A inactivation suggests the presence of other relevant genes in this region.
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Cultivating mindfulness in health care professionals: a review of empirical studies of mindfulness-based stress reduction (MBSR).
Complement Ther Clin Pract
PUBLISHED: 01-12-2009
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Demands faced by health care professionals include heavy caseloads, limited control over the work environment, long hours, as well as organizational structures and systems in transition. Such conditions have been directly linked to increased stress and symptoms of burnout, which in turn, have adverse consequences for clinicians and the quality of care that is provided to patients. Consequently, there exists an impetus for the development of curriculum aimed at fostering wellness and the necessary self-care skills for clinicians. This review will examine the potential benefits of mindfulness-based stress reduction (MBSR) programs aimed at enhancing well-being and coping with stress in this population. Empirical evidence indicates that participation in MBSR yields benefits for clinicians in the domains of physical and mental health. Conceptual and methodological limitations of the existing studies and suggestions for future research are discussed.
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Methylation of tumour suppressor gene promoters in the presence and absence of transcriptional silencing in high hyperdiploid acute lymphoblastic leukaemia.
Br. J. Haematol.
PUBLISHED: 01-06-2009
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Promoter methylation is a common phenomenon in tumours, including haematological malignancies. In the present study, we investigated 36 cases of high hyperdiploid (>50 chromosomes) acute lymphoblastic leukaemia (ALL) with methylation-specific multiplex ligase-dependent probe amplification to determine the extent of aberrant methylation in this subgroup. The analysis, which comprised the promoters of 35 known tumour suppressor genes, showed that 16 genes displayed abnormal methylation in at least one case each. The highest number of methylated gene promoters seen in a single case was thirteen, with all but one case displaying methylation for at least one gene. The most common targets were ESR1 (29/36 cases; 81%), CADM1 (IGSF4, TSLC1; 25/36 cases; 69%), FHIT (24/36 cases; 67%) and RARB (22/36 cases; 61%). Interestingly, quantitative reverse transcription-polymerase chain reaction showed that although methylation of the CADM1 and RARB promoters resulted in the expected pattern of downregulation of the respective genes, no difference could be detected in FHIT expression between methylation-positive and -negative cases. Furthermore, TIMP3 was not expressed regardless of methylation status, showing that aberrant methylation does not always lead to gene expression changes. Taken together, our findings suggest that aberrant methylation of tumour suppressor gene promoters is a common phenomenon in high hyperdiploid ALL.
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Acute B lymphoblastic leukaemia-propagating cells are present at high frequency in diverse lymphoblast populations.
EMBO Mol Med
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Leukaemia-propagating cells are more frequent in high-risk acute B lymphoblastic leukaemia than in many malignancies that follow a hierarchical cancer stem cell model. It is unclear whether this characteristic can be more universally applied to patients from non-high-risk sub-groups and across a broad range of cellular immunophenotypes. Here, we demonstrate in a wide range of primary patient samples and patient samples previously passaged through mice that leukaemia-propagating cells are found in all populations defined by high or low expression of the lymphoid differentiation markers CD10, CD20 or CD34. The frequency of leukaemia-propagating cells and their engraftment kinetics do not differ between these populations. Transcriptomic analysis of CD34(high) and CD34(low) blasts establishes their difference and their similarity to comparable normal progenitors at different stages of B-cell development. However, consistent with the functional similarity of these populations, expression signatures characteristic of leukaemia propagating cells in acute myeloid leukaemia fail to distinguish between the different populations. Together, these findings suggest that there is no stem cell hierarchy in acute B lymphoblastic leukaemia.
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YWHAE-FAM22 endometrial stromal sarcoma: diagnosis by reverse transcription-polymerase chain reaction in formalin-fixed, paraffin-embedded tumor.
Hum. Pathol.
