Quantitative analysis of CKS1B mRNA expression and copy number gain in patients with plasma cell disorders

2014 ◽  
Vol 53 (3) ◽  
pp. 110-117 ◽  
Author(s):  
Flavia Stella ◽  
Estela Pedrazzini ◽  
Edgardo Baialardo ◽  
Dorotea Beatriz Fantl ◽  
Natalia Schutz ◽  
...  
Keyword(s):  
Quantitative Analysis ◽  
Mrna Expression ◽  
Plasma Cell ◽  
Copy Number ◽  
Copy Number Gain ◽  
Plasma Cell Disorders

scholarly journals Increased TPSAB1 Copy Number in a Family With Elevated Basal Serum Levels of Tryptase

2021 ◽  
Vol 8 ◽  
Author(s):  
Laura Hernández-Hernández ◽  
Catalina Sanz ◽  
Elena Marcos-Vadillo ◽  
Asunción García-Sánchez ◽  
Esther Moreno ◽  
...  
Keyword(s):  
Gene Expression ◽  
Mrna Expression ◽  
Copy Number ◽  
Family Members ◽  
Copy Number Gain ◽  
Digital Pcr ◽  
Gene Dose ◽  
Control Subjects ◽  
Basal Serum ◽  
The Family

Background: Some recent familial studies have described a pattern of autosomal dominant inheritance for increased basal serum tryptase (BST), but no correlation with mRNA expression and gene dose have been reported.Objective: We analyzed TPSAB1 mRNA expression and gene dose in a four-member family with high BST and in two control subjects.Methods: Blood samples were collected from the family and control subjects. Complete morphologic, immunophenotypical, and molecular bone marrow mast cell (MC) studies were performed. mRNA gene expression and gene dose were performed in a LightCycler 480 instrument. Genotype and CNV were performed by quantitative real-time digital PCR (qdPCR).Results: CNV analysis revealed a hereditary copy number gain genotype (3β2α) present in all the family members studied. The elevated total BST in the family members correlated with a significant increase in tryptase gene expression and dose.Conclusions and Clinical Relevance: We present a family with hereditary α-tryptasemia and elevated BST which correlated with a high expression of tryptase genes and an increased gene dose. The family members presented with atypical MC-mediator release symptoms or were even asymptomatic. Clinicians should be aware that elevated BST does not always mean an MC disorder.

scholarly journals Integrated Epigenetic and Transcriptional Single Cell Analysis of t(11;14) Myeloma and Its BCL2 Dependency

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 725-725
Author(s):  
Noemie Leblay ◽  
Sungwoo Ahn ◽  
Ranjan Maity ◽  
Holly Lee ◽  
Elie Barakat ◽  
...  
Keyword(s):  
Transcription Factors ◽  
B Cell ◽  
Single Cell ◽  
Plasma Cell ◽  
Copy Number ◽  
Acquired Resistance ◽  
Copy Number Gain ◽  
Chromatin Accessibility ◽  
Immunoglobulin Gene ◽  
Cell Markers

