Gain of Chromosome 5q Predicts a Favorable Prognosis in Localized Renal Cell Carcinoma.

Approximately 65% of renal cell carcinomas (RCC) are diagnosed at a localized stage. We investigated the chromosome 5q gain impact on disease-free survival (DFS) in RCC patients. Overall, 676 patients with stages 1-2 RCC and having cytogenetic analysis were included. Gain of 5q was observed in 108 patients, more frequently in clear cell (ccRCC) than non-clear cell tumors. Gain of 5q is likely an independent prognostic factor since the concerned patients had a decreased recurrence risk in stages 1-2 RCC, confirmed in multivariable analysis. Detecting 5q gain could enhance recurrence risk assessment, allowing tailored post-surgery surveillance, and reducing unnecessary treatments.

A molecular interactome of the glioblastoma perivascular niche reveals integrin binding sialoprotein as a mediator of tumor cell migration.

Glioblastoma (GBM) is characterized by extensive microvascular hyperproliferation. In addition to supplying blood to the tumor, GBM vessels also provide trophic support to glioma cells and serve as conduits for migration into the surrounding brain, promoting recurrence. Here, we enrich CD31-expressing glioma vascular cells (GVCs) and A2B5-expressing glioma tumor cells (GTCs) from primary GBM and use RNA sequencing to create a comprehensive molecular interaction map of the secreted and extracellular factors elaborated by GVCs that can interact with receptors and membrane molecules on GTCs. To validate our findings, we utilize functional assays, including a hydrogel-based migration assay and in vivo mouse models to demonstrate that one identified factor, the little-studied integrin binding sialoprotein (IBSP), enhances tumor growth and promotes the migration of GTCs along the vasculature. This perivascular niche interactome will serve as a resource to the research community in defining the potential functions of the GBM vasculature.

Comparison of laboratory methods for the detection of neoplastic plasma cells in plasma cell dyscrasias.

AIMS: To compare the ability of immunohistochemistry (IHC), multiparameter flow cytometry (MFC) and fluorescence in situ hybridisation (FISH) to detect clonal plasma cells. We also attempted to outline a testing strategy for monitoring multiple myeloma patients. METHODS: A retrospective review was performed on 278 CD138+sorted FISH studies from November 2019 to December 2020 along with their concurrent IHC and MFC results. A p value was computed using McNemar's test for paired data. Association was calculated using the non-parametric Spearman correlation coefficient. RESULTS: Using the Mc Nemar's test for paired data, CD138+sorted FISH studies achieved the highest proportion of positive results and was significantly greater than MFC (63% vs 53%, p=0.01). FISH had more positive results than IHC, although this did not reach statistical significance (60% vs 57%, p=0.34). IHC and MFC had high correlation and high agreement (90.3% agreement, kappa=0.805, p<0.0001). CD138+sorted FISH studies achieved the highest proportion of positive results relative to IHC and MFC, indicating that it may be a reliable marker for clonal plasma cell detection. CONCLUSIONS: While CD138+sorted FISH is primarily used for prognostication, it may be employed as a single test for detection and monitoring clonality in certain scenarios. Further studies are needed to monitor the outcomes of patients with positive FISH and negative IHC and MFC. Additionally, there was high agreement between IHC and MFC, suggesting that performing both tests may not be necessary.

Post-irradiation pericardial malignant mesothelioma with deletion of p16: a case report.

Malignant mesotheliomas are rather uncommon neoplasms associated primarily with asbestos exposure; however, they may also arise as second primary malignancies after radiation therapy, with a latency period of 15-25 years. Numerous studies have reported an association between pleural malignant mesothelioma and chest radiation performed for other malignancies; on the other hand, post-irradiation mesotheliomas of the pericardium have been reported in only a few published cases to date, and no homozygous deletion of 9p21 has been described in such cases. We report the case of a 48-year-old man with a history of Hodgkin's lymphoma and no prior asbestos exposure who developed pericardial malignant epithelioid mesothelioma. We further discuss the cytologic, histologic, immunophenotypic, and fluorescence in situ hybridization findings in this case. To our knowledge, this is the first well-documented case of post-radiation pericardial malignant mesothelioma showing homozygous deletion of 9p21. Homozygous deletion of 9p21, the locus harboring the p16 gene, is present in post-irradiation pericardial malignant mesothelioma.

