Expression of TCF3 target genes defines a subclass of diffuse large B-cell lymphoma characterized by up-regulation of MYC target genes and poor clinical outcome following R-CHOP therapy.

TCF3 is a lymphopoietic transcription factor that acquires somatic driver mutations in diffuse large B-cell lymphoma (DLBCL). Hypothesizing that expression patterns of TCF3-regulated genes can inform clinical management, we found that unsupervised clustering analysis with 15 TCF3-regulated genes and eight additional ones resolved local DLBCL cases into two main clusters, denoted Groups A and B, of which Group A manifested inferior overall survival (OS, p = 0.0005). We trained a machine learning model to classify samples into the Groups based on expression of the 23 transcripts in an independent validation cohort of 569 R-CHOP-treated DLBCL cases. Group A overlapped with the ABC cell-of-origin subgroup but its prognostic power was superior. GSEA analysis demonstrated asymmetric expression of 30 gene sets between the Groups, pointing to biological differences. We present, validate and make available a novel method to assign DLBCL cases into biologically-distinct groups with divergent OS following R-CHOP therapy.

Evidence-based review of genomic aberrations in diffuse large B cell lymphoma, not otherwise specified (DLBCL, NOS): Report from the cancer genomics consortium lymphoma working group.

Diffuse large B cell lymphoma, not otherwise specified (DLBCL, NOS) is the most common type of non-Hodgkin lymphoma (NHL). The 2016 World Health Organization (WHO) classification defined DLBCL, NOS and its subtypes based on clinical findings, morphology, immunophenotype, and genetics. However, even within the WHO subtypes, it is clear that additional clinical and genetic heterogeneity exists. Significant efforts have been focused on utilizing advanced genomic technologies to further subclassify DLBCL, NOS into clinically relevant subtypes. These efforts have led to the implementation of novel algorithms to support optimal risk-oriented therapy and improvement in the overall survival of DLBCL patients. We gathered an international group of experts to review the current literature on DLBCL, NOS, with respect to genomic aberrations and the role they may play in the diagnosis, prognosis and therapeutic decisions. We comprehensively surveyed clinical laboratory directors/professionals about their genetic testing practices for DLBCL, NOS. The survey results indicated that a variety of diagnostic approaches were being utilized and that there was an overwhelming interest in further standardization of routine genetic testing along with the incorporation of new genetic testing modalities to help guide a precision medicine approach. Additionally, we present a comprehensive literature summary on the most clinically relevant genomic aberrations in DLBCL, NOS. Based upon the survey results and literature review, we propose a standardized, tiered testing approach which will help laboratories optimize genomic testing in order to provide the maximum information to guide patient care.

Validation, Implementation, and Clinical Impact of the Oncomine Myeloid Targeted-Amplicon DNA and RNA Ion Semiconductor Sequencing Assay.

The identification of clinically significant genes recurrently mutated in myeloid malignancies necessitates expanding diagnostic testing with higher throughput, such as targeted next-generation sequencing. We present validation of the Thermo Fisher Oncomine Myeloid Next-Generation Sequencing Panel (OMP), targeting 40 genes and 29 fusion drivers recurrently mutated in myeloid malignancies. The study includes data from a sample exchange between two Canadian hospitals demonstrating high concordance for detection of DNA and RNA aberrations. Clinical validation demonstrates high accuracy, sensitivity, and specificity of the OMP, with a lower limit of detection of 5% for single-nucleotide variants and 10% for insertions/deletions. Prospective sequencing was performed for 187 samples from 168 unique patients presenting with suspected or confirmed myeloid malignancy and other hematological conditions to assess clinical impact of identifying variants. Of detected variants, 48% facilitated or clarified diagnoses, 29% affected prognoses, and 25% had the potential to influence clinical management. Of note, OMP was essential to identifying patients with premalignant clonal states likely contributing to cytopenias. We also found that the detection of even a single variant by the OMP assay, versus 0 variants, was predictive of overall survival, independent of age, sex, or diagnosis (P = 0.03). This study demonstrates that molecular profiling of myeloid malignancies with the OMP represents a promising strategy to advance molecular diagnostics.

Objective quantification of BCL2 protein by multiplex immunofluorescence in routine biopsy samples of diffuse large B-cell lymphoma demonstrates associations with survival and BCL2 gene alterations.