Abstract Multiple myeloma is characterized by recurrent chromosomal translocations that involve the immunoglobulin gene enhancers and partners such as the cyclin D genes (CCND1, CCND2 or CCND3) or other genes like WHSC1 and MAF. t(11;14) results in upregulation of CCND1 with unique morphological, phenotypic markers, and drug sensitivity profiles with exquisite sensitivity to BCL2 inhibitors. It is evident now that this unique sensitivity profile is driven by the BH3-proapoptotic protein priming of BCL2 with high BCL2/MCL1 or BCL2/BCL2L1 ratios. However, the epigenetic mechanisms associated with t(11;14) and their impact on genes regulation and clinical response to venetoclax remains elusive. In the present study we compared the transcriptomics (scRNA-seq) and chromatin accessibility (scATAC-seq) of single plasma cells of MM patients with and without t(11;14) as well as pre- and post-venetoclax exposure in order to establish the epigenomic signature of t(11;14) and/or BCL2-sensitivity in myeloma. Serial BM aspirates (n=24) were collected from 15 relapsed or refractory myeloma patients (RRMM); harboring t(11;14) (n=6 pairs) and 9 without this translocation prior to initiation of salvage therapy and at time of relapse. All t(11;14) MM patients were treated with venetoclax. Unbiased chromatin accessibility and mRNA profiling of CD138 pos cells were performed using the chromium single cell ATAC and RNA-Seq 3' solution (10x Genomics), respectively. Cell Ranger, Seurat and ArchR were used for sample de-multiplexing, barcode processing, single-cell 3' gene, peaks counting, and data analysis. We first compared the scATAC-seq and scRNA-seq profiles found in CD138 pos MM cell isolated from patients harboring t(11;14) with the one obtained in patients without this translocation. As expected, t(11:14) patients had high chromatin accessibility at the CCND1 locus and high mRNA expression. Differentially accessible chromatin analysis identified 147518 peaks that were specific to t(11;14) patients. Of interest, motifs enrichment analysis of accessible peaks identified a "B cell-like" motifs signature with enriched TFs motifs such as TCF4 and PAX5 in t(11;14) patients compared to non t(11:14) enriched for IRF and STAT family of motifs. The integration of the scATAC-seq and scRNA-seq data confirmed the B cell signature of t(11;14) patients with upregulation of B cell markers such as MS4A1, VPREB3, CD79A, CD19, and down-regulation of plasma cell markers such as TDO2, EFEMP1, CD28, SLAMF7, and IL6R. Additionally, we found PAX1, PAX5, TCF3, TCF5, and SPI1 transcription factors to be highly expressed in t(11;14) while the non t(11:14) were enriched for IRF1-9 transcription factors. Of interest, the clustering analysis performed on scATAC-seq data identified 3 non t(11;14) patients with a chromatin accessibility profile similar to that of t(11;14) patients. They expressed B cell markers (PAX5, VPREB3 or FCRLA), overexpressed BCL2 and we are currently examining whether this B cell-like epigenetic signature determines sensitivity to venetoclax. In order to define the epigenetic contribution to the acquired resistance to venetoclax in t(11;14) myeloma, we compared the chromatin accessibility profiles of t(11;14) patients pre- vs. post-venetoclax treatment. Enriched motifs within accessible peaks differed significantly between pre- and post-venetoclax with RELA, REL, RELB and EGR1 motifs predominantly enrichmed in the pre-samples in contrast to JUN, JUNB, JUND and FOSL1/L2 motifs enrichment in the post-samples. Of note, integration analysis of scRNAseq (differentially expressed genes) and ATACseq data (differentially accessible peaks) identified MCL1 and ENSA (a gene 60 Kb centromeric to MCL1 on chr1q) as the top enriched genes and peaks in resistant samples suggesting that copy number gain at the MCL1 locus (which we confirmed by single cell CNV analysis) rather than epigenetic modifications is likely the main determinant of acquired resistant to venetoclax in t(11;14) MM. In the current study we have defined the epigenetic regulome and transcriptome associated with t(11;14) myeloma and its relatedness to B cell rather than plasma cell biology. Our studies also suggest that acquired resistance to venetoclax is largely driven by copy number gain at the MCL1 locus. Disclosures Bahlis: Karyopharm: Consultancy, Honoraria; Amgen: Consultancy, Honoraria; Takeda: Consultancy, Honoraria; Sanofi: Consultancy, Honoraria; Abbvie: Consultancy, Honoraria; Janssen: Consultancy, Honoraria; BMS/Celgene: Consultancy, Honoraria; GlaxoSmithKline: Consultancy, Honoraria; Genentech: Consultancy; Pfizer: Consultancy, Honoraria. Neri: BMS: Consultancy, Honoraria; Janssen: Consultancy, Honoraria; Sanofi: Consultancy, Honoraria; Amgen: Consultancy, Honoraria.

Exome Sequencing To Define A Genetic Signature Of Plasma Cells In Systemic AL Amyloidosis

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 3098-3098 ◽  
Author(s):  
Brian A Walker ◽  
Dorota Rowczienio ◽  
Eileen M Boyle ◽  
Christopher P Wardell ◽  
Sajitha Sachchithanantham ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Exome Sequencing ◽  
Plasma Cell ◽  
Copy Number ◽  
Monoclonal Gammopathy ◽  
Plasma Cells ◽  
Chromosomal Abnormalities ◽  
Light Chains ◽  
Al Amyloidosis ◽  
Plasma Cell Disorders