A rapid and reliable chromosome analysis method for products of conception using interphase nuclei.

BACKGROUND: Karyotype determination has a central role in the genetic workup of pregnancy loss, as aneuploidy (trisomy and monosomy) and polyploidy (triploidy and tetraploidy) are the cause in at least 50% of first trimester, 25% of second trimester, and 11% of third trimester miscarriages. There are several limitations with the current approaches of obtaining a karyotype using traditional cytogenetics, fluorescence in situ hybridization with a limited number of probes, and chromosomal microarray. These include culture failure, incomplete results, lower sensitivity, and longer reporting time. METHODS: To overcome current limitations, a novel molecular assay is developed with a Standard Resolution Interphase Chromosome Profiling probe set which is a variation of the recently developed High Resolution probe set. It generates a molecular karyotype that can detect all major changes commonly associated with pregnancy loss. Initial familiarization of signal patterns from the probe set was used, followed by validation of the method using 83 samples from miscarriages in a blind study from three different laboratories. Finally, the clinical utility of the method was tested on 291 clinical samples in two commercial reference laboratory settings on two different continents. RESULTS: The new molecular approach not only identified all the chromosome changes observed by current methods, but also significantly improved abnormality detection by characterizing derivative chromosomes and finding subtle subtelomeric rearrangements, balanced and unbalanced. All Robertsonian translocations were also detected. The abnormality rate was 54% on clinical samples from commercial laboratory 1 and 63% from laboratory 2. CONCLUSION: The attributes of this method make it an ideal choice for the genetic workup of miscarriages, namely (1) near 100% successful results, (2) greater sensitivity than conventional chromosome analysis or FISH panels, (3) rapid reporting time, and (4) favorable comparisons with chromosomal microarray.

Interphase Chromosome Profiling: A Method for Conventional Banded Chromosome Analysis Using Interphase Nuclei.

CONTEXT: - Chromosome analysis on bone marrow or peripheral blood samples fails in a small proportion of attempts. A method that is more reliable, with similar or better resolution, would be a welcome addition to the armamentarium of the cytogenetics laboratory. OBJECTIVE: - To develop a method similar to banded metaphase chromosome analysis that relies only on interphase nuclei. DESIGN: - To label multiple targets in an equidistant fashion along the entire length of each chromosome, including landmark subtelomere and centromere regions. Each label so generated by using cloned bacterial artificial chromosome probes is molecularly distinct with unique spectral characteristics, so the number and position of the labels can be tracked to identify chromosome abnormalities. RESULTS: - Interphase chromosome profiling (ICP) demonstrated results similar to conventional chromosome analysis and fluorescence in situ hybridization in 55 previously studied cases and obtained useful ICP chromosome analysis results on another 29 cases in which conventional methods failed. CONCLUSIONS: - ICP is a new and powerful method to karyotype peripheral blood and bone marrow aspirate preparations without reliance on metaphase chromosome preparations. It will be of particular value for cases with a failed conventional analysis or when a fast turnaround time is required.

De novo loss-of-function variants in STAG2 are associated with developmental delay, microcephaly, and congenital anomalies.

The cohesin complex is an evolutionarily conserved multi-subunit protein complex which regulates sister chromatid cohesion during mitosis and meiosis. Additionally, the cohesin complex regulates DNA replication, DNA repair, and transcription. The core of the complex consists of four subunits: SMC1A, SMC3, RAD21, and STAG1/2. Loss-of-function mutations in many of these proteins have been implicated in human developmental disorders collectively termed "cohesinopathies." Through clinical exome sequencing (CES) of an 8-year-old girl with a clinical history of global developmental delay, microcephaly, microtia with hearing loss, language delay, ADHD, and dysmorphic features, we describe a heterozygous de novo variant (c.205C>T; p.(Arg69*)) in the integral cohesin structural protein, STAG2. This variant is associated with decreased STAG2 protein expression. The analyses of metaphase spreads did not exhibit premature sister chromatid separation; however, delayed sister chromatid cohesion was observed. To further support the pathogenicity of STAG2 variants, we identified two additional female cases from the DECIPHER research database with mutations in STAG2 and phenotypes similar to our patient. Interestingly, the clinical features of these three cases are remarkably similar to those observed in other well-established cohesinopathies. Herein, we suggest that STAG2 is a dosage-sensitive gene and that heterozygous loss-of-function variants lead to a cohesinopathy.