Up-regulation of BCL2 in cases of diffuse large B-cell lymphoma (DLBCL) can confer treatment resistance. Quantitative immunofluorescence (QIF) histology allows objective quantification of protein-based biomarkers. We investigated the utility of QIF for evaluating BCL2 as a biomarker in DLBCL by quantifying BCL2 selectively in CD20-expressing lymphoma cells in biopsy samples from 116 cases of DLBCL in two cohorts one of which consisted of relapsed/refractory cases from a clinical trial. BCL2 protein by QIF correlated with BCL2 mRNA abundance and was associated with both translocation and copy number gain of the BCL2 gene. Elevated BCL2 protein expression by QIF, but not immunohistochemistry or mRNA quantification, was associated with inferior overall and relapse-free survival in the relapsed/refractory cohort. QIF is an effective means of quantifying BCL2 protein objectively in routine cancer biopsy specimens and shows promise for identifying relapsed/refractory DLBCL patients at risk of inferior outcomes after salvage therapy.

Simultaneous detection of lung fusions using a multiplex RT-PCR next generation sequencing-based approach: a multi-institutional research study.

BACKGROUND: Gene fusion events resulting from chromosomal rearrangements play an important role in initiation of lung adenocarcinoma. The recent association of four oncogenic driver genes, ALK, ROS1, RET, and NTRK1, as lung tumor predictive biomarkers has increased the need for development of up-to-date technologies for detection of these biomarkers in limited amounts of material. METHODS: We describe here a multi-institutional study using the Ion AmpliSeq™ RNA Fusion Lung Cancer Research Panel to interrogate previously characterized lung tumor samples. RESULTS: Reproducibility between laboratories using diluted fusion-positive cell lines was 100%. A cohort of lung clinical research samples from different origins (tissue biopsies, tissue resections, lymph nodes and pleural fluid samples) were used to evaluate the panel. We observed 97% concordance for ALK (28/30 positive; 71/70 negative samples), 95% for ROS1 (3/4 positive; 19/18 negative samples), and 93% for RET (2/1 positive; 13/14 negative samples) between the AmpliSeq assay and other methodologies. CONCLUSION: This methodology enables simultaneous detection of multiple ALK, ROS1, RET, and NTRK1 gene fusion transcripts in a single panel, enhanced by an integrated analysis solution. The assay performs well on limited amounts of input RNA (10 ng) and offers an integrated single assay solution for detection of actionable fusions in lung adenocarcinoma, with potential savings in both cost and turn-around-time compared to the combination of all four assays by other methods.

High-grade B-cell lymphoma with MYC and BCL2 rearrangements arising in a composite lymphoma.

BACKGROUND: We report the first case of composite lymphoma consisting of chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL), follicular lymphoma (FL) and high-grade B-cell lymphoma with MYC and BCL2 rearrangements within the same needle biopsy in which a clonal relationship between the FL and high-grade B-cell lymphoma components was demonstrated by molecular cytogenetics. CASE PRESENTATION: An 85-year-old man presented with masses in his neck and right groin. Cutting needle biopsy of the inguinal mass revealed the three lymphoma types which were morphologically, immunophenotypically and topographically distinct. Fluorescence in situ hybridization (FISH) identified an IGH-BCL2 rearrangement in both the FL and high-grade B-cell components while a MYC rearrangement was detected in the high-grade B-cell component alone. CONCLUSIONS: Our findings suggest that the high-grade lymphoma with MYC and BCL2 translocations evolved through transformation of the FL by a process that entailed acquisition of the MYC translocation. No clonal relationship between the FL and CLL/SLL components was evident since the IGH-BCL2 rearrangement was present in in the former but not the latter. This unique case of co-localized FL, CLL/SLL, and high-grade B-cell lymphoma contributes to our understanding of the clonal relationships that may exist between the components of composite lymphomas.

Noninvasive Prenatal Detection of Trisomy 21 by Targeted Semiconductor Sequencing: A Technical Feasibility Study.

OBJECTIVE: To develop an alternate noninvasive prenatal testing method for the assessment of trisomy 21 (T21) using a targeted semiconductor sequencing approach. METHODS: A customized AmpliSeq panel was designed with 1,067 primer pairs targeting specific regions on chromosomes 21, 18, 13, and others. A total of 235 samples, including 30 affected with T21, were sequenced with an Ion Torrent Proton sequencer, and a method was developed for assessing the probability of fetal aneuploidy via derivation of a risk score. RESULTS: Application of the derived risk score yields a bimodal distribution, with the affected samples clustering near 1.0 and the unaffected near 0. For a risk score cutoff of 0.345, above which all would be considered at "high risk," all 30 T21-positive pregnancies were correctly predicted to be affected, and 199 of the 205 non-T21 samples were correctly predicted. The average hands-on time spent on library preparation and sequencing was 19 h in total, and the average number of reads of sequence obtained was 3.75 million per sample. CONCLUSION: With the described targeted sequencing approach on the semiconductor platform using a custom-designed library and a probabilistic statistical approach, we have demonstrated the feasibility of an alternate method of assessment for fetal T21.