Abstract Systemic amyloid light chain amyloidosis (AL) is characterized by the deposition of immunoglobulin light chains as amyloid fibrils in different organs, where they form toxic protein aggregates. Most AL patients have relatively low levels of circulating free light chains and bone marrow plasmacytosis. The underlying disease is a plasma cell disorder, likely a monoclonal gammopathy, but limited data are available on the biology of the plasma cell clone underlying AL and existing studies have concentrated on chromosomal abnormalities. Many of the chromosomal abnormalities identified in AL are also seen in other plasma cell disorders, such as monoclonal gammopathy of undetermined significance (MGUS) and myeloma. These abnormalities include translocations involving the IGH locus, gains of 1q and deletions of 13q and 17p. Fluorescence in situhybridization studies have identified the translocation t(11;14) to be more frequent in AL and hyperdiploidy to be rare. The causal link between genetic changes in plasma cells and light chain instability remains unknown and progression to symptomatic myeloma is rare. We report the initial findings of the first exome sequencing to define the plasma cell signature in AL and compared this to MGUS and myeloma. CD138+ cells were selected using either EasySep (Stem Cell Technologies) or MACSort (Miltenyi) from the bone marrow of 18 AL patients and 5 MGUS patients. DNA was extracted from the CD138+ cells using the AllPrep kit (Qiagen). Non-involved DNA was isolated from peripheral white blood cells using the Flexigene kit (Qiagen). 200 ng DNA was subjected to exome sequencing using NEBNext kit (NEB) and SureSelect Human All Exon kit v5 and sequenced using 76-bp paired end reads. Fastq files were aligned to the reference genome using BWA and Stampy aligners. BAM files were recalibrated using the GATK and deduplicated using Picard. Paired tumour/normal BAMs were realigned together using the GATK indel realigner and SNVs were called using Mutect. Copy number data were estimated using the R package ExomeCNV. The median depth across all samples was 42x with 97% of the exome covered at 1x and 72% covered at 20x. Exome data to determine the cytogenetic groups of AL samples identified 42% hyperdiploid and 21% with t(11;14). The AL samples with t(11;14) did not contain any other copy number abnormalities. Exome sequencing on samples from patients with MGUS and myeloma was also performed to compare the genetic makeup and mutation spectrum of these well characterised plasma cell neoplasias with AL samples. MGUS samples had a median of 30 acquired nonsynonymous variants (range 24-189) and AL amyloidosis samples had a median of 17 acquired nonsynonymous variants (range 4-44). The AL samples had four recurrent mutations in PCMTD1 (n=3; L267F, P266S and M187I), C21orf33 (n=2; E72K), NLRP12 (n=2; L1018P, W959* ) and NRAS (n=2; Q61R, Q61H). In this small dataset, only 5 genes were mutated in both the MGUS and AL samples (DNMBP, FRG1, HIST1H1B, KRTAP4-11 and MCCC1). In order to assess the similarity (or differences) of plasma cells in AL to malignant plasma cells in general, we compared them to a random sampling of 20 multiple myeloma samples which had also been exome sequenced (median number of acquired nonsynonymous variants = 39 vs. 17 in AL samples). This revealed that the AL contained 21 mutated genes in common with the myeloma cohort, including DIS3 and NRAS. There were two DIS3 mutations in one AL sample at c.379D>E (p.D479E) and c.1999A>T (p.M667L), both of which were in the Ribonuclease II/R catalytic domain. Data on correlation of gene mutations and organ involvement in AL amyloidosis will be presented. We conclude that exome sequencing identifies a genetic signature of AL amyloidosis which is similar to other plasma cell disorders. This not only includes copy number abnormalities and translocations but also a similar number of nonsynonymous mutations to MGUS and fewer than the advanced myeloma samples. Study of further samples is in progress. Disclosures: No relevant conflicts of interest to declare.

HER2 activity in solid tumors.

2017 ◽  
Vol 35 (15_suppl) ◽  
pp. e23121-e23121
Author(s):  
Lara Ann Kujtan ◽  
Scott Morris ◽  
Janakiraman Subramanian
Keyword(s):  
Breast Cancer ◽  
Mrna Expression ◽  
Solid Tumors ◽  
Copy Number ◽  
Significant Proportion ◽  
Copy Number Gain ◽  
Tumor Type ◽  
Breast Cancers ◽  
Distinctive Pattern ◽  
Tumor Types