Label-free enumeration, collection and downstream cytological and cytogenetic analysis of circulating tumor cells.

Circulating tumor cells (CTCs) have a great potential as indicators of metastatic disease that may help physicians improve cancer prognostication, treatment and patient outcomes. Heterogeneous marker expression as well as the complexity of current antibody-based isolation and analysis systems highlights the need for alternative methods. In this work, we use a microfluidic Vortex device that can selectively isolate potential tumor cells from blood independent of cell surface expression. This system was adapted to interface with three protein-marker-free analysis techniques: (i) an in-flow automated image processing system to enumerate cells released, (ii) cytological analysis using Papanicolaou (Pap) staining and (iii) fluorescence in situ hybridization (FISH) targeting the ALK rearrangement. In-flow counting enables a rapid assessment of the cancer-associated large circulating cells in a sample within minutes to determine whether standard downstream assays such as cytological and cytogenetic analyses that are more time consuming and costly are warranted. Using our platform integrated with these workflows, we analyzed 32 non-small cell lung cancer (NSCLC) and 22 breast cancer patient samples, yielding 60 to 100% of the cancer patients with a cell count over the healthy threshold, depending on the detection method used: respectively 77.8% for automated, 60-100% for cytology, and 80% for immunostaining based enumeration.

Primitive Spindle Cell Neoplasm of Ileum with Extensive Heterotopic Cartilage, Presenting as Acute Abdomen in a 6-Day-Old Neonate.

Neonatal intestinal masses with spindle cell morphology have broad differential diagnoses and require a multidisciplinary approach to make the final diagnosis. Spindle cell masses with heterotopic cartilage in the gastrointestinal tract are very rare, and, to our knowledge, have not previously been reported in the neonate. Here we present a case of intestinal primitive spindle cell neoplasm with extensive heterotopic cartilage that manifested initially as acute abdomen in a 6-day-old term infant. Plain radiography demonstrated pneumoperitoneum, prompting diagnostic laparotomy that identified a perforated mass involving the midileum. Histologic and immunohistochemical examination demonstrated an infiltrative spindle cell tumor most compatible with infantile fibrosarcoma (IFS) by a process of exclusion, with nodules of mature heterotopic cartilage. Additional staging studies did not reveal any evidence of residual or metastatic disease. Recognition of this rare variant of IFS will aid in differentiation from other neonatal intestinal mesenchymal tumors.

An unusual initial presentation of mantle cell lymphoma arising from the lymphoid stroma of warthin tumor.

BACKGROUND: Warthin tumors presenting concomitantly with a lymphoma is vanishingly rare with only 15 reported cases in English literature. Herein, we report an unusual initial presentation of a mantle cell lymphoma involving the lymphoid stroma of a Warthin tumor. CASE PRESENTATION: A seventy-seven year old otherwise healthy gentleman with a 50-pack year smoking history presents with a slowly enlarging left cheek mass. CT scan of the neck demonstrated a left parotid gland tumor measuring 3.4 cm in greatest dimension. He underwent a left superficial parotidectomy, with subsequent histopathologic examination revealing a Warthin tumor with extensive expansion of the lymphoid stroma. Flow cytometric, immunohistochemical, and cytogenetic studies of the stromal component of the tumor confirmed the presence of a mantle cell lymphoma. Clinical staging demonstrated stage IVa disease, and was considered to be at low to intermediate risk due to the slow growth of the parotid lesion. The patient is undergoing close follow up with repeat PET-CT scans at six months. CONCLUSION: To the best of our knowledge, this is the first well documented collision tumor between mantle cell lymphoma and a Warthin tumor. This case also brings to light the significance of thorough evaluation of the lymphoid component of Warthin tumor.