The CDK inhibitor AT7519M in patients with relapsed or refractory chronic lymphocytic leukemia (CLL) and mantle cell lymphoma. A Phase II study of the Canadian Cancer Trials Group.

AT7519M is a small molecule inhibitor of cyclin-dependent kinases 1, 2, 4, 5, and 9 with in vitro activity against lymphoid malignancies. In two concurrent Phase II trials, we evaluated AT7519M in relapsed or refractory chronic lymphocytic leukemia (CLL) and mantle cell lymphoma (MCL) using the recommended Phase II dosing of 27 mg/m2 twice weekly for 2 of every 3 weeks. Primary objective was objective response rate (ORR). Nineteen patients were accrued (7 CLL, 12 MCL). Four CLL patients achieved stable disease (SD). Two MCL patients achieved partial response (PR), and 6 had SD. One additional MCL patient with SD subsequently achieved PR 9 months after completion of AT7519M. Tumor lysis syndrome was not reported. In conclusion, AT7519M was safely administered to patients with relapsed/refractory CLL and MCL. In CLL, some patients had tumor reductions, but the ORR was low. In MCL, activity was noted with ORR of 27%.

X-linked deafness-2 (DFNX2) phenotype associated with a paracentric inversion upstream of POU3F4.

PURPOSE: The authors report on a 7-year-old male, designated FR, who has severe sensorineural hearing loss. Features include a round face, hypertelorism, epicanthal folds, and flat nasal root. Although there were early developmental concerns regarding FR, all but his speech delay resolved when he was placed in an educational program that accommodated his hearing loss. Genetic studies were performed to investigate genetic causes for his hearing loss. METHOD: History, physical examination, audiologic assessment, and imaging were performed according to usual practice. FMR1,GJB2,GJB6, and POU3F4 genes were sequenced. Chromosomal studies consisted of karyotyping and breakpoint analysis by fluorescence in situ hybridization (FISH). RESULTS: Results from FMR1,GJB2,GJB6, and POU3F4 sequencing and echocardiography, electrocardiogram, and abdominal ultrasound were normal. A computed tomography (CT) scan revealed a large fundus of the internal auditory canals and absence of the bony partition between the fundus and the adjacent cochlear turns, with a widened modiolus bilaterally. FR's CT findings were consistent with those described in persons with X-linked deafness-2 (DFNX2) hereditary deafness. FR's karyotype was 46,inv(X)(q13q24),Y.ish inv(X)(XIST+)mat. FISH refined the breakpoints to inv(X)(q21.1q22.3). The Xq21.1 breakpoint was narrowed to a 25-kb region 450 kb centromeric to the DFNX2 gene, POU3F4. There are rare case reports of DFNX2 patients with chromosomal rearrangements positioned centromeric to POU3F4 and no mutations within the gene. CONCLUSION: Authors hypothesized that FR's hearing loss was caused by dysregulation of POU3F4 due to separation from regulatory elements affected by the inversion.

A framework for assessing global change risks to forest carbon stocks in the United States.

Among terrestrial environments, forests are not only the largest long-term sink of atmospheric carbon (C), but are also susceptible to global change themselves, with potential consequences including alterations of C cycles and potential C emission. To inform global change risk assessment of forest C across large spatial/temporal scales, this study constructed and evaluated a basic risk framework which combined the magnitude of C stocks and their associated probability of stock change in the context of global change across the US. For the purposes of this analysis, forest C was divided into five pools, two live (aboveground and belowground biomass) and three dead (dead wood, soil organic matter, and forest floor) with a risk framework parameterized using the US's national greenhouse gas inventory and associated forest inventory data across current and projected future Köppen-Geiger climate zones (A1F1 scenario). Results suggest that an initial forest C risk matrix may be constructed to focus attention on short- and long-term risks to forest C stocks (as opposed to implementation in decision making) using inventory-based estimates of total stocks and associated estimates of variability (i.e., coefficient of variation) among climate zones. The empirical parameterization of such a risk matrix highlighted numerous knowledge gaps: 1) robust measures of the likelihood of forest C stock change under climate change scenarios, 2) projections of forest C stocks given unforeseen socioeconomic conditions (i.e., land-use change), and 3) appropriate social responses to global change events for which there is no contemporary climate/disturbance analog (e.g., severe droughts in the Lake States). Coupling these current technical/social limits of developing a risk matrix to the biological processes of forest ecosystems (i.e., disturbance events and interaction among diverse forest C pools, potential positive feedbacks, and forest resiliency/recovery) suggests an operational forest C risk matrix remains elusive.

Nablus mask-like facial syndrome: deletion of chromosome 8q22.1 is necessary but not sufficient to cause the phenotype.