e23121 Background: ERBB2 is a member of the human epidermal growth factor receptor family. HER2 expression that is immunohistochemistry (IHC) 3+ or IHC 2+ with copy number gain is an effective predictor for treatment with trastuzumab in breast and gastroesophageal cancers. Here we present an analysis on HER2 activity measured by IHC, mRNA expression and copy number variation (CNV) across a variety of solid tumors. Methods: The study population consists of patients diagnosed with solid tumors (n = 856) that underwent a Paradigm Diagnostic Cancer Test during 2016. We then analyzed tumor types that tested positive for HER2 by IHC (3+), CNV and/or mRNA expression. Results: We identified 365 (43%) patients positive for HER2 by IHC, 258 (30%) had high HER2mRNA expression and 41 (5%) had amplification by CNV. Seventy-five patients were HER2 IHC 3+ or 2+/CNV positive. The proportion of HER2 IHC 3+ or 2+/CNV positive tumors in each tumor type was as follows: breast cancer 41 (18.5%), NSCLC 10 (8.1%), colorectal 6 (6.7%), esophago-gastric 3 (10%,) urothelial/bladder 3 (16%), biliary 2 (28.6%), ovarian cancer 2 (5.1%) and pancreas 1 (3.1%). Using copy number gain as the gold standard, across all tumor types, an IHC of 3+ had a 90.2% sensitivity and a 95.6% specificity. In the same analysis with breast cancers omitted, the sensitivity was 75%, and specificity 96.8%. Whereas high mRNA expression had a sensitivity of 97.6% and specificity of 73.3%, omitting breast cancers, the sensitivity was 93.8% and specificity 73.2%. Conclusions: HER2 activity was identified in a wide variety of solid tumors and a small but significant proportion of these tumors maybe candidates for treatment with HER2 inhibitors such as trastuzumab. Our analysis also identified that HER2 activity in breast cancers has a distinctive pattern which was not seen in other tumor types. HER2 IHC 3+ expression was much less sensitive among other tumor types compared to breast cancer. mRNA expression, while remaining sensitive among other tumor types, is not specific, even among breast cancer patients. Our analysis also identified that HER2 activity in breast cancers has a distinctive pattern which was not seen in other tumor types.

Glutathione S-transferase P1 mRNA expression in plasma cell disorders and its correlation with polymorphic variants and clinical outcome

2013 ◽  
Vol 37 (5) ◽  
pp. 671-674 ◽  
Author(s):  
Flavia Stella ◽  
Natalia Weich ◽  
Julieta Panero ◽  
Dorotea Beatriz Fantl ◽  
Natalia Schutz ◽  
...  
Keyword(s):  
Clinical Outcome ◽  
Mrna Expression ◽  
Plasma Cell ◽  
Glutathione S Transferase ◽  
Plasma Cell Disorders ◽  
Polymorphic Variants

scholarly journals Combination of Genome-Wide Polymorphisms and Copy Number Variations of Pharmacogenes in Koreans

2021 ◽  
Vol 11 (1) ◽  
pp. 33
Author(s):  
Nayoung Han ◽  
Jung Mi Oh ◽  
In-Wha Kim
Keyword(s):  
Copy Number ◽  
Genome Wide Association Study ◽  
Copy Number Gain ◽  
Copy Number Variations ◽  
Gene Gain ◽  
Single Nucleotide Variants ◽  
Single Nucleotide ◽  
Haplotype Blocks ◽  
Genome Wide ◽  
Control And Prevention

For predicting phenotypes and executing precision medicine, combination analysis of single nucleotide variants (SNVs) genotyping with copy number variations (CNVs) is required. The aim of this study was to discover SNVs or common copy CNVs and examine the combined frequencies of SNVs and CNVs in pharmacogenes using the Korean genome and epidemiology study (KoGES), a consortium project. The genotypes (N = 72,299) and CNV data (N = 1000) were provided by the Korean National Institute of Health, Korea Centers for Disease Control and Prevention. The allele frequencies of SNVs, CNVs, and combined SNVs with CNVs were calculated and haplotype analysis was performed. CYP2D6 rs1065852 (c.100C>T, p.P34S) was the most common variant allele (48.23%). A total of 8454 haplotype blocks in 18 pharmacogenes were estimated. DMD ranked the highest in frequency for gene gain (64.52%), while TPMT ranked the highest in frequency for gene loss (51.80%). Copy number gain of CYP4F2 was observed in 22 subjects; 13 of those subjects were carriers with CYP4F2*3 gain. In the case of TPMT, approximately one-half of the participants (N = 308) had loss of the TPMT*1*1 diplotype. The frequencies of SNVs and CNVs in pharmacogenes were determined using the Korean cohort-based genome-wide association study.

Liquid biopsy: an evolving paradigm for the biological characterisation of plasma cell disorders

Leukemia ◽  
2021 ◽  
Author(s):  
Sridurga Mithraprabhu ◽  
Maoshan Chen ◽  
Ioanna Savvidou ◽  
Antonia Reale ◽  
Andrew Spencer
Keyword(s):  
Plasma Cell ◽  
Liquid Biopsy ◽  
Plasma Cell Disorders
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