Erratum to: A case of B-cell acute lymphoblastic leukemia in a child with Down syndrome bearing a t(2;12)(p12;p13) involving ETV6 and biallelic IGH@ rearrangements.

[This corrects the article DOI: 10.1186/s40364-015-0036-1.].

A case of B-cell acute lymphoblastic leukemia in a child with Down syndrome bearing a t(2;12)(p12;p13) involving ETV6 and biallelic IGH@ rearrangements.

BACKGROUND: Rearrangements involving ETV6 (12p13) are among the most common structural abnormalities in pediatric B-cell acute lymphoblastic leukemia (B-ALL) and involve numerous partner genes. Additionally, the t(8;14)(q11.2;q32), which can result in the placement of CEBPD (8q11.2) near the regulatory regions of IGH@ (14q32) and consequent overexpression of CEPBD, occurs at a higher frequency in individuals with Down syndrome-associated ALL (DS-ALL) compared to both the general and pediatric population. The coexistence of cytogenetically detectable ETV6 abnormalities and t(8;14)(q11.2;q32) is a rare occurrence in B-ALL and has only been reported in a single case in the literature. FINDINGS: Herein, we present a case of B-ALL in a 9-year old male with Down syndrome in which conventional cytogenetic analysis revealed two reciprocal translocations: a t(8;14)(q11.2;q32) and a t(2;12)(p12;p13). Interphase and metaphase fluorescence in situ hybridization (FISH) analysis using break apart probes confirmed the involvement of IGH@ and ETV6 in these translocations, respectively. Additionally, interphase FISH revealed a clonal subpopulation bearing biallelic IGH@ rearrangements not observed by conventional cytogenetic analysis. CONCLUSIONS: To the best of our knowledge, this is the first reported case of B-ALL bearing an ETV6 translocation with a partner gene on the short arm of chromosome 2 confirmed by FISH. Additionally, it is the second reported case of t(8;14)(q11.2;q32)-ALL bearing a concomitant, cytogenetically detectable abnormality involving ETV6. This case provides insight into a novel translocation involving ETV6 as well as potentially unique and understudied mechanisms of clonal evolution in pediatric B-ALL.

A Cryptic t(11;14) Translocation in Mantle Cell Lymphoma Highlights the Importance of FISH.

Mantle cell lymphoma (MCL) is a mature B-cell neoplasm composed of monomorphic small to medium-sized atypical lymphocytes arising from naïve mantle zone B-cells, with a generally aggressive and incurable clinical course. The t(11;14)(q13;q32) between IGH@ and CCND1 is present in almost all cases of MCL. Secondary cytogenetic abnormalities are common, and have been associated in some cases with clinical progression. Variant and cryptic t(11;14) translocations have been reported as well. Herein, we present the case of an 80-year old woman with classical MCL, and a cryptic t(11;14) translocation detected by fluorescence in situ hybridization (FISH), and not by conventional cytogenetics. FISH on previously G-banded metaphases showed a cryptic CCND1-IGH@ fusion signal on a derivative chromosome 10, and another fusion signal on one of the abnormal copies of chromosome 11. Cases such as this highlight the importance of FISH studies as part of an algorithmic and multidisciplinary approach to diagnosis.

A (1;19) translocation involving TCF3-PBX1 fusion within the context of a hyperdiploid karyotype in adult B-ALL: a case report and review of the literature.