Nablus mask-like facial syndrome (NMLFS) has many distinctive phenotypic features, particularly tight glistening skin with reduced facial expression, blepharophimosis, telecanthus, bulky nasal tip, abnormal external ear architecture, upswept frontal hairline, and sparse eyebrows. Over the last few years, several individuals with NMLFS have been reported to have a microdeletion of 8q21.3q22.1, demonstrated by microarray analysis. The minimal overlapping region is 93.98-96.22 Mb (hg19). Here we present clinical and microarray data from five singletons and two mother-child pairs who have heterozygous deletions significantly overlapping the region associated with NMLFS. Notably, while one mother and child were said to have mild tightening of facial skin, none of these individuals exhibited reduced facial expression or the classical facial phenotype of NMLFS. These findings indicate that deletion of the 8q21.3q22.1 region is necessary but not sufficient for development of the NMLFS. We discuss possible genetic mechanisms underlying the complex pattern of inheritance for this condition.

HLA-DR(negative), CD34(negative) hypergranular acute myeloid leukemia with trisomy 6 and del(5)(q22q33): case report and review of the literature.

We report a unique pediatric case of hypergranular acute myeloid leukemia with myelodysplasia-related changes. The patient presented with moderate leukocytosis with neutrophilia with left-shift maturation and dysplasia, anemia, and multiple sclerotic bone lesions. The bone marrow was hypercellular with a predominance of myeloblast cells and/or abnormal promyelocytes with hypergranular cytoplasm. Flow cytometric immunophenotyping showed that the leukemic cells were positive for CD13, CD33, and myeloperoxidase, and negative for HLA-DR and CD34. Morphology and immunophenotyping were highly suggestive of acute promyelocytic leukemia. The classic t(15;17) or other RARα rearrangements were not detected by cytogenetic or molecular assays, ruling out acute promyelocytic leukemia. Standard cytogenetic analysis showed that the karyotype of the predominant clone was 47,XY,+6 with evidence of clonal evolution to 47,XY,+6,del(5)(q22q33). A literature and database review showed that trisomy 6 is a rare occurrence in hematological malignancies and, to our knowledge, has never been reported in association with del(5)(q22q33) in a child presenting with hypergranular acute myeloid leukemia with myelodysplasia-related changes. We present a current review of the literature and summarize the clinical features of 57 cases of trisomy 6 as the primary chromosomal abnormality in hematological disease.

Real food matters for health.

Human health, an enigma that has been a part of human existence from its origins, is founded on the intake of food. Holistic health is concerned with issues of the body, soul/mind, and spirit. Using concepts from ecology, this article explores the relationships between food, nutrients, whole food diets, and health. An analysis of studies linking whole food to health and depressive disorders are presented in an attempt to move food and diets into mainstream mental health discussions. The purpose of this article is to demonstrate that real food matters for the health of all and is of vital consequence for improving the chronic conditions plaguing modern society's mental health.

Expert to novice: clinicians learning new roles as clinical nurse educators.

The nursing shortage presents a challenge for both nursing education programs and clinical agencies to identify creative options for increasing the supply of nurse educators, especially those who can teach vital clinical skills to future nurses. Some clinicians have voiced that they would enjoy having opportunities to share their clinical expertise through teaching, but they lack preparation for this new role. Through written narratives of nurses enrolled in a Clinical Nurse Educator Academy, this study explored the research question: How can narratives of clinicians participating in a Clinical Nurse Educator Academy enhance understanding of the unique perspectives of these individuals as they prepare for roles as clinical nurse educators? Data consisted of 135 reflective narratives from 45 participants and were analyzed using van Manen's approach to human science research. One overarching pattern, "The Phenomenon of Learning to Teach:" and three themes,"Buckle Your Seatbelt," "Embracing the Novice," and "Mentoring in the Dark," emerged from the data. Implications for faculty and further research are offered.

Penetrating the dark silence.

This study articulates women's lifeworld experience of unexplained and unexpected fatigue. Interpretive phenomenology situated within the perspective of Maurice Merleau-Ponty provides the study framework. Using purposive sampling, the study investigated the unexplained fatigue of 12 women in a southeastern state. Data analysis of interviews revealed a lost voice within the phenomenon of fatigue, providing new insights into this diffuse and dehumanizing experience. Ethical considerations are addressed for recognizing, hearing, and caring for women living with disabling fatigue to extend the goals of Healthy People 2010 by addressing new ethical priorities for quality of life over the next decade.

Choosing to eat your way toward healthy aging.

As individuals age, they face many choices to promote healthy aging. One important consideration relates to nutritional choices. It is easy to become overwhelmed with the information in the media that suggests the superiority of some new diet or breakthrough dietary supplement. This article reminds us to focus on holistic approaches to nutrition on the premise that balance is paramount in our lives. As nurses, we need to guide others to make thoughtful choices for healthy aging.