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A subset of endometrial stromal sarcoma harbors t(10;17)(q23;p13), which results in the genetic fusion between YWHAE and 1 of 2 highly homologous FAM22 family members-FAM22A or FAM22B. In contrast to classic low-grade endometrial stromal sarcoma with JAZF1-SUZ12 fusions, YWHAE-FAM22 endometrial stromal sarcoma displays high-grade histologic features and is associated with more aggressive disease course. Ancillary fluorescence in situ hybridization assay demonstrating the presence of YWHAE rearrangement can be used to support the diagnosis, but the detection of fusion transcript would be the most definitive test. We describe here an optimized reverse transcription-polymerase chain reaction assay for detection of YWHAE-FAM22 fusion transcript in formalin-fixed and paraffin-embedded tumor samples. We studied a series of 6 YWHAE-FAM22 endometrial stromal sarcomas, 7 JAZF-SUZ12 endometrial stromal sarcomas, 3 JAZF1-PHF1/EPC1-PHF1 endometrial stromal sarcomas, 6 undifferentiated endometrial sarcomas, 4 uterine leiomyosarcomas, and 4 uterine adenosarcomas. All 6 YWHAE-FAM22 endometrial stromal sarcomas were confirmed by fluorescence in situ hybridization assay, whereas all non-YWHAE-FAM22 tumors were confirmed to lack YWHAE rearrangement by fluorescence in situ hybridization assay. The reverse transcription-polymerase chain reaction assay optimized for formalin-fixed and paraffin-embedded samples detected YWHAE-FAM22 fusion transcripts in all 6 YWHAE-FAM22 endometrial stromal sarcomas and none of the 24 non-YWHAE-FAM22 uterine sarcomas. These findings show that this reverse transcription-polymerase chain reaction assay is sensitive and specific for detection of YWHAE-FAM22 fusion transcript and can serve as a useful adjunct diagnostic assay to confirm the diagnosis of YWHAE-FAM22 endometrial stromal sarcoma in formalin-fixed and paraffin-embedded tumor samples.
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The diagnosis of endometrial carcinomas with clear cells by gynecologic pathologists: an assessment of interobserver variability and associated morphologic features.
Am. J. Surg. Pathol.
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The purposes of this study are to assess the level of interobserver variability in the diagnosis of endometrial carcinomas with clear cells by gynecologic pathologists based purely on their morphologic features and to comparatively describe the cases of putative clear cell carcinoma (CCC) with and without significant interobserver variability. A total of 35 endometrial carcinomas (1 slide per case) were reviewed by 11 gynecologic pathologists (median experience: 10 y) from 11 North American institutions. The cases were selected from the files of 3 institutions on the basis of the presence of at least focal clear cells and had previously been classified as a variety of histotypes at these institutions. Diagnoses were rendered in a blinded manner and without predetermined diagnostic criteria or categories. The ? values between any pair of observers ranged from 0.18 to 0.69 (combined 0.46), which was indicative of a "moderate" level of interobserver agreement for the group. Subgroups of "confirmed CCC" [cases diagnosed as such by at least 8 (73%) of the 11 observers, n=14] and "possible CCC" (cases diagnosed as CCC by ?1 but <8 observers, n=13) were compared with regard to a variety of semiquantified morphologic features. By combining selected morphologic features that displayed statistically significant differences between the 2 groups on univariate analyses, the following approximate morphologic profile emerged for the confirmed CCC group: papillae with hyalinized cores in ?33% of the lesion, clear cells in ?33% of the lesion, hyperchromasia in ?33% of the lesion, the absence of nuclear pseudostratification in >3 cells on the papillae, the absence of nuclear pseudostratification in glands in ?33% of the lesion, the absence of diffuse grade 3 nuclei, the absence of long and slender papillae in ?33% of the lesion, and glands and papillae lined by cuboidal to flat, noncolumnar cells. In a backward stepwise logistic regression analysis, features from the profile that predicted the confirmed CCC group included: (1) absence or minimality of diffuse sheets of grade 3 nuclei [P=0.025; 95% confidence interval (CI), 0.0266-0.363]; (2) absence or minimality of nuclear stratification in glands and papillae (P=0.040; 95% CI, -0.228 to -0.0054); and (3) glands and papillae lined by cuboidal to flat, noncolumnar cells (P=0.008; 95% CI, 0.0911-0.566). The 2 groups displayed significant overlap regarding a wide variety of features, and no single case displayed a full complement of potentially diagnostic features. Morphologic patterns associated with cases with very high levels of interobserver variability (defined as cases with ?4 different diagnoses rendered for them, n=9) included the near-exclusive or exclusively solid pattern of clear cells (3/9) and glandular/papillary proliferations whose only CCC-like feature was the presence of clear cells (2/9). In conclusion, the diagnosis of endometrial carcinomas with clear cells by gynecologic pathologists is associated with a moderate level of interobserver variability. However, there is a morphologic profile that characterizes cases that gynecologic pathologists more uniformly classify as CCC, and the presence of these features is supportive of a CCC diagnosis in an endometrial carcinoma with clear cells. Cases that display broad and significant qualitative deviations from the aforementioned profile should prompt the consideration of a diagnosis other than CCC.
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