BACKGROUND: The t(1;19)(q23;p13), which can result in the TCF3-PBX1 chimeric gene, is one of the most frequent translocations in B-acute lymphoblastic leukemia (B-ALL) and is observed in both adult and pediatric populations at an overall frequency of 6%. It can occur in a balanced or unbalanced form and as a sole abnormality is associated with an intermediate prognosis. Additionally, this translocation is observed in the context of hyperdiploid B-ALL, in which case it is associated with a poor prognosis. However, due to different translocation partner genes at chromosomes 1 and 19, distinct subtypes of hyperdiploid B-ALL with t(1;19)/der(19)t(1;19) are recognized based on the presence or absence of the TCF3-PBX1 fusion gene, but the cytogenetic and etiologic differences between the two remain understudied. FINDINGS: We report a case of an adult with a history of relapsed precursor B-ALL whose conventional cytogenetics showed an abnormal female karyotype with both hyperdiploidy and a t(1;19)(q23;p13). Fluorescence in situ hybridization (FISH) on previously G-banded metaphases using the LSI TCF3/PBX1 Dual Color, Dual Fusion Translocation Probe confirmed the presence of the TCF3-PBX1 gene fusion. CONCLUSIONS: This particular pattern with a TCF3-PBX1 fusion within the context of a hyperdiploid karyotype is seen in B-ALL and is usually associated with a poor outcome. This case is one of only a few cases with both hyperdiploidy and a confirmed TCF3-PBX1 fusion, demonstrating the importance of using FISH for proper molecular classification of these cases in order to distinguish them from those with hyperdiploidy but no TCF3-PBX1 fusion gene. Such molecular studies may provide insight into the precise differences between TCF3-PBX1 positive and negative hyperdiploid B-ALL bearing the t(1;19)(q23;p13).

An unusual karyotype in leiomyoma: Case report and literature review.

Vascular leiomyoma are uncommon, clinically benign smooth muscle tumors. Here we report a case of an otherwise typical leiomyoma with unusual cytogenetic changes including t(1;10). Reports from the existing literature suggest that approximately 40-50% of leiomyomas contain nonrandom chromosomal abnormalities of which a subset is tumor specific. t(12;14)(q15;q24) is one of the most common translocations, which occur in approximately 20% of karyotypes. Additional aberrations include del(7)(q22-q32), trisomy 12, and 6p21 rearrangements. Other recurrent abnormalities include monosomy 22, monosomy 10, del(10q), and structural rearrangements of chromosome 3.

Selective demethylation and altered gene expression are associated with ICF syndrome in human-induced pluripotent stem cells and mesenchymal stem cells.

Immunodeficiency, centromeric instability and facial anomalies type I (ICF1) syndrome is a rare genetic disease caused by mutations in DNA methyltransferase (DNMT) 3B, a de novo DNA methyltransferase. However, the molecular basis of how DNMT3B deficiency leads to ICF1 pathogenesis is unclear. Induced pluripotent stem cell (iPSC) technology facilitates the study of early human developmental diseases via facile in vitro paradigms. Here, we generate iPSCs from ICF Type 1 syndrome patient fibroblasts followed by directed differentiation of ICF1-iPSCs to mesenchymal stem cells (MSCs). By performing genome-scale bisulfite sequencing, we find that DNMT3B-deficient iPSCs exhibit global loss of non-CG methylation and select CG hypomethylation at gene promoters and enhancers. Further unbiased scanning of ICF1-iPSC methylomes also identifies large megabase regions of CG hypomethylation typically localized in centromeric and subtelomeric regions. RNA sequencing of ICF1 and control iPSCs reveals abnormal gene expression in ICF1-iPSCs relevant to ICF syndrome phenotypes, some directly associated with promoter or enhancer hypomethylation. Upon differentiation of ICF1 iPSCs to MSCs, we find virtually all CG hypomethylated regions remained hypomethylated when compared with either wild-type iPSC-derived MSCs or primary bone-marrow MSCs. Collectively, our results show specific methylome and transcriptome defects in both ICF1-iPSCs and differentiated somatic cell lineages, providing a valuable stem cell system for further in vitro study of the molecular pathogenesis of ICF1 syndrome. GEO accession number: GSE46030.

Novel three-dimensional in vitro models of ovarian endometriosis.

BACKGROUND: Endometriosis is characterized by the presence of functional endometrial tissue outside of the uterine cavity. It affects 1 in 10 women of reproductive age. This chronic condition commonly leads to consequences such as pelvic pain, dysmenorrhea, infertility and an elevated risk of epithelial ovarian cancer. Despite the prevalence of endometriosis and its impact on women's lives, there are relatively few in vitro and in vivo models available for studying the complex disease biology, pathophysiology, and for use in the preclinical development of novel therapies. The goal of this study was to develop a novel three-dimensional (3D) cell culture model of ovarian endometriosis and to test whether it is more reflective of endometriosis biology than traditional two dimensional (2D) monolayer cultures. METHODS: A novel ovarian endometriosis epithelial cell line (EEC16) was isolated from a 34-year old female with severe endometriosis. After characterization of cells using in vitro assays, western blotting and RNA-sequencing, this cell line and a second, already well characterized endometriosis cell line, EEC12Z, were established as in vitro 3D spheroid models. We compared biological features of 3D spheroids to 2D cultures and human endometriosis lesions using immunohistochemistry and real-time semi-quantitative PCR. RESULTS: In comparison to normal ovarian epithelial cells, EEC16 displayed features of neoplastic transformation in in vitro assays. When cultured in 3D, EEC16 and EEC12Z showed differential expression of endometriosis-associated genes compared to 2D monolayer cultures, and more closely mimicked the molecular and histological features of human endometriosis lesions. CONCLUSIONS: To our knowledge, this represents the first report of an in vitro spheroid model of endometriosis. 3D endometriosis models represent valuable experimental tools for studying EEC biology and the development of novel therapeutic approaches.

One in four individuals of African-American ancestry harbors a 5.5kb deletion at chromosome 11q13.1.

Cloning and sequencing of 5.5 kb deletion at chromosome 11q13.1 from the HeLa cells, tumorigenic hybrids and two fibroblast cell lines have revealed homologous recombination between AluSx and AluY resulting in the deletion of intervening sequences. Long-range PCR of the 5.5 kb sequence in 494 normal lymphocyte samples showed heterozygous deletion in 28.3% of African-American ancestry samples but only in 4.8% of Caucasian samples (p<0.0001). This observation is strengthened by the copy number variation (CNV) data of the HapMap samples which showed that this deletion occurs in 27% of YRI (Yoruba--West African) population but none in non-African populations. The HapMap analysis further identified strong linkage disequilibrium between 5 single nucleotide polymorphisms and the 5.5 kb deletion in people of African ancestry. Computational analysis of 175 kb sequence surrounding the deletion site revealed enhanced flexibility, low thermodynamic stability, high repetitiveness, and stable stem-loop/hairpin secondary structures that are hallmarks of common fragile sites.

Epstein-Barr virus patterns in US Burkitt lymphoma tumors from the SEER residual tissue repository during 1979-2009.

Burkitt lymphoma (BL) occurs at all ages, but the patterns of Epstein-Barr virus (EBV) positivity in relation to human immunodeficiency virus (HIV), immunoprofiles and age have not been fully explored. BL tissues from residual tissue repositories, and two academic centers in the United States were examined by expert hematopathologists for morphology, immunohistochemistry, MYC rearrangement, EBV-encoded RNA (EBER), and diagnosed according to the 2008 WHO lymphoma classification. Analysis was done using frequency tables, Chi-squared statistics, and Student's t-test. Of 117 cases examined, 91 were confirmed as BL. The age distribution was 26%, 15%, 19%, and 29% for 0-19, 20-34, 35-59, 60+ years, and missing in 11%. MYC rearrangement was found in 89% and EBER positivity in 29% of 82 cases with results. EBER positivity varied with age (from 13% in age group 0-19 to 55% in age group 20-34, and fell to 25% in age group 60+ years, p = 0.08); with race (56% in Blacks/Hispanics vs 21% in Whites/Asians/Pacific Islanders, p = 0.006); and by HIV status (64% in HIV positive vs 22% in HIV negative cases, p = 0.03). EBER positivity was demonstrated in about one-third of tumors and it was strongly associated with race and HIV status, and marginally with age-group.